Landscape and Selection of Vaccine Epitopes in SARS-CoV-2

Early in the pandemic, we designed a SARS-CoV-2 peptide vaccine containing epitope regions optimized for concurrent B cell, CD4+ T cell, and CD8+ T cell stimulation. The rationale for this design was to drive both humoral and cellular immunity with high specificity while avoiding undesired effects such as antibody-dependent enhancement (ADE). In this study, we combine computational prediction of T cell epitopes, recently published B cell epitope mapping studies, and epitope accessibility to select candidate peptide vaccines for SARS-CoV-2. We begin with an exploration of the space of possible T cell epitopes in SARS-CoV-2 with interrogation of predicted HLA-I and HLA-II ligands, overlap between predicted ligands, protein source, as well as concurrent human/murine coverage. Beyond MHC affinity, T cell vaccine candidates were further refined by predicted immunogenicity, viral source protein abundance, sequence conservation, coverage of high frequency HLA alleles and co-localization of CD4+ and CD8+ T cell epitopes. B cell epitope regions were chosen from linear epitope mapping studies of convalescent patient serum, followed by filtering to select regions with surface accessibility, high sequence conservation, spatial localization near functional domains of the spike glycoprotein, and avoidance of glycosylation sites. From 58 initial candidates, three B cell epitope regions were identified. By combining these B cell and T cell analyses, as well as a manufacturability heuristic, we propose a set of SARS-CoV-2 vaccine peptides for use in subsequent murine studies. The immunogenicity of the selected peptides was validated using ELISpot and ELISA following murine vaccination. We also curated a dataset of almost a thousand observed T-cell epitopes from convalescent COVID-19 patients across eight studies. Our vaccine selection process appears to be effective at predicting recurrent T-cell epitopes and strong T-cell responses were observed in mice following vaccination. Humoral responses were deficient, likely due to the unrestricted conformational space inhabited by linear vaccine peptides. Overall, we find our selection process and vaccine formulation to be appropriate for identifying T-cell epitopes and eliciting T-cell responses against those epitopes.新冠疫情大流行初期，我们设计了一款新型冠状病毒（SARS-CoV-2）肽疫苗(peptide vaccine)，其包含经优化的表位(epitope)区域，可同时刺激B细胞(B cell)、CD4+ T细胞(CD4+ T cell)与CD8+ T细胞(CD8+ T cell)。本设计的核心逻辑在于，在保证高特异性的同时诱导体液免疫(humoral immunity)与细胞免疫(cellular immunity)，并规避抗体依赖性增强(antibody-dependent enhancement, ADE)等不良效应。本研究整合了T细胞表位的计算预测、近期发表的B细胞表位定位(epitope mapping)研究结果，以及表位可及性分析，以筛选新型冠状病毒的候选肽疫苗。我们首先探索了新型冠状病毒中潜在T细胞表位的空间分布，对预测的人类白细胞抗原（human leukocyte antigen, HLA）I类与HLA-II类配体(ligand)、预测配体间的重叠区域、蛋白来源，以及同时覆盖人类与小鼠的表位覆盖度进行了解析分析。除主要组织相容性复合体(major histocompatibility complex, MHC)亲和力外，我们还通过预测免疫原性(immunogenicity)、病毒来源蛋白的丰度、序列保守性、高频HLA等位基因(allele)覆盖度，以及CD4+与CD8+ T细胞表位的共定位(co-localization)，进一步优化T细胞候选疫苗。B细胞表位区域则选自康复患者(convalescent patient)血清的线性表位(linear epitope)定位研究，随后通过筛选选取满足以下条件的区域：具备表面可及性、高序列保守性、定位于刺突糖蛋白(spike glycoprotein)功能域附近，且避开糖基化位点(glycosylation site)。从最初的58个候选区域中，我们最终确定了3个B细胞表位区域。结合上述B细胞与T细胞分析结果，以及可制造性启发式规则(manufacturability heuristic)，我们提出了一套新型冠状病毒疫苗肽序列，用于后续的小鼠研究。通过小鼠免疫后的酶联免疫斑点试验(ELISpot)与酶联免疫吸附试验(ELISA)验证，我们确认了所选肽段的免疫原性。此外，我们还整理构建了一套数据集，包含来自8项研究、共计近千个在新型冠状病毒肺炎（COVID-19）康复患者中观测到的T细胞表位。本研究的疫苗筛选流程可有效预测复现性T细胞表位，且免疫小鼠后可观测到强烈的T细胞免疫应答。但体液免疫应答存在不足，这可能是由于线性疫苗肽段所处的无约束构象空间所致。总体而言，我们的筛选流程与疫苗制剂(vaccine formulation)可有效识别T细胞表位，并诱导针对这些表位的T细胞免疫应答。