Purpose: Given that heterogeneous expression and variants of antigens on solid tumors are responsible for relapse after chimeric antigen receptor (CAR)-T cell therapy, we hypothesized that combinatorial targeting two tumor-associated antigens would lessen this problem and enhance the antitumor activity of T cells./nMethods: The co-expression level of CD70 and B7-H3 was analyzed in multiple tumor tissue samples. Further, two putative antigens were identified in The Cancer Genome Atlas and Gene Expression Profiling Interactive Analysis database. Two CD70 targeted CARs with different antigen binding domain, truncated CD27 and CD70 specific single-chain antibody fragment (scFv), were designed to screen a more suitable target-antigen binding moiety. Accordingly, we designed a bivalent tandem CAR (TanCAR) and further assessed the anti-tumor efficacy of TanCAR-T cells in vitro and in vivo./nResults: Our results indicated that co-expression of CD70 and B7-H3 was observed on multiple tumor types including kidney, breast, esophageal, liver, colon cancer, glioma as well as melanoma. The CD70 targeted CAR-T cells with binding moiety of CD70 specific scFv exhibit a higher affinity and antitumor effect against CD70⁺ tumor cells. TanCAR-T cells induced enhanced ability of cytolysis and cytokine release over unispecific CAR-T cells when encountering tumor cells expressing two target-antigens. Further, low doses of TanCAR-T cells could also effectively control the lung cancer and melanoma xenografts and improved overall survival of the treated animals./nConclusion: TanCAR-T cells targeting CD70 and B7-H3 exhibit enhanced antitumor functionality and improve the problem of antigenic heterogeneity and variant in the treatment against solid tumor and melanoma.

中文翻译：

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靶向CD70和B7-H3的串联CAR-T细胞对多种实体瘤表现出有效的临床前活性。

目的：鉴于实体肿瘤上抗原的异质表达和变体是嵌合抗原受体（CAR）-T细胞治疗后复发的原因，我们假设组合靶向两种与肿瘤相关的抗原可以减轻该问题并增强T的抗肿瘤活性细胞/ n方法：在多个肿瘤组织样本中分析了CD70和B7-H3的共表达水平。此外，在《癌症基因组图谱》和《基因表达谱分析互动分析》数据库中鉴定出两种推定抗原。设计具有截短的CD27和CD70特异性单链抗体片段（scFv）的两个具有不同抗原结合结构域的CD70靶向CAR，以筛选更合适的靶抗原结合部分。因此，我们设计了一个二价串联CAR（TanCAR）和进一步评估TanCAR-T细胞的抗肿瘤功效的体外和体内./n结果：我们的结果表明，在多种肿瘤类型中观察到了CD70和B7-H3的共表达，包括肾脏，乳腺癌，食道癌，肝癌，结肠癌，神经胶质瘤以及黑色素瘤。具有CD70特异性scFv结合部分的CD70靶向CAR-T细胞表现出对CD70 ⁺肿瘤细胞更高的亲和力和抗肿瘤作用。当遇到表达两种靶抗原的肿瘤细胞时，TanCAR-T细胞诱导的溶细胞能力和细胞因子释放高于单特异性CAR-T细胞。此外，低剂量TanCAR-T细胞也可以有效地控制肺癌和黑素瘤异种移植物和处理过的animals./n的改善的总体存活结论： 靶向CD70和B7-H3的TanCAR-T细胞显示出增强的抗肿瘤功能，并改善了针对实体瘤和黑色素瘤的抗原异质性和变异性问题。