Anti-obesity efficacy of LH-21, a cannabinoid CB(1) receptor antagonist with poor brain penetration, in diet-induced obese rats

Abstract

Background and purpose: Peripheral blockade of cannabinoid CB(1) receptors has been proposed as a safe and effective therapy against obesity, putatively devoid of the adverse psychiatric side effects of centrally acting CB(1) receptor antagonists. In this study we analysed the effects of LH-21, a peripherally acting neutral cannabinoid receptor antagonist with poor brain penetration, in an animal model of diet-induced obesity.

Experimental approach: To induce obesity, male Wistar rats were fed a high-fat diet (HFD; 60 kcal% fat) whereas controls received a standard diet (SD; 10 kcal% fat). Following 10 weeks of feeding, animals received a daily i.p. injection of vehicle or 3 mg·kg(-1) LH-21 for 10 days. Plasma and liver samples were used for biochemical analyses whereas visceral fat-pad samples were analysed for lipid metabolism gene expression using real-time RT-PCR. In addition, the potential of LH-21 to interact with hepatic cytochrome P450 isoforms and cardiac human Ether-à-go-go Related Gene (hERG) channels was evaluated.

Key results: LH-21 reduced feeding and body weight gain in HFD-fed animals compared with the control group fed SD. In adipose tissue, this effect was associated with decreased gene expression of: (i) leptin; (ii) lipogenic enzymes, including SCD-1; (iii) CB(1) receptors; and (iv) both PPARα and PPARγ. Although there were no significant differences in plasma parameters between HFD- and SD-fed rats, LH-21 did not seem to induce hepatic, cardiac or renal toxicity.

Conclusions and implications: These results support the hypothesis that treatment with the peripherally neutral acting CB(1) receptor antagonist, LH-21, may promote weight loss through modulation of visceral adipose tissue.

Figure 1

Effects of acute administration of LH-21 on cumulative food intake in male Wistar rats. Time course of the dose-response effect of LH-21 (1, 3, 10 and 30 mg·kg⁻¹, i.p.) at 30, 60, 120 and 240 min (A), and 24 h (B) on cumulative food intake (g·kg⁻¹ body weight). Columns are means ± SEM (n= 8 animals per group). Data were analysed by one- or two-way anova (treatment and time) and Bonferroni's post-hoc test. *P < 0.05, **P < 0.01 and ***P < 0.001 denote significant differences compared with the vehicle-treated group.

Figure 2

Effects on body weight and food intake in male Wistar rats fed SD or HFD, and following a 10 day treatment with LH-21. Cumulative body weight gain (A) and cumulative food intake (B) were evaluated weekly for 10 week exposure to SD or HFD. Points and columns are means ± SEM (n= 16 animals per group). Data were analysed by two-way anova (diet and time) and Bonferroni's post-hoc test. *P < 0.05, **P < 0.01 and ***P < 0.001 denote significant differences compared with the SD-fed group. Cumulative body weight gain (C, E) and cumulative food intake (D, F) in SD and HFD-fed animals, respectively, were evaluated after a 10 day exposure to vehicle or LH-21 (3 mg·kg⁻¹, daily, i.p.). Points and columns are means ± SEM (n= 8 animals per group). Data were analysed by two-way anova (diet and treatment) and Bonferroni's post-hoc. *P < 0.05, **P < 0.01 and ***P < 0.001 denote significant differences compared with the vehicle-treated group.

Figure 3

Effect of 10 day treatment with LH-21 on the gene expression of lipid metabolism-related proteins in visceral adipose tissue. Gene expression of PPARα and ACOX (Acox1) were determined as lipolytic molecules (A); and gene expression of PPARγ, FAS (Fasn) and SCD-1 (Scd1) were determined as lipogenic molecules (B) in SD and HFD-fed animals after 10 day exposure to vehicle or LH-21 (3 mg·kg⁻¹, i.p.). Columns are means ± SEM (n= 8 animals per group). Data were normalized with a ratio of housekeeping genes (Sp1 and Rpl19) and analysed by two-way anova (diet and treatment) using Bonferroni's post-hoc. ***P < 0.001 denotes significant differences compared with the vehicle-treated group with SD; ^#P < 0.05, ^##P < 0.01 and ^###P < 0.001 denote significant differences compared with the corresponding vehicle-treated group.

Figure 4

Effect of 10 day treatment with LH-21 on the gene expression of cannabinoid receptors in visceral adipose tissue. Gene expression of CB₁ (Cnr1) (A), CB₂ (Cnr2) (B), and GPR55 (C) were determined in SD and HFD-fed animals after 10 day exposure to vehicle or LH-21 (3 mg·kg⁻¹, i.p.). Columns are means ± SEM (n= 8 animals per group). Data were normalized with a ratio of housekeeping genes (Sp1 and Rpl19) and analysed by two-way anova (diet and treatment) using Bonferroni's post-hoc. *P < 0.05 denotes significant differences compared with the vehicle-treated group with SD; ^#P < 0.05, ^##P < 0.01 and ^###P < 0.001 denote significant differences compared with the corresponding vehicle-treated group.

Figure 5

Effect of 10 day treatment with LH-21 on the gene expression of adipocyte-derived hormones in visceral adipose tissue. Gene expression of leptin (Lep) (A) and adiponectin (Adipoq) (B) were determined in SD and HFD-fed animals after 10 day exposure to vehicle or LH-21 (3 mg·kg⁻¹, i.p.). Columns are means ± SEM (n= 8 animals per group). Data were normalized with a ratio of housekeeping genes (Sp1 and Rpl19) and analysed by one- and two-way anova (diet and treatment) using Bonferroni's post-hoc. *P < 0.05 denotes significant differences compared with the vehicle-treated group with SD; ^###P < 0.001 denotes significant differences compared with the corresponding vehicle-treated group.

Figure 6

Effects of acute administration of LH-21 on cumulative food intake in wild-type and PPARα-null mice. Time course of the effect of LH-21 (20 mg·kg⁻¹, i.p.) on cumulative food intake (g·kg⁻¹ body weight). Columns are means ± SEM (n= 9 animals per group). Data were analysed by two-way anova (treatment and time) and Bonferroni's post-hoc. *P < 0.05, **P < 0.01 and ***P < 0.001 denote significant differences compared with the wild-type vehicle-treated group; ^#P < 0.05, ^##P < 0.01 and ^###P < 0.001 denote significant differences compared with the PPARα-null vehicle-treated group.