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  5. 双打击淋巴瘤的分子特征、诊断与治疗进展综述

双打击淋巴瘤的分子特征、诊断与治疗进展综述

深度研究HealerEdge发表于 2025年11月03日 12:192阅读
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1. 双打击淋巴瘤的定义与流行病学特征

1.1 核心定义与分类演变

双打击淋巴瘤（Double-Hit Lymphoma, DHL）是一类侵袭性B细胞淋巴瘤，其核心特征为MYC癌基因与BCL2和/或BCL6癌基因同时发生重排 1。最初，DHL主要指弥漫大B细胞淋巴瘤（DLBCL）或具有DLBCL形态学特征的淋巴瘤，伴有MYC重排和BCL2重排。随着对淋巴瘤分子生物学认识的深入，2016年世界卫生组织（WHO）对淋巴造血组织肿瘤分类进行了更新，将MYC、BCL2和/或BCL6基因重排的侵袭性B细胞淋巴瘤归类为伴有MYC和BCL2和/或BCL6重排的高级别B细胞淋巴瘤（High-Grade B-cell Lymphoma with MYC and BCL2 and/or BCL6 Rearrangements, HGBL-DH/TH）1。

WHO 2022年分类和国际共识分类（International Consensus Classification, ICC）进一步细化了HGBL的定义。尽管两者在某些方面存在差异，但都强调了MYC与BCL2和/或BCL6基因重排在诊断中的关键作用 2。这类淋巴瘤通常表现出侵袭性的临床行为和不良预后，对传统化疗方案反应不佳 12。值得注意的是，MYC重排可涉及免疫球蛋白（Ig）基因座（如IgH）或非Ig基因座，且不同重排类型可能预示不同的预后 3。例如，伴有Ig基因座MYC重排的淋巴瘤预后通常较差 3。

与DHL需要明确基因重排不同，双表达淋巴瘤（Double-Expressor Lymphoma, DEL）是指在免疫组织化学（IHC）检测中同时过表达MYC蛋白和BCL2蛋白的DLBCL，无论是否存在基因重排 4。DEL在DLBCL中约占19-21.6% 45，通常表现为活化B细胞样（ABC）DLBCL亚型，与较差的预后相关 45。虽然DHL和DEL都预示着不良预后，但它们的分子基础存在显著差异：DHL是基于基因重排的诊断，具有明确的遗传学特征；而DEL是基于蛋白质表达水平的诊断，可能存在也可能不存在相应的基因重排。这种区别在临床实践中至关重要，因为基因重排的存在（即DHL）通常预示着比单纯蛋白过表达（即DEL）更为侵袭性的疾病进程和更差的治疗反应 2。例如，一项研究发现，结直肠DLBCL中一小部分病例属于DHL，表现出显著的侵袭性 6。此外，基因表达谱分析也揭示了DHL独特的生物学特征，DHITsig（double-hit signature）基因表达谱能够区分出伴有MYC和BCL2重排的GCB-DLBCL以及其他GCB-DLBCL，并与不良预后密切相关 7。

1.2 流行病学与疾病负担

双打击淋巴瘤（DHL）作为侵袭性B细胞淋巴瘤的一个亚型，其发病率在全球范围内呈现一定特征。尽管DHL在所有非霍奇金淋巴瘤（NHL）中占比不高，但其预后极差，因此对公共健康构成显著挑战。DHL是DLBCL中一个预后不良的亚群，其明确的发生率在文献中通常与DLBCL合并报告。DLBCL是全球最常见的非霍奇金淋巴瘤亚型，约占所有NHL的三分之一，各国发病率在20%至50%之间波动 8。根据美国癌症登记数据，DLBCL的年龄标准化发病率为每10万人7.2例 8。鉴于DLBCL的总体发病率，DHL虽然是其中一个较小的亚群，但其绝对病例数仍然不容忽视。

全球范围内，DLBCL的发病率呈现出一定的地域和时间趋势。在20世纪上半叶，NHL的发病率有所上升，但近期已趋于稳定 8。然而，一些历史发病率较低的地区，如亚洲，DLBCL的发病率有上升趋势，且由于发达国家人口老龄化，未来DLBCL发病率预计将进一步增加 8。这种趋势也可能间接影响DHL的流行病学，因为DHL是DLBCL的亚型。例如，一项来自台湾的研究发现，在282例DLBCL病例中，47例（16.7%）存在MYC易位，其中24例为DHL或三打击淋巴瘤（THL），这表明在特定地区DHL的发生率可能高于西方国家的平均水平 9。这项研究还指出，台湾地区三分之二的DHL/THL患者伴有BCL6基因重排，这与西方国家的数据有所不同，提示了地域差异性 9。

尽管目前尚未有全球疾病负担（GBD）研究直接针对DHL的特定数据，但考虑到DLBCL整体对生存质量的影响，DHL作为其中预后最差的亚型之一，其对患者生存质量和寿命的影响更为显著。非霍奇金淋巴瘤（NHL）通常会增加癌症诊断和死亡数量的近3% 10。随着DLBCL发病率的增长和患者寿命的延长，预计到2020年至2025年，美国和西欧的DLBCL和滤泡性淋巴瘤（FL）的事件和流行病例总数将增加 11。由于DHL对标准化疗方案的耐药性以及高复发率，其患者往往面临更短的生存期和更高的疾病相关死亡风险 12。这意味着DHL不仅对患者个体造成巨大的生理和心理负担，也对医疗系统和社会资源带来沉重压力。此外，DHL患者在诊断时常伴有高国际预后指数（IPI）评分、高乳酸脱氢酶（LDH）水平和结外侵犯等不良预后因素，进一步加剧了其疾病负担。对DHL流行病学特征的深入了解有助于优化筛查策略、制定更精准的治疗方案，并最终减轻其对全球健康的影响。

2. 分子生物学机制与病理特征

2.1 驱动基因与信号通路异常

双打击淋巴瘤（DHL）的侵袭性表型和不良预后根源于其独特的分子生物学机制，核心在于MYC与BCL2和/或BCL6癌基因的协同失调。这些基因的异常主要通过基因重排发生，但也涉及其他复杂的调控机制，共同驱动肿瘤细胞的增殖和生存。

MYC重排的分子机制

MYC癌基因的重排是DHL的标志性特征，其通常涉及MYC基因与高表达的免疫球蛋白（Ig）基因座（如IgH、Igκ、Igλ）或非Ig基因座（如MYC::PAX5、MYC::IGH等）发生易位 313。这些重排使得MYC基因脱离其正常的转录调控，置于强大的增强子（特别是超增强子）控制之下，导致MYC的组成型高表达 3。超增强子是一组密集排列的增强子，能够驱动基因在特定细胞类型中极高的表达水平。研究表明，MYC重排在DHL中具有不同的架构特征，包括较高频率的非Ig重排以及MYC::IGH重排的独特模式 3。这些独特的MYC重排模式可能受到DHL细胞中MYC伴侣基因座体细胞超突变水平高的影响，从而为这些重排的形成提供了更多机会 3。此外，非编码突变也可能通过超增强子重定向，影响MYC等基因的表达，导致淋巴瘤进展中蛋白质合成的失调 14。

MYC作为一个转录因子，在细胞增殖、生长、代谢和凋亡中发挥关键作用。MYC的异常激活导致细胞周期加速、蛋白质合成增加、代谢重编程，并阻断分化，从而赋予肿瘤细胞极强的增殖能力。

BCL2/BCL6重排的分子机制

除了MYC，BCL2或BCL6的重排也是DHL的关键驱动因素。

  • BCL2重排：BCL2基因重排通常导致其过度表达，主要通过抑制细胞凋亡来促进肿瘤细胞的存活。BCL2重排最常见的是t(14;18)易位，使得BCL2基因置于IgH增强子调控之下。BCL2蛋白作为一种抗凋亡蛋白，通过与BAX和BAK等促凋亡蛋白结合，阻止细胞色素c从线粒体释放，从而抑制细胞凋亡。
  • BCL6重排：BCL6是一个转录抑制因子，在生发中心（GC）B细胞的形成和功能中至关重要。BCL6重排常发生在GCB型DLBCL中，导致BCL6的异常表达，干扰其在B细胞分化中的正常作用。BCL6可以抑制BCL2的表达，通过与转录激活因子Miz1相互作用，结合BCL2启动子区并抑制Miz1诱导的BCL2表达 15。在滤泡性淋巴瘤（FL）和DLBCL中，BCL6对BCL2的抑制作用丧失，导致两种蛋白的病理性共表达，这可能是由于BCL2染色体易位、Miz1失调或BCL2启动子区体细胞突变等机制造成的 15。

协同致癌作用

MYC与BCL2/BCL6的协同作用是DHL恶性生物学行为的核心。

  • MYC与BCL2协同：MYC的持续高表达驱动细胞快速增殖，产生巨大的细胞死亡压力（如复制应激和DNA损伤）。BCL2的过表达通过抑制MYC诱导的细胞凋亡，解除这种限制，使得MYC驱动的细胞增殖得以持续，并促进肿瘤细胞存活 16。这种“增殖-存活”组合赋予了DHL细胞强大的生存优势和对传统化疗的抵抗性。
  • MYC与BCL6协同：BCL6的过度激活可以通过多种机制促进肿瘤发生。例如，BCL6通过抑制多个效应基因，包括那些参与细胞周期检查点、DNA损伤反应和细胞凋亡的基因，从而协同MYC促进细胞增殖。此外，BCL6在生发中心B细胞分化中的作用，也可能与MYC共同维持淋巴瘤细胞的“生发中心样”特征，使其保持未分化状态并逃避免疫监视。
  • 蛋白质合成失调：非编码突变导致超增强子重定向，影响PAX5/ZCCHC7基因对的表达，进而通过改变非编码5.8S核糖体RNA（rRNA）加工导致核仁失调，重新编程蛋白质合成，从而引发淋巴瘤蛋白质组的致癌性变化 14。这种蛋白质合成的失调与MYC的高表达密切相关，共同促进了肿瘤细胞的快速生长。

这些复杂的分子机制不仅解释了DHL的侵袭性，也为开发针对这些关键驱动因素的靶向治疗提供了理论基础。例如，针对MYC、BCL2或PI3Kδ（下游信号通路）的抑制剂正在研究中，以期克服现有治疗的局限性 1718。

2.2 肿瘤微环境与免疫逃逸

双打击淋巴瘤（DHL）的恶性进展和对治疗的抵抗不仅源于其内在的遗传学异常，也与肿瘤微环境（TME）的复杂调控及其介导的免疫逃逸机制密切相关。TME由肿瘤细胞、免疫细胞（如T细胞、B细胞、巨噬细胞、树突状细胞等）、间充质细胞、血管以及细胞外基质组成，这些组分通过复杂的相互作用网络共同影响肿瘤的发生、发展和对治疗的反应。

CD11c+树突状细胞（DCs）的改变

在DHL的TME中，CD11c+树突状细胞（Dendritic Cells, DCs）的数量和功能异常被认为是影响预后的重要因素。研究发现，DLBCL患者肿瘤微环境中CD11c+树突状细胞数量的减少与DHL/THL基因型显著相关，并且是独立的不良预后指标 19。CD11c+ DCs作为专业的抗原提呈细胞（APCs），在启动和调节抗肿瘤免疫反应中发挥关键作用 20。它们能够捕获、加工和提呈肿瘤抗原，激活T细胞，从而介导有效的抗肿瘤免疫。因此，CD11c+ DCs数量的减少可能削弱T细胞的活化，导致免疫监视功能下降，有利于肿瘤细胞的免疫逃逸。

进一步的研究揭示了CD11c+ DCs功能受损的潜在分子机制。例如，PIKfyve是一种在CD11c+细胞中表达的基因，其高表达与患者对免疫检查点抑制剂（ICB）治疗反应不佳有关 21。PIKfyve的敲除或特异性抑制剂（如apilimod）能够增强CD11c细胞的功能，包括通过选择性改变非经典NF-κB通路来促进T细胞免疫，并增强ICB在肿瘤模型中的疗效 21。这表明PIKfyve可能通过负向调节CD11c细胞功能，促进DHL的免疫逃逸。此外，PTPN22（一种酪氨酸磷酸酶）在DC中的缺失也能增强抗肿瘤免疫，提升抗PD-L1治疗效果，提示其在DCs中可能负向调节DC的增殖和抗原提呈能力 22。这些发现提示，靶向调控CD11c+ DCs的功能，有望成为DHL免疫治疗的新策略。

免疫检查点分子表达及肿瘤浸润淋巴细胞功能异常

DHL的TME中免疫检查点分子的表达模式与免疫逃逸密切相关。程序性死亡配体1（PD-L1）是重要的免疫检查点分子，其在肿瘤细胞或肿瘤微环境中的表达可抑制T细胞功能。然而，与非MYC/DH易位的LBCL相比，DHL患者肿瘤细胞的PD-L1蛋白和mRNA表达水平显著较低 23。这一发现可能提示，MYC和DH易位的LBCL患者可能从PD-L1/PD-1抑制剂治疗中获益较少 23。另有研究表明，高级别B细胞淋巴瘤（HGBL）中MYC和BCL2易位的肿瘤，其肿瘤相关巨噬细胞（TAMs）和PD-L1的表达也较低，这可能定义了一个“非炎症性”亚组，可能对免疫检查点阻断治疗不敏感 24。

尽管如此，部分研究仍发现PD-L1在DHL中存在表达，且PD-L1的阳性率在DHL中高于DLBCL-NOS，提示PMBCL和DHL本身可能对免疫治疗敏感 25。此外，IMPDH2在DHL细胞中的高表达与肿瘤微环境中T细胞浸润减少呈负相关，并且IMPDH2的高表达常伴随PD-L1的高表达 26。IMPDH2通过激活PI3K/AKT/mTOR信号通路促进淋巴瘤进展，并可能通过影响TME中的免疫细胞功能间接促进免疫逃逸 26。

除了PD-L1，DHL肿瘤细胞还可能通过上调其他免疫抑制分子或下调免疫刺激分子来促进免疫逃逸。例如，某些情况下，药物诱导的非整倍体/多倍体（DIAP）细胞在MYC/BCL2成瘾性DLBCL中被发现，这些细胞能够上调DNA损伤、DNA复制和免疫逃逸通路，从而导致治疗抵抗和疾病复发 27。这表明，肿瘤细胞自身的基因组不稳定性和对治疗的适应性改变也可能贡献免疫逃逸，例如通过改变细胞表面的分子表达，逃避T细胞的识别和杀伤。

总的来说，DHL的肿瘤微环境是一个高度复杂的生态系统，其中CD11c+ DCs数量的减少、PD-L1表达的异质性以及肿瘤细胞自身的免疫逃逸机制共同促进了DHL的侵袭性和对传统治疗的抵抗。深入理解这些相互作用机制，有望为开发针对DHL的有效免疫疗法提供新的靶点和策略。

2.3 组织形态与细胞起源

双打击淋巴瘤（DHL）的病理学特征反映了其高度侵袭性的生物学行为。在形态学上，DHL常表现出高级别淋巴瘤的特征，其细胞通常较大，核分裂象活跃，胞浆丰富，并伴有显著的核仁，形态多呈母细胞样（blastoid）或免疫母细胞样（immunoblastic）。母细胞样形态的肿瘤细胞具有圆形或卵圆形核，染色质细腻，核仁明显，类似淋巴母细胞。免疫母细胞样细胞则更大，胞浆更丰富，核呈偏位，核仁突出。这些高分级形态学特征与伯基特淋巴瘤（Burkitt lymphoma, BL）和弥漫大B细胞淋巴瘤（DLBCL）之间存在形态学重叠，这也是世界卫生组织（WHO）将这类淋巴瘤归类为“伴有MYC和BCL2和/或BCL6重排的高级别B细胞淋巴瘤”（HGBL-DH/TH）的原因之一。

在免疫表型方面，DHL通常起源于生发中心B细胞（GCB）亚型，这一特征可通过免疫组化（IHC）染色进行识别。经典的GCB免疫表型包括CD10阳性、BCL6阳性，且MUM1/IRF4阴性。CD10是生发中心B细胞的标志物，其阳性表达提示细胞来源于生发中心。BCL6是生发中心B细胞成熟和维持的关键转录因子 28。然而，DHL的免疫表型并非完全同质。虽然大多数DHL病例表现为GCB亚型 29，但也有部分病例可能显示为非GCB（如活化B细胞样，ABC）表型，但这相对较少见。此外，Ki-67增殖指数通常极高，反映了MYC驱动下细胞的高度增殖活性。

DHL与非特指型弥漫大B细胞淋巴瘤（DLBCL-NOS）在病理学上存在显著差异，尤其是在预后方面。DLBCL-NOS是一个异质性疾病群，根据细胞起源可分为GCB和ABC两大亚型，其预后和治疗反应各异 3031。虽然DLBCL-NOS中也可能出现MYC或BCL2的蛋白表达（即DEL），但仅当存在MYC和BCL2/BCL6基因重排时才被诊断为DHL 29。DHL的基因重排特征是其区别于DLBCL-NOS的关键。FISH（荧光原位杂交）是检测这些基因重排的金标准，是DHL诊断不可或缺的一部分 32。

形态学上，DLBCL-NOS的细胞形态多样，可表现为中心母细胞、免疫母细胞或间变性细胞等，但总体上其高分级形态特征和核分裂活性不如DHL突出。免疫组化方面，DLBCL-NOS的GCB亚型也表现为CD10+、BCL6+，但其MYC和BCL2蛋白表达水平或Ki-67指数可能低于DHL。重要的是，即使免疫组化表现为MYC和BCL2双表达（DEL），也必须通过FISH证实基因重排才能确诊为DHL。这种基因重排的存在使得DHL比DEL或DLBCL-NOS具有更具侵袭性的临床过程和更差的预后 29。例如，一项研究显示，DHL患者的4年总生存率仅为0%，远低于DLBCL-NOS（71.9%）和DEL（46.9%）患者 29。因此，准确区分DHL与DLBCL-NOS对于指导临床治疗和评估预后至关重要。

3. 诊断标准与检测技术

双打击淋巴瘤（DHL）的准确诊断对于及时采取恰当的治疗策略至关重要，因为其预后显著差于其他类型的侵袭性B细胞淋巴瘤。DHL的诊断是一个多维度、综合性的过程，涉及病理形态学、免疫组化以及分子遗传学检测的紧密结合。

3.1 多维度诊断流程

DHL的诊断流程通常遵循从形态学初筛到分子确诊的步骤，以确保识别出具有MYC和BCL2和/或BCL6基因重排的侵袭性B细胞淋巴瘤。

病理形态学（HE染色）

初步诊断始于组织活检样本的苏木精-伊红（HE）染色。DHL的形态学特征常表现为高级别B细胞淋巴瘤的典型表现，细胞体积大，核仁明显，核分裂活跃，胞浆丰富。细胞形态多样，可呈母细胞样（blastoid）或免疫母细胞样（immunoblastic），与伯基特淋巴瘤（Burkitt lymphoma, BL）和弥漫大B细胞淋巴瘤（DLBCL）存在形态学重叠。这些高度增殖的形态学特点提示了疾病的侵袭性，并促使进行进一步的免疫组化和分子检测。一些研究指出，BCLU/母细胞样形态与DHL/单打击淋巴瘤（SHL）具有显著相关性，这表明形态学观察在初筛中具有一定的价值 32。

免疫组化（IHC）

免疫组化是DHL诊断流程中的关键环节，用于评估相关蛋白的表达情况。

  • MYC和BCL2蛋白表达：通过IHC检测MYC和BCL2蛋白的过表达是筛查潜在DHL病例的重要步骤。通常设定MYC表达阈值为≥40%，BCL2表达阈值为≥50% 33。当同时检测到MYC和BCL2蛋白过表达时，即为双表达淋巴瘤（DEL）。DEL在DLBCL中约占20%-30% 34，尽管其预后通常较差，但并非所有DEL都伴有基因重排。一项研究表明，在40例DLBCL病例中，有11例（27.5%）为双表达淋巴瘤 33。MYC和BCL2蛋白的共同表达是预后不良的独立因素 3536。
  • Ki67增殖指数：Ki67是一种细胞增殖标志物，其高表达（通常>70%或>90%）提示细胞高度增殖，这是DHL的典型特征之一 19。Ki67增殖指数的评估有助于区分侵袭性更强的淋巴瘤。
  • 细胞起源标志物：为了确定细胞起源亚型（Cell-of-Origin, COO），通常还会检测CD10、BCL6和MUM1/IRF4。大多数DHL病例起源于生发中心B细胞（GCB）亚型，因此常表现为CD10阳性、BCL6阳性、MUM1阴性 3237。GCB表型与DHL基因型呈显著相关性，在筛选DHL/THL基因型时，GCB亚型被认为是重要的危险因素 19。

分子检测

分子检测是确诊DHL的决定性环节，用于证实关键癌基因的重排。

  • 荧光原位杂交（FISH）：FISH是检测MYC、BCL2和BCL6基因重排的金标准 1937。通过特异性探针标记这些基因位点，可以直接观察到染色体易位。DHL的诊断要求存在MYC基因重排以及BCL2和/或BCL6基因重排 37。MYC基因重排在DLBCL中发病率约为12.3% 35，而DHL（MYC和BCL2重排）的发生率约为6.6% 35，或5%-7% 34。不同研究中DHL的检出率有所差异，这可能与研究人群、检测方法和定义标准有关。
  • 二代测序（NGS）：NGS技术可以提供更全面的基因突变谱分析，包括单核苷酸变异（SNV）、拷贝数变异（CNV）和复杂的基因重排。虽然FISH仍是诊断基因重排的金标准，但NGS在揭示DHL的复杂分子背景，如伴随突变（TP53、EZH2、NOTCH1等）及其对预后和治疗反应的影响方面具有巨大潜力。例如，通过NGS可以发现超增强子易位等复杂机制，导致MYC的异常激活。

综合来看，DHL的诊断是一个迭代过程。形态学和免疫组化（特别是MYC/BCL2双表达和高Ki67指数，以及GCB表型）可作为高风险病例的初筛工具，对于符合这些标准的病例，必须进行FISH检测以确诊基因重排，从而实现DHL的精确诊断。对于资源有限的地区，基于形态学、IHC（MYC/BCL2共表达、GCB表型）的筛查模型具有较高的敏感性和特异性，可优化FISH检测的必要性评估 32。

3.2 筛查策略与资源适配

在资源有限的地区，进行全面的分子诊断（特别是荧光原位杂交，FISH）可能面临成本高昂、技术复杂和设备不足等挑战。因此，开发一种高效且经济的筛查策略，以识别最可能罹患双打击淋巴瘤（DHL）的患者，并优化FISH检测的必要性，对于改善这些地区的DHL诊断和治疗至关重要。

基于形态学特征的初筛

形态学检查是病理诊断的第一步，也是最基础的环节。在资源受限的环境中，经验丰富的病理医师可以利用苏木精-伊红（HE）染色切片对DHL高风险病例进行初步筛查。DHL细胞通常表现出高级别淋巴瘤的形态特征，例如大的、异型的淋巴细胞，核膜不规则，染色质粗糙，核仁明显，以及核分裂活跃。例如，高等级B细胞淋巴瘤（HGBL-NOS）具有高等级形态特征 38，提示形态学在初筛中具有潜在价值。因此，当观察到这些侵袭性形态时，应高度怀疑DHL的可能性，并将其列为后续更高级检测的优先对象。

免疫组化（IHC）在筛查中的应用

免疫组化（IHC）技术相对FISH而言，成本较低、操作简便，且在大多数病理实验室中均可开展。因此，IHC被广泛用作DHL的有效筛查工具，特别是用于识别双表达淋巴瘤（DEL）患者，从而进一步筛选需要进行FISH检测的病例。

  1. MYC和BCL2共表达（DEL）筛查：IHC检测MYC和BCL2蛋白的表达是识别高危DLBCL患者的重要手段。参考文献39通过反向蛋白质阵列（RPPA）评估了包括C-MYC和BCL2在内的多种生物标志物，发现C-MYC的高特异性有助于诊断。参考文献38提到在HGBL-NOS患者中，37%的患者具有MYC和BCL2双表达的免疫表型。尽管DEL不直接等同于DHL，但DEL本身代表着一个预后不良的群体。因此，在资源有限的情况下，可将所有DEL患者视为DHL的“高危疑似病例”，并优先安排FISH检测。这种策略能够显著提高FISH检测的效率，避免对所有DLBCL患者进行普遍筛查所带来的资源浪费。
  2. GCB表型筛查：大多数DHL起源于生发中心B细胞（GCB）亚型，其IHC特征通常为CD10阳性、BCL6阳性且MUM1阴性 40。一项研究表明，GCB表型与DHL基因型呈显著相关性，在筛选DHL/THL基因型时，GCB亚型被认为是重要的危险因素。因此，在DEL的基础上，结合GCB表型的判断，可以进一步缩小需要进行FISH检测的范围。例如，可以优先对GCB型DEL患者进行FISH检测，因为这部分患者发生DHL的可能性更高。

优化FISH检测的必要性评估

综合形态学、免疫组化（MYC/BCL2共表达和GCB表型），可以建立一个多指标的筛查模型，用于评估FISH检测的必要性：

  • 高风险组：如果患者同时符合以下条件：1）具有高级别形态特征；2）免疫组化显示MYC和BCL2高表达（DEL）；3）免疫表型为GCB型，则应立即进行MYC、BCL2和BCL6基因重排的FISH检测，以明确是否为DHL。这一组患者的FISH阳性率最高，临床上迫切需要确诊以指导治疗。
  • 中风险组：对于形态学不典型但IHC符合DEL标准，或GCB表型与DEL同时存在的患者，也应考虑进行FISH检测。虽然其DHL的阳性率可能低于高风险组，但仍然具有较高的临床意义。
  • 低风险组：对于IHC不符合DEL标准或没有高级别形态学特征的DLBCL患者，如果无其他高危临床指标，FISH检测的优先级可以降低。

这种分层筛查策略能够有效指导有限的诊断资源分配，确保最需要的患者能够及时获得准确的分子诊断，从而避免诊断延误和不必要的治疗。通过这种方法，即使在资源受限的环境下，也能尽可能地提高DHL的诊断效率和准确性，为患者提供更好的治疗机会。

3.3 分子分型与新型标志物

除了传统的形态学、免疫组化和FISH检测，分子分型和新型标志物的出现为双打击淋巴瘤（DHL）的诊断和风险分层提供了更深层次的理解。这些新型标志物旨在克服传统方法的局限性，特别是在识别FISH检测可能遗漏的具有高风险生物学特征的患者方面。

DHITsig（暗区基因表达谱）等新型分子标志物

基因表达谱（Gene Expression Profiling, GEP）技术已在识别弥漫大B细胞淋巴瘤（DLBCL）的分子亚型方面展现出巨大潜力，其中DHITsig（Double-Hit Signature，现已更名为“暗区基因表达谱”，DZsig）是一个重要的进展 41。DHITsig最初被描述为具有MYC和BCL2重排的高级别B细胞淋巴瘤（HGBCL-DH-BCL2）的特征性基因表达模式 42。然而，后续研究发现DZsig的表达范围超出了HGBCL-DH-BCL2，它能识别出预后不良的DLBCL亚组，包括那些通过常规FISH检测无法识别的患者 41。DZsig阳性DLBCL（DZsigpos-DLBCL）患者的两年总生存率显著低于非DZsig阳性的生发中心B细胞样（GCB）DLBCL患者 41。

DZsig的临床意义在于，它能够识别出即使没有明确的MYC和BCL2重排，但具有类似DHL生物学行为和不良预后的DLBCL患者 4142。这意味着，DZsig捕获的是一种特定的、侵袭性的分子表型，而不仅仅是基因重排的存在。例如，一项研究对20例DHITsig+但FISH检测MYC和/或BCL2无重排的GCB-DLBCL进行了全基因组测序，发现其中6例存在FISH检测“隐匿”的MYC或BCL2重排。此外，还发现其他潜在的MYC失调机制，如MYC或MIR17HG的拷贝数增加/扩增，以及PVT1启动子区域的局灶性缺失，这些都可能导致MYC过表达 42。这些发现强调了DZsig作为一种功能性标志物，能够更全面地反映MYC和BCL2通路的异常激活状态。

此外，一些新型的免疫组织化学（IHC）算法也被开发出来，旨在通过蛋白表达层面来模拟基因表达谱的分类。例如，一项研究提出了一个5标记的IHC算法（Bcl6、LMO2、c-MYC、GCET1和FOXP1）来区分伯基特淋巴瘤（BL）和DLBCL，这与DZsig有显著相关性。该IHC暗区算法能够支持BL的诊断，并识别出具有分子DZsig的MYC-BCL2 DHLs/THLs 43。这为资源有限的地区提供了一种成本效益高的方法来初步识别具有DZsig特征的患者。

传统FISH与基因表达谱在风险分层中的互补作用

传统FISH检测在DHL诊断中具有不可替代的核心地位，因为它直接证实了MYC和BCL2/BCL6基因重排的存在，这是DHL的定义性特征 37。然而，基因表达谱（特别是DZsig）与FISH在风险分层中扮演着互补的角色，而非互相取代。

  1. FISH的优势与局限：FISH的优势在于其高特异性和直接检测基因重排的能力。它是确诊DHL的金标准。然而，FISH可能存在“假阴性”，即某些复杂的或非典型的基因重排可能逃避常规探针的检测 42。此外，FISH仅关注特定的基因重排，无法全面反映MYC和BCL2通路异常激活的全部机制，例如拷贝数变异或转录后调控导致的过表达。
  2. 基因表达谱的优势与局限：GEP（如DZsig）的优势在于它能够整合多个基因的表达信息，提供肿瘤生物学行为的整体视图。它能够识别出FISH阴性但具有高风险基因表达模式的DLBCL患者，从而扩大了高风险人群的定义范围，使得更多需要强化治疗的患者能够被识别 4142。然而，GEP的缺点是成本较高，技术要求复杂，且通常需要新鲜冷冻组织，这在临床实践中存在一定挑战。
  3. 互补作用：理想的风险分层策略应结合FISH和GEP。对于FISH确诊的DHL患者，无疑属于高风险组。而对于FISH阴性的DLBCL患者，如果GEP显示为DZsig阳性，则应将其重新归类为高风险亚组，并考虑采取更积极的治疗策略 41。反之，一项研究发现，一小部分HGBCL-DH-BCL2缺乏DZsig表达，这些患者的分子特征与DZsig表达的HGBCL-DH-BCL2不同，并且与DLBCL-NOS具有相似的良好预后 41。这表明DZsig甚至可以在HGBCL-DH-BCL2内部进一步细化风险。因此，GEP可以作为FISH的补充，提高风险分层的精准度，避免过度治疗低风险患者或对高风险患者治疗不足。

总而言之，分子分型和新型标志物，尤其是DZsig，为DHL及相关侵袭性B细胞淋巴瘤的诊断和风险分层带来了革命性的改变。它们提供了比传统FISH更全面的生物学信息，并能识别出传统方法可能遗漏的预后不良患者。未来，结合FISH、GEP以及其他多组学数据，将有助于建立更精准的个体化治疗方案。

4. 预后评估与风险分层

双打击淋巴瘤（DHL）因其侵袭性的生物学特性，其预后评估和风险分层对于指导治疗方案的选择和改善患者生存至关重要。与非特指型弥漫大B细胞淋巴瘤（DLBCL-NOS）相比，DHL患者通常具有更差的预后，因此需要更精细的风险评估工具。

4.1 临床预后指标

在DHL的预后评估中，一系列传统的临床指标，如国际预后指数（IPI）、中枢神经系统国际预后指数（CNS-IPI）、乳酸脱氢酶（LDH）水平以及结外侵犯等，都被广泛研究并证实具有预测价值。

国际预后指数（IPI）与CNS-IPI

国际预后指数（IPI）是一个成熟且广泛使用的预后评分系统，综合了年龄、Ann Arbor分期、体能状态（ECOG PS）、乳酸脱氢酶（LDH）水平以及结外侵犯部位数量等因素。IPI评分越高，预后越差 44。对于DHL患者，由于其疾病本身的侵袭性，在诊断时往往具有较高的IPI评分，这进一步加剧了其不良预后 3745。例如，有研究显示，DHL患者在诊断时普遍存在高风险IPI评分，提示疾病已处于晚期且预后不佳 46。

