Introduction
Anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis (AAV) is a systemic autoimmune disease with poor prognosis that damages various organs and commonly occurs in the elderly [
1]. To induce remission, AAV is treated with high-dose glucocorticoid (GC) therapy combined with cyclophosphamide (CY), azathioprine (AZA), etc. [
2]. However, because these immunosuppressants are nonspecific and cause adverse reactions, development of therapeutic agents specific to the pathology of this disease is warranted. The efficacy of B cell depletion therapy has been demonstrated in AAV patients by the Rituximab in ANCA-Associated Vasculitis (RAVE) trial [
3] and the Randomized Trial of Rituximab Versus Cyclophosphamide for ANCA-Associated Renal Vasculitis (RITUXVAS) [
4] trial. Rituximab (RTX) has been shown to be as effective as CY for inducing remission in AAV patients. However, it is unknown how B cells actually affect disease activity and prognosis in AAV patients. Furthermore, although RTX, which is the first biologic drug approved for the treatment of AAV, has a specific action mechanism of depleting B cells, the differential use of RTX and CY remains a major clinical question.
The surface phenotypes of peripheral lymphocytes in patients with autoimmune disease are often considered to reflect affected tissue states. We have analyzed peripheral lymphocytes from patients with systemic lupus erythematosus (SLE) [
5], rheumatoid arthritis [
6], and psoriatic arthritis [
7], using 8-color flow cytometry. In these studies, we showed that analysis of peripheral lymphocytes provides valuable insight into the lineage, differentiation stage, activation state, etc., of lymphocytes associated with pathological conditions and can be an indicator for treatment selection.
Thus, the present study aimed to identify abnormalities in the differentiation of peripheral lymphocytes in AAV patients, analyze the clinical significance of these abnormalities, and investigate how to differentiate the use of RTX and CY according to abnormalities in lymphocytes.
Discussion
In the present study, which included patients with highly active AAV who were admitted to our hospital, we analyzed phenotypes of T and B cells before remission induction therapy to assess the clinical significance of these phenotypes and to differentiate the use of RTX and CY according to abnormal cell differentiation. Based on phenotyping of peripheral lymphocytes in this study, AAV patients exhibited a lower proportion of IgM+ unswitched memory B cells and a higher proportion of IgD−CD27− B cells. Thus, B cells were more differentiated in many patients with highly active AAV compared to those in HCs.
Regarding the association between peripheral B cells and clinical findings, it has been reported that peripheral CD5
+ B cells are associated with relapse after remission induction therapy with RTX [
10,
11]. However, because there has been no study comparing abnormal differentiation and clinical findings, it has remained unknown which lymphocyte phenotype is associated with prognosis. The present study revealed that B cell phenotypes were not associated with disease activity or clinical findings but were correlated with responses to treatment. As the number of highly differentiated B cells increased, patients were more resistant to treatment. However, when patients with increased class-switched memory B cells or IgD
−CD27
− B cells, defined as those with excessive B cell differentiation, were compared, no differences were observed in either baseline patient characteristics or T cell phenotypes. The presence of abnormal B cell phenotypes could not be predicted from ANCA levels, other baseline patient characteristics, or disease activity. In contrast, many patients with excessive B cell differentiation were resistant to treatment so that detection of excessive B cell differentiation may be important for predicting prognosis.
The above indicates that changes in prognosis mainly based on excessive B cell differentiation may be useful for differentiating between the use of RTX, which is used for B cell depletion therapy, and CY. The present study revealed that, in patients with excessive B cell differentiation, the concomitant use of RTX was more effective than that of conventional therapy and yielded improvements similar to that achieved by patients without excessive B cell differentiation. In addition to showing that RTX is effective in patients who are resistant to CY or experience relapse after treatment with CY [
12‐
15], various clinical studies have demonstrated that RTX is as effective as CY against AAV accompanied by serious organ involvement [
3,
16]. However, it is still unknown which patients should be treated with RTX and which with CY. The present study suggests that RTX is better indicated for patients with abnormal B cells. Moreover, this study also showed that the incidence of adverse events was lower in patients with excessive B cell differentiation who received a combination of RTX and GC than in those treated with concomitant conventional therapy, as GC doses could be reduced in the early stages. This may improve the survival rate in these patients.
Although the characteristics of B cells in AAV patients have been described in many reports, the importance of B cells remains controversial. Culton et al. [
17] and Tadema et al. [
18] reported that, compared with healthy individuals, AAV patients have more naive B cells (CD19
+CD27
−CD38
−/low) and fewer memory B cells (CD19
+CD27
+CD38
−/low). In contrast, Culton et al. [
17] reported that, compared with healthy individuals, AAV patients have more CD19
hi memory B cells, which have an elevated capacity to produce autoantibodies. Moreover, Lepse et al. [
19] reported that the proportion of regulatory B cells decreases in patients with highly active AAV. In the present study, phenotyping of peripheral lymphocytes revealed that AAV patients exhibited a low proportion of IgM+ unswitched memory B cells and a high proportion of IgD
−CD27
− B cells. Thus, in many patients with highly active AAV, B cells were more differentiated than in HCs, similar to a trend described in previous reports [
17,
18]. To date, several reports have been published concerning the relationship between peripheral blood B cells and responses of autoimmune disease to treatment. Lanzillotta et al. reported that in patients with IgG4-related disease, an increase in peripheral blood memory B cell count (CD19
+CD20
+CD27
+CD38
− cells) 6 months after the beginning of glucocorticoid treatment is associated with relapse [
20]. Yusof et al. reported that among RTX-treated patients with AAV, the length of time until relapse was longer in cases showing re-proliferation of peripheral blood naïve B cells (CD19
+CD27
− cells) rather than peripheral blood memory B cells (CD19
+CD27
+ B cells) 6 months after the beginning of RTX therapy for removal of peripheral blood B cells [
21]. These previous findings suggest the possibility that memory B cells, including class-switched memory B cells, are associated with relapse and resistance to treatment. These findings do not contradict the results of the present study.
The present study has several limitations. The RTX and IV-CY groups were not matched for baseline patient characteristics, and they were not compared in a double-blind manner. Thus, it cannot be ruled out that the selection of treatment was biased. However, no significant differences were observed between the RTX and IV-CY groups in terms of baseline patient characteristics, such as age, sex, disease duration, and disease activity, and the groups were compared for efficacy under the same conditions. The data from this study are preliminary in nature, because the number of cases with excessive B cell differentiation was small. Further studies on large number of patients are needed to confirm the utility of the reported findings.
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