Sierro-Martínez Belén, Escamilla-Gómez Virginia, Pérez-Ortega Laura, Guijarro-Albaladejo Beatriz, Hernández-Díaz Paola, de la Rosa-Garrido María, Lara-Chica Maribel, Rodríguez-Gil Alfonso, Reguera-Ortega Juan Luis, Sanoja-Flores Luzalba, Arribas-Arribas Blanca, Montiel-Aguilera Miguel Ángel, Carmona Gloria, Robles Maria Jose, Caballero-Velázquez Teresa, Briones Javier, Einsele Hermann, Hudecek Michael, Pérez-Simón Jose Antonio, García-Guerrero Estefanía
Instituto de Biomedicina de Sevilla, IBiS/Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Sevilla, Spain.
Unidad de Producción y Reprogramación Celular de Sevilla (UPRC)-Planta CTTC Campus Virgen del Rocío de Sevilla, Red Andaluza de diseño y traslación de Terapias Avanzadas, Seville, Spain.
Cell Oncol (Dordr). 2025 Feb;48(1):219-237. doi: 10.1007/s13402-024-00984-0. Epub 2024 Aug 27.
CAR therapy targeting BCMA is under investigation as treatment for multiple myeloma. However, given the lack of plateau in most studies, pursuing more effective alternatives is imperative. We present the preclinical and clinical validation of a new optimized anti-BCMA CAR (CARTemis-1). In addition, we explored how the manufacturing process could impact CAR-T cell product quality and fitness.
CARTemis-1 optimizations were evaluated at the preclinical level both, in vitro and in vivo. CARTemis-1 generation was validated under GMP conditions, studying the dynamics of the immunophenotype from leukapheresis to final product. Here, we studied the impact of the manufacturing process on CAR-T cells to define optimal cell culture protocol and expansion time to increase product fitness.
Two different versions of CARTemis-1 with different spacers were compared. The longer version showed increased cytotoxicity. The incorporation of the safety-gene EGFRt into the CARTemis-1 structure can be used as a monitoring marker. CARTemis-1 showed no inhibition by soluble BCMA and presents potent antitumor effects both in vitro and in vivo. Expansion with IL-2 or IL-7/IL-15 was compared, revealing greater proliferation, less differentiation, and less exhaustion with IL-7/IL-15. Three consecutive batches of CARTemis-1 were produced under GMP guidelines meeting all the required specifications. CARTemis-1 cells manufactured under GMP conditions showed increased memory subpopulations, reduced exhaustion markers and selective antitumor efficacy against MM cell lines and primary myeloma cells. The optimal release time points for obtaining the best fit product were > 6 and < 10 days (days 8-10).
CARTemis-1 has been rationally designed to increase antitumor efficacy, overcome sBCMA inhibition, and incorporate the expression of a safety-gene. The generation of CARTemis-1 was successfully validated under GMP standards. A phase I/II clinical trial for patients with multiple myeloma will be conducted (EuCT number 2022-503063-15-00).
靶向B细胞成熟抗原（BCMA）的嵌合抗原受体（CAR）疗法正在作为多发性骨髓瘤的治疗方法进行研究。然而，鉴于大多数研究中缺乏平台期，寻求更有效的替代方案势在必行。我们展示了一种新型优化抗BCMA CAR（CARTemis-1）的临床前和临床验证。此外，我们探讨了生产工艺如何影响CAR-T细胞产品质量和适用性。
在临床前水平的体外和体内评估CARTemis-1的优化情况。在良好生产规范（GMP）条件下验证CARTemis-1的生成，研究从白细胞分离术到最终产品的免疫表型动态变化。在此，我们研究了生产工艺对CAR-T细胞的影响，以确定最佳细胞培养方案和扩增时间，以提高产品适用性。
比较了具有不同间隔区的两种不同版本的CARTemis-1。较长版本显示出更高的细胞毒性。将安全基因表皮生长因子受体截短型（EGFRt）整合到CARTemis-1结构中可作为监测标志物。CARTemis-1不受可溶性BCMA的抑制，在体外和体内均呈现出强大的抗肿瘤作用。比较了用白细胞介素-2（IL-2）或IL-7/IL-15进行扩增的情况，结果显示用IL-7/IL-15扩增时增殖更多、分化更少且耗竭更少。按照GMP指南生产了连续三批CARTemis-1，均符合所有要求的规格。在GMP条件下生产的CARTemis-1细胞显示记忆亚群增加、耗竭标志物减少以及对骨髓瘤细胞系和原发性骨髓瘤细胞具有选择性抗肿瘤功效。获得最佳适用性产品的最佳释放时间点为>6天且<10天（第8 - 10天）。
CARTemis-1经过合理设计，以提高抗肿瘤功效、克服可溶性BCMA抑制并整合安全基因的表达。CARTemis-1的生成已在GMP标准下成功验证。将对多发性骨髓瘤患者开展一项I/II期临床试验（欧盟临床试验编号2022 - 503063 - 15 - 00）。
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