A 6-Month Study Comparing Efficacy, Safety, and Tolerability of the Preservative-free Fixed Combination of Tafluprost 0.0015% and Timolol 0.5% versus Each of Its Individual Preservative-Free Components

ORIGINAL RESEARCH A 6-Month Study Comparing Efficacy, Safety, and Tolerability of the Preservative-free Fixed Combination of Tafluprost 0.0015% and Timolol 0.5% versus Each of Its Individual Preservative-Free Components Norbert Pfeiffer •Carlo E. Traverso •Katrin Lorenz •Ville Saarela •Johanna Liinamaa • Hannu Uusitalo •Yury Astakhov •Ernest Boiko •Auli Ropo To view enhanced content go to www.advancesintherapy.com Received: August 6, 2014 / Published online: December 2, 2014 ÓThe Author(s) 2014. This article is published with open access at Springerlink.com ABSTRACT Introduction: The efficacy, safety and tolerability of the preservative-free (PF) fixed combination (FC) of tafluprost 0.0015% and timolol 0.5% (once daily) were compared to those of the individual components (PF tafluprost 0.0015% once daily and PF timolol 0.5% twice daily) in patients with open-angle glaucoma or ocular hypertension inadequately controlled on prior timolol or prostaglandin monotherapy for 6 months. Methods: A stratified, double-masked, randomized, multicenter phase III study was conducted. A total of 189 prior timolol users were randomized within the timolol stratum (TS) to receive either FC (n=95) or timolol 0.5% (TIM; n=94). Furthermore, a total of 375 prior prostaglandin analog (PGA) users were randomized within the prostaglandin stratum (PS) to receive either FC (n=188) or tafluprost 0.0015% (TAF; n=187). To be eligible for participation in the study, the patients were required to have an intraocular pressure (IOP) Trial Registration: ClinicalTrial.gov #NCT01292460. For the Preservative-free Tafluprost/Timolol Fixed Combination Study Group. The investigators who participated in the Preservativefree Tafluprost Fixed Combination Study Group are listed in Appendix. Electronic supplementary material The online version of this article (doi:10.1007/s12325-014-0163-3) contains supplementary material, which is available to authorized users. N. Pfeiffer (&)K. Lorenz Department of Ophthalmology, Mainz University Medical Center, Langenbeckstr. 1, 55101 Mainz, Germany e-mail: [email protected] C. E. Traverso Clinica Oculistica, Di.N.O.G.M.I., University of Genoa, IRCCS Azienda Ospedaliera Universitaria San Martino-IST, Genoa, Italy V. Saarela J. Liinamaa Department of Ophthalmology, Medical Research Center, Oulu University Hospital, University of Oulu, Oulu, Finland H. Uusitalo Department of Ophthalmology, University of Tampere and TAUH Eye Center, Tampere, Finland Y. Astakhov First Pavlov State Medical University of St. Petersburg, St. Petersburg, Russia E. Boiko Military Medical Academy, St. Petersburg, Russia A. Ropo Santen Oy, Clinical Research and Medical Affairs, Helsinki, Finland Adv Ther (2014) 31:1228–1246 DOI 10.1007/s12325-014-0163-3 of C22 mmHg when on timolol (TIM) or of C20 mmHg when on PGA in either treated eye at the screening and end-of-run-in visits. In addition to these, the study included visits at baseline, 2 and 6 weeks, 3 and 6 months and at a post-study visit. IOP was measured at 8 a.m., 10 a.m., 4 p.m., and 8 p.m. Results: In the TS, a significant reduction from baseline IOP was seen with FC and TIM throughout the study. Average diurnal IOP change from baseline at month 3 was -8.55 mmHg (32%) for FC and -7.35 mmHg (28%) for TIM. The model-based treatment difference (FC–TIM) was -0.885 mmHg [95% confidence interval (CI) -1.745 to -0.024; p=0.044] demonstrating the superiority of FC over TIM. In the PS, a significant reduction in IOP was seen with both FC and TAF throughout the study. The average diurnal IOP change from baseline at month 3 was -8.61 mmHg (33%) for FC and -7.23 