Antitumor activity of the combination of an HSP90 inhibitor and a PI3K/mTOR dual inhibitor against cholangiocarcinoma - PubMed (original) (raw)

. 2014 May 15;5(9):2372-89.

doi: 10.18632/oncotarget.1706.

Kun-Chun Chiang, Chi-Tung Cheng, Shih-Chiang Huang, Yeng-Yang Chen, Tsung-Wen Chen, Ta-Sen Yeh, Yi-Yin Jan, Hsi-Ming Wang, Jiang-Jie Weng, Peter Mu-Hsin Chang, Chun-Yu Liu, Chung-Pin Li, Yee Chao, Ming-Han Chen, Chi-Ying F Huang, Chun-Nan Yeh

Affiliations

Antitumor activity of the combination of an HSP90 inhibitor and a PI3K/mTOR dual inhibitor against cholangiocarcinoma

Ming-Huang Chen et al. Oncotarget. 2014.

Abstract

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.

PubMed Disclaimer

Figures

Figure 1

Figure 1. HSP90 and PTEN expression was correlated with survival in 78 patients with resectable MF-CCA

(A, B) Immunohistochemical staining of MF-CCA tumors with different intensity scores for HSP90 and PTEN expression; (C) The high-HSP90 group showed significantly worse overall survival (P < 0.001, left); The low-PTEN group showed significantly worse overall survival (P < 0.001, middle); The combined high-HSP90 and low-PTEN group showed the worst overall survival (P < 0.001, right).

Figure 2

Figure 2. The combination of NVP-AUY922 and NVP-BEZ235 synergistically induced apoptosis in CCA cell lines

(A) CGCCA and HuCCT1 cells were incubated with various concentrations (0, 8, 16, 32, 64, or 128 nM) of either NVP-AUY922 or NVP-BEZ235 for 72 h; (B) CGCCA and HuCCT1 cells were incubated with either NVP-AUY922 or NVP-BEZ235 or both NVP-AUY922 and NVP-BEZ235 at various concentrations for 72 h. The combination index (CI) < 1, CI = 1, or CI > 1 indicate synergism, an additive effect, or antagonism, respectively; (C) CGCCA and HuCCT1 cells were treated with 0.5 uM of either NVP-AUY922 or NVP-BEZ235 or a combination of these 2 for 72 h. The number of apoptotic cells measured using the TACS Annexin V-FITC apoptosis detection kit is represented as a percentage of the total events. (D) The immunoblots are analyses of cleaved poly (ADP-ribose) polymerase (PARP). β-Actin was used as the loading control.

Figure 3

Figure 3. NVP-AUY922 and NVP-BEZ235 functioned together to block the PI3K/Akt/mTOR signaling pathway

(A) Western blot analysis revealed the molecular signature of PI3K/mTOR inhibition induced by NVP-AUY922 in the CCA and HuCCT1 cell lines. Cell lysates from the CGCCA and HuCCT1 cell lines were treated with 0.5 uM NVP-AUY922 at various time points (0, 24, 48, 72 h); (B) Western blot analysis reveals the molecular signature induced by NVP-BEZ235 in CCA and HuCCT1 cell lines. Cell lysates from CGCCA and HuCCT1 cells were treated with 0.5 uM NVP-BEZ235 for various time points (0, 24, 48, and 72 h); (C) Western blot analysis revealed the molecular signature induced by NVP-BEZ235 and NVP-AUY922 treatment in CCA and HuCCT1 cell lines. Cell lysates from CGCCA and HuCCT1 cells were treated with 0.5 uM NVP-AUY922 and NVP-BEZ235 at various time points (0, 24, 48, and 72 h). β-Actin was used as the loading control.

Figure 4

Figure 4. NVP-AUY922 induced ER stress and mitochondrial damage, which was fueled by oxidative stress when combined with NVP-BEZ235

(A) CCA cells were incubated with 0.5 uM NVP-AUY922 for 0, 2, 4, 8, 16, and 24 h. Whole cell lysates were subjected to western blot analysis for HSP70, Grp94, Grp78, p-eIF2α, CHOP, IRE1α, and phosphor-JNK. β-Actin was used as the loading control; (B) CCA cells were incubated with 0.5 uM NVP-AUY922 for 48 h. The red and green color ratio of JC-1 reflects the change in the mitochondrial membrane potential (ΔΨm); (C) Relative levels of reduced glutathione (GSH) in CGCCA cell line treated with 0.5 uM NVP-AUY922 and NVP-BEZ235 alone or combined for 0, 24, and 48 h. (D) Reactive oxidative species (ROS) levels induced by 0.5 uM NVP-AUY922 and NVP-BEZ235 alone or combined for 0, 18, and 24 h in CGCCA cell line. (E) The model shows that NVP-AUY922 induces ER stress, which leads to mitochondrial damage, and ultimately to apoptosis. When combined with NVP-BEZ235 treatment, this process is fueled by oxidative stress. NVP-BEZ235 and NVP-AUY922 cooperate to induce apoptosis by vertically affecting the PI3K/Akt/mTOR signaling pathway at multiple nodes.

Figure 5

Figure 5. Detection of rat CCA by animal PET and changes in the tumor/liver SUV ratio

(A) Coronal views of fused CT and PET scans of control and experimental rats revealed the CCA-expressing areas of the liver in which the 18F-FDG uptake was higher than baseline at 2–5 wk after the experiment (i.e., wk 20, 22, and 25). (B) Change in the tumor-to-liver (T/L) ratio of SUV in the control and experiment groups at 2–5 wk after the experiment (i.e., wk 22 and 25).

Similar articles

Cited by

References

    1. Ustundag Y, Bayraktar Y. Cholangiocarcinoma: a compact review of the literature. World J Gastroenterol. 2008;14:6458–6466. - PMC - PubMed
    1. Khan SA, Thomas HC, Davidson BR, Taylor-Robinson SD. Cholangiocarcinoma. Lancet. 2005;366:1303–1314. - PubMed
    1. Patel T. Increasing incidence and mortality of primary intrahepatic cholangiocarcinoma in the United States. Hepatology. 2001;33:1353–1357. - PubMed
    1. Shaib Y, El-Serag HB. The epidemiology of cholangiocarcinoma. Semin Liver Dis. 2004;24:115–125. - PubMed
    1. Valle J, Wasan H, Palmer DH, Cunningham D, Anthoney A, Maraveyas A, Madhusudan S, Iveson T, Hughes S, Pereira SP, Roughton M, Bridgewater J. Cisplatin plus gemcitabine versus gemcitabine for biliary tract cancer. N Engl J Med. 2010;362:1273–1281. - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources