Improvement in treatment of experimental colitis in mice by using recombinant Lactococcus lactis with surface-displayed affibody against TNFα (THER4P.889)

TNFα is one of the most studied pro-inflammatory cytokines in the pathology of irritative bovel disease (IBD), including Crohn's disease and ulcerative colitis. The current therapy for the treatment of mid-to severe IBD is composed by corticosteroids and parenteral administration of monoclonal antibodies against TNFα with systemic side effects. In order to develop an oral administration of anti-TNFα agent, we have introduced an affibody against TNFα as a binding molecule. A recombinant Lactococcus lactis was developed by means of expression of anti-TNFα affibody as a fusion protein with three functional parts: signal Usp45 protein for the secretion to the microenvironment in gut, TNFα-binding affibody domain and peptidoglycan-bynding AcmA autolysin protein that enables attachment of peptidic construct to the surface of L. lactis. Such recombinant bacteria were characterized by the capability to bind Alexa Fluor 488-conjugated TNFα. By means of flow cytometry we clearly demonstrated a distinct shift in fluorescence between control cells and TNFα-binding cells. Furthermore, mice model with induced experimental colitis was used for animal studies. After oral administration of wild type L. lactis (control group) and recombinant anti-TNF α L. lactis, we observed significantly decrease in the concentration of TNFα in the stool as well as general mucosal improvement in group treated with recombinant anti-TNF lactic acid bacteria.