Prostate carcinoma (PCa) is by far the most common non-cutaneous malignancy in men in countries with high standard of living. Therefore, huge efforts are invested into the search of highly specific and selective markers of PCa. Prostate-specific membrane antigen (PSMA) is a prominent PCa marker due to the established correlation between cancer progression and PSMA expression levels. Among PSMA-specific reagents developed for diagnostics and therapy antibodies are of high interest due to high affinity and selectivity for PSMA. Compared to small molecule PSMA-specific inhibitors, the use of antibodies can result in significantly lower toxicity in non-target tissues such as salivary glands. Furthermore, protein engineering techniques enable generation of small recombinant derivatives or humanized variants of antibodies favorable for in vivo applications.
We have recently isolated and characterized in detail two new PSMA-specific antibodies - 5D3 and 5B1, with high affinity and selectivity for human PSMA. Using cancer cell lines and a mouse PCa xenograft model we have shown that both molecules are suitable for in vitro and in vivo applications and reveal approximately 10-times higher affinity for PSMA compared to the leading second-generation antibody J591. Our results confirm the antibodies to be promising candidates for the development of new PSMA-specific diagnostic and therapeutic tools. We currently produced and characterized Fab and single chain fragments derived from antibody 5D3. Analysis in vitro revealed that purified recombinant fragments retain functional binding site of the original 5D3 antibody. Moreover, in vivo imaging experiments run in mouse model of grafted PCa confirm that produced molecules are suitable for in vivo applications. To uncover interaction of PSMA with 5D3 antibody in detail, structural features of PSMA-5D3 complex were mapped by H/D exchange. Based on data from H/D exchange putative binding epitope was mapped to the PSMA surface. To describe interaction between PSMA and 5D3 antibody in more detail, we are undertaking structural studies of PSMA complexes with 5D3 fragments.