鉴于DHL患者中枢神经系统（CNS）受累的风险较高，CNS-IPI被引入以更精准地评估CNS复发风险。CNS-IPI结合了IPI的多个因素，并特别关注了骨髓侵犯、多发结外病变（尤其是高危部位如睾丸、肾上腺、乳腺、中枢神经系统等）等与CNS复发密切相关的指标 4748。DHL患者的CNS侵犯率较高，即便在接受CNS靶向治疗后，预后仍然极差 4546。因此，CNS-IPI对于识别需要CNS预防性治疗的高危DHL患者具有重要意义。

LDH水平

乳酸脱氢酶（LDH）是细胞代谢的产物，其升高通常反映了肿瘤细胞的高增殖活性和/或肿瘤负荷的增加。在DHL患者中，LDH水平升高是常见且具有显著预后价值的指标。高LDH水平常与晚期疾病、弥漫性骨髓侵犯、广泛结外侵犯以及不良总生存期（OS）和无进展生存期（PFS）相关联 374549。有研究指出，LDH水平升高是DLBCL患者发生CNS复发的独立预测因子 50。

结外侵犯

结外侵犯，尤其是多部位或特定高危部位的侵犯，是DHL预后不良的重要临床特征。DHL患者常表现出较高的结外侵犯率，包括骨髓、中枢神经系统（CNS）、胃肠道、睾丸、肾脏、肾上腺和乳腺等 454647。例如，一项针对DLBCL患者的分析发现，CNS侵犯是所有结外部位中预后最差的因素 44。骨髓侵犯同样与较差的预后相关 50。对于原发性乳腺DHL（PB-DHL）患者，肿瘤大小、风险分层和是否接受自体干细胞移植（ASCT）等因素均与预后显著相关 51。

高TMTV（总代谢肿瘤体积）对治疗抵抗的预警作用

正电子发射计算机断层显像（PET/CT）是评估淋巴瘤疾病负荷和治疗反应的重要影像学工具。总代谢肿瘤体积（Total Metabolic Tumor Volume, TMTV）是通过PET/CT量化肿瘤细胞代谢活性的指标，反映了肿瘤的整体代谢负荷。高TMTV被认为是DLBCL，包括DHL患者对免疫化疗抵抗的重要预警标志。GOYA研究对DLBCL患者的分析显示，非常高的TMTV与原发性化疗难治性疾病和12个月内疾病进展（POD12）密切相关，是独立于临床风险因素的预测指标 52。即使在考虑了BCL2/MYC基因重排状态后，TMTV仍然是原发性化疗难治性的显著预测因子 52。这表明，高TMTV不仅反映了肿瘤的侵袭性，也预示着对一线免疫化疗的反应不佳，可能需要更积极或不同的治疗策略。因此，基线TMTV的评估有助于在治疗前识别高风险患者，指导个性化治疗方案的制定。

4.2 分子预后标志物

在双打击淋巴瘤（DHL）的预后评估中，除了临床指标，分子和细胞层面的标志物也扮演着越来越重要的角色。这些分子预后标志物能够更深入地揭示疾病的生物学特性，为精准风险分层和个体化治疗提供依据。

MYC重排类型（IgH vs 非IgH）

MYC基因重排是DHL的定义性特征之一，其重排类型对预后具有显著影响。MYC基因可以与免疫球蛋白（Ig）基因座（如IgH、Igκ、Igλ）发生易位，也可以与非Ig基因座发生重排。MYC的易位（无论是与IgH还是非Ig基因座）通常预示着不良预后 53。一项研究发现，DHL患者的中位总生存期（4.5个月）显著低于伯基特淋巴瘤和国际预后指数匹配的弥漫大B细胞淋巴瘤患者 54。此外，MYC重排的复杂性还体现在，除了易位，MYC基因的拷贝数增加/扩增也被发现与患者较差的无进展生存期和总生存期显著相关 55。因此，详细分析MYC重排的类型和伴随的拷贝数变异，对于评估DHL患者的预后具有重要价值。

BCL2突变状态

BCL2基因的异常是DHL的另一关键特征，通常表现为基因重排导致的过表达。除了重排，BCL2基因的突变状态也被发现与DLBCL患者的预后相关。例如，一项研究发现，BCL2的拷贝数增加/扩增与患者较差的无进展生存期（PFS）和总生存期（OS）显著相关 55。特别是在伴有MYC基因改变的DHL中，BCL2的突变或过表达可能通过抑制细胞凋亡，进一步增强MYC驱动的细胞增殖能力，从而加剧疾病的侵袭性。因此，评估BCL2的突变状态，结合其重排情况，可以提供更全面的预后信息。

A-ALC（自体移植淋巴细胞计数）

自体移植淋巴细胞计数（Autograft Absolute Lymphocyte Count, A-ALC）是指患者在接受自体外周血造血干细胞移植（APBHSCT）后早期（通常是移植后特定时间点）的绝对淋巴细胞计数。A-ALC被认为是淋巴瘤患者接受APBHSCT后生存的预后因素。一项针对DHL/THL患者的研究显示，移植后A-ALC ≥0.5 × 10^9 细胞/公斤的患者具有显著优于A-ALC <0.5 × 10^9 细胞/公斤患者的总生存期（OS）和无进展生存期（PFS） 56。多变量分析进一步证实，A-ALC是DHL/THL患者OS和PFS的独立预测因子 56。这一发现提示，自体移植后的免疫重建状态，特别是淋巴细胞的数量，对于改善DHL/THL患者的长期生存至关重要。因此，在评估接受APBHSCT的DHL患者预后时，应将A-ALC纳入考量，并可能通过优化干细胞采集和移植后管理策略来提高A-ALC水平，从而改善患者结局。

此外，其他一些分子标志物，如XPO1的表达水平，也被发现与DHL的预后相关。高XPO1表达与DLBCL患者的不良预后显著相关，尤其是在BCL2过表达的患者中，并且在伴有MYC/BCL2双打击的高级别B细胞淋巴瘤（HGBCL-DH）细胞系中，XPO1抑制剂selinexor显示出较强的细胞毒性 57。这提示，XPO1可能作为DHL预后的一个生物标志物和潜在的治疗靶点。

综上所述，MYC重排类型、BCL2突变状态以及自体移植后的A-ALC等分子/细胞指标，为DHL的预后评估提供了多维度的视角。将这些分子预后标志物与临床预后指标相结合，能够构建更完善的风险分层模型，从而为DHL患者制定更精准的治疗方案。

4.3 治疗反应与早期复发预测

对于双打击淋巴瘤（DHL）患者，早期准确评估治疗反应和预测复发至关重要，这有助于及时调整治疗策略，改善患者预后。正电子发射计算机断层显像（PET/CT）和微小残留病（MRD）检测是当前评估治疗反应和预测早期复发的重要工具。

PET/CT代谢缓解深度

PET/CT在淋巴瘤的治疗反应评估中发挥着核心作用，它能够通过量化肿瘤的代谢活性来判断治疗效果。Lugano分类标准推荐使用PET/CT在治疗中和治疗结束后进行评估，并根据Deauville评分系统来评估代谢缓解的深度。GOYA研究是一项针对弥漫大B细胞淋巴瘤（DLBCL）患者的III期临床试验，虽然其主要目标是比较利妥昔单抗与obinutuzumab在R-CHOP方案中的疗效，但其数据为PET/CT在治疗反应评估中的作用提供了重要证据 52。

在该研究中，尽管未直接聚焦DHL，但其对DLBCL的分析结果具有借鉴意义。对于侵袭性强的淋巴瘤患者，即使在治疗结束时达到形态学上的完全缓解，代谢活性残留（即非完全代谢缓解）也可能预示着更高的复发风险。因此，PET/CT评估的代谢缓解深度，可能提示治疗抵抗或潜在的早期复发风险，需要更积极的随访或考虑巩固治疗。

此外，GOYA研究还强调了基线总代谢肿瘤体积（TMTV）与原发性化疗难治性疾病和12个月内疾病进展（POD12）的强关联性，即使在考虑了BCL2/MYC基因重排状态后，TMTV仍然是显著的预测因子 52。这意味着，基线时肿瘤的代谢负荷越大，患者对治疗的反应越差，早期复发的可能性也越高。因此，结合基线TMTV和治疗后的代谢缓解深度，可以更全面地评估DHL患者的治疗反应和复发风险。

微小残留病（MRD）检测

微小残留病（MRD）是指经过治疗后，体内仍存在少量肿瘤细胞，这些细胞无法通过常规影像学或形态学方法检测到，但可以通过高灵敏度的分子生物学技术检出。MRD检测在预测淋巴瘤患者早期复发方面展现出巨大潜力，尤其对于DHL这类预后不良的侵袭性淋巴瘤。

目前，MRD检测主要通过检测循环肿瘤DNA（ctDNA）实现，ctDNA是从肿瘤细胞释放到血液中的DNA片段，携带有肿瘤特异性突变或基因重排信息。高通量测序（NGS）是检测ctDNA中MRD的主要技术。

  • ctDNA在预测早期复发中的应用：多项研究表明，ctDNA水平与DLBCL患者的预后和复发密切相关。治疗前ctDNA水平升高被认为是PFS和OS的独立预后因素 58。在治疗过程中，ctDNA水平的动态变化可用于评估治疗反应。例如，在初始两个周期治疗后ctDNA水平显著下降的患者（分子应答者）具有更好的PFS和OS 58。重要的是，治疗结束后的ctDNA检测被证明是检测高风险复发患者微小残留病的敏感指标，即使这些患者在影像学上已达到完全缓解（CR） 58。一项研究发现，ctDNA阳性的患者复发风险是ctDNA阴性患者的6.47倍 58。
  • IgH/IgK基因重排的监测：在DLBCL患者中，监测免疫球蛋白重链（IGH）和免疫球蛋白κ轻链（IGK）基因的可变区、多样区和连接区（VDJ）重排在ctDNA中的动态变化，是预测疾病进展的有效方法 59。NGS技术能够识别组织样本中的主要V(D)J克隆重排，并在外周血ctDNA中追踪这些克隆，从而实现MRD的监测。研究显示，加入IGK克隆的监测可以提高MRD的检测率，从42.9%增加到58.0% 59。此外，治疗中期和治疗结束时的NGS MRD监测与影像学结果相比，在预测PFS方面表现出更好的性能。值得注意的是，NGS MRD监测能够在影像学发现复发之前，提前数月预测疾病复发 59。IGH和IGK克隆清除率越快，预后越好 59。

综上所述，PET/CT的代谢缓解深度评估和ctDNA介导的MRD检测，为DHL患者的治疗反应评估和早期复发预测提供了强大的工具。将这些先进的检测技术整合到临床实践中，有望实现更精准的风险分层，并指导个体化的巩固或挽救治疗策略，从而改善DHL患者的临床结局。

5. 治疗策略与临床实践

5.1 一线治疗：强化化疗与方案优化

双打击淋巴瘤（DHL）因其侵袭性强、预后差的特点，使得传统标准治疗方案R-CHOP（利妥昔单抗、环磷酰胺、多柔比星、长春新碱、泼尼松）的疗效欠佳。因此，一线治疗策略的重点在于强化化疗方案，以期克服其固有的耐药性，提高缓解率和长期生存。目前，临床实践中主要对比R-CHOP与DA-EPOCH-R（剂量调整的依托泊苷、泼尼松、长春新碱、环磷酰胺、多柔比星联合利妥昔单抗）等强化方案的疗效差异，并探索基于患者IPI评分和分子风险的分层治疗。

R-CHOP与DA-EPOCH-R等强化方案的疗效差异

针对DHL患者，R-CHOP方案已被证实疗效不佳。多项回顾性研究和临床观察表明，DHL患者接受R-CHOP治疗后，通常疾病进展快，中位无进展生存期（PFS）和总生存期（OS）显著短于非DHL患者 6061。R-CHOP的失败率高，复发后OS中位仅8.6个月 61。因此，临床上开始探索更强化的治疗方案，其中DA-EPOCH-R是最受关注的选择之一。

DA-EPOCH-R方案通过持续输注化疗药物，并根据患者的血细胞计数进行剂量调整，旨在实现最大耐受剂量的化疗，以期克服DHL的耐药性 6062。多项研究，包括回顾性分析和前瞻性临床试验，已评估了DA-EPOCH-R在DHL患者中的疗效：

  • DA-EPOCH-R的潜在优势：一些研究认为，DA-EPOCH-R方案能够为DHL患者带来更好的预后 60。一项研究比较了CD5阳性DHL患者中DA-EPOCH-R与R-CHOP的疗效，结果显示DA-EPOCH-R组的2年PFS和OS均显著优于R-CHOP组（PFS：63.6% vs 45.4%；OS：67.4% vs 47.8%）。尤其是在CD5阳性DHL患者中，DA-EPOCH-R相比R-CHOP，PFS有明显改善（85.7% vs 23.0%），尽管OS未达到统计学显著差异 63。另一项针对MYC和BCL2双打击淋巴瘤的研究也指出，DA-EPOCH-R能够改善患者结局 64。
  • DA-EPOCH-R的挑战与局限：尽管DA-EPOCH-R被认为是DHL的“标准”强化方案 62，但其疗效并非总是明确优于R-CHOP。一些回顾性分析显示，DA-EPOCH-R在DHL/THL患者中的生存结局与历史估计值相当 62。一项多中心分析发现，DA-EPOCH-R的剂量调整实践对DHL/THL患者的生存结局没有显著影响，累积化疗剂量、剂量升级或剂量调整依从性均未与OS或PFS显著相关 62。这表明DA-EPOCH-R的实施复杂性较高，且在真实世界中的疗效可能受到多种因素影响，如剂量调整的依从性、毒性管理等。
  • 其他强化方案：除了DA-EPOCH-R，其他更密集的方案，如与伯基特淋巴瘤（Burkitt lymphoma）类似的方案（如CODOX-M/IVAC），也被用于DHL的治疗。这些方案通常毒性较大，可能不适用于所有患者。

基于IPI/分子风险的分层治疗

鉴于DHL的高度异质性和对传统化疗的抵抗性，基于IPI评分和分子风险（如MYC/BCL2重排类型、共表达状态等）进行分层治疗是当前研究和临床实践的重要方向。

  • 高危IPI患者：对于诊断时IPI评分较高的DHL患者，通常被认为是极高危人群，需要更积极的治疗策略。在这种情况下，DA-EPOCH-R或更强化的方案通常被推荐作为一线治疗选择，以期获得更好的初始缓解。然而，即使是强化方案，高IPI患者的预后仍然不佳，提示可能需要额外的巩固治疗或新型疗法的介入。

  • 分子风险分层：

    • MYC/BCL2重排状态：确诊为DHL的患者（MYC和BCL2/BCL6重排），无论IPI评分如何，都应被视为高危，并考虑强化治疗。
    • 双表达淋巴瘤（DEL）：对于IHC检测为MYC和BCL2双表达，但FISH检测无基因重排的DEL患者，虽然其预后差于非DEL的DLBCL，但可能优于DHL。因此，对于DEL患者是否需要与DHL相同的强化治疗，仍存在争议。一些研究建议，DEL患者在接受DA-EPOCH-R治疗后可能获得良好疗效，尤其是在DEL队列中，所有患者均实现持久完全代谢缓解 64。
    • 新型分子标志物：未来，分子高分级（MHG）等通过基因表达谱定义的分子亚型可以识别出即使无基因重排但具有DHL样生物学行为的DLBCL患者 65。这类患者可能也需要强化治疗，从而实现更精准的风险分层。

  • 利妥昔单抗维持治疗：对于高危患者，尤其是在初始治疗后达到完全缓解但仍存在高复发风险的患者，利妥昔单抗维持治疗可能是一个考虑选项。利妥昔单抗作为抗CD20单克隆抗体，在DLBCL中已被证实能够提高PFS和OS。然而，对于DHL患者，其维持治疗的有效性仍需进一步的大规模临床研究证实。

总之，DHL的一线治疗策略正从“一刀切”向“精准分层”转变。DA-EPOCH-R作为强化化疗方案，在部分DHL患者中显示出优势，但仍需更深入的研究来明确其最佳适用人群和疗效的普遍性。未来，结合IPI、分子遗传学特征和新型分子标志物，将有助于为DHL患者制定更个体化的治疗方案，以期最大程度地改善临床结局。

5.2 复发/难治性DHL的免疫治疗

对于复发/难治性（R/R）双打击淋巴瘤（DHL）患者，传统挽救性化疗的疗效极其有限，预后极差。近年来，以CAR-T细胞疗法和双特异性抗体为代表的免疫治疗为这部分患者带来了新的希望。这些新型疗法通过利用或激活患者自身的免疫系统来识别和清除肿瘤细胞，展现出显著的临床疗效，但同时也伴随着特有的毒副作用。

CAR-T细胞疗法：突破性的治疗进展

CAR-T细胞疗法（嵌合抗原受体T细胞疗法）通过基因工程改造患者自身的T细胞，使其表达能够特异性识别肿瘤细胞表面抗原（如CD19）的嵌合抗原受体。CD19靶向的CAR-T疗法已在R/R DLBCL（包括DHL在内）中取得了革命性的进展，并获批用于临床。

  • 疗效分析：
    • 高缓解率：CAR-T疗法，特别是阿西卡他基因西洛来赛（axi-cel）、替沙瑞妥基因来赛（tisa-cel）和利索卡他基因马仑西（liso-cel）等CD19靶向产品，在重度预处理的R/R DLBCL患者中，包括DHL亚群，均显示出持久且前所未有的完全缓解（CR）率，并具有治愈潜力 66。一项多中心回顾性研究显示，CAR-T疗法可以克服DHL和双表达淋巴瘤（DEL）的不良预后影响 67。研究中，DHL患者（n=80）和非DHL患者（n=328）接受CAR-T治疗后，两组的无进展生存期（PFS）差异无统计学意义（HR 0.8, 95%CI 0.5-1.3, p=0.35） 67。这意味着CAR-T疗法在DHL患者中能够取得与非DHL患者相似的疗效，显著改善了这部分患者的预后。另一项真实世界数据显示，复发性DHL/THL患者接受ASCT后真实世界总生存期（rwOS）不如非DHL/THL患者，但CAR-T治疗后这种差异并未显现 68。
    • 不同产品间的差异：在标准治疗的挽救方案和ASCT的比较中，axi-cel和liso-cel相比tisa-cel在长期控制方面表现出优越性 66。对于不适合ASCT的R/R DLBCL患者，liso-cel显示出有利的风险-收益比 66。
  • 毒副作用（CRS/神经毒性）：CAR-T疗法主要的毒副作用包括细胞因子释放综合征（CRS）和免疫效应细胞相关神经毒性综合征（ICANS）。CRS表现为发热、低血压、缺氧等全身炎症反应；ICANS则可引起谵妄、癫痫、失语等神经系统症状。这些不良事件虽然可能严重，但通常是可控的 66。早期研究中，axi-cel的毒性相对较高，但在重度预处理患者中其疗效也可能更优 66。
  • 局限性：尽管CAR-T疗效显著，但仍有25%至50%的R/R DLBCL患者未能达到缓解 69。此外，CAR-T疗法的实施成本高昂，且存在并发症和报销问题，是其广泛应用的主要障碍 66。部分患者在CAR-T治疗后可能出现疾病复发，且复发后的DHL患者预后极差 67。复发机制包括CD19抗原丢失或下调 70、CAR-T细胞持久性差以及肿瘤微环境的免疫抑制等。

双特异性抗体：CD20×CD3靶向策略

双特异性抗体是一种新兴的免疫治疗手段，能够同时结合T细胞上的CD3受体和肿瘤细胞上的CD20抗原，从而桥接T细胞和肿瘤细胞，激活T细胞介导的肿瘤杀伤。

  • 莫苏奈妥珠单抗（Mosunetuzumab）：作为一种CD20×CD3双特异性抗体，莫苏奈妥珠单抗在R/R B细胞淋巴瘤（包括DLBCL）中展现出良好的安全性和抗肿瘤活性 71。在一项针对R/R滤泡性淋巴瘤（FL）的II期研究中，固定疗程的莫苏奈妥珠单抗治疗使54%的患者（60.0% [95% CI 49.1-70.2]）获得完全缓解 72。虽然该研究主要针对FL，但其在DLBCL等侵袭性B细胞淋巴瘤中也显示出活性。
  • 疗效和安全性：莫苏奈妥珠单抗的毒副作用可控，最常见的是CRS，但多为1-2级，且主要发生在第一个周期 7172。其他常见不良事件包括中性粒细胞减少、低磷血症等 72。其固定疗程的给药方式和相对较低的毒性，使其有望作为门诊治疗方案，方便患者。
  • 与其他双特异性抗体：除了莫苏奈妥珠单抗，其他CD20×CD3双特异性抗体如glofitamab、epcoritamab等也已在R/R DLBCL中显示出显著疗效。这些药物通常在CAR-T治疗失败或不适用CAR-T的患者中发挥作用，提供了一种非细胞疗法的替代方案。

CAR-T与双特异性抗体的比较与未来展望

CAR-T细胞疗法和双特异性抗体各有优势，且在R/R DHL的治疗中具有互补性。CAR-T疗法通常能提供更持久的缓解，但成本高昂、技术复杂且毒副作用相对较重。双特异性抗体则具有即用型、可控性好、毒性相对较低的特点，更易于推广。未来，两者可能在不同的治疗阶段或患者群体中发挥各自的优势。例如，放射治疗可作为CAR-T治疗前的桥接策略 73。此外，新型CAR产品（如靶向CD22或CD30）以及与其他药物（如BCL2抑制剂）的联合治疗，有望进一步提高R/R DHL患者的治疗应答率和持久性。

5.3 靶向治疗与新型药物探索

双打击淋巴瘤（DHL）因其独特的分子病理特征和对传统免疫化疗的抵抗性，促使研究人员积极探索基于其分子机制的靶向治疗策略。这些新型药物旨在特异性作用于DHL细胞的关键信号通路或生物学特征，以期提高疗效并降低毒副作用。

BCL2抑制剂（维奈克拉）

BCL2基因重排导致的BCL2蛋白过表达是DHL的核心特征之一，它通过抑制细胞凋亡促进肿瘤细胞的存活。因此，BCL2抑制剂成为DHL靶向治疗的重要方向。维奈克拉（Venetoclax）是一种高效、选择性的BCL2抑制剂，通过模拟BH3结构域与BCL2蛋白结合，解除对促凋亡蛋白（如BAX、BAK）的抑制，从而恢复细胞凋亡。

  • 疗效：在复发/难治性DLBCL，特别是BCL2过表达的DLBCL患者中，维奈克拉单药或联合其他药物已显示出一定的疗效。尽管DHL患者通常对维奈克拉单药治疗的反应率较低，但将其与传统化疗（如R-EPOCH）或新型靶向药物（如BTK抑制剂、CD20抗体）联合使用时，有望提高疗效。一项研究表明，贝伐珠单抗（一种VEGF-A抑制剂）与BCL2抑制剂维奈克拉联合使用，对B细胞非霍奇金淋巴瘤（包括SU-DHL-6细胞系）具有协同抗肿瘤作用，可显著诱导细胞死亡并缩小肿瘤大小和重量 74。这提示，通过多靶点联合可能克服单药治疗的局限性。
  • 局限性：维奈克拉在DLBCL中的疗效受多种因素影响，包括BCL2蛋白表达水平、MYC重排状态以及共存的基因突变（如TP53突变）等。对于具有MYC和BCL2双重打击的DHL患者，由于其存在MYC驱动的快速增殖和BCL2介导的凋亡抑制，单用维奈克拉可能不足以克服其高度侵袭性。

BTK抑制剂（Pirtobrutinib）

布鲁顿酪氨酸激酶（BTK）是B细胞受体（BCR）信号通路中的关键激酶，在B细胞的增殖、分化和存活中发挥重要作用。BTK抑制剂已在多种B细胞淋巴瘤中取得成功，但对于DHL的疗效仍在探索中。Pirtobrutinib（formerly LOXO-305）是一种新型的、高选择性的、可逆的非共价BTK抑制剂。

  • 疗效：Pirtobrutinib主要用于治疗对共价BTK抑制剂产生耐药或不耐受的B细胞淋巴瘤患者。在BRUIN I/II期临床研究中，Pirtobrutinib在既往接受过共价BTK抑制剂治疗的慢性淋巴细胞白血病（CLL）/小淋巴细胞淋巴瘤（SLL）和套细胞淋巴瘤（MCL）患者中显示出良好的安全性和有效性，总缓解率（ORR）分别为62%和52% 75。虽然DHL并非其主要研究人群，但作为一种侵袭性B细胞淋巴瘤，DHL患者可能从BTK抑制剂中获益，尤其是在BCR信号通路被激活的亚群中。研究表明，Pirtobrutinib在对先前BTK抑制剂不耐受的B细胞恶性肿瘤患者中表现出良好的安全性、耐受性和有效性 76。此外，Pirtobrutinib在里氏转化（Richter transformation，常见为DLBCL形式）患者中也显示出令人鼓舞的安全性与活性，其中多数患者曾接受过里氏转化导向治疗，包括共价BTK抑制剂 77。这提示Pirtobrutinib可能为重度预处理的DHL患者提供新的治疗选择。
  • 优势：Pirtobrutinib的非共价结合机制使其能够克服由BTK C481S突变引起的共价BTK抑制剂耐药，且具有更低脱靶毒性，从而改善患者的耐受性 7578。

ADC药物（Loncastuximab Tesirine）

抗体-药物偶联物（Antibody-Drug Conjugate, ADC）是一种将靶向抗体与强效细胞毒药物偶联而成的新型抗肿瘤药物，它能够特异性地将化疗药物递送到肿瘤细胞，从而减少对正常组织的损伤。Loncastuximab Tesirine是一种靶向CD19的ADC，已获批用于治疗R/R DLBCL。

  • 疗效：在LOTIS-2关键II期临床试验中，Loncastuximab Tesirine在经重度预处理的R/R DLBCL患者中展现出显著的单药抗肿瘤活性和可接受的安全性，包括伴有高风险特征（如双打击、三打击、转化型或原发难治性DLBCL）的患者。总缓解率（ORR）达到48.3%，其中完全缓解（CR）率为24.1% 79。长期随访数据显示，该药物可带来持久的缓解，特别是在CR患者中，24个月的总生存率和无进展生存率分别达到68.2%和72.5% 80。
  • 意义：Loncastuximab Tesirine为重度预处理的R/R DLBCL（包括DHL）患者提供了新的治疗选择，尤其对于那些不适合或对CAR-T疗法无应答的患者。CD19在大多数B细胞淋巴瘤中高表达，使其成为ADC的理想靶点。

MYC靶向（如CAMKII抑制剂）

MYC癌基因的异常激活是DHL的核心驱动力，但直接靶向MYC蛋白本身一直是一个巨大的挑战，因为MYC缺乏易于结合的小分子口袋。因此，研究方向转向了靶向MYC的下游效应器或其上游调控通路。

  • CAMKII抑制剂：钙/钙调蛋白依赖性蛋白激酶II（CAMKII）是MYC信号通路中的一个关键介质。研究表明，CAMKII在淋巴瘤细胞中可能发挥重要作用，抑制CAMKII活性可以干扰MYC的下游效应，从而抑制淋巴瘤细胞的增殖。目前，CAMKII抑制剂在DHL中的临床研究仍处于早期阶段，但有望成为间接靶向MYC的潜在策略。
  • 其他MYC靶向策略：包括抑制MYC转录、翻译或蛋白稳定性的药物，以及靶向MYC结合伴侣（如MAX、Miz1）的策略都在积极探索中。例如，通过干扰MYC-MAX异二聚体的形成或活性来抑制MYC的功能。

表观遗传调控（BCL6激酶重定位）

BCL6是DHL的另一个关键驱动基因，其在生发中心B细胞增殖和分化中发挥重要作用。表观遗传修饰，如组蛋白乙酰化和甲基化，在BCL6的表达和功能调控中扮演关键角色。

  • BCL6激酶重定位：BCL6的活性受多种机制调控，包括磷酸化修饰。特定激酶的异常激活或定位可能导致BCL6功能失调，从而促进淋巴瘤发生。靶向这些激酶或其介导的BCL6异常调控，可能为DHL提供新的治疗机会。
  • 表观遗传药物：组蛋白去乙酰化酶抑制剂（HDACi）和EZH2抑制剂等表观遗传药物已在淋巴瘤治疗中显示出活性。这些药物通过改变染色质结构和基因表达，可能间接影响MYC和BCL6的表达和功能。例如，EZH2抑制剂在GCB型DLBCL中显示出疗效，而大部分DHL起源于GCB。

新型药物探索的未来方向

除了上述药物，DHL的靶向治疗还包括PI3K/AKT/mTOR信号通路抑制剂、CD47阻断剂以及细胞周期抑制剂等。未来的研究将更加注重多靶点联合治疗，以期在抑制肿瘤增殖的同时，克服耐药性并激活宿主抗肿瘤免疫反应。同时，基于患者特定的分子特征（如特定的基因突变、拷贝数变异、基因表达谱）进行精准分层，将有助于指导靶向药物的选择，实现真正意义上的个体化治疗。

5.4 造血干细胞移植的定位

造血干细胞移植（Hematopoietic Stem Cell Transplantation, HSCT）在双打击淋巴瘤（DHL）的治疗中扮演着重要角色，尤其是在高危患者的一线巩固治疗以及复发/难治性（R/R）患者的挽救治疗中。自体造血干细胞移植（auto-HSCT）和异基因造血干细胞移植（allo-HSCT）各有其适应症和局限性。

自体造血干细胞移植（auto-HSCT）

auto-HSCT长期以来是治疗复发/难治性弥漫大B细胞淋巴瘤（DLBCL）的标准方法，对于DHL患者，其作用定位则更为复杂。

  • 一线缓解后巩固：对于新诊断的DHL患者，由于其侵袭性强和预后不良，即使一线强化化疗（如DA-EPOCH-R）后达到完全缓解（CR），也可能面临高复发风险。在这种情况下，auto-HSCT作为巩固治疗的价值一直备受关注。美国移植和细胞治疗学会（ASTCT）的临床实践指南指出，对于接受R-CHOP或类似方案治疗的DHL/THL患者，在实现CR后可以考虑auto-HSCT巩固治疗 81。然而，对于接受强化诱导治疗的DHL/THL患者，ASTCT指南认为auto-HSCT巩固的必要性较小，即不推荐进行巩固性auto-HSCT 81。这表明，auto-HSCT作为一线巩固策略，可能更适用于那些未接受最强化诱导治疗的DHL患者，或是在接受强化诱导后仍存在高危因素的患者。
  • 复发/难治性DHL的挽救治疗：对于R/R DHL患者，如果对挽救性化疗敏感并达到完全缓解（CR）或部分缓解（PR），auto-HSCT通常被视为一种有效的挽救治疗策略。例如，利妥昔单抗联合ICE方案（RICE）作为二线治疗在自体干细胞移植前，能够显著提高复发/难治性DLBCL患者的完全缓解率，从而改善移植后的无进展生存率 82。然而，随着CAR-T细胞疗法等新型治疗手段的出现，auto-HSCT在R/R DLBCL（包括DHL）二线治疗中的地位正在发生变化。ASTCT指南推荐，对于原发难治或早期复发（≤12个月）的DLBCL患者，首选CAR-T疗法；而对于晚期复发（>12个月）且对挽救性化疗敏感的患者，auto-HSCT仍是推荐的巩固治疗方案 81。这提示，在DHL患者中，auto-HSCT可能更适用于晚期复发且对挽救治疗敏感的患者。

A-ALC对移植后生存的影响

自体移植绝对淋巴细胞计数（Autograft Absolute Lymphocyte Count, A-ALC）是指在自体造血干细胞移植后早期（如移植后第15天或30天）外周血中测得的绝对淋巴细胞计数。A-ALC被认为是预测淋巴瘤患者auto-HSCT后生存预后的重要因素。一项针对77例DHL/THL患者的研究显示，A-ALC ≥0.5 × 10^9 细胞/公斤的患者在移植后具有显著更长的总生存期（OS）和无进展生存期（PFS） 56。多变量分析进一步证实，A-ALC是DHL/THL患者OS和PFS的独立预测因子 56。这一发现强调了移植后早期免疫重建状态的重要性，提示在进行auto-HSCT时，不仅要收集足够的CD34+细胞，还要关注淋巴细胞的采集量，以优化移植后的免疫恢复，从而改善DHL/THL患者的长期预后 56。

