Rinkoski, Tommy A.

Radiation exposure to the eye during cancer treatment can lead to ocular radiation injury (ORI), a devastating condition that can have a profound and permanent impact on vision-related quality of life. Rodent models do not have adequate ocular anatomy to accurately simulate human ORI, and modeling in non-human primates is limited by logistical and ethical concerns. To improve future translational research investigating ways to treat or prevent ORI, we developed protocols for a tree shrew model of ORI. Northern tree shrews (Tupaia belangeri) were obtained by our laboratory. Custom housing and handling methods were developed, including custom body suits to maintain the tree shrew’s body temperature during procedures. Radiation delivery was optimized to accurately deliver radiation, and imaging was performed to observe fundus changes from ORI. Optimization of tree shrew handling, housing, anesthesia approaches, radiation delivery, and clinically-relevant ocular imaging permitted successful induction and assessment of ORI in tree shrews. With these protocols, tree shrews can be used as a highly relevant model organism with key anatomic features similar to humans to study ORI.

Aims To determine the associations between measures of oestrogen exposure and Fuchs endothelial corneal dystrophy (FECD) severity. Methods Clinic-based cross-sectional study of 32 postmenopausal women and 11 men with mild or severe FECD, age>55 years. Participants completed questionnaires for data on demographics, anthropometric factors, medical history and potential risk factors for FECD. Women completed an additional reproductive history questionnaire used to calculate the months of lifetime oestrogen exposure. Slit-lamp biomicroscopy, specular microscopy, corneal Scheimpflug tomography and laboratory testing ( TCF4 repeat expansion quantification, total estradiol, free estradiol, sex hormone binding globulin (SHBG), calcaneal bone density) were performed. Logistic regression models were developed to predict FECD severity based on three-way interactions of each oestrogen exposure measure, sex and TCF4 genotype. Results 43 patients were enrolled in the study (mild FECD: 17 women, 3 men; severe FECD: 15 women, 8 men). Serum-free estradiol was higher in the severe compared with mild FECD group (0.21±0.2 vs 0.09±0.1 pg/mL; p=0.046). When stratified by sex, men showed no significant associations between oestrogen measures and FECD severity. However, in women, the odds of severe FECD were increased with more months of lifetime oestrogen exposure (all women; log OR (95% credible interval): 1.3 (0.14 to 4.3)), higher concentrations of free estradiol (all women; 2.1 (0.0049 to 10)), greater % free estradiol (only women without TCF4 repeat expansion; 1.3 (0.16 to 3.8)) and higher concentrations of SHBG (only women with TCF4 repeat expansion; 2.2 (0.45 to 9.1)). Conclusions While the application of these data is constrained by the limited number of participants, a clinic-based sample, small number of men compared with women and single-point measures of serum hormone measures, these data suggest that FECD severity is associated with higher oestrogen exposures in women but not men and may be dependent on TCF4 repeat expansion status.

Purpose: To evaluate pharmacokinetic parameters and ocular hypotensive effects of cromakalim prodrug 1 (CKLP1) in normotensive large animal models. Methods: Optimal CKLP1 concentration was determined by dose response and utilized in short- (5–8 days) and long-term (60 days) evaluation in hound dogs ( n = 5) and African Green Monkeys ( n = 5). Blood pressure was recorded 3–5 times per week with a tail cuff. Concentrations of CKLP1 and the parent compound levcromakalim were assessed in hound dog plasma and select tissues by LC-MS/MS after bilateral ocular treatment with CKLP1 for 8 days. Pharmacokinetic parameters were calculated from days 1, 4, and 8 data. After necropsy, histology was assessed in 43 tissue samples from each animal. Results: In hound dogs and African Green monkeys, 10 mM CKLP1 (optimal concentration) significantly lowered intraocular pressure (IOP) by 18.9% ± 1.1% and 16.7% ± 6.7%, respectively, compared with control eyes ( P < 0.05). During treatment, no significant change in systolic or diastolic blood pressure was observed in either species ( P > 0.1). Average values for half-life of CKLP1 was 295.3 ± 140.4 min, C max, 10.5 ± 1.6 ng/mL, and area under the concentration vs. time curve (AUC last ) 5261.4 ± 918.9 ng·min/mL. For levcromakalim, average values of half-life were 96.2 ± 27 min, C max 1.2 ± 0.2 ng/mL, and AUC last 281.2 ± 110.8 ng·min/mL. No significant pathology was identified. Conclusions: CKLP1 lowered IOP in hound dogs and African green monkeys with no effect on systemic blood pressure. Ocular topical treatment of CKLP1 showed excellent tolerability even after extended treatment periods.

Significance There is a need for a robust and efficient eukaryotic polypeptide display platform for the discovery and affinity maturation of antibodies and other scaffolds, offering a powerful addition to the existing display technologies that use microorganisms. This study is significant in that it demonstrates just such a technology, avian leukosis virus (ALV) polypeptide display, supplying an innovative and substantive departure from the status quo of antibody display that uses platforms based on microorganisms. The use of eukaryotic cells provides the assurance of maximizing the proper folding, posttranslational modifications, and display of the single-chain Fv antibodies and the virus fusion proteins in virions and on the cell surface. ALV display offers virus and cell display platforms in a eukaryotic expression system.