Therapeutic Targeting of Immune Evasion from the MICA - NKG2D Pathway

项目概要 MICA 和 MICB (MICA/B) 是多种人类癌症中经常表达的应激蛋白 是基因组损伤的结果，但很少由健康细胞表达。 MICA/B 作为配体 所有细胞毒性淋巴细胞表达的 NKG2D 受体，包括 CD8 T 细胞、 T 细胞、NKT 细胞和 NK 细胞，能够识别和消除应激和转化的细胞。 MICA/B 的蛋白水解脱落是 在许多人类癌症中，NKG2D 介导的肿瘤免疫是一种主要的免疫逃避机制。这个 脱落过程涉及通过二硫键异构酶 ERp5 的作用展开 MICA/B α3 结构域 它使得 MICA/B 能够被 ADAM 和 MMP 家族的蛋白酶切割。这是不可行的 使用小分子抑制剂抑制体内脱落，因为相关蛋白酶具有广泛的底物 特殊性。我们开发了一种通过设计空间阻断抗体来抑制 MICA/B 脱落的方法 MICA/B α3 域中的脱落位点。这些抗体通过多种方式有效抑制 MICA/B 脱落 一组人类癌细胞系，从而显着增加这些刺激的细胞表面密度 NKG2D 配体。因此，MICA/B 抗体可诱导 NK 细胞强烈杀死人类肿瘤细胞。 这些抗体还在小鼠转移模型中诱导免疫。单细胞 RNA-seq 数据表明 MICA/B 抗体诱导转移浸润 NK 细胞向活化且具有细胞毒性的 NK 细胞发生显着转变 状态。我们还在人源化小鼠模型中验证了这些抗体，其中人类 NK 细胞靶向 由人类肿瘤细胞形成的转移。 许多人类癌症因 MHC I 类缺失而对检查点封锁的免疫疗法产生抗药性 表达。然而，先天性介导的抗肿瘤免疫不需要 MHC I 类蛋白表达。 T 细胞群（NKT 细胞、T 细胞）和均表达 NKG2D 受体的 NK 细胞。主要目标是 该项目旨在开发 MICA/B 抗体作为 MHC I 类缺陷肿瘤细胞的治疗策略， 对传统的 CD8 T 细胞有抵抗力。我们开发了一种综合方法来研究这一重要的 完全免疫功能小鼠模型（目标 1）以及人源化小鼠模型和人类的问题 肿瘤转移（目标 2）。在目标 1 中，我们将研究先天 T 细胞和 NK 细胞群对 MICA/B 抗体介导针对自发转移的免疫。在目标 2 中，我们将进行深入的 对人黑色素瘤中表达 NKG2D 的先天 T 细胞和 NK 细胞群进行单细胞 RNA 测序分析 转移。我们还将使用人源化小鼠模型来开发已确定的联合疗法 增强 MICA/B 表达的癌症治疗药物，因此可能与 MICA/B 产生协同作用 抗体。这些研究将通过开发新的方法显着推进癌症免疫治疗领域的发展 针对对当前免疫疗法有抵抗力的人类癌症的方法。

Project Summary MICA and MICB (MICA/B) are stress proteins that are frequently expressed by diverse types of human cancer as a consequence of genomic damage, but are rarely expressed by healthy cells. MICA/B serve as ligands for the NKG2D receptor expressed by all cytotoxic lymphocytes, including CD8 T cells,  T cells, NKT cells and NK cells, enabling recognition and elimination of stressed and transformed cells. Proteolytic shedding of MICA/B is a major immune evasion mechanism from NKG2D-mediated tumor immunity in many human cancers. This shedding process involves unfolding of the MICA/B 3 domain by the action of the disulfide isomerase ERp5 which enables MICA/B cleavage by proteases belonging to the ADAM and MMP families. It is not feasible to inhibit shedding in vivo with small molecule inhibitors because the relevant proteases have broad substrate specificities. We developed an approach to inhibit MICA/B shedding by designing antibodies that sterically block the shedding site in the MICA/B 3 domain. These antibodies potently inhibit MICA/B shedding by a diverse panel of human cancer cell lines and thereby substantially increase the cell surface density of these stimulatory NKG2D ligands. As a consequence, MICA/B antibodies induce strong killing of human tumor cells by NK cells. These antibodies also induce immunity in mouse models of metastasis. Single-cell RNA-seq data show that a MICA/B antibody induces a striking shift among metastasis-infiltrating NK cells to an activated and cytotoxic state. We have also validated these antibodies in a humanized mouse model in which human NK cells target metastases formed by human tumor cells. Many human cancers are resistant to immunotherapy with checkpoint blockade through loss of MHC class I expression. However, MHC class I protein expression is not required for anti-tumor immunity mediated by innate T cell populations (NKT cells, T cells) and NK cells that all express the NKG2D receptor. The major goal of this project is to develop MICA/B antibodies as a therapeutic strategy for MHC class I deficient tumor cells that are resistant to conventional CD8 T cells. We have developed an integrated approach to study this important question in fully immunocompetent mouse models (Aim 1) as well as humanized mouse models and human tumor metastases (Aim 2). In Aim 1, we will examine the contribution of innate T cell and NK cell populations to MICA/B antibody mediated immunity against spontaneous metastases. In Aim 2, we will perform an in depth single-cell RNA-seq analysis of NKG2D-expressing innate T cell and NK cell population in human melanoma metastases. We will also use a humanized mouse model to develop combination therapies with established cancer therapeutics that enhance MICA/B expression and may therefore act synergistically with MICA/B antibodies. These studies will significantly advance the cancer immunotherapy field by developing novel approaches to target human cancers resistant to current immunotherapies.