This was a 4-week multicenter, randomized, double-masked, dose-ranging, parallel-group study comparing three higher concentrations of latanoprost (75, 100, and 125 μg/mL) with latanoprost 50 μg/mL. This study was conducted in accordance with the International Conference on Harmonization Guideline for Good Clinical Practice and the Declaration of Helsinki and the protocol approved by the independent ethics committees/institutional review boards responsible for each participating institution. Written informed consent was obtained from all subjects prior to enrollment in the study.
Ethics committees/IRBS included Ministerio da Saude Hospitais Civis de Lisboa, Lisbon, Portugal; Ethics Committee of Institute of Aviation Medicine, Prague, Czech Republic; South East Sydney Area Health Research Ethics Committee, Sydney, Australia; Comite Consultatif de Protection des Personnes dans la Recherche Biomedicale, Hopital Tarnier, Paris, France; AIBILI Associacao para Investigacao Biomedica e Inovacao em Luz e Imagem Azinhaga de Santa Coma, Coimbra, Portugal; Solihull Local Research Ethics Committee, Birmingham, UK; East Birmingham Local Research Ethics Committee Birmingham Heartlands Hospital, Birmingham, UK; AKADIMOS Ophthalmology Centre of Northern Greece, Thessaloniki, Greece; Ethics Committee of OSL, Prague, Czech Republic; Aga Khan University Hospital IRB, Karachi, Pakistan; Unidada Local de Saude de Matosinhos, Matosinhos, Portugal; Research Ethics Committee Glasgow Royal Infirmary, Glasgow, Scotland, UK; Sunderland Local Research Ethics Committee, Sunderland, UK; The Layton Rahmatulla Benevolent Trust Free Eye & Cancer Hospital, Lahore, Pakistan; The Ethics Committee of the Faculty of Medicine Cholalongkom University, Bangkok, Thailand; The Ethical Review Committee on Research Involving Human Subjects, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand; eticka komise of the General Faculty Hospital, Charles University, Prague, Czech Republic; Royal Adelaide Hospital Research Ethics Committee, Adelaide, Australia; Medical Ethics Committee, Dow Medical College and Civil Hospital, Karachi, Pakistan; University Hospital Brno, Brno, Czech Republic; PGMI/Service Hospital Ethics Committee, Lahore, Pakistan.
Male or female subjects ≥18 years of age with POAG or ocular hypertension requiring unilateral or bilateral administration of IOP-lowering treatment, including subjects who were naïve to such treatment, were enrolled. It was recommended, but not required, that subjects have an IOP ≥17 mmHg and ≤36 mm in at least one eye at screening to be eligible. After a washout period in subjects on prior medications (see “Treatment and assessments” below) subjects were reassessed for eligibility and required to have an IOP ≥24 mmHg and ≤36 mmHg in at least one eye at 8 a.m. at the baseline visit. Exclusion criteria included: closed/barely open anterior chamber angle or a history of acute angle closure; history of discontinued prostaglandin IOP-lowering treatment, unless the reason for discontinuation was participation in a clinical study; ocular surgery or argon laser trabeculoplasty in one or both eyes within 3 months prior to the screening visit; current use of contact lenses; use or anticipated requirement during the study period of topical medication known to affect IOP; anticipated need to modify systemic medication known to affect IOP during the study period; any condition in one or both eyes that would prevent reliable applanation tonometry; ocular inflammation/infection in one or both eyes within 1 month prior to the screening visit or during the washout period; known hypersensitivity to benzalkonium chloride or to any other component of the study medication; other abnormal ocular conditions or symptoms that prevented the subject from participating in the study, according to the investigator’s judgment; participation in any other ocular or systemic clinical study within 1 month prior to the screening visit; other severe, acute, or chronic medical or psychiatric condition or laboratory abnormality that, in the investigator’s judgment, could have increased the risk associated with study participation or study drug administration, or may have interfered with the interpretation of the study results; uncontrolled systemic disease; women of childbearing potential who were not using adequate contraceptive methods or who were pregnant or nursing.
Treatments and assessments
Subjects were assessed for eligibility at a screening visit that took place up to 4 weeks prior to the baseline visit. At screening, ocular and medical histories and previous and concomitant medications were documented; ophthalmoscopy, refraction, biomicroscopy, and a slit lamp examination (to evaluate hyperemia) were performed; and best-corrected visual acuity (BCVA) and IOP were measured. Conjunctival hyperemia was graded according to a 4-point photographic reference scale ranging from 0 (none) to 3 (severe). Throughout the study, IOP was measured in both eyes (the right eye preceding the left eye) using a calibrated Goldmann applanation tonometer. The measurement was done by two study personnel (an operator and a recorder) as recommended by the guidelines from the Eye Care Technology Forum . Measurements were repeated in the same eye twice consecutively. If the difference in measurements was ≤2 mmHg, the mean of these measurements was recorded. If the difference in measurements was >2 mmHg, a third measurement was made and the median of the three measurements was recorded. IOP was measured after other ophthalmic examinations but prior to pupil dilation.
Subjects who were receiving IOP-lowering treatment and who met the eligibility criteria entered into a washout period, the duration of which depended on their current IOP treatment (4 weeks for prostaglandin analogs; 3 weeks for beta-blockers [alone or in combination] or oral carbonic anhydrase inhibitors; 2 weeks for epinephrine and dipivefrin or adrenergic agonists; and 1 week for topical carbonic anhydrous inhibitors, pilocarpine, carbachol and aceclidine). Subjects could attend the study site for safety monitoring during the washout period at the discretion of the investigator and/or the subject’s choice. For subjects not receiving IOP-lowering medications, the screening visit could be combined with the baseline visit.
