Neoantigen targeted dendritic cell vaccine generates durable T cell responses exhibiting the full spectrum of differentiation states in NSCLC patients

Background. Dendritic cell (DC)-based neoantigen vaccination holds potential as a safe and effective adjuvant therapy for patients with early-stage, resectable NSCLC, a tumor type typically characterized by high mutational loads. DCs have the unique ability to elicit robust antitumoral T-cell responses, while neoantigens are ideal targets to elicit high-affinity T cell responses with exquisite tumor specificity. Here, we present the results of a phase I clinical trial in which a novel DC vaccine targeting neoantigens was evaluated in six patients with early stage, resected NSCLC. Methods. Tumor samples were subjected to a comprehensive neoantigen identification approach encompassing genomics, transcriptomics and immunopeptidomics. Using genomics and transcriptopmics data, tumor-specific antigens were identified by a bioinformatics approach. Additionally, immunopeptidomics was performed by immunoprecipitation of human MHC class I molecule followed by immunopeptides enrichment and LC-MS/MS analysis. Two immunopeptidomics screens were performed. In the first immunopeptidomics screen, patient-derived tumors were analyzed to uncover neoepitopes specific for the patient. In the second, screen, patient-derived EBV-immortilized B cell lines overexpressing the selected neoepitopes were analyzed to verify that the predicted neoepitopes can bind to the HLA haplotypes of the patients. For anti-tumor vaccination, patients underwent leukapheresis for the manufacturing of monocyte-derived DCs loaded with neoantigens (Neo-mDCs) according to a four-day protocol. Neo-mDCs were injected intravenously following an intrapatient dose escalation scheme. Primary endpoint of the trial was safety. Secondary endpoints were feasibility, immunogenicity, and relapse-free survival. Results. In the first immunopeptidomics screen, one neoepitopes derived was identified from the tumor of one patient. In the second immunopeptidomics screen, several predicted neoepitopes were confirmed to be presented on the HLA haplotypes of the patients by analyzing the patient-derived EBV-immortilized B cell lines. Additionally, the vaccine was demonstrated to be feasible and safe. T cell responses were observed in 5 of 6 vaccinated patients and were dominated by CD8+ T cells, which could be detected ex vivo at high frequencies >1.5 years after the last dose. Furthermore, single cell analysis indicated that the CD8+ T cell responsive population was polyclonal and exhibited the near entire spectrum of T cell differentiation states, including a naïve-like state associated with long lasting memory but excluding exhausted cell states. Three of six vaccinated patients experienced disease relapse. Conclusion. Neo-mDC vaccination is safe and feasible. Vaccination induces large populations of neoantigen-specific T-cell responses containing long lasting memory and effector cells in early-stage NSCLC patients, suggesting clinical potential.

背景。基于树突状细胞（Dendritic cell, DC）的新抗原疫苗，有望成为早期可切除非小细胞肺癌（non-small cell lung cancer, NSCLC）患者安全有效的辅助治疗手段，这类肿瘤通常以高突变负荷为特征。树突状细胞具备诱导强效抗肿瘤T细胞应答的独特能力，而新抗原则是诱导具有高度肿瘤特异性的高亲和力T细胞应答的理想靶点。本研究报道了一项I期临床试验的结果，该试验针对6例早期可切除非小细胞肺癌患者，评估了一款靶向新抗原的新型树突状细胞疫苗。

方法。研究人员对肿瘤样本采用涵盖基因组学、转录组学和免疫肽组学的综合新抗原鉴定策略。借助基因组学与转录组学数据，通过生物信息学方法鉴定肿瘤特异性抗原。此外，免疫肽组学分析采用先免疫沉淀人MHC I类分子，随后富集免疫肽段并进行液相色谱-串联质谱（LC-MS/MS）分析的流程。本次研究共开展两次免疫肽组学筛选：第一次筛选针对患者来源的肿瘤样本，以发掘患者特异性新表位；第二次筛选则针对过表达所选定新表位的患者来源EB病毒转化B细胞系（EBV-immortalized B cell lines），以验证预测的新表位可结合患者的HLA单倍型。针对抗肿瘤疫苗接种，患者接受白细胞分离术，以按照四天方案制备负载新抗原的单核细胞来源树突状细胞（neoantigen-loaded monocyte-derived dendritic cells, Neo-mDCs）。随后按照患者内剂量递增方案静脉注射Neo-mDCs。本试验的主要终点为安全性，次要终点包括可行性、免疫原性以及无复发生存期。

结果。在第一次免疫肽组学筛选中，研究人员从1例患者的肿瘤样本中鉴定出1种新表位。在第二次免疫肽组学筛选中，通过分析患者来源的EB病毒转化B细胞系，证实多种预测的新表位可呈递于患者的HLA单倍型。此外，该疫苗被证实具备可行性与安全性。6例接种患者中有5例观察到T细胞应答，且应答以CD8+ T细胞为主，末次接种后超过1.5年仍可在体外以较高频率检测到该类细胞。进一步的单细胞分析显示，CD8+ T细胞应答群体呈多克隆性，并覆盖了T细胞分化状态的几乎全部谱系，包括与长期记忆相关的初始样状态，但不包括耗竭性细胞状态。6例接种患者中有3例出现疾病复发。

结论。负载新抗原的单核细胞来源树突状细胞疫苗具备安全性与可行性。该疫苗可在早期非小细胞肺癌患者体内诱导大量新抗原特异性T细胞应答，这类应答包含长期记忆细胞与效应细胞，提示其具有临床应用潜力。