Antitumor effects of regorafenib and sorafenib in preclinical models of hepatocellular carcinoma

Abstract

The purpose of this study was to investigate the antitumor activity of regorafenib and sorafenib in preclinical models of HCC and to assess their mechanism of action by associated changes in protein expression in a HCC-PDX mouse model. Both drugs were administered orally once daily at 10 mg/kg (regorafenib) or 30 mg/kg (sorafenib), which recapitulate the human exposure at the maximally tolerated dose in mice. In a H129 hepatoma model, survival times differed significantly between regorafenib versus vehicle (p=0.0269; median survival times 36 vs 27 days), but not between sorafenib versus vehicle (p=0.1961; 33 vs 28 days). Effects on tumor growth were assessed in 10 patient-derived HCC xenograft (HCC-PDX) models. Significant tumor growth inhibition was observed in 8/10 models with regorafenib and 7/10 with sorafenib; in four models, superior response was observed with regorafenib versus sorafenib which was deemed not to be due to lower sorafenib exposure. Bead-based multiplex western blot analysis was performed with total protein lysates from drug- and vehicle-treated HCC-PDX xenografts. Protein expression was substantially different in regorafenib- and sorafenib-treated samples compared with vehicle. The pattern of upregulated proteins was similar with both drugs and indicates an activated RAF/MEK/ERK pathway, but more proteins were downregulated with sorafenib versus regorafenib. Overall, both regorafenib and sorafenib were effective in mouse models of HCC, although several cases showed better regorafenib activity which may explain the observed efficacy of regorafenib in sorafenib-refractory patients.

Keywords: HCC; antitumor activity; preclinical pharmacology; regorafenib; sorafenib.

Conflict of interest statement

CONFLICTS OF INTEREST SB, MK, LF, FTH, TL, MT, and DZ are full-time employees of Bayer AG. SK was a full-time employee of Natural and Medical Sciences Institute at the University Tubingen at the time of the study and is now employed by Rentschler Biopharma SE, Germany. QJ and QG are full time employees of WuXi AppTec, Shanghai, China.

Figures

Figure 1. Survival of mice carrying orthotopic H129 liver tumors without treatment or treated with vehicle, regorafenib, or sorafenib

Treatments with regorafenib (A) and sorafenib (B) were performed in separate studies. The survival times are measured in days.

Figure 2. Tumor growth inhibition of 10 HCC-PDX models treated with vehicle, regorafenib, or sorafenib

Models are sorted according to mean tumor volumes of the vehicle-treated groups. Detailed numbers are provided in Supplementary Tables 1 and 2. *p<0.05; **p<0.01; †p<0.001; ††p<0.0001; no label = not significant; error bars indicate standard deviation. HCC-PDX, patient-derived hepatocellular carcinoma xenograft.

Figure 3. Tumor growth curves of three HCC-PDX models representing different responses to regorafenib and sorafenib treatment

Tumor-bearing mice were treated orally for 28 days (dotted line) with regorafenib at 10 mg/kg, sorafenib at 30 mg/kg, or with vehicle. Asterisks mark time points when animals were lost/sacrificed. (A) Responder (model 5); (B) differential responder (model 10); (C) non-responder (model 20). Error bars indicate standard deviations. HCC-PDX, patient-derived hepatocellular carcinoma xenograft.

Figure 4. Effects of regorafenib or sorafenib treatment on selected protein analytes in HCC-PDX model 189 by a bead-based multiplex western blot analysis

Proteins showing a significant change (p

Figure 5

Fold changes of selected protein analytes (pooled) between (A) regorafenib and (B) sorafenib versus vehicle-treated HCC-PDX 189 xenografts. Components of interest are labeled with open circles and names. Data points above the horizontal orange bar are statistically significant (p<0.05). Vertical orange bars show the cut-offs for downregulation (fold change less than –1) and upregulation (fold change greater than 1). HCC-PDX, patient-derived hepatocellular carcinoma xenograft.

Figure 5

Fold changes of selected protein analytes (pooled) between (A) regorafenib and (B) sorafenib versus vehicle-treated HCC-PDX 189 xenografts. Components of interest are labeled with open circles and names. Data points above the horizontal orange bar are statistically significant (p<0.05). Vertical orange bars show the cut-offs for downregulation (fold change less than –1) and upregulation (fold change greater than 1). HCC-PDX, patient-derived hepatocellular carcinoma xenograft.