mmHg (28%) for TAF. The model-based treatment difference (FC–TAF) was -1.516 mmHg (95% CI -2.044 to -0.988; p\0.001) demonstrating the superiority of FC over TAF. In the TS, related ocular adverse events (AEs) were more frequent for patients treated with FC compared to TIM (16.8% versus 6.4%), whereas related non-ocular AEs were more frequent with TIM compared to FC (2.1% versus 0.0%). In the PS, AEs were similarly distributed between FC and TAF. The frequency of conjunctival hyperemia of FC was low (6.4%). Conclusion: The preservative-free fixed combination of tafluprost and timolol provided a substantial and significant IOP reduction in both strata. The IOP reduction was superior to both tafluprost 0.0015% and timolol 0.5% when given as monotherapies. Overall, the study treatments were safe and well tolerated. Funding: Santen Oy, Tampere, Finland. Keywords: Glaucoma; Fixed combination; Preservative free; Preservatives; Prostaglandins; Tafluprost; Timolol INTRODUCTION The medical treatment of ocular hypertension and glaucoma is focused on reducing intraocular pressure (IOP) to reach and maintain the individual target IOP. However, for many patients, a single medication is not sufficient in this respect [1,2]. If the target IOP cannot be achieved using a monotherapy, a combination of drugs with different mechanisms of action is recommended [3]. In recent years, the use of fixed combination (FC) glaucoma medications in patients with glaucoma or ocular hypertension has substantially increased. Fixed combinations contain two medications in a single bottle, which may be more convenient for the patient because fewer instillations and bottles are used, and thus likely improves compliance and adherence because of the simpler treatment regimen [4]. Multiple topical therapies may also be associated with a higher incidence of side effects [5]. Increased exposure to preservatives may have untoward effects on the ocular surface and may lead to a higher incidence of ocular signs and symptoms, and poor compliance [4,6–8]. Benzalkonium chloride (BAK) is the most widely used preservative in IOP-lowering ophthalmic preparations. However, it has been demonstrated in a variety of experimental and clinical studies to be pro-apoptotic and pro-inflammatory, to damage tear film layer, corneal epithelium and corneal nerves, and has a negative impact on the number of conjunctival goblet cells [9–13]. Ocular surface disease (OSD) and dry eye syndrome Adv Ther (2014) 31:1228–1246 1229 are frequently detected in patients treated with preserved glaucoma medications [14,15]. It has been shown that OSD is related to the number of preserved eye drops used, the prolonged use of preserved medication and the total benzalkonium chloride (BAK) exposure [13–16]. The use of preservative-free (PF) formulations may avoid these negative effects of BAK [17–19]. Tafluprost ophthalmic solution 0.0015% (Taflotan TM , Saflutan TM , Tapros TM , Santen, Osaka, Japan) was the first prostaglandin analog (PGA) that became available in a PF formulation. The IOPlowering efficacy of this PF formulation is comparable to that of latanoprost [20–22]. PGA/timolol FCs are frequently used in glaucoma management. Both active ingredients of these fixed combinations have a different mode of action [3]. These fixed combinations provide an IOP reduction of approximately 30–37% and have to be instilled only once daily [23,24]. The additivity and safety of tafluprost and timolol administered in a non-fixed combination (NFC) have been demonstrated in two randomized, double-masked, parallel-group, multicenter clinical studies [25,26]. Thus, PF prostaglandin/timolol FC for the treatment of patients with glaucoma and ocular hypertension may have potential benefits for patients using fixed combinations of a prostaglandin analog and timolol. The