Mechanisms and small-molecule targeting of SWI/SNF activity in neuroblastoma

摘要 神经母细胞瘤(NB)是一种起源于神经脊细胞的侵袭性儿科恶性肿瘤。Nb是 是最常见的儿童实体肿瘤之一，37%的患者被诊断为婴儿。患者患有高- 尽管强化治疗可能导致几种转阴，但风险型NB的五年存活率约为50% 结果，包括低体重、听力损失和神经损伤、死亡或其他后遗症。为了提高 对于这些儿科患者的治疗，需要新的方法。SMARCA4(BRG1)，主要的ATPase SWI/SNF复合体，已被确定为对NB的致癌依赖性，具有频繁的基因扩增 在晚期疾病中。不幸的是，靶向SWI/SNF ATPase活性的适宜性和 它对NB的抑制作用仍然知之甚少。我们已经使用了新的快速反应工具来瞄准 SMARCA4在我们的初步数据中发现，它的失活导致了NB活力的严重丧失 独立于MYCN或其他细胞遗传学特征的细胞。我们发现SMARCA4的失活 在G1-S边界附近诱导细胞死亡，与复制应激或细胞周期失调一致。在这 我们试图：(1)定位SWI/SNF复合体对DNA的直接细胞周期特异性效应 可及性、转录和细胞死亡；(2)确定细胞周期放松调控和复制的机制 SWI/SNF抑制诱导的应激反应；以及(3)在体外鉴定肿瘤特异性协同药物组合，并 在活体内验证其疗效。我们将使用一组目前临床上使用的药物来治疗高危NB，以 在2D细胞和3D球体中检查SWI/SNF抑制的治疗协同作用，并检查它们的组合 在神经母细胞瘤免疫能力强的小鼠模型中。我们确定联合疗法的策略 会被告知一个带有人类神经脊和外周神经元的反屏幕，以识别药物 对非肿瘤细胞毒性最小的组合。我们的研究将确定主要途径 受SWI/SNF抑制的影响，NB细胞的复制应激和细胞周期失调有关。我们的发现将 此外，还可以识别和验证在体内加强SWI/SNF阻断的治疗方法。因为 小分子SWI/SNF抑制剂对小鼠有很好的耐受性，我们的翻译研究代表了一个重要的 临床前步骤，可能导致对50%无反应的高危NB患者进行临床试验。

Abstract Neuroblastoma (NB) is an aggressive pediatric malignancy originating from cells of neural crest origin. NB is among the most frequent pediatric solid tumors, with 37% of patients diagnosed as infants. Patients with high- risk NB have a five-year survival rate of ~50% despite intensive therapy that can cause several negative outcomes, including low body weight, hearing loss and nerve damage, death, or other sequelae. To improve treatment for these pediatric patients, new approaches are needed. SMARCA4 (BRG1), the primary ATPase of SWI/SNF complexes, has been identified as an oncogenic dependency for NB, with frequent gene amplifications in advanced disease. Unfortunately, the suitability of targeting SWI/SNF ATPase activity and the mechanisms of its inhibition in NB have remained poorly understood. We have employed new fast-acting tools to target SMARCA4 in our preliminary data, which revealed that its inactivation induces profound loss of viability of NB cells independently of MYCN or other cytogenetic features. We have found that inactivation of SMARCA4 induces cell death near the G1-S boundary, consistent with replication stress or cell-cycle dysregulation. In this proposal, we seek to: (1) Map the direct cell cycle-specific effects of SWI/SNF complexes towards DNA accessibility, transcription, and cell death; (2) Identify the mechanisms of cell cycle deregulation and replication stress induced by SWI/SNF inhibition; and (3) Identify tumor-specific synergistic drug combinations in vitro and validate their efficacy in vivo. We will employ a panel of drugs currently used clinically to treat high-risk NB, to examine therapeutic synergy with SWI/SNF inhibition in 2D cells, 3D spheroids, and examine their combination in a rigorous immune-competent mouse model of neuroblastoma. Our strategy to identify combination therapies will be informed by a counter-screen with human neural crest and peripheral neurons, to identify drug combinations that show minimal toxicity to non-tumor cells. Our studies will identify the principal pathways influenced by SWI/SNF inhibition in NB related to replication stress and cell cycle dysregulation. Our findings will furthermore enable identification and validation of therapies that potentiate SWI/SNF blockade in vivo. Because small-molecule SWI/SNF inhibitors are well tolerated by mice, our translational studies represent an important preclinical step that may lead to clinical trials for the 50% of unresponsive high-risk NB patients.