In situ radioimmunotherapy to maximize the engagement of conventional type 1 dendritic cells against non-T cell-inflamed tumors

已知放射疗法（RT）对肿瘤细胞施加直接的细胞毒性作用;然而，最近的研究表明， 揭示其对肿瘤免疫原性的影响，从而影响RT的总体结果。 单独使用通常不足以克服免疫抑制性肿瘤微环境（TME）， RT的增强免疫刺激作用正在深入研究中。到目前为止，大部分的重点都是 在RT后进行免疫调节，特别是与免疫检查点抑制剂组合。之甚少 已知RT前TME的操作如何影响免疫原性和治疗效果， RT.越来越多的证据表明，依赖Batf 3的传统1型树突状细胞（cDC 1）很少 在肿瘤髓样区室中发现的这些细胞具有交叉呈递肿瘤相关的 抗原（TAA）的CD 8 + T细胞，并作为“主调节器”的T细胞反应在癌症。我们假设 原位诱导和激活cDC 1增强了RT的治疗效果和免疫原性。 为了验证这一假设，我们开发了一种组合的原位放射免疫疗法， 1）Flt 3L动员cDC 1至TME; 2）RT促进癌细胞的免疫原性死亡和成熟 和3）双重TLR 3/CD 40刺激以激活负载抗原的cDC 1，用于引发肿瘤特异性CD 8 + T细胞 细胞我们使用多个免疫原性差的肿瘤同基因原位小鼠模型的新数据 对抗PD-L1疗法不敏感揭示原位放射免疫疗法诱发从头适应性T细胞应答 其特征在于新的克隆型和干细胞样Tcf 1 + Slamf 6+表型， 抗PD-L1抗体，介导持久的完全反应，并产生肿瘤特异性全身性 免疫记忆令人信服的证据表明，RT的免疫原性可以通过原位免疫增强。 然而，原位放射免疫治疗对cDC 1的免疫调节作用 远处转移性肿瘤仍不清楚。cDC 1引发CD 4 + T细胞以及CD 8 + T细胞，但CD 4 + T细胞的作用与CD 8 + T细胞不同。 原位放射免疫治疗中的T细胞仍然难以捉摸。此外，仍不清楚是否在原地 放射免疫疗法克服了人非T细胞炎性肿瘤中的不良T细胞浸润。本提案中 我们将阐明CD 4 + T细胞在增强原位放射免疫治疗抗肿瘤疗效中的作用（目的 1）。此外，我们将寻求更好地了解免疫调节作用的机制， 原位放射免疫治疗靶向非放射性远处转移肿瘤（目标2）。在目标3中，我们将 寻求确定在患有不可切除和转移性乳腺癌的患者中人类TME的改变 用原位放射免疫疗法治疗。这些研究将增加必要的机械理解RT如何 和免疫系统的相互作用，并提供洞察原位放射免疫治疗的临床潜力 对抗对抗PD-L1治疗不敏感的非T细胞炎症肿瘤。

Radiation therapy (RT) is known to exert direct cytotoxic effects on tumor cells; however, recent research is revealing its influence on the immunogenicity of tumors, thus affecting the overall outcome of RT. While RT alone is usually insufficient to overcome the immunosuppressive tumor microenvironment (TME), strategies to boost immune-stimulating effects of RT are under intensive investigation. To date, most of the focus has been placed on immunomodulation after RT, in particular in combination with immune checkpoint inhibitors. Little is known about how manipulation of the TME before RT can impact on immunogenicity and therapeutic efficacy of RT. A growing body of evidence reveals that Batf3-dependent conventional type 1 dendritic cells (cDC1) rarely found within the tumor myeloid compartment have the important capacity of cross-presenting tumor-associated antigens (TAA) to CD8+ T cells, and act as `master regulators' for the T cell response in cancer. We hypothesize that in situ induction and activation of cDC1 enhances the therapeutic efficacy and immunogenicity of RT. To test this hypothesis, we developed a combinatorial in situ radioimmunotherapy comprised of in situ administration of: 1) Flt3L to mobilize cDC1 to the TME; 2) RT to promote immunogenic death of cancer cells and maturation of DC; and 3) dual TLR3/CD40 stimulation to activate antigen-loaded cDC1 for priming of tumor-specific CD8+ T cells. Our new data using multiple syngeneic orthotopic murine models of poorly immunogenic tumors insensitive to anti-PD-L1 therapy reveal that in situ radioimmunotherapy elicits de novo adaptive T cell responses that are characterized by novel clonotypes and stem-like Tcf1+ Slamf6+ phenotypes, renders tumors responsive to anti-PD-L1 antibody, mediates durable complete responses, and develops tumor-specific systemic immunological memory. Compelling evidence suggests that immunogenicity of RT can be enhanced by in situ induction and activation of cDC1; however, immunomodulatory effect of in situ radioimmunotherapy against distant metastatic tumors remains unclear. cDC1 prime CD4+ T cells as well as CD8+ T cells, but the role of CD4+ T cells in in situ radioimmunotherapy remains elusive. In addition, it remains unknown whether in situ radioimmunotherapy overcomes poor T-cell infiltration in human non-T cell-inflamed tumors. In this proposal we will elucidate the roles of CD4+ T cells in augmenting antitumor efficacy of in situ radioimmunotherapy (Aim 1). Additionally, we will seek to better understand the mechanisms underlying the immunomodulatory effect of in situ radioimmunotherapy targeting non-irradiated distant metastatic tumors (Aim 2). Finally, in Aim 3, we will seek to determine the alteration of the human TME in patients with unresectable and metastatic breast cancer treated with in situ radioimmunotherapy. These studies will add essential mechanistic understanding to how RT and the immune system interact, and provide insight into the clinical potential of in situ radioimmunotherapy against non-T cell-inflamed tumors insensitive to anti-PD-L1 therapy.