Inhibition of properdin is more effective than other inhibitors of the complement system at preventing cell lysis in an in vitro model of atypical hemolytic uremic syndrome

Abstract

The alternative pathway (AP) of the complement system can be activated excessively in several inflammatory diseases, particularly when there is a defect in regulatory components of the complement system. For instance, defects of complement regulatory proteins such as factor H (FH) and lack of CD55 and CD59 are associated with atypical hemolytic uremic syndrome (aHUS) and paroxysmal nocturnal hemoglobinuria (PNH), respectively. Both diseases lead to severe prothrombotic pathologies in which the damage is mediated mainly by the AP. Thus, strategies to protect host cells by effectively inhibiting AP complement-mediated attack are necessary. Here we have developed and characterized monoclonal antibodies against properdin (clones 6E11A4, 3A3E1, 6E9E6, and 1G6D2), the only known positive regulator of complement, to study their effects in preventing lysis in an in-vitro model of aHUS. We determined that our antibodies recognize both human and rabbit properdin, and the non-inhibitory antibodies recognize distinct epitopes as determined in a binding competition assay. More importantly, the data indicate that the inhibitory anti-properdin monoclonal antibodies had lower IC50 values than all other complement inhibitors tested, including Soliris, which is currently used to treat patients with this disease. We have also standardized a highly sensitive sandwich ELISA, using monoclonal antibody pairs, with a lower properdin detection limit than commercially available kits (20 pg/ml). Further studies aimed at determining the therapeutic value of inhibiting properdin in human inflammatory diseases are warranted.