present study in patients with openangle glaucoma or ocular hypertension was designed to compare the IOP-lowering efficacy, safety and tolerability of the PF FC of tafluprost 0.0015% (TAF)/timolol 0.5% (TIM) ophthalmic solution to those of its individual components, namely, PF tafluprost 0.0015% and PF timolol 0.5%. METHODS Study Design This was a stratified, double-masked, randomized, multinational, multicenter phase III study conducted in 60 centers and 10 countries. The study was registered with ClinicalTrial.gov #NCT01292460, and was reviewed and approved by the independent ethics committees of each participating country. Prior to enrollment, written informed consent was obtained from all patients. The study was conducted in accordance with the current good clinical practice (GCP) requirements. All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national), and with the Helsinki Declaration of 1975, as revised in 2000 and 2008. A power analysis was performed to justify the number of patients enrolled in the study. Study Population Male and female patients, aged 18 years and above, diagnosed with either ocular hypertension or open-angle glaucoma (primary open-angle, pseudoexfoliative or pigmentary glaucoma) in one or both eyes were enrolled in the study. All patients were required to be treated either with a PGA [prostaglandin stratum (PS)] or with timolol 0.5% (TIM) [timolol stratum (TS)] monotherapy for at least 2 weeks before the screening visit. To be eligible for study participation, the patients were required to have IOP in either treated eye of C22 mmHg at any time of the day during the screening visit for prior timolol users (TS) or C20 mmHg for prior prostaglandin analog 1230 Adv Ther (2014) 31:1228–1246 users (PS). Other inclusion criteria were an increase of at least 2 mmHg in the average diurnal IOP (measured at 8:00 a.m., 10:00 a.m., 4:00 p.m and 8:00 p.m.) at the baseline visit as compared to the average diurnal IOP at the endof-run-in period 2 weeks after the screening visit with either open-label timolol 0.5% (TS) or tafluprost 0.0015% (PS) treatment and a best corrected visual acuity no worse than ?0.6 logarithm of the minimal angle of resolution (logMAR) in both eyes. Exclusion criteria were pregnancy and planned pregnancy for the study period; nursing; corneal abnormalities or other conditions preventing reliable applanation tonometry; prior refractive eye surgery; hypersensitivity or contraindication to tafluprost or timolol; prior filtration surgery or any other ocular surgery including intraocular laser procedures within 6 months prior to screening in the eye(s) to be treated; advanced visual field defects in either eye or anticipated progression during the study period; risk for angle closure (\2 grade anterior chamber angle width according to Schaffer’s classification); use of contact lenses at screening or during the study; and inability to safely discontinue the use of ocular hypotensive medications during the wash-out period. Study Visits and Treatment The study visits included screening, end-of-runin, baseline, 2 and 6 weeks, 3 and 6 months, and post-study (Fig. 1). At the screening visit, all eligible patients were assigned according to their prior treatment regimen [timolol 0.5% (TIM) or PGA] to receive either PF timolol 0.5% (TS) twice daily (8:00 a.m. and 8:00 p.m.) or PF tafluprost 0.0015% (PS) once daily (8:00 a.m.) for 2 weeks. The run-in period was followed by a wash-out period of at least 4 weeks. At the baseline visit, the TS patients were randomized to receive either PF timolol 0.5% twice daily (8:00 a.m. and 8:00 p.m.), or PF FC tafluprost 0.0015%/timolol 0.5% at 8:00 a.m. and vehicle at 8:00 p.m. in the affected eye(s). Accordingly, the PS patients were randomized to receive PF tafluprost 0.0015% once daily (8:00 a.m.) or