TREK-1 effects on IOP in DEX-induced mouse OHT model

PRIMARY OUTCOME

intraocular pressure reduction

EXPECTED OUTCOMES

- 1. DEX treatment will induce significant IOP elevation (18-25 mmHg) compared to baseline (8-12 mmHg) and vehicle-treated fellow eyes - 2. Single ML-402 0.3% treatment will reduce elevated IOP by 20-35% within 1-2 hours of administration - 3. Peak IOP reduction will occur 2-4 hours post-ML-402 administration with effects lasting 6-8 hours - 4. Chronic ML-402 treatment will provide sustained IOP reduction (15-25% below DEX-elevated levels) throughout treatment period - 5. Dose-response relationship will show ED50 for ML-402 between 0.1-0.3% concentration with maximal effects at 0.3-1.0% - 6. TREK-1 antagonist spadin will block ML-402-induced IOP reduction by 70-90%, confirming mechanism specificity - 7. ML-402 treatment will not cause significant ocular surface toxicity or anterior segment inflammation based on clinical examination

SUCCESS CRITERIA

- • Successful induction of ocular hypertension in >80% of DEX-treated eyes with IOP >15 mmHg consistently - • Achieve statistically significant IOP reduction with ML-402 treatment (p<0.05) compared to vehicle control - • Demonstrate dose-dependent IOP lowering effects with at least 3 concentrations showing progressive responses - • ML-402 effects must be blocked by TREK-1 antagonist (>50% inhibition) to confirm channel-specific mechanism - • Reproducible IOP measurements with coefficient of variation <15% and consistent tonometer calibration - • Complete study without significant dropouts due to complications or technical failures (>85% completion rate) - • Document absence of ocular toxicity through ophthalmoscopic examination and maintain normal corneal clarity

PROTOCOL

**Phase 1: Animal Preparation and DEX-Induced OHT Model** -- Days 1-14 C57BL/6 mice (8-12 weeks, n=10 per group) housed in standard conditions. Baseline IOP measurements using rebound tonometer (TonoLab) under light isoflurane anesthesia. DEX-induced OHT model: topical dexamethasone acetate 0.1% drops (5 μL) applied twice daily to right eyes for 14 days. Left eyes receive vehicle (saline) as controls. Daily IOP monitoring at consistent times (10-11 AM) to track development of ocular hypertension. **Phase 2: ML-402 Treatment Preparation** -- Days 15-16 ML-402 (TREK-1 agonist) dissolved in sterile saline with 0.1% DMSO. Prepare fresh solutions daily. Treatment concentrations: Vehicle control (saline + 0.1% DMSO), ML-402 0.1%, ML-402 0.3%. Osmolality verified (290-320 mOsm) and pH adjusted to 7.0-7.4. Sterility confirmed by plating on agar plates. **Phase 3: Acute IOP Response to ML-402** -- Days 17-18 After 14 days of DEX treatment and confirmed IOP elevation (>15 mmHg), administer single topical ML-402 dose (5 μL) to hypertensive eyes. IOP measurements at: baseline (pre-drop), 30 minutes, 1h, 2h, 4h, 6h, and 8h post-treatment. Use multiple measurements per timepoint (n=5 readings averaged) with coefficient of variation <10%. **Phase 4: Chronic ML-402 Treatment Protocol** -- Days 19-25 Chronic treatment phase: ML-402 drops administered twice daily for 7 days while continuing DEX treatment. IOP measured daily at consistent times. Monitor for signs of ocular irritation or systemic effects. Include washout period assessment (days 26-28) to evaluate duration of effects. **Phase 5: Dose-Response and Mechanism Validation** -- Days 26-30 Conduct dose-response study with ML-402 concentrations: 0.03%, 0.1%, 0.3%, 1.0%. Include TREK-1 antagonist control (spadin 0.1% + ML-402 0.3%) to confirm mechanism. Measure IOP response magnitude and duration for each concentration. Calculate ED50 for IOP reduction. **Phase 6: Statistical Analysis and Safety Assessment** -- Days 31-35 Primary endpoint: percent IOP reduction from elevated baseline. Statistical analysis using repeated measures ANOVA with post-hoc Tukey tests. Safety assessment: ophthalmoscopy for corneal damage, anterior chamber inflammation, lens changes. Histological examination of anterior segment tissues. Power analysis validation for observed effect sizes.

LINKED HYPOTHESES

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[TREK-1 effects on IOP in DEX-induced mouse OHT model](http://scidex.ai/artifact/exp-3df21b8b-ac54-47e4-b733-3cf1fed9bf2d)

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