Preclinical Pharmacokinetic Interaction and Histopathological Analyses of Hedyotis diffusa on Sorafenib in Rats

Abstract

Sorafenib is one of the most effective target therapeutic agents for patients with late-stage hepatocellular carcinoma. To seek possible alternative adjuvant agents to enhance the efficacy and improve the side effect of sorafenib, Hedyotis diffusa, one of the most prescribed phytomedicines for treating liver cancer patients in Taiwan, was evaluated in this work. We hypothesized that H. diffusa extract is a safety herb combination on the pharmacokinetic and pharmacodynamic effects of sorafenib. We designed treatments of sorafenib in combination with or without H. diffusa extract to examine its pharmacokinetic properties and effects on liver inflammation. The HPLC-photodiode-array method was designed for monitoring the plasma level and pharmacokinetic parameter of sorafenib in rat plasma. The pharmacokinetic results demonstrated that the area under the curve of sorafenib (10 mg/kg, p.o.) in combination with various doses of H. diffusa formulation (1, 3, and 10 g/kg, p.o.) for 5 consecutive days were 5560 ± 1392, 7965 ± 2055, 7271 ± 1371, and 8821 ± 1705 min μg/mL, respectively, no significant difference when compared with sorafenib treatment alone. Furthermore, the hepatic activity in rats administered with sorafenib with/without H. diffusa extract was quantitatively scored by modified hepatic activity index grading. H. diffusa extract in the range of 1 to 10 g/kg per day did not elicit significant herb-induced hepatotoxicity in rats, based on the histopathological study. Consequently, our findings provided positive safety outcomes for the administration of sorafenib in combination with the phytomedicine H. diffusa.

Figure 1

UHPLC chromatograms of (A) blank plasma, (B) blank plasma spiked with sorafenib (5 μg/mL), and (C) the plasma sample (sorafenib; 10 μg/mL) at 2 h after sorafenib (10 mg/kg, p.o.) administration; 1: retention time of the internal standard (diethylstilbestrol; 10 μg/mL): 3.1 min, 2: retention time of sorafenib: 7.1 min.

Figure 2

(A) Time–concentration curve of sorafenib in rat plasma after oral administration at a dose of group A1 (10 mg/kg, p.o., ●), group A2 (20 mg/kg, p.o., ○), and group A3 (40 mg/kg, p.o., ▼). Data expressed as mean ± S.D.; (B) Time–concentration profile of sorafenib in rat plasma after single-dose oral administration (group A1, 10 mg/kg, p.o., ●); pretreatment with H. diffusa extract (1 g/kg, p.o.) for 5 consecutive days, followed by a single dose of sorafenib (10 mg/kg, p.o.) at day 5 (group A4, ○; n = 6); pretreatment with H. diffusa extract (3 g/kg, p.o.) for 5 consecutive days followed by a single dose of sorafenib (10 mg/kg, p.o.) at day 5 (group A5, ▼; n = 6); and pretreatment with H. diffusa extract (10 g/kg, p.o.) for 5 consecutive days followed by a single dose of sorafenib (10 mg/kg, p.o.) at day 5 (group A6, Δ; n = 6). Data are expressed as mean ± S.D.; n = 6. HD: H. diffusa extract.

Figure 3

Histopathological analyses of the rat liver tissue following administration of sorafenib alone or in combination with H. diffusa extract. (A) Group N, Control group; normal rat liver; (B) Group B1, sorafenib (10 mg/kg, p.o.); (C) Group B2, sorafenib (10 mg/kg, p.o.), following pretreatment with H. diffusa extract (1 g/kg/day, p.o., for 5 consecutive days); (D) Group B3, sorafenib (10 mg/kg, p.o.), following pretreatment with H. diffusa extract (3 g/kg/day, p.o., for 5 consecutive days); (E) Group B4, sorafenib (10 mg/kg, p.o.), following pretreatment with H. diffusa extract (10 g/kg/day, p.o., for 5 consecutive days); (F) Group B5, sorafenib (10 mg/kg, p.o.; consecutive 2 weeks); (G) Group B6, sorafenib (10 mg/kg, p.o.; consecutive 2 weeks), following pretreatment with H. diffusa extract (1 g/kg/day, p.o., for consecutive 2 weeks); (H) Group B7, sorafenib (10 mg/kg, p.o.; consecutive 2 weeks), following pretreatment with H. diffusa extract (3 g/kg/day, p.o., for consecutive 2 weeks); and (I) Group B8, sorafenib (10 mg/kg, p.o.; consecutive 2 weeks), following pretreatment with H. diffusa extract (10 g/kg/day, p.o., for consecutive 2 weeks). 1: focal and periportal inflammation; 2: portal inflammation; 3: focal hemorrhage, congestion, and necrosis; 4: focal lytic necrosis of hepatocytes.

Figure 4

Rat plasma AST and ALT levels after a single dose of sorafenib (10 mg/kg, p.o.) and pretreatment with H. diffusa formulation (group B2, 1 g/kg, p.o., n = 6; group B3, 3 g/kg, p.o., n = 6; and group B4, 10 g/kg, p.o., n = 6) for 2 weeks, followed by a single dose of sorafenib (10 mg/kg, p.o.). Blood samples were taken at 0, 3, 6, 12, and 24 h after the oral administration of the single sorafenib dose or after 2 weeks of pretreatment regimen of co-administration of sorafenib and H. diffusa formulation.