Frequent topical administration of hypotensive eye drops in glaucoma patients may lead to the development of dry eye disease (DED) symptoms, because of tear film destabilization and the adverse effects associated with antiglaucoma formulations. To address all this, in the current study preservative-free latanoprost-loaded (0.005 % w/v) synthetic phosphatidylcholine (1,2-dioleoyl-sn-glycero-3-phosphocholine 0.75 % w/v, 1,2-dimyristoyl-sn-glycero-3-phosphocholine 0.25 % w/v) liposomes dispersed in the mucoadhesive polymer hyaluronic acid (0.2 % w/v), containing the osmoprotective ingredients betaine (0.40 % w/v) and leucine (0.90 % w/v) (LAT-HA-LIP), have been prepared and further characterised. Permeation and retention evaluations on a validated ex vivo porcine eye model revealed that the active metabolite latanoprost acid was quantified only starting from LAT-HA-LIP once passing conjunctiva, sclera and choroid compared to the marketed latanoprost (0.005 % w/v) benchmark (MF). The liposomal formulation outperformed MF when applied to the corneal tissue. Additionally, distribution and interactions within corneal and scleral tissues were investigated by means of two-photon microscopy with liposomal formulations containing coumarin-6. Furthermore, acute and chronic tolerance studies on rabbits revealed no signs of discomfort or ocular damage. Schirmer's test, tear osmolarity, tear breakup time (TBUT) and fluorescence staining evaluated through the Oxford grading scale, were assessed as DED diagnostic parameters over a 25-day monitoring period
LAT-HA-LIP consistently maintained levels comparable to physiological solution (0.9 % w/v NaCl) used as control, with a slight increase of TBUT values from day 15 (6.00 ± 0.63 s for control, 7.00 ± 0.78 s for LAT-HA-LIP at day 15, p = 0.0066). A daily topical application of LAT-HA-LIP for 15 consecutive days, effectively lowered IOP in a sustained way (2.51-3.88 mmHg mean IOP reduction over the 5-15-day period). These results highlight the clinical relevance of the proposed technological platform, able to provide IOP reduction during the simulated long-term administration and simultaneous ocular surface protection with potential for the treatment of glaucoma.