Targeting immunoproteasome-mediated antigen presentation in colorectal cancer immunotherapy

摘要 免疫系统可以将健康细胞与肿瘤细胞区分开，因为后者表达肿瘤相关蛋白。 抗原（TAAs）。在CD 8 + T细胞介导的免疫应答的背景下，TAA的识别通过以下方式发生： TAAs通过主要组织相容性I类（MHC-I）在肿瘤细胞上的呈递及其与T细胞的相互作用 细胞受体（TCR）。削弱这一事件将最终减少或消灭CD 8 + T细胞 介导的肿瘤细胞毒性。然而，抗原提呈的减少或丧失是免疫缺陷的常见和必需的。 肿瘤细胞逃避免疫识别和破坏的机制，包括基因组缺失， MHC-I基因、抗原呈递相关基因的转录抑制与肿瘤的失调 通过26 S免疫蛋白酶体进行抗原加工。尽管最近基于直接免疫的免疫疗法取得了成功， 由于免疫检查点蛋白（PD-1，CTLA-4和PD-L1）的阻断，大多数实体瘤患者 没有持久的临床获益。肿瘤对免疫检查点的抵抗机制 阻断（ICB）涉及低水平的肿瘤浸润淋巴细胞、抑制性免疫细胞（TCFs，MDSC 和巨噬细胞）和不利的肿瘤微环境。抗原加工过程中的畸变， 呈递机制（APM）基因与不良临床结果相关。因此，治疗干预 将是一种有前途的治疗方法，以提高对肿瘤抗原呈递的反应性。 免疫检查点阻断。在一项从草药中筛选小分子化合物的研究中， 细胞介导的细胞毒性，我们确定了阿替卡林I（ATT-I），显着促进肿瘤抗原 在人和小鼠结肠直肠癌细胞的递呈中， CD 8 + T细胞。在同基因小鼠结直肠肿瘤模型中，ATT-I与PD-1抗体的组合治疗 显著增强免疫检查点阻断疗法的功效。我们建议， 免疫蛋白酶体与ATT-I的结合促进肿瘤抗原呈递，增强T细胞的细胞毒性， 提高了肿瘤对免疫疗法的反应。在本申请中，我们将1）确定分子 ATT-I促进结直肠癌细胞中抗原加工和呈递的机制; 2）评估 ATT-I治疗对肿瘤内免疫细胞浸润、细胞毒性T淋巴细胞活性及其 与结直肠肿瘤中肿瘤微环境的相互作用; 3）评估ATT-I在结直肠肿瘤中的治疗活性， 使用基因工程增强免疫检查点阻断免疫疗法（ICB）的功效 结肠直肠癌小鼠模型和人离体结肠直肠肿瘤类器官模型。

Abstract The immune system can distinguish healthy cells from tumor cells, as the latter expresses tumor associated antigens (TAAs). In the context of CD8+ T cell-mediated immune responses, recognition of TAAs occurs through the presentation of TAAs via major histocompatibility class I (MHC-I) on tumor cells and their interaction with T- cell receptor (TCR) on the CD8+ T cells. Impairing this event will ultimately reduce or annihilate the CD8+ T-cell mediated tumor cytotoxicity. However, reduction or loss of antigen presentation is a frequent and essential mechanism used by tumor cells to escape immune recognition and destruction, including genomic deletion of MHC-I genes, transcriptional suppression of antigen presentation-associated genes, and dysregulation of tumor antigen processing by 26S immunoproteasome. Despite the recent success in immunotherapies based on direct blockade of immune checkpoint proteins (PD-1, CTLA-4 and PD-L1), a majority of patients with solid tumors do not experience durable clinical benefits. The mechanisms of tumor resistance against immune checkpoint blockade (ICB) involve low levels of tumor infiltrating lymphocytes, suppressive immune cells (Tregs, MDSCs and macrophages), and adverse tumor microenvironment. Aberrations in the antigen processing and presentation machinery (APM) genes correlate with poor clinical outcomes. Therefore, therapeutic intervention of tumor antigen presentation would be a promising therapeutic approach to improve responsiveness towards immune checkpoint blockade. In a screen for small-molecule compounds from herbal medicine that potentiate T cell-mediated cytotoxicity, we identified atractylenolide I (ATT-I) that significantly promotes tumor antigen presentation of both human and mouse colorectal cancer cells and thereby enhances the cytotoxic response of CD8+ T cells. In syngeneic mouse colorectal tumor models, treatment of ATT-I in combination with PD-1 antibody markedly enhanced the efficacy of immune checkpoint blockade therapy. We propose that targeting the function of immunoproteasome with ATT-I promotes tumor antigen presentation, empowers T-cell cytotoxicity, and thus elevates the tumor response to immunotherapy. In this application, we will 1) determine the molecular mechanism by which ATT-I promotes antigen processing and presentation in colorectal cancer cells; 2) evaluate the effect of ATT-I treatment on intra-tumor immune cell infiltration, activity of cytotoxic T lymphocytes, and their interaction with tumor microenvironment in colorectal tumors; 3) assess the therapeutic activity of ATT-I in enhancing the efficacy of immune checkpoint blockade immunotherapy (ICB) using genetically engineered mouse models of colorectal cancer and human ex vivo colorectal tumor organoid models.