**Background:** Dry eye disease (DED) is a highly prevalent multifactorial condition characterized by tear film instability, hyperosmolarity, and ocular surface inflammation. Current treatments, including lubricants and anti-inflammatory agents like cyclosporine and corticosteroids, have limited efficacy and significant adverse effects, leading to poor adherence. Sesquiterpene lactones, particularly helenalin from Arnica montana, are potent inhibitors of NF-κB signaling, a key regulator of ocular surface inflammation. This study evaluated the safety, tolerability, and clinical efficacy of a topical ophthalmic formulation containing helenalin and hyaluronic acid (HA) in healthy volunteers and patients with mild-to-moderate DED.
**Methods:** The study comprised two parts: a phase I safety/tolerability study in 24 healthy volunteers (mean age 36.4 ± 9.82 years, 54.16% male) who applied one drop of the formulation three times daily for 21 days, and a phase II efficacy study in 48 patients with mild-to-moderate DED (mean age 53.1 ± 9.82 years in Group 1, 52.5 ± 13.15 in Group 2; 50% male in each group). Patients were randomized to receive either the helenalin-HA formulation (Group 1) or a commercially available HA 0.04% lubricant (Group 2) for 28 days. Outcomes included OSDI score, non-invasive tear breakup time (NIBUT), non-invasive average breakup time (NIAvg-BUT), Schirmer's test, MMP-9 positivity (InflammaDry®), conjunctival impression cytology (CIC) graded by Nelson system, ocular surface staining score (OSS), and meibomiography. A crossover phase without washout assessed MMP-9 changes after switching treatments.
**Key Results:** In the phase I study, no serious adverse events occurred; transient mild burning (12.5%), tearing (8.33%), and blurred vision (4.16%) resolved within 10 minutes. No significant changes in BCVA, IOP, or corneal endothelial cell density were observed. In the phase II study, both groups showed significant improvement in OSDI (Group 1: 21.31 ± 3.21 to 10.58 ± 4.21; Group 2: 21.74 ± 15 to 11.23 ± 8.95) and NIBUT (Group 1: 8.39 ± 5.86 to 14.53 ± 4.53 s; Group 2: 8.43 ± 4.82 to 13.83 ± 5.69 s). Only Group 1 showed significant improvement in NIAvg-BUT (10.46 ± 4.19 to 14.24 ± 3.66 s) and Schirmer's test (15.83 ± 5.4 to 20.05 ± 4.37 mm). MMP-9 positivity decreased from 100% to 25% in Group 1 (p < 0.05) versus 100% to 87.5% in Group 2. CIC normalized in all Group 1 patients (33.3% abnormal at baseline to 0% at 1 month; p = 0.0078), while Group 2 showed no change (29.1% abnormal at both time points). OSS improved significantly in Group 1 (p = 0.0367) but not in Group 2. Meibomiography showed notable improvement in all Group 1 patients. After crossover, MMP-9 positivity in Group 1 increased from 25% to 91.6% after stopping the formulation, while in Group 2 it decreased from 87.5% to 20.8% after switching to the helenalin formulation.
**Clinical Implications:** The helenalin-HA formulation demonstrated excellent safety and tolerability, with rapid and significant improvements in both subjective symptoms and objective signs of DED, including tear film stability, tear secretion, and ocular surface inflammation. The reduction in MMP-9 positivity and normalization of conjunctival morphology suggest effective anti-inflammatory action, likely mediated by NF-κB pathway inhibition. The crossover results provide mechanistic evidence of time-dependent anti-inflammatory effects. This formulation may offer a novel, well-tolerated therapeutic option for mild-to-moderate DED, addressing the limitations of current anti-inflammatory treatments. However, the small sample size, single-center design, and short follow-up warrant larger, longer-term studies to confirm efficacy and elucidate molecular mechanisms.