??PK mediated melanoma stem cell oncolysis through non-redundant programmed cell

描述(申请人提供)：黑色素瘤是第六种最常见的癌症，约占所有新诊断的恶性肿瘤的5%。这种发病率和死亡率高的癌症在很大程度上对现有的治疗方法具有抵抗力。这至少部分归因于癌症干细胞(CSC)，它们越来越被认为是复发和治疗耐药疾病的主要来源。溶瘤病毒是一种很有前途的新疗法，具有靶向CSC的潜力。早期临床试验表明，病毒疗法耐受性良好，但其疗效受到溶细胞活性差、肿瘤穿透不足以及无法在静止的肿瘤细胞中复制的限制。目前的努力试图改善复制，但基于诱导程序性细胞死亡(PCD)的病毒治疗策略尚不可用。单纯疱疹病毒(HSV)是一种特别有吸引力的溶瘤病毒，因为它具有广泛的宿主谱，其基因组不整合到宿主基因组中从而排除插入突变，可用抗病毒药物来防止不利的病毒复制，并且病毒逃避免疫，允许再次治疗。根据我们最近的发现，HSV-2基因ICP10PK(Delta PK)缺失的HSV-2突变株可导致黑色素瘤细胞死亡。黑色素瘤细胞中激活的RAS信号通路提供了选择性的Delta PK复制，但肿瘤分解是由多个非冗余的PCD通路介导的。这些途径包括钙蛋白酶和caspase活性增加，在黑色素瘤中具有肿瘤抑制样功能的表观遗传沉默基因H11上调，自噬相关蛋白Beclin 1的表达增强，以及炎症性caspase-1(热下垂)的激活。Delta PK是一种特别有前景的病毒疗法，因为它可以诱导Th1细胞，Th1细胞可以覆盖抑制黑色素瘤的环境，而且它在人类中的耐受性很好。拟议的研究试图更好地阐明Delta PK诱导/激活的死亡途径之间的关系，并确定它是否可以杀死黑色素瘤干细胞。其具体目的是：(1)阐明自噬和上睑下垂在Delta PK诱导的黑色素瘤溶解中的潜在作用。重点将集中在这些交替的死亡途径和已建立的蛋白酶途径之间的串扰，以及(Ii)表征Delta PK在体外和体内感染和杀死黑色素瘤干细胞的潜力。工作假说是，Delta PK通过自噬和下睑下垂增强PCD的诱导，Delta PK通过这些途径中的一个或多个感染和杀死黑色素瘤干细胞。

DESCRIPTION (provided by applicant): Melanoma is the sixth most common cancer and accounts for approximately ~5% of all newly diagnosed malignancies. This high morbidity and mortality cancer is largely resistant to available therapies. This is due at least in part, to cancer stem cells (CSC) that are increasingly recognized as the primary source of recurrent and therapeutically resistant disease. Oncolytic viruses are a promising new therapeutic that has the potential to target CSC. Early clinical trials indicate that virotherapy is tolerated well but its efficacy is limited by poor cytolytic activity, insufficient tumor penetration and failures to replicate in quiescent tumor cells. Current efforts seek to improve replication but virotherapy strategies based on the induction of programmed cell death (PCD) are not available. Herpes simplex virus (HSV) is a particularly attractive oncolytic virus because it has a broad host spectrum, its genome does not integrate into the host genome thereby precluding insertion mutagenesis, antiviral drugs are available to safeguard against unfavorable virus replication and the virus evades immunity, allowing re-treatment. The proposed studies follow on our recent findings that an HSV-2 mutant deleted in the HSV-2 gene ICP10PK (delta PK) induces melanoma cell death. The activated Ras signaling pathways in melanoma cells provide for selective delta PK replication but oncolysis is mediated by multiple non-redundant PCD pathways. These pathways include increased calpain and caspase activities, upregulation of the epigenetically silenced gene H11 that has a tumor suppressor-like function in melanoma, enhanced expression of the autophagy-related protein Beclin 1, and activation of the inflammatory caspase-1 (pyroptosis). Delta PK is a particularly promising virotherapy because it elicits Th1 cells, which override the suppressive melanoma milieu and it is tolerated well in humans. The proposed studies seek to better elucidate the relationship between the death pathways induced/activated by delta PK and determine whether it can kill melanoma stem cells. The specific aims are: (i) To elucidate the potential contributions of autophagy and pyroptosis to delta PK induced melanoma oncolysis. Focus will be on the cross-talk between these alternate death pathways and established protease pathways and (ii) To characterize delta PK's potential to infect and kill melanoma stem cells both in vitro and in vivo. The working hypothesis is that PCD induction by delta PK is augmented by autophagy and pyroptosis and delta PK infects and kills melanoma stem cells through one or more of these pathways.