The use of rituximab (RTX), an anti-CD20 monoclonal antibody (Ab), in refractory myasthenia gravis (MG) is associated with a better response in patients with Abs to the muscle-specific tyrosine kinase (MuSK) than in other MG subgroups. Anti-MuSK Abs are mostly IgG4 with proven pathogenicity and positive correlation with clinical severity. The rapid and sustained response to RTX may be related to MuSK Ab production by short-lived Ab-secreting cells derived from specific CD20 + B cells. Here, we investigated the long-term effects of RTX in nine refractory MuSK-MG patients with a follow-up ranging from 17 months to 13 years. In patients' sera, we titrated MuSK-specific IgG (MuSK-IgG) and MuSK-IgG4, along with total IgG and IgG4 levels. Optimal response to RTX was defined as the achievement and maintenance of the status of minimal manifestations (MM)-or-better together with a ≥ 50% steroid reduction, withdrawal of immunosuppressants, and no need for plasma-exchange or intravenous immunoglobulin. After a course of RTX, eight patients improved, with optimal response in six, while only one patient did not respond. At baseline, MuSK-IgG and MuSK-IgG4 serum titers were positive in all patients, ranging from 2.15 to 49.5 nmol/L and from 0.33 to 46.2 nmol/L, respectively. MuSK Abs mostly consisted of IgG4 (range 63.80-98.86%). RTX administration was followed by a marked reduction of MuSK Abs at 2-7 months and at 12-30 months (p < 0.02 for MuSK-IgG and p < 0.01 for MuSK-IgG4). In patients with a longer follow-up, MuSK Ab titers remained suppressed, paralleling clinical response. In the patient who achieved long-term complete remission, MuSK-IgG4 was no longer detectable within 2 years, while MuSK-IgG remained positive at very low titers up to 10 years after RTX. In the patient who did not respond, MuSK-IgG and MuSK-IgG4 remained unchanged. In this patient series, total IgG and IgG4 transiently decreased (p < 0.05) at 2-7 months after RTX. The different trends of

Hepatitis C virus (HCV) causes mixed cryoglobulinemia (MC) by driving clonal expansion of IgMCD27 B cells. These cells display both the features of anergy induced by continual engagement of the B-cell receptor (BCR), such as high expression of phosphorylated extracellular signal-regulated kinase (pERK) and reduced lifespan, and of virus-specific exhaustion, such as CD21 phenotype and a defective response to ligation of BCR and Toll-like receptor 9 (TLR9). MC usually regresses after eradication of HCV with interferon, whose immunomodulatory activity might contribute to this effect. We investigated the phenotypic and functional changes in clonal B cells of MC patients with sustained virologic responses to direct-acting antivirals (DAAs), which lack immunomodulatory properties. We found that high pERK expression and accelerated apoptosis revert within 4 weeks after beginning therapy, whereas clonal B cells unresponsive to TLR9 stimulation persist for at least 24 weeks, although they may partially rescue normal CD21 expression. Thus, similar to mouse models, features of anergy in MC B cells rapidly revert after disengagement from HCV, whereas virus-specific exhaustion imparts a durable inhibitory imprint on cell function. Treatment of HCV MC with DAAs provides a valuable tool for untangling the molecular mechanisms of anergy and exhaustion in human B cells.

Abstract. Cancer cells need to become motile in order to escape the primary tumor and move to distant areas to form metastasis. They move as single cells or as a group, following different stimuli, including growth factors. Among them, insulin-like growth factor-1 (IGF-1) and epidermal growth factor (EGF) and their receptors have been implicated in the development and progression of human breast carcinoma. In this report, we provide evidence that the tyrosine kinase Src is responsible for migration promoted by both IGF-1 and EGF in MDA-MB-231 and MCF7 cells, although with a different effect. Moreover, both IGF-1 and EGF induce reorganization of actin cytoskeleton in lamellipodia and membrane ruffles in a time-and Src-dependent manner. Furthermore, we analyzed the tyrosine phosphorylation status of the actin-binding protein cortactin upon growth factor stimulation, showing that even the activation of cortactin is time-and Src-dependent. In addition, immunofluorescence analysis with anti-paxillin antibody reveals that, after treatment with growth factors, tyrosine phosphorylated cortactin is localized on the plasma membrane in correspondence of focal adhesions. Collectively, our findings suggest a crucial role for Src-mediated activation of cortactin in cell migration, reorganization of actin cytoskeleton and phosphotyrosine cortactin localization to the focal adhesions in human breast cancer cell lines upon both IGF-1 and EGF stimulation.