肿瘤细胞治疗研究
英文原题：Humanized CD70-targeted CAR-T cells with reduced fratricide demonstrate potent antitumor activity against AML and RCC.
嵌合抗原受体（CAR）-T细胞疗法在B细胞恶性肿瘤中取得了显著成功，但由于缺乏理想的靶抗原，其在急性髓系白血病（AML）和肾细胞癌（RCC）中的应用仍面临挑战。
嵌合抗原受体（CAR）-T细胞疗法在B细胞恶性肿瘤中已取得显著成功，但由于缺乏理想的靶抗原，其在急性髓系白血病（AML）和肾细胞癌（RCC）中的应用仍面临挑战。CD70在AML和RCC肿瘤细胞上高表达，而在正常组织中其表达主要局限于活化的淋巴细胞，使其成为有前景的免疫治疗靶点。然而，活化T细胞上的CD70表达可引发CD70靶向CAR-T细胞的相互杀伤，这对其制备和疗效构成重大挑战。以临床阶段抗CD70抗体ARGX-110的类似物作为参照，我们从免疫小鼠的噬菌体展示库中进行筛选，鉴定出高亲和力抗CD70抗体，随后用其构建了一组第二代CAR。功能表征鉴定出先导候选物A174-CAR-T，其具有高CAR表达、强体外抗肿瘤活性和低相互杀伤。我们进一步对其单链可变区进行人源化，生成A174-hu1-CAR。与亲本构建体相比，A174-hu1-CAR-T细胞表现出增强的扩增和显著降低的CD70表面表达，与改善的顺式掩蔽（即同一细胞上的表面屏蔽）相一致。分子对接和表面静电势分析提示，人源化优化了scFv-CD70结合界面，这与CAR表面表达增加共同促成了掩蔽增强和相互杀伤减少。在功能上，A174-hu1-CAR-T细胞保持了强效细胞毒性，并显示出增殖性和中枢记忆T细胞亚群的富集。在异种移植模型中，A174-hu1-CAR-T输注对AML和RCC均表现出强效抗肿瘤疗效，且安全性特征良好。总之，我们开发了一种人源化CD70靶向CAR-T细胞疗法A174-hu1-CAR-T，其具有降低的自相残杀作用和强效抗肿瘤活性，为进一步转化开发提供了临床前基础。
Chimeric antigen receptor (CAR)-T cell therapy has achieved remarkable success in B-cell malignancies, but its application in acute myeloid leukemia (AML) and renal cell carcinoma (RCC) remains challenging due to the lack of ideal target antigens. CD70 is highly expressed on AML and RCC tumor cells, while its expression in normal tissues is largely restricted to activated lymphocytes, making it a promising immunotherapeutic target. However, CD70 expression on activated T cells can trigger fratricide in CD70-targeted CAR-T cells, posing a major challenge for their preparation and efficacy. Using an analog of the clinical-stage anti-CD70 antibody ARGX-110 as a reference, we screened a phage display library from immunized mice and identified high-affinity anti-CD70 antibodies, which were then used to construct a panel of second-generation CARs. Functional characterization identified a lead candidate, A174-CAR-T, with high CAR expression, strong in vitro antitumor activity, and low fratricide. We further humanized its single-chain variable fragment to generate A174-hu1-CAR. Compared with the parental construct, A174-hu1-CAR-T cells exhibited enhanced expansion and markedly reduced surface expression of CD70, consistent with improved cis-masking (i.e., surface shielding on the same cell). Molecular docking and surface electrostatic potential analyses suggested that humanization optimized the scFv-CD70 binding interface, which together with increased CAR surface expression contributed to the enhanced masking and reduced fratricide. Functionally, A174-hu1-CAR-T cells maintained potent cytotoxicity and showed enrichment of proliferative and central memory T-cell subsets. In xenograft models, A174-hu1-CAR-T infusion demonstrated potent antitumor efficacy against both AML and RCC, with a favorable safety profile. Overall, we developed a humanized CD70-targeted CAR-T cell therapy, A174-hu1-CAR-T, with reduced fratricide and potent antitumor activity, providing a preclinical foundation to support further translational development.
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