异基因造血干细胞移植（allo-HSCT）

allo-HSCT通常在auto-HSCT失败或患者不适合auto-HSCT的情况下考虑，特别是对于高危R/R DHL患者。allo-HSCT的优势在于其潜在的移植物抗肿瘤效应（Graft-versus-Lymphoma, GvL），能够清除体内残留的肿瘤细胞。然而，allo-HSCT的风险也更高，包括移植物抗宿主病（GvHD）、感染和非复发死亡率。目前，allo-HSCT在DHL中的确切地位仍不明确，通常作为最后一道防线或在特定高危亚组中考虑。

移植时机的优化

移植时机的选择对于DHL患者至关重要。

  • 早期移植：对于高危新诊断DHL患者，在一线化疗达到缓解后尽早进行巩固性auto-HSCT可能有助于延长无病生存期。然而，由于DHL对化疗的固有耐药性，即使早期巩固，部分患者仍可能复发。
  • 挽救性移植：对于R/R DHL患者，在接受挽救性化疗后达到敏感缓解再进行auto-HSCT是关键。对于那些对挽救性化疗不敏感或早期复发的患者，CAR-T疗法已成为优先选择 81。

总而言之，造血干细胞移植在DHL的治疗中仍具有重要价值。auto-HSCT可作为一线强化治疗后高危患者的巩固手段，以及晚期复发且对挽救性化疗敏感患者的挽救方案。A-ALC作为预测auto-HSCT后生存的独立预后因子，为优化移植策略提供了新的视角。随着新型免疫疗法的发展，HSCT在DHL治疗中的具体定位和与其他疗法的序贯使用，将是未来研究的重要方向。

6.1 分类与诊断的争议

双打击淋巴瘤（DHL）的分类与诊断在过去十年中经历了显著演变，并随着WHO（世界卫生组织）和ICC（国际共识分类）最新版本的发布，一些争议点和分歧仍然存在，对临床实践产生重要影响。核心争议之一在于MYC/BCL6重排是否应与经典的MYC/BCL2重排一同纳入“双打击淋巴瘤”的统一分类，以及WHO-5和ICC在高级别B细胞淋巴瘤（HGBL）定义上的差异。

MYC/BCL6重排的分类争议

传统的DHL定义主要指MYC和BCL2基因同时发生重排的侵袭性B细胞淋巴瘤。然而，MYC和BCL6同时重排的淋巴瘤（通常被称为“双打击淋巴瘤”）其生物学行为和预后与MYC/BCL2重排型DHL存在异质性，引发了是否应将其统一归类的讨论。

  • 生物学异质性：有研究指出，MYC/BCL6重排的淋巴瘤在临床和病理学上表现出一定的异质性，可能与MYC/BCL2重排的DHL不完全相同。例如，一些MYC/BCL6重排的病例可能表现出更接近弥漫大B细胞淋巴瘤（DLBCL）的特征，而不是典型的侵袭性DHL表型。这种异质性使得对其进行统一分类变得复杂，并可能影响治疗决策。
  • WHO-5与ICC的分歧：
    • WHO-5的立场：在WHO造血和淋巴组织肿瘤分类的第5版（WHO-HAEM5）中，明确将DHL定义为“伴有MYC和BCL2重排的DLBCL/HGBL，伴或不伴BCL6重排” 8384。这意味着WHO-5主要强调MYC和BCL2重排的核心地位，而将MYC和BCL6重排的病例划归到DLBCL，NOS（非特指型弥漫大B细胞淋巴瘤）或HGBL，NOS（非特指型高级别B细胞淋巴瘤）中，具体取决于其形态学特征 838485。WHO-5的这种分类逻辑是基于MYC/BCL2重排所表现出的更统一且更差的预后 2。
    • ICC的立场：国际共识分类（ICC）则采取了略有不同的方法。除了认可WHO-5定义的MYC/BCL2重排型DHL，ICC还将“伴有MYC和BCL6重排的HGBL”作为一个“临时实体”予以承认 8385。ICC认为，虽然这类淋巴瘤在生物学上可能存在异质性，但其MYC和BCL6重排的特征仍值得独立评估，这反映了对其潜在临床意义的关注。

这种分类上的分歧在临床实践中造成了一定的困扰。例如，一个具有MYC和BCL6重排的淋巴瘤，根据WHO-5可能被诊断为DLBCL，NOS或HGBL，NOS，而根据ICC则可能被视为一个独立的临时实体。不同的诊断可能导致对患者风险评估和治疗方案选择上的差异，尤其是在临床试验的入组标准和研究结果的比较方面。

HGBL定义对临床实践的影响

高级别B细胞淋巴瘤（HGBL）的概念最初在2016年WHO分类中引入，旨在涵盖那些形态学上介于DLBCL和伯基特淋巴瘤（BL）之间，或具有MYC和BCL2/BCL6重排的侵袭性B细胞淋巴瘤 86。WHO-5和ICC对HGBL的定义进一步细化，但仍存在差异，这些差异对临床实践的影响主要体现在以下几个方面：

  1. 诊断标准的不一致性：由于WHO-5和ICC对DHL和HGBL的定义存在差异，特别是MYC/BCL6重排的归类问题，导致病理医生在出具诊断报告时可能需要注明所遵循的分类体系，以避免混淆。这增加了诊断的复杂性，并可能影响医生对诊断报告的解读。
  2. 治疗选择的困境：DHL和HGBL通常需要比标准R-CHOP方案更强化的治疗，如DA-EPOCH-R方案。然而，对于MYC/BCL6重排的淋巴瘤，如果根据WHO-5被归类为DLBCL，NOS，临床医生可能会倾向于采用DLBCL的标准治疗方案，而不是更激进的DHL强化方案。这种不确定性可能导致部分患者未获得最佳的初始治疗。例如，一项研究指出，强化化疗（如DA-EPOCH-R）在DHL中的疗效优于R-CHOP，如果MYC/BCL6重排的病例未被识别为需要强化治疗，可能会影响患者预后。
  3. 临床试验设计与结果解释：分类的差异也影响了临床试验的设计。不同的试验可能采用不同的诊断标准入组患者，这使得比较不同试验的结果变得困难。此外，由于诊断标准的不一致，某些亚群患者可能无法被准确纳入到针对DHL或HGBL的特定临床试验中，从而影响新疗法的开发和评估。
  4. 预后评估的挑战：准确的分类对于预后评估至关重要。例如，WHO-5将MYC/BCL2重排作为DHL的核心特征，是因为其预后明显劣于其他DLBCL亚型。如果将预后不同的MYC/BCL6重排病例与MYC/BCL2重排病例统一归类，可能会稀释特定基因重排组合的预后信息，使得预后模型不够精确。反之，如果将MYC/BCL6重排病例降级分类，可能导致部分患者未能充分认识到其疾病的侵袭性。

总体而言，WHO-5和ICC在分类上的分歧，特别是对于MYC/BCL6重排的定位，反映了对这些侵袭性淋巴瘤生物学复杂性的持续探索。未来的研究需要进一步阐明MYC/BCL6重排的独特生物学特征和临床意义，以期达成更统一的诊断标准和更精准的治疗策略，最终改善患者的预后。

6.2 精准治疗的个体化探索

双打击淋巴瘤（DHL）的高度异质性和传统治疗效果的局限性，使得精准治疗成为改善患者预后的关键方向。随着分子诊断技术的发展，基于分子亚群特征（如基因重排、突变谱、基因表达谱）制定靶向联合方案，并利用多组学数据指导临床试验，正成为DHL个体化治疗探索的前沿。

基于分子亚群的靶向联合方案

DHL患者的分子特征远不止MYC和BCL2/BCL6的重排。深入分析这些患者的分子景观，可以识别出更多可靶向的驱动基因和信号通路，从而设计更具针对性的联合治疗方案。

  1. DZsig（暗区基因表达谱）阳性/阴性策略：
    • DZsig阳性患者：如前所述，DZsig（或称DHITsig）能够识别出预后不良的DLBCL亚组，包括那些FISH检测可能阴性但具有类似DHL生物学行为的患者 。DZsig阳性的DLBCL患者具有更高的侵袭性，对标准R-CHOP治疗反应差。对于DZsig阳性患者，即使FISH检测为阴性，也应考虑强化治疗方案，例如DA-EPOCH-R，甚至进一步结合新型靶向药物。此外，DZsig阳性的患者可能对MYC和BCL2通路的抑制剂更为敏感，例如联合使用BCL2抑制剂（如维奈克拉）和MYC相关通路抑制剂（如CAMKII抑制剂或BTK抑制剂）的策略。
    • DZsig阴性但DHL基因重排阳性患者：有研究发现，一小部分HGBCL-DH-BCL2（即经典的DHL）缺乏DZsig表达，这些患者的分子特征和预后可能与DZsig阳性的DHL不同，甚至与DLBCL-NOS具有相似的良好预后 。这提示DZsig可能有助于在经典的DHL中进一步分层，识别出预后相对较好、可能无需过度强化的患者。这部分患者可能对标准强化治疗（如DA-EPOCH-R）反应良好，甚至可以考虑在某些情况下降低治疗强度，以减少毒副作用。
  2. TP53突变状态：TP53是重要的抑癌基因，其突变在DHL中常见，并与极差的预后相关。对于伴有TP53突变的DHL患者，传统化疗效果更差。这类患者可能需要更积极的策略，例如联合MDM2抑制剂（如Idasanutlin）或新型化疗药物，或考虑更早进行细胞疗法（如CAR-T）。
  3. BCL2突变和表达水平：除了基因重排，BCL2蛋白的高表达或某些特定突变也可能影响维奈克拉的敏感性。通过对BCL2突变谱和表达水平的精确评估，可以筛选出对维奈克拉更敏感的患者，并指导维奈克拉与其他药物（如BTK抑制剂、CD20抗体）的联合使用。
  4. NOTCH2突变：近期研究发现，DLBCL相关的NOTCH2突变能够逃避泛素依赖性降解，导致蛋白稳定和致癌性RAS信号通路激活，从而促进化疗耐药。靶向NOTCH2的抑制剂（如γ-分泌酶抑制剂Nirogacestat）与AKT抑制剂（如Ipatasertib）联合使用，在CHOP耐药的DLBCL肿瘤中具有协同作用，提示了NOTCH2作为DHL潜在靶点的可能 87。这为具有特定NOTCH2突变的DHL患者提供了新的治疗方向。

多组学（基因组+转录组+蛋白组）指导的精准治疗临床试验

多组学技术整合了基因组学（DNA测序）、转录组学（RNA测序）和蛋白质组学（蛋白质定量）等多个层面的数据，能够全面揭示肿瘤的分子图谱，为精准治疗临床试验提供前所未有的洞察力。

  1. 全面理解DHL的生物学异质性：通过多组学分析，可以识别出DHL不同亚型之间细微的分子差异，例如MYC重排的精确断裂点、BCL2和BCL6的伴随突变、肿瘤微环境的免疫浸润特征以及信号通路的激活状态。这有助于更精确地定义治疗敏感或耐药的分子亚群。
  2. 识别新的治疗靶点：基因组测序可以发现新的驱动突变和基因拷贝数变异；转录组测序可以揭示异常激活的信号通路和差异表达的基因；蛋白质组学则可以直接观察蛋白质水平的改变，并评估药物靶点的可及性。例如，通过多组学分析可以识别DHL特有的高表达蛋白，为开发新型ADC药物或双特异性抗体提供靶点。
  3. 预测药物反应和耐药机制：多组学数据有助于构建复杂的生物信息学模型，预测患者对特定药物（如BTK抑制剂、BCL2抑制剂、免疫检查点抑制剂）的反应，并在治疗早期识别潜在的耐药机制。例如，通过分析治疗前后肿瘤组织的转录组变化，可以发现肿瘤细胞为逃避药物作用而激活的代偿性通路，从而指导后续的联合治疗或序贯治疗。
  4. 指导篮式/伞式临床试验：多组学分析为“篮式”或“伞式”临床试验提供了基础。在篮式试验中，不同肿瘤类型但具有相同分子特征的患者被纳入同一个治疗组，接受靶向该分子特征的药物治疗。在伞式试验中，单一肿瘤类型（如DHL）的患者被根据其分子特征分为多个亚组，每个亚组接受不同的靶向治疗。这种设计能够加速新药的开发和验证，特别是在罕见且异质性高的肿瘤类型中。
  5. 循环肿瘤DNA（ctDNA）在多组学中的应用：ctDNA的动态监测不仅可以用于MRD检测和早期复发预测，还可以通过其携带的基因组信息，实时反映肿瘤的进化和耐药克隆的出现。结合ctDNA的基因组和转录组分析（如液体活检中的mRNA），可以实现对治疗过程中肿瘤分子特征的非侵入性、实时监测，为动态调整治疗方案提供依据。

总之，DHL的精准治疗正在从基于单一基因重排的定义，向整合多层面分子信息的复杂多组学模型演进。通过不断深入了解DHL的分子生物学，并利用先进的分析技术，未来的治疗策略将更加个体化，有望显著改善患者的预后，实现真正的精准医学。

6.3 新型疗法与转化研究

随着对双打击淋巴瘤（DHL）复杂分子生物学和肿瘤微环境的深入理解，以及免疫治疗领域的飞速发展，针对DHL的新型疗法和转化研究正展现出巨大的潜力。这些研究不仅致力于提高治疗应答率和持久缓解，更着眼于克服现有疗法的局限性，开发出更安全、更有效、更易于推广的治疗方案。

双特异性抗体与CAR-NK细胞

  1. 双特异性抗体（Bispecific Antibodies）：除了前文提到的CD20×CD3双特异性抗体，如莫苏奈妥珠单抗（mosunetuzumab）和格罗菲他马布（glofitamab）等，未来将有更多新型双特异性抗体被开发出来，靶向DHL细胞表面的其他特异性抗原或免疫检查点分子。这些抗体通过同时结合肿瘤细胞和免疫效应细胞，或同时阻断两个免疫抑制通路，实现更强大的抗肿瘤活性。其优势在于“即用型”特性、生产相对简便以及毒副作用可控，使其成为CAR-T细胞疗法的有力补充，尤其适用于CAR-T治疗失败或不适用的患者。未来的研究将探索不同靶点组合的双特异性抗体，以及与化疗、靶向治疗或细胞疗法的联合应用，以期进一步提高疗效。

  2. CAR-NK细胞（Chimeric Antigen Receptor-Natural Killer Cells）：CAR-T细胞疗法虽然在R/R DHL中表现出色，但其高昂的成本、复杂的制备过程以及潜在的严重毒副作用（如细胞因子释放综合征[CRS]和免疫效应细胞相关神经毒性综合征[ICANS]）限制了其广泛应用。CAR-NK细胞作为下一代细胞免疫疗法，有望克服这些挑战。

    • 优势：NK细胞是先天免疫系统的关键组成部分，具有广谱的抗肿瘤活性。与CAR-T细胞相比，CAR-NK细胞具有以下优势：
      • 安全性更高：CAR-NK细胞通常不会引起严重的CRS和ICANS，且移植物抗宿主病（GvHD）风险极低，这使得其可以作为异基因“通用型”疗法，无需进行HLA配型 8889。例如，一项I/II期临床试验显示，源自脐带血的CD19 CAR-NK细胞治疗11名R/R CD19阳性癌症患者，未观察到CRS、神经毒性或GvHD，且73%的患者获得缓解，其中7例完全缓解 88。
      • 生产更简便：CAR-NK细胞可以从健康供者的脐带血中大规模制备，实现“即用型”供应，降低了生产成本和时间 90。
      • 多靶点效应：NK细胞本身具有多种活化和抑制受体，除了CAR介导的特异性杀伤，还能通过其天然识别机制（如识别MHC-I下调的细胞）发挥抗肿瘤作用。
    • 临床进展：目前，CD19靶向的CAR-NK细胞疗法已在临床试验中展现出良好前景 9091。一项I期临床试验针对8名复发/难治性大B细胞淋巴瘤患者，重复输注CD19-BBz CAR-NK细胞，结果显示无剂量限制性毒性、CRS、神经毒性或GvHD，且30天总缓解率达62.5%，其中50%的患者达到完全缓解 91。未来，CAR-NK细胞疗法将进一步优化CAR结构、提高体内持久性，并探索与其他疗法的联合应用，以期为DHL患者提供更安全、可及的细胞免疫治疗选择。

溶瘤病毒（Oncolytic Viruses）

溶瘤病毒是一类能够选择性感染并裂解肿瘤细胞，同时激活宿主抗肿瘤免疫反应的病毒。在DHL中，溶瘤病毒可以靶向感染淋巴瘤细胞，并通过病毒复制直接杀伤肿瘤。更重要的是，病毒裂解肿瘤细胞后释放的肿瘤相关抗原和危险信号分子，能够招募并激活免疫细胞（如T细胞和NK细胞），增强全身性的抗肿瘤免疫。目前，溶瘤病毒与免疫检查点抑制剂、化疗或靶向治疗的联合应用正处于积极探索中，以期发挥协同效应。例如，溶瘤病毒可以通过改变肿瘤微环境，使其从“冷肿瘤”转变为“热肿瘤”，从而增强免疫检查点抑制剂的疗效。

基于肿瘤微环境调控的联合策略开发

肿瘤微环境（TME）在DHL的免疫逃逸中扮演着关键角色。靶向调控TME，特别是激活CD11c+树突细胞等免疫细胞，是未来转化研究的重要方向。

  1. CD11c+树突细胞激活：研究表明，DHL肿瘤微环境中CD11c+树突细胞的减少与不良预后相关 19。因此，通过多种策略激活或增加CD11c+ DCs的数量和功能，有望逆转免疫抑制状态，增强抗肿瘤免疫。

    • FLT3L：FMS样酪氨酸激酶3配体（FLT3L）能够促进DC的增殖和分化。联合使用FLT3L与CAR-T细胞疗法或免疫检查点抑制剂，可能通过增加DCs数量来增强抗肿瘤免疫。
    • TLR激动剂：Toll样受体（TLR）激动剂能够直接激活DCs，促进其抗原提呈和细胞因子分泌。
    • PIKfyve抑制剂：PIKfyve是一种在CD11c+细胞中表达的基因，其高表达与免疫检查点抑制剂治疗效果不佳相关。抑制PIKfyve（例如通过apilimod）能够增强CD11c+细胞功能，促进T细胞免疫，并增强ICB的疗效。这一策略为靶向活化DHL中的树突细胞提供了新的可能。
    • 溶瘤病毒和肿瘤疫苗：溶瘤病毒裂解肿瘤细胞和肿瘤疫苗能够释放大量肿瘤抗原，被DCs摄取和提呈，从而激活特异性T细胞反应。

  2. 抑制免疫抑制细胞和分子：除了激活免疫效应细胞，抑制TME中的免疫抑制细胞（如调节性T细胞[Tregs]、髓源性抑制性细胞[MDSCs]和肿瘤相关巨噬细胞[TAMs]）以及免疫抑制分子（如PD-1/PD-L1、CTLA-4）也是重要的策略。针对DHL的TME特征，开发选择性抑制MDSCs或TAMs的药物，或将免疫检查点抑制剂与其他免疫刺激剂联合应用，有望取得突破。例如，靶向巨噬细胞的药物可以与免疫检查点阻断剂联用，以提高抗肿瘤免疫反应。

  3. 肿瘤外泌体的调控：淋巴瘤细胞分泌的外泌体（Exosomes）在TME中发挥重要作用，它们不仅含有致癌分子（如c-Myc、Bcl-2），还能通过促进血管生成、免疫抑制和细胞增殖来促进肿瘤生长 92。靶向抑制淋巴瘤来源外泌体的产生或其功能，可能成为新的治疗策略。例如，一项研究表明，DLBCL来源的外泌体可作为免疫抑制介质，诱导T细胞凋亡并上调PD-1，同时促进基质细胞增殖和血管生成，从而促进肿瘤生长 92。因此，开发抑制外泌体生成或阻断其信号通路的药物，有望协同其他疗法，增强DHL的治疗效果。

总而言之，未来针对DHL的新型疗法和转化研究将是一个多维度、综合性的领域。通过不断开发下一代细胞疗法（如CAR-NK）、优化双特异性抗体、探索溶瘤病毒的潜力，并结合对肿瘤微环境的精细调控策略（如激活CD11c+DCs和抑制外泌体作用），有望为DHL患者带来更安全、高效且持久的治疗方案，最终改善这一侵袭性淋巴瘤的临床结局。

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参考文献

1Current Treatment of Burkitt Lymphoma and High-Grade B-Cell Lymphomas.PubMed

P Connor Johnson, Jeremy S Abramson
Oncology (Williston Park). 2022 Aug 10;36(8):499-505. doi: 10.46883/2022.25920970.
PURPOSE OF REVIEW: This article reviews the current data and future directions in the management of Burkitt lymphoma (BL) and high-grade B-cell lymphoma (HGBL). RECENT FINDINGS: BL is a rare, mature B-cell lymphoma molecularly defined by translocation of the proto-oncogene MYC. Multiple intensive combination chemoimmunotherapy regimens have demonstrated excellent efficacy in this disease, although treatment toxicity remains a challenge in many patients. Double-hit lymphoma (DHL) represents HGBL with translocations of the oncogene MYC along with either BCL2 or BCL6, or both. In 2016, the World Health Organization update of this classification was revised to a new entity defined by cytogenetics: HGBL with MYC and BCL2 and/or BCL6 rearrangements. Recent prospective data using dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, and rituximab has demonstrated encouraging treatment efficacy in these patients. HGBL, not otherwise specified (NOS) is a heterogeneous, aggressive, mature B-cell lymphoma that does not meet criteria for BL, DHL, or diffuse large B-cell lymphoma NOS. Therapy for this entity is not well established. SUMMARY: The aggressive B-cell lymphomas BL, DHL, and HGBL, NOS are unique diseases with specific pathogenesis and biology. Insights into the molecular biology of these diseases have enabled new classifications and personalization of therapy.

2The high-grade B-cell lymphomas: double hit and more.PubMed

Andrew J Davies
Blood. 2024 Dec 19;144(25):2583-2592. doi: 10.1182/blood.2023020780.
Both the 2022 World Health Organization Classification of Hematolymphoid Tumors, 5th Edition and the International Consensus Classification of lymphoma have refined the way we now approach high-grade B-cell lymphoma (HGBL) with MYC and BCL2 and/or BCL6 rearrangements moving the previous generation of classification a step forward. The unifying biology of MYC/BCL2 tumors has become clearer and their inferior prognosis confirmed compared with those with morphologic similar phenotypes but lacking the classifcation defining cytogenetic abnormalities. Fluorescent in situ hybridization testing has now become largely population based, and we have learned much from this. We can readily define molecular categories and apply these widely to clinical practice. Uncertainty has, however, been shed on the place of MYC/BCL6 translocations in defining a common disease group of double hit lymphoma due to biological heterogeneity. We have enhanced our knowledge of outcomes and the role of therapy intensification to overcome chemotherapy resistance in HGBL. For those patients failed by initial induction chemotherapy, immunotherapy approaches, including chimeric antigen receptor T-cell therapies, are improving outcomes. Novel inhibitors, targeting dysregulated oncogenic proteins, are being explored at pace. The rare, but difficult, diagnostic classification HGBL (not otherwise specified) remains a diagnosis of exclusion with limited data on an optimal clinical approach. The days of talking loosely of double- and triple-hit lymphoma are numbered as biology and outcomes may not be shared. This review synergizes the current data on biology, prognosis, and therapies in HGBL.

3Motive and opportunity: MYC rearrangements in high-grade B-cell lymphoma with MYC and BCL2 rearrangements (an LLMPP study).PubMed

Laura K Hilton, Brett Collinge, Susana Ben-Neriah, et al.
Blood. 2024 Aug 1;144(5):525-540. doi: 10.1182/blood.2024024251.
Rearrangements that place the oncogenes MYC, BCL2, or BCL6 adjacent to superenhancers are common in mature B-cell lymphomas. Lymphomas with diffuse large B-cell lymphoma (DLBCL) or high-grade morphology with both MYC and BCL2 rearrangements are classified as high-grade B-cell lymphoma with MYC and BCL2 rearrangements ("double hit"; HGBCL-DH-BCL2) and are associated with aggressive disease and poor outcomes. Although it is established that MYC rearrangements involving immunoglobulin (IG) loci are associated with inferior outcomes relative to those involving other non-IG superenhancers, the frequency of and mechanisms driving IG vs non-IG MYC rearrangements have not been elucidated. Here, we used custom targeted capture and/or whole-genome sequencing to characterize oncogene rearrangements across 883 mature B-cell lymphomas including Burkitt lymphoma, follicular lymphoma, DLBCL, and HGBCL-DH-BCL2 tumors. We demonstrate that, although BCL2 rearrangement topology is consistent across entities, HGBCL-DH-BCL2 have distinct MYC rearrangement architecture relative to tumors with single MYC rearrangements or with both MYC and BCL6 rearrangements (HGBCL-DH-BCL6), including both a higher frequency of non-IG rearrangements and different architecture of MYC::IGH rearrangements. The distinct MYC rearrangement patterns in HGBCL-DH-BCL2 occur on the background of high levels of somatic hypermutation across MYC partner loci in HGBCL-DH-BCL2, creating more opportunity to form these rearrangements. Furthermore, because 1 IGH allele is already disrupted by the existing BCL2 rearrangement, the MYC rearrangement architecture in HGBCL-DH-BCL2 likely reflects selective pressure to preserve both BCL2 and B-cell receptor expression. These data provide new mechanistic explanations for the distinct patterns of MYC rearrangements observed across different lymphoma entities.

4Recent advancements in double-expressor lymphoma: novel therapeutic approaches and prospects.PubMed

Yuejian Zhuo, Dongdong Zhang
Oncologist. 2025 Jun 4;30(6). doi: 10.1093/oncolo/oyaf085.
Double-expressor lymphoma (DEL) is a newly identified special subtype of diffuse large B-cell lymphoma (DLBCL), which is predominantly found in the activated B-cell-like (ABC) subtype of DLBCL. Characterized by concurrent overexpression of BCL2 and MYC, DEL is associated with poorer prognosis. Standard chemoimmunotherapy can achieve clinical cure in nearly 70% of DLBCL cases. DEL mainly presents with intermediate-to-high-risk international prognostic index scores, advanced stage at diagnosis, and may involve specific chromosomal rearrangements, mainly influencing older patients. These factors are interconnected and contribute to less favorable treatment outcomes. We review emerging drugs and clinical trial data potentially effective against DEL, formulating treatment recommendations based on evidence levels to provide a theoretical foundation for the clinical treatment of DEL.

5Frequency Of Double Expressor Lymphoma In A Tertiary Care Hospital.PubMed

Masroor Hassan, Maria Tasneem Khattak, Muhammad Ashraf Qamar, et al.
J Ayub Med Coll Abbottabad. 2021 Jan-Mar;33(1):44-48.
BACKGROUND: Accounting for 30% cases of all lymphoid neoplasms, Diffuse large B cell lymphoma (DLBCL) is the commonest lymphoma worldwide. It occurs over a wide age range and has diverse morphology, immunophenotype and clinical outcome. Objectives of the study were to determine the frequency of Double expressor lymphoma (DEL) in a tertiary care hospital. METHODS: This descriptive cross-sectional study was carried out in the Department of Histopathology, Rehman Medical Institute Peshawar from June 1st to December 1st, 2018. A total of 88 newly diagnosed cases of diffuse large B-cell lymphoma (DLBCL); diagnosed on incisional or excisional biopsies were included in the study by non-probability consecutive sampling. Statistical analysis was carried out using SPSS-23. Quantitative variables were calculated as mean±SD. Qualitative variables were computed as frequency and percentages. Post stratification chi-square test was applied keeping p value equal or less than 0.05 as significance. RESULTS: In our 88 cases of DLBCL, 56 (63.6%) were males and 32 (36.4%) were females. Age of patients ranged from 15yrs to 84yrs. Mean age was 50.8±15.2SD. Activated B-cell like (ABC) subtype of DLBCL constitute 51 cases (58%) while 37 cases (42%) were of germinal centre B-cell like (GCB) subtype. Nineteen cases (21.6%) were of DEL. Cervical node was the commonest site of involvement (n=17, 19.3%) followed by stomach (n=10, 11.4%) and tonsil (n=6, 6.8%). Out of 19 cases of DEL, 17 cases (89.5%) were of ABC type. DEL was found to have significant correlation with ABC subtype of DLBCL (p=0.002). DEL had no correlation with gender (p=0.6), age (p=0.27), Mib-1 (p=0.36) and tumour site (p=0.42). CONCLUSIONS: The frequency of DEL in our study was 21.6% which is comparable to other studies who used similar cut-offs for c-Myc and BCL2 and similar criteria of inclusion as in our study. Significant association was found between DEL and ABC subtype of DLBCL.

6Colorectal diffuse large B-cell lymphoma: molecular subclassification and prognostic significance of immunoglobulin gene translocation.PubMed

Yoshifumi Hori, Hidetaka Yamamoto, Yui Nozaki, et al.
Hum Pathol. 2020 Feb;96:67-78. doi: 10.1016/j.humpath.2019.09.003. Epub 2019 Nov 15.
Primary colorectal diffuse large B-cell lymphoma (DLBCL) is rare, and its clinicopathological and genetic features are poorly understood. The aim of our study was to elucidate the frequency and prognostic significance of molecular subgroups in colorectal DLBCL. We examined 25 cases of colorectal lymphoma with DLBCL-like morphology and classified them into germinal center B-cell like (GCB)/non-GCB subgroups by immunohistochemistry (IHC) for CD10, bcl-6 and MUM1, or into double-expressor (DE)/non-DE subgroups by IHC for bcl-2 and c-myc. Translocations involving BCL2, BCL6, MYC, IGH, IGK, IGL, and MALT1 were also investigated using break-apart fluorescence in situ hybridization (FISH). The 25 cases were classified into two entities-DLBCL, not otherwise specified (NOS) (n = 23; 92%) and high grade B-cell lymphoma, double hit (n = 2; 8%)-according to the recent WHO classification. None of them showed histological evidence of Epstein-Barr virus infection or high-grade transformation from low grade B-cell lymphoma. Ten cases were GCB-type and four cases were DE-type, but these subtypes did not contribute to clinicopathological differences. Translocations involving BCL2, BCL6, MYC, IGH, IGK, IGL, and MALT1 were detected in 3 (12%), 3 (12%), 10 (40%), 14 (56%), 3 (12%), 3 (12%), and 0 (0%) of 25 cases, respectively. Of note, the presence of IGH translocation was significantly associated with better overall survival (P = .0053) and progression free survival (P = .0259). Similarly, the translocation involving at least one of the IGs (IGH, IGK, and/or IGL) was associated with more favorable prognosis in DLBCLs or even in DLBCL, NOS. This is the first report to reveal that a small subset of colorectal DLBCL corresponds to double-hit lymphoma. In addition, translocations involving at least one of the IGs may be a favorable prognostic factor in colorectal DLBCL. Testing the translocation involving rearrangement of IGs as well as MYC and BCL2/BCL6 may thus be useful for diagnosis and prognosis.

7Double-Hit Gene Expression Signature Defines a Distinct Subgroup of Germinal Center B-Cell-Like Diffuse Large B-Cell Lymphoma.PubMed

Daisuke Ennishi, Aixiang Jiang, Merrill Boyle, et al.
J Clin Oncol. 2019 Jan 20;37(3):190-201. doi: 10.1200/JCO.18.01583. Epub 2018 Dec 3.
PURPOSE: High-grade B-cell lymphoma with MYC and BCL2 and/or BCL6 rearrangements (HGBL-DH/TH) has a poor outcome after standard chemoimmunotherapy. We sought to understand the biologic underpinnings of HGBL-DH/TH with BCL2 rearrangements (HGBL-DH/TH- BCL2) and diffuse large B-cell lymphoma (DLBCL) morphology through examination of gene expression. PATIENTS AND METHODS: We analyzed RNA sequencing data from 157 de novo germinal center B-cell-like (GCB)-DLBCLs, including 25 with HGBL-DH/TH- BCL2, to define a gene expression signature that distinguishes HGBL-DH/TH- BCL2 from other GCB-DLBCLs. To assess the genetic, molecular, and phenotypic features associated with this signature, we analyzed targeted resequencing, whole-exome sequencing, RNA sequencing, and immunohistochemistry data. RESULTS: We developed a 104-gene double-hit signature (DHITsig) that assigned 27% of GCB-DLBCLs to the DHITsig-positive group, with only one half harboring MYC and BCL2 rearrangements (HGBL-DH/TH- BCL2). DHITsig-positive patients had inferior outcomes after rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone immunochemotherapy compared with DHITsig-negative patients (5-year time to progression rate, 57% and 81%, respectively; P < .001), irrespective of HGBL-DH/TH- BCL2 status. The prognostic value of DHITsig was confirmed in an independent validation cohort. DHITsig-positive tumors are biologically characterized by a putative non-light zone germinal center cell of origin and a distinct mutational landscape that comprises genes associated with chromatin modification. A new NanoString assay (DLBCL90) recapitulated the prognostic significance and RNA sequencing assignments. Validating the association with HGBL-DH/TH- BCL2, 11 of 25 DHITsig-positive-transformed follicular lymphomas were classified as HGBL-DH/TH- BCL2 compared with zero of 50 in the DHITsig-negative group. Furthermore, the DHITsig was shared with the majority of B-cell lymphomas with high-grade morphology tested. CONCLUSION: We have defined a clinically and biologically distinct subgroup of tumors within GCB-DLBCL characterized by a gene expression signature of HGBL-DH/TH- BCL2. This knowledge has been translated into an assay applicable to routinely available biopsy samples, which enables exploration of its utility to guide patient management.

