Prostaglandin analogues (PG), beta-blockers (BB) or their combination (PG+BB) are used primarily to reduce the intraocular pressure (IOP) pathologically associated with glaucoma. Since, fibrosis of the trabecular meshwork (TM) is a major aetiological factor in glaucoma, we studied the effect of these drugs on fibrosis-associated gene expression in TM of primary glaucoma patients. In the present study, TM and iris of primary open-angle (n = 32) and angle-closure (n = 37) glaucoma patients were obtained surgically during trabeculectomy and categorized based on the type of IOP-lowering medications use as PG, BB or PG+BB. mRNA expression of pro-fibrotic and anti-fibrotic genes was quantified using qPCR in these tissues. The gene expression levels of pro-fibrotic genes were significantly lower in PG+BB as compared to other groups. These observations and underlying signalling validated in vitro in human TM cells also showed reduced fibrotic gene and protein expression levels following PG+BB treatment. In conclusion, it is observed that PG+BB combination rather than their lone use renders a reduced fibrotic status in TM. This further suggests that IOP-lowering medications, in combination, would also modulate fibrosis-associated molecular changes in the TM, which may be beneficial for maintaining aqueous out-flow mechanisms over the clinical treatment duration. © 2020 Narayana Nethralaya Foundation. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd