Molecular ontology of drug tolerant persisters in HER2 positive breast cancer - Resubmission - 1

针对“驱动癌基因”的“靶向治疗”在许多肿瘤的治疗中发挥关键作用， 包括由编码受体酪氨酸激酶HER 2的基因扩增引起的乳腺癌 (HER2+ BC）。然而，在转移性HER 2 + BC中，甚至对抗癌药物组合也发生耐药性。 药物，并防止其出现是必不可少的转化缓解治愈。“耐药坚持者” （DTP）在暴露于酪氨酸激酶抑制剂（TKI）和其他抗肿瘤药物后， 静止（休眠）状态通过可逆的，非遗传机制。DTP与耐药性有关， EGFRmut非小细胞肺癌（NSCLC）、BRAFmut黑色素瘤和其他癌症，但在 在HER 2 + BC中的作用尚未彻底探索。我们发现用HER 2-TKI治疗的HER 2 + BC细胞系 （拉帕替尼、图卡替尼）产生两种类型的DTP，其具有管腔或间充质转录组。作为 在其他细胞系中，一部分HER 2 + BC细胞在退出有丝分裂后通过G 0期随机转运， 直接进入G1。值得注意的是，管腔DTP独特地产生于这些瞬时G 0细胞（“前G 0细胞”）。 DTP”），其也表达DTP基因的子集。HER 2 + BC是高度增殖的，但与其他肿瘤一样， 他们也有大量的G 0（Ki 67-）细胞，我们对TKI治疗的样本进行了初步分析， HER 2 + BC患者符合从G 0细胞产生DTP。因此，与目前的模型相比， 认为药物耐受性，静止状态是由TKI和其他靶向药物诱导的。 我们的数据表明，前DTP可能已经存在于药物初治患者中， 人口 我们假设这种短暂的G 0/前DTP细胞的染色质具有全局不同的 组织，引发他们诱导完整的DTP转录组，并成为耐药后， TKI暴露。这种“G 0选择/诱导”模型是否适用于间充质样HER 2 + DTP、DTP 由其他药剂诱导的，和/或更广泛地至真正的HER 2肿瘤的恶性肿瘤仍然不清楚。因为DTPs 包括潜在的细胞库，用于接种对HER 2 TKI和其他 靶向/常规疗法，描绘其个体发生机制可能揭示新的癌症 脆弱性和最终的新疗法我们专注于G 0细胞中基因调控的独特特征 也可能与肿瘤休眠有关。该提案加入了表观遗传学/染色体专家的行列 SKOK和癌症生物学/细胞信号传导（NEEL）来解决这些及时的相关问题 具体而言，我们将：（1）表征HER 2-TKI pre-DTP中的染色质结构和基因调控， （2）检验来自间充质细胞的HER 2-TKI DTP的假设， HER 2 + BC系也来自G 0样pre-DTP，并具有缩合蛋白II组织的染色质，和（3）测试 拉帕替尼DTP在HER 2-TKI DTP体内模型中的病理生理学意义。

SUMMARY: “Targeted therapies” against “driver oncogenes” play key roles in the therapy of many tumors, including breast cancers caused by amplification of the gene encoding the receptor tyrosine kinase HER2 (HER2+ BC). In metastatic HER2+ BC, however, resistance occurs even to combinations of anti-cancer drugs, and preventing its emergence is essential for converting remissions to cures. “Drug-tolerant persisters” (DTPs) survive exposure to tyrosine kinase inhibitors (TKIs) and other anti-neoplastics in a non-proliferative, quiescent (dormant) state via reversible, non-genetic mechanisms. DTPs are implicated in resistance of EGFRmut non-small cell lung cancer (NSCLC), BRAFmut melanoma, and other cancers, but a potential role in in HER2+ BC had not been explored thoroughly. We found that HER2+ BC lines treated with HER2-TKIs (lapatinib, tucatinib) give rise to two types of DTPs, which have luminal or mesenchymal transcriptomes. As in other cell lines, a fraction of HER2+ BC cells transit stochastically through G0 after exiting mitosis, instead of proceeding directly into G1. Remarkably, luminal DTPs arose uniquely from these transient G0 cells (“pre- DTPs”), which also express a subset of DTP genes. HER2+ BCs are highly proliferative, yet like other tumors, they also have significant numbers of G0 (Ki67-) cells, and our initial analyses of samples from TKI-treated HER2+ BC patients comport with DTP generation from G0 cells. Thus, in contrast to current models, which contend that the drug-tolerant, quiescent state is induced by TKIs and other targeted therapies/chemotherapies, our data indicate that pre-DTPs might already be present in the drug-naïve population. We hypothesize that the chromatin of such transient, G0/pre-DTP cells has a globally distinct organization that primes them to induce the complete DTP transcriptome and become drug-tolerant upon TKI exposure. Whether this “G0 selection/induction” model applies to mesenchymal-like HER2+ DTPs, DTPs induced by other agents, and/or more broadly to bona fide HER2 tumors remains unclear. Because DTPs comprise a potential cellular reservoir for seeding stable resistance to HER2 TKIs and other targeted/conventional therapies, delineating their ontogenetic mechanisms could reveal novel cancer vulnerabilities and ultimately, new therapies. Our focus on the unique features of gene regulation in G0 cells could also have implications for tumor dormancy. This proposal joins experts in epigenetics/chromosome organization (SKOK) and cancer biology/cell signaling (NEEL) to address these timely, relevant questions Specifically, we will: (1) Characterize chromatin architecture and gene regulation in HER2-TKI pre-DTPs and DTPs from luminal-like HER2+ BC lines, (2) Test the hypothesis that HER2-TKI DTPs from mesenchymal HER2+ BC lines also arise from G0-like pre-DTPs and have condensin II-organized chromatin, and (3) Test the pathophysiological significance of lapatinib DTPs in in vivo models of HER2-TKI DTPs.