8Epidemiology and etiology of diffuse large B-cell lymphoma.PubMed

Sophia S Wang
Semin Hematol. 2023 Nov;60(5):255-266. doi: 10.1053/j.seminhematol.2023.11.004. Epub 2023 Nov 27.
As the most common non-Hodgkin lymphoma subtype, diffuse large B-cell lymphoma (DLBCL) incidence patterns generally parallel that for NHL overall. Globally, DLBCL accounts for a third of all NHLs, ranging between 20% and 50% by country. Based on United States (U.S.) cancer registry data, age-standardized incidence rate for DLBCL was 7.2 per 100,000. DLBCL incidence rises with age and is generally higher in males than females; in the U.S., incidence is highest among non-Hispanic whites (9.2/100,000). Like NHL incidence, DLBCL incidence rose in the first half of the 20th century but has largely plateaued. However, there is some evidence that incidence rates are rising in areas of historically low rates, such as Asia; there are also estimates for rising DLBCL incidence in the near future due to the changing demographics in developed countries whose aging population is growing. Established risk factors for DLBCL include those that result in severe immune deficiency such as HIV/AIDS, inherited immunodeficiency syndromes, and organ transplant recipients. Factors that lead to chronic immune dysregulations are also established risk factors, and include a number of autoimmune conditions (eg, Sjögren syndrome, systemic lupus erythematosus, rheumatoid arthritis), viral infections (eg, HIV, KSHV/HHV8, HCV, EBV), and obesity. Family history of NHL/DLBCL, personal history of cancer, and multiple genetic susceptibility loci are also well-established risk factors for DLBCL. There is strong evidence for multiple environmental exposures in DLBCL etiology, including exposure to trichloroethylene, benzene, and pesticides and herbicides, with recent associations noted with glyphosate. There is also strong evidence for associations with other viruses, such as HBV. Recent estimates suggest that obesity accounts for nearly a quarter of DLBCLs that develop, but despite recent gains in the understanding of DLBCL etiology, the majority of disease remain unexplained. An understanding of the host and environmental contributions to disease etiology, and concerted efforts to expand our understanding to multiple race/ethnic groups, will be essential for constructing clinically relevant risk prediction models and develop effective strategies for disease prevention.

9High-Grade B-Cell Lymphoma (HGBL) with and and/or Rearrangements Is Predominantly BCL6-Rearranged and BCL6-Expressing in Taiwan.PubMed

Cheng-Chih Tsai, Yung-Cheng Su, Oluwaseun Adebayo Bamodu, et al.
Cancers (Basel). 2021 Mar 31;13(7):1620. doi: 10.3390/cancers13071620.
This study investigated the epidemiological and clinical peculiarities of and rearrangement in patients with high grade B-cell lymphoma (HGBL) from Taiwan, compared with data from Western countries. Two hundred and eighty-two DLBCL cases from Taipei Medical University-affiliated hospitals ( = 179) and Tri-Service General Hospital ( = 103) were enrolled for this study. From the 282, 47 (16.7%) had translocation; 24 of these harbored concurrent and/or translocation (double-hit, DH or triple-hit, TH). Twelve DH-HGBL cases had simultaneous and translocations, 8 harbored and rearrangement, while the remaining 4 patients exhibited TH. Together, 66.7% of DH/TH-HGBL patients were rearrangement positive. Among these -rearranged DH/TH-HGBL patients, only 6 (37.5%) overexpressed MYC and BCL6 proteins simultaneously, indicating that MYC-BCL6 co-overexpression may not be plausible surrogate biomarker for screening -rearranged DH-HGBL. By the end of year 5, all patients with TH-HGBL, DH-HGBL and all but one DH-HGBL cases had expired or were lost to follow-up. Progression-free survival (PFS) was longer for the non-DH/TH-HGBL group compared with the DH/TH-HGBL group. While the patients with DH-HGBL were lost to follow-up by day 800, their remaining TH-HGBL and DH-HGBL peers exhibited very poor PFS, regardless of age strata. More so, patients with rearrangement were 5.5-fold more likely associated with extranodal involvement compared with their BCL2-rearranged peers. Moreover, ~60.0% of the -rearranged DH-HGBL cases were non-GCB, suggesting that including screening for rearrangement in patients with the non-GCB phenotype may aid medical decision-making and therapeutic strategy. Contrary to contemporary data from western countries, 2 in every 3 patients with DH/TH-HGBL in Taiwan harbor rearrangement. Consistent with present findings, we recommend mandatory screening for rearrangement in patients with aggressive HGBL in Taiwan.

10Epidemiology of Non-Hodgkin's Lymphoma.PubMed

Krishna C Thandra, Adam Barsouk, Kalyan Saginala, et al.
Med Sci (Basel). 2021 Jan 30;9(1):5. doi: 10.3390/medsci9010005.
Non-Hodgins's lymphoma (NHL) is the most common hematological malignancy worldwide, accounting for nearly 3% of cancer diagnoses and deaths. NHL is the seventh most prevalent cancer and has the sixth highest mortality among cancers in the US. NHL accounts for 4% of US cancer diagnoses, and incidence has increased 168% since 1975 (while survival has improved 158%). NHL is more common among men, those >65 years old, and those with autoimmune disease or a family history of hematological malignancies. NHL is a heterogenous disease, with each subtype associated with different risk factors. Marginal zone lymphoma (MZL) is strongly associated with Sjogren's syndrome (SS) and Hashimoto's thyroiditis, while peripheral T-cell lymphoma (PTCL) is most associated with celiac disease. Occupational exposures among farm workers or painters increases the risk of most of the common subtypes. Prior radiation treatment, obesity, and smoking are most highly associated with diffuse large B-cell lymphoma (DLBCL), while breast implants have been rarely associated with anaplastic large cell lymphoma (ALCL). Infection with Epstein-Barr Virus (EBV) is strongly associated with endemic Burkitts lymphoma. HIV and human herpes virus 8 (HHV-8), is predisposed to several subtypes of DLBCL, and human T-cell lymphoma virus (HTLV-1) is a causative agent of T-cell lymphomas. Obesity and vitamin D deficiency worsen NHL survival. Atopic diseases and alcohol consumption seem to be protective against NHL.

11Epidemiology of diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) in the United States and Western Europe: population-level projections for 2020-2025.PubMed

Gena Kanas, Wenzhen Ge, Ruben G W Quek, et al.
Leuk Lymphoma. 2022 Jan;63(1):54-63. doi: 10.1080/10428194.2021.1975188. Epub 2021 Sep 11.
Diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) treatments have been rapidly evolving for patients treated in later lines of therapy (LoT). Country-specific cancer registry data for the US and Western Europe (WE) were combined with physician survey results to project the incidence, prevalence, and number of DLBCL and FL patients eligible for and treated by LoT between 2020 and 2025. The total number of incidents and prevalent cases of DLBCL and FL is expected to increase between 2020 and 2025 in the US and WE. 56% and 53% of the third line plus (3L+) eligible DLBCL patients and 60% and 55% of eligible FL patients initiated treatment in the US and WE, respectively. Further research is warranted to understand the reasons behind the high proportion of treatment eligible patients who do not initiate treatment, and potential differences between countries, especially in the 3L + settings.

12A Clinician's Approach to Double-Hit Lymphoma: Identification, Evaluation, and Management.PubMed

Ashley D Staton, Jonathon B Cohen
J Oncol Pract. 2016 Mar;12(3):232-8. doi: 10.1200/JOP.2015.009647.
Double-hit lymphomas have concurrent rearrangements of CMYC and BCL2 or occasionally BCL6. Although double-hit lymphomas are a part of the mature B-cell lymphoma lineage, they have an aggressive clinical course that is complicated by an extremely poor response to standard therapy for aggressive non-Hodgkin lymphoma. Overall survival is short for many patients with double-hit lymphomas, which reinforces the importance of identifying appropriate therapies for these patients. Fortunately, recent reports have demonstrated improved outcomes with the use of intensive induction therapies. This article discusses the biology, therapeutic considerations, treatment opinions, possible role of autologous stem-cell transplant, and need for ongoing clinical trials for this subgroup of patients with lymphoma.

13Non-IG::MYC in diffuse large B-cell lymphoma confers variable genomic configurations and MYC transactivation potential.PubMed

Chunye Zhang, Ellen Stelloo, Sharon Barrans, et al.
Leukemia. 2024 Mar;38(3):621-629. doi: 10.1038/s41375-023-02134-1. Epub 2024 Jan 6.
MYC translocation occurs in 8-14% of diffuse large B-cell lymphoma (DLBCL), and may concur with BCL2 and/or BCL6 translocation, known as double-hit (DH) or triple-hit (TH). DLBCL-MYC/BCL2-DH/TH are largely germinal centre B-cell like subtype, but show variable clinical outcome, with IG::MYC fusion significantly associated with inferior survival. While DLBCL-MYC/BCL6-DH are variable in their cell-of-origin subtypes and clinical outcome. Intriguingly, only 40-50% of DLBCL with MYC translocation show high MYC protein expression (>70%). We studied 186 DLBCLs with MYC translocation including 32 MYC/BCL2/BCL6-TH, 75 MYC/BCL2-DH and 26 MYC/BCL6-DH. FISH revealed a MYC/BCL6 fusion in 59% of DLBCL-MYC/BCL2/BCL6-TH and 27% of DLBCL-MYC/BCL6-DH. Targeted NGS showed a similar mutation profile and LymphGen genetic subtype between DLBCL-MYC/BCL2/BCL6-TH and DLBCL-MYC/BCL2-DH, but variable LymphGen subtypes among DLBCL-MYC/BCL6-DH. MYC protein expression is uniformly high in DLBCL with IG::MYC, but variable in those with non-IG::MYC including MYC/BCL6-fusion. Translocation breakpoint analyses of 8 cases by TLC-based NGS showed no obvious genomic configuration that enables MYC transactivation in 3 of the 4 cases with non-IG::MYC, while a typical promoter substitution or IGH super enhancer juxtaposition in the remaining cases. The findings potentially explain variable MYC expression in DLBCL with MYC translocation, and also bear practical implications in its routine assessment.

14Noncoding mutations cause super-enhancer retargeting resulting in protein synthesis dysregulation during B cell lymphoma progression.PubMed

Rebecca J Leeman-Neill, Dong Song, Jonathan Bizarro, et al.
Nat Genet. 2023 Dec;55(12):2160-2174. doi: 10.1038/s41588-023-01561-1. Epub 2023 Dec 4.
Whole-genome sequencing of longitudinal tumor pairs representing transformation of follicular lymphoma to high-grade B cell lymphoma with MYC and BCL2 rearrangements (double-hit lymphoma) identified coding and noncoding genomic alterations acquired during lymphoma progression. Many of these transformation-associated alterations recurrently and focally occur at topologically associating domain resident regulatory DNA elements, including H3K4me3 promoter marks located within H3K27ac super-enhancer clusters in B cell non-Hodgkin lymphoma. One region found to undergo recurrent alteration upon transformation overlaps a super-enhancer affecting the expression of the PAX5/ZCCHC7 gene pair. ZCCHC7 encodes a subunit of the Trf4/5-Air1/2-Mtr4 polyadenylation-like complex and demonstrated copy number gain, chromosomal translocation and enhancer retargeting-mediated transcriptional upregulation upon lymphoma transformation. Consequently, lymphoma cells demonstrate nucleolar dysregulation via altered noncoding 5.8S ribosomal RNA processing. We find that a noncoding mutation acquired during lymphoma progression affects noncoding rRNA processing, thereby rewiring protein synthesis leading to oncogenic changes in the lymphoma proteome.

15BCL6 suppression of BCL2 via Miz1 and its disruption in diffuse large B cell lymphoma.PubMed

Masumichi Saito, Urban Novak, Erich Piovan, et al.
Proc Natl Acad Sci U S A. 2009 Jul 7;106(27):11294-9. doi: 10.1073/pnas.0903854106. Epub 2009 Jun 23.
The BCL6 proto-oncogene encodes a transcriptional repressor that is required for germinal center (GC) formation and whose deregulation by genomic lesions is implicated in the pathogenesis of GC-derived diffuse large B cell lymphoma (DLBCL) and, less frequently, follicular lymphoma (FL). The biological function of BCL6 is only partially understood because no more than a few genes have been functionally characterized as direct targets of BCL6 transrepression activity. Here we report that the anti-apoptotic proto-oncogene BCL2 is a direct target of BCL6 in GC B cells. BCL6 binds to the BCL2 promoter region by interacting with the transcriptional activator Miz1 and suppresses Miz1-induced activation of BCL2 expression. BCL6-mediated suppression of BCL2 is lost in FL and DLBCL, where the 2 proteins are pathologically coexpressed, because of BCL2 chromosomal translocations and other mechanisms, including Miz1 deregulation and somatic mutations in the BCL2 promoter region. These results identify an important function for BCL6 in facilitating apoptosis of GC B cells via suppression of BCL2, and suggest that blocking this pathway is critical for lymphomagenesis.

16U-2932: two clones in one cell line, a tool for the study of clonal evolution.PubMed

H Quentmeier, R M Amini, M Berglund, et al.
Leukemia. 2013 Apr;27(5):1155-64. doi: 10.1038/leu.2012.358. Epub 2012 Dec 7.
Genetic heterogeneity is common in tumors, explicable by the development of subclones with distinct genetic and epigenetic alterations. We describe an in vitro model for cancer heterogeneity, comprising the diffuse large B-cell lymphoma cell line U-2932 which expresses two sets of cell surface markers representing twin populations flow-sorted by CD20 vs CD38 expression. U-2932 populations were traced to subclones of the original tumor with clone-specific immunoglobulin IgVH4-39 hypermutation patterns. BCL6 was overexpressed in one subpopulation (R1), MYC in the other (R2), both clones overexpressed BCL2. According to the combined results of immunoglobulin hypermutation and cytogenetic analysis, R1 and R2 derive from a mother clone with genomic BCL2 amplification, which acquired secondary rearrangements leading to the overexpression of BCL6 (R1) or MYC (R2). Some 200 genes were differentially expressed in R1/R2 microarrays including transcriptional targets of the aberrantly expressed oncogenes. Other genes were regulated by epigenetic means as shown by DNA methylation analysis. Ectopic expression of BCL6 in R2 variously modulated new candidate target genes, confirming dual silencing and activating functions. In summary, stable retention of genetically distinct subclones in U-2932 models tumor heterogeneity in vitro permitting functional analysis of oncogenes against a syngenic background.

17Design, synthesis and biological evaluation of novel selective PI3Kδ inhibitors containing pyridopyrimidine scaffold.PubMed

Mengyan Ma, Yifan Feng, San-Qi Zhang, et al.
Future Med Chem. 2023 Aug;15(16):1491-1509. doi: 10.4155/fmc-2023-0149. Epub 2023 Aug 11.
In our study compounds with pyrido[3,2-]pyrimidine and pyrido[3,4-]pyrimidine were designed, synthesized and evaluated for their biological activity against hematologic tumors. The biological activity of compounds was evaluated by ADP-Glo Luminescence assay, MTT [3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide] assay, western blotting and flow cytometry, respectively. Compounds , and containing pyrido[3,2-]pyrimidine inhibited phosphoinositide 3-kinase-δ (PI3Kδ) at subnanomolar levels and had good δ-isoform selectivity. , and showed significant inhibitory effects against SU-DHL-6 cells and effectively inhibited Akt phosphorylation in a good concentration-dependent manner. induced apoptosis and caused cell cycle arrest in SU-DHL-6 cells. Docking studies showed that , and bound tightly to PI3Kδ through key hydrogen bonding interactions. This study suggests that employing pyrido[3,2-]pyrimidine can facilitate the design of novel potent and selective PI3Kδ inhibitors.

18Identification of a potent and selective phosphatidylinositol 3-kinase δ inhibitor for the treatment of non-Hodgkin's lymphoma.PubMed

Wei-Qiong Zuo, Rong Hu, Wan-Li Wang, et al.
Bioorg Chem. 2020 Dec;105:104344. doi: 10.1016/j.bioorg.2020.104344. Epub 2020 Oct 8.
PI3Kδ has proved to be an effective target for anti-lymphoma drugs. However, the application of current approved PI3Kδ inhibitors has been greatly limited due to their specific immune-mediated toxicity and increased risk of infection, it is necessary to develop more PI3Kδ inhibitors with new scaffold. In this study, SAR study with respect to piperazinone-containing purine derivatives led to the discovery of a potent and selective PI3Kδ inhibitor, 4-(cyclobutanecarbonyl)-1-((2-(2-ethyl-1H-benzo[d]imidazol-1-yl)-9-methyl-6-morpholino-9H-purin-8-yl)methyl)piperazin-2-one (WNY1613). WNY1613 exhibits good antiproliferative activity against a panel of non-Hodgkin's lymphoma (NHL) cell lines by inducing cancer cell apoptosis and inhibiting the phosphorylation of PI3K and MAPK downstream components. In addition, it can also prevent the tumor growth in both SU-DHL-6 and JEKO-1 xenograft models without observable toxicity. WNY1613 thus could be developed as a promising candidate for the treatment of NHL after subsequent extensive pharmacodynamics and pharmacokinetics investigation.

19Decreased CD11c-positive dendritic cells in the tumor microenvironment predict double-hit/triple-hit genotype and survival in diffuse large B-cell lymphoma.PubMed

Chang-Tsu Yuan, Shih-Sung Chuang, Pei-Yuan Cheng, et al.
J Pathol Clin Res. 2022 Sep;8(5):436-447. doi: 10.1002/cjp2.283. Epub 2022 Jun 17.
Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma and is a potentially curable disease. However, it is heterogenous, and the prognosis is poor if the tumor cells harbor fusions involving MYC and BCL2 or MYC and BCL6 (double-hit [DH] lymphoma), or fusions involving all three genes (triple-hit [TH] lymphoma). Fluorescence in situ hybridization is currently the gold standard for confirming the presence of DH/TH genotypes. However, the test is laborious and not readily available in some laboratories. Germinal center B (GCB) signatures and dual expression of MYC and BCL2 are commonly used as initial screening markers (traditional model) in clinical practice. Our study proposes immunohistochemical markers for more conveniently and accessibly screening DH/TH genotypes in DLBCL. We retrospectively reviewed the clinical and pathological parameters of patients with DLBCL. We assessed the proliferative index, apoptotic index, and tumor microenvironment (TME), with regard to T cells and CD11c(+) dendritic cells, in formalin-fixed paraffin-embedded tissue. We then generated a decision tree as a screening algorithm to predict DH/TH genotypes and employed decision curve analysis to demonstrate the superiority of this new model in prediction. We also assessed the prognostic significance of related parameters. Our study revealed that GCB subtypes, a Ki67 proliferative index higher than 70%, and BCL2 expression were significantly associated with DH/TH genotypes. Decreased CD11c(+) dendritic cells in the TME indicated additional risk. Our proposed screening algorithm outperformed a traditional model in screening for the DH/TH genotypes. In addition, decreased CD11c(+) dendritic cells in the DLBCL TME were an independent unfavorable prognosticator. In conclusion, we provide a convenient, well-performing model that predicts DH/TH genotypes in DLBCL. The prognostic significance of CD11c(+) dendritic cells in the TME might influence the classification and development of immunotherapy for DLBCL in the future.

20Ultraviolet Radiation-Induced Tolerogenic Dendritic Cells in Skin: Insights and Mechanisms.PubMed

Gelare Ghajar-Rahimi, Nabiha Yusuf, Hui Xu
Cells. 2025 Feb 18;14(4):308. doi: 10.3390/cells14040308.
Ultraviolet (UV) radiation has profound effects on the immune system, including the induction of tolerogenic dendritic cells (DCs), which contribute to immune suppression and tolerance. This review explores the roles of conventional CD11c⁺ DCs, as well as cutaneous Langerhans cells and CD11b⁺ myeloid cells, in UV-induced immune modulation. Two key mechanisms underlying the immunosuppressive relationship between UV and DCs are discussed: the inactivation of DCs and the induction of tolerogenic DCs. DCs serve as a critical link between the innate and adaptive immune systems, serving as professional antigen-presenting cells. In this context, we explore how UV-induced DCs influence the activity of specific T cell subsets, including regulatory T lymphocytes and T helper cells, and shape immune outcomes. Finally, we highlight the implications of UV-induced tolerogenic DCs in select dermatologic pathologies, including cutaneous lupus, polymorphic light eruption, and skin cancer. Understanding the mechanisms by which UV radiation alters DC function offers insights into the complex interplay between environmental factors and immune regulation, providing potential avenues for preventive and therapeutic intervention in UV-induced skin diseases.

21PIKfyve, expressed by CD11c-positive cells, controls tumor immunity.PubMed

Jae Eun Choi, Yuanyuan Qiao, Ilona Kryczek, et al.
Nat Commun. 2024 Jun 28;15(1):5487. doi: 10.1038/s41467-024-48931-9.
Cancer treatment continues to shift from utilizing traditional therapies to targeted ones, such as protein kinase inhibitors and immunotherapy. Mobilizing dendritic cells (DC) and other myeloid cells with antigen presenting and cancer cell killing capacities is an attractive but not fully exploited approach. Here, we show that PIKFYVE is a shared gene target of clinically relevant protein kinase inhibitors and high expression of this gene in DCs is associated with poor patient response to immune checkpoint blockade (ICB) therapy. Genetic and pharmacological studies demonstrate that PIKfyve ablation enhances the function of CD11c cells (predominantly dendritic cells) via selectively altering the non-canonical NF-κB pathway. Both loss of Pikfyve in CD11c cells and treatment with apilimod, a potent and specific PIKfyve inhibitor, restrained tumor growth, enhanced DC-dependent T cell immunity, and potentiated ICB efficacy in tumor-bearing mouse models. Furthermore, the combination of a vaccine adjuvant and apilimod reduced tumor progression in vivo. Thus, PIKfyve negatively regulates the function of CD11c cells, and PIKfyve inhibition has promise for cancer immunotherapy and vaccine treatment strategies.

22Dendritic cell-intrinsic PTPN22 negatively regulates antitumor immunity and impacts anti-PD-L1 efficacy.PubMed

Santiago Acero-Bedoya, Emily F Higgs, Anna C Martinez, et al.
J Immunother Cancer. 2024 Oct 26;12(10):e009588. doi: 10.1136/jitc-2024-009588.
BACKGROUND: Individuals with a loss-of-function single-nucleotide polymorphism in the gene encoding PTPN22 have an increased risk for autoimmune diseases, and patients with cancer with such alleles may respond better to checkpoint blockade immunotherapy. Studies in PTPN22 knockout (KO) mice have established it as a negative regulator of T cell responses in cancer models. However, the role of PTPN22 in distinct immune cell compartments, such as dendritic cells (DCs), remains undefined. METHODS: We developed a novel PTPN22 conditional KO (cKO) mouse model that enables specific deletion in CD11c DCs by crossing to CD11c-Cre transgenic mice. Antitumor immunity was characterized using the B16.SIY and MC38.SIY cancer models and immune profiles of relevant tissues were evaluated by spectral flow cytometry. Antigen uptake, processing, and presentation, as well as DC proliferation to Flt3L, were characterized ex vivo. RESULTS: Deletion of PTPN22 in DCs resulted in augmented antitumor immunity in multiple syngeneic tumor models. Tumor antigen-specific CD8 T cells were increased in the tumor microenvironment (TME) of PTPN22 cKO mice and improved tumor control was CD8 T cell-dependent. Augmented T cell priming was also detected at early time points in the draining lymph nodes, and these effects were correlated with an increased number of proliferating CD103 DCs, also seen in the TME. In vitro studies revealed increased DC proliferation in response to Flt3L, as well as increased antigen processing and presentation. PTPN22 cKO mice bearing MC38 parental tumors showed combinatorial benefit with anti-PD-L1 therapy. CONCLUSIONS: Deletion of PTPN22 in DCs is sufficient to drive an augmented tumor antigen-specific T cell response, resulting in enhanced tumor control. PTPN22 negatively regulates DC proliferation and antigen processing and presentation. Our work argues that PTPN22 is an attractive therapeutic target for cancer immunotherapy and highlights the potential to modulate antitumor immunity through the manipulation of DC signaling.

23PD-L1 expression is low in large B-cell lymphoma with MYC or double-hit translocation.PubMed

Mette Vestergaard Elbaek, Mette Ølgod Pedersen, Marie Fredslund Breinholt, et al.
Hematol Oncol. 2019 Oct;37(4):375-382. doi: 10.1002/hon.2664. Epub 2019 Aug 25.
In large B-cell lymphoma (LBCL), MYC translocation and MYC/BCL2 or MYC/BCL6 double hit (DH) are associated with poor prognosis, and there is an unmet need for novel treatment targets in this patient group. Treatments targeting the PD-L1/PD-1 pathway are still poorly elucidated in LBCL. PD-L1 expression might predict response to treatment targeting the PD-L1/PD-1 pathway. We therefore investigated the relationship between PD-L1 protein and mRNA expression levels and MYC and DH translocation in LBCL. We detected MYC, BCL2, and BCL6 translocation by fluorescent in situ hybridization in tissue samples from 130 patients randomly selected from two cohorts of patients with LBCL: 49 patients with MYC translocation of whom 36 had DH and 81 without MYC translocation. PD-L1 protein expression was detected by immunohistochemistry (IHC) in tissue samples from 77 patients and PD-L1 mRNA expression by next-generation RNA sequencing (NGS) in another 77 patients. Twenty-four patients overlapped, ie, were analysed with both IHC and NGS. Nonparametric tests were performed to evaluate intergroup differences. PD-L1 protein expression level was significantly lower in patients with MYC (n = 42, median = 3.3%, interquartile range [IQR] 0.0-10.8) or DH translocations (n = 31, median = 3.3%, IQR 0.0-10.0) compared with patients with no MYC (n = 35, median = 16.7%, IQR 3.3-30.0) or no DH translocations (n = 46, 13.3%, IQR 2.5-30.0), P = .004 and P ≤ .001, respectively. PD-L1 mRNA expression was also significantly lower in patients with MYC or DH translocations, P = .001 and P = .006, respectively. Higher PD-L1 protein and mRNA expression levels were associated with non-germinal centre (GC) type compared with germinal centre B-cell (GCB)-type diffuse LBCL (DLBCL), P = .004 and P = .002, respectively. In conclusion, we report an association between low PD-L1 expression and MYC and DH translocation in patients with LBCL. Our findings may indicate that patients with MYC or DH translocation may benefit less from treatment with PD-L1/PD-1-inhibitors compared with patients without these translocations. This should be evaluated in larger, prospective, consecutive trials.

24High-grade B-cell lymphomas with MYC and BCL2 translocations lack tumor-associated macrophages and PD-L1 expression: A possible noninflamed subgroup.PubMed

Marie F Breinholt, Douglas V N P Oliveira, Tobias W Klausen, et al.
Hematol Oncol. 2021 Aug;39(3):284-292. doi: 10.1002/hon.2839. Epub 2021 Feb 8.
We investigated the intratumoral source of PD-L1 expression and the infiltration of tumor-associated macrophages (TAMs) in large B-cell lymphomas (LBCLs) with or without MYC-translocation, as well as possible correlations to BCL2-and BCL6-translocations and cell of origin (COO). One-hundred and twenty-six patient samples were studied in a cohort enriched for MYC-translocated tumors with 34 samples carrying this translocation. Demonstration of intratumoral distribution and cellular source of PD-L1 was enabled by immunohistochemical (IHC) dual staining specifically highlighting PD-L1 expression in lymphoma B-cells with antibodies against PD-L1 and PAX5. Additional IHC with antibodies against CD68 and CD163 identified TAMs. We found that CD68-positive TAMs were the main source of PD-L1 protein expression in contrast to lymphoma B cells which rarely expressed PD-L1. Semiquantitative IHC demonstrated a significant correlation between CD68 and PD-L1 protein expression. Unsupervised hierarchical analysis of PD-L1, CD68, and CD163 IHC data subsequently demonstrated three potential clusters defined by expression of the three biomarkers. Cluster A consisted of patient samples with significantly lower expression of PD-L1, CD68, and CD163, but also significantly higher prevalence of BCL2-translocation and MYC-BCL2-double-hit (DH) compared to the other two clusters. In cluster C we found a significant accumulation of BCL6 translocated tumors. This cluster in contrast had the highest protein expression of PD-L1, CD68, and CD163. Cluster B tumors had an intermediate expression of the three biomarkers, but no accumulation of the specific genetic translocations. Our data, which were based on morphological analysis, immunophenotyping and genotyping by fluorescence in situ hybridization were in line with new concepts of LBCL taxonomy integrating genetic, phenotypical, and immunological characteristics with identification of new subgroups where MYC translocation and MYC-BCL2 DH may identify a noninflamed subtype. These findings may furthermore hold significant predictive value especially regarding immune checkpoint blockade therapy, but further molecular characterization should be done to substantiate this hypothesis.

25Comparison of PD-L1 detection assays and corresponding significance in evaluation of diffuse large B-cell lymphoma.PubMed

Sixia Huang, Lin Nong, Li Liang, et al.
Cancer Med. 2019 Jul;8(8):3831-3845. doi: 10.1002/cam4.2316. Epub 2019 May 31.
The expression of programmed cell death ligand 1 (PD-L1) is a biomarker for immunotherapy, but approved detection method is absent in diffuse large B-cell lymphoma (DLBCL). Here, we performed three methods including immunohistochemistry (IHC) (clone SP263 and SP142), RNAscope, and fluorescence in situ hybridization (FISH) to evaluate PD-L1 status on a cohort of DLBCL including 94 of DLBCL-NOS, 25 of primary mediastinal large B-cell lymphoma (PMBCL) and 7 of double-hit lymphoma (DHL). SP263 with 25% for immune cell (IC) or combined cell and SP142 with 10% for tumor cell (TC), 20% for both of IC and combined cell were proved to have corresponding survival prognostic. Combined showed comparable prognostic value with TC and IC . SP263 and SP142 showed strong concordance (k = 0.788) with combined rates of 33.3% (42/126) and 34.9% (44/126), respectively. In DLBCL-NOS, TC by SP263 preferred to non-GCB and immunoblastic variant DLBCL-NOS (P = 0.029 and P = 0.004). Combined (SP263 and SP142) were associated with more than one extranodal site involved (P = 0.006, P = 0.042), higher ECOG PS scores (P = 0.001, P < 0.001), high IPI risk (P = 0.012, P = 0.005), and poor treatment response (P = 0.095, P = 0.002). IC by SP263 and SP142 were both independent risk factors (P = 0.027, P = 0.037). 9p24.1 locus amplification and gain were identified in 4.3% and 7.6% DLBCL-NOS and indicated shorter overall survival (P = 0.004). Positive rate of PD-L1 by RNAscope was 36.5%, while no clinical significance shown. PD-L1 positive rates were all higher in PMBCL and DHL than in DLBCL-NOS by SP263, SP142, RNAscope, and FISH (P = 0.001, P < 0.001, P = 0.005 and P < 0.001, respectively). In conclusion, combined PD-L1 expression by IHC was potentially reliable and convenient as a predicting biomarker. SP263 staining was easier to evaluate and recognized more PD-L1-stained cells, but SP142 presented a better prognostic indicator. FISH and RNAscope could be used as supplementary assays. PMBCL itself was a sensitive cohort for immunotherapy.

