Complement system activation after allogeneic bone marrow transplantation may early predict the development of acute gastrointestinal GvHD

The Complement system consists of several serum proteins and receptors that belong to the innate immune response. It is activated by three possible pathways leading to the production of pro-inflammatory factors and the formation of the membrane attack complex. As it is known to be implicated in the physiopathology of several immune-mediated diseases, we investigated its role in GVHD in both humans and a mouse model. Thirty nine patients allografted for diverse haematological malignancies in our institution following myeloablative conditioning (n=22) or reduced intensity conditioning (RIC) (n=17) entered prospectively the study. Extensive exploration of the Complement system was carried out before and sequentially after allogeneic haematopoietic stem cell transplantation (HSCT). An activation of the classical pathway, as defined by a decrease of C3 and C4 proteins below normal values and of at least 50% of their pre-HSCT levels, was observed in 10/39 patients. Such activation was found in 9/22 (40%) patients who received myeloablative conditioning and in 1/17 (5.8%) patients allografted with RIC. This phenomenon occurred during the 4 first weeks after HSCT in 7 patients, at two months in one case and after withdrawal of immunosuppression in 2 patients. We observed a strong association between Complement activation and the development of acute gastrointestinal (GI) GVHD. Nine of the 10 patients who activated the Complement system (90%) developed GI GVHD within two weeks following the activation. By contrast only 4 of the 29 who did not activate the system, all of whom had received a RIC, developed GI GVHD (13.8%) (p<0.001). No correlation was found with the use of monoclonal antibodies, the degree of HLA and sex disparity between donor and recipient or the CMV status before the graft. We then analyzed Complement activation in a parent (C57BL/6, H–2b) to F1 [(C57BL/6×DBA2), H–bd] GVHD mouse model. Serum C3 Complement fraction and serum albumin protein (SAP) were measured sequentially after HSCT in lethally irradiated (9.75 Gy) mice reconstituted with either syngeneic or allogeneic bone marrow cells and splenocytes (107 and 13×106 cells/recipient, respectively). In comparison to naïve non HSCT recipient mice, the C3/SAP ratio was increased up to three times from Day 4 to 28 in syngeneic HSCT recipients (p=0.0002), reflecting the inflammatory response induced by the conditioning regimen without Complement activation in this conditioned non-GVHD control group. By contrast, the C3/SAP ratio was significantly decreased from Day 7 to 28 after HSCT in allogeneic recipients of HSCT in comparison to that of naïve (p=0.0005) and of syngeneic recipients (p=0.0002), indicating an activation of the Complement system in allografted mice that uniformly developed GI GVHD by Day 20 after HSCT. In conclusion, our results show that allogeneic HSCT induces Complement activation in both humans and mice, as a consequence of the immune alloresponse and not as a direct effect of the conditioning regimen. They also suggest that Complement C3 and C4 dosage might be useful in early prediction of acute gastrointestinal GVHD in humans. Ongoing studies are investigating the potential protective effect of pharmacological inhibitors of Complement activation on the development of GVHD in our mouse model.