Specific Targets for Pancreatic Cancer Therapy

描述（由申请人提供）：胰腺肿瘤是美国第五大最常见的癌症，每年新诊断病例约为48,000例，年死亡率超过99%。其病因在很大程度上是未知的，目前没有有效的治疗方法。在胰腺癌中，一些重要的存活分子如EGFR、NF-kB、COX-2和Bcl-xL被高度激活。NF-kB调控Bcl-xL、COX-2、VEGF、MMP-9、uPAR等多种参与肿瘤进展、生存、血管生成和侵袭的基因。抗凋亡Bcl-xL蛋白的高表达促进细胞存活，并在胰腺癌进展导致耐药过程中发挥关键作用。在90%的胰腺癌中发现了它的表达。环氧化酶-2 （COX-2）酶在胰腺癌中的过度表达也被证明有助于生长、转移和化疗耐药。因此，抑制NF-kB、Bcl-2/Bcl-xL和COX-2应该作为胰腺癌的一种新的治疗策略。我们的合作研究者设计了非肽、药物样、细胞渗透性强效小分子抑制剂（SMI），可以结合Bcl-2和Bcl-xL的口袋并阻断/破坏它们的抗凋亡功能。这些SMI在体外结合实验中进行了测试，发现它们是Bak BH3肽与Bcl-2和Bcl-xL结合的有效抑制剂。我们还发现染料木黄酮通过转录失活NF-kB并下调Bcl-xL、VEGF、MMP-9和uPAR。此外，塞来昔布对COX-2活性和Akt/NF-kB的抑制可能对杀死胰腺癌细胞有潜在的作用。我们发现人类胰腺癌细胞系具有显著的高表达Bcl-XL、NF-kB和COX-2。因此，我们假设通过SMI靶向Bcl-2/Bcl-xL生存通路，并用染料木素和塞来昔布灭活nf - kb诱导的Bcl-2/BcI-xL/COX-2可能是治疗胰腺癌的新治疗策略。这项工作的重要之处是NF-kB通过转录调控Bcl-xL的表达，因此染料木素灭活NF-kB和SMI治疗会导致Bcl-xL的降低。此外，添加塞来昔布可能通过抑制NF-kB以及COX-2的转录和酶活性来诱导胰腺癌细胞死亡。我们的具体目标是：1)研究SMI促进人胰腺癌细胞凋亡的分子机制；2)利用人胰腺癌异种移植模型检测SMI的抗肿瘤活性；3)在体外和体内研究染料木素抑制Bcl-xL转录是否能协同SMI诱导的凋亡。4)研究塞来昔布抑制COX-2对染料木黄酮处理和smi处理的胰腺癌细胞和SCID小鼠异种移植模型中Akt/NF-kB活化的影响。靶向Bcl-2/Bcl-xL、NF-kB和COX-2可能成为胰腺癌新的治疗方法。

DESCRIPTION (provided by applicant): Pancreatic neoplasia is the fifth most common cancer in the United States with >28,000 newly diagnosed cases per year with an annual mortality rate of greater than 99%. Its etiology is largely unknown and no curative treatment is presently available. In pancreatic cancer, several important survival molecules such as EGFR, NF-kB, COX-2, and Bcl-xL are highly activated. NF-kB regulates many genes involved in tumor progression, survival, angiogenesis and invasion such as Bcl-xL, COX-2, VEGF, MMP-9 and uPAR. High expression of anti-apoptotic Bcl-xL protein promotes cell survival and plays a key role in pancreatic cancer progression leading to drug resistance. Its expression was found in 90% of pancreatic cancer. The overexpression of the cyclooxygenase-2 (COX-2) enzyme in pancreatic cancer has also been shown to contribute to growth, metastasis, and chemoresistance. Therefore, inhibitions of NF-kB, Bcl-2/Bcl-xL, and COX-2 should serve as a novel treatment strategy for pancreatic cancer. Our co-investigator has designed non-peptide, drug-like, cell permeable potent small molecule inhibitors (SMI) that bind to the pocket of Bcl-2 and Bcl-xL and block/disrupt their anti-apoptotic function. These SMI were tested in an in vitro binding assay and found to be potent inhibitor of the binding of Bak BH3 peptide to Bcl-2 and Bcl-xL. We have also shown that genistein inactivates NF-kB and down regulates Bcl-xL, VEGF, MMP-9, and uPAR by transcriptional inactivation. Furthermore, the inhibition of COX-2 activity and Akt/NF-kB by celecoxib could be potentially useful in killing pancreatic cancer cells. We have found that human pancreatic cancer cell lines are notable for high expression of Bcl-XL, NF-kB, and COX-2. Therefore, we hypothesize that targeting the Bcl-2/Bcl-xL survival pathway by SMI and inactivating NF-kB-induced Bcl-2/BcI-xL/COX-2 generation by genistein and celecoxib could be novel therapeutic strategies for the treatment of pancreatic cancer. Of importance to this work is that NF-kB transcriptionally regulates the expression of Bcl-xL, hence inactivation of NF-kB by genistein and treatment with SMI will lead to decreased Bcl-xL. Moreover, the addition of celecoxib may be very useful for inducing pancreatic cancer cell death by inhibiting NF-kB as well as transcription and enzyme activity of COX-2. Our Specific Aims are: 1) To investigate the molecular mechanisms by which SMI promote apoptosis in human pancreatic cancer cell lines, 2) To determine anti-tumor activity of SMI using human pancreatic cancer xenograft model, 3) To determine whether transcriptional repression of Bcl-xL by genistein will synergize SMI-induced apoptosis in vitro and in vivo, and 4) To determine the effects of inhibition of COX-2 by celecoxib on Akt/NF-kB activation in genistein-treated and SMI-treated pancreatic cancer cells in vitro and in SCID mouse xenograft models. Targeting Bcl-2/Bcl-xL, NF-kB and COX-2 could be novel therapeutic treatment of pancreatic cancer.