Tina T. Wong

Genome-wide association analyses identify three new susceptibility loci for primary angle closure glaucoma

Primary angle closure glaucoma (PACG) is a major cause of blindness worldwide. We conducted a genome-wide association study including 1,854 PACG cases and 9,608 controls across 5 sample collections in Asia. Replication experiments were conducted in 1,917 PACG cases and 8,943 controls collected from a further 6 sample collections. We report significant associations at three new loci: rs11024102 in PLEKHA7 (per-allele odds ratio (OR) = 1.22; P = 5.33 × 10−12), rs3753841 in COL11A1 (per-allele OR = 1.20; P = 9.22 × 10−10) and rs1015213 located between PCMTD1 and ST18 on chromosome 8q (per-allele OR = 1.50; P = 3.29 × 10−9). Our findings, accumulated across these independent worldwide collections, suggest possible mechanisms explaining the pathogenesis of PACG.

The Relationship of Intraocular Pressure with Age, Systolic Blood Pressure, and Central Corneal Thickness in an Asian Population

PURPOSE To describe the distribution of intraocular pressure (IOP) and its cross-sectional relationship to age, systolic blood pressure (sBP), and central corneal thickness (CCT) in an Asian population. METHODS This was a population-based, cross-sectional study of 3280 Malay subjects (78.7% response) aged 40 to 80 years residing in Singapore. The participants had a standardized interview, examination, and ocular imaging at a centralized study clinic. IOP was measured with Goldmann applanation tonometry (GAT) before pupil dilation, CCT measurements were obtained with an ultrasound pachymeters, and sBP was taken with participants seated after 5 minutes of rest with an automatic blood pressure monitor. RESULTS IOP increased with age to the sixth decade, after which a decrease in IOP was seen with further increase in age, resulting in an inverted U pattern. sBP increased linearly with age whereas CCT decreased linearly with age. In regression models, age, CCT, and sBP were all significant determinants of IOP (P < 0.001 for all three). In younger persons aged 40 to 59 years, both CCT and sBP were positively associated with IOP (P < 0.001 for both), but in older persons of 60 to 80 years, only age and sBP had a positive association with of IOP (P = 0.001 for age, P < 0.001 for sBP). CONCLUSIONS Age, CCT and sBP are significant determinants of IOP in persons aged 40 to 80 years, with CCT being a more important determinant in younger persons. The opposing effects of age-specific changes in sBP and CCT interact to lead to a relatively flat profile of IOP with age, possibly with a subtle inverted U-shaped relationship.

The Singapore 5-Fluorouracil Trabeculectomy Study: Effects on Intraocular Pressure Control and Disease Progression at 3 Years

PURPOSE To report 3-year results of a randomized, controlled trial comparing the use of a single application of 5-fluorouracil (5-FU) with placebo in trabeculectomy surgery. DESIGN Prospective, randomized, double-blinded treatment trial. PARTICIPANTS Two hundred forty-three Asian patients with primary open-angle or primary angle-closure glaucoma undergoing primary trabeculectomy. METHODS One eye of each patient was randomized to receive either intraoperative 5-FU or normal saline (placebo) during trabeculectomy. MAIN OUTCOME MEASURES Primary outcome measure was the level of intraocular pressure (IOP). Secondary outcomes were progression of visual field loss, rates of adverse events, and interventions after surgery. RESULTS Of the 288 eligible patients, 243 were enrolled and 228 completed 3 years follow-up; 120 patients received 5-FU and 123 received placebo. Trial failure, according to predefined IOP criteria, was lower in the 5-FU group compared with the placebo group, although the difference was only significant with a failure criterion of IOP >17 mmHg (P = 0.0154). There was no significant difference in progression of optic disc and/or visual field loss over 36 months between 5-FU and placebo (relative risk [RR], 0.67; 95% confidence interval [CI], 0.34-1.31; P = 0.239). Uveitis occurred more often in the 5-FU-treated group (14/115 [12%] vs 5/120 [4%]; P = 0.032). CONCLUSIONS This is the first masked, prospective, randomized trial reporting the effect of adjunctive 5-FU in trabeculectomy surgery in an East Asian population. The trial shows that an increased success rate can be achieved for several years after a single intraoperative treatment with 5-FU. We conclude that 5-FU is relatively safe and can be routinely used in low-risk East Asian patients. FINANCIAL DISCLOSURE(S) The authors have no proprietary or commercial interest in any materials discussed in this article.

