Department of Medical Oncology, Kanazawa Medical University, 1-1 Daigaku, Uchinada, Ishikawa 920-0293, Japan.
J Gastroenterol. 2012 Mar;47(3):321-33. doi: 10.1007/s00535-011-0484-9. Epub 2011 Nov 1.
Pancreatic cancer is obstinate and resistant to gemcitabine, a standard chemotherapeutic agent for the disease. We previously showed a therapeutic effect of glycogen synthase kinase-3β (GSK3β) inhibition against gastrointestinal cancer and glioblastoma. Here, we investigated the effect of GSK3β inhibition on pancreatic cancer cell sensitivity to gemcitabine and the underlying molecular mechanism.
Expression, phosphorylation, and activity of GSK3β in pancreatic cancer cells (PANC-1) were examined by Western immunoblotting and in vitro kinase assay. The combined effect of gemcitabine and a GSK3β inhibitor (AR-A014418) against PANC-1 cells was examined by isobologram and PANC-1 xenografts in mice. Changes in gene expression in PANC-1 cells following GSK3β inhibition were studied by cDNA microarray and reverse transcription (RT)-PCR.
PANC-1 cells showed increased GSK3β expression, phosphorylation at tyrosine 216 (active form), and activity compared with non-neoplastic HEK293 cells. Administration of AR-A014418 at pharmacological doses attenuated proliferation of PANC-1 cells and xenografts, and significantly sensitized them to gemcitabine. Isobologram analysis determined that the combined effect was synergistic. DNA microarray analysis detected GSK3β inhibition-associated changes in gene expression in gemcitabine-treated PANC-1 cells. Among these changes, RT-PCR and Western blotting showed that expression of tumor protein 53-induced nuclear protein 1, a gene regulating cell death and DNA repair, was increased by gemcitabine treatment and substantially decreased by GSK3β inhibition.
The results indicate that GSK3β inhibition sensitizes pancreatic cancer cells to gemcitabine with altered expression of genes involved in DNA repair. This study provides insight into the molecular mechanism of gemcitabine resistance and thus a new strategy for pancreatic cancer chemotherapy.
胰腺癌对吉西他滨（一种治疗该疾病的标准化疗药物）具有顽固性和耐药性。我们之前曾表明糖原合酶激酶-3β（GSK3β）抑制对胃肠道癌和神经胶质瘤的治疗作用。在这里，我们研究了 GSK3β 抑制对胰腺癌细胞对吉西他滨敏感性的影响及其潜在的分子机制。
通过 Western 免疫印迹和体外激酶测定法检查胰腺癌细胞（PANC-1）中 GSK3β 的表达、磷酸化和活性。通过棋盘实验和 PANC-1 异种移植瘤在小鼠中检查吉西他滨和 GSK3β 抑制剂（AR-A014418）对 PANC-1 细胞的联合作用。通过 cDNA 微阵列和逆转录（RT）-PCR 研究 GSK3β 抑制后 PANC-1 细胞中基因表达的变化。
与非肿瘤性 HEK293 细胞相比，PANC-1 细胞显示出增加的 GSK3β 表达、酪氨酸 216 处的磷酸化（活性形式）和活性。药理剂量的 AR-A014418 给药可减弱 PANC-1 细胞和异种移植物的增殖，并显著增强它们对吉西他滨的敏感性。棋盘分析确定联合作用具有协同作用。DNA 微阵列分析检测到吉西他滨处理的 PANC-1 细胞中与 GSK3β 抑制相关的基因表达变化。在这些变化中，RT-PCR 和 Western 印迹显示，肿瘤蛋白 53 诱导核蛋白 1 的表达增加，该基因调节细胞死亡和 DNA 修复，而 GSK3β 抑制则大大降低了该基因的表达。
结果表明，GSK3β 抑制通过改变参与 DNA 修复的基因的表达使胰腺癌细胞对吉西他滨敏感。本研究为吉西他滨耐药性的分子机制提供了深入了解，并为胰腺癌化疗提供了新策略。
