Characterization of GLPG0492, a selective androgen receptor modulator, in a mouse model of hindlimb immobilization

Abstract

Background: Muscle wasting is a hallmark of many chronic conditions but also of aging and results in a progressive functional decline leading ultimately to disability. Androgens, such as testosterone were proposed as therapy to counteract muscle atrophy. However, this treatment is associated with potential cardiovascular and prostate cancer risks and therefore not acceptable for long-term treatment. Selective Androgen receptor modulators (SARM) are androgen receptor ligands that induce muscle anabolism while having reduced effects in reproductive tissues. Therefore, they represent an alternative to testosterone therapy. Our objective was to demonstrate the activity of SARM molecule (GLPG0492) on a immobilization muscle atrophy mouse model as compared to testosterone propionate (TP) and to identify putative biomarkers in the plasma compartment that might be related to muscle function and potentially translated into the clinical space.

Methods: GLPG0492, a non-steroidal SARM, was evaluated and compared to TP in a mouse model of hindlimb immobilization.

Results: GLPG0492 treatment partially prevents immobilization-induced muscle atrophy with a trend to promote muscle fiber hypertrophy in a dose-dependent manner. Interestingly, GLPG0492 was found as efficacious as TP at reducing muscle loss while sparing reproductive tissues. Furthermore, gene expression studies performed on tibialis samples revealed that both GLPG0492 and TP were slowing down muscle loss by negatively interfering with major signaling pathways controlling muscle mass homeostasis. Finally, metabolomic profiling experiments using 1H-NMR led to the identification of a plasma GLPG0492 signature linked to the modulation of cellular bioenergetic processes.

Conclusions: Taken together, these results unveil the potential of GLPG0492, a non-steroidal SARM, as treatment for, at least, musculo-skeletal atrophy consecutive to coma, paralysis, or limb immobilization.

Figure 1

GLPG0492 reversed immobilization-induced muscle atrophy in a dose-dependent manner. Normalized gastrocnemius weight from both immobilized (Panel A) and contralateral (Panel B) legs was assessed at day 7 in intact or immobilized mice receiving increasing doses of GLPG0492 (0.3, 3, 10 mg/kg/day) or TP (1 mg/kg/day) or CTL immobilized (ethanol/corn oil, 5/95 v/v). Panel C: Normalized prostate weight at day 7, from immobilized mice receiving increasing doses of GLPG0492 (0.3, 3, 10 mg/kg/day) or TP or CTL vehicle (n = 10 per group; #p < 0.05, ##p < 0.01, ###p < 0.001 vs. Intact (white bar); *p < 0.05, **p < 0.01, ***p < 0.001 vs. CTL immobilized (black bar)).

Figure 2

GLPG0492 reduced muscle fibers atrophy induced by hindlimb immobilization. Panel A: representative immuno-fluorescence staining of gastrocnemius cryosections for myosin light chain and laminin discriminating slow-twitch fibers (red star) from fast-twitch fibers (white star). TP and GLPG0492 were tested at 1 and 10 mg/kg/day, respectively. Panel B: FCSA was determined at day 7 in intact and immobilized mice receiving increasing doses of GLPG0492 (0.3, 3, 10 mg/kg/day) or TP (1 mg/kg/day) or CTL vehicle. (n = 10 per group; #p < 0.05, ##p < 0.01, ###p < 0.001 vs. intact; ***p < 0.001 vs. CTL immobilized). Panel C: Cross-sectional area (CSA) of gastrocnemius muscle fast and slow fibers distribution (n = 10 per group; *p < 0.05, **p < 0.01, ***p < 0.001 vs. CTL immobilized).

Figure 3

GLPG0492 selectively modulated key pathways involved in muscle atrophy. Gene expression levels were monitored at day 7 in tibialis muscle samples from intact or immobilized mice receiving either GLPG0492 (10 mg/kg/day) or TP (1 mg/kg/day) or vehicle. Figure 3a: Atrogin-1, MurF1, LC3, IGFI, FoxO1 and Figure 3b: Myogenin, IL1B, PGC-1α (n = 10 per group; ##p < 0.01, ###p < 0.001 vs. Intact; *p < 0.05, **p < 0.01, ***p < 0.001 GLPG0492-treated vs. CTL immobilized).

Figure 4

GLPG0492 increased plasma metabolite levels linked to TCA cycle, β - oxidation and amino acids metabolism. Metabolic profiling by 1H-NMR of serum samples from intact or immobilized mice receiving either GLPG0492 (10 mg/kg/day) or vehicle was performed at day 7. (n = 5 per group; #p < 0.05 Intact vs. Immobilized; *p < 0.05, **p < 0.01 vs. CTL immobilized).