Novel findings obtained by single chain Fv antibody library establishment from tumor infiltrating B lymphocytes of human breast carcinomas and melanomas

3328 Introduction : Human monoclonal antibodies against tumor-associated ganglioside antigens have the potential for tumor therapeutic reagents. We report here a novel strategy to obtain such antibodies. We aimed to investigate and compare B cells infiltrating breast carcinomas and melanomas (TIL-B), in terms of tumor antigen specific binding potential. We questioned the characteristics of these different tumor types and how the immunoglobuline repertoire of their TIL-B parallels the distinguished features.Description of the work: Different types of breast carcinoma and melanoma tumor tissues were processed for further cellular and molecular studies. Heavy (VH) and light chain (Vκ, Vλ) immunoglobulin variable region genes were amplified. Single chain Fv (scFv) antibody fragments were constructed and phage display libraries generated. We compared our previously developed method to other techniques. The effectiveness of the selection process in respect of tumor associated ganglioside specific antibody fragments was tested against various tumor cell membranes. Selected tumor binder antibody fragments of breast cancer and melanoma TIL-B libraries were investigated in cell ELISA, immunofluorescence FACS analysis, confocal microscopy. DNA sequence analysis and biochemical investigations were done to define the targeted tumor associated antigens. Summary of results: Heavy and light chain immunoglobulin variable region genes could be amplified from any of the investigated tissues. This way single chain Fv antibody fragment phage libraries could be generated and specific tumor binders could be selected.With our technique we obtained human antibody fragmenst with unique GD3 ganglioside specificities on breast carcinomas and melanomas. The anti-GD3 antibody pattern was selective for the cancerous tissues and showed apoptotic effects in different tumor cell cultures. Conclusion: Our results suggest that B cells accumulated in breast carcinomas or melanomas have the potential to reveal key tumor associated antigens. Acknowledgements:Supporting grants (Fulbright Research Scholarship 26.0911, OTKA T048933, OTKA T030380, NATO CLG 978639, Rajko Medenica Research Foundation). Key words: antibody fragments, breast carcinomas, ganglioside, melanoma