Rituximab maintenance therapy for patients with antineutrophil cytoplasmic antibody-associated vasculitis in Japan.

OBJECTIVES: We examined the efficacy and safety of rituximab (RTX) maintenance therapy for patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) in Japan. METHODS: We conducted a retrospective study using a multi-center cohort database of vasculitis patients. All maintenance treatment courses were divided into three groups: a RTX group, a group treated with other immunosuppressant drugs (IS) and a group receiving glucocorticoid monotherapy (GC). The primary endpoint was the comparison of relapse-free survival after 1 year. We also analyzed the occurrence of severe adverse events (SAEs) to assess safety. RESULTS: We included 123 courses of 107 patients (RTX n = 14, IS n = 64, GC n = 45). Twelve of 14 in the RTX group patients were diagnosed with granulomatosis with polyangiitis (GPA). The relapse-free survival of RTX maintenance therapy was comparable to that in the other groups (p = .122). After 1 year of treatment, the RTX group was administered lower steroid doses and one-third of them could withdraw corticosteroid. The overall incidence of SAE was 0.54/patient-year in the RTX group, 0.39/patient-year in the IS group and 0.34/patient-year in the GC group. CONCLUSION: RTX maintenance therapy could be effective and safe in Japanese GPA patients.

Shoulder ultrasound and serum lactate dehydrogenase predict inadequate response to glucocorticoid treatment in patients with polymyalgia rheumatica.

We aimed to identify predictors of inadequate response to glucocorticoid (GC) treatment in patients with polymyalgia rheumatica (PMR). We retrospectively studied 32 patients as a derivation cohort and 24 patients as a validation cohort. The patients were divided into two groups according to the response to GC treatment: GC-responders and GC-inadequate responders (GC-IRs). We compared laboratory data and bilateral shoulder ultrasound findings between the groups. Receiver operating characteristic (ROC) analysis was performed to determine the optimal cutoff value of candidate predictors of treatment response; the predictors were examined using multivariate logistic analysis. Gray-scale ultrasound findings of long head of the biceps (LHB) tenosynovitis and subacromial/subdeltoid (SAD) bursitis were scored semiquantitatively (0-3). A total gray-scale score (TGSS) was calculated as the sum of the gray-scale scores. In the derivation cohort, serum lactate dehydrogenase (LDH) levels and TGSS were significantly higher in GC-IRs than in GC-responders. On ROC analysis, the cutoff values of serum LDH levels ≥ 175 IU/ml and TGSS ≥ 5 were found to be the candidate predictors. Multivariate logistic analysis revealed an independent association of both the predictors with inadequate response to GC treatment. In the validation cohort, patients with one or both predictors exhibited a higher incidence of inadequate response to GC treatment. These findings indicate that the severities of LHB tenosynovitis and SAD bursitis evaluated using ultrasound and serum LDH levels are independent predictors of inadequate response to GC treatment in patients with PMR. Treatment adjustment based on prediction model may allow precise treatment of patients with PMR.

Activation of central/effector memory T cells and T-helper 1 polarization in malignant melanoma patients treated with anti-programmed death-1 antibody.

Human anti-programmed death-1 (PD-1) antibody possesses the capability to revitalize host T cells and has been an effective therapy for metastatic malignant melanoma (MM). The precise subsets of T cells predominantly activated by anti-PD-1, however, have not yet been clarified. In this study, peripheral blood mononuclear cells obtained from MM patients scheduled to receive anti-PD-1 (nivolumab) therapy, and healthy subjects (HS), were systematically examined on flow cytometry to identify changes in the proportion of immune cell subsets. Compared with HS, MM patients prior to therapy had an increased proportion of activated CD8+ T cells with effector memory phenotypes (Tem), and PD-1 positive subsets of CD4+ central memory T cells (Tcm) and T-helper (Th)17 cells. After a single course of anti-PD-1 therapy, MM patients had an increase in activated Tem and Tcm subsets of CD4+ and CD8+ T cells, and activated Th1 plus T-helper follicular 1 cells. There was no consistent change in the proportion of Tfh cells, B cells, natural killer cells, or dendritic cells. The observed activated phenotypes were attenuated during the course of therapy, but regulatory T cells belonging to the CD3+CD4+CD45RO+CD25high fraction increased at disease progression. Taken together, anti-PD-1 therapy modulates systemic immune reactions and exerts anti-tumor effects, not only by revitalizing Tem and Tcm of CD4+ and CD8+ T cells, but also via a shift to a Th1 phenotype.

