Targeting Lung Tissue Resident NK Cells with Inhaled IL-15 and TIGIT Blockade in Osteosarcoma

项目总结 细胞因子免疫疗法，如IL-2和最近的IL-15，已被用于刺激内源性 细胞毒性自然杀伤(NK)和T细胞对晚期癌症的反应和支持过继转移 淋巴因子激活的杀伤细胞。虽然一些患者已经取得了应答，但IL-2和IL-2的使用 15已受到适度益处的阻碍，存在严重全身毒性的风险，特别是在给予 静脉注射(IV)。与静脉注射不同，吸入IL-15疗法具有集中释放的优点 在限制全身暴露和潜在毒性的同时，对肺部这一经常发生严重转移性疾病的部位进行了监测。 骨肉瘤(OSA)是一种侵袭性的原发骨癌，有很高的肺转移倾向，并且 在过去的40年里，肺转移患者的预后一直停滞不前。在我们最近完成的 (手稿准备中)吸入重组人白介素15治疗犬自发性肺转移的首次同类试验 OSA和黑色素瘤，我们观察到37%的临床益处(1个完全缓解，1个部分缓解，5个稳定 19只狗在60天内患病)。在这里，我们建议评估吸入反应的免疫机制。 IL-15在小鼠和人转移性OSA模型中的表达，假设一种独特的组织驻留于NK 肺部的群体具有高度的细胞毒性和增殖反应，与记忆- 类似于能够增强抗肿瘤反应的表型。在目标1中，我们将分析肺部居民NK K7M2和LM8阻塞性睡眠呼吸暂停综合征模型中的细胞(CD3-NKp46+CD49a+)，假设吸入了由 雾化吸入可刺激NK介导的抗OSA肺转移的抗肿瘤反应。匹配的人类 接受转移切除术的OSA患者的血液、非肿瘤肺和肿瘤样本将被 用于验证这些小鼠研究，特别是人类肺NK细胞表现出高度增强的特征 细胞毒性和效应功能可用于侵袭免疫抑制的OSA转移瘤 吸入IL-15的设置。在目标2中，我们将评估联合治疗。T细胞免疫受体与 免疫球蛋白和免疫受体酪氨酸抑制基序结构域(TIGIT)已成为一种关键的 NK细胞耗竭的标志，它限制了多种癌症类型的抗肿瘤反应，包括肉瘤。 因此，我们将联合吸入IL-15和使用抗TIGIT单抗的TIGIT阻断来评估NK 表型和功能、向肺肿瘤的转移、持久性和治疗后肿瘤的消退 老鼠模型。考虑到联合免疫治疗的可能性，毒性也将被仔细评估 引发免疫病理和细胞因子风暴。转移性阻塞性睡眠呼吸暂停综合征是一种罕见且难以治疗的癌症。尽管 在其他癌症中取得巨大成功，针对PD-1、PD-L1和CTLA-4的免疫治疗显示出极小的疗效 在OSA，强调了对这些侵袭性癌症进行新的免疫疗法的必要性。这项提议将 评估使用吸入IL-15刺激肺部组织驻留NK细胞的新策略，以最大限度地提高 作用部位的治疗性暴露，同时限制全身暴露和毒性。

PROJECT SUMMARY Cytokine immunotherapies, such as IL-2 and more recently IL-15, have been used to stimulate endogenous cytotoxic natural killer (NK) and T cell responses against advanced cancers and to support adoptively transferred lymphokine-activated killer cells. While responses have been achieved in some patients, the use of IL-2 and IL- 15 has been hampered by modest benefits with the risk of severe systemic toxicity, particularly when given intravenously (IV). Distinct from IV delivery, inhaled IL-15 therapy offers the advantages of concentrated delivery to the lungs, a frequent site of gross metastatic disease, while limiting systemic exposure and potential toxicity. Osteosarcoma (OSA) is an aggressive primary bone cancer with a high propensity for lung metastases, and outcomes for patients with lung metastases have been stagnant for the past 4 decades. In our recently completed (manuscript in preparation) first-in-kind trial of inhaled rhIL-15 in dogs with spontaneous lung metastases from OSA and melanoma, we observed clinical benefit in 37% (1 complete response, 1 partial response, and 5 stable disease at 60 days in 19 dogs). Here, we propose to evaluate the immune mechanisms of response to inhaled IL-15 in mouse and human models of metastatic OSA, hypothesizing that a unique tissue resident NK population in the lungs has heightened cytotoxic and proliferative responses consistent with a memory- like phenotype which enables augmented anti-tumor responses. In Aim 1, we will analyze lung resident NK cells (CD3-NKp46+CD49a+) in K7M2 and LM8 models of OSA, hypothesizing that inhaled IL-15 delivered by nebulizer will stimulate NK-mediated anti-tumor responses against OSA lung metastases. Matched human blood, non-tumor-bearing lung, and tumor samples from patients with OSA undergoing metastasectomy will be used to validate these mouse studies, in particular that human lung NK cells show features of heightened cytotoxicity and effector function which can be harnessed to infiltrate immunosuppressive OSA metastases in the setting on inhaled IL-15. In Aim 2, we will evaluate combination therapy. T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domains (TIGIT) has emerged as a critical marker of NK cell exhaustion which limits anti-tumor responses in multiple cancer types, including sarcomas. Thus, we will combine inhaled IL-15 with TIGIT blockade using anti-TIGIT monoclonal antibodies to assess NK phenotype and function, trafficking to lung tumors, persistence, and tumor regression following treatment in mouse models. Toxicity will also be carefully assessed given the potential for combination immunotherapy to precipitate immunopathology and cytokine storm. Metastatic OSA is a rare and difficult-to-treat cancer. Despite great success in other cancers, immunotherapy targeting PD-1, PD-L1, and CTLA-4 has shown minimal efficacy in OSA, underscoring the need for novel immunotherapies for these aggressive cancers. This proposal will evaluate novel strategies to stimulate tissue-resident NK cells in the lungs using inhaled IL-15 to maximize the therapeutic exposure at the site of action while limiting systemic exposure and toxicity.