Lipid Metabolism and KSHV-associated Lymphoma Pathogenesis

描述(由申请人提供)：卡波西肉瘤相关疱疹病毒(KSHV)是主要发生在免疫受损宿主中的癌症的病原体，包括感染艾滋病毒的患者。其中一种被称为原发性渗出性淋巴瘤(PEL)的克隆性B细胞恶性肿瘤与显著的死亡率有关，目前还没有有效的治疗方法来减缓PEL的进展或延长生存期。PEL的发病机制涉及KSHV基因的表达和特定信号转导通路的激活，这些信号转导通路诱导肿瘤细胞增殖和延长生存时间。因此，针对KSHV特异性激活的这些通路的新型化合物的开发可能会为PEL提供更有效和更安全的治疗选择。具有生物活性的鞘磷脂在许多生理过程中起着重要的作用，但与鞘氨醇激酶(SK)和1-磷酸鞘氨醇(S1P)的生成有关的鞘磷脂代谢异常也与肿瘤的进展有关。我们已经产生了一种活性化合物ABC294640，它特异性地靶向多种SK亚型，并在体外和体内诱导肿瘤细胞死亡的同时有效地削弱S1P介导的信号转导。了解到ABC294640抑制与PEL发病相关的信号转导，我们进行了初步研究，以确定ABC294640是否抑制PEL细胞的生长。我们的体外研究表明，ABC294640选择性地抑制KSHV感染的淋巴瘤细胞的生长，但不抑制KSHV阴性的淋巴瘤细胞的生长。这些数据补充了我们的其他数据，在这些数据中，ABC294640诱导感染KSHV的原代细胞凋亡，但不诱导未感染的细胞凋亡。此外，ABC294640诱导了神经酰胺类脂质部分的剂量依赖性增加，已知神经酰胺类脂部分可诱导其他癌细胞系的凋亡。ABC294640还上调了促凋亡的JNK活性，同时降低了PEL细胞的结构性核因子-kB活性，这两个事件以前与启动癌细胞凋亡有关。最后，ABC294640诱导PEL细胞中KSHV基因的裂解表达--这一过程可导致细胞死亡。因此，我们假设ABC294640通过神经酰胺的积累和/或诱导KSHV基因的裂解表达和下游促凋亡信号，优先诱导PEL细胞的凋亡和细胞死亡，并且ABC294640抑制体内PEL的进展。为了评估全身给药ABC294640减少体内PEL形成的可行性，并更好地了解ABC294640如何选择性地靶向KSHV感染的细胞，我们提出了三个目标：1)合成足够数量的ABC294640用于体内研究；2)评估ABC294640抑制免疫缺陷小鼠PEL形成的能力；以及3)确定ABC294640抑制KSHV感染细胞生长和存活的机制。通过这些研究的完成，我们希望进行临床试验，以评估ABC294640治疗HIV感染的PEL患者的有效性和安全性。我们的其他机制研究也应该为未来的研究提供进展，以解决ABC294640在选择性靶向病毒感染细胞中的作用。 公共卫生相关性：与卡波西肉瘤相关疱疹病毒(KSHV/HHV8)相关的肿瘤，包括原发性渗出性淋巴瘤(PEL)，是与艾滋病毒相关的疾病之一，预示着艾滋病毒感染患者的不良预后。目前的PEL治疗方法并不有效，而且会产生明显的毒性。因此，在抑制PEL细胞生长的动物模型中进行有希望的化合物的临床前测试，并选择性地针对KSHV感染的细胞，将为在未来的PEL临床试验中测试这些化合物提供基础。

DESCRIPTION (provided by applicant): The Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of cancers arising preferentially within immune compromised hosts, including patients infected with HIV. One of these cancers, a clonal B-cell malignancy known as primary effusion lymphoma (PEL), is associated with significant mortality, and there are currently no effective therapies for reducing the progression of PEL or prolonging survival. Pathogenesis of PEL involves KSHV gene expression and activation of specific signal transduction pathways which induce proliferation and prolong survival of tumor cells. Therefore, the development of novel compounds targeting these pathways specifically activated by KSHV may result in more effective and safe treatment options for PEL. Bioactive sphingolipids play an important role in many physiologic processes, but aberrant sphingolipid metabolism related to the action of sphingosine kinase (SK) and the generation of sphingosine-1-phosphate (S1P) also contribute to cancer progression. We have generated an active compound, ABC294640, that specifically targets multiple SK isoforms and effectively impairs S1P- mediated signal transduction while inducing tumor cell death in vitro and in vivo. With the knowledge that ABC294640 inhibits signal transduction relevant to PEL pathogenesis, we performed preliminary studies to determine whether ABC294640 inhibits PEL cell growth. Our in vitro studies revealed that ABC294640 selectively inhibits KSHV-infected lymphoma cell growth but not the growth of KSHV-negative lymphoma cells. These data compliment our additional data wherein ABC294640 induces apoptosis in KSHV-infected primary cells but not uninfected cells. Moreover, ABC294640 induced a dose-dependent increase in ceramides-lipid moieties known to induce apoptosis in other cancer cell lines. ABC294640 also upregulated proapoptotic JNK activation while reducing constitutive NF-kB activation in PEL cells, two events previously implicated in the initiation of cancer cell apoptosis. Finally, ABC294640 induced lytic KSHV gene expression in PEL cells-a process known to cause cell death. Therefore, we hypothesize that ABC294640 induces apoptosis and cell death preferentially in PEL cells through the accumulation of ceramides and/or the induction of lytic KSHV gene expression and downstream proapoptotic signaling, and that ABC294640 reduces progression of PEL in vivo. To evaluate the feasibility of reducing PEL formation in vivo with the systemic administration of ABC294640 and to better understand how ABC294640 selectively targets KSHV-infected cells, we propose three aims: 1) to synthesize sufficient amounts of ABC294640 for in vivo studies; 2) to evaluate the ability of ABC294640 to inhibit PEL formation in immune deficient mice; and 3) to determine mechanisms for ABC294640 reduction of KSHV-infected cell growth and survival. Through the completion of these studies, we hope to move toward the performance of clinical trials to evaluate the efficacy and safety of ABC294640 administration in HIV-infected patients with PEL. Our additional mechanistic studies should also provide inroads for future studies to address the role of ABC294640 in selectively targeting virus-infected cells. PUBLIC HEALTH RELEVANCE: Tumors etiologically linked to the Kaposi's sarcoma-associated herpesvirus (KSHV/HHV8), including primary effusion lymphoma (PEL), are among those HIV-associated illnesses which portend a poor prognosis for HIV-infected patients. Current PEL therapies are not efficacious and incur significant toxicity. Therefore, preclinical testing of promising compounds in animal models which inhibit PEL cell growth and which selectively target KSHV-infected cells would provide a basis for testing these compounds in future clinic trials for PEL.