PIK3C2B inhibition improves function and prolongs survival in myotubular myopathy animal models
Nesrin Sabha, … , Eva L. Feldman, James J. Dowling
Nesrin Sabha, … , Eva L. Feldman, James J. Dowling
Published August 22, 2016
Citation Information: J Clin Invest. 2016;126(9):3613-3625. https://doi.org/10.1172/JCI86841.
View: Text | PDF
Research Article Muscle biology

PIK3C2B inhibition improves function and prolongs survival in myotubular myopathy animal models

Abstract

Myotubular myopathy (MTM) is a devastating pediatric neuromuscular disorder of phosphoinositide (PIP) metabolism resulting from mutations of the PIP phosphatase MTM1 for which there are no treatments. We have previously shown phosphatidylinositol-3-phosphate (PI3P) accumulation in animal models of MTM. Here, we tested the hypothesis that lowering PI3P levels may prevent or reverse the MTM disease process. To test this, we targeted class II and III PI3 kinases (PI3Ks) in an MTM1-deficient mouse model. Muscle-specific ablation of Pik3c2b, but not Pik3c3, resulted in complete prevention of the MTM phenotype, and postsymptomatic targeting promoted a striking rescue of disease. We confirmed this genetic interaction in zebrafish, and additionally showed that certain PI3K inhibitors prevented development of the zebrafish mtm phenotype. Finally, the PI3K inhibitor wortmannin improved motor function and prolonged lifespan of the Mtm1-deficient mice. In all, we have identified Pik3c2b as a genetic modifier of Mtm1 mutation and demonstrated that PIK3C2B inhibition is a potential treatment strategy for MTM. In addition, we set the groundwork for similar reciprocal inhibition approaches for treating other PIP metabolic disorders and highlight the importance of modifier gene pathways as therapeutic targets.

Authors

Nesrin Sabha, Jonathan R. Volpatti, Hernan Gonorazky, Aaron Reifler, Ann E. Davidson, Xingli Li, Nadine M. Eltayeb, Claudia Dall’Armi, Gilbert Di Paolo, Susan V. Brooks, Ana Buj-Bello, Eva L. Feldman, James J. Dowling

×

Figure 4

Temporal knockout of Pik3c2b at both early and late stages of disease rescues the Mtm1 disease phenotype.

Options: View larger image (or click on image) Download as PowerPoint
Temporal knockout of Pik3c2b at both early and late stages of disease re...
(A) RT-PCR showing excision of floxed Pik3c2b. (B) Western blot reveals undetectable PIK3C2B protein in cKO muscle. (C) Early-TAM-cKO (induced by tamoxifen treatment starting at 21 days) or late-TAM-cKO (starting at 30 days) of Pik3c2b confers a survival benefit to Mtm1 KO mice. Mtm1 KO alone plus TAM median survival = 42 days, while both early-TAM-cKO (n = 3) and late-TAM-cKO (n = 7) animals live at least 100 days, with the oldest survivors more than 160 days old. (D) Body weights of early-TAM-cKO (n = 3), late-TAM-cKO (n = 7), and Mtm1 KO mice (n = 4) are initially similar to those of Mtm1 KOs, but from 51 days of age onward they have weights similar to WT. (E) Late-TAM-cKOs appear similar (thought slightly smaller) than WTs (age = 160 days). (F) Grip strength measured at 28 and 100 days of age. At 28 days of age, Mtm1 KOs have a significant decrease in mean grip strength (from Figure 1D). After induction of cKO at 30 days, however, grip strength of late-TAM-cKO (n = 4), as measured at 100 days of age, equals that of WT (94% ± 3.1% vs. WT 100% ± 7.5% [P = 0.38]). (G) Late-TAM-cKOs have normalized treadmill behavior. As demonstrated in Figure 1E, Mtm1 KOs have reduced average propulsion time at 28 days (77± 7.2 ms). However, conditional KO of Pik3c2b in Mtm1 KOs starting at 30 days results in normal propulsion at 100 days of age (119 ± 8 ms, n = 4, vs. 125 ± 0.7 ms for WT, n = 3, P = 0.27). (H) Early-TAM-cKOs have normal myofiber size and structure and an absence of central nuclei. Late-TAM-cKO muscle structure is also relatively restored, with slight reductions in myofiber size and small increases in central nuclei (arrows). Scale bars: 20 μm. Comparisons were done using ANOVA followed by Dunnett’s multicomparison test.