Rapid phenotypic screening for anti-fibrotic activity

Our innovative assay uses In-Cell ELISA to monitor the inhibition of fibrotic activity by your test compound.

Fibrosis, the accumulation of extracellular matrix proteins due to chronic inflammation, is a common underlying pathology in several diseases. It accounts for 42% of deaths in developed countries.

Common examples are idiopathic pulmonary fibrosis, hepatic fibrosis (cirrhosis), kidney fibrosis, heart fibrosis, scleroderma, Dupuytren’s contracture and Peyronie’s disease, although any organ in the body can be affected. Pathophysiology is complex and multi-factorial. Four drugs are approved for the treatment of fibrosis, yet more than 100 fibrotic disorders still lack effective disease-modifying treatments.

Figure 1: Immunocytochemical staining of fibroblasts for α-SMA expression. Burn scar cells were treated with 10 ng/mL TGF-β1 for 72 h. Cell nuclei were stained blue using DAPI and α-SMA stained green. Images subject to 100X magnification.

Our phenotypic screening method detects anti-fibrotic activity in any small molecule compound or biological agent. Primary human fibroblasts cultured in vitro can be transformed into myofibroblasts, an indicator of fibrotic activity in vivo. The change in cell phenotype is monitored by In-Cell ELISA of phenotypic markers.

Inhibition of the transformation by a test compound is evidence of anti-fibrotic activity. This novel, rapid and robust assay can be used to screen a large library of potential drug candidates at low cost.

Figure 2: Outline of our phenotypic assay. Patient derived skin fibroblasts are seeded into 96 or 384-well plates. After 24 hours, skin fibroblasts are treated with 10ng/mL TGF-β1 plus test compounds. After 72 hours, myofibroblast transformation can be quantified by measuring α-SMA expression.

Figure 3: In-cell ELISA image output. Fibroblasts treated with 10ng/mL TGF-β1 with a range of concentrations from two compounds, including negative and positive controls in columns 1 and 2. Green colour denotes expression of α-SMA, red denotes DRAQ5 nuclear stain.

Figure 4: Quantification of assay output. Example of a concentration response curve produced by a compound previously tested in our lab. Data points plotted as n = 3.

We have already screened 1,953 FDA-approved drugs using this assay in multiple tissue types (1 -3), and have identified a drug combination through one of these screens, which has now entered clinical use as an off-label prescription. This is direct proof of translation (4). See the ARU Fibrosis Research Group page for more information about our research.

Diagram showing our current progress. Our Peyronie's Disease assay is available as an off-label prescription, we are sourcing backing for a clinical trial of our hypertrophic scars assay, we are at the hit validation stage for our intra-abdominal adhesions and pulmonary fibrosis projects, and are developing an assay for cardiac fibrosis.

Screening assay service

Our screening assay is available as a service.

Assays are conducted on 96-well or 384-well plates. Any compound showing a positive effect can be further investigated by constructing concentration-response curves. Promising candidate compounds can then be taken forward to have their anti-fibrotic activity confirmed in functional phenotypic assays, such as total ECM production, collagen contraction, and epithelial-mesenchymal transition.

Get in touch

For more information, contact Dr Marc Kuiper at [email protected] and/or Dr Kirsty McKenzie at [email protected]

References

1. Ilg, M. M., Lapthorn, A. R., Ralph, D. J. and Cellek, S. (2022) 'Phenotypic screening of 1,953 FDA-approved drugs reveals 26 hits with potential for repurposing for Peyronie's disease', PLOS One, 17(12), e0277646. Available at: https://doi.org/10.1371/journal.pone.0277646

2. Lapthorn, A. R., Ilg, M. M., Sullivan, J. V., Dziewulski, P. and Cellek, S. (2022) 'Phenotypic screening identifies hydroxypyridone anti-fungals as novel medicines for the prevention of hypertrophic scars', European Journal of Pharmacology, 937, 175374. Available at: https://doi.org/10.1016/j.ejphar.2022.175374

3. Lapthorn, A. R., Harding-Fox, S. L., Feltham, K. M., Ilg, M. M. and Cellek, S. (2025) 'Discovery of novel anti-fibrotics through phenotypic screening', Drug Discovery Today, 30(9), 104450. Available at: https://doi.org/10.1016/j.drudis.2025.104450

4. Ralph, D. J, and Cellek, S. (2026) 'Combination therapy for acute Peyronie' s disease: a success story of translation from bench to bedside, Nature Reviews Urology. Available at: https://doi.org/10.1038/s41585-026-01171-6