Department of Molecular Medicine, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran.
Department of Immunology, Hybridoma Lab, Pasteur Institute of Iran, Tehran, Iran.
J Cell Biochem. 2019 Jun;120(6):10787-10795. doi: 10.1002/jcb.28370. Epub 2019 Jan 22.
Adoptive transfer of T cells expressing chimeric antigen receptors (CARs) is considered to be a novel anticancer therapy. To date, in most cases, single-chain variable fragments (scFvs) of murine origin have been used in CARs. However, this structure has limitations relating to the potential immunogenicity of mouse antigens in humans and the relatively large size of scFvs. For the first time, we used camelid nanobody (VHH) to construct CAR T cells against prostate specific membrane antigen (PSMA). The nanobody against PSMA (NBP) was used to show the feasibility of CAR T cells against prostate cancer cells. T cells were transfected, and then the surface expression of the CAR T cells was confirmed. Then, the functions of VHH-CAR T cell were evaluated upon coculture with prostate cancer cells. At the end, the cytotoxicity potential of NBPII-CAR in T cells was approximated by determining the cell surface expression of CD107a after encountering PSMA. Our data show the specificity of VHH-CAR T cells against PSMA cells (LNCaP), not only by increasing the interleukin 2 (IL-2) cytokine (about 400 pg/mL), but also the expression of CD69 by almost 38%. In addition, VHH-CAR T cells were proliferated by nearly 60% when cocultured with LNCaP, as compared with PSMA negative prostate cancer cell (DU-145), which led to the upregulation of CD107a in T cells upto 31%. These results clearly show the possibility of using VHH-based CAR T cells for targeted immunotherapy, which may be developed to target virtually any tumor-associated antigen for adoptive T-cell immunotherapy of solid tumors.
嵌合抗原受体 (CAR) T 细胞的过继转移被认为是一种新型的抗癌疗法。迄今为止，在大多数情况下，嵌合抗原受体 (CAR) 中使用的是鼠源单链可变片段 (scFv)。然而，这种结构存在与人类中鼠抗原的潜在免疫原性以及 scFv 相对较大的尺寸相关的局限性。我们首次使用骆驼科纳米抗体 (VHH) 构建针对前列腺特异性膜抗原 (PSMA) 的 CAR T 细胞。针对 PSMA 的纳米抗体 (NBP) 用于展示针对前列腺癌细胞的 CAR T 细胞的可行性。转染 T 细胞，然后确认 CAR T 细胞的表面表达。然后，在与前列腺癌细胞共培养时评估 VHH-CAR T 细胞的功能。最后，通过确定遇到 PSMA 后 T 细胞表面 CD107a 的表达来近似 NBPII-CAR 在 T 细胞中的细胞毒性潜力。我们的数据表明 VHH-CAR T 细胞针对 PSMA 细胞 (LNCaP) 的特异性，不仅通过增加白细胞介素 2 (IL-2) 细胞因子 (约 400pg/mL)，而且通过几乎 38%的 CD69 表达。此外，与 PSMA 阴性前列腺癌细胞 (DU-145) 共培养时，VHH-CAR T 细胞增殖了近 60%，这导致 T 细胞中 CD107a 的上调高达 31%。这些结果清楚地表明了使用基于 VHH 的 CAR T 细胞进行靶向免疫治疗的可能性，这可能被开发用于针对实体瘤的过继 T 细胞免疫治疗的几乎任何肿瘤相关抗原。