Nanomedicine for glaucoma: liposomes provide sustained release of latanoprost in the eye

Purpose: To report the development and therapeutic evaluation of a liposomal nanocarrier for sustained release of latanoprost, in the rabbit eye. Methods: We fabricated latanoprost-loaded egg-phosphatidylcholine (EggPC) liposomes using the film hydration technique. The delivery vehicles were nano-sized (Z avg = 109 ± 18 nm), had a narrow poly dispersity index (PDI = 0.19 ± 0.04), and a very high loading efficiency (94% ± 5%). Based on in vitro data, we evaluated this formulation for lowering intraocular pressure (IOP) in rabbit eyes. Following a single subconjunctival injection of the latanoprost loaded formulation, the eyes were clinically monitored and the IOP recorded. Results: Latanoprost-loaded EggPC liposomes demonstrated a high drug/lipid mole ratio of 0.181, remained stable for at least 6 months on storage (4°C), and at least 1 month at 25°C. A slow and sustained release of 60% of latanoprost was achieved by 14 days in the in vitro release study. The same formulation demonstrated a greater sustained IOP lowering effect compared with daily administration of topical latanoprost beyond 90 days (4.8 ± 1.5 vs 2.5 ± 0.9 mmHg; P < 0.001). No signs of inflammation were evident in the eyes from slit-lamp examination analysis. Conclusion: The loading required for a long-term sustained delivery of latanoprost for up to 90 days in the rabbit eyes was achieved with EggPC liposomes. A single injection of latanoprost-loaded EggPC liposomes can lower the IOP for up to 90 days, with a greater IOP lowering effect than daily topical administration of latanoprost.

Proteomic profiling of inflammatory signaling molecules in the tears of patients on chronic glaucoma medication.

PURPOSE To identify the tear proteins associated with the long-term use of glaucoma medication by using proteomic analysis and to compare these proteins to those previously reported in primary dry eye disease. METHODS Eighteen patients treated with topical antiglaucoma medications and 10 normal age-matched subjects with no prior topical treatment were recruited for the study. Tears were collected by using Schirmers strip and analyzed by iTRAQ (isobaric tag for relative and absolute quantitation) for tear proteins by mass spectrometry. Conjunctival samples were collected and RNA expression determined by PCR. RESULTS Of the 124 identified tear proteins (99% confidence, ProtScore ≥ 2.0), we found that the tear levels of S100-A8, S100-A9, mammaglobin B, and 14-3-3 ζ/δ were significantly increased in the medicated group compared with levels in the nonmedicated group (P < 0.05). For S100-A9, mammaglobin B, and 14-3-3 ζ/δ, use of topical medication for less than 1 year did not reach statistical significance compared with that in the nonmedicated group. Eyes on topical medication for less than 1 year showed a decrease in proline-rich 4 protein tear level (P = 0.0049) compared to nonmedicated group. The tear proteins detected in the medicated group differed from those in the primary dry eye group. CONCLUSIONS Treatment with topical antiglaucoma medications for longer than 1 year may start to induce ocular surface inflammation. The inflammatory tear protein profile present in chronically medicated glaucomatous eyes appears to be different from that found in primary dry eye. Identification of tear proteins specific to medicated glaucomatous eyes will help to specifically develop targeted screening modalities and therapeutic agents different from current conventional dry eye management.

Combined treatment with bevacizumab and 5-fluorouracil attenuates the postoperative scarring response after experimental glaucoma filtration surgery.

