Drug repurposing has been shown to bring safe medications to new patient populations, as recently evidenced by the COVID-19 pandemic. We investigated whether we could use phenotypic screening to repurpose drugs for the treatment of Peyronie’s disease (PD). PD is a fibrotic disease characterised by continued myofibroblast presence and activity leading to formation of a plaque in the penile tunica albuginea (TA) that can cause pain during erection, erectile dysfunction, and penile deformity. PD affects 3–9% of men with treatment options limited to surgery or injection of collagenase which can only be utilised at late stages after the plaque is formed. Currently there are no approved medications that can be offered to patients presenting with early disease before the formation of the plaque. Drug repurposing may therefore be the ideal strategy to identify medical treatments to address this unmet medical need in early PD. We used primary human fibroblasts from PD patients in a phenotypic screening assay that measures TGF-β1-induced myofibroblast transformation which is the main cellular phenotype that drives the pathology in early PD. A library of FDA-approved 1,953 drugs was screened in duplicate wells at a single concentration (10 μM) in presence of TGF-β1. The myofibroblast marker α-SMA was quantified after 72h incubation. A positive control of SB-505124 (TGF-β1 receptor antagonist) was included on each plate. Hits were defined as showing >80% inhibition, whilst retaining >80% cell viability. 26 hits (1.3%) were identified which were divided into the following main groups: anti-cancer drugs, anti-inflammation, neurology, endocrinology, and imaging agents. Five of the top-ten drugs that increase myofibroblast-transformation appear to act on VEGFR. This is the first phenotypic screening of FDA-approved drugs for PD and our results suggest that it is a viable method to predict drugs with potential for repurposing to treat early PD.
Introduction: Hypertrophic scarring after burn injuries occurs in approximately 91% of all burn patients worldwide. The main pathology underlying hypertrophic scarring is fibrosis and one of the hallmarks of dermal fibrosis is the transformation of fibroblasts to alpha-smooth muscle actin (a-SMA) expressing myofibroblasts. We have developed a high-throughput screening (HTS) assay to identify novel drugs that can inhibit transformation of fibroblasts to myofibroblasts and therefore can be developed for treatment of burn scarring. Materials & Methods: Fibroblasts were derived from hypertrophic scars of burn patients. Exposure to TGF-b1 was used to induce transformation to myofibroblasts. Quantification of a-SMA and other markers was performed using immunocytochemistry (ICC), real-time qPCR (RT-qPCR) and Western blotting. The In-Cell ELISA (ICE) method was used to develop the HTS assay, where a-SMA expression and cell numbers were quantified. Results: Fibroblast identity of primary patient-derived cells was confirmed by the expression of vimentin and the absence of desmin and cytokeratin-14 expression. TGF-b1 was shown to induce myofibroblast transformation in these cells, with a 3-fold increase in expression of a-SMA. The HTS assay was able to reliably quantify TGF-b1 induced myofibroblast transformation, with early optimization of the screening assay yielding a Z factor greater than 0.5. Conclusions: The assay has been validated for HTS and will be used to screen an FDA-approved drug library of 1,900 compounds, to see if any can be repurposed as a treatment for burn scars. This is advantageous as the drugs are already deemed safe in patients. It is also likely that this approach will prevent the need for multiple surgical interventions, saving both time and money for clinicians and patients alike.
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