Lee Hyunjoo, Haque Shabirul, Gupta Rashmi, Kolitz Jonathan E, Allen Steven L, Rai Kanti, Chiorazzi Nicholas, Mongini Patricia K A
The Feinstein Institutes for Medical Research, Northwell Health, Manhasset, NY 11030, USA.
Department of Medicine, Zucker School of Medicine at Hofstra/Northwell, Hempstead, NY 11549, USA.
Lymphatics. 2024 Jun;2(2):50-78. doi: 10.3390/lymphatics2020005. Epub 2024 Mar 28.
CLL B cells express elevated pro-survival BCL2, and its selective inhibitor, venetoclax, significantly reduces leukemic cell load, leading to clinical remission. Nonetheless, relapses occur. This study evaluates the hypothesis that progressively diminished BCL2 protein in cycling CLL cells within patient lymph node niches contributes to relapse. Using CFSE-labeled, purified CLL populations known to respond with vigorous cycling in d6 cultures stimulated with TLR9-activating ODN (oligodeoxynucleotide) + IL15, we show that BCL2 protein progressively declines during consecutive cell divisions. In contrast, MCL1 and survivin are maintained/slightly elevated during cycling. Delayed pulsing of quiescent and activated CLL cultures with selective inhibitors of BCL2 or survivin revealed selective targeting of noncycling and cycling populations, respectively, raising implications for therapy. To address the hypothesis that BCL2-repressive miRs (), encoded in Chr13, are mechanistically involved, we compared BCL2 protein levels within ODN + IL15-stimulated CLL cells, with/without del(13q), yielding results suggesting these miRs contribute to BCL2 reduction. In support, within ODN-primed CLL cells, an IL15-driven STAT5/PI-3K pathway (required for vigorous cycling) triggers elevated p53 TF protein known to directly activate the locus. Furthermore, IL15 signaling elicits the repression of mRNA within 24 h. Additional comparisons of del(13q)+ and del(13q)-/- cohorts for elevated p53 TF expression during cycling suggest that a documented -mediated negative feedback loop for p53 synthesis is active during cycling. Findings that robust CLL cycling associates with progressively decreasing BCL2 protein that directly correlates with decreasing venetoclax susceptibility, combined with past findings that these cycling cells have the greatest potential for activation-induced cytosine deaminase (AICDA)-driven mutations, suggest that venetoclax treatment should be accompanied by modalities that selectively target the cycling compartment without eliciting further mutations. The employment of survivin inhibitors might be such an approach.
慢性淋巴细胞白血病（CLL）B细胞表达升高的促生存蛋白BCL2，其选择性抑制剂维奈托克可显著降低白血病细胞负荷，从而导致临床缓解。尽管如此，仍会出现复发情况。本研究评估了这样一种假说：患者淋巴结微环境中循环的CLL细胞内BCL2蛋白逐渐减少是导致复发的原因。使用经羧基荧光素二乙酸琥珀酰亚胺酯（CFSE）标记的、纯化的CLL细胞群体，已知这些细胞在用Toll样受体9（TLR9）激活寡脱氧核苷酸（ODN）+白细胞介素15（IL15）刺激的第6天培养物中会剧烈循环，我们发现BCL2蛋白在连续细胞分裂过程中逐渐下降。相比之下，髓细胞白血病-1（MCL1）和生存素在循环过程中保持不变/略有升高。用BCL2或生存素的选择性抑制剂对静止和活化的CLL培养物进行延迟脉冲处理，分别揭示了对非循环和循环细胞群体的选择性靶向作用，这对治疗具有启示意义。为了验证位于13号染色体上编码的BCL2抑制性微小RNA（miR）在机制上是否参与其中这一假说，我们比较了在有或无13q缺失的情况下，经ODN + IL15刺激的CLL细胞内的BCL2蛋白水平，结果表明这些miR有助于BCL2的减少。作为支持，在经ODN预处理的CLL细胞中，由IL15驱动的信号转导和转录激活因子5（STAT5）/磷脂酰肌醇-3激酶（PI-3K）通路（剧烈循环所必需）会触发已知可直接激活该基因座的p53转录因子蛋白水平升高。此外，IL15信号在24小时内会引发该基因mRNA的抑制。对13q缺失阳性和13q缺失阴性队列在循环过程中p53转录因子表达升高情况的进一步比较表明，记录在案的p53合成的介导负反馈环在循环过程中是活跃的。有力的CLL循环与BCL2蛋白逐渐减少相关，而BCL2蛋白减少又与维奈托克敏感性降低直接相关，再结合过去的研究结果，即这些循环细胞具有由激活诱导的胞嘧啶脱氨酶（AICDA）驱动的突变的最大潜力，这表明维奈托克治疗应辅以选择性靶向循环细胞区室而不引发进一步突变的方法。使用生存素抑制剂可能就是这样一种方法。
