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Elucidation of the Anti-Fibrotic Activity of a Repurposed Drug for Idiopathic Pulmonary Fibrosis
J Reardon - 2022
… , has been identified as a potential novel therapy for IPF. Spironolactone is an off-patent
drug for … Thus, we aimed to access its suitability as an IPF pharmacological
treatment. The … This thesis investigated the anti-fibrotic activity of spironolactone …
Elucidation of the Anti-Fibrotic Activity of a Repurposed Drug for Idiopathic Pulmonary Fibrosis
Idiopathic pulmonary fibrosis (IPF) is a chronic interstitial lung disease with significant mortality and morbidity, with few effective and well tolerated pharmacological treatments. Currently, IPF is the most common idiopathic interstitial pneumonia with a median survival time of 2-4 years following diagnosis. Two anti-fibrotic therapies, pirfenidone and nintedanib, are FDA-approved for IPF treatment. However, the therapeutics are cost prohibitive in numerous countries and induce significant adverse side effects that are difficult for patients to tolerate. Therefore, the demand for a novel, effective, well tolerated anti-fibrotic therapeutic remains. To this end, spironolactone, an established mineralocorticoid receptor (MR) antagonist, has been identified as a potential novel therapy for IPF. Spironolactone is an off-patent drug for fibrotic activity reduction, clinically safe, and a cost-effective option for widespread, scalable implementation.
The overall goal of this research project was to investigate the major anti-fibrotic signalling pathways and subsequent pharmacological effects of spironolactone. Thus, we aimed to access its suitability as an IPF pharmacological treatment. The investigation of the pharmacological activity and mechanisms of action of spironolactone as an anti-fibrotic is necessary in order for us to proceed to phase 1 clinical studies.
This thesis investigated the anti-fibrotic activity of spironolactone on IPF-derived fibroblasts and alveolar epithelial cells in a ‘Scar-in-a-jar’ fibrogenesis assay. A specific focus undertaken was on evaluating pro-fibrotic markers as possible therapeutic targets and investigation of the classical mechanism of spironolactone. Foremost, the efficacy of spironolactone in reducing collagen deposition from pulmonary fibroblasts was successfully demonstrated on Wi38 cells. IPF-derived fibroblasts displayed a significant reduction of collagen deposition after treatment with spironolactone alone and after combined therapies with spironolactone and pirfenidone. Spironolactone alone successfully demonstrated its potential in reversing collagen tissue formation. Significant reductions in COL1A1 and ACTA2 gene expression were observed following combination therapy exposure, in comparison to untreated controls, displaying potential for spironolactone in reducing myofibroblast activation. This study indicated that spironolactone is producing its anti-fibrotic activity via a MR-independent signalling pathway.
Having established a suitable in vitro dose of spironolactone for alveolar epithelial cells, spironolactone, and combination therapy with pirfenidone reduced A549 cell α-SMA production. The study indicates that spironolactone has the potential to reduce epithelial- mesenchymal transition (EMT), as observed in IPF pathogenesis. A significant outcome of EMT in alveolar epithelial cells is the collective migration of cells which aids the aberrant wound healing response in IPF. Employing a scratch migration assay we observed that spironolactone alone and combination therapy reduced cell migration. Additionally, we observed that the gene expression of ACTA2 and FGFR1, pro-fibrotic genes were reduced in our rapid fibrogenesis model after treatment with spironolactone. The study indicated that the anti- fibrotic activity of spironolactone on alveolar epithelial cells is through an MR-independent pathway.
In conclusion, this thesis demonstrates the anti-fibrotic activity of spironolactone in IPF- derived fibroblasts and alveolar epithelial cells, thus suggesting further exploratory work in an in vitro fibrogenesis model. Taken together, the findings in this research study indicate the potential of spironolactone to proceed to preclinical studies, with the long-term goal of repurposing spironolactone as an IPF pharmacological treatment.
スピロノラクトン, ミネラルコルチコイド受容体 (MR) 拮抗薬
IPF の潜在的な新しい治療法
>目標: 主要な抗線維性シグナル伝達経路とそれに続くスピロノラクトンの薬理効果を調査
>IPFの薬理学的治療としての適合性にアクセスすることを目的。薬理学的活性と作用機序。
>「Scar-in-a-jar」線維形成アッセイで、IPF由来の線維芽細胞および肺胞上皮細胞に対するスピロノラクトンの抗線維化活性を調査。行われた特定の焦点は、可能な治療標的としてのプロ線維症マーカーの評価と、スピロノラクトンの古典的なメカニズムの調査でした。
>肺線維芽細胞からのコラーゲン沈着の減少におけるスピロノラクトンの有効性が、Wi38 細胞でうまく実証された。
>IPF由来線維芽細胞は、スピロノラクトン単独での治療後、およびスピロノラクトンとピルフェニドンとの併用療法後に、コラーゲン沈着の有意な減少を示した.
>スピロノラクトンだけでも、コラーゲン組織の形成を逆転させる可能性があることが実証された。 COL1A1 および ACTA2 遺伝子発現の有意な減少が、未治療の対照と比較して併用療法曝露後に観察され、筋線維芽細胞の活性化を減少させるスピロノラクトンの可能性を示す。スピロノラクトンがMRに依存しないシグナル伝達経路を介してその抗線維化活性を生み出していることを示した。
>肺胞上皮細胞、スピロノラクトン、およびピルフェニドンとの併用療法に適した in vitro 用量のスピロノラクトンを確立すると、A549 細胞の α-SMA 産生が減少。スピロノラクトンがIPFの病因で観察されるように、上皮間葉転換（EMT）を減少させる可能性があることを示す。肺胞上皮細胞におけるEMTの重要な結果は、IPFにおける異常な創傷治癒反応を助ける細胞の集団移動。スクラッチ遊走アッセイを使用して、スピロノラクトン単独および併用療法が細胞遊走を減少させる。スピロノラクトンで処理後、急速な線維形成モデルで線維化促進遺伝子である ACTA2 および FGFR1 の遺伝子発現が減少。肺胞上皮細胞に対するスピロノラクトンの抗線維化活性がMR非依存性経路によるもの。
>結論: IPF 由来の線維芽細胞および肺胞上皮細胞におけるスピロノラクトンの抗線維化活性を示しており、in vitro 線維形成モデルにおけるさらなる探索的研究を示唆。この調査研究の結果は、スピロノラクトンを IPF 薬理学的治療として転用するという長期的な目標を持って、前臨床研究に進むスピロノラクトンの可能性を示す。
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特発性肺線維症( idiopathic pulmonary fibrosis (IPF) )関連の学術的情報収集してシェアしています。Google Scholar SearchのUpdateを定期的に掲載しています。GoogleのAIが一定の重み付けはしているとは思いますが、玉石混交です。
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