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Abstract：
Ovarian cancer (OC) is a highly aggressive gynecological malignancy prevalent worldwide. Most OC cases are typically diagnosed at advanced stages, which has led to a 5-year overall survival rate of less than 35% following conventional treatment. Furthermore, immune checkpoint inhibitor therapy has shown limited efficacy in the treatment of patients with OC, and CAR-T therapy has also demonstrated modest results owing to inadequate T cell infiltration. Therefore, novel strategies must be developed to enhance T cell persistence and trafficking within the OC tumor microenvironment.
In this study, we developed a novel adoptive T-cell therapy for ovarian cancer based on a chimeric antigen receptor structure. We used a ligand-receptor binding motif to enhance the therapeutic effect of targeting CA125. Since mesothelin can naturally bind to CA125 with high affinity, we concatenated the core-binding fragment of mesothelin with the 4-1BB and CD3ζ signal fragments to assemble a novel CA125-targeting chimeric receptor (CR). The CAR structure targeting CA125 derived from the 4H11 antibody was also constructed. CR- and CAR-encoding RNA were electroporated into T cells to evaluate their antitumor activity both in vitro and in vivo.
While CR-T or CAR-T cells exhibited moderate activity against two ovarian cancer cell lines, T cells co-expressing CR and CAR exhibited a superior killing effect compared to T cells expressing either CR or CAR alone. Furthermore, upon interaction with ovarian tumors, the ability of CR and CAR T cells to release activation markers and functional cytokines increased significantly. Similarly, CR and CAR co-expressing T cells persistently controlled the growth of transplanted ovarian cancer tumors in NSG mice and significantly prolonged the overall survival of tumor-challenged mice. Transcriptome sequencing revealed that the survival and cytotoxicity of T cells co-expressing CR and CAR were significantly altered compared with those of T cells expressing either CR or CAR.
Our findings demonstrate that CA125 targeting CR and CAR can synergistically kill ovarian cancer cells, indicating that CA125 targeting by the two binding motifs simultaneously in tumors may improve the therapeutic outcomes of ovarian cancer treatment.
In this study, we developed a novel adoptive T-cell therapy for ovarian cancer based on a chimeric antigen receptor structure. We used a ligand-receptor binding motif to enhance the therapeutic effect of targeting CA125. Since mesothelin can naturally bind to CA125 with high affinity, we concatenated the core-binding fragment of mesothelin with the 4-1BB and CD3ζ signal fragments to assemble a novel CA125-targeting chimeric receptor (CR). The CAR structure targeting CA125 derived from the 4H11 antibody was also constructed. CR- and CAR-encoding RNA were electroporated into T cells to evaluate their antitumor activity both in vitro and in vivo.
While CR-T or CAR-T cells exhibited moderate activity against two ovarian cancer cell lines, T cells co-expressing CR and CAR exhibited a superior killing effect compared to T cells expressing either CR or CAR alone. Furthermore, upon interaction with ovarian tumors, the ability of CR and CAR T cells to release activation markers and functional cytokines increased significantly. Similarly, CR and CAR co-expressing T cells persistently controlled the growth of transplanted ovarian cancer tumors in NSG mice and significantly prolonged the overall survival of tumor-challenged mice. Transcriptome sequencing revealed that the survival and cytotoxicity of T cells co-expressing CR and CAR were significantly altered compared with those of T cells expressing either CR or CAR.
Our findings demonstrate that CA125 targeting CR and CAR can synergistically kill ovarian cancer cells, indicating that CA125 targeting by the two binding motifs simultaneously in tumors may improve the therapeutic outcomes of ovarian cancer treatment.
摘要：
背景：卵巢癌（OC）是全球流行的高度侵袭性妇科恶性肿瘤。大多数OC病例通常在晚期诊断，这导致常规治疗后5年总生存率低于35%。此外，免疫检查点抑制剂疗法在治疗OC患者中显示出有限的疗效，由于T细胞浸润不足，CAR-T疗法也显示出适度的结果。因此,必须开发新的策略来增强OC肿瘤微环境中T细胞的持久性和运输。
方法：在本研究中，我们基于嵌合抗原受体结构开发了一种新的卵巢癌过继性T细胞疗法.我们使用配体-受体结合基序来增强靶向CA125的治疗效果。由于间皮素可以高亲和力天然结合CA125，我们将间皮素的核心结合片段与4-1BB和CD3ζ信号片段连接在一起，以组装一种新型的CA125靶向嵌合受体（CR）。还构建了靶向源自4H11抗体的CA125的CAR结构。将CR-和CAR-编码RNA电穿孔到T细胞中以评估它们在体外和体内的抗肿瘤活性。
结果：虽然CR-T或CAR-T细胞对两种卵巢癌细胞系表现出中等活性，与单独表达CR或CAR的T细胞相比,共表达CR和CAR的T细胞表现出优异的杀伤作用。此外，与卵巢肿瘤相互作用后，CR和CART细胞释放活化标志物和功能性细胞因子的能力显著增强。同样，CR和CAR共表达T细胞持续控制NSG小鼠移植卵巢癌肿瘤的生长，并显著延长肿瘤攻击小鼠的总生存期。转录组测序显示,与表达CR或CAR的T细胞相比,共表达CR和CAR的T细胞的存活和细胞毒性显著改变。
结论：我们的研究结果表明，靶向CR和CAR的CA125可以协同杀死卵巢癌细胞，这表明在肿瘤中通过两个结合基序同时靶向CA125可能改善卵巢癌治疗的治疗结果。
方法：在本研究中，我们基于嵌合抗原受体结构开发了一种新的卵巢癌过继性T细胞疗法.我们使用配体-受体结合基序来增强靶向CA125的治疗效果。由于间皮素可以高亲和力天然结合CA125，我们将间皮素的核心结合片段与4-1BB和CD3ζ信号片段连接在一起，以组装一种新型的CA125靶向嵌合受体（CR）。还构建了靶向源自4H11抗体的CA125的CAR结构。将CR-和CAR-编码RNA电穿孔到T细胞中以评估它们在体外和体内的抗肿瘤活性。
结果：虽然CR-T或CAR-T细胞对两种卵巢癌细胞系表现出中等活性，与单独表达CR或CAR的T细胞相比,共表达CR和CAR的T细胞表现出优异的杀伤作用。此外，与卵巢肿瘤相互作用后，CR和CART细胞释放活化标志物和功能性细胞因子的能力显著增强。同样，CR和CAR共表达T细胞持续控制NSG小鼠移植卵巢癌肿瘤的生长，并显著延长肿瘤攻击小鼠的总生存期。转录组测序显示,与表达CR或CAR的T细胞相比,共表达CR和CAR的T细胞的存活和细胞毒性显著改变。
结论：我们的研究结果表明，靶向CR和CAR的CA125可以协同杀死卵巢癌细胞，这表明在肿瘤中通过两个结合基序同时靶向CA125可能改善卵巢癌治疗的治疗结果。
