Development and characterization of anti-fibrotic natural compound similars with improved effectivity

Abstract

Cardiac fibroblasts constitute the major cell type of the murine and human heart. Once activated, they contribute to an excessive deposition of extracellular matrix (ECM) leading to cardiac fibrosis and subsequently organ dysfunction. With the exception of the pulmonary drugs, nintedanib and pirfenidone, drugs specifically targeting anti-fibrotic pathways are scarce. We recently performed large library screenings of natural occurring compounds and identified first lead structures with anti-fibrotic properties in vitro and in vivo. In line, we now aimed to improve efficacy of these anti-fibrotic lead structures by combining in vitro validation studies and in silico prediction. Next to this combined approach, we performed large OMICs-multi-panel-based mechanistic studies. Applying human cardiac fibroblasts (HCF), we analysed 26 similars of the initially identified anti-fibrotic lead molecules bufalin and lycorine and determined anti-proliferative activity and potential toxicity in an array of in vitro and ex vivo studies. Of note, even at lower concentrations, certain similars were more effective at inhibiting HCF proliferation than nintedanib and pirfenidone. Additionally, selected similars showed low cytotoxicity on human iPS-derived cardiomyocytes and anti-fibrotic gene regulation in human ex vivo living myocardial slices. Further, array and RNA sequencing studies of coding and non-coding RNAs in treated HCFs revealed strong anti-fibrotic properties, especially with the lycorine similar lyco-s (also known as homoharringtonine), that led to a nearly complete shutdown of ECM production at concentrations 100-fold lower than the previously identified anti-fibrotic compound lycorine without inducing cellular toxicity. We thus identified a new natural compound similar with strong anti-fibrotic properties in human cardiac fibroblasts and human living heart tissue potentially opening new anti-fibrotic treatment strategies.

The Molecular Circadian Clock Is a Target of Anti-cancer Translation Inhibitors

“…We noted a similarity score (88% of the digoxin-like score) for the anti-cancer compound HHT which exerts its beneficial anti-leukemic activity by binding to ribosomes, thereby disabling the elongation of nascent peptide chains (Tujebajeva et al, 1989). As this FDA-approved anti-cancer drug is associated with cardiovascular complications (Kantarjian et al, 2013) and also has anti-fibrotic properties in heart (Kreutzer et al, 2022), it prompted us to further characterize HHT activity on REV-ERBα protein expression in cardiomyocytes.…”

Myosin expression and contractile function are altered by replating stem cell–derived cardiomyocytes

“…Wild-type hiPSC-CMs Phoenix were cultivated and differentiated into CMs followed by purification using a modified protocol as previously described ( Chatterjee et al, 2021 ; Haase et al, 2017 ; Kreutzer et al, 2022 ). Briefly, differentiation was initiated at 75–80% confluency with 5 μM CHIR99021 (Merck) in cardio differentiation medium (500 ml RPMI 1640 medium, GlutaMAX Supplement, HEPES [Life Technologies], 250 mg human recombinant albumin [Sigma-Aldrich], 100 mg L -ascorbic acid 2-phosphate sesquimagnesium salt hydrate [Sigma-Aldrich]) in 12-well plates (Greiner) coated with Geltrex (Invitrogen, Thermo Fisher Scientific).…”

Defining cardiac fibrosis complexity and regulation towards therapeutic development

“…A source of these may be natural compounds, with many showing promising antifibrotic functions. 189 Indeed, p38 kinase signalling and cardiac fibrosis can be inhibited by matrine, 190 a plant alkaloid with few adverse effects. Additionally, regulation of other post-translational modifications may offer therapeutic benefits; modulation of HSP90 S-nitrosylation 41 and CD147 glycosylation 191 significantly altered cardiac fibrosis levels in vivo.…”

“…However, many kinase and phosphatase inhibitors exhibit poor selectivity, inefficiency, and subsequent cardiotoxicity 187,188 ; refinement or selection of specific or pleiotropically beneficial compounds is required. A source of these may be natural compounds, with many showing promising antifibrotic functions 189 . Indeed, p38 kinase signalling and cardiac fibrosis can be inhibited by matrine, 190 a plant alkaloid with few adverse effects.…”