DNA damage response and cancer immunity

我们实验室的首要目标是确定如何使用dna损伤反应抑制剂(Ddri)。 在增强放射治疗后抗肿瘤免疫反应的同时增强对癌细胞的杀伤作用 (XRT)。DNA损伤反应(DDR)是一种信号系统，它整合了DNA修复途径和 细胞周期，以维护基因组的稳定性。除了激活细胞周期检查点和细胞中的DNA修复 经XRT处理后，DDR限制了非应激细胞的起始放电，并延迟了细胞周期的转变。当骑自行车的时候- 依赖的激酶是细胞周期加速器，DDR激酶是细胞周期的刹车，在这个类比中，DDRI 刹车失灵，加速失控。在这里，我们将确定如何在CD8+T中重新连接DDR 细胞在S期适应大量和伴随的DNA复制和转录。我们还将 确定DDRI对癌症和免疫细胞的影响。我们假设ATR激酶抑制剂能诱导 导致核糖核苷错误结合到基因组中并由此产生嵌合体的起源激发 XRT后RNA-DNA片段与I型干扰素依赖免疫记忆的关系在癌症中检验我们的假说 和免疫细胞，我们已经产生了一种创新的可移植癌症模型。Mcm4Chaos3/Chaos3 小鼠携带Mcm4突变，破坏复制解旋酶的稳定。Mcm4Chaos3/Chaos3来源的细胞 MICE的原产地许可减少了60%。我们已经产生了Mcm4Chaos3/Chaos3 B16癌细胞，可以 移植到Mcm4wt/wt和Mcm4Chaos3/Chaos3小鼠体内。这将允许我们将ATR的功能分离出来 限制起源于调节癌细胞和免疫细胞复制分叉修复的信号。在目标1中， 我们将定义细胞周期动力学，并确定ATR抑制剂如何在免疫和癌症中导致DNA损伤 体外培养的细胞。在目标2中，我们将定义细胞周期动力学，并确定ATR抑制剂是否诱导DNA 体内免疫细胞和1型干扰素的损伤。在目标3中，我们将确定ATR抑制剂 与XRT结合，通过影响免疫和/或产生持久反应和免疫记忆 癌细胞。这个项目的成功完成将定义DDR如何在CD8+T细胞中重新连接到 加速细胞周期转变，适应大量和伴随的DNA复制和转录 S期，G1期缩短，约占细胞周期的70%。这些研究具有重要意义，因为 检查点阻断和过继T细胞转移的目的是诱导CD8+T细胞快速分裂。 该项目的成功完成将确定DDRI的组合和序列，这些组合和序列会加剧癌症 小鼠肿瘤模型在增强抗肿瘤免疫应答的同时杀伤细胞 XRT。这些研究非常有意义，因为我们使用了目前正在进行的115项临床试验的DDRI和XRT，其中 用于治疗50%的癌症患者，60%的癌症患者有治疗意图。

The overarching goal of our laboratory is to determine how DNA damage response inhibitors (DDRi) can be used to potentiate cancer cell killing while concurrently increasing anti-tumor immune responses after radiation therapy (XRT). The DNA Damage Response (DDR) is a signaling system that integrates DNA repair pathways and the cell cycle to safeguard genome stability. In addition to activating cell cycle checkpoints and DNA repair in cells treated with XRT, the DDR limits origin firing and delays cell cycle transitions in unstressed cells. While cyclin- dependent kinases are cell cycle accelerators, DDR kinases are cell cycle brakes and, in this analogy, DDRi disable the brakes, causing unchecked acceleration. Here we will determine how the DDR is rewired in CD8+ T cells to accommodate massive and concomitant DNA replication and transcription in S phase. We will also determine the impact of DDRi in cancer and immune cells. We hypothesize that ATR kinase inhibitors induce origin firing that causes ribonucleosides to be mis-incorporated into the genome, and that this generates chimeric RNA-DNA fragments and type I IFN-dependent immunologic memory after XRT. To test our hypothesis in cancer and immune cells, we have generated an innovative transplantable model of cancer. The Mcm4Chaos3/Chaos3 mouse carries a mutation in Mcm4 that destabilizes the replicative helicase. Cells derived from Mcm4Chaos3/Chaos3 mice have a 60% reduction in origin licensing. We have generated Mcm4Chaos3/Chaos3 B16 cancer cells that can be transplanted into Mcm4wt/wt and Mcm4Chaos3/Chaos3 mice. This will allow us to separate the function of ATR that limits origin firing from that which mediates the repair of replication forks in cancer and immune cells. In Aim 1, we will define cell cycle kinetics and determine how ATR inhibitors induce DNA damage in immune and cancer cells in vitro. In Aim 2, we will define cell cycle kinetics and determine whether ATR inhibitors induce DNA damage in immune cells and type 1 interferons in vivo. In Aim 3, we will determine whether ATR inhibitors combine with XRT to generate durable responses and immunologic memory through effects on immune and/or cancer cells. Successful completion of this project will define how the DDR is rewired in CD8+ T cells to accelerate cell cycle transitions and accommodate massive and concomitant DNA replication and transcription in S phase which, accounts for ~70% of the cell cycle as G1 is abridged. These studies are highly significant as the objective of checkpoint blockade and adoptive T cell transfer is to induce rapid division in CD8+ T cells. Successful completion of this project will identify combinations and sequences of DDRi that potentiate cancer cell killing while concurrently increasing anti-tumor immune responses in mouse models of cancer treated with XRT. These studies are highly significant as we use DDRi that are currently in 115 clinical trials and XRT which is used to treat >50% of cancer patients, >60% with curative intent.