DEVELOPMENT OF STANDARDS FOR FACTORS VII AND IX

因素 VII 和 IX 的标准制定

• 批准号: 3748294
• 负责人: M WEINSTEIN
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3748294
• 关键词: chemical stability; chromatography; coagulation factor IX; coagulation factor VIII; drug preservation; drug quality /standard; monoclonal antibody; recombinant proteins; western blottings

New national standards for factor VIII and factor IX must be manufactured to replace diminishing supplies of MEGA, the factor VIII standard, and FN2, a factor IX standard. The standards must be stabile, have assay characteristics that are compatible with all currently licensed preparations, and give equivalent one-stage plasma and chromogenic test results in order to harmonize with the recommendations of the European Pharmacopoeia. To meet these requirements, we evaluated source material supplied by four manufacturers. Factor VIII source material included two recombinant products, two monoclonal antibody-purified preparations, and two products purified by conventional chromatography. Factor IX bulks included one product purified by monoclonal antibody, two by chromatography, and one by fractional precipitation. Factor VIII was evaluated by plasma and chromogenic substrate tests; factor IX was assayed in plasma. The short term stability of both factors was measured after they underwent a thaw, freeze, thaw cycle, similar to conditions that will occur during the manufacture of the standards. The structural integrity of the factors was evaluated by Western blots. After re thawing, 3 of the 6 factor VIII preparations had plasma potencies within 25% of their initial labeled values. One preparation lost 70% of its activity. Two of the three stabile preparations had chromogenic test results within 15% of their plasma values. After incubation at 30oC for 150 hours, most samples maintained their potency within 10% of initial values. Western blot results were similar for all factor VIII products. Regarding factor IX, the thaw-freeze-thaw cycle had little effect on activity. Two factor IX products were stabile when incubated at 30oC for up to 75 hours, while the others lost activity. Only one factor IX product showed no degradation on a Western blot. We conclude that two products are suitable sources for the factor VIII standard, and one for the factor IX standard.

必须制造新的国家第VIII和第IX因子标准 要取代Mega的供应减少，VIII标准和 FN2，一个因素IX标准。 标准必须稳定，有测定 与当前所有许可的所有特征 准备工作，并进行等效的一阶段等离子体和发色测试 结果是为了与欧洲的建议协调 药典。为了满足这些要求，我们评估了原始资料 由四个制造商提供。 VIII源材料包括两个 重组产物，两个单克隆抗体纯化的制剂，以及 通过常规色谱纯化的两种产品。 因子IX批量 包括一种由单克隆抗体纯化的产品，两种 色谱法，一个通过分数沉淀。 因子VIII是 通过血浆和发色底物测试评估；因子IX是 在血浆中测定。测量了这两个因素的短期稳定性 他们融化后，冻结，解冻周期，类似于条件 这将在制造标准期间发生。 结构 通过蛋白质印迹评估因素的完整性。 RE之后 解冻，6个因子VIII制剂中有3个具有等离子体的功能 其初始标记值的25％。 一项准备工作损失了70％ 活动。 三个稳定制剂中的两个进行了成色 结果在其血浆值的15％之内。 在30oC下孵育后 150小时，大多数样品在初始的10％以内保持了效力 值。所有因子VIII产品的蛋白质印迹结果都相似。 关于因子IX，解冻冻结周期对 活动。 当在30oC下孵育时，两个因子IX产品是稳定的 长达75小时，而其他人则失去了活动。 只有一个因素IX 产品在蛋白质印迹上没有降解。 我们得出的结论是两个 产品是VIII标准因素的合适来源，一个用于 因子IX标准。