Unveiling the Therapeutic Potential: A Peptide Drug Conjugate's Influence on Transcription and Translation in Ocular...

Fibrosis is a major challenge in glaucoma management, particularly following surgical interventions like trabeculectomy. In this study, we explored the therapeutic potential of a fibrosis-modulating peptide (VRF2019) and its mitomycin conjugate by evaluating their transcriptomic impact on primary Tenon fibroblast cells isolated from post-trabeculectomy tissues. To the best of our knowledge, this is the first comprehensive transcriptomic mapping of glaucoma-associated fibrotic fibroblasts, incorporating targeted drug delivery mechanisms. Fibroblasts were subjected to three conditions: Control (untreated), peptide treatment, conjugate treatment, and gene expression changes were analyzed through pairwise comparisons. Peptide-treated cells exhibited dysregulation in 231 transcripts, whereas conjugate-treated cells demonstrated more pronounced changes with 553 dysregulated transcripts. Comparisons between peptide- and conjugate-treated cells revealed 267 differentially expressed transcripts. Bioinformatic analysis highlighted key pathways affected by the treatments, including upregulation of IL-17, TNF-alpha, NF-kappa B, and Hippo signaling pathways, as well as suppression of protein phosphorylation and cell communication pathways. Kinase profiling revealed distinct and overlapping dysregulations across the conditions, with TGFBR1, EPHA2, and NUAK2 upregulated in peptide-treated cells, while conjugate treatment downregulated kinases such as PIM1, RET, and CAMKK1. Weighted gene coexpression network analysis identified significant hub genes, including TGFBR1 in the peptide-treated cells and TNK1 in conjugate-treated cells, elucidating key molecular players in fibrosis modulation. These findings demonstrate that the peptide and its conjugate exert significant antifibrotic effects by modulating transcriptional, signaling, and apoptotic pathways. The conjugate exhibited enhanced regulation of extracellular signaling and fibrotic pathways, suggesting its superior therapeutic potential. This study provides a foundational transcriptomic framework for advancing targeted therapies in glaucoma-associated fibrosis.