26Single-cell RNA sequencing in double-hit lymphoma: IMPDH2 induces the progression of lymphoma by activating the PI3K/AKT/mTOR signaling pathway.PubMed

Guangcan Gao, Qingfeng Xue, Jing He, et al.
Int Immunopharmacol. 2023 Dec;125(Pt A):111125. doi: 10.1016/j.intimp.2023.111125. Epub 2023 Oct 29.
BACKGROUND: IMPDH2 is the rate-limiting enzyme of the de novo GTP synthesis pathway and has a key role in tumors; however, the specific mechanism underlying IMPDH2 activity in diffuse large B cell lymphoma (DLBCL) is still undetermined. This study aims to explore the potential mechanism of IMPDH2 in DLBCL, and its possible involvement in double-hit lymphoma (DHL), i.e., cases with translocations involving MYC and BCL2 and/or BCL6. METHODS: Using single-cell sequencing and bioinformatics analysis to screen for IMPDH2. Exploring the differential expression of IMPDH2 and its correlation with prognosis through multiplexed immunofluorescence analysis. Using CCK8, EdU, clone formation assay, and animal model to analyze biological behavior changes after inhibiting IMPDH2. Explaining the potential mechanism of IMPDH2 in DLBCL by Western blot and multiplexed immunofluorescence. RESULTS: Prognostic risk model was constructed by single-cell sequencing, which identified IMPDH2 as a DHL-related gene. IMPDH2 was highly expressed in cell lines and tissues, associated with poor patient prognosis and an independent prognostic factor. In vitro and in vivo experiments showed that IMPDH2 inhibition significantly inhibited DHL cell proliferation. Flow cytometry showed apoptosis and cycle arrest. Western blot results suggested that c-Myc regulated the activation of PI3K/AKT/mTOR signaling pathway by IMPDH2 to promote tumor development in DHL. Moreover, multiplex immunofluorescence revealed decreased T-cell infiltration within the tumor microenvironment exhibiting concurrent high expression of IMPDH2 and PD-L1. CONCLUSIONS: Our results suggest that IMPDH2 functions as a tumor-promoting factor in DHL. This finding is expected to generate novel insights into the pathogenesis of these patients, thereby identifying potential therapeutic targets.

27Drug-induced aneuploidy and polyploidy is a mechanism of disease relapse in MYC/BCL2-addicted diffuse large B-cell lymphoma.PubMed

Shariful Islam, Andrew L Paek, Michael Hammer, et al.
Oncotarget. 2018 Nov 13;9(89):35875-35890. doi: 10.18632/oncotarget.26251.
UNLABELLED: Double-hit (DH) or double-expresser (DE) lymphomas are high-grade diffuse large B-cell lymphomas (DLBCL) that are mostly incurable with standard chemo-immunotherapy due to treatment resistance. The generation of drug-induced aneuploid/polyploid (DIAP) cells is a common effect of anti-DLBCL therapies (e.g. vincristine, doxorubicin). DIAP cells are thought to be responsible for treatment resistance, as they are capable of re-entering the cell cycle during off-therapy periods. Previously we have shown that combination of alisertib plus ibrutinib plus rituximab can partially abrogate DIAP cells and induce cell death. Here, we provide evidence that DIAP cells can re-enter the cell cycle and escape cell death during anti-DLBCL treatment. We also discuss MYC/BCL2 mediated molecular mechanism that underlie treatment resistance. We isolated aneuploid/polyploid populations of DH/DE-DLBCL cells after treatment with the aurora kinase (AK) inhibitor alisertib. Time-lapse microscopy of single polyploid cells revealed that following drug removal, a subset of these DIAP cells divide and proliferate by reductive cell divisions, including multipolar mitosis, meiosis-like nuclear fission and budding. Genomic, proteomic, and kinomic profiling demonstrated that alisertib-induced aneuploid/polyploid cells up-regulate DNA damage, DNA replication and immune evasion pathways. In addition, we identified amplified receptor tyrosine kinase and T-cell receptor signaling, as well as MYC-mediated dysregulation of the spindle assembly checkpoints . We infer that these factors contribute to treatment resistance of DIAP cells. These findings provide opportunities to develop novel DH/DE-DLBCL therapies, specifically targeting DIAP cells. KEY POINTS: ● MYC mediated upregulation of TPX2, KPNA2 and RanGAP1 dysregulate the spindle assembly checkpoint in drug-induced polyploid cells.● Drug-induced polyploid cells re-enter the cell cycle via multipolar mitosis, fission or budding, a mechanism of disease relapse.

28A role for in constraining germinal center B cell formation.PubMed

Xingxing Shao, Xin Liu, Hai Qi
Cell Insight. 2023 Jan 28;2(2):100078. doi: 10.1016/j.cellin.2023.100078. eCollection 2023 Apr.
Germinal center is a transient lymphoid tissue structure in which B cells undergo affinity maturation and differentiate into memory B cells and plasma cells. GC formation depends on B cell expression of BCL6, a master transcription regulator of the GC state. Bcl6 expression is under elaborate control by external signals. HES1 plays important roles in T-cell lineage commitment, although little is known about its potential roles in GC formation. Here we report that B-cell-specific HES1 deletion causes a significant increase in GC formation, leading to increased production of plasma cells. We further provide evidence that HES1 inhibits BCL6 expression in a bHLH domain-dependent manner. Our study suggests a new layer of regulation of GC initiation mediated by HES1 and, by inference, Notch signals .

29Double Hit and Double Expresser Diffuse Large B Cell Lymphoma Subtypes: Discrete Subtypes and Major Predictors of Overall Survival.PubMed

Anurag Mehta, Ajita Verma, Garima Gupta, et al.
Indian J Hematol Blood Transfus. 2020 Oct;36(4):627-634. doi: 10.1007/s12288-019-01248-w. Epub 2020 Jan 8.
Double hit lymphomas (DHL) and double expresser lymphomas (DEL) are subsets of diffuse large B cell lymphomas (DLBCL) which are being increasingly recognised as cause of treatment failure. This emphasizes the need for their separation from other DLBCL cases in order to prognosticate and administer more aggressive treatment to this set of patients. The present study was conducted with the aim to identify the DHL/DEL patients and study their distinctive clinicopathological profile and overall survival. This retrospective analysis involved 172 cases of DLBCL sub-classified on the basis of cell of origin. Immunohistochemical (IHC) analysis for and CD10 was performed. Rearrangement studies were performed using break apart Fluorescent in situ hybridization. Overall survival (OS) was also evaluated. Distinctive clinical and pathological features of DHL and DEL were identified. Rearrangement study by FISH revealed seven cases of DHL (+ &/or rearrangement). Also, 20 patients (11.6%) showed a concurrent expression of and oncoproteins (DEL) on IHC. Most (6/7) DHL patients were double expressors also. The DHL patients demonstrated a significant association with female gender, high serum LDH levels (> 750 U/L) and GCB phenotype. DEL patients contrarily predominated amongst males, had intermediate LDH levels (251-500 U/L) and non GCB phenotype. The OS of the patients was 63.8% at 4 years. The OS of the DLBCL, DEL and DHL patients was 71.9%, 46.9%, and 0%, respectively at 4 years ( value 0.010). In case of DEL subtype, factors such as age < 60 years (66.7%), male sex (60.8%), nodal localization (52.5%), early disease stage (84.6%), low IPI score (60%), absence of B symptoms (50%), LDH < 250 U/L (80%) and GCB phenotype (53.3%) were associated with better OS. Further, the OS of DHL cases was 0% at 4 years. Double hit and double expresser lymphomas have poor prognostic outcomes and should be separated from DLBCL. All DELs should be tested for DHLs and especially those with immunoblastic morphology. DHL and DEL subtypes delineate the subtypes with inferior OS and reinstate the need for aggressive interventions.

30Diffuse large B-cell lymphoma.PubMed

Shaoying Li, Ken H Young, L Jeffrey Medeiros
Pathology. 2018 Jan;50(1):74-87. doi: 10.1016/j.pathol.2017.09.006. Epub 2017 Nov 20.
Diffuse large B cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma worldwide, representing approximately 30-40% of all cases in different geographic regions. Patients most often present with a rapidly growing tumour mass in single or multiple, nodal or extranodal sites. The most common type of DLBCL, designated as not otherwise specified, represents 80-85% of all cases and is the focus of this review. There are also rare types of lymphoma composed of large B-cells, in aggregate about 15-20% of all neoplasms that are sufficiently distinctive to recognise separately. DLBCL not otherwise specified (referred to henceforth as DLBCL) is a heterogeneous entity in terms of clinical presentation, genetic findings, response to therapy, and prognosis. A major advance was the application of gene expression profiling (GEP) to the study of DLBCL which further clarified this heterogeneity and provided a rationale for subdividing cases into groups. The most popular system divides cases of DLBCL according to cell-of-origin into germinal centre B-cell like (GCB) and activated B-cell like (ABC) subtypes, with about 10-15% of cases being unclassifiable. Patients with the GCB subtype usually have better prognosis than patients with the ABC subtype. Although cell-of-origin is useful for predicting outcome, the GCB and ABC subtypes remain heterogeneous, with better and worse prognostic subsets within each group. Next generation sequencing (NGS) analysis of DLBCL has facilitated global identification of numerous and diverse genetic abnormalities in these neoplasms and has shown that GCB and ABC tumours have different mutation profiles. Although the therapy of patients with DLBCL is an active area of research, the current 5-year overall survival rate is 60-70% using standard-of-care frontline therapy. A precision medicine approach for the design of new therapies based on molecular findings in DLBCL is likely the best path forward. As pathologists, our role has expanded beyond diagnosis. We must perform a complete work-up of DLBCL cases. In addition to our traditional role in establishing the diagnosis, we need to analyse markers that provide information regarding prognosis and potential therapeutic targets. We also must ensure that adequate tissue is triaged for molecular studies which are essential for designing therapy regimens, particularly in the setting of disease relapse.

31Diffuse large B-cell lymphoma: 2019 update on diagnosis, risk stratification, and treatment.PubMed

Yang Liu, Stefan Klaus Barta
Am J Hematol. 2019 May;94(5):604-616. doi: 10.1002/ajh.25460.
DISEASE OVERVIEW: Diffuse large B-cell lymphoma (DLBCL) is the most common type of aggressive non-Hodgkin lymphoma originating from the germinal center, and it represents a heterogeneous group of diseases with variable outcomes that are differentially characterized by clinical features, cell of origin (COO), molecular features, and most recently, frequently recurring mutations. DIAGNOSIS: DLBCL is ideally diagnosed from an excisional biopsy of a suspicious lymph node, which shows sheets of large cells that disrupt the underlying structural integrity of the follicle center and stain positive for pan-B-cell antigens, such as CD20 and CD79a. COO is determined by immunohistochemical stains, while molecular features such as double-hit or triple-hit disease are determined by fluorescent in situ hybridization analysis. Commercial tests for frequently recurring mutations are currently not routinely used to inform treatment. RISK STRATIFICATION: Clinical prognostic systems for DLBCL, including the rituximab International Prognostic Index, age-adjusted IPI, and NCCN-IPI, use clinical factors for the risk stratification of patients, although this does not affect the treatment approach. Furthermore, DLBCL patients with non-germinal center B-cell (GCB)-like DLBCL (activated B-cell like and unclassifiable) have a poorer response to up-front chemoimmunotherapy (CI) compared to patients with GCB-like DLBCL. Those with c-MYC-altered disease alone and in combination with translocations in BCL2 and/or BCL6 (particularly when the MYC translocation partner is immunoglobulin) respond poorly to up-front CI and salvage autologous stem cell transplant at relapse. RISK-ADAPTED THERAPY: This review will focus on differential treatment of DLBCL up-front and at the time of relapse by COO and molecular features.

32Screening Strategy for Detecting Double-Hit Lymphoma in a Resource-Limited Setting.PubMed

Balamurugan Thirunavukkarasu, Amanjit Bal, Gaurav Prakash, et al.
Appl Immunohistochem Mol Morphol. 2022 Jan 1;30(1):49-55. doi: 10.1097/PAI.0000000000000967.
AIM: High-grade B-cell lymphomas with MYC and BCL2 and/or BCL6 rearrangements [double-hit lymphomas (DHL)] are aggressive lymphomas. Current literature recommends fluorescent in-situ hybridization analysis (FISH) in all cases of diffuse large B-cell lymphoma (DLBCL) to identify cases of DHL. However, this approach is not feasible in a resource-limited setting. We analyzed cases of de novo high-grade B-cell non-Hodgkin lymphoma using histomorphology, immunohistochemistry and FISH to identify which cases need to undergo FISH testing in a resource-limited setting. MATERIALS AND METHODS: Cases of de novo high-grade B-cell non-Hodgkin lymphoma that included DLBCL, not otherwise specified and B-cell lymphoma unclassifiable (BCLU) with features intermediate between DLBCL and Burkitt lymphoma diagnosed over a period of 5 years were analyzed by Hans algorithm, MYC, BCL2, and Ki67. MYC, BCL2, and BCL6 break apart FISH was tested in selected cases. RESULTS: One hundred and nine cases were obtained, of which 102 had DLBCL morphology and 7 had BCLU/blastoid morphology. BCL2 expression was noted in 48 cases (44%), MYC in 33 cases (30.3%) and MYC/BCL2 co-expression in 24 cases (22%). FISH testing could be done in 42 consecutive cases, of which 5 cases had MYC and BCL2 co-rearrangement (11.9%) (double-hit) and 2 cases showed rearrangement for only MYC (4.7%) (single-hit). Single-hit lymphoma/DHL showed significant independent positive correlation with BCLU/blastoid morphology, CD10 expression, germinal center B-cell phenotype, and MYC/BCL2 co-expression. The sensitivity and specificity of each parameters include BCLU/blastoid morphology (42% vs. 94%), CD10 positive (50% vs. 88%), germinal center B-cell phenotype (57% vs. 82%), MYC/BCL2 co-expression (85% vs. 80%). Selected candidates for FISH (any one of the above parameters) using this strategy showed a sensitivity and specificity of 100% and 68%, respectively (P=0.001). CONCLUSION: We propose a highly sensitive screening strategy for detection of MYC/BCL2 rearrangement in high-grade B-cell lymphoma in a resource-limited setting (pending validation in a larger cohort).

33An immunohistochemical study of double-expressor lymphomas and its correlation with cell of origin.PubMed

Anuradha Ananthamurthy
J Cancer Res Ther. 2023 Apr;19(Supplement):S0. doi: 10.4103/jcrt.jcrt_587_21.
BACKGROUND: Diffuse large B cell Lymphomas (DLBCL) that co express C MYC and BCL 2 are known as 'double expressor lymphomas' and are believed to have a worse prognosis than other diffuse large B cell lymphomas. This was a study to assess the frequency of double expressor lymphomas in our cohort of DLBCL. AIMS AND OBJECTIVES: The aims of this study were to assess the frequency of double expression of C MYC and BCL 2 in cases of DLBCL and to correlate the same with clinicopathological parameters including cell of origin, namely germinal centre type versus non germinal centre type. MATERIALS AND METHODS: This was a retrospective observational study Immunostaining for MYC antibody and BCL2 were performed using the standard polymer/DAB technique. 40% for MYC and 50% for BCL2 were taken as cut off values.Chi square analysis was used to compare the variables, and a p value of less than 0.05 was taken as statistically significant. RESULTS: Of 40 cases studied, 11 (27.5%) were double expressors. There was no significant correlation of double expression with gender, site (nodal versus extra nodal), cell of origin namely germinal centre/non germinal centre types and Ki67 index when compared to the other group which did not show double expression. CONCLUSIONS: Immunohistochemistry is a useful technique to detect double expressor lymphomas which are known to have an aggressive course. Cell of origin did not show significant correlation with double expression in our study.

34Double hit and double expressors in lymphoma: Definition and treatment.PubMed

Peter A Riedell, Sonali M Smith
Cancer. 2018 Dec 15;124(24):4622-4632. doi: 10.1002/cncr.31646. Epub 2018 Sep 25.
Emerging biologic subsets and new prognostic markers are significantly and adversely affecting curability after standard chemoimmunotherapy for aggressive B-cell lymphomas. The identification of concurrent MYC and B-cell CLL/lymphoma 2 (BCL2) deregulation, whether at a genomic or protein level, has opened a new era of investigation within the most common subtype of aggressive B-cell lymphomas. Double-hit lymphoma (DHL), defined as a dual rearrangement of MYC and BCL2 and/or B-cell CLL/lymphoma 6 (BCL6) genes, is an uncommon subset accounting for 5% to 7% of all diffuse large B-cell lymphomas (DLBCLs), and long-term survivors are rare. Double-expressor lymphoma (DEL), defined as overexpression of MYC and BCL2 proteins not related to underlying chromosomal rearrangements, is not a distinct entity in the current World Health Organization classification but accounts for 20% to 30% of DLBCL cases and also has poor outcomes. There are many practical considerations related to identifying, determining the prognosis of, and managing DHL and DEL.

35[Clinical characteristics and prognosis of concurrent positive t(14; 18) and myc gene rearrangement in diffuse large B cell lymphoma].PubMed

H W Zhang, Z W Chen, L Y Wang, et al.
Zhonghua Zhong Liu Za Zhi. 2016 Mar 23;38(3):206-10. doi: 10.3760/cma.j.issn.0253-3766.2016.03.009.
OBJECTIVE: To study the incidence of positive t(14; 18) and myc gene rearrangement, and the clinical features and prognosis of concurrent positive t(14; 18) and myc gene rearrangement "double-hit lymphoma" (DHL) in diffuse large B cell lymphoma. METHODS: The positive t(14; 18) and myc gene rearrangement in 106 cases of DLBCL were analyzed using interphase fluorescent in situ hybridization (FISH) technique. The expression of myc and bcl-2 proteins was determined by immunohistochemistry. The relationship of positive t(14; 18) and myc gene rearrangement with clinical features, pathogenesis and prognosis for the patients was analyzed. SPSS 16.0 software was used for statistical analysis. RESULTS: Among the 106 cases, there were 27 (25.5%) cases with positive t(14; 18) and 13 (12.3%) cases with myc gene rearrangement, and 7 cases (6.6%) of DLBCL with concurrent t(14; 18)-positive and myc gene rearrangement. A relationship was observed between positive t(14; 18) and myc gene rearrangement (P=0.019). The follow-up data showed that the 7 DHL patients were in age of 52-84 years, the International Prognostic Index (IPI) scores were 3 in two cases, 4 in four cases and 5 in one case, and the ECOG scores were 3 in all the 7 cases. Four patients had bone marrow involvement and were combined with leukemia. The survival time ranged from 0.5 to 6 months, with a median survival of 4 months. The univariate analysis showed that B symptom, Ann Arbor stage, ECOG score, LDH level, IPI score, immunophenotype, bcl-2 protein expression, myc protein expression, and myc gene rearrangement were all associated with poor prognosis (P<0.05 for all). The multivariate analysis using a COX proportional hazard model confirmed that ECOG score, bcl-2 protein expression, myc protein expression, myc gene rearrangement, and immunophenotype were independent prognostic factors affecting survival (P<0.05 for all), among them, the myc gene rearrangement was the strongest prognostic factor (OR=4.337, P<0.001). CONCLUSIONS: "Double-hit" DLBCL is rare and can be mainly identified only by molecular detection. Perhaps positive t(14; 18) and myc gene rearrangement play concurrent role in its "double-hit" pathogenesis. DHL are highly invasive, and most of DHL patients have poor prognosis. Further studies of larger case number are required to determine the pathologic features and the therapeutic strategy of this subgroup.

36The prognostic role of Bcl-2, Ki67, c-MYC and p53 in diffuse large B-cell lymphoma.PubMed

Ana Maria Pătraşcu, Ionela Rotaru, Lavinia Olar, et al.
Rom J Morphol Embryol. 2017;58(3):837-843.
Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous group of lymphoid malignancies, which counts for more than a third of non-Hodgkin's lymphoma cases. The aim of the current study is to evaluate the prognostic role of several immunohistochemical (IHC) markers involved in the pathological process of DLBCL. This is a retrospective analysis of the 97 de novo DLBCL patients admitted between January 2007 and December 2016 in the Department of Hematology, "Filantropia" Municipal Hospital, Craiova, Romania. The expression of Bcl-2, Ki67, c-MYC and p53 was assessed by immunohistochemistry. A significant level of association was observed between high prognostic index values and Bcl-2, Ki67, c-MYC and p53 positive cases. Moreover, overall survival and disease-free survival were higher in patients with negative expression for these markers. Bcl-2, Ki67, c-MYC and p53 could make important diagnostic and therapeutic targets; therefore, their routine assessment should be mandatory.

37Double hit lymphoma: contemporary understanding and practices.PubMed

Eashwar Somasundaram, Jeremy S Abramson
Leuk Lymphoma. 2025 Jan;66(1):26-33. doi: 10.1080/10428194.2024.2412295. Epub 2024 Oct 8.
Double-hit lymphoma (DHL) is a high-risk subtype of large B-cell lymphoma, defined by concurrent rearrangements and . The diagnosis is confirmed through histologic and immunophenotypic examination and fluorescence hybridization (FISH) to demonstrate the rearrangements. DHL morphology ranges from DLBCL to high-grade B-cell lymphoma which can resemble Burkitt lymphoma and is almost always germinal center B-cell like (GCB). Prognosis is influenced by elevated lactate dehydrogenase (LDH), advanced stage, and extranodal involvement, among other factors. Treatment outcomes vary, but intensive chemotherapy regimens such as dose-adjusted EPOCH-R have shown the most promising results, though low-risk cases do occur and may do well with less intensive treatments. Recent therapeutic advances such as CAR-T cells and bispecific antibodies offer promise for patients with relapsed/refractory disease. This review synthesizes data from recent literature to provide a comprehensive analysis of the molecular underpinnings, diagnostic criteria, prognostic factors, and therapeutic strategies for DHL.

38High-grade B-cell lymphoma, not otherwise specified: a multi-institutional retrospective study.PubMed

Adam S Zayac, Daniel J Landsburg, Mitchell E Hughes, et al.
Blood Adv. 2023 Nov 14;7(21):6381-6394. doi: 10.1182/bloodadvances.2023009731.
In this multi-institutional retrospective study, we examined the characteristics and outcomes of 160 patients with high-grade B-cell lymphoma, not otherwise specified (HGBL-NOS)-a rare category defined by high-grade morphologic features and lack of MYC rearrangements with BCL2 and/or BCL6 rearrangements ("double hit"). Our results show that HGBL-NOS tumors are heterogeneous: 83% of patients had a germinal center B-cell immunophenotype, 37% a dual-expressor immunophenotype (MYC and BCL2 expression), 28% MYC rearrangement, 13% BCL2 rearrangement, and 11% BCL6 rearrangement. Most patients presented with stage IV disease, a high serum lactate dehydrogenase, and other high-risk clinical factors. Most frequent first-line regimens included dose-adjusted cyclophosphamide, doxorubicin, vincristine, and etoposide, with rituximab and prednisone (DA-EPOCH-R; 43%); rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP; 33%); or other intensive chemotherapy programs. We found no significant differences in the rates of complete response (CR), progression-free survival (PFS), or overall survival (OS) between these chemotherapy regimens. CR was attained by 69% of patients. PFS at 2 years was 55.2% and OS was 68.1%. In a multivariable model, the main prognostic factors for PFS and OS were poor performance status, lactate dehydrogenase >3 × upper limit of normal, and a dual-expressor immunophenotype. Age >60 years or presence of MYC rearrangement were not prognostic, but patients with TP53 alterations had a dismal PFS. Presence of MYC rearrangement was not predictive of better PFS in patients treated with DA-EPOCH-R vs R-CHOP. Improvements in the diagnostic criteria and therapeutic approaches beyond dose-intense chemotherapy are needed to overcome the unfavorable prognosis of patients with HGBL-NOS.

39Utility of a Reverse Phase Protein Array to Evaluate Multiple Biomarkers in Diffuse Large B-Cell Lymphoma.PubMed

Masaki Suzuki, Atsushi Muroi, Masanori Nojima, et al.
Proteomics Clin Appl. 2020 Jan;14(1):e1900091. doi: 10.1002/prca.201900091. Epub 2019 Nov 27.
PURPOSE: Diffuse large B-cell lymphoma (DLBCL), the most common non-Hodgkin lymphoma, is a heterogeneous lymphoma with different clinical manifestations and molecular alterations, and several markers are currently being measured routinely for its diagnosis, subtyping, or prognostication by immunohistochemistry (IHC). Here, the utility of a reverse-phase-protein-array (RPPA) as a novel supportive tool to measure multiple biomarkers for DLBCL diagnosis is validated. EXPERIMENTAL DESIGN: The expression of seven markers (CD5, CD10, BCL2, BCL6, MUM1, Ki-67, and C-MYC) is analyzed by RPPA and IHC using 37 DLBCL tissues, and the correlation between the two methods is determined. To normalize tumor content ratio in the tissues, the raw RPPA values of each marker are adjusted by that of CD20 or PAX-5. RESULTS: The CD20-adjusted data for CD5, MUM1, BCL2, Ki-67, and C-MYC has better correlation with IHC results than PAX-5-adjusted data. Receiver operating characteristic (ROC) analysis reveals that CD5, MUM1, BCL2, and C-MYC exhibit a better sensitivity and specificity >0.750. Furthermore, the CD20-adjusted C-MYC value strongly correlates with that of IHC, and has a particularly high specificity (0.882). CONCLUSIONS AND CLINICAL RELEVANCE: Although further investigation using a large number of DLBCL specimens needs to be conducted, these results suggest that RPPA could be applicable as a supportive tool for determining lymphoma prognosis.

40Confirmation of the molecular classification of diffuse large B-cell lymphoma by immunohistochemistry using a tissue microarray.PubMed

Christine P Hans, Dennis D Weisenburger, Timothy C Greiner, et al.
Blood. 2004 Jan 1;103(1):275-82. doi: 10.1182/blood-2003-05-1545. Epub 2003 Sep 22.
Diffuse large B-cell lymphoma (DLBCL) can be divided into prognostically important subgroups with germinal center B-cell-like (GCB), activated B-cell-like (ABC), and type 3 gene expression profiles using a cDNA microarray. Tissue microarray (TMA) blocks were created from 152 cases of DLBCL, 142 of which had been successfully evaluated by cDNA microarray (75 GCB, 41 ABC, and 26 type 3). Sections were stained with antibodies to CD10, bcl-6, MUM1, FOXP1, cyclin D2, and bcl-2. Expression of bcl-6 (P <.001) or CD10 (P =.019) was associated with better overall survival (OS), whereas expression of MUM1 (P =.009) or cyclin D2 (P <.001) was associated with worse OS. Cases were subclassified using CD10, bcl-6, and MUM1 expression, and 64 cases (42%) were considered GCB and 88 cases (58%) non-GCB. The 5-year OS for the GCB group was 76% compared with only 34% for the non-GCB group (P <.001), which is similar to that reported using the cDNA microarray. Bcl-2 and cyclin D2 were adverse predictors in the non-GCB group. In multivariate analysis, a high International Prognostic Index score (3-5) and the non-GCB phenotype were independent adverse predictors (P <.0001). In summary, immunostains can be used to determine the GCB and non-GCB subtypes of DLBCL and predict survival similar to the cDNA microarray.

41Molecular determinants of clinical outcomes in a real-world diffuse large B-cell lymphoma population.PubMed

Waleed Alduaij, Brett Collinge, Susana Ben-Neriah, et al.
Blood. 2023 May 18;141(20):2493-2507. doi: 10.1182/blood.2022018248.
Molecular heterogeneity of diffuse large B-cell lymphoma (DLBCL) underlies the variable outcomes achieved with immunochemotherapy. However, outcomes of gene expression profiling (GEP)-defined molecular subgroups in a real-world DLBCL population remain unknown. Here we examined the prevalence and outcomes of molecular subgroups in an unselected population of 1149 patients with de novo DLBCL in British Columbia, Canada. Evaluable biopsies were profiled by fluorescence in situ hybridization (FISH), immunohistochemistry, and digital GEP to assign cell-of-origin and the so-called "double-hit signature" (DHITsig)-a signature originally described as being characteristic for high-grade B-cell lymphoma with MYC and BCL2 rearrangements (HGBCL-DH-BCL2). DHITsig was expressed in 21% of 431 germinal center B-cell-like (GCB)-DLBCL and all 55 Burkitt lymphomas examined. Reflecting this latter finding, DHITsig has been renamed the "dark zone signature" (DZsig). DZsigpos-DLBCL, non-DZsigpos GCB-DLBCL and activated B-cell-like (ABC)-DLBCL were associated with a 2 year overall survival of 57%, 89%, and 71%, respectively. 62% of DZsigpos tumors were negative for HGBCL-DH-BCL2 by FISH, but were associated with outcomes similar to HGBCL-DH-BCL2. A small group of HGBCL-DH-BCL2 that lacked DZsig expression had different molecular features compared with DZsig-expressing HGBCL-DH-BCL2 and were associated with favorable outcomes comparable to DLBCL, not otherwise specified. DZsigpos and ABC-DLBCL had a shorter diagnosis-to-treatment interval (DTI) than GCB-DLBCL, with this metric being associated with outcome. In conclusion, DZsig expression extends beyond HGBCL-DH-BCL2 and captures a poor-prognosis DLBCL subgroup with short DTI, including patients unidentifiable by routine FISH testing, that should be considered for treatment intensification or novel therapies in prospective trials.

42The double-hit signature identifies double-hit diffuse large B-cell lymphoma with genetic events cryptic to FISH.PubMed

Laura K Hilton, Jeffrey Tang, Susana Ben-Neriah, et al.
Blood. 2019 Oct 31;134(18):1528-1532. doi: 10.1182/blood.2019002600.
High-grade B-cell lymphomas with MYC and BCL2 and/or BCL6 rearrangements (HGBL-DH/THs) include a group of diffuse large B-cell lymphomas (DLBCLs) with inferior outcomes after standard chemoimmunotherapy. We recently described a gene expression signature that identifies 27% of germinal center B-cell DLBCLs (GCB-DLBCLs) as having a double-hit-like expression pattern (DHITsig) and inferior outcomes; however, only half of these cases have both MYC and BCL2 translocations identifiable using standard breakapart fluorescence in situ hybridization (FISH). Here, 20 DHITsig+ GCB-DLBCLs apparently lacking MYC and/or BCL2 rearrangements underwent whole-genome sequencing. This revealed 6 tumors with MYC or BCL2 rearrangements that were cryptic to breakapart FISH. Copy-number analysis identified 3 tumors with MYC and 6 tumors with MIR17HG gains or amplifications, both of which may contribute to dysregulation of MYC and its downstream pathways. Focal deletions of the PVT1 promoter were observed exclusively among DHITsig+ tumors lacking MYC translocations; this may also contribute to MYC overexpression. These results highlight that FISH fails to identify all HGBL-DH/THs, while revealing a range of other genetic mechanisms potentially underlying MYC dysregulation in DHITsig+ DLBCL, suggesting that gene expression profiling is more sensitive for identifying the biology underlying poor outcomes in GCB-DLBCL.

43An immunohistochemical germinal center B-cell dark zone signature identifies Burkitt lymphoma and molecular high-grade B-cell lymphomas.PubMed

Xiaoxian Zhao, Alexandra Balmaceda, Via S Abiera, et al.
Am J Clin Pathol. 2025 Oct 4;164(4):559-566. doi: 10.1093/ajcp/aqaf074.
OBJECTIVE: We hypothesized that a set of immunohistochemistry (IHC) stains could be used to distinguish Burkitt lymphoma (BL), the quintessential B-cell lymphoma with a germinal center B-cell (GCB) dark zone (DZ) expression signature, from diffuse large B-cell lymphoma, not otherwise specified (DLBCL, NOS). This might also be applicable to high-grade B-cell lymphomas (HGBCLs) with MYC and BCL2 rearrangements (double-hit lymphomas [DHLs]) and triple-hit lymphomas (THLs). METHODS: A 5-marker IHC algorithm was designed from gene lists that distinguish physiologic DZ from light zone GCBs. RESULTS: In training and validation cohorts, we distinguished BL from DLBCL, NOS with high sensitivity and specificity. Because DHLs/THLs are enriched for the gene expression DZ signature (DZsig), we evaluated 19 DHLs/THLs and 4 HGBCLs, NOS. Most (83%) cases were IHC DZ. The NanoString DLBCL90 assay was performed on 34 cases to correlate IHC DZ results with the molecular DZsig. The IHC DZ call was significantly associated with the DZsig (P = .0011). The sensitivity and specificity of IHC to recognize DZsig+ cases among DLBCL, NOS and DHLs with BCL2 rearrangements/THLs were 91% and 100%, respectively. CONCLUSIONS: The IHC DZ algorithm can support a diagnosis of BL and identifies MYC-BCL2 DHLs/THLs with a molecular DZsig.

44Development and validation of a prognostic nomogram for diffuse large B-cell lymphoma: a multicenter cohort study from Northern China.PubMed

Juya Cui, Tao Wang, Junyan Zhang, et al.
Clin Transl Oncol. 2025 Nov;27(11):4269-4279. doi: 10.1007/s12094-025-03955-5. Epub 2025 May 23.
OBJECTIVE: This study aimed to refine the prognostic stratification provided by the International Prognostic Index (IPI) for patients diagnosed with diffuse large B-cell lymphoma (DLBCL) in a northern Chinese cohort and to construct a more precise prognostic nomogram to facilitate individualized treatment strategies and improve survival outcomes. METHODS: Clinical data from 1,364 patients diagnosed with DLBCL between January 2000 and December 2019 were retrospectively analyzed across multiple centers in Beijing and Shanxi, China. A prognostic nomogram model for overall survival (OS) was developed incorporating the following variables: age, disease stage, Eastern Cooperative Oncology Group (ECOG) performance status, lactate dehydrogenase (LDH) levels, involvement of various extranodal sites, and disease progression within 6 months (POD6). Model performance was evaluated through receiver-operating characteristic (ROC) curve analysis, calibration curves, concordance index (C-index), and area under the curve (AUC) metrics. RESULTS: The developed nomogram demonstrated consistent predictive capacity, with AUC values exceeding 0.7 at 1-, 3-, and 5-year time points. Calibration plots indicated close alignment with the ideal reference line, with slopes approximating 1, supporting the model's predictive accuracy and clinical relevance. POD6 was identified as the most significant high-risk factor associated with decreased OS (hazard ratio [HR] = 5.13, 95% confidence interval [CI]: 1.03-2.51, p < 0.0001). Among the IPI components, all except ECOG performance status and extranodal involvement remained significant. Notably, central nervous system involvement exhibited the strongest adverse prognostic effect among extranodal sites (HR = 1.61, 95% CI: 1.03-2.51, p = 0.035). CONCLUSION: A novel prognostic nomogram was established for patients with DLBCL, offering improved prognostic accuracy compared to the traditional IPI. This model presents clinical utility in supporting personalized management and treatment planning. CLINICAL REGISTRATION NUMBER: Not applicable.

45BCL2 and MYC dual-hit lymphoma/leukemia.PubMed

Naoto Tomita
J Clin Exp Hematop. 2011;51(1):7-12. doi: 10.3960/jslrt.51.7.
Translocation of the BCL2 gene on the chromosome band 18q21.3 results in consistent expression of the Bcl2 protein, an apoptosis inhibitor. BCL2 usually translocates to the immunoglobulin (IG) heavy chain (IGH) gene as t(14;18)(q32;q21.3) and rarely to IG light chain (IGK, IGL) loci as t(2;18)(p11;q21.3) or t(18;22)(q21.3;q11). The t(14;18) translocation is observed in 70-95% of follicular lymphoma cases and 20-30% of diffuse large B-cell lymphoma (DLBCL) cases. The MYC gene on chromosome band 8q24 acts as an accelerator of cell proliferation. MYC translocates to 14q32/IGH as t(8;14)(q24;q32) or less commonly to 2p11/IGK as t(2;8)(p11;q24) or 22q11/IGL as t(8;22)(q24;q11). The 8q24/MYC translocation is detected in nearly all Burkitt lymphoma (BL) and up to 10% of DLBCL cases. Both translocations rarely occur in an identical cell and this lymphoid malignancy is termed BCL2 and MYC dual-hit lymphoma/leukemia (DHL). The pathological diagnosis in most cases of DHL with BCL2-IG and MYC-IG translocation is B-cell lymphoma, unclassifiable, with features intermediate between DLBCL and BL, although DLBCL is most common in DHL with BCL2-IG and MYC-nonIG translocation. The frequency of DHL with BCL2 and MYC translocation is estimated at around 2% of all B-cell malignancies. The condition is characterized by elevated serum lactate dehydrogenase levels, the presence of B symptoms, bone marrow involvement, advanced disease stage, extranodal involvement, and central nervous system (CNS) involvement at presentation or disease progression. Despite treatment strategies including CNS-targeted therapy, the prognosis for DHL is extremely poor. In this review, the current knowledge of the clinicopathological status of DHL is summarized and discussed.

46Challenges and Opportunities for High-grade B-Cell Lymphoma With MYC and BCL2 and/or BCL6 Rearrangement (Double-hit Lymphoma).PubMed

Dongfeng Zeng, Aakash Desai, Fangfang Yan, et al.
Am J Clin Oncol. 2019 Mar;42(3):304-316. doi: 10.1097/COC.0000000000000427.
The most common subtype of non-Hodgkin lymphoma, diffuse large B-cell lymphoma, is cured in approximately two thirds of patients after initial therapy. The remaining one-third of patients who suffer relapse or become refractory have very poor survival outcomes despite salvage chemotherapy with or without stem cell transplantation. A considerable proportion of relapsed or refractory large B cells belong to the WHO subtype known as high-grade B-cell lymphoma with rearrangement of MYC and BCL2 and/or BCL6, also known as double-hit lymphoma (DHL). Most DHL patients present with Ann Arbor's stage III/IV, a comparatively higher rate of extranodal involvement including bone marrow and central nervous system infiltration, high levels of lactate dehydrogenase, and an elevated Ki67 expression in the tumor cells. Newer therapeutic approaches, including targeted therapy against BCL2, MYC, or other associated pathways, are needed. In addition, recent therapies that harness the immune system, such as checkpoint inhibitors and chimeric antigen receptor T-cell therapy, are changing the paradigm of treatment for non-Hodgkin lymphoma and could impact the outcome of DHL.

47Clinical features and prognostic factors of intravascular large B-cell lymphoma: a cohort study of 20 patients from 2018 to 2024.PubMed

Yaqing Li, Suxiao Li, Xiyang Liu, et al.
Oncologist. 2025 Jun 4;30(6). doi: 10.1093/oncolo/oyaf156.
BACKGROUND: Intravascular large B-cell lymphoma (IVLBCL) is a rare, aggressive lymphoma characterized by lymphoma cells proliferating within small blood vessels, lacking an extravascular mass. Its low incidence complicates diagnosis and treatment. METHODS: This study analyzed 20 IVLBCL patients diagnosed between 2018 and 2024, aiming to describe their clinical presentations, diagnostic procedures, treatment, and outcomes. RESULTS: All patients had stage IV disease with poor ECOG performance status and high-risk IPI scores (100% ≥ 4). Common symptoms included fever (65%), impaired consciousness (30%), and respiratory issues (25%). Laboratory findings revealed cytopenias (60% anemia, 60% thrombocytopenia, and 35% leukocytopenia), elevated lactate dehydrogenase (20/20, 100%), C-reactive protein (14/14, 100%), and erythrocyte sedimentation rate (11/12, 91.7%). Five of six patients had extreme IL-10 elevation. Imaging studies showed a high incidence of adrenal gland involvement (9/20, 45%), reproductive system involvement (7/20, 35%), central nervous system (CNS) involvement (6/20, 30%), and splenomegaly (9/20, 45%). Nineteen of 20 patients were non-GCB (CD10-MUM1+). Seven of 14 patients tested positive for CD5. Eighteen of 20 patients received CD20 antibody plus chemotherapy as first-line therapy, with complete responses (CRs) achieved in 7 of 18 patients. After a median follow-up of 42.4 months, the 1-year and 2-year overall survival rates were 71.3% and 65.4%, respectively. Unfavorable risk factors included central nervous system (CNS) and bone marrow (BM) involvement, and elevated creatinine. CONCLUSION: The study highlights the high incidence of adrenal and reproductive system involvement in IVLBCL, with CNS, BM, and elevated creatinine contributing to rapid disease progression.

48Impact of rituximab on incidence of and risk factors for central nervous system relapse in patients with diffuse large B-cell lymphoma: a systematic review and meta-analysis.PubMed

Jing Zhang, Bobin Chen, Xiaoping Xu
Leuk Lymphoma. 2014 Mar;55(3):509-14. doi: 10.3109/10428194.2013.811239. Epub 2013 Jul 29.
The impact of rituximab (R) on the incidence of central nervous system (CNS) relapse of diffuse large B-cell lymphoma (DLBCL) is unclear. We performed a meta-analysis to explore the risk factors and assess the association between R-chemotherapy (R-chemo) and CNS relapse. We searched MEDLINE, PubMed, EMBASE and OVID for eligible studies. Published group statistics were extracted from each study for analysis; individual patient data from each study were not accessed. Fixed- or random-effects models were used to estimate the odds ratio (OR) with a 95% confidence interval (CI). Eight studies were identified. The OR for R-chemo compared with identical chemotherapy was 0.70 (95% CI 0.54-0.91). Stage III/IV (OR 2.25, 95% CI 1.64-3.08), International Prognostic Index (IPI) > 1 (OR 2.62, 95% CI 1.59-4.33), performance status (PS) > 1 (OR 1.67, 95% CI 1.23-2.27), elevated lactate dehydrogenase (LDH) (OR 2.23, 95% CI 1.54-3.22), bone marrow involvement (OR 2.85, 95% CI 1.99-4.07), more than one extranodal involvement (OR 2.61, 95% CI 1.93-3.54), presence of B symptoms (OR 1.87, 95% CI 1.37-2.56) and testicular involvement (OR 3.83, 95% CI 1.84-7.97) were associated with increased risks of CNS relapse. This meta-analysis demonstrated a lower incidence of CNS relapse of DLBCL in the rituximab era. The risk of CNS relapse can be assessed by stage, IPI, PS, LDH, presence of B symptoms, number of extranodal sites, bone marrow and testicular involvement.

49Clinical characteristics and outcomes of newly diagnosed patients with HIV-associated aggressive B-cell NHL in China.PubMed

Chaoyu Wang, Jun Liu, Haike Lei, et al.
J Cell Mol Med. 2022 Oct;26(19):5067-5077. doi: 10.1111/jcmm.17534. Epub 2022 Sep 3.
Little is known about the incidence, clinical characteristics and prognostic factors in HIV associated lymphoma as these are less common than HIV-negative lymphoma in China. Currently, there are no standard guidelines for treatment of these patients. Therefore, we performed a study to analyse the clinical characteristics and outcomes of newly diagnosed HIV-associated aggressive B-cell non-Hodgkin's lymphoma (NHL) patients in Chongqing University Cancer Hospital (CUCH). Totally 86 newly diagnosed HIV-associated aggressive B-cell NHL patients in CUCH, southwest China, from July 2008 to August 2021, were analysed. In the entire cohort, median age was 48 years (range, 23-87 years), and more patients were male (87.2%). Most patients had elevated lactate dehydrogenase (LDH) (82.6%), advanced ann arbor stage (80.2%) and high IPI score (IPI score, 3-5) (62.7%) at diagnosis. Median CD4+ T-cell count at diagnosis was 191/μl (range, 4-1022), 84 patients (97.7%) were on combination antiretroviral therapy (cART) at lymphoma diagnosis. In DLBCL patients, cox multivariate analysis showed that age ≥ 60 (HR = 2.251, 95%CI 1.122-4.516; p = 0.012), elevated LDH (HR = 4.452, 95%CI 1.027-19.297; p = 0.041) and received less than two cycles of chemotherapy (HR = 0.629, 95%CI 0.589-1.071; p = 0.012) were independent risk factors for adverse prognosis based on PFS. Age ≥ 60 (HR = 3.162, 95%CI 1.500-6.665; p = 0.002) and received less than two cycles of chemotherapy (HR = 0.524, 95%CI 0.347-0.791; p = 0.002) were also independent risk factor for adverse prognosis based on OS. In BL patients, cox multivariate analysis showed that elevated LDH and received less than two cycles of chemotherapy were independent risk factors for adverse prognosis. In the DLBCL group, median PFS times in the received rituximab and no received rituximab groups were not reached and 12 months, respectively (p = 0.006). Median OS times were not reached and 36 months, respectively (p = 0.021). In the BL group, median PFS times in the received rituximab and no received rituximab groups were not reached and 4.8 months, respectively (p = 0.046). Median OS times were not reached and 10.1 months, respectively (p = 0.035). Overall, these data indicated that standardized anti-lymphoma therapy and rituximab administration were significantly associated with improved outcomes in patients with HIV-associated DLBCL and BL.

50Risk factors for poor treatment outcome and central nervous system relapse in diffuse large B-cell lymphoma with bone marrow involvement.PubMed

Keun-Wook Lee, Jongyoun Yi, In Sil Choi, et al.
Ann Hematol. 2009 Sep;88(9):829-38. doi: 10.1007/s00277-008-0682-6. Epub 2009 Jan 27.
Although several studies have described the prognostic implication of bone marrow (BM) involvement (BMI) in lymphoma, studies focused on BM-involved diffuse large B-cell lymphoma (DLBCL) are very rare and small-sized. This study was performed to examine the prognostic impact of morphologic findings of BMI by lymphoma and risk factors for central nervous system (CNS) relapse in BM-involved DLBCL. Between 1993 and 2005, 675 patients were diagnosed with DLBCL, and 88 patients who had BMI at initial diagnosis were eligible for this study. The median overall survival (OS) and failure-free survival (FFS) of 88 patients were 36.6 and 20.1 months, respectively. When three variables from BM morphologic findings (the pattern of BM infiltration, extent of BMI by lymphoma, and percentage of large cells in the infiltrate) were simultaneously included into multivariate model, the increased extent of BMI by lymphoma (> or =10%) in BM area was the only negative prognostic factor, independent of the International Prognostic Index (IPI). Patients with both lower IPI scores and less extent of BMI showed an excellent prognosis with chemotherapy alone (5-year OS and FFS rates, 80% and 69%). However, morphologic BM features were not independent predictive factors for CNS recurrences. An increased lactate dehydrogenase (LDH) level at initial diagnosis was the only independent predictive factor for CNS relapse. Further efforts should be directed toward finding optimal treatment modalities based on the IPI and the extent of BMI by lymphoma. CNS prophylaxis may be considered only in patients with initial elevated LDH levels.

51Primary breast double-hit lymphoma management and outcomes: a real-world multicentre experience.PubMed

Tingting Zhang, Yuanfeng Zhang, Hairong Fei, et al.
Cancer Cell Int. 2021 Sep 17;21(1):498. doi: 10.1186/s12935-021-02198-y.
BACKGROUND: Primary breast double-hit lymphoma (PB-DHL) is a rare, highly aggressive malignancy that poses challenges regarding accurate diagnosis and selecting optimal treatment regimens. METHODS: We retrospectively reviewed 48 cases of patients diagnosed with PB-DHL in six academic centres between June 2014 and June 2020 in China. Study-specific data were recorded, including treatment options, therapeutic evaluation, prognostic factors and relapse patterns, and the overall survival (OS) and progression-free survival (PFS) were evaluated. RESULTS: In total, 48 patients were enrolled, with 14 patients treated with DA-EPOCH-R/MA (rituximab, dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, alternating with high-dose methotrexate and cytarabine), 18 patients treated with DA-EPOCH-R (rituximab, dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin), and 16 patients treated with R-HyperCVAD (rituximab, hyperfractionated cyclophosphamide, vincristine, doxorubicin, dexamethasone, alternating with cytarabine plus methotrexate). The overall 5-year OS and PFS rates were 41.7% (95% confidence interval [CI], 27.6-56.8%) and 37.5% (95% CI, 24.0-52.6%), respectively. Of the three treatment regimens, the 5-year OS was higher in DA-EPOCH-R/MA group than in the DA-EPOCH-R or R-HyperCVAD subgroups (57.1% vs. 38.9% vs. 31.3%; P = 0.016), as was the 5-year PFS (50.0% vs. 38.9% vs. 25.0%; P = 0.035). Autologous stem cell transplantation (ASCT) prolonged the OS and PFS compared with non-ASCT patients (5-year OS: 72.2% vs. 23.3%; P < 0.001; 5-year PFS: 72.2% vs. 16.7 %, P < 0.001). Multivariate analysis identified tumour size, risk stratification, treatment with DA-EPOCH-R/MA, breast irradiation, and ASCT as significant prognostic factors. CONCLUSIONS: DA-EPOCH-R/MA is a promising regimen for PB-DHL, and breast irradiation yields complementary benefits for prognosis. ASCT significantly decreased disease relapse, providing a potential curative PB-DHL intervention and justifying ASCT as first-line therapy for young patients. More effective treatment strategies for PB-DHL patients remain encouraging.

52Baseline Total Metabolic Tumor Volume is Prognostic for Refractoriness to Immunochemotherapy in DLBCL: Results From GOYA.PubMed

Irene Canales Ruiz, Maurizio Martelli, Laurie H Sehn, et al.
Clin Lymphoma Myeloma Leuk. 2022 Aug;22(8):e804-e814. doi: 10.1016/j.clml.2022.04.010. Epub 2022 Apr 14.
INTRODUCTION: A good response to initial therapy is key to maximizing survival in patients with diffuse large B-cell lymphoma (DLBCL), but patients with chemorefractory disease and early progression have poor outcomes. PATIENTS AND METHODS: Data from the GOYA study in patients with DLBCL who received first-line rituximab or obinutuzumab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) were analyzed. Positron emission tomography/computed tomography (PET/CT)-derived characteristics associated with total metabolic tumor volume (TMTV) and clinical risk factors for primary chemorefractory disease and disease progression within 12 months (POD12) were explored. RESULTS: Of those patients fulfilling the criteria for analysis, 108/1126 (10%) were primary chemorefractory and 147/1106 (13%) had POD12. Primary chemorefractory and POD12 status were strongly associated with reduced overall survival. After multivariable analysis of clinical and imaging-based risk factors by backward elimination, only very high TMTV (quartile [Q] 1 vs. Q4 odds ratio [OR]: 0.45; P = .006) and serum albumin levels (low vs. normal OR of 1.86; P = .004) were associated with primary chemorefractoriness. After additionally accounting for BCL2/MYC translocation in a subset of patients, TMTV and BCL2/MYC double-hit status remained as significant predictors of primary chemorefractoriness (Q1 vs. Q4 OR: 0.32, P = .01 and double-hit vs. no-hit OR of 4.47, P = .02, respectively). Risk factors including very high TMTV, high sum of the product of the longest diameters (SPD), geographic region (Asia), short time since diagnosis, extranodal involvement and low serum albumin were retained for POD12. CONCLUSION: PET-derived TMTV has prognostic value in identifying patients at risk of early treatment failure.

53Double hit diffuse large B-cell lymphomas: diagnostic and therapeutic challenges.PubMed

Jonathan W Friedberg
Chin Clin Oncol. 2015 Mar;4(1):9. doi: 10.3978/j.issn.2304-3865.2015.03.02.
Although diffuse large B-cell lymphoma (DLBCL) is curable with standard chemoimmunotherapy, over 30% of patients with advanced stage disease experience refractory disease or progression. Recent studies suggest that rearrangement of the myc oncogene occurs in approximately 10% of patients with DLBCL, and confers a very poor prognosis, particularly when there is concomitant rearrangement of bcl-2, a condition referred to as "double hit DLBCL". Using immunohistochemistry, up to 30% of patients have evidence of increased expression of myc, which occurs in both activated B-cell and germinal center type DLBCL. When bcl-2 is also positive by immunohistochemistry, prognosis is also poor. There are no randomized studies guiding treatment for patients with double hit DLBCL, but new datasets are emerging suggesting a possible role for dose-adjusted EPOCH infusional chemotherapy with rituximab. This review will conclude with a survey of novel agents which may be rationally incorporated into chemotherapy platforms for this high risk subset of DLBCL.

54B-cell lymphomas with concurrent IGH-BCL2 and MYC rearrangements are aggressive neoplasms with clinical and pathologic features distinct from Burkitt lymphoma and diffuse large B-cell lymphoma.PubMed

Matija Snuderl, Olga K Kolman, Yi-Bin Chen, et al.
Am J Surg Pathol. 2010 Mar;34(3):327-40. doi: 10.1097/PAS.0b013e3181cd3aeb.
B-cell lymphomas with concurrent IGH-BCL2 and MYC rearrangements, also known as "double-hit" lymphomas (DHL), are rare neoplasms characterized by highly aggressive clinical behavior, complex karyotypes, and a spectrum of pathologic features overlapping with Burkitt lymphoma (BL), diffuse large B-cell lymphoma (DLBCL) and B-lymphoblastic lymphoma/leukemia (B-LBL). The clinical and pathologic spectrum of this rare entity, including comparison to other high-grade B-cell neoplasms, has not been well defined. We conducted a retrospective analysis of clinical and pathologic features of 20 cases of DHL seen at our institution during a 5-year period. In addition, we carried out case-control comparisons of DHL with BL and International Prognostic Index (IPI)-matched DLBCL. The 11 men and 9 women had a median age of 63.5 years (range 32 to 91). Six patients had a history of grade 1 to 2 follicular lymphoma; review of the prior biopsy specimens in 2 of 5 cases revealed blastoid morphology. Eighteen patients had Ann Arbor stage 3 or 4 disease and all had elevated serum lactate dehydrogenase (LDH) levels at presentation. Extranodal disease was present in 17/20 (85%), bone marrow involvement in 10/17 (59%) and central nervous system (CNS) disease in 5/11 (45%). Nineteen patients were treated with combination chemotherapy, of whom 18 received rituximab and 14 received CNS-directed therapy. Fourteen patients (70%) died within 8 months of diagnosis. Median overall survival in the DHL group (4.5 mo) was inferior to both BL (P=0.002) and IPI-matched DLBCL (P=0.04) control patients. Twelve DHL cases (60%) were classified as B-cell lymphoma, unclassifiable, with features intermediate between DLBCL and BL, 7 cases (35%) as DLBCL, not otherwise specified, and 1 case as B-LBL. Distinguishing features from BL included expression of Bcl2 (P<0.0001), Mum1/IRF4 (P=0.006), Ki-67 <95% (P<0.0001), and absence of EBV-EBER (P=0.006). DHL commonly contained the t(8;22) rather than the t(8;14) seen in most BL controls (P=0.001), and exhibited a higher number of chromosomal aberrations (P=0.0009). DHL is a high-grade B-cell neoplasm with a poor prognosis, resistance to multiagent chemotherapy, and clinical and pathologic features distinct from other high-grade B-cell neoplasms. Familiarity with the morphologic and immunophenotypic spectrum of DHL is important in directing testing to detect concurrent IGH-BCL2 and MYC rearrangements when a karyotype is unavailable. The aggressive clinical behavior and combination of genetic abnormalities seen in these cases may warrant categorization as a separate entity in future classifications and call for novel therapeutic approaches.

55Prognostic value of and genetic alterations for diffuse large B-cell lymphoma patients treated with R-CHOP.PubMed

Yan Qin, Haizhu Chen, Peng Liu, et al.
Cancer Biol Med. 2021 Nov 23;19(6):893-909. doi: 10.20892/j.issn.2095-3941.2021.0193.
OBJECTIVE: Limited data about the prognostic significance of mutations and copy number variations in diffuse large B-cell lymphoma (DLBCL) are available. This study aimed to comprehensively describe genetic alterations in DLBCL patients, and examine correlation of , and other genetic alterations with outcomes in patients treated with R-CHOP. METHODS: Probe capture-based high-resolution sequencing was performed on 191 patients diagnosed with DLBCL. MYC, BCL2, and BCL6 protein expressions were detected by immunohistochemistry. RESULTS: The presence of alterations significantly correlated with poor progression-free survival (PFS) (5-year PFS: 13.7% 40.8%; = 0.003) and overall survival (OS) (5-year OS: 34.0% 70.9%; = 0.036). Importantly, patients who harbored gain/amplifications () also had a remarkably inferior 5-year PFS (11.1% 38.3%; < 0.001) and OS (22.1% 69.6%; = 0.009). In contrast, neither mutations nor translocations were significantly prognostic for survival. Multivariable analyses showed that the presence of alterations, especially , mutations, and International Prognostic Index (IPI) were significantly associated with inferior PFS and OS. Novel prognostic models for OS were constructed based on 3 risk factors, including alterations (Model 1) or (Model 2), mutations, and IPI, to stratify patients into 4 risk groups with different survival outcomes. CONCLUSIONS: This study showed that DLBCL patients treated with R-CHOP, alterations, especially and mutations were significantly associated with inferior outcomes, which were independent of the IPI. The novel prognostic models we proposed predicted outcomes for DLBCL patients treated with R-CHOP, but further validation of the prognostic models is still warranted.

56Impact of autograft-absolute lymphocyte count on survival in double/triple hit lymphomas post-autologous stem cell transplantation.PubMed

Luis F Porrata, David J Inwards, Stephen M Ansell, et al.
Leuk Lymphoma. 2022 Oct;63(10):2436-2443. doi: 10.1080/10428194.2022.2064988. Epub 2022 Apr 28.
The autograft absolute lymphocyte count (A-ALC) ≥0.5 × 10 cells/kg is a survival prognostic factor for lymphoma patients undergoing autologous peripheral blood hematopoietic stem cell transplantation (APBHSCT). However, the A-ALC has not be tested as prognostic factor against double hit/triple hit lymphomas (DHL/THL). Thus, we set up to investigate if A-ALC is a prognostic factor for overall survival (OS) and progression-free survival (PFS) for DHL/THL post-APBHSCT. From January 2012 until December 2020, we identified 77 DHL/THL patients treated with APBHSCT. All patients required to have the diagnosis of DHL/THL by FISH for rearrangements of , and With a median follow-up of 20.4 months (range, 0.4-94.5 months), DHL/THL patients infused with A-ALC ≥0.5 x 10 cells/kg experienced superior OS (HR = 0.251, 95%CI 0.117-0.539, < 0.0004) and PFS (HR = 0.347, 95%CI 0.160-0.753, < 0.007). Multivariate analysis showed that A-ALC was an independent predictor for OS (HR =0.119, 95%CI 0.030-0.473, < 0.003) and PFS (HR = 0.400, 95%CI 0.189-0.850, < 0.02). Our study showed that A-ALC is a prognostic factor for survival in DHL/THL. Our current practice for lymphoma patients is to collect enough stem cell but also A-ALC to improve clinical outcomes post-APBHSCT.

57XPO1 expression worsens the prognosis of unfavorable DLBCL that can be effectively targeted by selinexor in the absence of mutant p53.PubMed

Manman Deng, Mingzhi Zhang, Zijun Y Xu-Monette, et al.
J Hematol Oncol. 2020 Nov 4;13(1):148. doi: 10.1186/s13045-020-00982-3.
The XPO1 inhibitor selinexor was recently approved in relapsed/refractory DLBCL patients but only demonstrated modest anti-DLBCL efficacy, prompting us to investigate the prognostic effect of XPO1 in DLBCL patients and the rational combination therapies in high-risk DLBCL. High XPO1 expression (XPO1) showed significant adverse prognostic impact in 544 studied DLBCL patients, especially in those with BCL2 overexpression. Therapeutic study in 30 DLBCL cell lines with various molecular and genetic background found robust cytotoxicity of selinexor, especially in cells with BCL2-rearranged (BCL2-R) DLBCL or high-grade B-cell lymphoma with MYC/BCL2 double-hit (HGBCL-DH). However, expression of mutant (Mut) p53 significantly reduced the cytotoxicity of selinexor in overall cell lines and the BCL2-R and HGBCL-DH subsets, consistent with the favorable impact of XPO1 observed in Mut-p53-expressing patients. The therapeutic effect of selinexor in HGBCL-DH cells was significantly enhanced when combined with a BET inhibitor INCB057643, overcoming the drug resistance in Mut-p53-expressing cells. Collectively, these data suggest that XPO1 worsens the survival of DLBCL patients with unfavorable prognostic factors such as BCL2 overexpression and double-hit, in line with the higher efficacy of selinexor demonstrated in BCL2-R DLBCL and HGBCL-DH cell lines. Expression of Mut-p53 confers resistance to selinexor treatment, which can be overcome by combined INCB057643 treatment in HGBCL-DH cells. This study provides insight into the XPO1 significance and selinexor efficacy in DLBCL, important for developing combination therapy for relapsed/refractory DLBCL and HGBCL-DH.

58Clinical implications of circulating tumor DNA in predicting the outcome of diffuse large B cell lymphoma patients receiving first-line therapy.PubMed

Miaomiao Li, Lan Mi, Chunyang Wang, et al.
BMC Med. 2022 Oct 25;20(1):369. doi: 10.1186/s12916-022-02562-3.
BACKGROUND: Circulating tumor DNA (ctDNA) has been proven to be a promising tumor-specific biomarker in solid tumors, but its clinical utility in risk stratification and early prediction of relapse for diffuse large B cell lymphoma (DLBCL) has not been well explored. METHODS: Here, using a lymphoma-specific sequencing panel, we assessed the prognostic and predictive utilities of ctDNA measurements before, during, and after first-line therapy in 73 Chinese DLBCL patients. RESULTS: The pretreatment ctDNA level serving as an independent prognostic factor for both progression-free survival (PFS, adjusted HR 2.47; p = 0.004) and overall survival (OS, adjusted HR 2.49; p = 0.011) was confirmed in our cohort. Furthermore, the patients classified as molecular responders who presented a larger decrease in ctDNA levels after the initial two treatment cycles had more favorable PFS (unreached vs. 6.25 months; HR 5.348; p = 0.0015) and OS (unreached vs. 25.87; HR 4.0; p = 0.028) than non-responders. In addition, interim ctDNA clearance may be an alternative noninvasive method of positron emission tomography and computed tomography (PET-CT) for predicting better PFS (HR 3.65; p = 0.0033) and OS (HR 3.536; p = 0.016). We also demonstrated that posttreatment ctDNA was a sensitive indicator for detecting minimal residual disease (MRD) in patients with a high risk of recurrence (HR 6.471; p = 0.014), who were otherwise claimed to achieve radiographic CR (complete remission). CONCLUSIONS: CtDNA is a promising noninvasive tool for prognosis prediction, response assessment, and early relapse prediction of first-line treatment in DLBCL patients.

59Prognostic relevance of immunoglobulin heavy chain rearrangement and immunoglobulin kappa light chain rearrangement in patients with diffuse large B cell lymphoma.PubMed

Jie Wang, Sha Zhao, Ting Niu, et al.
Oncologist. 2025 Mar 10;30(3). doi: 10.1093/oncolo/oyaf016.
PURPOSES: Evidence has demonstrated that monitoring of the variable, diversity, and joining gene segments (VDJ) rearrangement of immunoglobulin (Ig) gene in the circulating tumor DNA (ctDNA) is highly valuable in predicting the prognosis of patients with diffuse large B cell lymphoma (DLBCL). In this study, we investigated the role of both Ig heavy chain (IGH) and Ig kappa light chain (IGK) gene rearrangements detected in ctDNA samples in predicting DLBCL progression. METHODS: Next-generation sequencing (NGS) was used to identify the dominant V(D)J clonotypic rearrangement in tissue samples of 33 DLBCL patients. Minimal residual disease (MRD) was monitored at the interim and end of the treatment, as well as the follow-up time by tracking the dominant V(D)J clonotypic rearrangement (defined as the "NGS MRD" method) in the peripheral blood (PB) ctDNA samples. The nomogram was established to predict the 12-month and 24-month progression-free survival (PFS) probability. RESULTS: Prior to treatment, the dominant clones identified in the tissue samples could be retrieved in tissue-matched PB of 26 (78.8%, 26/33) patients. The addition of IGK clones to IGH clones increased the MRD detection rate from 42.9% to 58.0% in the total series. NGS MRD and imaging scans showed poor concordance at the interim of treatment (Kappa = 0.24) and the follow-up time (Kappa = 0.28), and fair concordance at the end of treatment (Kappa = 0.46). However, we confirmed that the interim NGS MRD monitoring demonstrated improved prognostic performance compared to imaging scans, and both NGS MRD monitoring and imaging scans served as valuable prognostic factors for PFS at the end of treatment. Notably, NGS MRD monitoring predicted disease relapse in 3 patients prior to imaging scans. Furthermore, we found that both the faster IGH and IGK clone clearance rates were associated with favorable prognosis. The nomogram model identified IGH and IGK clone clearance rates, together with the interim NGS MRD result were the important predictors of 12-month and 24-month progression of DLBCL. CONCLUSIONS: MRD monitoring via NGS of Ig for both IGH and IGK is a promising noninvasive tool for prognosis prediction and early relapse prediction of DLBCL patients.

