Identification of Collagen-Suppressive Agents in Keloidal Fibroblasts Using a High-Content, Phenotype-Based Drug Screen

Abstract

Keloids are characterized by excessive extracellular collagen and exaggerated scarring. Large-volume lesions can be functionally debilitating, therapeutically intractable, and psychologically devastating. A key barrier to translational momentum for novel antikeloid agents is the lack of a faithful high-content screen. We devised, to our knowledge, a previously unreported phenotype-based assay that measures secreted collagen by keloidal fibroblasts in tissue hypoxic conditions (1% oxygen). Four keloidal fibroblasts and 1 normal dermal fibroblast line were exposed to 199 kinase inhibitors. Of 199 kinase inhibitors, 41 (21%) and 71 (36%) increased and decreased the $\overline{\text{CI}}$ _norm (mean collagen inhibition normalized to viability) by more than 10%, respectively. The most collagen suppressive agents were CGP60474 ($\overline{\text{CI}}$ _norm = 0.36), KIN001-244 ($\overline{\text{CI}}$ _norm = 0.55), and RAF265 ($\overline{\text{CI}}$ _norm = 0.58). The top candidate, CGP60474, consistently abolished collagens I and VII production, exhibited minimal global toxicity, and induced a fivefold increase in phosphorylated extracellular signal–regulated kinase. This proof-of-concept high-content screen can identify drugs that appear to target critical keloidal pathophysiology—collagen secretion.