Nab-paclitaxel is effective against intrahepatic cholangiocarcinoma via disruption of desmoplastic stroma

Abstract

Intrahepatic cholangiocarcinoma (IH-CCA) is the second predominant hepatic malignancy worldwide. However, effective treatment strategies for IH-CCA have not yet been developed. Nab-paclitaxel may be an effective drug against IH-CCA, a type of desmoid-like tumor, and its antitumor effects may be attributable to its ability to disrupt the cancer-associated fibroblasts. In the present study, MTT and Annexin-V apoptosis detection kits were used to evaluate the efficacy of paclitaxel and nab-paclitaxel against human cholangiocarcinoma KKU-100 and KKU-213 cell lines. A rat model of thioacetamide-induced spontaneous desmoplastic IH-CCA was used to compare the treatment response of four different drug regimens: Control, paclitaxel, nab-paclitaxel and gemcitabine/oxaliplatin. Positron emission tomography and immunofluorescence analysis were used to measure the tumor volume and to study the resected tumor, respectively. In vitro, paclitaxel and nab-paclitaxel induced anti-proliferative effects in KKU-100 and KKU-M213 cells. With regards to the treatment regimes, only nab-paclitaxel and gemcitabine/oxaliplatin induced antitumor effects in the rat model of thioacetamide-induced IH-CCA. The immunofluorescence study indicated that nab-paclitaxel was more efficient in disrupting cancer-associated fibroblasts than paclitaxel. In conclusion, nab-paclitaxel is effective against IH-CCA owing to its ability to markedly disrupt the desmoplastic stroma.

Keywords: cancer-associated fibroblast; cholangiocarcinoma; desmoplastic; nab-paclitaxel; α-smooth muscle actin.

Figure 1.

Spontaneous TAA-induced IH-CCA model. The schematic diagram illustrates the administration of TAA to rats for 16 weeks for the development of spontaneous IH-CCA. TAA (300 mg/l) was continuously administered to rats by oral gavage. After 10 weeks of administration, the animals were sacrificed and cholangiolar viability with atypical ductal cells was evaluated in the liver tissues. Continuous feeding of the rats with TAA for a further 6 weeks results in the development of cholangiocarcinoma with surrounding cancer-associated fibroblasts. TAA, thioacetamide; IH-CCA, intrahepatic cholangiocarcinoma.

Figure 2.

Nab-paclitaxel induced cholangiocarcinoma cell apoptosis and inhibited tumor cell viability. The antitumor effect of paclitaxel or nab-paclitaxel was observed in two CCA cell lines. (A) KKU-100 and KKU-213 cell lines were cultured with or without paclitaxel or nab-paclitaxel at indicated concentrations (0, 5, 10, 20, 40, 80, 160 or 320 nM) for 72 h. Cell viability was evaluated using an MTT assay and data are presented as the mean ± standard deviation of 3 independent experiments. (B) KKU-100 and KKU-213 cells were treated with or without nab-paclitaxel at indicated concentrations (0, 5, 10 or 20 nM) for 48 h. Apoptotic cells were measured using the TACS Annexin V-FITC apoptosis detection kit and are represented as a percentage of the total cell death.

Figure 3.

Nab-paclitaxel induced in vivo antitumor effects. (A) The spontaneous TAA-induced IH-CCA model was used for drug treatments. All pre-treatment groups of rats exhibited ¹⁸F-FDG-PET-detectable tumors following administration of TAA for 20 weeks. (B) Alterations in SUV 2 and 4 weeks post-treatment with paclitaxel, nab-paclitaxel and gemcitabine/oxaliplatin (G+O), respectively. *P<0.05. TAA, thioacetamide; IH-CCA, intrahepatic cholangiocarcinoma; NS, not significant; SUV, standardized uptake value.

Figure 4.

CAFs were disrupted by nab-paclitaxel in IH-CCA. (A) In order to study the effects of the treatments on CAFs, tissues from rats treated with the indicated agents were stained with α-SMA (red) and quantitated for the presence of activated and total fibroblasts. (B) The confocal microscopic analysis of α-SMA (red) and 4′6-diamidino-2-phenylindole (blue) immunofluorescence staining of total and activated CAFs in tumors treated with paclitaxel and nab-paclitaxel. CAFs, cancer-associated fibroblasts; IH-CCA, intrahepatic cholangiocarcinoma; α-SMA, α-smooth muscle actin.