At the baseline visit, examinations performed at screening were repeated except for refraction; ocular symptoms and adverse events were assessed; and iris/en face photographs were taken (2 sets for each eye; one retained at the study center and one sent to a central reading center). Baseline IOP was measured at 8 a.m. and 4 p.m. (± 1 h). Subjects were randomized to receive latanoprost 50, 75, 100, or 125 μg/mL in a 1:1:1:1 ratio with a computerized randomization list using the random permuted blocks method. A multiple of the block size (n = 8) was distributed to each study center, where the investigator assigned subject numbers consecutively. Subjects were instructed to instill one drop of study medication in one or both eyes in the evening of the baseline visit between 7 p.m. and 9 p.m. and every evening during the entire treatment period with the last dose administered in the evening of the day preceding the week 4 visit. All concentrations of latanoprost contained the same concentration of benzalkonium chloride (0.2 mg/ml).
Follow-up visits occurred at weeks 1, 2, 3, and 4. At each visit, biomicroscopy was performed; BCVA, conjunctival hyperemia, and IOP were evaluated; and ocular symptoms, adverse events, and concomitant medications were recorded. At least 12 h must have elapsed between administration of the evening dose of study medication and measurement of the morning IOP. At week 4, ophthalmoscopy was performed and iris/en face photographs were repeated. Subjects were contacted by telephone 1 week after the week 4 visit to elicit additional information concerning adverse events.
Adverse events, both observed and volunteered, were monitored by investigators throughout the study. The severity of events (mild, moderate, or severe) and the investigator’s opinion concerning whether the event was related to study drug were noted. Serious adverse events were those that were life-threatening, required inpatient hospitalization/prolongation of hospitalization, caused persistent or significant disability/incapacity, or resulted in congenital anomaly/birth defect or death. Adverse events were followed until they resolved or stabilized. Adverse events were considered treatment-emergent if they occurred between baseline and week 4 or if they were present at baseline but increased in severity and/or seriousness during the study.
Endpoints and analyses
Efficacy analyses were performed for both the study eye and the worse eye. The study eye was defined as any eye meeting all inclusion and no exclusion criteria; if both eyes were eligible, the mean value of the IOP recording in both eyes at a given time point was included in analyses. The worse eye was the study eye with the highest baseline 8 a.m. IOP; data from the right eye were used in subjects with two study eyes with equal 8 a.m. IOP levels at baseline. The intention-to-treat (ITT) population included all subjects who received at least one dose of study medication; separate ITT analyses were performed using observed data and using the last observation carried forward (LOCF) method to impute missing IOP data. The per-protocol (PP) population included all subjects who completed the study treatment period without major protocol violations.
The primary efficacy endpoint was the change in IOP from baseline to week 4 (day 28; 8 a.m. and 4 p.m.) in the worse eye, and the primary analysis of this endpoint was conducted in the PP population. Pairwise comparisons of latanoprost 75 μg/mL, 100 μg/mL and 125 μg/mL versus latanoprost 50 μg/mL were made using analysis of covariance (ANCOVA) with baseline IOP as the covariate and center and dose group as factors. A point estimate (least square mean), p-value and a two-sided 90% confidence interval (where the upper limit corresponded to the one-sided test of 5%) for the adjusted treatment differences were calculated. If the treatment effect was not significant, no further models were fitted. If the treatment effect was significant, the model was refitted with terms for treatment by covariate interactions to assess the robustness of the treatment effect across covariates. Standardized residuals were plotted for all models fitted to validate model assumptions. To test the robustness of the results of the primary analysis, analyses were repeated in the PP population for study eyes and in ITT population for both worse and study eyes.
For both worse and study eyes, secondary efficacy endpoints included IOP change from baseline to weeks 1, 2, and 3, analyzed using the PP population, and percentage change in IOP at 8 a.m. 4 weeks postbaseline, analyzed using both the PP and the ITT populations. The statistical significance of pairwise comparisons between latanoprost 50 μg/mL and each higher concentration was tested at each visit and time point using the same model as for the primary efficacy analysis. These efficacy comparisons were one-sided and performed at the 5% level of significance; no adjustment for multiple comparisons was made. The influence of potential prognostic factors (ie, age, gender) on the change in IOP was evaluated using frequency tables for the PP population on the worse eye.
Safety analyses were based on both eyes using the ITT population. Adverse events were classified by system organ class and preferred term using the Medical Dictionary for Regulatory Activities (MedDRA) version 2.3, and treatment-emergent adverse events were tabulated by treatment group. The frequency of cells/flare in the anterior chamber and ocular symptoms was tabulated as was the change from baseline in iris pigmentation and eyelash growth. Hyperemia was assessed using the same scale that was used in the phase III studies of latanoprost (25,28), with 0 = None (no hyperemia), 1 = Mild (reddening of the palpebral or bulbar conjunctiva). 2 = Moderate (bright reddening of palpebral or bulbar conjunctiva) and 3 = Severe (deep, bright, and diffuse reddening of palpebral or bulbar conjunctiva).
The sample size of 240 subjects was based on clinical judgment rather than on formal power calculations. Assuming a standard deviation of 4 mmHg, a study recruiting 50 evaluable subjects per treatment arm would provide 80% power to detect a difference of 2.0 mmHg in the change from baseline IOP (pairwise comparison of higher latanoprost concentrations vs latanoprost 50 μg/mL) at 28 days at the 5% level of significance (one-sided test). Furthermore, the corresponding power would be 58% to detect a difference of 1.5 mmHg.