PERSONALIZED THERAPY FOR p16-DEFICIENT MELANOMA

项目摘要 仅在美国，黑色素瘤每年就造成超过10，000例死亡，并且目前针对黑色素瘤的治疗方法是： 晚期黑素瘤不能提供持久临床益处。因此，改进的分子 对黑色素瘤的了解对于开发有效的疗法是至关重要的。根据癌症基因组 Atlas（TCGA）黑色素瘤数据超过45%的黑色素瘤缺乏肿瘤抑制因子p16。然而，没有具体的 为p16缺陷型黑色素瘤提供个性化治疗选择。因此，我们进行了一次创新的 大规模可药用基因组RNAi筛选并鉴定细胞分裂周期7（CDC 7）激酶为候选物 这是p16缺陷型黑色素瘤生存所必需的。shRNA诱导的CDC 7敲低被抑制 体内p16缺陷型黑色素瘤肿瘤和转移。我们的中心假设是p16缺陷 黑色素瘤的存活依赖于CDC 7，并且CDC 7抑制对于治疗黑色素瘤具有治疗价值。 p16缺陷型黑素瘤总体目标是确定遗传和药理学的影响， CDC 7对黑色素瘤生长和转移的抑制作用，并了解为什么CDC 7对于 p16缺陷型黑素瘤具体而言，在目标1中，我们将确定遗传和药理学的影响， CDC 7对黑色素瘤生长和转移的抑制。为此，将确定CDC 7在 p16缺陷背景下的肿瘤生长和转移。此外，使用XL 413，小分子 CDC 7抑制剂，单独或与BRAFV 600 E抑制剂维罗非尼组合（用于黑色素瘤共携带 BRAFV 600 E突变和p16缺陷），我们将确定单独的CDC 7的药理学抑制是否 或与BRAFV 600 E抑制剂组合可用于有效治疗P16缺陷型黑素瘤。 vivo.该实验方法将利用黑色素瘤肿瘤生长和转移的小鼠模型， 利用从耶鲁大学SPORE获得的已建立的和黑素瘤患者来源的短期黑素瘤培养物， 皮肤癌在目标2中，我们将确定为什么CDC 7对p16缺陷型黑色素瘤的生存是必要的。 使用全转录组基因表达谱和串联亲和纯化和质谱- 基于蛋白质组学方法，我们发现p16缺陷的黑色素瘤细胞显示未折叠的激活， CDC 7可能通过抑制蛋白质应答（UPR）途径而维持UPR途径的活性。 转录因子HOXA 1促进黑色素瘤肿瘤生长和转移。因此，我们将确定 CDC 7如何通过HOXA 1调节p16缺陷黑素瘤细胞中的UPR通路。此外，我们将测试 CDC 7缺失诱导的p16缺陷型黑色素瘤细胞的肿瘤生长和转移抑制是否可以 通过恢复HOXA 1表达和UPR通路活性来拯救。总的来说，我们希望能发现一个 一种新的CDC 7驱动的可药物化的遗传脆弱性途径，可以靶向有效的个性化治疗。 p16缺陷型黑色素瘤的治疗，包括高度侵袭性转移性黑色素瘤。

Project Summary Melanoma account for over 10,000 deaths annually in the United States alone and current therapies for advanced-stage melanoma do not provide durable clinical benefit. Therefore, improved molecular understanding of melanoma is essential for developing effective therapies. According to The Cancer Genome Atlas (TCGA) melanoma data over 45% melanoma lack tumor suppressor p16. However, there is no specific personalized treatment option available for p16-deficient melanoma. Therefore, we performed an innovative large-scale druggable genome RNAi screen and identified Cell division cycle 7 (CDC7) kinase as a candidate that is necessary for the survival of p16-deficient melanoma. shRNA-induced knockdown of CDC7 inhibited p16-deficient melanoma tumors and metastases in vivo. Our central hypothesis is that p16-deficient melanoma depends upon CDC7 for their survival and CDC7 inhibition will be of therapeutic value for treating p16-deficient melanoma. The overall objective is to determine the effect of genetic and pharmacological inhibition of CDC7 on melanoma tumor growth and metastasis and understand why CDC7 is necessary for p16-deficient melanoma. Specifically, in Aim 1, we will determine the effect of genetic and pharmacological inhibition of CDC7 on melanoma tumor growth and metastasis. To this end, will determine the role of CDC7 in tumor growth and metastasis in the context of p16-deficiency. Additionally, using XL413, a small molecule inhibitor of CDC7, alone or in combination with BRAFV600E inhibitor vemurafenib (for melanoma co-harboring both BRAFV600E mutation and p16-deficiency), we will determine if pharmacological inhibition of CDC7 alone or in combination with BRAFV600E inhibitors can be employed to effectively treat p16-deficient melanoma in vivo. The experimental approach will utilize mouse models of melanoma tumor growth and metastasis and will utilize established and melanoma patient-derived short-term melanoma cultures available from Yale SPORE in Skin Cancer. In Aim 2, we will determine why CDC7 is necessary for the survival of p16-deficient melanoma. Using transcriptome-wide gene expression profiling and Tandem Affinity Purification and mass spectrometry- based proteomics approaches, we find that the p16-deficient melanoma cells show activation of Unfolded Protein Response (UPR) pathway and that CDC7 maintains the activity of UPR pathway potentially via transcription factor HOXA1 to promote melanoma tumor growth and metastasis. Therefore, we will determine how CDC7 regulates UPR pathway via HOXA1 in p16-deficient melanoma cells. Additionally, we will test whether CDC7-loss induced tumor growth and metastasis inhibition of p16-deficient melanoma cells can be rescued by restoring HOXA1 expression and the UPR pathway activity. Collectively, we expect to uncover a novel CDC7-driven druggable genetic vulnerability pathway that can be targeted for effective personalized therapy of p16-deficient melanoma, including for highly aggressive metastatic melanoma.