The PI3K/Akt/mTOR pathway is overactivated and heat shock protein (HSP) 90 is overexpressed in common cancers. We hypothesized that targeting both pathways can kill intrahepatic cholangiocarcinoma (CCA) cells. HSP90 and PTEN protein expression was evaluated by immunohistochemical staining of samples from 78 patients with intrahepatic CCA. CCA cell lines and a thioacetamide (TAA)-induced CCA animal model were treated with NVP-AUY922 (an HSP90 inhibitor) and NVP-BEZ235 (a PI3K/mTOR inhibitor) alone or in combination.Both HSP90 overexpression and loss of PTEN were poor prognostic factors in patients with intrahepatic CCA. The combination of the HSP90 inhibitor NVP-AUY922 and the PI3K/mTOR inhibitor NVP-BEZ235 was synergistic in inducing cell death in CCA cells. A combination of NVP-AUY922 and NVP-BEZ235 caused tumor regression in CCA rat animal model. This combination not only inhibited the PI3K/Akt/mTOR pathway but also induced ROS, which may exacerbate the vicious cycle of ER stress. Our data suggest simultaneous targeting of the PI3K/mTOR and HSP pathways for CCA treatment.

BACKGROUND. Cholangiocarcinoma (CCA) is an aggressive tumor with a poor prognosis. There is no standard therapy for CCA, and novel drugs for treating refractory CCA need to be identified. METHODS. The authors hypothesized that, if a drug could reverse the gene expression signature of CCA, then it may inhibit the carcinogenesis of CCA and, hence, would be a potential therapeutic agent. Thus, the gene expression signatures from patients with CCA were queried using the bioinformatic method Connectivity Map, resulting in the enrichment of heat‐shock protein 90 (HSP90) inhibitors with therapeutic potentials. RESULTS. Two HSP90 inhibitors, 17‐AAG (tanespimycin) and the synthetic diarylisoxazole amide resorcinol NVP‐AUY922, demonstrated potent antiproliferative activity in in vitro studies. In a thioacetamide‐induced animal model, NVP‐AUY922 also had antitumor activity and resulted in objective tumor regression. In addition, NVP‐AUY922 reduced the expression of client oncoproteins involved in CCA oncogenesis and inhibited downstream proteins of both the phosphatidylinositol 3‐kinase catalytic subunit α/v‐akt murine thymoma viral oncogene homolog 1 protein kinase (PIK3/AKT) pathway and the v‐Ki‐ras2 Kirsten rat sarcoma viral oncogene/mitogen‐activated protein kinase (KRAS/MAPK) pathway. CONCLUSIONS. Preclinical data from the current study suggest that NVP‐AUY922 may be an effective treatment option for patients with CCA. Cancer 2013. © 2012 American Cancer Society.

Cholangiocarcinoma (CCA) is a devastating disease with very poor prognosis due to late diagnosis and resistance to traditional chemotherapies and radiotherapies. Herein, thioacetamide (TAA)‐induced rat CCA model and CGCCA cell line were used; we aim to study the cytogenetic features during tumoral development of CCA and uncover the mystery regarding carcinogenesis of CCA. The Array comparative genomic hybridization analysis, in silico method, gene knockdown, Western blot, cell count proliferation assay, clonogenecity assay, and IHC staining were applied in this study. Array comparative genomic hybridization analysis was performed on all different TAA‐induced phases of rat tissues to reveal the certain pattern, +2q45, +Xq22, −12p12, have been identified for the tumor early stage, where involve the gene TNNI3K. In addition, 16 genes and 3 loci were associated with rapid tumor progression; JAK‐STAT signaling pathway was highly correlated to late stage of CCA. In silico database was used to observe TNNI3K was highly express at tumor part compared with normal adjacent tissue in CCA patients from TCGA dataset. Furthermore, the growth of TNNI3K‐knockdown SNU308 and HuCCT1 cells decreased when compared with cells transfected with an empty vector cell demonstrated by proliferation and colonogenecity assay. Besides, over expression of TNNI3K was especially confirmed on human CCA tumors and compared with the intrahepatic duct stone bile duct tissues and normal bile duct tissues (P < 0.001). Our findings might uncover the mystery regarding carcinogenesis of CCA, and provide the potential genetic mechanism to the clinicians some ideas for the patients’ treatment.