Transcriptional Regulation of Peripheral Immune Cells as a Modulator of Melanoma Immunotherapy Resistance and Toxicity

项目摘要黑色素瘤是美国第五大最常见的癌症，虽然早期- 分期黑色素瘤是有利的，晚期疾病长期以来被认为是不治之症。免疫检查点抑制剂 (ICIS)使转移性黑色素瘤的治疗发生了革命性的变化；然而，仍有约40%的患者没有反应 在接受ICI治疗的患者中，约80%的患者发生免疫相关不良事件(IrAEs)。识别 可靠的治疗前生物标记物对患者进行分层以达到最大疗效同时将毒性降至最低是当务之急 临床任务，因为现有的肿瘤或肿瘤/免疫替代物有限。最近的研究表明，基线 宿主免疫由影响T细胞激活和分化的特定调控网络控制， 包括CD8+T细胞(ICI的主要靶点)。积累的数据表明，调控基因组( 由转录因子、增强子和其他元件(如lncRNAs)组成的调节组)控制着 细胞毒性T细胞的转录重编程，在宿主抗肿瘤免疫反应中起关键作用。 我们和其他人已经报道，调节组通过与以下相关的潜在遗传变异而丰富 自身免疫风险，这种遗传易感性与ICI的结果相关。基于这些耐人寻味 根据观察结果，我假设外周免疫细胞的基线“调节体”状态是由 遗传基因组和转录因子的相互作用，代表了ICI抗性和 对irAEs的敏感性。我推测这些转录调控网络(TRN)控制着T细胞的活动， 潜在地揭示了降低毒性和增强ICI疗效的生物途径。对于F99阶段，我将 建立一种获取治疗前外周CD8+T细胞(前pCD8+)活性的基因组方法 ICI治疗后转移性黑色素瘤患者ICI疗效的替代性研究 百时美施贵宝领导的一项大型临床试验的患者。我会辨认转录签名 与前pCD8+细胞的ICI结果相关；单细胞rna-seq将提供对免疫细胞的洞察 ICI上下文中的子类型(目标1.1)。结合转座酶可及染色质(ATAC)-seq的检测， RNA序列和全基因组测序，我将确定CD8+特异性调节组TRN为ICI的调节器 晚期黑色素瘤患者的预后(目标1.2)。对于K00阶段，我将把这个假设扩展到 探索治疗前后单细胞水平的变化，与特定免疫细胞的变化相关 有ICI反应的类型和使用黑色素瘤患者PBMC的irAEs(目标2)。我还将开发模型链接 ICI相关的单细胞PBMC细胞类型和具有特定肿瘤新表位的转录特征 治疗靶向。使用这些生物标记物识别最有可能对ICI有反应的患者也可能指向 为更有效和更低毒性的治疗提供新的生物途径，进一步改善ICI患者 黑色素瘤和潜在的其他癌症的结果。

PROJECT SUMMARY Melanoma is the fifth most common cancer in the US and while the prognosis for early- stage melanoma is favorable, late-stage disease was long considered incurable. Immune checkpoint inhibitors (ICIs) have revolutionized metastatic melanoma treatment; nevertheless, ~40% of patients still do not respond to therapy and ~80% of ICI-treated patients experience immune-related adverse events (irAEs). Identifying reliable pre-treatment biomarkers stratifying patients for maximal efficacy while minimizing toxicity is an urgent clinical task, as existing tumor or tumor/immune surrogates are limited. Recent research suggests that baseline host immunity is controlled by specific regulatory networks impacting T-cell activation and differentiation, including CD8+ T cells (the primary targets of ICI). Accumulated data show that the regulatory genome (the “regulome”), composed of transcription factors, enhancers, and other elements (e.g. lncRNAs), controls the transcriptional reprogramming of cytotoxic T-cells, playing a critical role in host anti-tumor immune response. We and others have reported that the regulome is enriched by underlying genetic variation associated with autoimmune risk and that this genetic susceptibility associates with ICI outcomes. Based on these intriguing observations, I hypothesize that baseline “regulome” states of peripheral immune cells, determined by the interplay of inherited genomic and transcriptomic factors, represent novel biomarkers of ICI resistance and susceptibility to irAEs. I postulate that these transcriptional regulatory networks (TRNs) govern T cell activity, potentially revealing biological pathways to reduce toxicity and enhance ICI efficacy. For the F99 phase, I will develop a genomic approach capturing TRNs of pre-treatment peripheral CD8+ T-cell (pre-pCD8+) activity as surrogates of ICI efficacy in metastatic melanoma using samples from ICI-treated metastatic melanoma patients from a large clinical trial led by Bristol-Myers Squibb. I will identify transcriptional signatures associated with ICI outcomes in pre-pCD8+ cells; single-cell RNA-seq will offer insight into immune cell subtypes in the ICI context (Aim 1.1). Combining Assay for Transposase-Accessible Chromatin (ATAC)-seq, RNA seq, and whole-genome sequencing, I will determine CD8+-specific regulome TRNs as modulators of ICI outcomes in advanced melanoma patients (Aim 1.2). For the K00 phase, I will expand this hypothesis to explore pre- and post-treatment changes at the single-cell level, correlating alterations in specific immune cell types with ICI response and irAEs using melanoma patient PBMCs (Aim 2). I will also develop models linking ICI-associated single-cell PBMC cell types and transcriptomic signatures with specific tumor neo-epitopes for therapeutic targeting. Identifying patients most likely to respond to ICI using these biomarkers may also point to novel biological pathways for more efficacious and less toxic treatments, further improving ICI patient outcomes in melanoma and potentially other cancers.