Functional interleukin-33 receptors are expressed in early progenitor stages of allergy-related granulocytes.

Interleukin-33 (IL-33) induces T helper type 2 (Th2) cytokine production and eosinophilia independently of acquired immunity, leading to innate immunity-mediated allergic inflammation. Allergy-related innate myeloid cells such as eosinophils, basophils and mast cells express the IL-33 receptor (IL-33R), but it is still unknown how IL-33 regulates allergic inflammation involving these cells and their progenitors. Here, we revealed that the functional IL-33R was expressed on eosinophil progenitors (EoPs), basophil progenitors (BaPs) and mast cell progenitors (MCPs). In the presence of IL-33, these progenitors did not expand, but produced a high amount of Th2 and pro-inflammatory cytokines such as IL-9, IL-13, IL-1ß and IL-6. The amount of cytokines produced by these progenitors was greater than that by mature cells. In vivo, IL-33 stimulated the expansion of EoPs, but it was dependent upon the elevated serum IL-5 that is presumably derived from type 2 innate lymphoid cells that express functional IL-33R. These data collectively suggest that EoPs, BaPs and MCPs are not only the sources of allergy-related granulocytes, but can also be sources of allergy-related cytokines in IL-33-induced inflammation. Because such progenitors can differentiate into mature granulocytes at the site of inflammation, they are potential therapeutic targets in IL-33-related allergic diseases.

CIN85 is required for Cbl-mediated regulation of antigen receptor signaling in human B cells.

The aberrant regulation of B-cell receptor (BCR) signaling allows unwanted B cells to persist, thereby potentially leading to autoimmunity and B-cell malignancies. Casitas B-lineage lymphoma (Cbl) proteins suppress BCR signaling; however, the molecular mechanisms that control Cbl function in human B cells remain unclear. Here, we demonstrate that CIN85 (c-Cbl interacting protein of 85 kDa) is constitutively associated with c-Cbl, Cbl-b, and B-cell linker in B cells. Experiments using CIN85-overexpressing and CIN85-knockdown B-cell lines revealed that CIN85 increased c-Cbl phosphorylation and inhibited BCR-induced calcium flux and phosphorylation of Syk and PLCγ2, whereas it did not affect BCR internalization. The Syk phosphorylation in CIN85-overexpressing and CIN85-knockdown cells was inversely correlated with the ubiquitination and degradation of Syk. Moreover, CIN85 knockdown in primary B cells enhanced BCR-induced survival and growth, and increased the expression of BcLxL, A1, cyclin D2, and myc. Following the stimulation of BCR and Toll-like receptor 9, B-cell differentiation- associated molecules were up-regulated in CIN85-knockdown cells. Together, these results suggest that CIN85 is required for Cbl-mediated regulation of BCR signaling and for downstream events such as survival, growth, and differentiation of human B cells.

Clinical course of patients with rheumatoid arthritis who continue or discontinue biologic therapy after hospitalization for infection: a retrospective observational study.

BACKGROUND: To analyse the subsequent clinical course of patients with rheumatoid arthritis (RA) who either continued or discontinued biologic agents after hospitalization for infections. METHODS: We retrospectively reviewed the clinical records of 230 RA patients with 307 hospitalizations for infections under biologic therapy between September 2008 and May 2014 in 15 institutions for up to 18 months after discharge. The risks of RA flares and subsequent hospitalizations for infections from 61 days to 18 months after discharge were evaluated. RESULTS: Survival analyses indicated that patients who continued biologic therapy had a significantly lower risk of RA flares (31.4% vs. 60.6%, P < 0.01) and a slightly lower risk of subsequent infections (28.7% vs. 34.5%, P = 0.37). Multivariate analysis showed that discontinuation of biologic therapy, diabetes, and a history of hospitalization for infection under biologic therapy were associated with RA flares. Oral steroid therapy equivalent to prednisolone 5 mg/day or more and chronic renal dysfunction were independent risk factors for subsequent hospitalizations for infections. CONCLUSIONS: Discontinuation of biologic therapy after hospitalization for infections may result in RA flares. Continuation of biologic therapy is preferable, particularly in patients without immunodeficiency.

The Association of Airway Comorbidities With the Clinical Phenotypes and Outcomes of Patients With Antineutrophil Cytoplasmic Autoantibody-associated Vasculitis.

