Nuclear Factor-kappaB in Ovarian Cancer

NF-kB通路促进癌细胞的存活。我对卵巢癌的研究开始于描述这种疾病中NF-kB的激活状态和生物学相关性。转录因子的NF-κ B家族广泛表达。NF-kB信号通路与卵巢癌有关，但NF-kB信号通路在卵巢癌中的意义和机制尚不清楚。有先例表明NF-kB是癌症中的关键信号传导机制。我最初假设NF-kB通路在卵巢癌中过度激活，具有更强的侵袭性。NF-κ B通路参与卵巢癌细胞的体外增殖和细胞因子分泌，并导致卵巢癌细胞株的化疗耐药性。因此，我试图确定NF-κ B通路蛋白在原发性卵巢癌组织中的表达模式和预后相关性。我证明了诊断时NF-κ B亚单位p50的过度表达传达了这些患者的不良结局。NF-kB在卵巢癌中的生物学相关性是在我的实验室建立的。在证明了卵巢癌中NF-kB机制的协调存在后，我试图调节其活性。标记NF-κ B抑制剂（IkB），以便在IkB激酶（IKK）特异性诱导磷酸化后通过蛋白酶体降解。因此，靶向抑制IKK可以分离NF-kB作为卵巢癌发病机制。抑制IKKb可影响卵巢癌细胞系的生长、粘附、侵袭和细胞因子分泌。我开发了一个IKKb信号在卵巢癌中的基因表达签名，使用IKKb的药理学和遗传操作。基于卵巢癌特异性靶基因的已知功能，该特征使我们能够深入了解NF-kB在卵巢癌中的结果，并使我能够探测已建立的卵巢癌数据库，以估计NF-kB信号传导对卵巢癌女性生存的相对影响。较高的NF-kB活性传达了更差的结果，表明IKKb的调节可能使肿瘤显示靶基因表达升高的患者受益。这项工作的一个关键发现是NF-κ B信号的组织特异性。在卵巢癌中实验定义的9个基因签名与我之前在多发性骨髓瘤中确定的11个基因完全不同。我们与小分子IKKb抑制剂联合进行了一项全球RNAi致敏筛选，以寻找增强毒性的相互作用。 我们的筛选鉴定出了半胱天冬酶8。我们的合作者在淋巴瘤中进行了类似的筛查，发现了IKKa。因此，我假设caspase 8在卵巢癌中比在B细胞淋巴瘤中更活跃，我使用亚致死浓度的IKK B抑制剂和caspase 8 shRNA验证了IKK B和caspase 8的协同性。我开始研究caspase 8在卵巢癌中的功能。首先，我询问了半胱天冬酶8的酶活性是否是与IKKb协同作用所必需的。在IKKb抑制剂的存在下，胱天蛋白酶8裂解活性的肽抑制剂不影响卵巢癌细胞的活力，表明胱天蛋白酶8的这种功能对于其与IKKb的合作不是必需的。这一发现表明，caspase 8在卵巢癌中激活NF-κ B方面发挥着不同的功能。

The NF-kB pathway promotes survival of cancer cells. My research in ovarian cancer began with characterizing the activation state and biological relevance of NF-kB in this disease. The NF-kB family of transcription factors is ubiquitously expressed. NF-kB signaling has been implicated in ovarian cancer, but the significance and mechanism of NF-kB signaling in ovarian cancer is unknown. There is precedent to propose that NF-kB is a critical signaling mechanism in cancer. I initially hypothesized that the NF-kB pathway is over-activated in ovarian cancers with more aggressive behavior. The NF-kB pathway was implicated in ovarian cancer proliferation and cytokine secretion in vitro, and contributed to chemoresistance of ovarian cancer cell lines. I therefore sought to determine the expression patterns and prognostic associations of NF-kB pathway proteins in primary ovarian cancer tissues. I demonstrated that overexpression of the NF-kB subunit p50 at diagnosis conveyed poor outcome in these patients. The biological relevance of NF-kB in ovarian cancer was established in my laboratory. Having demonstrated the coordinate presence of NF-kB machinery in ovarian cancers, I sought to modulate its activity. Inhibitors of NF-kB (IkBs) are tagged for degradation through the proteasome upon specific inducible phosphorylation by IkB kinases (IKKs). Therefore, targeted inhibition of IKKs could isolate NF-kB as a mechanism for ovarian cancer pathogenesis. A subset of ovarian cancer cell lines was affected by inhibition of IKKb in properties of growth, adhesion, invasion and cytokine secretion. I developed a gene expression signature of IKKb signaling in ovarian cancer using both pharmacologic and genetic manipulation of IKKb. This signature gave insight into the results of NF-kB in ovarian cancer, based on known functions of the ovarian cancer-specific target genes, and allowed me to probe established ovarian cancer databases in order to estimate the relative impact of NF-kB signaling on the survival of women with ovarian cancer. Higher NF-kB activity conveyed a worse outcome, suggesting that modulation of IKKb might benefit patients whose tumors showed elevated target gene expression. A key discovery from this work was the tissue specificity of NF-kB signaling. The 9-gene signature experimentally defined in ovarian cancer was completely different from the 11 genes I previously identified in multiple myeloma. We performed a global RNAi sensitization screen in combination with a small molecule IKKb inhibitor looking for interactions that enhanced toxicity. Our screen identified caspase 8. A similar screen in lymphoma, performed by our collaborators, found IKKa. Therefore, I hypothesize that caspase 8 is more active in ovarian cancer, in contrast to B cell lymphomas.I validated the cooperativity of IKKb and caspase 8 using a sub-lethal concentration of IKKb inhibitor with caspase 8 shRNA. I began to examine the context of caspase 8 function in ovarian cancer. First, I asked whether caspase 8 enzymatic activity was required for synergy with IKKb. A peptide inhibitor of caspase 8 cleavage activity did not affect ovarian cancer cell viability in the presence of IKKb inhibitor, indicating that this function of caspase 8 was not necessary for its cooperation with IKKb. This finding suggests that caspase 8 plays a different function to activate NF-kB in ovarian cancer.