PF tafluprost 0.0015%/timolol 0.5% FC once daily (8:00 a.m.) in the affected eye(s) (Fig. 2). Unilateral dosing of the study medication was also allowed when the contralateral eye required no treatment. All study medications were packed in identical single-dose containers and pouches for masking. Efficacy Variables Diurnal IOP measurements were made by calibrated Goldmann applanation tonometry at 8:00 a.m., 10:00 a.m., 4:00 p.m. and 8:00 p.m. during all visits other than screening and post-study; at these visits, a single IOP measurement was obtained at any time of the day. The primary evaluation of IOP was based on the worse eye and all patients who received at least one dose of the study medications and had at least one post-baseline measurement available [intention to treat (ITT) dataset]. If both eyes met the IOP criteria at baseline the eye with the higher IOP at the 8:00 a.m. IOP measurement was regarded as the worse eye. In case both eyes had the same IOP at baseline, the right eye was designated as the study eye. The primary efficacy variable was the change from baseline in the average diurnal IOP at month 3. The primary statistical objective was to demonstrate that the PF FC tafluprost 0.0015%/timolol 0.5%, administered once daily in the morning is superior to both individual components of the FC: PF tafluprost 0.0015% (TAF) administered once daily in the morning (8:00 a.m.) and PF timolol 0.5% (TIM) administered twice daily (8:00 a.m. and 8:00 Adv Ther (2014) 31:1228–1246 1231 p.m.). Secondary efficacy variables included the proportion of responders at month 3 (defined as change from baseline IOP of 20% or more in increments of 5%), change from baseline in mean diurnal IOP at weeks 2 and 6 and month 6, and change from baseline in the time-wise IOPs at 8:00 a.m., 10:00 a.m., 4:00 p.m., and 8:00 p.m. at weeks 2 and 6 and months 3 and 6. Safety and Tolerability The analysis of the safety and tolerability data was based on all the patients who received at least one dose of the respective study medication and had a subsequent safety measurement. Ocular and non-ocular adverse events (AEs) were recorded at each visit. Slit lamp examinations were conducted at each visit and the findings were graded on a scale from 0 to 3 (0: normal; 1: mild; 2: moderate; 3: severe). Central corneal thickness (CCT) was measured at screening and at month 6 by ultrasound pachymetry. The severity of conjunctival hyperemia was assessed from baseline through month 6 using a set of reference photographs (redness grading) and a 5-point scale (0: none; 1: mild; 2: moderate; 3: severe; 4: very severe). Ophthalmoscopic examinations were conducted via dilated pupil at screening and at months 3 and 6, and included the evaluation of vitreous, retina, and optic nerve head. Visual field testing was performed at screening and post-study. Drop discomfort was evaluated at weeks 2 and 6 and months 3 and 6 on a 4-point scale (0: none, 1: mild, 2: moderate, 3: severe). Resting blood pressure and heart rate were measured twice during the day at 8:00 a.m. and 10:00 a.m. from baseline visit to month 6. Statistical Methods A repeated-measures analysis of the covariance (RM ANCOVA) model was used to evaluate the primary efficacy variable within the TS and PS. A two-sided 95% confidence interval (CI) for the mean difference estimated from the model was used for the evaluation of the superiority hypothesis. Superiority was achieved if the upper limit of the 95% CI (TS: FC–TIM; PS: FC–TAF) was less than 0 mmHg or the corresponding pvalue was less than 0.05. The last observation carried forward (LOCF) method was used for discontinued patients in the ITT dataset at month 3. A Cochran–Mantel– Haenszel (CMH) test was used for the comparison of non-continuous variables between the two treatment groups. The planned sample size was 220 enrolled subjects (110 per arm) in the TS and 380 enrolled subjects (190 per arm) in the PS. These sample Screening visit Visit 1 End-of-run-in visit Visit 2 At least 14 days after screening Baseline visit Prostaglandin stratum (PS): prior PGA users End-of-run-in visit Baseline visit Visit 3 Follow-up visits: Week 2, Week 6, Month 3, Month 6 Day 0 Visits 4–7 Follow up visits: Week 2, Week 6, Month 3, Month 6 Post study visit Post study visit Visit 8 Timolol stratum (TS): prior timolol users Wash-out period Fig. 1 An overview of scheduled study visits for both prostaglandin stratum and timolol stratum. PGA prostaglandin analog, PS prostaglandin stratum, TS timolol stratum 1232 Adv Ther (2014) 31:1228–1246 sizes provided the power of 90% based on differences of 2.0 mmHg for the TS and 1.5 mm Hg for the PS, a standard deviation (SD) of 4.0 mmHg for change in IOP, a drop-out rate of 20% and a two-sided 5% type I error. A smaller drop-out rate than anticipated was seen during the study, thus fewer patients could be randomized in the study without a loss in statistical power. RESULTS In total, 711 subjects were screened for the study. Of these, 564 patients with open or ocular hypertension who met the inclusion criteria were enrolled: 189 patients were randomized and treated in the TS (n=95 patients in the FC arm and n=94 in the TIM arm) and another 375 patients were randomized and treated in the PS (n=188 patients in the FC arm and 187 patients in the TAF arm) (Fig. 2). The treatment arms within both strata were comparable with regard to demographic characteristics and baseline IOP: the majority of patients were female and the mean age was 65.7 ±10.27 years. Primary open-angle glaucoma was the most frequent diagnosis (73.9%), followed by ocular hypertension (22.3%), pseudoexfoliative glaucoma (2.8%) and pigmentary glaucoma (0.9%). In the TS and PS, mean baseline IOPs were similar for both arms at all time points (8:00 a.m., 10:00 a.m., 4:00 p.m. and 8:00 p.m.), and ranged between 25.9 and 26.9 mmHg in the TS and 25.5 and 26.6 mmHg in the PS, respectively (Table 1). A total of 172 patients (91.0%) completed the 6-month study period in the TS: 86 patients (FC 90.5%; TIM 91.5%) in both treatment arms. The most common primary reasons for termination in the TS were: patient request (3 patients for both FC and TIM), AEs (5 patients for FC and 1 patient for TIM) and uncontrolled IOP (1 patient for FC and 2 patients for TIM). In the PS, a total of 348 patients (92.8%) completed the 6-month study period: 175 Prior medical therapy Timolol or PGA Monotherapy Screened: n= 711r Prior timolol users Timolol stratum (TS) Screened: n= 223 Prior PGA users PGA stratum (PS) Screened: n= 488 Patients randomized n= 189 Patients randomized n= 375 FC treatment n=95 TIM treatment n = 94 FC treatment n= 188 TAF treatment n= 187 PF fixed combination (once daily) at 8:00 a.m. and vehicle at 8:00 p.m. PF timolol 0.5% (twice daily) at 8:00 a.m. and 8:00 p.m. PF fixed combination (once daily) at 8:00 a.m. and vehicle at 8:00 p.m. PF tafluprost 0.0015% (once daily) at 8:00 a.m. and vehicle at 8:00 p.m. Fig. 2 Stratification, number of patients, and medical treatment. FC preservative-free fixed combination tafluprost 0.0015%/timolol 0.5%. PGA prostaglandin analog, PS prostaglandin stratum, TAF monotherapy preservative-free tafluprost 0.0015%, TIM monotherapy preservative-free timolol 0.5%, TS timolol stratum Adv Ther (2014) 31:1228–1246 1233 patients (93.1%) in the FC arm and 173 patients (92.5%) in the TAF arm. The most common primary reasons for termination in the PS were: uncontrolled IOP (5 patients for FC and 8 patients for TAF), AEs (6 patients for FC and 2 patients for TAF) and patient request (2 patients for FC and 3 patients for TAF). Only 3 patients had no post-baseline efficacy