Endocrine tissue molecular pathways dysregulated by immune checkpoint inhibitors causing ICI-triggered adverse events

项目摘要/摘要 程序化死亡配体-1(PD-L1)是一种免疫检查点抑制物(ICI)，与其受体相互作用， 程序性死亡-1(PD-1)抑制T细胞激活。癌细胞适应于操纵这一机制，通过 过表达PD-L1以逃避免疫激活和攻击。因此，取消Pd-L1/Pd-1相互作用 单抗是一种治疗多种癌症的有效策略。然而，它是 并发免疫相关不良事件(IrAEs)，包括内分泌不良反应(EAES)。甲状腺功能障碍， 包括破坏性甲状腺炎和甲状腺功能减退症，是与PD-L1/PD-1相关的最常见的EAES 封锁，影响高达21%的患者。因为他们的机制尚不清楚，目前这些情况 对症治疗，因此，患者通常很难管理，EAES可能会干扰 接受癌症治疗。因此，需要针对导致EAES的机制的新的治疗方法。我们 已经发现，PD-L1在甲状腺中表达，在甲状腺细胞中发挥内在功能。我们已经产生了 PD-L1表达下调或上调的人甲状腺细胞系，并证明PD-L1- L1参与了细胞生存机制，以应对炎性细胞因子诱导的应激。对Pd-的抑制 甲状腺细胞中的L1使其更容易受到炎症诱导的细胞应激和凋亡的影响 细胞因子。PD-L1在甲状腺细胞中的内在功能及其在ICI甲状腺炎发生发展中的作用 以前从未被研究过。在这项提案中，我们将剖析PD-L1在甲状腺中的免疫非依赖性功能 细胞及其在ICI-甲状腺炎发生发展中的作用。我们的假设是阻断甲状腺细胞中的PD-L1 通过抗PD-L1治疗，触发PD-L1控制的促生存细胞内信号的失调， 使甲状腺细胞容易受到细胞因子的破坏。在目标1中，我们将执行细胞和分子 PD-L1表达上调和下调的甲状腺细胞系的细胞内解剖研究 PD-L1的功能通过评估：(1)PD-L1内在活性的分子介体(例如，传递的信号 通过PD-L1胞浆结构域)；(2)PD-L1在细胞对炎症应激反应中的作用； (3)PD-L1活性的下游效应子。在目标2中，我们将使用一种新的抗PD-L1小鼠模型 我们开发的诱导性甲状腺炎测试：(1)炎性细胞因子如何影响抗PD-L1的发展 诱导的甲状腺炎；以及(2)在这个小鼠模型中，通过恢复 甲状腺细胞的自噬。综上所述，我们提出了一种治疗ICI甲状腺炎的“两次打击”模型，即PD-PD- L1阻断可触发：(1)激活甲状腺内T细胞和炎性细胞因子分泌；(2) 甲状腺细胞内信号的失调使其容易受到炎性细胞因子的损害。在……里面 在这项建议中，我们将剖析PD-L1在甲状腺细胞中的非免疫作用，并扩大对 抗PD-L1免疫治疗的“脱靶”效应。我们的研究有望帮助设计有针对性的机制- 针对ICI相关的甲状腺和其他器官不良事件的基础治疗。

Project Summary/Abstract Programed death ligand-1 (PD‐L1) is an immune checkpoint inhibitor (ICI) that interacts with its receptor, programmed death‐1 (PD‐1) to inhibit T‐cell activation. Cancer cells adapted to manipulate this mechanism, by overexpressing PD-L1 to evade immune activation and attack. Therefore, abrogation of PD-L1/PD-1 interaction by monoclonal antibodies (MAb’s) is an effective therapeutic strategy for several cancers. However, it is complicated by immune related adverse events (irAEs), including endocrine AEs (eAEs). Thyroid dysfunction, including destructive thyroiditis and hypothyroidism, are the most common eAEs associated with PD-L1/PD-1 blockade, affecting up to 21% of patients. Because their mechanisms are not known, currently these conditions are treated symptomatically and as a result, patients are often difficult to manage, and the eAEs can interfere with the cancer treatment. Therefore, new therapies are needed that target the mechanisms causing eAEs. We have found that PD-L1, expressed in the thyroid, exerts intrinsic functions in thyroid cells. We have generated human thyroid cell lines with either downregulated or increased PD-L1 expression and demonstrated that PD- L1 engages cell survival mechanisms in response to inflammatory cytokine-induced stress. Suppression of PD- L1 in thyroid cells rendered them more susceptible to cellular stress and apoptosis induced by inflammatory cytokines. The intrinsic functions of PD-L1 in thyrocytes and their role in the development of ICI thyroiditis have not been studied before. In this proposal we will dissect the immune-independent functions of PD-L1 in thyroid cells and their role in the development of ICI-thyroiditis. Our hypothesis is that blockade of PD-L1 in thyroid cells through anti-PD-L1 therapies, triggers dysregulation of pro-survival intracellular signals controlled by PD-L1, rendering thyrocytes vulnerable to cytokine mediated destruction. In Aim 1 we will perform cellular and molecular studies in thyroid cell lines with upregulated and downregulated PD-L1 expression to dissect the intracellular functions of PD-L1 by assessing: (1) Molecular mediators of PD-L1 intrinsic activity (e.g., signals delivered through the PD-L1 intracytoplasmic domain); (2) The role of PD-L1 in cellular responses to inflammatory stress; and (3) The downstream effectors of PD-L1 activity. In Aim 2 we will use a new mouse model of anti-PD-L1 induced thyroiditis we developed to test: (1) How inflammatory cytokines affect the development of anti-PD-L1 induced thyroiditis; and (2) The possibility of reversing/suppressing thyroiditis in this mouse model by restoring autophagy in thyroid cells. In summary, we are proposing a “two-hit” model for ICI thyroiditis whereby PD- L1 blockade triggers both: (1) Activation of intra-thyroidal T-cells & inflammatory cytokine secretion; and (2) Dysregulation of thyrocyte intra-cellular signals making them susceptible to inflammatory cytokine damage. In this proposal we will dissect the nonimmune roles for PD-L1 in thyroid cells and extend the understating of the “off-target” effects of anti-PD-L1 immunotherapy. Our studies will hopefully help design targeted mechanism- based therapies for ICI associated adverse events in the thyroid and other organs.