60Emerging Strategies in Treating Double Hit Lymphomas.PubMed

Chadi Nabhan, Anthony R Mato
Clin Lymphoma Myeloma Leuk. 2017 Sep;17(9):563-568. doi: 10.1016/j.clml.2017.06.017. Epub 2017 Jun 21.
Double hit lymphomas (DHLs) are a new category in the World Health Organization newest classification for lymphoid malignancies. DHL encompasses various histologies of lymphomas where the MYC oncogene and either BCL2 or BCL6 oncogenes are present concomitantly. Several observational studies and retrospective series have demonstrated that patients with DHL carry a poor prognosis and respond less and for a shorter duration to standard R-CHOP (rituximab, cyclophosphamide, vincristine, adriamycin, and prednisone). These studies have also proposed that dose intensification (with Burkitt-like regimens such as DA-EPOCH-R [dose-adjusted rituximab, etoposide, vincristine, Adriamycin, cyclophosphamide, and prednisone]) might offer patients with DHL better outcomes and improved prognosis. In this timely review, we discuss incidence of DHL, testing implications of MYC translocation, current treatment strategies, and future directions. Understanding this entity and its therapeutic consequences is essential to improve patients' outcomes.

61Outcomes of Patients With Double-Hit Lymphoma Who Achieve First Complete Remission.PubMed

Daniel J Landsburg, Marissa K Falkiewicz, Joseph Maly, et al.
J Clin Oncol. 2017 Jul 10;35(20):2260-2267. doi: 10.1200/JCO.2017.72.2157. Epub 2017 May 5.
Purpose Patients with double-hit lymphoma (DHL) rarely achieve long-term survival following disease relapse. Some patients with DHL undergo consolidative autologous stem-cell transplantation (autoSCT) to reduce the risk of relapse, although the benefit of this treatment strategy is unclear. Methods Patients with DHL who achieved first complete remission following completion of front-line therapy with either rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) or intensive front-line therapy, and deemed fit for autoSCT, were included. A landmark analysis was performed, with time zero defined as 3 months after completion of front-line therapy. Patients who experienced relapse before or who were not followed until that time were excluded. Results Relapse-free survival (RFS) and overall survival (OS) rates at 3 years were 80% and 87%, respectively, for all patients (n = 159). Three-year RFS and OS rates did not differ significantly for autoSCT (n = 62) versus non-autoSCT patients (n = 97), but 3-year RFS was inferior in patients who received R-CHOP compared with intensive therapy (56% v 88%; P = .002). Three-year RFS and OS did not differ significantly for patients in the R-CHOP or intensive therapy cohorts when analyzed by receipt of autoSCT. The median OS following relapse was 8.6 months. Conclusion In the largest reported series, to our knowledge, of patients with DHL to achieve first complete remission, consolidative autoSCT was not associated with improved 3-year RFS or OS. In addition, patients treated with R-CHOP experienced inferior 3-year RFS compared with those who received intensive front-line therapy. When considered in conjunction with reports of patients with newly diagnosed DHL, which demonstrate lower rates of disease response to R-CHOP compared with intensive front-line therapy, our findings further support the use of intensive front-line therapy for this patient population.

62A multi-center analysis of the impact of DA-EPOCH-R dose-adjustment on clinical outcomes of patients with double/triple-hit lymphoma.PubMed

Matthew J Cortese, Wei Wei, Sebastian Cerdeña, et al.
Leuk Lymphoma. 2023 Jan;64(1):107-118. doi: 10.1080/10428194.2022.2140281. Epub 2022 Nov 2.
Patients with double- and triple-hit lymphomas (DHL/THL) have inferior outcomes with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP), and higher-intensity regimens such as dose-adjusted (DA)-EPOCH-R are standard. Dose-intensification of DA-EPOCH-R is guided by hematologic toxicity, without conclusive benefit for DHL/THL patients. To determine if cumulative doses of DA-EPOCH-R or compliance with dose adjustment impacts survival, we retrospectively evaluated detailed clinical data from 109 adult (age ≥18 years) patients with DHL/THL treated with ≥4 cycles of induction DA-EPOCH-R from 2014 to 2019 at six centers. A comprehensive multivariate analysis was performed. Survival outcomes for the entire cohort were comparable to historical estimates for DHL/THL treated with this regimen (median follow-up 27.9 months). Overall survival (OS) and progression-free survival (PFS) were not significantly associated with cumulative chemotherapy dose, dose escalation, or compliance with dose adjustment. Heterogeneous dosing practices were observed. Prospective investigation is warranted to evaluate the practice of dose adjustment of R-EPOCH for patients with DHL/THL.

63Prognostic analysis of CD5 expression in double-hit diffuse large B-cell lymphoma and effectiveness comparison in patients treated with dose-adjusted EPOCH plus rituximab/R-CHOP regimens.PubMed

Fangwen Zhang, Ling Li, Lei Zhang, et al.
Blood Lymphat Cancer. 2019 Aug 19;9:33-43. doi: 10.2147/BLCTT.S216292. eCollection 2019.
OBJECTIVES: To compare the efficacy of rituximab, dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide and doxorubicin (DA-EPOCH-R) with traditional rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) regimens in CD5+ double-hit lymphoma (DHL) and to evaluate prognostic factors. METHODS: We retrospectively studied 139 patients with newly diagnosed DHL/THL diffuse large B-cell lymphoma (including 20 cases CD5+ and 119 cases CD5-), 87 cases were MYC/BCL2 DHL, 30 cases were MYC/BCL6 DHL, 22 cases were THL. MYC, BCL2 and BCL6 rearrangements were examined by fluorescence in-situ hybridization. CD5 is detected by immunohistochemistry (IHC). RESULTS: The objective response rate (ORR) difference between CD5+ and CD5- was significant (80.0% vs 63.8%, =0.003). The median follow-up time was 18 months (range: 4-39 months). Progression-free survival (PFS) of CD5+ group was significantly worse than that of CD5- (28.1% vs 59.0%, =0.028), while no significant difference was observed in overall survival (OS) (32.1% vs 59.9%, =0.057). Compared with the two regimens, the 2-year survival rate of DA-EPOCH-R group was significantly superior than that of R-CHOP (63.6% vs 45.4%, =0.034 for PFS; 67.4% vs 47.8%, =0.038 for OS). Besides, CD5+ patients receiving DA-EPOCH-R had survival benefits compared with R-CHOP in PFS (85.7% vs 23.0%, =0.029), but there was no statistical difference in OS (87.7% vs 34.4.0%, =0.064). However, in DA-EPOCH-R protocol, there was no significant difference between CD5+ DHL (MYC/BCl2 and MYC/BCL6) and triple-hit lymphoma (=0.776 for PFS; =0.728 for OS). Multivariate analysis showed that CD5+ treatment regimen and disease stage were independent prognostic factors. CONCLUSION: Our retrospective study shows that CD5+ has a poorer prognosis than CD5- patients. Based on its improved lifetime and good tolerance on CD5+ patients, which is expected to become the first-line treatment for high-risk DLBCL types based on more clinical research.

64Phase 1 study of lenalidomide plus dose-adjusted EPOCH-R in patients with aggressive B-cell lymphomas with deregulated MYC and BCL2.PubMed

James K Godfrey, Chadi Nabhan, Theodore Karrison, et al.
Cancer. 2019 Jun 1;125(11):1830-1836. doi: 10.1002/cncr.31877. Epub 2019 Feb 1.
BACKGROUND: Dual translocation of MYC and BCL2 or the dual overexpression of these proteins in patients with aggressive B-cell lymphomas (termed double-hit lymphoma [DHL] and double-expressor lymphoma [DEL], respectively) have poor outcomes after chemoimmunotherapy with the combination of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP). Retrospective reports have suggested improved outcomes with dose-intensified regimens. In the current study, the authors conducted a phase 1 study to evaluate the feasibility, toxicity, and preliminary efficacy of adding lenalidomide to dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin with rituximab (DA-EPOCH-R) in patients with DHL and DEL. METHODS: The primary objective of the current study was to determine the maximum tolerated dose of lenalidomide in combination with DA-EPOCH-R. A standard 3+3 design was used with lenalidomide administered on days 1 to 14 of each 21-day cycle (dose levels of 10 mg, 15 mg, and 20 mg). Patients attaining a complete response after 6 cycles of induction therapy proceeded to maintenance lenalidomide (10 mg daily for 14 days every 21 days) for 12 additional cycles. RESULTS: A total of 15 patients were enrolled, 10 of whom had DEL and 5 of whom had DHL. Two patients experienced dose-limiting toxicities at a lenalidomide dose of 20 mg, consisting of grade 4 sepsis. The maximum tolerated dose of lenalidomide was determined to be 15 mg. The most common nonhematologic grade ≥3 adverse events included thromboembolism (4 patients; 27%) and hypokalemia (2 patients; 13%) (toxicities were graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events [version 4.0]). The preliminary efficacy of the regimen was encouraging, especially in the DEL cohort, in which all 10 patients achieved durable and complete metabolic responses with a median follow-up of 24 months. CONCLUSIONS: The combination of lenalidomide with DA-EPOCH-R appears to be safe and feasible in patients with DHL and DEL. These encouraging results have prompted an ongoing phase 2 multicenter study.

65Molecular High-Grade B-Cell Lymphoma: Defining a Poor-Risk Group That Requires Different Approaches to Therapy.PubMed

Chulin Sha, Sharon Barrans, Francesco Cucco, et al.
J Clin Oncol. 2019 Jan 20;37(3):202-212. doi: 10.1200/JCO.18.01314. Epub 2018 Dec 3.
PURPOSE: Biologic heterogeneity is a feature of diffuse large B-cell lymphoma (DLBCL), and the existence of a subgroup with poor prognosis and phenotypic proximity to Burkitt lymphoma is well known. Conventional cytogenetics identifies some patients with rearrangements of MYC and BCL2 and/or BCL6 (double-hit lymphomas) who are increasingly treated with more intensive chemotherapy, but a more biologically coherent and clinically useful definition of this group is required. PATIENTS AND METHODS: We defined a molecular high-grade (MHG) group by applying a gene expression-based classifier to 928 patients with DLBCL from a clinical trial that investigated the addition of bortezomib to standard rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP) therapy. The prognostic significance of MHG was compared with existing biomarkers. We performed targeted sequencing of 70 genes in 400 patients and explored molecular pathology using gene expression signature databases. Findings were validated in an independent data set. RESULTS: The MHG group comprised 83 patients (9%), with 75 in the cell-of-origin germinal center B-cell-like group. MYC rearranged and double-hit groups were strongly over-represented in MHG but comprised only one half of the total. Gene expression analysis revealed a proliferative phenotype with a relationship to centroblasts. Progression-free survival rate at 36 months after R-CHOP in the MHG group was 37% (95% CI, 24% to 55%) compared with 72% (95% CI, 68% to 77%) for others, and an analysis of treatment effects suggested a possible positive effect of bortezomib. Double-hit lymphomas lacking the MHG signature showed no evidence of worse outcome than other germinal center B-cell-like cases. CONCLUSION: MHG defines a biologically coherent high-grade B-cell lymphoma group with distinct molecular features and clinical outcomes that effectively doubles the size of the poor-prognosis, double-hit group. Patients with MHG may benefit from intensified chemotherapy or novel targeted therapies.

66The Improving Outcomes in Relapsed-Refractory Diffuse Large B Cell Lymphoma: The Role of CAR T-Cell Therapy.PubMed

Vibor Milunović, Dora Dragčević, Martina Bogeljić Patekar, et al.
Curr Treat Options Oncol. 2025 Jun;26(6):445-464. doi: 10.1007/s11864-025-01305-9. Epub 2025 Apr 28.
Diffuse large B cell lymphoma, not otherwise specified (DLBCL-NOS) is the most common aggressive lymphoma and can be cured with CHOP-R immunochemotherapy in 60% of cases. The second-line therapy includes salvage regimens followed by autologous stem cell transplantation (ASCT), which offers a cure to a minority of patients due to limitations in efficacy and eligibility. These data present the unmet need in the field, and this review article focuses on how second-generation chimeric antigen receptor T (CAR T) cell therapy targeting CD19 antigen may improve the outcomes with relapsed/refractory DLBCL. In heavily pretreated patients, who have dismal outcomes with conventional therapy, all three approved products-tisangenlecleucel (tisa-cel), axicabtagene ciloleucel (axi-cel), and lisocabtagene maraleucel (liso-cel) have shown durable, unprecedented complete responses with the potential for cure. When compared to salvage regimens and ASCT as the standard of care, axi-cel and liso-cel, unlike tisa-cel, have demonstrated superiority in long-term control. In ASCT-ineligible r/r DLBCL, liso-cel has shown a favourable benefit-risk ratio. Regarding safety, two adverse events of interest have emerged: cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome, both of which are manageable. Real-world evidence reflects the results of pivotal trials while favouring axi-cel in heavily pretreated patients, albeit with higher toxicity. The main barrier to the implementation of this treatment modality is the cost associated with the process of CAR T therapy, along with complications and reimbursement issues. However, the barriers can be overcome, and CAR T therapy has the potential to become the standard of care in relapsed/refractory DLBCL. Furthermore, with advances in the scientific engineering of CAR products and the understanding of novel treatment modalities currently being tested in clinical trials, we believe that targeted cellular therapy will become the future of relapsed/refractory DLBCL treatment.

67Double hit & double expressor lymphomas: a multicenter analysis of survival outcomes with CD19-directed CAR T-cell therapy.PubMed

Reem Karmali, Geoffrey Shouse, Pallawi Torka, et al.
Blood Cancer J. 2025 Mar 26;15(1):43. doi: 10.1038/s41408-025-01250-8.
Double-hit (DHL) and double expressor (DEL) DLBCL have poor prognosis with standard therapy but CART may overcome this poor prognostic impact. In this multicenter retrospective study, we sought to confirm this observation by evaluating survival outcomes among patients with relapsed/refractory DHL and DEL treated with CART and evaluate outcomes of relapse post-CART. A total of 408 adult patients with relapsed/refractory DLBCL from 13 academic centers were included based on the availability of DHL and DEL. All 408 patients were included in the DHL (n = 80) vs non-DHL (n = 328) analysis, while 333 patients were included in the analysis of DHL (n = 80) vs DEL (n = 74) vs non (n = 179). On MVA, there were no differences for PFS for DHL vs non-DHL (HR 0.8, 95%CI 0.5-1.3, p = 0.35) or DHL vs DEL vs other (three-way p value, p = 0.5). Response rates and toxicities were similar among groups. Patients with DEL had the highest relapse rates post-CART, while DHL had the worst overall survival after CART relapse. In sum, our data support the notion that CART cell therapy can overcome the poor prognostic impact of DHL and DEL DLBCL in the relapsed/refractory setting. Additionally, patients with DHL that relapse after CART have a very poor prognosis.

68Patterns of Utilization and Outcomes of Autologous Stem Cell Transplantation and Chimeric Antigen Receptor T-Cell Therapy in Relapsed or Refractory Diffuse Large B-cell Lymphomas with MYC and BCL2 and/or BCL6 Rearrangements.PubMed

Sean Patrick Bliven, Lauren Shea, Susan Bal, et al.
Clin Lymphoma Myeloma Leuk. 2022 Nov;22(11):825-834. doi: 10.1016/j.clml.2022.06.011. Epub 2022 Jun 29.
BACKGROUND: Patients with Diffuse Large Bcell Lymphoma (DLBCL) with MYC and BCL2 and/or BCL6 gene rearrangements [double-hit lymphoma/triple-hit lymphoma (DHL/THL)] have poor prognosis in the relapsed/refractory setting. METHODS: We utilized a real-world deidentified database of DLBCL patients and report patterns of therapy utilization in relapsed/refractory DLBCL. We used log-rank test to compare real-world overall survival (rwOS) among DHL and non-DHL subgroups for CAR Tcell therapy or ASCT respectively, stratified for prior lines of therapy. RESULTS: Of all 7,877 patients with DLBCL, 367 patients had DHL while 6113 had non-DHL. Second line chemotherapy was administered to 147 DHL patients and 1517 non-DHL. 1393 were excluded, including 934 with unknown DHL/THL status. Approximately 47% received salvage intent chemotherapy in the DHL subgroup, of which 19% patients eventually received ASCT, while 34% received salvage intent chemotherapy in the non-DHL/THL group with 32% receiving ASCT. DHL/THL status negatively influenced median rwOS for patients who underwent ASCT in the second-line while it was associated with numerically inferior but without statistically significant rwOS among patients that underwent CAR Tcell therapy on multivariable analysis. CONCLUSION: rwOS of relapsed DHL/THL is inferior to non-DHL/THL. Fewer patients with DHL/THL were able to proceed with ASCT after salvage chemotherapy compared to non-DHL/THL. ASCT as second-line therapy for relapsed DHL/THL had worse rwOS than for non-DHL/THL, consistent with the natural history of DHL/THL. This difference was not seen for CAR Tcell therapy, which combined with promising results from clinical trials, suggests a greater role for CAR T-cell therapy in relapsed/refractory DHL.

69CD19 CAR-T treatment shows limited efficacy in r/r DLBCL with double expression and TP53 alterations.PubMed

Bin Xue, Yifan Liu, Jie Zhou, et al.
Cytotherapy. 2024 Dec;26(12):1465-1471. doi: 10.1016/j.jcyt.2024.07.011. Epub 2024 Jul 25.
OBJECT: Autologous CD19 chimeric antigen receptor T-cell therapy (CAR-T) significantly modifies the natural course of chemorefractory diffuse large B-cell lymphoma (DLBCL). However, 25% to 50% of patients with relapsed/refractory DLBCL still do not achieve remission. Therefore, investigating new molecular prognostic indicators that affect the effectiveness of CAR-T for DLBCL and developing novel combination therapies are crucial. METHODS: Data from 73 DLBCL patients who received CD19 CAR-T (Axi-cel or Relma-cel) were retrospectively collected from Shanghai Tongji Hospital of Tongji University, The Second Affiliated Hospital Zhejiang University School of Medicine, and The Affiliated People's Hospital of Ningbo University. Prior to CD19 CAR-T-cell transfusions, the patients received fludarabine and cyclophosphamide chemotherapy regimen. RESULTS: Our study revealed that relapsed/refractory diffuse large B-cell lymphoma (r/r DLBCL) patients with both Double-expression (MYC > 40% and BCL2 > 50%) and TP53 alterations tend to have a poorer clinical prognosis after CAR-T therapy, even when CAR-T therapy is used in combination with other therapies. However, CAR-T therapy was found to be effective in patients with only TP53 alterations or DE status, suggesting that their prognosis is in line with that of patients without TP53 alterations or DE status. CONCLUSIONS: Our study suggests that r/r DLBCL patients with both DE status and TP53 alterations treated with CAR-T therapy are more likely to have a poorer clinical prognosis. However, CAR-T therapy has the potential to improve the prognosis of patients with only TP53 alterations or DE status to be similar to that of patients without these abnormalities.

70Transdifferentiation of diffuse large B-cell lymphoma to a poorly differentiated neoplasm following CAR T-cell therapy.PubMed

Nandan Padmanabha, Matthew J Weinstock, Sean Xu, et al.
J Hematop. 2024 Sep;17(3):149-153. doi: 10.1007/s12308-024-00592-9. Epub 2024 Jun 15.
Chimeric antigen receptor T-cell (CAR-T) therapy is a recent advancement in precision medicine with promising results for patients with relapsed or refractory B-cell malignancies. However, rare post-therapy morphologic, immunophenotypic, and genomic alterations can occur. This study is to present a case of a patient with diffuse large B-cell lymphoma (DLBCL) who underwent anti-CD19 CAR-T therapy with disease in the uterus that showed transdifferentiation to a poorly differentiated malignant neoplasm that failed to express any lineage specific markers. In immunohistochemistry, fluorescence in situ hybridization (FISH) and targeted next-generation sequencing (NGS) were utilized to fully characterize the diagnostic DLBCL sample in comparison to the poorly differentiated neoplasm of the uterus. Analysis of the diagnostic DLBCL and the poorly differentiated neoplasm demonstrated evidence of a clonal relationship as well as revealing acquisition of mutations associated with CAR-T resistance. Furthermore, downregulation of B-cell associated antigens was observed, underscoring a mechanistic link to CAR-T evasion as well as demonstrating diagnostic confusion. This case illustrates the utility of employing multiple diagnostic modalities in elucidating a pathologic link between a B-cell lymphoma and poorly differentiated neoplasm following targeted therapy.

71Single-Agent Mosunetuzumab Shows Durable Complete Responses in Patients With Relapsed or Refractory B-Cell Lymphomas: Phase I Dose-Escalation Study.PubMed

Lihua E Budde, Sarit Assouline, Laurie H Sehn, et al.
J Clin Oncol. 2022 Feb 10;40(5):481-491. doi: 10.1200/JCO.21.00931. Epub 2021 Dec 16.
PURPOSE: Mosunetuzumab is a bispecific antibody targeting CD20 and CD3 that redirects T cells to engage and eliminate malignant B cells and is being developed for relapsed or refractory (R/R) B-cell non-Hodgkin lymphomas (B-NHLs). METHODS: This first-in-human trial (ClinicalTrials.gov identifier: NCT02500407) evaluated the safety and tolerability and efficacy of mosunetuzumab in patients with R/R B-NHL and established the recommended phase II dose. Data from dose escalation are presented. Single-agent mosunetuzumab was administered intravenously in 3-week cycles, at full dose in cycle 1 day 1 (group A) or with ascending (step-up) doses during cycle 1 on days 1, 8, and 15 (group B), for eight or 17 cycles on the basis of tumor response. RESULTS: Two hundred thirty patients were enrolled. Doses up to 2.8 mg and 60 mg were assessed in groups A and B, respectively; maximum tolerated dose was not exceeded. In group B (n = 197), common adverse events (≥ 20% of patients) were neutropenia (28.4%), cytokine release syndrome (27.4%), hypophosphatemia (23.4%), fatigue (22.8%), and diarrhea (21.8%). Cytokine release syndrome was mostly low-grade (grade ≥ 3: 1.0%) and mainly confined to cycle 1. Across the doses investigated (group B), best overall response rates were 34.9% and 66.2% in patients with aggressive and indolent B-NHL, respectively, and complete response rates were 19.4% and 48.5%. Among patients with a complete response, the median duration of response was 22.8 months (95% CI, 7.6 to not estimable) and 20.4 (95% CI, 16 to not estimable) in patients with aggressive and indolent B-NHL, respectively. CONCLUSION: Mosunetuzumab, administered with step-up dosing, has a manageable safety profile and induces durable complete responses in R/R B-NHL. The expansion stage of the study is ongoing at the dose level of 1/2/60/60/30 mg selected for further study.

72Safety and efficacy of mosunetuzumab, a bispecific antibody, in patients with relapsed or refractory follicular lymphoma: a single-arm, multicentre, phase 2 study.PubMed

Lihua E Budde, Laurie H Sehn, Matthew Matasar, et al.
Lancet Oncol. 2022 Aug;23(8):1055-1065. doi: 10.1016/S1470-2045(22)00335-7. Epub 2022 Jul 5.
BACKGROUND: Mosunetuzumab is a CD20 × CD3 T-cell-engaging bispecific monoclonal antibody that redirects T cells to eliminate malignant B cells. In a phase 1 study, mosunetuzumab was well tolerated and active in patients with relapsed or refractory B-cell lymphoma. We, therefore, aimed to evaluate the safety and anti-tumour activity of fixed-duration mosunetuzumab in patients with relapsed or refractory follicular lymphoma who had received two or more previous therapies. METHODS: We conducted a single-arm, multicentre, phase 2 study at 49 centres in seven countries (Australia, Canada, Germany, South Korea, Spain, UK, and USA). All patients were aged 18 years or older with histologically confirmed follicular lymphoma (grade 1-3a) and an Eastern Cooperative Oncology Group performance status of 0-1. Patients had disease that was relapsed or refractory to two or more previous lines of treatment, including an anti-CD20 therapy and an alkylating agent. Intravenous mosunetuzumab was administered in 21-day cycles with cycle 1 step-up dosing: 1 mg on cycle 1 day 1, 2 mg on cycle 1 day 8, 60 mg on cycle 1 day 15 and cycle 2 day 1, and 30 mg on day 1 of cycle 3 and onwards. Patients with a complete response by investigator assessment using the International Harmonisation Project criteria completed treatment after cycle 8, whereas patients with a partial response or stable disease continued treatment for up to 17 cycles. The primary endpoint was independent review committee-assessed complete response rate (as best response) in all enrolled patients; the primary efficacy analysis compared the observed IRC-assessed complete response rate with a 14% historical control complete response rate in a similar patient population receiving the pan class I PI3K inhibitor copanlisib. Safety was assessed in all enrolled patients. This study is registered with ClinicalTrials.gov, number NCT02500407, and is ongoing. FINDINGS: Between May 2, 2019, and Sept 25, 2020, we enrolled 90 patients. As of the data cutoff date (Aug 27, 2021), the median follow-up was 18·3 months (IQR 13·8-23·3). According to independent review committee assessment, a complete response was recorded in 54 patients (60·0% [95% CI 49·1-70·2]). The observed complete response rate was significantly higher than the historical control complete response rate with copanlisib of 14% (p<0·0001), thereby meeting the primary study endpoint. Cytokine release syndrome was the most common adverse event (40 [44%] of 90 patients) and was predominantly grade 1 (23 [26%] of 90) and grade 2 (15 [17%]), and primarily confined to cycle 1. The most common grade 3-4 adverse events were neutropenia or neutrophil count decreased (24 [27%] of 90 patients), hypophosphataemia (15 [17%]), hyperglycaemia (seven [8%]), and anaemia (seven [8%]). Serious adverse events occurred in 42 (47%) of 90 patients. No treatment-related grade 5 (ie, fatal) adverse event occurred. INTERPRETATION: Fixed-duration mosunetuzumab has a favourable safety profile and induces high rates of complete remissions, allowing potential administration as an outpatient regimen, in patients with relapsed or refractory follicular lymphoma and two or more previous therapies. FUNDING: F Hoffmann-La Roche and Genentech.

73Radiation Therapy as a Bridging Strategy for CAR T Cell Therapy With Axicabtagene Ciloleucel in Diffuse Large B-Cell Lymphoma.PubMed

Austin J Sim, Michael D Jain, Nicholas B Figura, et al.
Int J Radiat Oncol Biol Phys. 2019 Dec 1;105(5):1012-1021. doi: 10.1016/j.ijrobp.2019.05.065. Epub 2019 Jun 5.
PURPOSE: Axicabtagene ciloleucel (axi-cel) is a CD19-directed chimeric antigen receptor (CAR) T-cell therapy for relapsed or refractory diffuse large B-cell lymphoma. Bridging therapy may be required for lymphoma control during the manufacturing interval between collection of autologous T cells and final CAR T product administration. The optimal bridging therapy is not known and patients are often chemorefractory. We present a case series of patients receiving radiation as a bridge to axi-cel. METHODS AND MATERIALS: Between December 2017 and October 2018, 12 patients were intended to receive bridging radiation before axi-cel. The group was characterized by highly aggressive disease including 6 of 12 with "double hit" lymphoma and 6 of 12 with disease ≥10 cm in diameter. All patients received 2 to 4 Gy/fraction to a median dose of 20 Gy (range, 6-36.5 Gy). Half of patients received either 30 Gy in 10 fractions or 20 Gy in 5 fractions. Seven patients received concurrent chemotherapy. Eleven patients underwent axi-cel infusion and one did not. Median follow-up was 3.3 months (range, 1.1-12.0 months). RESULTS: No significant toxicities were identified during bridging radiation, and no patient experienced in-field progression of disease before axi-cel infusion. One patient experienced abdominal pain, which resolved after dose reduction. Two patients had out-of-field progression of disease during the bridging period. After axi-cel infusion, 3 of 11 patients (27%) experienced severe cytokine release syndrome or neurotoxicity. At 30 days, the objective response rate was 81.8% (11 of 12 evaluable; 1 stable disease, 1 out-of-field progression), with complete response in 27% (3 of 11). At last follow-up, the best objective response rate was 81.8%, with a complete response attained in 45% (5 of 11). Lymphocyte counts decreased slightly in 10 of 12 patients during radiation (median, 0.25 k/uL). CONCLUSIONS: Radiation (with or without concurrent chemotherapy) can be safely administered as a bridge to axi-cel in high-risk lymphoma. Caution should be taken if irradiation is started before apheresis, and lymphocyte counts should be monitored closely throughout. Future investigation is warranted to optimize the use of bridging radiation before CAR T therapy.

74Bevacizumab synergises with the BCL 2 inhibitor venetoclax to effectively treat B-cell non-Hodgkin's lymphoma.PubMed

Li Wang, Shaoyong Peng, Weipeng Sun, et al.
Eur J Haematol. 2019 Sep;103(3):234-246. doi: 10.1111/ejh.13279. Epub 2019 Jul 3.
OBJECTIVES: Our present study has shown a potential role for VEGF-A-mediated autocrine signalling to promote survival and proliferation of SU-DHL-6 cells, but the cells could not undergo apoptosis but rather decrease proliferation after bevacizumab treatment. Therefore, we would like to further study the antitumour efficacy of venetoclax (BCL2 inhibitor) in combination with bevacizumab in B-cell NHL. METHODS: The human cytokine antibody array, RT-qPCR, Western blot, ELISA, apoptosis assay and xenografted mouse model et al were used. RESULTS: We described a unique phenomenon that SU-DHL-6 cells showed cell density-dependent survival and growth. Then, we suggested the expression of VEGF-A was positively correlated with the cell density using a human cytokine antibody array and indicated an important role of VEGF-A in the survival and proliferation of SU-DHL-6 cells. Additionally, xenografted SU-DHL-6 cells formed tumours in mice that grew in response to VEGF stimulation. GEO data set also suggested that high VEGF-A expression reflected poor prognosis. The combination therapy with bevacizumab and navitoclax could significantly induce of cell death in vitro and reduce the tumour size and weight with well tolerated in vivo. CONCLUSIONS: Our findings propose a novel combined strategy in which bevacizumab synergises with the BCL2 inhibitor venetoclax that is effective against B-cell NHL.

75Pirtobrutinib in relapsed or refractory B-cell malignancies (BRUIN): a phase 1/2 study.PubMed

Anthony R Mato, Nirav N Shah, Wojciech Jurczak, et al.
Lancet. 2021 Mar 6;397(10277):892-901. doi: 10.1016/S0140-6736(21)00224-5.
BACKGROUND: Covalent Bruton's tyrosine kinase (BTK) inhibitors are efficacious in multiple B-cell malignancies, but patients discontinue these agents due to resistance and intolerance. We evaluated the safety and efficacy of pirtobrutinib (working name; formerly known as LOXO-305), a highly selective, reversible BTK inhibitor, in these patients. METHODS: Patients with previously treated B-cell malignancies were enrolled in a first-in-human, multicentre, open-label, phase 1/2 trial of the BTK inhibitor pirtobrutinib. The primary endpoint was the maximum tolerated dose (phase 1) and overall response rate (ORR; phase 2). This trial is registered with ClinicalTrials.gov, NCT03740529. FINDINGS: 323 patients were treated with pirtobrutinib across seven dose levels (25 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, and 300 mg once per day) with linear dose-proportional exposures. No dose-limiting toxicities were observed and the maximum tolerated dose was not reached. The recommended phase 2 dose was 200 mg daily. Adverse events in at least 10% of 323 patients were fatigue (65 [20%]), diarrhoea (55 [17%]), and contusion (42 [13%]). The most common adverse event of grade 3 or higher was neutropenia (32 [10%]). There was no correlation between pirtobrutinib exposure and the frequency of grade 3 treatment-related adverse events. Grade 3 atrial fibrillation or flutter was not observed, and grade 3 haemorrhage was observed in one patient in the setting of mechanical trauma. Five (1%) patients discontinued treatment due to a treatment-related adverse event. In 121 efficacy evaluable patients with chronic lymphocytic leukaemia (CLL) or small lymphocytic lymphoma (SLL) treated with a previous covalent BTK inhibitor (median previous lines of treatment 4), the ORR with pirtobrutinib was 62% (95% CI 53-71). The ORR was similar in CLL patients with previous covalent BTK inhibitor resistance (53 [67%] of 79), covalent BTK inhibitor intolerance (22 [52%] of 42), BTK C481-mutant (17 [71%] of 24) and BTK wild-type (43 [66%] of 65) disease. In 52 efficacy evaluable patients with mantle cell lymphoma (MCL) previously treated with covalent BTK inhibitors, the ORR was 52% (95% CI 38-66). Of 117 patients with CLL, SLL, or MCL who responded, all but eight remain progression-free to date. INTERPRETATION: Pirtobrutinib was safe and active in multiple B-cell malignancies, including patients previously treated with covalent BTK inhibitors. Pirtobrutinib might address a growing unmet need for alternative therapies for these patients. FUNDING: Loxo Oncology.