data; thus, the ITT datasets included 188 and 373 patients in the TS and PS, respectively: in the TS, 95 patients treated with the FC and 93 treated with TIM, in the PS 188 patients treated with FC and 185 patients treated with TAF. Efficacy in Prior Timolol Users (TS) A clinically and statistically significant reduction of IOP from baseline was seen with Table 1 Demographic and baseline characteristics by treatment group Timolol stratum (TS) Prostaglandin stratum (PS) FC (n595) TIM (n594) FC (n5188) TAF (n5187) Gender, n(%) Male 44 (46.3) 36 (38.3) 70 (37.2) 61 (32.6) Female 51 (53.7) 58 (61.7) 118 (62.8) 126 (67.4) Age (years) Mean 64.9 67.4 65.4 65.4 SD 10.3 9.2 10.7 10.3 Range 23–84 40–87 27–85 25–87 Central corneal thickness (lm) Right Left Right Left Right Left Right Left Mean 555 556 554 554 542 541 544 544 SD 36.1 37.4 39.6 43.0 33.6 33.6 34.7 35.7 Range 460–659 475–665 337–640 282–631 442–646 437–635 445–648 448–637 Ocular diagnosis, worse eye n(%) Ocular hypertension 25 (26.3) 23 (24.5) 39 (20.7) 39 (20.9) Primary open-angle glaucoma 69 (72.6) 69 (73.4) 140 (74.5) 139 (74.3) Pseudoexfoliative glaucoma 0 (0.0) 2 (2.1) 7 (3.7) 7 (3.7) Pigmentary glaucoma 1 (1.1) 0 (0.0) 2 (1.1) 2 (1.1) Baseline intraocular pressure, mmHg (mean ±SD) ITT dataset 8:00 a.m. 26.94 (2.84) 26.55 (2.32) 26.35 (3.16) 26.59 (3.21) 10:00 a.m. 26.50 (2.33) 26.32 (2.28) 25.93 (3.22) 26.10 (3.01) 4:00 p.m. 26.32 (2.08) 25.97 (2.36) 25.63 (3.17) 25.59 (3.13) 8:00 p.m. 26.19 (2.30) 25.87 (2.64) 25.54 (3.17) 25.61 (2.74) FC preservative-free fixed combination tafluprost 0.0015%/timolol 0.5%, ITT intention to treat, SD standard deviation, TAF monotherapy preservative-free tafluprost 0.0015%, TIM monotherapy preservative-free timolol 0.5% 1234 Adv Ther (2014) 31:1228–1246 both treatment regimens throughout the study, and low IOP levels were maintained in both treatment arms up to the 6-month visit. Timewise differences at all visits were clearly in favor of the FC: FC lowered IOP between 7.1 and 9.0 mmHg (within group p\0.001 at all time points, ITT dataset). In comparison, TIM 0.5% twice daily lowered IOP between 6.5 and 8.1 mmHg (within group p\0.001 at all time points, ITT dataset) (Fig. 3). At month 3, the time-wise differences (FC–TIM) ranged on average from -1.50 to -0.85 mmHg (Fig. 4), and the estimated average treatment difference (FC–TIM) from the primary RM ANCOVA model for the ITT LOCF dataset was -0.885 mmHg with a 95% CI from -1.745 to -0.024 mmHg (p=0.044), thus superiority of FC over TIM was achieved (Table 2). The sensitivity analysis without using the baseline IOP as a covariate (RM ANOVA) confirmed these results: the treatment difference at month 3 was -1.105 mmHg with a 95% CI from -1.995 to -0.215 mmHg (p=0.015). IOP levels for all individual patients in the TS at baseline and at month 3 and the corresponding box-whisker plots of mean diurnal IOPs (mmHg) at month 3 versus baseline are shown in Fig. 5. It should be noted that one outlier had a substantial effect on the result (Fig. 5a). The results for all secondary efficacy variables were well in line with the primary efficacy variable: the proportion of responders (reduction of mean IOP of 20, 25, 30, and 35%) was in favor of the FC in each response category. A decrease of 35% or more was seen in 33.7% in the FC treatment arm and 23.7% in the TIM treatment arm (p=0.081; CMH-test; ITT LOCF dataset). It should be noted that TS was not properly powered for responder analyses. Furthermore, all estimated time-wise treatment differences (FC–TIM) were clearly in favor of the FC and on average around 1 mmHg; the favorable average difference was sustained up to month 6 (p=0.017; RM ANCOVA; ITT dataset). Efficacy in Prior PGA users (PS) A clinically and statistically significant reduction of IOP from baseline was seen with both treatment