In silico-driven drug repurposing strategy identifies Aloxistatin for the treatment of human heart failure

Introduction & Aim: Fibrotic diseases affect more than 100 million people worldwide, with myocardial and pulmonary fibrosis accounting for the majority. Currently available pharmacological treatment options are limited to drugs that merely slow down the progressive pathological remodeling of the extracellular matrix, leading to detrimental tissue stiffness. This study investigates drug repurposing as a therapy to improve fibrotic disease outcomes.

Methods: In silico drug screening identified drugs interacting with deregulated genes in heart failure patients. Human cardiac fibroblasts (HCF) were treated to validate its anti-fibrotic potential by examining wound healing capacity, cell proliferation and viability. Anti-fibrotic potential was further studied on ultrathin (300µm) rat ex vivo living myocardial slices (LMS). Additionally, we quantified the secretion of the pro-fibrotic miR-21 after Aloxistatin treatment.

Results: In silico analysis of RNA-sequencing datasets highlighted Aloxistatin as an inhibitor of upregulated cathepsin G in ischemic and dilated cardiomyopathy patients (GSE116250) introducing a promising candidate for therapeutic validation. Treatment of HCF with Aloxistatin triggered decreased migratory activity and inhibited proliferation in vitro. Gene expression analysis revealed downregulation of pro-fibrotic markers CTGF, POSTN, and ACTA2 indicating a potent underlying anti-fibrotic molecular mechanism. First cardiotoxic evaluation by Aloxistatin treatment in LMS underlined sustained contraction performance. Analysis of supernatant from cultured LMS disclosed lower secretion levels of pro-fibrotic FN1 after Aloxistatin treatment.

Conclusion: Herein, our in silico-driven drug repurposing strategy has highlighted anti-fibrotic potential of Aloxistatin in the context of myocardial biology. In line, we postulate that Aloxistatin could counteract fibrotic cardiac tissue remodeling by inhibiting e.g. upregulated cathepsin G. Functional analysis on HCF demonstrated inhibited migratory and proliferative activity after treatment, which was validated by repressed gene expression of pro-fibrotic markers. Translational studies in rat LMS confirmed non-cardiotoxic behavior. Further investigations in diseased LMS are planned to analyze fibrosis-related gene expression after Aloxistatin treatment. We conclude that in silico-based approaches can be translated to LMS validation offering an elegant novel model system studying cardiac drug efficacy.