Glycoconjugation of single chain antibodies (scFv) with various molecules for cancer diagnosis and treatment: To extend our bioconjugation work towards the immunoliposome formulation, we have taken up single chain antibodies (scFv) against CD22 and HER-2 proteins. The cDNAs of scFv against CD22 and HER-2 proteins were obtained from Dr. Dimitrovs group. These cDNAs were manipulated such that the single chain antibodies expressed in E. coli have a C-terminal 17 amino acid tag peptide. Although both the scFv antibodies share high protein sequence similarity, the anti HER-2 scFv is expressed very poorly, thus only micro gram quantities can be obtained where as the anti CD22 scFv has been expressed very well and tens of milligram quantity of refolded protein can be obtained. These proteins will be glycosylated using ppGalNAc-T and bioconjugated with fluro probes or MRI contrast agents or with lipid moieties for formulation of immunoliposomes for targeted drug delivery.
Synthesis of lipids carrying aminooxy or alkyne group for linking with a glycoprotein that has a sugar moiety linked with an orthogonal reactive group: The lipid molecule 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) was converted into either DPPE-aminooxy or DPPE-alkyne derivatives for conjugation with scFV that carry sugar moiety with an orthogonal reactive group. The scFV molecules carrying lipid molecules will next used (in a collaborative project with Dr. Blumenthals and Dr. Dimitrovs groups), for the formulation of liposomes for the targeted drug delivery.