regimens throughout the study, and low IOP levels were maintained in both treatment arms up to 6 months. Time-wise differences at all visits were clearly in favor of the FC: FC lowered IOP between 8.2 and Table 2 Overall treatment differences in IOP at month 3 (ITT LOCF dataset)—TS and PS. For the TS, the effect of the outlier is shown separately RM ANCOVA ITT dataset (month 3) PF FC versus monotherapy PF TIM (primary with outlier) PF FC versus monotherapy PF TIM (without outlier) PF FC versus monotherapy PF TAF Difference (mmHg) -0.885 -1.136 -1.516 Upper 95% CI -0.024 -0.379 -0.988 Lower 95% CI -1.745 -1.897 -2.044 pvalue 0.044 0.004 \0.001 CI confidence interval, FC preservative-free fixed combination tafluprost 0.0015%/timolol 0.5%, IOP intraocular pressure, ITT intention to treat, LOCF last observation carried forward, PF preservative free, PS prostaglandin stratum, RM ANCOVA repeat measures analysis of the covariance, TAF monotherapy preservative-free tafluprost 0.0015%, TIM monotherapy preservative-free timolol 0.5%, TS timolol stratum Adv Ther (2014) 31:1228–1246 1235 9.0 mmHg (within group p\0.001 at all time points, ITT dataset). In comparison, TAF lowered IOP between 6.8 and 7.4 mmHg (within group p\0.001 at all time points, ITT dataset) (Fig. 6). At month 3, the time-wise differences (FC–TAF) ranged on average from -1.76 to -1.03 mmHg (Fig. 4) and the estimated average treatment difference (FC– TAF) from the primary RM ANCOVA model for the ITT LOCF dataset was -1.516 mmHg with a 95% CI from -2.044 to -0.988 mmHg (p\0.001), thus the superiority of FC over TAF was achieved (Table 2). The sensitivity analysis without the baseline covariate (RM ANOVA) confirmed these results: the treatment difference at month 3 was -1.402 mmHg with a 95% CI from -1.996 to -0.807 mmHg (p\0.001). IOP levels for all individual patients in the PS at baseline and at month 3 and the corresponding box-whisker plots of mean diurnal IOPs (mmHg) at month 3 versus baseline are shown in Fig. 7. The results for all secondary efficacy variables were well in line with the primary efficacy variable. The proportion of responders (reduction of mean IOP of 20, 25, 30, and 35%) was in favor of the FC: in each response category, there was a clear and statistically significant advantage in favor of the FC. IOP reductions of C30% from baseline were achieved in 61.8% and 37.9% of patients in the FC and TAF treatment arm, respectively (p\0.001; CMH-test; ITT LOCF dataset), and IOP reductions of C35% from baseline was achieved in 38.2% and 23.6% of patients in the FC and TAF treatment arm, respectively (p=0.002; CMH-test; ITT LOCF dataset. Furthermore, all estimated time-wise treatment differences (FC–TAF) were clearly in favor of the FC, on average around 1.5 mmHg, and the favorable average difference was sustained up to 6 months (p\0.001; RM ANCOVA; ITT dataset). Ocular and Non-ocular Adverse Events In the TS, 94 related and non-related AEs (54 ocular, 40 non-ocular) were reported by 43 Fig. 3 Changes of mean (SD) intraocular pressure (IOP) from baseline at weeks 2 and 6, and months 3 and 6 at 8 a.m., 10 a.m., 4 p.m. and 8 p.m. in the TS. Worse eye analysis in the ITT dataset. FC preservative-free fixed combination tafluprost 0.0015%/timolol 0.5%, IOP intraocular pressure, ITT intention to treat, SD standard deviation, TIM monotherapy preservative-free timolol 0.5%, TS timolol stratum 1236 Adv Ther (2014) 31:1228–1246 to those achieved after administering the individual components, PF tafluprost 0.0015% and PF timolol 0.5%. The PF FC tafluprost/ timolol was well tolerated and safe, and associated with a low prevalence of conjunctival hyperemia. ACKNOWLEDGMENTS Sponsorship and article processing charges for this study were provided by Santen Oy, Tampere, Finland. The sponsor participated in the design