Study of the cancer antigen EMMPRIN and isolation of therapeutic human antibodies
Hellenic Pasteur Institute
Extracellular matrix degradation is an important mechanism for tumor cell invasion and metastasis and is mainly accomplished by specific enzymes called metalloproteinases. It has been shown that protein EMMPRIN regulates the expression of metalloproteinases and is associated with poor prognosis. The extracellular domain of EMMPRIN, (ECD), contains 2 structural domains (Ig1 and Ig2) with a high degree of glycosylation.
The main interest of our study was focused on:
a) Elucidation of the exact region of the extracellular domain of EMMPRIN that is responsible for the induction of metalloproteinases and clarify whether glycosylation is required for this process, and
b) to use this domain in order to isolate human therapeutic antibodies that will be able to inhibit EMMPRIN activity.
To achieve our first goal, we initially cloned and expressed the extracellular domains coding for ECD, Ig1 and Ig2, in two heterologous expression systems, so that the recombinant proteins will be produced in their glycosylated and nonglycosylated forms. Then, we tested the ability of these proteins to stimulate the expression of metalloproteinases from dermal fibroblasts by using gelatin zymography. Our data showed that the glycosylated ECD and Ig2 domains were able to induce metalloproteinase levels significantly higher than their non glycosylated counterparts, whereas for the Ig1 domain there was no considerable change between the two forms. These results have lead to the conclusion that the glycosylated Ig2 extracellular region of EMMPRIN is critically implicated in the activation of metalloproteinases, a fact that has not been mentioned to any bibliographical reports until today.
As for the second goal, the isolation of human therapeutical antibodies, we used an antibody library previously constructed in our lab from the B lymphocytes of a breast cancer patient. We have found that the isolated human antibody in the presence of the ECD of EMMPRIN showed high specificity for the antigen, but it was incapable of inhibiting metalloproteinase levels from dermal fibroblasts after incubation with the ECD of EMMPRIN. On the contrary, this antibody can provoke a slight induction of metalloproteinases after 24 hours of incubation. The role of such antibodies to metastatic cancer remains to be elucidated.
Final report (in Greek)
Avgi Mamalaki, Senior Researcher, Hellenic Pasteur Institute
Adriana Papadimitropoulou, Postgraduate Collaborator, Hellenic Pasteur Institute
Petros Heliades, Research Associate, Hellenic Pasteur Institute
Dimitra Makatsori, Research Assistant, Hellenic Pasteur Institute
Maria Samoili, Biologist