Kote Sachin, Pirog Artur, Bedran Georges, Alfaro Javier, Dapic Irena
International Centre for Cancer Vaccine Science, University of Gdansk, 80-308 Gdansk, Poland.
Cancers (Basel). 2020 Feb 26;12(3):535. doi: 10.3390/cancers12030535.
Neoantigen-based immunotherapies promise to improve patient outcomes over the current standard of care. However, detecting these cancer-specific antigens is one of the significant challenges in the field of mass spectrometry. Even though the first sequencing of the immunopeptides was done decades ago, today there is still a diversity of the protocols used for neoantigen isolation from the cell surface. This heterogeneity makes it difficult to compare results between the laboratories and the studies. Isolation of the neoantigens from the cell surface is usually done by mild acid elution (MAE) or immunoprecipitation (IP) protocol. However, limited amounts of the neoantigens present on the cell surface impose a challenge and require instrumentation with enough sensitivity and accuracy for their detection. Detecting these neopeptides from small amounts of available patient tissue limits the scope of most of the studies to cell cultures. Here, we summarize protocols for the extraction and identification of the major histocompatibility complex (MHC) class I and II peptides. We aimed to evaluate existing methods in terms of the appropriateness of the isolation procedure, as well as instrumental parameters used for neoantigen detection. We also focus on the amount of the material used in the protocols as the critical factor to consider when analyzing neoantigens. Beyond experimental aspects, there are numerous readily available proteomics suits/tools applicable for neoantigen discovery; however, experimental validation is still necessary for neoantigen characterization.
基于新抗原的免疫疗法有望比当前的标准治疗方法改善患者预后。然而，检测这些癌症特异性抗原是质谱领域的重大挑战之一。尽管免疫肽的首次测序在几十年前就已完成，但如今从细胞表面分离新抗原所使用的方案仍多种多样。这种异质性使得不同实验室和研究之间难以比较结果。从细胞表面分离新抗原通常通过温和酸洗脱（MAE）或免疫沉淀（IP）方案进行。然而，细胞表面存在的新抗原数量有限带来了挑战，需要具有足够灵敏度和准确性的仪器来进行检测。从少量可用的患者组织中检测这些新肽限制了大多数研究的范围，只能进行细胞培养。在此，我们总结了提取和鉴定主要组织相容性复合体（MHC）I类和II类肽的方案。我们旨在根据分离程序的适用性以及用于新抗原检测的仪器参数来评估现有方法。我们还将重点关注方案中使用的材料量，这是分析新抗原时需要考虑的关键因素。除了实验方面，有许多现成的蛋白质组学套件/工具可用于新抗原发现；然而，新抗原表征仍需要进行实验验证。
