免疫检查点阻断（ICB）已成为多种实体肿瘤的标准治疗。已研究多种联合治疗方法以提高治疗效果。抗血管生成抑制剂与ICB的组合已在多种癌症中显示出疗效。为了改进对这些治疗方法的协同作用的机制理解，我们对ipilimumab和bevacizumab治疗的长期反应患者的血清进行了人类蛋白质芯片的血清学筛选。我们发现针对EDIL3的高滴度抗体反应与良好的临床预后相关。EDIL3是一种细胞外蛋白，已被确认为多种恶性肿瘤的不良预后标志物。我们的肿瘤免疫功能障碍与排除分析预测了EDIL3与细胞毒性免疫细胞浸润的免疫排除标记相关，并导致ICB的非应答。在我们的研究中，预测肿瘤相关成纤维细胞（CAFs）是免疫排除相关细胞中EDIL3的来源。此外，通过TCGA-SKCM和CheckMate 064数据分析，发现EDIL3水平与全面成纤维细胞TGF-β反应增加、血管生成标志物富集以及上皮-间质转化的诱导相关。我们的体外研究验证了患者来源的CAFs中EDIL3过表达和TGF-β调节。在患者的治疗前血清样本中，EDIL3的循环水平与VEGF的循环水平相关，并且像VEGF一样，EDIL3增强了患者来源的肿瘤内皮细胞（TEC）的血管生成能力。在机制上，3D微流体培养和2D细胞外穿膜实验证实了EDIL3介导的LFA-1/ICAM-1相互作用破坏可能是T细胞排除的可能机制。我们建议将EDIL3作为改善免疫细胞经内皮迁移和ICB治疗效果的潜在靶点。

Immune checkpoint blockade (ICB) has become the standard of care for several solid tumors. Multiple combinatorial approaches have been studied to improve therapeutic efficacy. The combination of anti-angiogenesis agents and ICB has demonstrated efficacy in several cancers. To improve the mechanistic understanding of synergies with these treatment modalities, we performed serologic screens of human protein array using sera from long-term responding patients treated with ipilimumab and bevacizumab. We discovered a high-titer antibody response against EDIL3 that correlated with favorable clinical outcomes. EDIL3 is an extracellular protein, identified as a marker of poor prognosis in various malignancies. Our Tumor Immune Dysfunction and Exclusion analysis predicted that EDIL3 is associated with immune exclusion signatures for cytotoxic immune cell infiltration and non-response to ICB. In our study, cancer-associated fibroblasts (CAFs) were predicted as source of EDIL3 in immune exclusion-related cells. Furthermore, TCGA-SKCM and CheckMate 064 data analyses correlated high levels of EDIL3 with increased pan-fibroblast TGF-β response, enrichment of angiogenic signatures, and induction of epithelial-to-mesenchymal transition. Our in vitro studies validated EDIL3 overexpression and TGF-β regulation in patient-derived CAFs. In pre-treatment serum samples from patients, circulating levels of EDIL3 were associated with circulating levels of VEGF, and like VEGF, EDIL3 increased the angiogenic abilities of patient-derived tumor endothelial cells (TEC). Mechanistically, 3-D microfluidic cultures and 2D transmigration assays with TEC endorsed EDIL3-mediated disruption of LFA-1/ICAM-1 interaction as possible means of T cell exclusion. We propose EDIL3 as a potential target for improving the transendothelial migration of immune cells and efficacy of ICB therapy.