Hepatitis C virus (HCV)-associated mixed cryoglobulinemia (MC) is a benign lymphoproliferative disorder characterized by the expansion of a B cell clone producing IgM endowed with rheumatoid factor (RF) activity that forms cold-precipitable immune complexes (cryoglobulins) causing small vassel vasculits. The treatment for HCV-associated MC is the eradication of the virus that is now achieved in almost 100% of patients with the new direct acting antivirals (DAAs). Recent data have shown that DAA induce a complete clinical response of MC in 71-90% of patients after HCV eradication and, thus, 10-29% of HCV patients fail to recover from cryoglobulinemic vasculitis or relapse after initial response. Persistence of cryoglobulinemia after the clearance of HCV appears to be due to the persistence of pathogenic B-cell clones. Therefore additional treatment with the B cell depleting monoclonal antibody rituximab (RTX) might increase vasculitis¿response.
The purpose of this project is to study the efficacy of combined treatment of RTX with DAA for the treatment of HCV-associated MC and to analyze whether, after removal of viral stimulation together with B cell depletion, recovery of B cell homeostasis is achieved. To this aim circulating B cell clones will be investigated by flow cytometry and at a molecular level before therapy and throughout the follow-up.
In addition B cell clones will be investigated at the functional level (at baseline or in patients treated with only DAA) in order to individualize alternative stimuli that, after HCV clearance, might be responsible for their persistence. To this aim in vitro studies will investigate whether immune complexes induce a proliferative response in RF-positive B cell clones.
These studies might provide new insights in the pathogenic mechanisms involved in the persistence or relapse of MC after HCV eradication that might also be responsible for lymphoma evolution, a frequent complication of MC patients.
Symptomatic MC vasculitis bears the highest health care costs among the extrahepatic manifestations of HCV infection, and affects 4.9% of patients with chronic HCV infection (Younossi Z, Gastroenterology. 2016). In addition, MC is the leading cause of chronic and end-stage renal disease among HCV-infected patients without decompensated cirrhosis. DAA-based antiviral therapy yields 94-100% SVR in patients with HCV-associated mixed cryoglobulinemia, but complete clinical response of vasculitis in only 71-90% of them (Gragnani L. et al. Hepatology 2016). Furthermore, achieving complete response of vasculitis requires several months after antiviral therapy. RTX yields rapid although transient clinical responses in these patients (Visentini M. et al. Autoimmun Rev 2011). Recent international guidelines on treatment of HCV-related MC vasculitis (Zignego AL et al. Autoimmun Rev. 2017) state that, although data about combined RTX plus DAA-based therapy are still scarce, this is a promising approach that warrants confirmatory studies. The results of our study will provide new data on the efficacy of combined therapy with DAA and RTX for the treatment of MC. At our referral centre for MC more than 300 patients with MC are followed and in the last 3 years we have treated more that 100 patients with DAA. These patients provide an historical group suitable for comparing the results of this study applying combined therapy for MC.
It has been postulated that persistence or relapse of MC vasculits after HCV eradication might be due to the persistence of the B cell clone, expanded in some patients with MC. The use of RTX, as B cell depleting monoclonal antibody, together with the removal, with the DAA, of the chronic antigenic stimulation provided by HCV, might in principal restore B cell homeostasis and interrupt further cryoglobulins production. This would result in a stable and complete clinical response of MC vasculits. Here we offer the possibility to provide evidence of this pathogenic model. In addition we will analyze if the persistence of the B cell clone, observed in MC after HCV eradication, might be due to circulating immune complexes able to bind rheumatoid factor bearing BCR in clonal B cells in MC, providing evidence that HCV is not the only stimulus able to induce B cell expansion. These data, in a more wide sense, might also shed light in the pathogenesis of other low-grade HCV-negative lymphoproliferative disorders, suggesting that other triggers, as immune complexes might provide chronic stimulation.