Death Receptor Signaling in Pancreatic Cancer: Mechanisms and Therapeutic Targets

描述（由申请人提供）： 胰腺癌是美国癌症死亡的第四大原因，仍然是我们退伍军人的重要健康问题。自1975年以来，胰腺癌患者的5年生存率仅从2%提高到6%。我们和其他人已经证明，调节死亡受体激活的外源性凋亡途径为癌症治疗提供了新的途径。死亡受体4或5（DR 4/5）的重组配体或激动性抗体，包括DR 5激动性抗体TRA-8，已在临床前研究中显示诱导几种类型癌症的细胞凋亡，并且目前正在实体瘤（包括胰腺肿瘤）的几个I-III期临床试验中进行测试。然而，胰腺癌对TRA-8和其他DR配体或激动性抗体的抗性是有效治疗的主要障碍。 在过去的8年里，我们一直在寻找克服癌细胞对DR诱导的凋亡的抵抗的有效试剂。我们已经证明，调节死亡诱导信号复合物（DISC）中的组分决定了DR所传递的下游存活和凋亡信号，从而决定了癌细胞对DR诱导的凋亡的敏感性。最近，我们鉴定了DR 5相关DISC的一个新组分，聚（ADP-核糖）聚合酶1（PARP 1）。PARP 1表达水平与胰腺癌细胞对TRA-8诱导的凋亡的抗性相关。PARP 1的抑制使耐药胰腺癌细胞在体外对TRA-8诱导的细胞凋亡敏感;并增强了TRA-8治疗体内耐药胰腺肿瘤的功效。相反，TRA-8敏感细胞中PARP 1的过表达使它们对TRA-8更具抗性。PARP 1主要在细胞核中起DNA修复的作用。在人胰腺癌组织和细胞中，我们鉴定了PARP 1的细胞质定位。我们证明PARP 1通过新的机制调节DR 5诱导的凋亡信号，通过促进聚（ADP-核糖）（pADPr）修饰caspase-8抑制caspase-8的活化。另一方面，PARP 1抑制减少了DISC对钙调蛋白（CaM）和Src的募集，我们先前报道过它们介导胰腺癌细胞中死亡受体诱导的存活途径。因此，我们假设PARP 1的抑制促进死亡受体诱导的凋亡信号并抑制存活信号，从而增强胰腺癌对TRA-8治疗的敏感性。所提出的研究将表征PARP 1在以下方面的功能：1）调节死亡受体介导的凋亡信号; 2）调节死亡受体介导的存活信号;和3）胰腺癌小鼠模型中的TRA-8治疗。本研究将定义PARP 1调节死亡受体介导的凋亡和存活信号通路的新功能和分子机制，这不仅将阐明PARP 1在基本的外源性凋亡机制中的作用，而且还提供了新的分子见解，促进进一步的研究，将这些发现转化为我们退伍军人患者护理的治疗策略。

DESCRIPTION (provided by applicant): Pancreatic cancer is the fourth leading cause of cancer death in the United States, and remains an important health problem for our Veterans. Since 1975, the 5-year survival rate for pancreatic cancer patients has only improved from 2% to 6%. We and others have demonstrated that modulating death receptor-activated extrinsic apoptosis pathways provides novel avenues for cancer therapy. Recombinant ligands or agonistic antibodies for death receptor 4 or 5 (DR4/5), including DR5 agonistic antibody TRA-8, have been shown to induce apoptosis of several types of cancers in preclinical studies and are currently being tested in several phase I-III clinical trals for solid tumors, including pancreatic tumors. However, resistance of pancreatic cancer to TRA-8, and other DR ligands or agonistic antibodies, are major hurdles for effective therapy. Over the past 8 years, we have been seeking effective reagents that overcome the resistance of cancer cells to DR-induced apoptosis. We have demonstrated that modulating the components in the Death-Inducing Signaling Complex (DISC) determines the downstream survival and apoptosis signals conveyed by the DRs, and thus the sensitivity of cancer cells to DR-induced apoptosis. Recently, we identified a new component of the DR5-associated DISC, poly(ADP-ribose) polymerase 1 (PARP1). PARP1 expression levels correlated with the resistance of pancreatic cancer cells to TRA-8-induced apoptosis. Inhibition of PARP1 sensitized resistant pancreatic cancer cells to TRA-8-induced apoptosis in vitro; and enhanced the efficacy of TRA-8 therapy of resistant pancreatic tumors in vivo. In contrast, overexpression of PARP1 in TRA-8-sensitive cells rendered them more resistant to TRA-8. PARP1 is known to function mainly in the nucleus for DNA repair. In human pancreatic cancer tissues and cells, we identified cytoplasmic localization of PARP1. We demonstrated that PARP1 regulated DR5-induced apoptotic signals through novel mechanisms, by promoting poly(ADP-ribose) (pADPr) modification of caspase-8 that inhibited caspase-8 activation. On the other hand, PARP1 inhibition decreased DISC-recruitment of calmodulin (CaM) and Src, which we previously reported to mediate death receptor-induced survival pathways in pancreatic cancer cells. Therefore, we hypothesize that inhibition of PARP1 promotes death receptor-induced apoptotic signals and inhibits survival signals, thus enhancing the sensitivity of pancreatic cancers to TRA-8 therapy. The proposed studies will characterize the function of PARP1 in 1) regulating death receptor-mediated apoptotic signals; 2) regulating death receptor- mediated survival signals; and 3) TRA-8 therapy in mouse models of pancreatic cancer. The present studies will define the novel function and molecular mechanisms underlying PARP1 regulation of death receptor- mediated apoptotic and survival signaling pathways, which will not only elucidate the role of PARP1 in the fundamental extrinsic apoptotic machinery, but also provide novel molecular insights facilitating further studies to translate these findings into therapeutic strategies for patient care for our Veterans.