结膜下注射表达 Stanniocalcin-1 的腺相关病毒可持续降低小鼠眼压

IF 3.2 Q1 OPHTHALMOLOGY Ophthalmology science Pub Date : 2024-07-31 DOI:10.1016/j.xops.2024.100590
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引用次数: 0

摘要

目的研究结膜下给药单链腺相关病毒 2 号血清型(ssAAV2-STC-1-FLAG)以表达带有 FLAG 标记的斯坦尼钙素-1(stanniocalcin-1),作为一种新型的持续性(眼压)降低药物,同时减少眼表副作用。方法正常血压的 C57BL/6J 小鼠的一只眼球结膜下注射 ssAAV2-STC-1-FLAG(2 μL;6 × 109 病毒基因组 [VGs]),另一只眼球注射相同体积和浓度的 ssAAV2-绿色荧光蛋白(GFP)或相同体积的磷酸盐缓冲盐水。给眼压过高的 DBA/2J 小鼠结膜下注射 6 × 109 VGs 的 ssAAV2-STC-1-FLAG 或 ssAAV2-GFP。每周向 C57BL/6J 小鼠结膜穹窿注射地塞米松诱导类固醇介导的眼压过高,然后向小鼠结膜下注射 6 × 109 VGs 的 ssAAV2-STC-1-FLAG 或 ssAAV2-GFP。前列腺素 F 受体基因敲除小鼠结膜下注射 6 × 109 VGs ssAAV2-STC-1-FLAG 或磷酸盐缓冲盐水。还构建了不含 FLAG 标记的相同载体(ssAAV2-STC-1),并在血压正常的 C57BL/6J 小鼠中进行了评估。所有实验均使用 Tonolab 眼压计评估眼压。主要结果测量眼压评估结果结膜下注射 ssAAV2-STC-1-FLAG 能显著降低正常血压小鼠注射后 10 周的眼压。最大眼压降幅出现在注射后第 3 周(17.4%;17.1 ± 0.8 vs. 14.1 ± 0.8 mmHg,P < 0.001)。在降低眼压的作用减弱后,第二次注射可恢复降眼压作用。在实验期间,结膜下注射 ssAAV2-STC-1-FLAG 可降低 DBA/2J 小鼠(16.9%;17.8 ± 2.0 vs. 14.8 ± 0.9 mmHg,P <;0.001)和类固醇介导的眼压升高小鼠(20.0%;19.0 ± 0.6 vs. 15.2 ± 0.7 mmHg,P <;0.001)的眼压。野生型小鼠(15.9%)和FP受体基因敲除小鼠(15.7%)的眼压降低程度与同侧眼相似。一种相关的构建物也以类似的方式降低了没有FLAG标记的小鼠的眼压。结论用载体系统结膜下递送STC-1转基因可能是一种持续降低眼压和改善眼部耐受性的新型治疗策略,同时也避免了现有药物的每日用药要求。
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Subconjunctival Administration of an Adeno-Associated Virus Expressing Stanniocalcin-1 Provides Sustained Intraocular Pressure Reduction in Mice

Purpose

To investigate subconjunctival administration of a single-stranded, adeno-associated virus, serotype 2, engineered to express stanniocalcin-1 with a FLAG tag (ssAAV2-STC-1-FLAG) as a novel sustained (IOP) lowering agent with a reduced ocular surface side effect profile.

Design

In vivo preclinical investigation in mice.

Subjects

C57BL/6J, DBA/2J, prostaglandin F (FP) receptor knockout mice.

Methods

Normotensive C57BL/6J mice were treated with a subconjunctival injection of ssAAV2-STC-1-FLAG (2 μL; 6 × 109 viral genomes [VGs]) in 1 eye and the same volume and concentration of ssAAV2-green fluorescent protein (GFP) or the same volume of phosphate-buffered saline in the fellow eye. Ocular hypertensive DBA/2J mice were subconjunctivally injected with 6 × 109 VGs of ssAAV2-STC-1-FLAG or ssAAV2-GFP. Steroid-mediated ocular hypertension was induced in C57BL/6J mice with weekly injections of dexamethasone into the conjunctival fornix, and mice were then injected subconjunctivally with 6 × 109 VGs of ssAAV2-STC-1-FLAG or ssAAV2-GFP. Prostaglandin F receptor knockout mice were injected subconjunctivally with 6 × 109 VGs of ssAAV2-STC-1-FLAG or phosphate-buffered saline. An identical vector was constructed without the FLAG tag (ssAAV2-STC-1) and evaluated in normotensive C57BL/6J mice. Intraocular pressure was assessed using the Tonolab tonometer for all experiments. Tumor necrosis factor alpha (TNFα), a marker of ocular surface inflammation, was compared between subconjunctivally delivered ssAAV2-STC-1-FLAG and other treatments including daily topical latanoprost.

Main Outcome Measures

Intraocular pressure assessment.

Results

Subconjunctival delivery of ssAAV2-STC-1-FLAG significantly reduced IOP for 10 weeks post injection in normotensive mice. Maximal IOP reduction was seen at week 3 postinjection (17.4%; 17.1 ± 0.8 vs. 14.1 ± 0.8 mmHg, P < 0.001). After the IOP-lowering effect had waned, a second injection restored the ocular hypotensive effect. Subconjunctivally delivered ssAAV2-STC-1-FLAG lowered IOP in DBA/2J mice (16.9%; 17.8 ± 2.0 vs. 14.8 ± 0.9 mmHg, P < 0.001) and steroid-mediated ocular hypertensive mice (20.0%; 19.0 ± 0.6 vs. 15.2 ± 0.7 mmHg, P < 0.001) over the experimental period. This construct also reduced IOP to a similar extent in wild-type (15.9%) and FP receptor knockout (15.7%) mice compared with the fellow eye. A related construct also lowered IOP without the FLAG tag in a similar manner. Reduction in conjunctival TNFα was seen when comparing subconjunctivally delivered ssAAV2-STC-1-FLAG to daily topical latanoprost.

Conclusions

Subconjunctival delivery of the STC-1 transgene with a vector system may represent a novel treatment strategy for sustained IOP reduction and improved ocular tolerability that also avoids the daily dosing requirements of currently available medications.

Financial Disclosure(s)

Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

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Ophthalmology science
Ophthalmology science Ophthalmology
CiteScore
3.40
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审稿时长
89 days
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Barriers to Extracting and Harmonizing Glaucoma Testing Data: Gaps, Shortcomings, and the Pursuit of FAIRness Severity Scale of Diabetic Macular Ischemia Based on the Distribution of Capillary Nonperfusion in OCT Angiography Editorial Board Table of Contents Cover
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