Gale Andrew J, Bhat Vikas, Pellequer Jean-Luc, Griffin John H, Mosnier Laurent O, Von Drygalski Annette
Department of Molecular and Experimental Medicine, The Scripps Research Institute, 10550 N Torrey Pines Rd, La Jolla, California, USA.
Avelas Biosciences, La Jolla, California, USA.
Pharm Res. 2016 Jun;33(6):1517-26. doi: 10.1007/s11095-016-1895-3. Epub 2016 Mar 9.
Activated (super)Factor V ((super)FVa) is a novel engineered FV with excellent prohemostatic efficacy. (Super)FVa has three APC cleavage site mutations and an interdomain disulfide bond. Stability, pharmacokinetics, and immunogenic and thrombogenic potential are reported here.
Stability and circulating half-life were determined after incubation in buffer and human plasma, and after injection into FVIII-deficient mice. Immunogenicity potential was assessed by B- and T-cell specific epitope prediction and structural analysis using surface area and atomic depth computation. Thrombogenic potential was determined by quantification of lung fibrin deposition in wild-type mice after intravenous injection of (super)FVa (200 U/kg), recombinant human (rh) Tissue Factor (0.4-16 pmol/kg), rhFVIIa (3 mg/kg) or saline.
FVa retained full activity over 30 h in buffer, the functional half-life in human plasma was 4.9 h, and circulating half-life in FVIII-deficient mice was ~30 min. Predicted immunogenicity was not increased compared to human FV. While rh Tissue Factor, the positive control, resulted in pronounced lung fibrin depositions (mean 121 μg/mL), (super)FVa did not (6.7 μg/mL), and results were comparable to fibrin depositions with rhFVIIa (7.6 μg/mL) or saline (5.6 μg/mL).
FVa has an appropriate safety and stability profile for further preclinical development as a prohemostatic against severe bleeding.
活化（超级）因子V（（超级）FVa）是一种新型工程化因子V，具有出色的促止血功效。（超级）FVa有三个活化蛋白C（APC）切割位点突变和一个结构域间二硫键。本文报道了其稳定性、药代动力学以及免疫原性和致血栓形成潜力。
在缓冲液和人血浆中孵育后，以及注射到VIII因子缺乏的小鼠体内后，测定其稳定性和循环半衰期。通过B细胞和T细胞特异性表位预测以及使用表面积和原子深度计算的结构分析来评估免疫原性潜力。通过静脉注射（超级）FVa（200 U/kg）、重组人（rh）组织因子（0.4 - 16 pmol/kg）、rhFVIIa（3 mg/kg）或生理盐水后，对野生型小鼠肺纤维蛋白沉积进行定量，以确定致血栓形成潜力。
FVa在缓冲液中30小时内保持全部活性，在人血浆中的功能半衰期为4.9小时，在VIII因子缺乏的小鼠体内循环半衰期约为30分钟。与人类因子V相比，预测的免疫原性没有增加。阳性对照rh组织因子导致明显的肺纤维蛋白沉积（平均121μg/mL），而（超级）FVa则没有（6.7μg/mL），其结果与rhFVIIa（7.6μg/mL）或生理盐水（5.6μg/mL）的纤维蛋白沉积结果相当。
FVa具有适当的安全性和稳定性，适合作为针对严重出血的促止血剂进行进一步的临床前开发。
