Dysthe Matthew, Navin Ishwar, van Leeuwen Dayenne, Pineda Josue, Baumgartner Corrine, Rooney Cliona M, Parihar Robin
Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas, USA.
Immunology and Microbiology, Baylor College of Medicine, Houston, Texas, USA.
J Immunother Cancer. 2025 Jul 23;13(7):e010672. doi: 10.1136/jitc-2024-010672.
BACKGROUND: Adoptive transfer of chimeric antigen receptor (CAR)-expressing natural killer (NK) cells has demonstrated success against hematological malignancies. Efficacy against solid tumors has been limited by poor NK cell survival and function in the suppressive tumor microenvironment (TME). To enhance efficacy against solid tumors, stimulatory cytokines have been incorporated into CAR-NK cell therapeutic approaches. However, current cytokine strategies have limitations, including systemic toxicities, exogenous dependencies, and unwanted TME bystander effects. Here, we aimed to overcome these limitations by modifying CAR-NK cells to express a constitutively active interleukin (IL)-7 receptor, termed C7R, capable of providing intrinsic CAR-NK cell activation that does not rely on or produce exogenous signals nor activate bystander cells. METHODS: We examined persistence, antitumor function, and transcriptional profiles of CAR-NK cells coexpressing C7R in a novel tumor immune microenvironment (TiME) co-culture system and against hematologic and solid tumor xenografts in vivo. RESULTS: Peripheral blood NK cells expressing a CAR directed against the solid tumor antigen GD2 and modified with C7R demonstrated enhanced tumor killing and persistence in vitro compared with CAR-NK cells without cytokine support and similar functions to CAR-NK cells supplemented with recombinant IL-15. C7R.CAR-NK cells exhibited enhanced survival and proliferation within neuroblastoma TiME xenografts in vivo but produced poor long-term tumor control compared with CAR-NK cells supplemented with IL-15. Similar results were seen using C7R-expressing CD19.CAR-NK cells against CD19+leukemia xenografts. Gene expression analysis revealed that chronic signaling via C7R induced a transcriptional signature consistent with intratumor stressed NK cells with blunted effector function. We identified gene candidates associated with chronic cytokine-stressed NK cells that could be targeted to reduce CAR-NK cell stress within the solid TME. CONCLUSION: C7R promoted CAR-NK cell survival in hostile TMEs independent of exogenous signals but resulted in poor antitumor function in vivo. Our data reveals the detrimental role of continuous IL-7 signaling in CAR-NK cells and provides insights into proper application of cytokine signals when attempting to enhance CAR-NK cell antitumor activity.
背景：表达嵌合抗原受体（CAR）的自然杀伤（NK）细胞的过继性转移已证明对血液系统恶性肿瘤有效。在抑制性肿瘤微环境（TME）中，NK细胞存活和功能不佳限制了其对实体瘤的疗效。为提高对实体瘤的疗效，刺激细胞因子已被纳入CAR-NK细胞治疗方法中。然而，目前的细胞因子策略存在局限性，包括全身毒性、对外源性物质的依赖性以及不必要的TME旁观者效应。在此，我们旨在通过改造CAR-NK细胞以表达一种组成型活性白细胞介素（IL）-7受体（称为C7R）来克服这些局限性，该受体能够提供不依赖于外源性信号、不产生外源性信号且不激活旁观者细胞的内在CAR-NK细胞激活。 方法：我们在一种新型肿瘤免疫微环境（TiME）共培养系统中以及在体内针对血液学和实体瘤异种移植模型，检测了共表达C7R的CAR-NK细胞的持久性、抗肿瘤功能和转录谱。 结果：与没有细胞因子支持的CAR-NK细胞相比，表达针对实体瘤抗原GD2的CAR并经C7R改造的外周血NK细胞在体外表现出增强的肿瘤杀伤能力和持久性，并且与补充重组IL-15的CAR-NK细胞具有相似的功能。C7R.CAR-NK细胞在体内神经母细胞瘤TiME异种移植模型中表现出增强的存活和增殖能力，但与补充IL-15的CAR-NK细胞相比，长期肿瘤控制效果较差。使用表达C7R的CD19.CAR-NK细胞针对CD19+白血病异种移植模型也观察到了类似结果。基因表达分析显示，通过C7R的慢性信号传导诱导了一种与肿瘤内应激NK细胞一致的转录特征，其效应功能减弱。我们鉴定出了与慢性细胞因子应激NK细胞相关的基因候选物，这些候选物可作为靶点以减轻实体TME内CAR-NK细胞的应激。 结论：C7R在不依赖外源性信号的情况下促进了CAR-NK细胞在敌对TME中的存活，但在体内导致了较差的抗肿瘤功能。我们的数据揭示了持续的IL-7信号在CAR-NK细胞中的有害作用，并为在试图增强CAR-NK细胞抗肿瘤活性时正确应用细胞因子信号提供了见解。
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