PI3Kα inhibition blocks osteochondroprogenitor specification and the hyper-inflammatory response to prevent heterotopic ossification

Reviewer #1 (Public Review):

Summary:
In the present study, the authors examined the possibility of using phosphatidyl-inositol kinase 3-kinase alpha (PI3Ka) inhibitors for heterotopic ossification (HO) in fibrodysplasia ossificans progressiva (FOP). Administration of BYL719, a chemical inhibitor of PI3Ka, prevented HO in a mouse model of FOP that expressed a mutated ACVR1 receptor. Genetic ablation of PI3Ka (p110a) also suppressed HO in mice. BYL719 blocked osteochondroprogenitor specification and reduced inflammatory responses, such as pro-inflammatory cytokine expression and migration/proliferation of immune cells. The authors claimed that inhibition of PI3Ka is a safe and effective therapeutic strategy for HO.

Strengths:
This manuscript reports an interesting finding that BYL719 inhibits HO in a mouse model of FOP.

Weaknesses:
The molecular mechanisms of BYL719 were still unclear because BYL719 affected multiple events and many types of cells. Additional experimental data would be needed to show more clearly how PI3Ka regulates HO.

Reviewer #2 (Public Review):

Summary:
The authors in this study previously reported that BYL719, an inhibitor of PI3Kα, suppressed heterotopic ossification in mice model of a human genetic disease, fibrodysplasia ossificans progressive, which is caused by the activation of mutant ACVR1/R206H by Activin A. The aim of this study is to identify the mechanism of BYL719 for the inhibition of heterotopic ossification. They found that BYL719 suppressed heterotopic ossification in two ways: one is to inhibit the specification of precursor cells for chondrogenic and osteogenic differentiation and the other is to suppress the activation of inflammatory cells.

Strengths:
This study is based on the authors' previous reports and the experimental procedures including the animal model are established. In addition, to confirm the role of PI3Kα, the authors used the conditional knock-out mice of the subunit of PI3Kα. They clearly demonstrated the evidence indicating that the targets of PI3Kα are not members of TGFBR by a newly established experimental method.

Weaknesses:
Overall, the presented data were closely related to those previously published by the authors' group or others, and there were very few new findings.
Heterotopic ossification in the mice model was not stable and was inappropriate for scientific evaluation.
The method for chondrogenic differentiation was not appropriate, and the scientific evidence of successful differentiation was lacking.
The design of the gene expression profile comparison was not appropriate and failed to obtain the data for the main aim of this study.
The experiments of inflammatory cells were performed in cell lines without ACVR1/R206H mutation, and therefore the obtained data were not precisely related to the inflammation in FOP.