Shanghai Yihao Biological Technology Co., Ltd, Shanghai, 200231, China.
Department of General Surgery, Changhai Hospital, Shanghai, 200433, China.
Exp Cell Res. 2021 Dec 1;409(1):112886. doi: 10.1016/j.yexcr.2021.112886. Epub 2021 Oct 19.
Chimeric antigen receptor (CAR) T cells have been successfully used for the treatment of hematological malignancies including acute and chronic lymphoblastic leukemia. However, results of CAR T cell projects in solid tumors have been less impressive to date, partly because of immunosuppressive tumor microenvironment (TME). It is widely known that high adenosine production is an important factor causing tumor-induced immunosuppression in TME, and adenosine mediates the suppression of anti-tumor T cell responses via binding and signaling through adenosine 2a receptor (A2aR). Previous studies have shown that adenosine generated by cancer cells significantly inhibits T cell anti-tumor activity through binding and then activating adenosine 2A receptors (A2aRs) of T cells. Based on the previous work, in our study, we evaluated whether A2aR disruption by shRNA could enhance the anti-tumor function of anti-mesothelin (MSLN) CAR T cells both in vitro and in vivo. For this goal above, we used MSLN-positive human ovarian serous carcinoma cells (SKOV3) and human colon cancer cells (HCT116) as target cancer cells while MSLN-negative human ovarian cancer cells (ES2) as non-target cancer cells. We observed that targeting cell-intrinsic A2aR through shRNA overexpression caused significant A2aR disruption in CAR T cells and profoundly increased CAR T cell efficacy in both CAR T cell cytokine production and cytotoxicity towards MSLN-positive cancer cells in vitro. More importantly, in SKOV3 xenograft mouse models, anti-MSLN CAR-T cells significantly reduced the tumor burden compared with non-transduced T cells, and the anti-tumor activity of A2aR-disrupted anti-MSLN CAR-T cells was stronger than that of wild-type anti-MSLN CAR-T cells. Altogether, our study showed enhanced anti-tumor efficacy caused by shRNA-mediated A2aR disruption in anti-MSLN CAR T cells both in vitro and in vivo, which proved that shRNA-mediated modification of gene expression might be an excellent strategy for improving CAR T cell function in immunosuppressive tumor microenvironment (TME) and could potentially improve the outcome of treatment in clinical trials.
嵌合抗原受体 (CAR) T 细胞已成功用于治疗包括急性和慢性淋巴细胞白血病在内的血液系统恶性肿瘤。然而，到目前为止，CAR T 细胞项目在实体瘤中的结果并不令人印象深刻，部分原因是免疫抑制性肿瘤微环境 (TME)。众所周知，高腺苷产生是导致 TME 中肿瘤诱导免疫抑制的一个重要因素，腺苷通过与腺苷 2a 受体 (A2aR) 结合和信号转导来介导抗肿瘤 T 细胞反应的抑制。先前的研究表明，癌细胞产生的腺苷通过与 T 细胞上的腺苷 2A 受体 (A2aR) 结合并激活其功能，显著抑制 T 细胞的抗肿瘤活性。基于先前的工作，在我们的研究中，我们评估了通过 shRNA 破坏 A2aR 是否可以增强抗间皮素 (MSLN) CAR T 细胞在体外和体内的抗肿瘤功能。为此，我们使用 MSLN 阳性人卵巢浆液性癌细胞 (SKOV3) 和人结肠癌细胞 (HCT116) 作为靶癌细胞，而 MSLN 阴性人卵巢癌细胞 (ES2) 作为非靶癌细胞。我们观察到，通过 shRNA 过表达靶向细胞内固有 A2aR 会导致 CAR T 细胞中 A2aR 明显破坏，并显著增加 CAR T 细胞在体外产生细胞因子和对 MSLN 阳性癌细胞的细胞毒性方面的疗效。更重要的是，在 SKOV3 异种移植小鼠模型中，与非转导 T 细胞相比，抗 MSLN CAR-T 细胞显著降低了肿瘤负担，并且 A2aR 破坏的抗 MSLN CAR-T 细胞的抗肿瘤活性强于野生型抗 MSLN CAR-T 细胞。总之，我们的研究表明，shRNA 介导的 A2aR 破坏在体外和体内均可增强抗 MSLN CAR T 细胞的抗肿瘤疗效，证明 shRNA 介导的基因表达修饰可能是改善 CAR T 细胞在免疫抑制性肿瘤微环境 (TME) 中功能的一种极好策略，并有可能提高临床试验中的治疗效果。