PURPOSE This study evaluated the use of combined bevacizumab with 5-fluorouracil (5-FU) on postoperative scarring and bleb survival after experimental glaucoma filtration surgery in comparison to the agents alone. METHODS Filtration surgery was performed on 26 female New Zealand White rabbits. The rabbits were allocated to one of four treatments: 5-FU combined with bevacizumab, 5-FU alone, bevacizumab alone, or phosphate buffered saline (PBS). The subconjunctival injections were administered immediate postoperatively and weekly for 3 weeks. Clinical assessment and bleb photography were performed. Histologic staining determined the presence of subconjunctvial fibrosis and mRNA expression of collagen I and fibronectin in the tissue was quantified. RESULTS Bevacizumab in combination with 5-FU resulted in a greater antifibrotic effect compared with monotherapy with 5-FU or bevacizumab alone, as evidenced by the attenuation in fibronectin and mature collagen I expression and deposition (P < 0.05). In addition, this was associated with a 100% bleb survival at day 28 in the combined treatment group compared with monotherapy (50% bevacizumab [P < 0.05] and 25% 5-FU [P < 0.001]). Conjunctival vascularity significantly reduced with bevacizumab treatment both alone and in combination with 5-FU. CONCLUSIONS The results provide compelling evidence that combined bevacizumab and 5-FU offers superior antifibrotic effect over monotherapy in a model of glaucoma filtration surgery, while prolonging bleb survival at the same time. A synergistic effect is suggested to be present.

Confirmation of the presence of uveal effusion in Asian eyes with primary angle closure glaucoma: an ultrasound biomicroscopy study.

OBJECTIVE To confirm the presence of uveal effusion in the eyes of Asian patients with primary angle closure glaucoma (PACG) using ultrasound biomicroscopy (UBM). METHODS In this observational case series, 70 patients with PACG (28 untreated patients with newly diagnosed PACG and 42 patients who had undergone previous laser iridotomy and were being monitored) and 12 patients with acute primary angle closure (APAC) were recruited. Eyes of patients with newly diagnosed PACG and APAC underwent UBM before and after laser iridotomy, whereas eyes of patients with treated PACG underwent UBM at enrollment. Uveal effusion was defined as a clear space between the choroid and sclera and was graded as follows: grade 0, none; grade 1, slitlike; grade 2, bandlike; and grade 3, obvious. RESULTS Overall, uveal effusion was found in 11 of 70 eyes with PACG (15.7%; 95% confidence interval, 8.8%-26.2%) and in 3 of 12 eyes with APAC (25%; 95% confidence interval, 8.0%-53.4%). For patients with newly diagnosed PACG, uveal effusion was found in 4 of 28 eyes (14.2%; 95% confidence interval, 5.1%-32.1%) before laser iridotomy; 2 eyes had effusion after laser iridotomy. When present, the effusion was grade 1 in PACG eyes and grade 2 or 3 in APAC eyes. CONCLUSIONS Uveal effusion was present in a significant proportion of Asian eyes with PACG and APAC, confirming a recent report of this finding.

Sustained Release of an Anti-Glaucoma Drug: Demonstration of Efficacy of a Liposomal Formulation in the Rabbit Eye

