Neutralization of the biological activity of glycosylated and non glycosylated hGM CSF by monoclonal antibodies

OGGERO EBERHARDT, M.; FRANK, R.; KRATJE, R. B.; ETCHEVERRIGARAY, M.

Tylösand. Suecia

Congreso; 17th Meeting of the European Society for Animal Cell Technology (ESACT); 2001

European Society for Animal Cell Technology

Human Granulocyte Macrophage Colony Stimulating Factor (hGM CSF) is a glycoprotein having clinical utility by enhancing the rate of hematopoietic recovery after cancer chemoterapy. Nevertheless, there is increasing evidence of spontaneous human anti GM CSF antibodies associated with chronic infections, autoimmune diseases or simply their ocurrence in normal sera that could compromise the clinical efficacy of the therapy. Here, we show a bioassay based on the inhibition of the biological activity of glycosylated and non glycosylated GM CSF using three monoclonal antibodies (MAbs) against E.coli derived GM CSF (MAbs 1B8, CC5B5 and 7E10). Our previous studies indicated the recognition of two receptor binding sites by these MAbs. The sites were mapped using sets of overlapping peptides as well as generic hexapeptide libraries prepared by SPOT synthesis technique and it was demonstrated that MAb 1B8 and CC5B5 are able to bind to regions of potential N-glycosylation. Depending on the use of the glycosylated or non glycosylated GM CSF, different neutralizing capacity of the MAbs was observed. We compared the amounts of MAbs needed to inhibit the 50% of the biological activity of both GM CSFs. Lower quantity of MAbs 1B8 and CC5B5 was necessary to neutralize the non glycosylated cytokine. Contrarily, MAb 7E10 showed a completely distinct neutralization ability, considering that higher amount of this MAb was needed to produce the same inhibition of the non glycosylated GM CSF with regard to the glycosylated one. The specificity of the antibodies present in patients´sera should play an important role in the failure or success of GM CSF clinical trials. Competition experiments between our MAbs and sera from potential patients to be treated with GM-CSF, should enable us to predict which version of the cytokine could be more effective.