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） 在放射治疗后增强癌细胞杀伤同时增加抗肿瘤免疫应答 （XRT）. DNA损伤反应（DDR）是一种整合DNA修复途径的信号系统， 细胞周期以保护基因组稳定性。除了激活细胞周期检查点和细胞中的DNA修复外， 用XRT处理后，DDR限制了无应激细胞中的起源放电并延迟了细胞周期转变。而细胞周期蛋白- 依赖性激酶是细胞周期加速器，DDR激酶是细胞周期制动器，并且在这种类比中，DDRi 使刹车失灵导致加速失控在这里，我们将确定如何在CD 8 + T中重新布线DDR 细胞适应大量的和伴随的DNA复制和转录在S期。我们还将 确定DDRi在癌症和免疫细胞中的影响。我们假设ATR激酶抑制剂诱导 导致核糖核苷被错误掺入基因组的起源点火，这产生了嵌合的 XRT后RNA-DNA片段和I型IFN依赖性免疫记忆为了验证我们在癌症中的假设 和免疫细胞，我们已经创造了一个创新的癌症移植模型。Mcm4Chaos3/Chaos3 小鼠携带Mcm 4中的突变，该突变使复制解旋酶不稳定。来源于Mcm 4Chaos 3/Chaos 3的细胞 老鼠的原产地许可减少了60%。我们已经产生了Mcm 4Chaos 3/Chaos 3 B16癌细胞， 移植到Mcm 4 wt/wt和Mcm 4Chaos 3/Chaos 3小鼠中。这将使我们能够分离ATR的功能， 限制了癌症和免疫细胞中介导复制叉修复的起始点。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诱导DNA损伤， 体外细胞在目标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抑制剂是否 联合收割机与XRT结合，通过对免疫和/或 癌细胞该项目的成功完成将定义DDR如何在CD 8 + T细胞中重新布线， 加速细胞周期转换并适应大量和伴随的DNA复制和转录 S期占细胞周期的70%左右，G1期缩短。这些研究非常重要， 检查点阻断和过继性T细胞转移的目的是诱导CD 8 + T细胞的快速分裂。 该项目的成功完成将确定增强癌症的DDRi组合和序列 细胞杀伤，同时增加抗肿瘤免疫应答 XRT。这些研究非常重要，因为我们使用的DDRi目前正在115项临床试验中，而XRT 用于治疗>50%的癌症患者，>60%具有治愈目的。

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.