Glaucoma, a leading cause of irreversible blindness, results from elevated intraocular pressure (IOP) due to impaired aqueous humor outflow, primarily at the trabecular meshwork (TM). Rho-associated protein kinase (ROCK) promotes TM stiffness by inducing actomyosin contraction and stabilizing actin filaments. Here, we report four novel aromatic compounds, nocarnaphthols A–D (1–4​), isolated from a marine-derived Nocardiopsis sp. SCY18. Nocarnaphthols A (1) and C (3) potently inhibited ROCK1/2 with IC₅₀ values of 6.38–9.65 μM. In human TM cells, these compounds reduced phosphorylation of myosin light chain and cofilin, promoting cytoskeletal relaxation. Barrier restoration in a TGF-β2-induced fibrotic TM model was confirmed by trans-endothelial electrical resistance. In vivo, nocarnaphthol A (0.03% and 0.1%, twice daily) was topically administered to 7-month-old DBA/2J mice, a well-established genetic model of age-related glaucoma, for 3 months. IOP dropped by 30% and 26% at 0.03% and 0.1%, respectively, by week 1—exceeding netarsudil and latanoprost (18%)—and normalized earlier than all controls. No contralateral IOP change or systemic toxicity (e.g., body weight loss) was observed throughout the treatment period. These findings highlight nocarnaphthols A as a promising dual ROCK inhibitor with potent cellular and in vivo efficacy, offering a novel chemical scaffold for glaucoma therapeutics.