Tumor suppressor vulnerability conferred by aneuploid loss of haploinsufficient metallothionein genes

摘要 癌症很少通过单基因药物靶向治疗。肿瘤的进化是对 选择压力精确肿瘤学通常针对单个基因产物，而该基因只是成为 会发生变异虽然个别基因可能会发生突变，但肿瘤生物学仍然是 限制于每种癌症类型的特定通路失调。我们以前创造了尖端的 生物信息学工具，以更好地了解哪些受约束的途径作为肿瘤抑制因子， 癌基因，由于在分子途径水平的协同基因失调。非整倍体是导致 不幸的是，每一次非整倍体事件都改变了预测的驱动因素， 基因和未知乘客基因。肿瘤中非整倍体变化的原因研究仍然很困难，如果 用目前的细胞生物学和遗传学工具来研究并非不可能。不过，我们发现了一种独特的， 通常被抑制（70%的高级别浆液性卵巢癌有单等位基因丢失，与 平均减少的表达），这是服从良好控制的基础科学实验的途径： 镉反应途径它由11个高度同源的金属硫蛋白基因排列在一个单一的 染色体位点金属硫蛋白螯合大量的细胞内Zn 2+和环境遗传毒性Cd 2 + 离子。金属硫蛋白基因座的缺失与染色体不稳定性有关，并发生在肿瘤的早期 阵我们的体外试验表明，受控的金属硫蛋白抑制导致DNA损伤升高。 然而，金属硫蛋白作为肿瘤抑制因子和作为癌症细胞和分子调节因子的作用还不清楚。 生物学基本上是未知的。本项目将建立金属硫蛋白的特异性肿瘤抑制表型 并确定这种非整倍体途径是否将作为多倍体途径的代表性例子， 基因脆弱性可以更好地实现下一代癌症疗法。我们将（1）在体内表征 金属硫蛋白基因缺失在卵巢癌自发性肿瘤形成中的作用，（2）发展控制 基因抑制模型能够抑制所有11个基因，包括通过合成的基于dead-Cas9的 转录因子，（3）确定哪种镉依赖性和镉非依赖性金属硫蛋白- 受调节的分子途径传递肿瘤抑制功能，以及（4）发现最佳的药物类别， 选择性杀死低金属硫蛋白细胞。该项目创建的遗传工具将使因果调查成为可能。 为未来的项目提供完整的分子途径。综合来看，这项创新的研究计划将直接测试 非整倍体抑制途径如何促进肿瘤发生， 在整个肿瘤发展过程中药理学上可靶向的脆弱性。

Abstract Cancer will rarely be cured through pharmacologic targeting of single genes. Tumors evolve in response to selection pressure. Precision oncology often targets a single gene product, and that gene simply becomes mutated once the drug is administered. Although individual genes may mutate, tumor biology is nonetheless constrained to dysregulating specific pathways for each cancer type. We have previously created cutting-edge bioinformatic tools to better understand which constrained pathways are acting as tumor suppressors and oncogenes, due to collaborative gene dysregulation at the molecular pathway level. Aneuploidy is a major cause of molecular pathway changes in cancer and unfortunately each aneuploid event alters both predicted driver genes and unknown passenger genes. Investigation of causal aneuploid changes in a tumor remains difficult, if not impossible, to study with current cell biology and genetic tools. However, we have discovered a unique, commonly suppressed (70% of high-grade serous ovarian cancers have a monoallelic loss, correlating with average reduced expression), pathway which is amenable to well-controlled basic science experimentation: the cadmium response pathway. It is composed of 11 highly homologous metallothionein genes arrayed on a single chromosomal locus. Metallothioneins sequester the bulk of intracellular Zn2+ and environmental genotoxic Cd2+ ions. The loss of the metallothionein locus is associated with chromosome instability and occurs early in tumor formation. Our in vitro assays show controlled metallothionein suppression results in elevated DNA damage. However, the role of metallothioneins as tumor suppressors and as regulators of cancer cellular and molecular biology is largely unknown. This project will establish specific tumor suppressor phenotypes of metallothioneins in ovarian cancer and determine if this aneuploid pathway will serve as a representative example of how multi- genic vulnerabilities can better enable next-generation cancer therapies. We will (1) characterize in vivo the effects of metallothionein gene loss in spontaneous tumor formation in ovarian cancer, (2) develop controlled models of gene suppression enabling suppression of all 11 genes, including by a synthetic dead-Cas9-based transcription factor, (3) determine which cadmium-dependent and cadmium-independent metallothionein- regulated molecular pathways convey tumor suppressor functions, and (4) discover drug classes which best selectively kill low-metallothionein cells. Genetic tools created by this project will enable causal investigation of entire molecular pathways for future projects. Taken together, this innovative research program will directly test how an uncharacterized aneuploid-suppressed pathway contributes to oncogenesis and remains a pharmacologically targetable vulnerability throughout tumor development.