76Pirtobrutinib monotherapy in Bruton tyrosine kinase inhibitor-intolerant patients with B-cell malignancies: results of the phase I/II BRUIN trial.PubMed

Nirav N Shah, Michael Wang, Lindsey E Roeker, et al.
Haematologica. 2025 Jan 1;110(1):92-102. doi: 10.3324/haematol.2024.285754.
Bruton tyrosine kinase inhibitors (BTKi) have transformed the treatment of B-cell malignancies, but intolerance has often led to their discontinuation. The phase I/II BRUIN study evaluated pirtobrutinib, a highly selective non-covalent (reversible) BTKi, in patients with relapsed / refractory B-cell malignancies (clinicaltrials.gov 03740529). Pirtobrutinib was investigated in 127 patients with intolerance to at least one prior BTKi therapy in the absence of progressive disease. The most common adverse event (AE) leading to BTKi discontinuation was cardiac disorders (N=40, 31.5%), specifically atrial fibrillation (N=30, 23.6%). The median follow-up was 17.4 months and the median time on pirtobrutinib was 15.3 months. The most common reasons for pirtobrutinib discontinuation were progressive disease (26.8%), AE (10.2%) or death (5.5%). The most frequent treatment-emergent AE were fatigue (39.4%) and neutropenia (37.0%). Among patients who discontinued a prior BTKi for a cardiac issue, 75% had no recurrence of their cardiac AE. No patient discontinued pirtobrutinib for the same AE that led to discontinuation of the prior BTKi. In 78 chronic lymphocytic / small lymphocytic lymphoma (CLL/SLL) and 21 mantle cell lymphoma (MCL) patients intolerant to prior BTKi, overall response rate to pirtobrutinib was 76.9% and 81.0%, respectively. Median progression-free survival for CLL/SLL was 28.4 months but was not estimable for MCL. These results suggest that pirtobrutinib was safe, well-tolerated, and an efficacious option in patients with prior BTKi-intolerance.

77Pirtobrutinib, a highly selective, non-covalent (reversible) BTK inhibitor in patients with B-cell malignancies: analysis of the Richter transformation subgroup from the multicentre, open-label, phase 1/2 BRUIN study.PubMed

William G Wierda, Nirav N Shah, Chan Y Cheah, et al.
Lancet Haematol. 2024 Sep;11(9):e682-e692. doi: 10.1016/S2352-3026(24)00172-8. Epub 2024 Jul 18.
BACKGROUND: Richter transformation usually presents as an aggressive diffuse large B-cell lymphoma, occurs in up to 10% of patients with chronic lymphocytic leukaemia, has no approved therapies, and is associated with a poor prognosis. Pirtobrutinib has shown promising efficacy and tolerability in patients with relapsed or refractory B-cell malignancies, including those who progress on covalent Bruton tyrosine kinase (BTK) inhibitors. This study aims to report the safety and activity of pirtobrutinib monotherapy in a subgroup of patients with Richter transformation from the multicentre, open-label, phase 1/2 BRUIN study. METHODS: This analysis included adult patients (aged ≥18 years) with histologically confirmed Richter transformation, an Eastern Cooperative Oncology Group performance status score of 0-2, and no limit of previous therapies, with patients receiving first-line treatment added in a protocol amendment (version 9.0, Dec 15, 2021). Pirtobrutinib 200 mg was administered orally once a day in 28-day cycles. The primary endpoint of phase 1 of the BRUIN trial as a whole, which has been previously reported, was to establish the recommended phase 2 dose for pirtobrutinib monotherapy and the phase 2 primary endpoint was overall response rate. Safety and activity were measured in all patients who received at least one dose of pirtobrutinib monotherapy. This BRUIN phase 1/2 trial was registered with ClinicalTrials.gov and is closed to enrolment (NCT03740529). FINDINGS: Between Dec 26, 2019, and July 22, 2022, 82 patients were enrolled, of whom five were enrolled during phase 1 and 77 during phase 2. All but one patient received a starting dose of 200 mg pirtobrutinib once a day as the recommended phase 2 dose. The remaining patient received 150 mg pirtobrutinib once a day, which was not escalated to 200 mg. The median age of patients was 67 years (IQR 59-72). 55 (67%) of 82 patients were male and 27 (33%) were female. Most patients were White (65 [79%] of 82). 74 (90%) of 82 patients received at least one previous Richter transformation-directed therapy. Most patients (61 [74%] of 82) had received previous covalent BTK inhibitor therapy for chronic lymphocytic leukaemia or Richter transformation. The overall response rate was 50·0% (95% CI 38·7-61·3). 11 (13%) of 82 patients had a complete response and 30 (37%) of 82 patients had a partial response. Eight patients with ongoing response electively discontinued pirtobrutinib to undergo stem-cell transplantation. The most common grade 3 or worse adverse event was neutropenia (n=19). There were no treatment-related deaths. INTERPRETATION: Pirtobrutinib shows promising safety and activity among patients with Richter transformation, most of whom received previous Richter transformation-directed therapy, including covalent BTK inhibitors. These data suggest that further investigation is warranted of pirtobrutinib as a treatment option for patients with relapsed or refractory Richter transformation after treatment with a covalent BTK inhibitor. FUNDING: Loxo Oncology.

78Bruton Tyrosine Kinase Inhibitors in B-Cell Malignancies: Their Use and Differential Features.PubMed

Matt Shirley
Target Oncol. 2022 Jan;17(1):69-84. doi: 10.1007/s11523-021-00857-8. Epub 2021 Dec 14.
Starting with the first-in-class agent ibrutinib, the development of Bruton tyrosine kinase (BTK) inhibitors has led to dramatic improvements in the management of B-cell malignancies. Subsequently, more-highly selective second-generation BTK inhibitors (including acalabrutinib, zanubrutinib, tirabrutinib and orelabrutinib) have been developed, primarily with an aim to reduce off-target toxicities. More recently, third-generation agents including the non-covalent BTK inhibitors pirtobrutinib and nemtabrutinib have entered later-stage clinical development. BTK inhibitors have shown strong activity in a range of B-cell malignancies, including chronic lymphocytic leukaemia/small lymphocytic lymphoma, mantle cell lymphoma, Waldenström's macroglobulinaemia and marginal zone lymphoma. The agents have acceptable tolerability, with adverse events generally being manageable with dosage modification. This review article summarises the evidence supporting the role of BTK inhibitors in the management of B-cell malignancies, including highlighting some differential features between agents.

79Loncastuximab tesirine in relapsed or refractory diffuse large B-cell lymphoma (LOTIS-2): a multicentre, open-label, single-arm, phase 2 trial.PubMed

Paolo F Caimi, Weiyun Ai, Juan Pablo Alderuccio, et al.
Lancet Oncol. 2021 Jun;22(6):790-800. doi: 10.1016/S1470-2045(21)00139-X. Epub 2021 May 11.
BACKGROUND: Patients with relapsed or refractory diffuse large B-cell lymphoma (DLBCL) who do not respond to or who have progressive disease after salvage therapies have a poor prognosis. Loncastuximab tesirine is a CD19-directed antibody-drug conjugate with encouraging phase 1 single-agent antitumour activity and acceptable safety in non-Hodgkin lymphoma. We aimed to evaluate the antitumour activity and safety of loncastuximab tesirine in patients with relapsed or refractory DLBCL. METHODS: We did a multicentre (28 hospital sites in the USA, UK, Italy, and Switzerland), open-label, single-arm, phase 2 trial (LOTIS-2) in patients aged 18 years or older with relapsed or refractory DLBCL after two or more multiagent systemic treatments, who had measurable disease and Eastern Cooperative Oncology Group performance status 0-2. Eligible patients received loncastuximab tesirine intravenously on day 1 of each 21-day cycle, at 150 μg/kg for two cycles, then 75 μg/kg thereafter, for up to 1 year or until disease relapse or progression, unacceptable toxicity, death, major protocol deviation, pregnancy, or patient, investigator, or sponsor decision. The primary endpoint was overall response rate assessed by central review. Primary antitumour activity and safety analyses were done in the as-treated population (patients who received at least one dose of loncastuximab tesirine), when all responding patients had at least 6 months of follow-up after initial documented response. Enrolment is complete. This trial is registered with ClinicalTrials.gov, NCT03589469. FINDINGS: Between Aug 1, 2018, and Sept 24, 2019, 184 patients were assessed for eligibility and 145 (79%) were enrolled and received at least one dose of loncastuximab tesirine, including patients with high-risk characteristics for poor prognosis, such as double-hit, triple-hit, transformed, or primary refractory DLBCL. 70 of 145 patients had complete or partial response (overall response rate 48·3% [95% CI 39·9-56·7]); 35 had complete response and 35 had partial response. The most common grade 3 or higher treatment-emergent adverse events were neutropenia (37 [26%] of 145 patients), thrombocytopenia (26 [18%]), and increased gamma-glutamyltransferase (24 [17%]). Serious adverse events were reported in 57 (39%) of 145 patients. Treatment-emergent adverse events with a fatal outcome occurred in eight (6%) of 145 patients; none were considered related to loncastuximab tesirine. INTERPRETATION: Loncastuximab tesirine has substantial single-agent antitumour activity and produces durable responses with an acceptable safety profile, potentially offering a new therapeutic option for heavily pretreated patients with relapsed or refractory DLBCL. FUNDING: ADC Therapeutics.

80Loncastuximab tesirine in relapsed/refractory diffuse large B-cell lymphoma: long-term efficacy and safety from the phase II LOTIS-2 study.PubMed

Paolo F Caimi, Weiyun Z Ai, Juan Pablo Alderuccio, et al.
Haematologica. 2024 Apr 1;109(4):1184-1193. doi: 10.3324/haematol.2023.283459.
Therapies that demonstrate durable, long-term responses with manageable safety and tolerability are needed for patients with relapsed/refractory diffuse large B-cell lymphoma (R/R DLBCL). Loncastuximab tesirine (loncastuximab tesirine-lpyl [Lonca]), an anti-CD19 antibody conjugated to a potent pyrrolobenzodiazepine dimer, demonstrated single-agent antitumor activity in the pivotal phase II LOTIS-2 study in heavily pretreated patients with R/R DLBCL. Here we present updated efficacy and safety analyses from LOTIS-2, performed for all patients and in subsets of patients with a complete response (CR), including patients with CR who were event-free (no progressive disease or death) for ≥1 year and ≥2 years from cycle 1, day 1 of treatment. Lonca was administered every 3 weeks (0.15 mg/kg for 2 cycles; 0.075 mg/kg for subsequent cycles). As of the final data cutoff (September 15, 2022; median follow-up: 7.8 months [range, 0.3-42.6]), 70 of 145 (48.3%) patients achieved an overall response. Thirty-six (24.8%) patients achieved CR, of which 16 (44%) and 11 (31%) were event-free for ≥1 year and ≥2 years, respectively. In the all-treated population, the median overall survival was 9.5 months; the median progression-free survival was 4.9 months. Among patients with CR, median overall survival and progression-free survival were not reached, with 24-month overall and progression-free survival rates of 68.2% (95% CI: 50.0-81.0) and 72.5% (95% CI: 48.2-86.8), respectively. No new safety concerns were detected. With additional follow-up, Lonca continued to demonstrate durable, long-term responses with manageable safety and tolerability in patients with CR (clinicaltrials gov. Identifier: NCT03589469).

81ASTCT Clinical Practice Recommendations for Transplantation and Cellular Therapies in Diffuse Large B Cell Lymphoma.PubMed

Narendranath Epperla, Ambuj Kumar, Syed A Abutalib, et al.
Transplant Cell Ther. 2023 Sep;29(9):548-555. doi: 10.1016/j.jtct.2023.06.012. Epub 2023 Jul 5.
Autologous hematopoietic cell transplantation (auto-HCT) has long been the standard approach for patients with relapsed/refractory (R/R) chemosensitive diffuse large B cell lymphoma (DLBCL). However, the advent of chimeric antigen receptor (CAR) T cell therapy has caused a paradigm shift in the management of R/R DLBCL patients, especially with the recent approval of CD19-directed CAR-T therapy in the second-line setting in high-risk groups (primary refractory and early relapse [≤12 months]). Consensus on the contemporary role, optimal timing, and sequencing of HCT and cellular therapies in DLBCL is lacking; therefore, the American Society of Transplantation and Cellular Therapy (ASTCT) Committee on Practice Guidelines undertook this project to formulate consensus recommendations to address this unmet need. The RAND-modified Delphi method was used to generate 20 consensus statements with a few key statements as follows: (1) in the first-line setting, there is no role for auto-HCT consolidation for patients achieving complete remission (CR) following R-CHOP (rituximab, cyclophosphamide, adriamycin, vincristine, and prednisone) or similar therapy in non-double-hit/triple-hit cases (DHL/THL) and in DHL/THL cases receiving intensive induction therapies, but auto-HCT may be considered in eligible patients receiving R-CHOP or similar therapies in DHL/THL cases; (2) auto-HCT consolidation with thiotepa-based conditioning is standard of care for eligible patients with primary central nervous system lymphoma achieving CR with first-line therapy; and (3) in the primary refractory and early relapse setting, the preferred option is CAR-T therapy, whereas in late relapse (>12 months), consolidation with auto-HCT is recommended for patients achieving chemosensitivity to salvage therapy (complete or partial response), and CAR-T therapy is recommended for those not achieving remission. These clinical practice recommendations will serve as a tool to guide clinicians managing patients with newly diagnosed and R/R DLBCL.

82Rituximab and ICE as second-line therapy before autologous stem cell transplantation for relapsed or primary refractory diffuse large B-cell lymphoma.PubMed

Tarun Kewalramani, Andrew D Zelenetz, Stephen D Nimer, et al.
Blood. 2004 May 15;103(10):3684-8. doi: 10.1182/blood-2003-11-3911. Epub 2004 Jan 22.
Patients with relapsed or primary refractory diffuse large B-cell lymphoma (DLBCL) who achieve complete response (CR) before autologous stem cell transplantation (ASCT) generally have better outcomes than those who achieve only partial response (PR). We investigated whether adding rituximab to the ifosfamide-carboplatin-etoposide (ICE) chemotherapy regimen (RICE) could increase the CR rate of patients with DLBCL under consideration for ASCT. Thirty-six eligible patients were treated with RICE, and 34 received all 3 planned cycles. The CR rate was 53%, significantly better than the 27% CR rate (P =.01) achieved among 147 similar consecutive historical control patients with DLBCL treated with ICE; the PR rate was 25%. Febrile neutropenia was the most frequent grade 3 or 4 nonhematologic toxicity; it occurred in 7.5% of delivered cycles. No patient had RICE-related toxicity that precluded ASCT. The median number of CD34(+) cells per kilogram mobilized was 6.3 x 10(6). Progression-free survival rates of patients who underwent transplantation after RICE were marginally better than those of 95 consecutive historical control patients who underwent transplantation after ICE (54% vs 43% at 2 years; P =.25). RICE appears to induce very high CR rates in patients with relapsed and refractory DLBCL; however, further studies are necessary to determine whether this treatment regimen will improve outcomes after ASCT.

83High-grade B-cell lymphomas: Double hit and non-double hit.PubMed

Lianqun Qiu, L Jeffrey Medeiros, Shaoying Li
Hum Pathol. 2025 Feb;156:105700. doi: 10.1016/j.humpath.2024.105700. Epub 2024 Nov 26.
The classification of high-grade B-cell lymphoma (HGBL) has continuously evolved over past decades. These neoplasms, as currently defined, represent about 2% of all non-Hodgkin lymphomas and patients with these neoplasms are often refractory or relapsed following standard therapy. The 5th edition of the World Health Organization classification of hematologic neoplasms (WHO-HAEM5) has refined the classification of HGBL and recognizes two types: (1) Diffuse large B-cell lymphoma (DLBCL)/HGBL with MYC and BCL2 rearrangements, with or without BCL6 rearrangements; and (2) HGBL, not otherwise specified (HGBL-NOS). WHO-HAEM5 excluded DLBCL/HGBL with concurrent MYC and BCL6 rearrangements from this category and reclassified them into DLBCL or HGBL-NOS categories respectively based on morphology. The International Consensus Classification (ICC) takes a slightly different approach. In addition to recognizing the two WHO-HAEM5 categories, they recognize HGBL with concurrent MYC and BCL6 rearrangements as a provisional entity. In this review, we provide an update of HGBL and its subgroups, focusing on their clinicopathologic features, diagnosis, molecular genetic features, and pathogenesis. Our diagnostic approach and caveats for differential diagnosis are also discussed with an emphasis on the differential diagnosis with B lymphoblastic leukemia/lymphoma.

84Large B-Cell Lymphomas in the 5th Edition of the WHO-Classification of Haematolymphoid Neoplasms-Updated Classification and New Concepts.PubMed

Katrin S Kurz, Michaela Ott, Sabrina Kalmbach, et al.
Cancers (Basel). 2023 Apr 13;15(8):2285. doi: 10.3390/cancers15082285.
The family/class of the large B-cell lymphomas (LBCL) in the 5th edition of the World Health Organization (WHO) classification of haematolymphoid tumors (WHO-HAEM5) features only a few major changes as compared to the 4th edition. In most entities, there are only subtle changes, many of them only representing some minor modifications in diagnostic terms. Major changes have been made in the diffuse large B-cell lymphomas (DLBCL)/high-grade B-cell lymphomas (HGBL) associated with and and/or rearrangements. This category now consists of and rearranged cases exclusively, while the double hit lymphomas now constitute genetic subtypes of DLBCL, not otherwise specified (NOS) or of HGBL, NOS. Other major changes are the conceptual merger of lymphomas arising in immune-privileged sites and the description of LBCL arising in the setting of immune dysregulation/deficiency. In addition, novel findings concerning underlying biological mechanisms in the pathogenesis of the different entities are provided.

85[Classification of aggressive B-cell lymphomas : News and open questions].PubMed

Andreas Rosenwald, Thomas Menter, Stefan Dirnhofer
Pathologie (Heidelb). 2023 May;44(3):166-172. doi: 10.1007/s00292-023-01187-4. Epub 2023 Mar 14.
The 5th edition of the WHO classification (WHO-HAEM5) and the International Consensus Classification (ICC) show a broad consensus in the categorization of aggressive, large B‑cell lymphomas with expected minor impact only on the daily diagnostic routine. The changes compared to the 2017 revised WHO-HAEM4R are moderate and include updated names of entities, sharpened diagnostic criteria, and upgrades from provisional to definite entities. The definition of the most common aggressive B‑cell lymphoma, diffuse large B‑cell lymphoma (DLBCL), not otherwise specified (NOS), remains unchanged, and both classifications strongly encourage subtyping into germinal center B‑like (GCB) or the activated B‑like (ABC or non-GCB) DLBCL. DLBCL, NOS, should be separated from other large B‑cell lymphomas including large B‑cell lymphoma with IRF4 rearrangement (upgraded to a definite entity in both classifications) and large-cell/high-grade B‑cell lymphomas with 11q aberration. Aggressive B‑cell lymphomas with MYC and BCL2 rearrangements form a molecularly distinct group and are listed as definite entities in both classifications. This is in contrast to the more heterogeneous group of aggressive B‑cell lymphomas with MYC and BCL6 rearrangements that are recognized as a provisional entity in the ICC, while they fall into the DLBCL, NOS, or the HGBL, NOS, groups in the WHO-HAEM5.

86How I diagnose high-grade B-cell lymphoma.PubMed

Erika M Moore, Sarah E Gibson
Am J Clin Pathol. 2025 Apr 19;163(4):487-500. doi: 10.1093/ajcp/aqae158.
OBJECTIVES: High-grade B-cell lymphoma (HGBL), introduced in the 2016 World Health Organization (WHO) revised fourth edition classification, included cases defined by MYC and BCL2 and/or BCL6 rearrangements or by high-grade morphology. Diagnostic criteria and nomenclature for these lymphomas were refined in the 2022 WHO fifth edition (WHO-5) classification and International Consensus Classification (ICC). This review describes our approach to the diagnosis of HGBL. METHODS: Two cases are presented illustrating how we diagnose HGBL, including 1 case harboring MYC and BCL6 rearrangements and a second showing TdT expression in an HGBL with MYC and BCL2 rearrangements. The ways in which these cases are distinguished from other lymphomas with high-grade features and the appropriate nomenclature using WHO-5 and ICC classifications are emphasized. RESULTS: An HGBL diagnosis requires integration of morphology, immunophenotype, and genetics and exclusion of other lymphomas with high-grade morphology, including Burkitt lymphoma, B-lymphoblastic leukemia/lymphoma (B-LBL/ALL), and blastoid mantle cell lymphoma. A diagnosis of HGBL/large B-cell lymphoma with 11q aberration should also be considered in certain patient populations. CONCLUSIONS: High-grade B-cell lymphomas are subclassified based on morphologic and genetic features. There are differences in the nomenclature and definition of these lymphomas in the WHO-5 and ICC classifications. Distinguishing HGBLs from other mature B-cell lymphomas and B-LBL/ALL is critical so that patients receive appropriate treatment.

87DLBCL-associated NOTCH2 mutations escape ubiquitin-dependent degradation and promote chemoresistance.PubMed

Nan Zhou, Jaewoo Choi, Grant Grothusen, et al.
Blood. 2023 Sep 14;142(11):973-988. doi: 10.1182/blood.2022018752.
Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma. Up to 40% of patients with DLBCL display refractory disease or relapse after standard chemotherapy treatment (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone [R-CHOP]), leading to significant morbidity and mortality. The molecular mechanisms of chemoresistance in DLBCL remain incompletely understood. Using a cullin-really interesting new gene (RING) ligase-based CRISPR-Cas9 library, we identify that inactivation of the E3 ubiquitin ligase KLHL6 promotes DLBCL chemoresistance. Furthermore, proteomic approaches helped identify KLHL6 as a novel master regulator of plasma membrane-associated NOTCH2 via proteasome-dependent degradation. In CHOP-resistant DLBCL tumors, mutations of NOTCH2 result in a protein that escapes the mechanism of ubiquitin-dependent proteolysis, leading to protein stabilization and activation of the oncogenic RAS signaling pathway. Targeting CHOP-resistant DLBCL tumors with the phase 3 clinical trial molecules nirogacestat, a selective γ-secretase inhibitor, and ipatasertib, a pan-AKT inhibitor, synergistically promotes DLBCL destruction. These findings establish the rationale for therapeutic strategies aimed at targeting the oncogenic pathway activated in KLHL6- or NOTCH2-mutated DLBCL.

88Use of CAR-Transduced Natural Killer Cells in CD19-Positive Lymphoid Tumors.PubMed

Enli Liu, David Marin, Pinaki Banerjee, et al.
N Engl J Med. 2020 Feb 6;382(6):545-553. doi: 10.1056/NEJMoa1910607.
BACKGROUND: Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy has shown remarkable clinical efficacy in B-cell cancers. However, CAR T cells can induce substantial toxic effects, and the manufacture of the cells is complex. Natural killer (NK) cells that have been modified to express an anti-CD19 CAR have the potential to overcome these limitations. METHODS: In this phase 1 and 2 trial, we administered HLA-mismatched anti-CD19 CAR-NK cells derived from cord blood to 11 patients with relapsed or refractory CD19-positive cancers (non-Hodgkin's lymphoma or chronic lymphocytic leukemia [CLL]). NK cells were transduced with a retroviral vector expressing genes that encode anti-CD19 CAR, interleukin-15, and inducible caspase 9 as a safety switch. The cells were expanded ex vivo and administered in a single infusion at one of three doses (1×10, 1×10, or 1×10 CAR-NK cells per kilogram of body weight) after lymphodepleting chemotherapy. RESULTS: The administration of CAR-NK cells was not associated with the development of cytokine release syndrome, neurotoxicity, or graft-versus-host disease, and there was no increase in the levels of inflammatory cytokines, including interleukin-6, over baseline. The maximum tolerated dose was not reached. Of the 11 patients who were treated, 8 (73%) had a response; of these patients, 7 (4 with lymphoma and 3 with CLL) had a complete remission, and 1 had remission of the Richter's transformation component but had persistent CLL. Responses were rapid and seen within 30 days after infusion at all dose levels. The infused CAR-NK cells expanded and persisted at low levels for at least 12 months. CONCLUSIONS: Among 11 patients with relapsed or refractory CD19-positive cancers, a majority had a response to treatment with CAR-NK cells without the development of major toxic effects. (Funded by the M.D. Anderson Cancer Center CLL and Lymphoma Moonshot and the National Institutes of Health; ClinicalTrials.gov number, NCT03056339.).

89CAR-T: What Is Next?PubMed

Yi-Ju Chen, Bams Abila, Yasser Mostafa Kamel
Cancers (Basel). 2023 Jan 21;15(3):663. doi: 10.3390/cancers15030663.
The year 2017 was marked by the Food and Drug Administration (FDA) approval of the first two chimeric antigen receptor-T (CAR-T) therapies. The approved indications were for the treatment of relapsed or refractory diffuse large B-cell lymphoma (DLBCL) and for the treatment of patients up to 25 years of age with acute lymphoblastic leukemia (ALL) that is refractory or in a second or later relapse. Since then, extensive research activities have been ongoing globally on different hematologic and solid tumors to assess the safety and efficacy of CAR-T therapy for these diseases. Limitations to CAR-T therapy became apparent from, e.g., the relapse in up to 60% of patients and certain side effects such as cytokine release syndrome (CRS). This led to extensive clinical activities aimed at overcoming these obstacles, so that the use of CAR-T therapy can be expanded. Attempts to improve on efficacy and safety include changing the CAR-T administration schedule, combining it with chemotherapy, and the development of next-generation CAR-T therapies, e.g., through the use of CAR-natural killer (CAR-NK) and CAR macrophages (CAR-Ms). This review will focus on new CAR-T treatment strategies in hematologic malignancies, clinical trials aimed at improving efficacy and addressing side effects, the challenges that CAR-T therapy faces in solid tumors, and the ongoing research aimed at overcoming these challenges.

90Safety, efficacy and determinants of response of allogeneic CD19-specific CAR-NK cells in CD19 B cell tumors: a phase 1/2 trial.PubMed

David Marin, Ye Li, Rafet Basar, et al.
Nat Med. 2024 Mar;30(3):772-784. doi: 10.1038/s41591-023-02785-8. Epub 2024 Jan 18.
There is a pressing need for allogeneic chimeric antigen receptor (CAR)-immune cell therapies that are safe, effective and affordable. We conducted a phase 1/2 trial of cord blood-derived natural killer (NK) cells expressing anti-CD19 chimeric antigen receptor and interleukin-15 (CAR19/IL-15) in 37 patients with CD19 B cell malignancies. The primary objectives were safety and efficacy, defined as day 30 overall response (OR). Secondary objectives included day 100 response, progression-free survival, overall survival and CAR19/IL-15 NK cell persistence. No notable toxicities such as cytokine release syndrome, neurotoxicity or graft-versus-host disease were observed. The day 30 and day 100 OR rates were 48.6% for both. The 1-year overall survival and progression-free survival were 68% and 32%, respectively. Patients who achieved OR had higher levels and longer persistence of CAR-NK cells. Receiving CAR-NK cells from a cord blood unit (CBU) with nucleated red blood cells ≤ 8 × 10 and a collection-to-cryopreservation time ≤ 24 h was the most significant predictor for superior outcome. NK cells from these optimal CBUs were highly functional and enriched in effector-related genes. In contrast, NK cells from suboptimal CBUs had upregulation of inflammation, hypoxia and cellular stress programs. Finally, using multiple mouse models, we confirmed the superior antitumor activity of CAR/IL-15 NK cells from optimal CBUs in vivo. These findings uncover new features of CAR-NK cell biology and underscore the importance of donor selection for allogeneic cell therapies. ClinicalTrials.gov identifier: NCT03056339 .

91Safety and feasibility of 4-1BB co-stimulated CD19-specific CAR-NK cell therapy in refractory/relapsed large B cell lymphoma: a phase 1 trial.PubMed

Wen Lei, Hui Liu, Wenhai Deng, et al.
Nat Cancer. 2025 May;6(5):786-800. doi: 10.1038/s43018-025-00940-3. Epub 2025 Apr 18.
Chimeric antigen receptor (CAR)-modified NK (CAR-NK) cells are candidates for next-generation cancer immunotherapies. Here we generated CD19-specific CAR-NK cells with 4-1BB and CD3ζ signaling endo-domains (CD19-BBz CAR-NK) by transduction of cord blood-derived NK cells using baboon envelope pseudotyped lentiviral vectors and demonstrated their antitumor activity in preclinical B cell lymphoma models in female mice. We next conducted a phase 1 dose-escalation trial involving repetitive administration of CAR-NK cells in 8 patients with relapsed/refractory large B cell lymphoma (NCT05472558). Primary end points were safety, maximum tolerated dose, and overall response rate. Secondary end points included duration of response, overall survival, and progression-free survival. No dose-limiting toxicities occurred, and the maximum tolerated dose was not reached. No cases of cytokine release syndrome, neurotoxicity, or graft-versus-host disease were observed. Results showed an overall response rate of 62.5% at day 30, with 4 patients (50%) achieving complete response. The median progression-free survival was 9.5 months, and the median overall survival was not reached. A post hoc exploratory single-cell RNA sequencing analysis revealed molecular features of CAR-NK cells associated with therapeutic efficacy and efficacy-related immune cell interaction networks. This study met the pre-specified end points. In conclusion, CD19-BBz CAR-NK cells were feasible and therapeutically safe, capable of inducing durable response in patients with B cell lymphoma.

92Dual effect of DLBCL-derived EXOs in lymphoma to improve DC vaccine efficacy in vitro while favor tumorgenesis in vivo.PubMed

Zhenzhen Chen, Liangshun You, Lei Wang, et al.
J Exp Clin Cancer Res. 2018 Aug 13;37(1):190. doi: 10.1186/s13046-018-0863-7.
BACKGROUND: Exosomes derived from tumor cells (TEXs) are involved in both immune suppression, angiogenesis, metastasis and anticancer stimulatory, but the biological characteristics and role of diffuse large B cell lymphoma (DLBCL)-derived exosomes have been less investigated. METHODS: Exosomes (EXOs) were isolated from OCI-LY3, SU-DHL-16, and Raji cells and biological characteristics of EXOs were investigated using electron microscopy, flow cytometry analysis, and Western blot analysis. The protein expression of EXOs was determined by an antibody array. Next, the communication between EXOs and lymphoma cell, stromal cell, dendritic cells (DCs), and T cells was evaluated. Finally, effect of DLBCL TEXs on tumor growth in vivo was investigated. RESULTS: We demonstrated that EXOs derived from DLBCL cell lines displayed malignancy molecules such as c-Myc, Bcl-2, Mcl-1, CD19, and CD20. There was a different protein expression pattern between DLBCL TEXs and Burkitt lymphoma TEXs. DLBCL TEXs were easily captured by DCs and lymphoma cells, and mainly acted as an immunosuppressive mediator, evidenced by induction of apoptosis and upregulation of PD-1 in T cells. Furthermore, the TEXs stimulated not only cell proliferation, migration of stromal cells but also angiogenesis. As a result, the TEXs promoted tumor growth in vivo. On other hand, DLBCL TEXs did not induce apoptosis of DCs. After pulsed with the TEXs, DCs could stimulate clonal expansion of T cells, increase the secretion of IL-6 and TNFα, and decrease the production of immunosuppressive cytokine IL-4 and IL-10. The T cells from tumor bearing mice immunized by TEX were shown to possess superior antilymphoma potency relative to immunization of tumor lysates. CONCLUSIONS: This study provides the framework for novel immunotherapies targeting TEXs in DLBCL.