Topical medication remains the first line treatment of glaucoma; however, sustained ocular drug delivery via topical administration is difficult to achieve. Most drugs have poor penetration due to the multiple physiological barriers of the eye and are rapidly cleared if applied topically. Currently, daily topical administration for lowering the intra-ocular pressure (IOP), has many limitations, such as poor patient compliance and ocular allergy from repeated drug administration. Poor compliance leads to suboptimal control of IOP and disease progression with eventual blindness. The delivery of drugs in a sustained manner could provide the patient with a more attractive alternative by providing optimal therapeutic dosing, with minimal local toxicity and inconvenience. To investigate this, we incorporated latanoprost into LUVs (large unilamellar vesicles) derived from the liposome of DPPC (di-palmitoyl-phosphatidyl-choline) by the film hydration technique. Relatively high amounts of drug could be incorporated into this vesicle, and the drug resides predominantly in the bilayer. Vesicle stability monitored by size measurement and DSC (differential scanning calorimetry) analysis showed that formulations with a drug/lipid mole ratio of about 10% have good physical stability during storage and release. This formulation demonstrated sustained release of latanoprost in vitro, and then tested for efficacy in 23 rabbits. Subconjunctival injection and topical eye drop administration of the latanoprost/liposomal formulation were compared with conventional daily administration of latanoprost eye drops. The IOP lowering effect with a single subconjunctival injection was shown to be sustained for up to 50 days, and the extent of IOP lowering was comparable to daily eye drop administration. Toxicity and localized inflammation were not observed in any treatment groups. We believe that this is the first demonstration, in vivo, of sustained delivery to the anterior segment of the eye that is safe and efficacious for 50 days.

Sustained Drug Release in Nanomedicine: A Long-Acting Nanocarrier-Based Formulation for Glaucoma

Therapeutic nanomedicine has concentrated mostly on anticancer therapy by making use of the nanosize for targeted therapy. Such nanocarriers are not expected to have sustained release of the bioactive molecule beyond a few days. There are other conditions where patients can benefit from sustained duration of action following a single instillation, but achieving this has been difficult in nanosized carriers. An important prerequisite for sustained delivery over several months is to have sufficiently high drug loading, without disruption or changes to the shape of the nanocarriers. Here we report on successful development of a drug-encapsulated nanocarrier for reducing intraocular pressure in a diseased nonhuman primate model and explain why it has been possible to achieve sustained action in vivo. The drug is a prostaglandin derivative, latanoprost, while the carrier is a nanosized unilamellar vesicle. The mechanistic details of this unique drug-nanocarrier combination were elucidated by isothermal titration calorimetry. We show, using Cryo-TEM and dynamic light scattering, that the spherical shape of the liposomes is conserved even at the highest loading of latanoprost and that specific molecular interactions between the drug and the lipid are the reasons behind improved stability and sustained release. The in vivo results clearly attest to sustained efficacy of lowering the intraocular pressure for 120 days, making this an excellent candidate to be the first truly sustained-release nanomedicine product. The mechanistic details we have uncovered should enable development of similar systems for other conditions where sustained release from nanocarriers is desired.

Biocompatibility and biodegradation studies of subconjunctival implants in rabbit eyes.

Sustained ocular drug delivery is difficult to achieve. Most drugs have poor penetration due to the multiple physiological barriers of the eye and are rapidly cleared if applied topically. Biodegradable subconjunctival implants with controlled drug release may circumvent these two problems. In our study, two microfilms (poly [d,l-lactide-co-glycolide] PLGA and poly[d,l-lactide-co-caprolactone] PLC were developed and evaluated for their degradation behavior in vitro and in vivo. We also evaluated the biocompatibility of both microfilms. Eighteen eyes (9 rabbits) were surgically implanted with one type of microfilm in each eye. Serial anterior-segment optical coherence tomography (AS-OCT) scans together with serial slit-lamp microscopy allowed us to measure thickness and cross-sectional area of the microfilms. In vitro studies revealed bulk degradation kinetics for both microfilms, while in vivo studies demonstrated surface erosion kinetics. Serial slit-lamp microscopy revealed no significant inflammation or vascularization in both types of implants (mean increase in vascularity grade PLGA50/50 12±0.5% vs. PLC70/30 15±0.6%; P = 0.91) over a period of 6 months. Histology, immunohistochemistry and immuno-fluorescence also revealed no significant inflammatory reaction from either of the microfilms, which confirmed that both microfilms are biocompatible. The duration of the drug delivery can be tailored by selecting the materials, which have different degradation kinetics, to suit the desired clinical therapeutic application.