of the study, conducting of the study, data collection, data management, data analysis, data interpretation, and the preparation, review, and approval of the manuscript. Medical writing and editorial assistance were provided by Friedemann Kimmich, eyecons, Pfinztal, Germany and funded by Santen Oy. All authors had full access to all of the data in this study and take complete responsibility for the integrity of the data and accuracy of the data analysis. All named authors meet the ICMJE criteria for authorship for this manuscript, take responsibility for the integrity of the work as a whole, and have given final approval to the version to be published. Conflict of interest. Norbert Pfeiffer has received research grants and speaker honorariums from Santen Oy, Tampere, Finland, and has received an unrestricted research fund from the Sir Emeka Offor Foundation (Abuja, Nigeria). Carlo Traverso, Katrin Lorenz, Ville Saarela, Johanna Liinamaa, Hannu Uusitalo, Yury Astakhov and Ernest Boiko have received research grants and speaker honorariums from Santen Oy, Tampere, Finland. Auli Ropo is an employee of Santen Oy, Tampere, Finland. Compliance with ethics guidelines. All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2000 and 2008. Informed consent was obtained from all patients for being included in the study. Open Access. This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and the source are credited. APPENDIX The following investigators participated in this multicenter clinical trial: The Preservative-free Tafluprost Fixed Combination Study Group: Hannu Uusitalo (Tampere, Finland), Kai Kaarniranta (Kuopio, Finland), Ville Saarela (Oulu, Finland), Norbert Pfeiffer (Mainz, Germany), Ulrich Bartz-Schmidt (Tuebingen, Germany), Stephan Dunker (Troisdorf, Germany), Thomas Hamacher (Starnberg, Germany), Klaas Heidemann (Leer, Germany), Gu ¨nter Hofmann (Schweinfurt, Germany), Ines Lanzl (Prien, Germany), Karin Oehmig (Chemnitz, Germany), Gernot Petzold (Kulmbach, Germany), Ulrich Richter (Regensburg, Germany), Ulrich Thelen (Muenster, Germany), Axel Zehe (Dresden, Germany), Marie-Luise Scherzer (Regenstrauf, Germany), Nathalie Collignon (Lie `ge, Belgium), U.F. Tegelberg (Goes, Netherlands), Carlo Enrico Traverso (Genoa, Italy), Mirella Blini (Milan, Italy), Emilio Campos (Bologna, Italy), Marco Centofanti (Rome, Italy), Federico Adv Ther (2014) 31:1228–1246 1243 Grignolo (Turin, Italy), Gianluca Manni (Rome, Italy), Enrico Martini (Sassuolo, Italy), Nicola Pescosolido (Rome, Italy), Luciano Quaranta (Brescia, Italy), Odilia Vattovani (Trieste, Italy), Giuseppe Ravalico (Trieste, Italy), Luca Rosetti (Milan, Italy), Vincenzo Russo (Foggia, Italy), Adolfo Sebastiani (Ferrara, Italy), Hana Garzozi (Haifa, Israel), Orna Geyer Haifa, Israel), Shimon Kurtz (Tel Aviv, Israel), Moshe Lusky (Petach Tikva, Israel), Ronit Nesher (Kfar-Saba, Israel), Miriam Zalish (Rehovot, Israel), Simon Longstaff (Sheffield, United Kingdom), Rupert Bourne (Huntingdon, United Kingdom), Timothy Manners (York, United Kingdom), Velota Sung (Birmingham, United Kingdom), Jo ´sef Kaluzny (Bydgoszcz, Poland), Danuta Karczewicz (Szczecin, Poland), Marta MisiukHojlo (Wroclaw, Poland), Jerzy Nawrocki (Lodz, Poland), Wanda Romaniuk (Katowice, Poland), Malgorzata Siewierska (Krako ´w, Poland), Edward Wylegala (Bielsko-Biala, Poland), Tomasz Zarnowski (Lublin, Poland), Kuldar Kaljurand (Tartu, Estonia), Kai Noor (Tallinn, Estonia), Kadri Tammeaid (Tallinn, Estonia), Natalie Feldmann (Kohtla-Jaerve, Estonia), Krista Turman (Tallin, Estonia), Evgeniy Egorov (Moscow, Russia), Valeriy Erichev (Moscow, Russia), Ernest Boiko (St. Petersburg, Russia), Yury Astakhov (St. Petersburg, Russia), Vladimir Alexeev (St. Petersburg, Russia), Alla Ryabteva (Moscow, Russia). 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