Altiok Eda I, Browne Shane, Khuc Emily, Moran Elizabeth P, Qiu Fangfang, Zhou Kelu, Santiago-Ortiz Jorge L, Ma Jian-Xing, Chan Matilda F, Healy Kevin E
Department of Bioengineering, University of California, Berkeley, California, United States of America.
Centre for Research in Medical Devices (CÚRAM), National University of Ireland Galway, Galway, Ireland.
PLoS One. 2016 Jun 3;11(6):e0155990. doi: 10.1371/journal.pone.0155990. eCollection 2016.
Current anti-VEGF drugs for patients with diabetic retinopathy suffer from short residence time in the vitreous of the eye. In order to maintain biologically effective doses of drug for inhibiting retinal neovascularization, patients are required to receive regular monthly injections of drug, which often results in low patient compliance and progression of the disease. To improve the intravitreal residence time of anti-VEGF drugs, we have synthesized multivalent bioconjugates of an anti-VEGF protein, soluble fms-like tyrosine kinase-1 (sFlt) that is covalently grafted to chains of hyaluronic acid (HyA), conjugates that are termed mvsFlt. Using a mouse corneal angiogenesis assay, we demonstrate that covalent conjugation to HyA chains does not decrease the bioactivity of sFlt and that mvsFlt is equivalent to sFlt at inhibiting corneal angiogenesis. In a rat vitreous model, we observed that mvsFlt had significantly increased intravitreal residence time compared to the unconjugated sFlt after 2 days. The calculated intravitreal half-lives for sFlt and mvsFlt were 3.3 and 35 hours, respectively. Furthermore, we show that mvsFlt is more effective than the unconjugated form at inhibiting retinal neovascularization in an oxygen-induced retinopathy model, an effect that is most likely due to the longer half-life of mvsFlt in the vitreous. Taken together, our results indicate that conjugation of sFlt to HyA does not affect its affinity for VEGF and this conjugation significantly improves drug half-life. These in vivo results suggest that our strategy of multivalent conjugation could substantially improve upon drug half-life, and thus the efficacy of currently available drugs that are used in diseases such as diabetic retinopathy, thereby improving patient quality of life.
目前用于治疗糖尿病视网膜病变患者的抗血管内皮生长因子（VEGF）药物在眼玻璃体内的停留时间较短。为了维持抑制视网膜新生血管形成的生物有效剂量的药物，患者需要每月定期注射药物，这往往导致患者依从性较低且疾病进展。为了提高抗VEGF药物在玻璃体内的停留时间，我们合成了一种抗VEGF蛋白的多价生物共轭物，即可溶性fms样酪氨酸激酶-1（sFlt），它与透明质酸（HyA）链共价连接，这种共轭物被称为mvsFlt。使用小鼠角膜血管生成试验，我们证明与HyA链的共价连接不会降低sFlt的生物活性，并且mvsFlt在抑制角膜血管生成方面与sFlt相当。在大鼠玻璃体模型中，我们观察到与未共轭的sFlt相比，mvsFlt在2天后的玻璃体内停留时间显著增加。计算得出sFlt和mvsFlt在玻璃体内的半衰期分别为3.3小时和35小时。此外，我们表明在氧诱导的视网膜病变模型中，mvsFlt在抑制视网膜新生血管形成方面比未共轭形式更有效，这种效果很可能是由于mvsFlt在玻璃体内的半衰期更长。综上所述，我们的结果表明sFlt与HyA的共轭不会影响其对VEGF的亲和力，并且这种共轭显著提高了药物半衰期。这些体内结果表明，我们的多价共轭策略可以大幅提高药物半衰期，从而提高目前用于治疗糖尿病视网膜病变等疾病的现有药物的疗效，进而改善患者生活质量。
