Heterogeneous nucleation-controlled particulate formation of recombinant human platelet-activating factor acetylhydrolase in pharmaceutical formulation
Heterogeneous nucleation-controlled particulate formation of recombinant human platelet-activating factor acetylhydrolase in pharmaceutical formulation
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DOI：
10.1002/jps.20237
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发表时间：
2005-02-01
影响因子：
3.8
通讯作者：
Randolph, TW
中科院分区：
文献类型：
--
作者：
Chi, EY;Weickmann, J;Randolph, TW
临床大量制备重组人血小板活化因子乙酰水解酶(rhPAF-AH)冻干制剂。在与注射用无菌水重组形成水溶液(10 MM柠檬酸钠，7.5 w/v%蔗糖，0.1 w/v%Pluronic-F68，pH 6.5)后，在室温下，几个可见的缓慢生长的颗粒在数小时内稳定形成。为了探讨这一现象的机制，重组后立即通过过滤去除所有蛋白质聚集体和外源颗粒。在室温下孵育20天，这些过滤的样品中没有可见的聚集体形成。相反，当加入纳米亲水二氧化硅颗粒时，它们可以快速而广泛地聚集重组人PAF-AH。在浓度较低的Pluronic-F68浓度为0.01%的溶液中，这种影响加剧。即使在rhPAF-AH吸附可逆的条件下也会发生聚集，并且没有引起蛋白质二级和三级结构的可检测到的变化。降低rhPAF-AH在纳米二氧化硅颗粒上的吸附程度(例如，添加Pluronic-F68)或亲和力(例如，增加溶液pH)被发现在减少聚集方面是有效的。将聚集态的rhPAF-AH接种于rhPAF-AH制剂中，未观察到加速聚集现象。这些结果表明，rhPAF-AH的聚集过程是通过非均相成核控制的机制进行的，溶液中存在的外源颗粒作为种子在其上吸附、成核并生长成大的聚集体。(C)2004年Wiley-Liss，Inc.和美国药剂师协会。
Clinical lots of recombinant human platelet-activating factor acetylhydrolase (rhPAF-AH) were prepared in a lyophilized formulation. After reconstitution with sterile water for injection to form an aqueous solution (10 MM sodium citrate, 7.5 w/v% sucrose, and 0.1 w/v% Pluronic-F68, pH 6.5), a few visible, slowly growing particles formed consistently within hours at room temperature. To investigate the mechanism of this phenomenon, immediately after reconstitution, all protein aggregates and exogenous particles were removed by filtration. During 20 days incubation at room temperature, no visible aggregates formed in these filtered samples. In contrast, when nano-sized hydrophilic silica particles were added, they seeded rapid and extensive aggregation of rhPAF-AH. This effect was exacerbated in solutions containing a lower Pluronic-F68 concentration at 0.01%. Aggregation occurred even under conditions where rhPAF-AH adsorption was reversible, and induced no detectable changes to protein secondary and tertiary structures. Decreasing the extent (e.g., adding Pluronic-F68) or affinity (e.g., increasing solution pH) of rhPAF-AH adsorption on nano-sized silica particles was found to be effective at reducing aggregation. Accelerated aggregation was not observed when rhPAF-AH formulation was seeded with aggregated rhPAF-AH. These results show that rhPAF-AH aggregation proceeds through a heterogeneous nucleation-controlled mechanism, where exogenous particles present in solution serve as seeds on which rhPAF-AH adsorb, nucleate, and grow into large aggregates. (C) 2004 Wiley-Liss, Inc. and the American Pharmacists Association.