# High-throughput 3D phenotypic screening identifies repurposed MEK inhibitors as drivers of chondrogenesis for cartilage regeneration

## TL;DR

This study screens drugs for their ability to promote cartilage formation, finding that MEK inhibitors like Trametinib could help regenerate cartilage.

## Contribution

The study introduces MEK inhibitors as novel candidates for cartilage regeneration through high-throughput 3D screening.

## Key findings

- Trametinib significantly alters cell morphology and promotes a chondrogenic-like shape.
- Molecular analysis shows upregulation of Collagen II and aggrecan, key cartilage markers.
- Trametinib enhances cell migration and actin organization consistent with chondrogenic differentiation.

## Abstract

Chondrogenesis is essential for cartilage repair and regeneration, particularly in treating osteoarthritis and cartilage injuries. While conventional therapies rely heavily on growth factors, recent interest has turned toward drug repurposing strategies involving small-molecule inhibitors. This study aims to evaluate the chondrogenic potential of selected bioactive compounds, with a particular focus on Trametinib, a MEK inhibitor.

A library of 55 bioactive compounds was screened using high-content imaging and a 3D hydrogel model that mimics the native cartilage microenvironment. Cellular morphology, migration, and cytoskeletal organization were assessed to identify chondrogenic phenotypes. Trametinib, along with Panobinostat, SAHA, and Brefeldin A, was further evaluated via dose-response analyses and molecular assays to determine their impact on chondrogenic differentiation.

Trametinib was identified as a potent modulator of chondrogenesis-related cellular phenotypes. It significantly altered cell morphology, promoted a chondrogenic-like shape, and enhanced cell migration. Changes in actin organization were quantified using SER-Spot and SER-Ridge metrics, showing patterns consistent with chondrogenic differentiation. Molecular analysis revealed upregulation of Collagen II and aggrecan, key markers of cartilage formation.

These findings support the potential of MEK inhibitors like Trametinib, and other selected bioactive compounds, as promising agents for cartilage regeneration. Their repurposing could offer innovative therapeutic strategies for treating cartilage-related disorders, including osteoarthritis.