Cell type-directed Tim-3 targeting in melanoma

项目总结 免疫检查点阻断在转移性癌症患者中引起了前所未有的临床反应 黑色素瘤和其他癌症。在癌症治疗试验中一个有希望的新检查点是T- 细胞免疫球蛋白和粘蛋白结构域3(TIM-3)。TIM-3阻断逆转T细胞损伤，从而 重振抗肿瘤T细胞免疫。然而，我们发现TIM-3抑制剂，包括那些在临床上的 试验，不仅靶向T细胞-TIM-3，而且对树突状细胞(DC)上的TIM-3具有不同的亲和力， 巨噬细胞(MΦS)、NK细胞和黑色素瘤细胞。因此，临床上的益处可能不完全依赖于 T细胞-TIM-3，但也抑制这些额外的TIM-3表达的细胞类型。作为支持，封锁T- 细胞TIM-3被抑制，黑色素瘤导向的TIM-3抑制促进小鼠黑色素瘤的生长 模型，从而抵消了TIM-3治疗的预期疗效。一致地，强制表达TIM-3在 黑色素瘤细胞抑制肿瘤的发生、转移的形成和增殖途径的活性。我们的 因此，初步研究确定黑色素瘤细胞固有的、DC、MΦ和NK细胞TIM-3是意想不到的变量 和/或治疗结果的潜在混杂因素。他们进一步强调了定义治疗性的必要性 TIM-3抗体(Ab)在特定细胞类型水平上的反应后果。TIM-3蛋白熊 多种N-和O-糖结构，它们的组成、大小和电荷在细胞间差异很大 这可能解释了我们在细胞之间发现的Tim-3抗体克隆反应性的显著差异 类型。例如，TIM-3临床试验候选药物TSR-022与T细胞和黑色素瘤密切结合，但不是 NK、DC或MΦ-TIM-3，而其他TIM-3抗体对T细胞、MΦS、DC和/或NK具有高亲和力， 但不是黑色素瘤细胞。值得注意的是，糖链修饰方案将抑制物结合转移到所需的T细胞- TIM-3识别和黑色素瘤-TIM-3反应性降低。我们的初步数据强调了对 剖析免疫与黑色素瘤细胞-TIM-3糖基表位、抗体亲和力、信号和免疫生物学。结果 将通过验证优先针对免疫细胞的方案来优化TIM-3的治疗效果-TIM-3 多糖，同时避免不必要的黑色素瘤细胞-TIM-3的阻断。我们的目标是1)定义细胞类型- 相关的TIM-3糖链、配体、抗体亲和力和信号网络，2)检查免疫细胞与 黑色素瘤-现有TIM-3拮抗剂的内在效应及其与解释治疗益处的相关性， 以及3)确定新的TIM-3靶向策略，以加强免疫细胞-TIM-3抑制。我们将使用国家- 最新的TIM-3功能和糖链修饰策略的得失，具有可变组织结构的TIM-3抑制剂- 相关亲和力、免疫和黑色素瘤模型系统以确定特定细胞类型的TIM-3功能 和糖分子靶标。我们的倡议还实施了临床肿瘤生物显微镜从患者接受 免疫检查点抑制剂。总之，这些研究将为下一代生物标记物和 区分免疫和癌细胞的治疗方式-TIM-3，以获得最佳的免疫治疗结果。

PROJECT SUMMARY Immune checkpoint blockade has elicited unprecedented clinical responses in patients with metastatic melanoma and other cancers. A promising new checkpoint under investigation in cancer therapeutic trials is T- cell immunoglobulin and mucin domain 3 (Tim-3). Tim-3 blockade reverses T-cell impairment, thereby reinvigorating antitumor T-cell immunity. However, we found that Tim-3 inhibitors, including those in clinical trials, not only target T-cell-Tim-3, but also have varying affinity for Tim-3 on dendritic cells (DCs), macrophages (MΦs), NK and melanoma cells. Clinical benefit might thus not exclusively rely on antagonism of T-cell-Tim-3, but also on inhibition of these additional Tim-3-expressing cell types. In support, blockade of T- cell-Tim-3 suppressed, while melanoma-directed Tim-3 inhibition enhanced tumor growth in murine melanoma models, thereby counteracting desired efficacy of Tim-3 therapy. Consistently, enforced expression of Tim-3 on melanoma cells suppressed tumorigenesis, metastasis formation, and proliferative pathway activity. Our preliminary studies thus identify melanoma cell-intrinsic, DC-, MΦ-, and NK-cell-Tim-3 as unexpected variables and/or potential confounders of treatment outcome. They further highlight the need to define therapeutic consequences of Tim-3 antibody (Ab) responses at the level of specific cell types. The Tim-3 protein bears multiple N- and O-glycostructures that differ dramatically in composition, size, and charge between cell lineages and which might explain the marked variations in Tim-3 Ab clone reactivity we found between cell types. For example, the clinical Tim-3 trial candidate, TSR-022, avidly bound T-cell- and melanoma-, but not NK-, DC-, or MΦ-Tim-3, while other Tim-3 Abs showed high affinity for Tim-3 on T-cells, MΦs, DCs, and/or NK, but not melanoma cells. Notably, glycan-modifying regimens shifted inhibitor binding towards desired T-cell- Tim-3 recognition and reduced melanoma-Tim-3 reactivity. Our preliminary data highlights the critical need for dissecting immune- vs. melanoma cell-Tim-3 glyco-epitopes, Ab affinity, signaling, and immunobiology. Results will help optimize Tim-3 therapeutic efficacy by validating regimens that preferentially target immune cell-Tim-3 glycans, while avoiding unwanted blockade of melanoma cell-Tim-3. Our aims are to 1) define cell type- associated Tim-3 glycan moieties, ligands, Ab affinities, and signaling networks, 2) examine immune cell- vs. melanoma-intrinsic effects of existing Tim-3 antagonists and their relevance to interpreting therapeutic benefit, and 3) identify new Tim-3 targeting strategies that accentuate immune cell-Tim-3 inhibition. We will use state- of-the-art gain and loss of Tim-3 function and glycan-modifying strategies, Tim-3 inhibitors with variable tissue- associated affinities, and immune and melanoma model systems to define cell type-specific Tim-3 functions and glycomolecular targets. Our initiative also implements clinical tumor biospecimens from patients receiving immune checkpoint inhibitors. Together, these studies will pave the way for next generation biomarkers and treatment modalities that discriminate immune- from cancer cell-Tim-3 for optimized immunotherapy outcomes.