Purification of clinical-grade disulfide stabilized antibody fragment variable—Pseudomonas exotoxin conjugate (dsFv-PE38) expressed in Escherichia coli View Full Text


Ontology type: schema:ScholarlyArticle     


Article Info

DATE

2013-01

AUTHORS

Hua Jiang, Yueqing Xie, Andrew Burnette, John Roach, Steven L. Giardina, Toby T. Hecht, Stephen P. Creekmore, Gautam Mitra, Jianwei Zhu

ABSTRACT

Immunotoxins are rationally designed cancer targeting and killing agents. Disulfide stabilized antibody Fv portion-toxin conjugates (dsFv-toxin) are third generation immunotoxins containing only the antibody fragment variable portions and a toxin fused to the V(H) or V(L). Pseudomonas exotoxin fragment (PE-38) is a commonly used toxin in immunotoxin clinical trials. dsFv-toxin purification was previously published, but the recovery was not satisfactory. This report describes the development of a cGMP production process of the dsFv-toxin that incorporated a novel purification method. The method has been successfully applied to the clinical manufacturing of two dsFv-PE38 immunotoxins, MR1-1 targeting EGFRvIII and HA22 targeting CD22. The two subunits, V(L) and V(H) PE-38 were expressed separately in Escherichia coli using recombinant technology. Following cell lysis, inclusion bodies were isolated from the biomass harvested from fermentation in animal source component-free media. The dsFv-toxin was formed after denaturation and refolding, and subsequently purified to homogeneity through ammonium sulfate precipitation, hydrophobic interaction and ion-exchange chromatography steps. It was shown, in a direct comparison experiment using MR1-1 as model protein, that the recovery from the new purification method was improved three times over that from previously published method. The improved recovery was also demonstrated during the clinical production of two dsFv-PE38 immunotoxins-MR1-1 and HA22. More... »

PAGES

621-632

Identifiers

URI

http://scigraph.springernature.com/pub.10.1007/s00253-012-4319-2

DOI

http://dx.doi.org/10.1007/s00253-012-4319-2

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1042060722

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/22890777


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Download the RDF metadata as:  json-ld nt turtle xml License info

HOW TO GET THIS DATA PROGRAMMATICALLY:

JSON-LD is a popular format for linked data which is fully compatible with JSON.

curl -H 'Accept: application/ld+json' 'https://scigraph.springernature.com/pub.10.1007/s00253-012-4319-2'

N-Triples is a line-based linked data format ideal for batch operations.

curl -H 'Accept: application/n-triples' 'https://scigraph.springernature.com/pub.10.1007/s00253-012-4319-2'

Turtle is a human-readable linked data format.

curl -H 'Accept: text/turtle' 'https://scigraph.springernature.com/pub.10.1007/s00253-012-4319-2'

RDF/XML is a standard XML format for linked data.

curl -H 'Accept: application/rdf+xml' 'https://scigraph.springernature.com/pub.10.1007/s00253-012-4319-2'


 

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