Optimal BET inhibitor combination therapies in triple negative breast cancer

项目摘要 三阴性乳腺癌（TNBC）是乳腺癌的一种主要亚型，其与通常的不良预后相关。 结果，诊断时年龄较小，预后比其他亚型差。它目前没有选择， 靶向治疗和化疗仍然是唯一的药理学选择;因此，更好的治疗是 迫切需要。我们最近证明了一类表观遗传靶向药物BET的潜力， 抑制剂，作为TNBC中有前途的新疗法。然而，迅速发展的耐药性需要 进一步研究，以确定如何有效地管理这些药物，并防止耐药性。治疗 已知原型BET抑制剂JQ 1诱导细胞状态表型变化，其可 介导阻力。TNBC中高度的肿瘤内异质性也可能有助于发展 抗药性和治疗失败。因此，需要更好地了解肿瘤群体是如何 受选择性治疗压力的影响。此外，有效的联合治疗必须 制定并优化其管理，以实现更持久的反应。我假设 紫杉醇化疗和PD-L1免疫治疗的双联和三联治疗，以及 组合给药的顺序将对治疗功效和肿瘤发生率产生影响。 进化在目标1中，我将研究响应于JQ 1联合治疗的群体动力学， 紫杉醇，使用用高复杂性DNA文库条形码化的TNBC细胞系来追踪单个亚克隆 在治疗期间，在培养物和免疫缺陷小鼠的异种移植物中。在目标2中，我将研究最佳 通过在异种移植物中测试两种顺序的伴随和序贯疗法来施用JQ 1与紫杉醇 用条形码细胞，以评估治疗方案之间的群体动力学差异， 单细胞RNA-seq的细胞反应差异。然后，我将使用EvoSeq，一种分离细胞的方法， 在具有条形码水平特异性的群体内，以检索具有差异选择的条形码的细胞， 治疗时间表，并检查他们是否易患耐药性，以及是否预处理 一种药物改变了对第二种药物的敏感性。在目标3中，我将描述异质性的变化， 从JQ 1、紫杉醇和抗PD-L1抗体的三联组合中，在测试所有单药后， 在免疫活性小鼠中，MMTV-PyMT细胞中的三重组合和三重组合。本研究结果 将导致更深入地了解肿瘤内异质性如何改变治疗反应，并建立 如何最佳地组合联合收割机BET抑制剂。这项研究也将有直接的翻译影响，在通知 临床试验的合理设计，如果成功，将导致TNBC患者的生存率提高。

PROJECT SUMMARY Triple negative breast cancer (TNBC) is a major subtype of breast cancer that is associated with generally poor outcome, younger age at diagnosis, and worse prognosis than other subtypes. It currently has no options for targeted therapy, and chemotherapy remains the only pharmacologic option; thus, better treatments are urgently needed. We recently demonstrated the potential of a class of epigenetic targeted agents, BET inhibitors, as a promising new therapy in TNBC. However, the rapid development of resistance necessitates further study to determine how to effectively administer these drugs and to prevent resistance. Treatment with the prototypical BET inhibitor, JQ1, is known to induce phenotypic changes in cellular state, which may mediate resistance. The high degree of intratumor heterogeneity in TNBC may also contribute to development of resistance and treatment failure. Therefore, a better understanding is needed of how tumor populations are affected by the selective pressures of treatment. Furthermore, effective combination therapies must be developed and their administrations optimized in order achieve a more durable response. I hypothesize that double and triple combination therapies with paclitaxel chemotherapy and PD-L1 immunotherapy, as well as the order in which combinations are administered, will have an effect on treatment efficacy and tumor evolution. In Aim 1, I will investigate population dynamics in response to JQ1 combination therapy with paclitaxel, using TNBC cell lines barcoded with a high-complexity DNA library to track individual subclones during treatment, both in culture and in xenografts in immunodeficient mice. In Aim 2, I will investigate optimal administration of JQ1 with paclitaxel by testing concomitant and sequential therapy in both orders in xenografts with barcoded cells, in order to assess differences in population dynamics between treatment schedules and differences in cellular response with single cell RNA-seq. I will then use EvoSeq, a method to isolate cells from within a population with barcode-level specificity, to retrieve cells with differentially selected barcodes between treatment schedules and examine whether they are predisposed to resistance and whether pretreatment with one drug alters sensitivity to the second drug. In Aim 3, I will characterize changes in heterogeneity that result from the triple combination of JQ1, paclitaxel, and anti-PD-L1 antibody, after testing all single agents, double combinations, and triple combination in MMTV-PyMT cells in immunocompetent mice. The results of this study will lead to a deeper understanding of how intratumor heterogeneity modifies treatment response and establish how to optimally combine BET inhibitors. This research will also have direct translational impact in informing the rational design of clinical trials and, if successful, would lead to improved survival for patients with TNBC.