کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2020921 1069217 2011 11 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Development of a GMP Phase III purification process for VB4-845, an immunotoxin expressed in E. coli using high cell density fermentation
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Development of a GMP Phase III purification process for VB4-845, an immunotoxin expressed in E. coli using high cell density fermentation
چکیده انگلیسی

VB4-845 is a recombinant immunotoxin comprised of an anti-epithelial cell adhesion molecule (EpCAM) scFv fused to a truncated form of the bacterial toxin, Pseudomonas exotoxin A. VB4-845, purified from TB fed-batch fermentation, showed clinical efficacy when administered locally to treat non-muscle invasive bladder cancer (NMIBC) and squamous cell carcinomas of the head and neck (SCCHN). Here, we describe the implementation of an Escherichia coli high cell density (HCD) cultivation and purification process for VB4-845. HCD cultivation was a prerequisite for achieving higher yields necessary for Phase III clinical trials and commercialization. Using this process, the VB4-845 titer in the supernatant was increased by 30-fold over the original TB fed-batch cultivation. To obtain clinical grade material, a process involving a five-step column purification procedure was implemented and led to an overall recovery of ∼40%. VB4-845 purity of >97% was achieved after the first three columns following the removal of low-molecular weight product-related impurities and aggregates. Endotoxins were effectively separated from VB4-845 on the Q-columns and by washing the Ni-column with a detergent buffer while host cell proteins were removed using ceramic hydroxyapatite. Comparability studies demonstrated that the purified product from the Phase III process was identical to the Phase II reference standard produced using TB fed-batch fermentation.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Protein Expression and Purification - Volume 78, Issue 1, July 2011, Pages 27–37
نویسندگان
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