Belgaumi A F, Patrick C C, Deitcher S R
Department of Hematology/Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA.
Am J Hematol. 1999 Sep;62(1):13-8. doi: 10.1002/(sici)1096-8652(199909)62:1<13::aid-ajh3>3.0.co;2-x.
Minipumps may facilitate cost-effective and convenient continuous infusion (CI) therapy for severe hemophilia A. This study evaluated the in vitro sterility, ability to support bacterial growth, and specific activity stability of a recombinant factor VIII (FVIII; Bioclate, Centeon) delivered by simulated CI at a variety of temperatures and after the addition of heparin or antibiotic. Closed system CIs of Bioclate (89.5 IU/ml) with and without heparin were sampled and cultured over a 6 day period. Bioclate (53.7 IU/ml) with and without heparin or vancomycin was inoculated with 102-105 CFU/ml of S. aureus, S. epidermidis, Escherichia coli, E. cloacae, or Y. enterocolitica and assessed by quantitative culture after 1 and 3 days. The stability of Bioclate (50, 100, and 250 IU/ml) at three temperatures (21 degrees C, 37 degrees C, and 39 degrees C) with and without heparin or vancomycin was tested over a period of 28 days. FVIII activity was measured in triplicate by a chromogenic assay (Coamatic Factor VIII, Chromogenix) and purity evaluated by Western blot. No bacterial growth was detected during CI of FVIII for up to 6 days. Following bacterial inoculation, there was rapid growth (>3 log increase) of all tested bacterial species except S. aureus which only displayed a 1 log expansion at 3 days. The addition of heparin containing 9.45 microg/U benzyl alcohol had no effect on bacterial growth. The addition of vancomycin caused a modest suppression of S. aureus growth but not of E. coli. Diluent alone did not support bacterial growth. Neither concentration, increased temperature, nor the addition of heparin or vancomycin had a significant effect on FVIII activity stability. Samples retained >75% baseline activity for between 3 and 7 days, except the infusion of Bioclate 50 IU/ml plus heparin maintained at 21 degrees C which remained stable for 28 days. Western blot analysis supported the activity assay findings. Standard and concentrated preparations of Bioclate are suitable for CI when delivered by the MiniMed 404-SP minipump. Because of the observed nutritive capability of this FVIII concentrate for sustaining bacterial growth, any contamination could result in systemic infection.
微型泵可能有助于为重度甲型血友病患者提供经济高效且便捷的持续输注（CI）治疗。本研究评估了通过模拟CI在多种温度下以及添加肝素或抗生素后，重组凝血因子VIII（FVIII；百因止，Centeon公司）的体外无菌性、支持细菌生长的能力以及比活性稳定性。对含肝素和不含肝素的百因止（89.5 IU/ml）封闭系统CI进行采样，并在6天内进行培养。将含肝素和不含肝素或万古霉素的百因止（53.7 IU/ml）接种102 - 105 CFU/ml的金黄色葡萄球菌、表皮葡萄球菌、大肠杆菌、阴沟肠杆菌或小肠结肠炎耶尔森菌，在1天和3天后通过定量培养进行评估。在28天内测试了含肝素和不含肝素或万古霉素的百因止（50、100和250 IU/ml）在三个温度（21℃、37℃和39℃）下的稳定性。通过发色法（Coamatic Factor VIII，Chromogenix公司）一式三份测量FVIII活性，并通过蛋白质印迹法评估纯度。在FVIII的CI过程中，长达6天未检测到细菌生长。细菌接种后，除金黄色葡萄球菌在3天时仅显示1个对数级的生长外，所有测试细菌种类均快速生长（>3个对数级增加）。添加含9.45 μg/U苯甲醇的肝素对细菌生长无影响。添加万古霉素对金黄色葡萄球菌的生长有适度抑制作用，但对大肠杆菌无抑制作用。单独的稀释剂不支持细菌生长。浓度、温度升高以及添加肝素或万古霉素对FVIII活性稳定性均无显著影响。样品在3至7天内保持>75%的基线活性，除了在21℃下保存的输注百因止50 IU/ml加肝素的样品在28天内保持稳定。蛋白质印迹分析支持活性测定结果。当通过美敦力404 - SP微型泵进行输注时，百因止的标准制剂和浓缩制剂适用于CI治疗。由于观察到这种FVIII浓缩物具有支持细菌生长的营养能力，任何污染都可能导致全身感染。
