DataSheet_2_PRAME and CTCFL-reactive TCRs for the treatment of ovarian cancer.pdf

Recurrent disease emerges in the majority of patients with ovarian cancer (OVCA). Adoptive T-cell therapies with T-cell receptors (TCRs) targeting tumor-associated antigens (TAAs) are considered promising solutions for less-immunogenic ‘cold’ ovarian tumors. In order to treat a broader patient population, more TCRs targeting peptides derived from different TAAs binding in various HLA class I molecules are essential. By performing a differential gene expression analysis using mRNA-seq datasets, PRAME, CTCFL and CLDN6 were selected as strictly tumor-specific TAAs, with high expression in ovarian cancer and at least 20-fold lower expression in all healthy tissues of risk. In primary OVCA patient samples and cell lines we confirmed expression and identified naturally expressed TAA-derived peptides in the HLA class I ligandome. Subsequently, high-avidity T-cell clones recognizing these peptides were isolated from the allo-HLA T-cell repertoire of healthy individuals. Three PRAME TCRs and one CTCFL TCR of the most promising T-cell clones were sequenced, and transferred to CD8+ T cells. The PRAME TCR-T cells demonstrated potent and specific antitumor reactivity in vitro and in vivo. The CTCFL TCR-T cells efficiently recognized primary patient-derived OVCA cells, and OVCA cell lines treated with demethylating agent 5-aza-2′-deoxycytidine (DAC). The identified PRAME and CTCFL TCRs are promising candidates for the treatment of patients with ovarian cancer, and are an essential addition to the currently used HLA-A*02:01 restricted PRAME TCRs. Our selection of differentially expressed genes, naturally expressed TAA peptides and potent TCRs can improve and broaden the use of T-cell therapies for patients with ovarian cancer or other PRAME or CTCFL expressing cancers.

卵巢癌（OVCA）患者中，多数病例出现复发性疾病。以肿瘤相关抗原（TAAs）为靶点的T细胞受体（TCRs）疗法，被视为治疗免疫原性较低的‘冷’型卵巢肿瘤的颇有希望的解决方案。为了治疗更广泛的病患群体，迫切需要更多靶向不同TAAs来源肽段，并与多种HLA I类分子结合的TCRs。通过使用mRNA-seq数据集进行差异基因表达分析，我们选定了PRAME、CTCFL和CLDN6作为严格意义上的肿瘤特异性TAAs，它们在卵巢癌中的表达水平较高，而在所有健康组织的风险组织中至少低20倍。在原发性卵巢癌患者样本和细胞系中，我们证实了这些TAAs的表达，并在HLA I类配体组中鉴定出天然表达的TAA来源肽段。随后，从健康个体的异基因HLA T细胞库中分离出高亲和力的T细胞克隆，这些克隆能够识别这些肽段。对最具潜力的T细胞克隆中的三个PRAME TCR和一个CTCFL TCR进行测序，并将其转移到CD8+ T细胞中。PRAME TCR-T细胞在体外和体内均表现出强大的、特异性的抗肿瘤反应。CTCFL TCR-T细胞能够有效识别源自原发性患者的卵巢癌细胞，以及用去甲基化剂5-aza-2′-deoxycytidine（DAC）处理的卵巢癌细胞系。所识别的PRAME和CTCFL TCRs是治疗卵巢癌患者的有希望候选者，并且是当前使用的HLA-A*02:01限制性PRAME TCRs的重要补充。我们选择差异表达的基因、天然表达的TAA肽段和高效的TCRs，可以改善并拓宽T细胞疗法在卵巢癌或其他PRAME或CTCFL表达型癌症患者中的应用。