Journal of Vaccines & Vaccination

Early stage development of cancer vaccine based on Her1 extracellular domain: Impact of process changes in biological activity

10^th Euro Global Summit and Expo on Vaccines & Vaccination

June 16-18, 2016 Rome, Italy

Katia Garcia Duardo

Center of Molecular Immunology, Cuba

Posters & Accepted Abstracts: J Vaccines Vaccin

Abstract:

Vaccine preparations based on the extracellular domain of Her1 (Her1-ECD) have demonstrated, both in vitro and in vivo, a potent anti-metastatic effect on EGFR+ Lewis lung carcinoma model, while associated side effects were absent. The Her1-ECD is a glycoprotein with a molecular weight of 105 kDa and has 11 potential sites for N-glycosylation. Currently, in Cuba this vaccine has being evaluated in patients with refractory prostate cancer. Her1-ECD molecule (used for the Cuban clinical trials are obtained from culture supernatant of HEK 293 transfectomes used the culture medium PFHM/PFCHO and is purified using immunoaffinity chromatography. Several studies have demonstrated that glycosylation can affect the protein folding, stability, regulates protein half-life, immunogenecity, biological activity and other functions. In this work, the N-glycosylation of Her1-ECD was preliminarily characterized by SDS-PAGE, glycan differentiation by lectin and normal phase chromatography. Afterwards, the biological activity of the glycosylated and fully deglycosylated Her1-ECD protein was compared. In addition, a comparability study was performed, in order to evaluate the impact of process changes (different culture media) in the physicochemical and biological characteristics of Her1-ECD protein. Such study was carrying out using different techniques such as: SDS-PAGE, SEC-HPLC, isoelectric point, peptide mapping, mass spectrometry, SCX-HPLC, oligosaccharide map, ELISA and flow cytometric. As principal results were obtained that N-glycosylation profile of Her1-ECD is composed of high mannose, hybrid and complex N-glycans types and Her1-ECD glycosylation modifies the humoral immune response, measured as antibody titers, recognition of EGFR in A431 cell line and cell cycle arrest. On the other hand, comparability studies results indicated that media changes has an effect in the degree of sialylation of the protein but does not affect its biological activity (measured as titers of Abs and recognition for A431 cell line).

Biography :

Email: katiag@cim.sld.cu