Preclinical characterization of the JAK/STAT inhibitor SGI-1252 on skeletal muscle function, morphology, and satellite cell content

<div><p>Background</p><p>Recent studies have highlighted the JAK/STAT signaling pathway in the regulation of muscle satellite cell behavior. Herein we report preclinical studies designed to characterize the effects of a novel JAK/STAT inhibitor on plantar flexor skeletal muscle function, morphology, and satellite cell content.</p><p>Methods</p><p>The compound, SGI-1252, was administered orally (400mg/kg) in a 10% dextrose solution to wild type mice (n = 6) 3 times per week for 8 weeks. A control group (n = 6) received only the dextrose solution.</p><p>Results</p><p>SGI-1252 was well tolerated, as animals displayed similar weight gain over the 8-week treatment period. Following treatment, fatigue in the gastrocnemius-soleus-plantaris complex was greater in the SGI-1252 mice during a 300 second tetanic contraction bout (p = 0.035), though both the rate of fatigue and maximal force production were similar. SGI-1252 treated mice had increased type II myofiber cross-sectional area (1434.8 ± 225.4 vs 1754.7 ± 138.5 μm²), along with an increase in wet muscle mass (125.45 ± 5.46 vs 139.6 ± 12.34 mg, p = 0.032) of the gastrocnemius relative to vehicle treated mice. SGI-1252 treatment reduced gastrocnemius STAT3 phosphorylation 53% (94.79 ± 45.9 vs 44.5 ± 6.1 MFI) and significantly increased the concentration of Pax7⁺ satellite cells (2589.2 ± 105.5 vs 2859.4 ± 177.5 SC/mm³) in the gastrocnemius. SGI-1252 treatment suppressed MyoD (p = 0.013) and Myogenin (p<0.0001) expression in human primary myoblasts, resulting in reduced myogenic differentiation (p = 0.039).</p><p>Conclusions</p><p>Orally delivered SGI-1252 was well tolerated, attenuates skeletal muscle STAT3 activity, and increases satellite cell content in mouse gastrocnemius muscle, likely by inhibiting myogenic progression.</p></div

Proliferation and expression of myogenic regulatory factors MyoD and myogenin.

<p>(A) Proliferation was assessed using 5-ethynil-2'-deoxyuridine (EdU⁺) following 6, 24, 48, and 72 hours in a growth medium. Myogenic regulatory factors (B) MyoD⁺ and (C) Myogenin⁺ nuclei were assessed following 24 and 72 hours in a differentiation medium. Representative images of (D) MyoD⁺ and (E) Myogenin⁺ nuclei at 24 hours. Scale bar = 50μm. * Significance p < 0.05.</p

Muscle characteristics.

<p>(A) Gastrocnemius wet muscle mass, (B) Proportion of type I and II myofibers, (C) type I and (D) type II myofiber cross-sectional area (mCSA) with (E) Representative images of controls and SGI-1252 gastrocnemius type I (bright green) and type II (dark) myofibers. White square in images denotes the boundaries of the enlarged image. Scale bar = 200 μm and 50 μm respectively. * Significance of p < 0.05.</p

Body mass.

<p>Wild-type mice were weighed and treated 3 days a week for 8 weeks with either SGI-1252 or a vehicle solution. Data are means ± SD.</p

Functional characteristics.

<p>(A) Maximal specific force and (B) fatigue were assessed to determine functional characteristics of the gastrocnemius-plantaris-soleus complex. Data are presented as means ± S.D. * Significance of p < 0.05.</p

STAT3 and satellite cell concentrations.

<p>(A) Phosphorylated-STAT3 displayed as a percentage of the total STAT3 concentration. (B) Histological quantification of Pax7⁺ satellite cells per volume of tissue with a (C) representative image from control and SGI-1252 treated mice. White arrows indicate the location of Pax7⁺ cells. Scale bar = 50 μm. * Significance p < 0.05.</p