Vector and strain system for the in vivo conversion of antibody fragments into IgG molecules

DESCRIPTION provided by applicant Current methods for converting an antibody fragment for example scFv Fab Fabandapos etc involves subcloning from an M phagemid or yeast display vector by either restriction enzyme digestion or PCR followed by ligation into or more vectors to produce the heavy and light chains transformation into E coli and subsequent DNA sequencing validation The proposed scFv andgt IgG method using recombineering changes this paradigm for subcloning antibody fragments to produce IgG molecules into a simple transduction of a properly modified phagemid or yeast display vector into a genetically modified strain of E coli harboring a specially modified IgG expression shuttle plasmid The costs and labor are reduced from approximately $ $ for traditional subcloning and DNA sequencing analysis to under $ per clone conversion Additionally automation equipment is unnecessary and the low error rate of recombineering is expected to obviate the need for DNA sequence validation AxioMx is developing a pipeline for the rapid discovery less than two weeks of recombinant Abs Completion of the objectives of this proposal will allow researchers to develop and quickly assemble IgG molecules which could be useful for high throughput proteome analysis diagnostics and immunotherapeutics The ability to clone immuno pools from phage yeast and other display technologies while keeping the heavy and light chains linked is a significant advantage to the proposed method We expect this method to increase the efficient production of better antibodies with implications for both diagnostics and therapeutics PUBLIC HEALTH RELEVANCE The market for antibodies in research is approximately $ B with the market for custom antibodies andgt $ M and expected to grow at least until C Bird lead analyst Frost and Sullivan report Strategic Analysis of the Research Antibody Market March Most recombinant antibodies are screened in display as antibody fragments Current methods for converting an antibody fragment for example scFv Fab Fabandapos etc involves subcloning from an M phagemid or yeast display vector by either restriction enzyme digestion or PCR followed by ligation into or more vectors to produce the heavy and light chains transformation into E coli and subsequent DNA sequencing validation The proposed method will reduce the costs and labor from $ $ for traditional subcloning including DNA sequencing analysis to under $ per clone conversion Additionally the need for automation equipment will be limited even in high throughput mode And the low error rate of recombineering is expected to obviate the need for DNA sequence validation Finally unlike almost all other methods the recombineering method links the heavy and light chains together So that transformation of the novel strain of E coli with a pool of display clones will keep together within the bacterium the heavy and light chains that bind to the target Clones will be able to be tested in mammalian cells faster and cheaper