OBJECTIVE: We investigated the association of airway comorbidities with the clinical phenotypes and outcomes of myeloperoxidase (MPO)-antineutrophil cytoplasmic antibodies (ANCA)-positive ANCA-associated vasculitis (AAV). METHODS: An AAV patient multicenter cohort trial was established in 13 hospitals in western Japan between 2012 and 2018. We examined 143 of the new-onset MPO-ANCA-positive AAV patients. Their clinical characteristics and comorbidities at disease onset were compared based on clinical phenotypes. Multivariate analysis was performed to identify factors predictive of remission and death. RESULTS: Twenty-seven cases with granulomatosis with polyangiitis (GPA), 10 with eosinophilic GPA (EGPA), 81 with microscopic polyangiitis (MPA), and 25 with unclassified AAV were identified. The average age of MPO-ANCA-positive patients was 71.4 years. Comorbidity (87.4%) and airway comorbidity (70.6%) were frequently observed in these patients. Examination of the clinical phenotypes revealed that the cases of GPA were frequently accompanied by infectious airway comorbidity (upper airway disease, bronchiectasis, pulmonary infections), and most of the cases of MPA and unclassified AAV were accompanied by fibrotic interstitial lung disease (fILD) or emphysema. Among MPO-ANCA-positive patients, infectious airway comorbidity was predictive of both remission (HR 1.58, P = 0.03) and mortality (HR 2.64, P = 0.04), and fILD was predictive of mortality (HR 7.55, P = 0.008). The combination of infectious airway comorbidities and fILD caused the worst survival outcomes in patients. CONCLUSION: MPO-ANCA-positive AAV was frequently accompanied by airway comorbidities. In addition to fILD, infectious airway comorbidities were closely associated with those clinical phenotypes and outcomes.

IgG anti-hinge antibodies against IgG4 F(ab')2 fragments generated using pepsin are useful diagnostic markers for rheumatoid arthritis: implications of the possible roles of metalloproteinases and IgG subclasses in generating immunogenic hinge epitopes.

BACKGROUND: Pepsin agglutinators, discovered over 50 years ago, have been recently referred to as anti-hinge antibodies (AHAs) because of their reaction with the IgG hinge epitope. AHAs have different reactivity for each hinge epitope generated by each protease that cleaves the hinge region at different sites. Moreover, AHAs have different reactivity against different hinge epitopes derived from each IgG subclass even when the same protease is used. Since the expression of matrix metalloproteinase-3 (MMP-3) is enhanced in rheumatoid arthritis (RA), AHA production could also be increased. The purpose of this study was to determine whether the levels of AHAs against IgG hinge epitopes produced by MMP-3 are elevated in RA. METHODS: The serum levels of IgG or IgA AHAs against the IgG1/IgG4 F(ab')2 fragments, generated by either MMP-3 or pepsin, were measured using ELISA in 111 patients with RA and 81 healthy controls (HC). Receiver operating characteristic (ROC) analysis was used for obtaining optimal cutoff values and cutoff values indicating high specificity (> 95%) of the AHA. The targeted epitope of a specific AHA was investigated through inhibition ELISA. RESULTS: Seven AHAs were statistically higher in RA patients than in HC, except IgG AHA against IgG1 F(ab')2, which was generated by MMP-3 proteolytic cleavage. The areas under the ROC curve were 0.66-0.80, although the sensitivities at high specificity were low (5.4-24.3%). The cumulative number of positive AHAs in each individual was statistically higher in RA patients than in HC, suggesting the extreme extent of AHA repertoires in RA. Inhibition studies revealed that IgG AHAs against IgG4 F(ab')2 fragments generated by pepsin cross-reacted with IgG1 F(ab')2 fragments generated by pepsin. Multivariate logistic regression analysis identified the IgG AHA against IgG4 F(ab')2 fragments generated by pepsin as an independent variable for RA diagnosis, even in RA patients who were negative for both RF and ACPA (odds ratio 1.18, 95% CI 1.06-1.32; P = 0.003). Additional experiments using non-RA patients finally strengthened the diagnostic utility. CONCLUSION: In RA patients, we observed diversification and amplification of AHA repertoires and diagnostic utility of the specific AHA against IgG4 F(ab')2 fragments generated by pepsin but not MMP-3.

Generation mechanism of RANKL(+) effector memory B cells: relevance to the pathogenesis of rheumatoid arthritis.

BACKGROUND: The efficacy of B cell-depleting therapies for rheumatoid arthritis underscores antibody-independent functions of effector B cells such as cognate T-B interactions and production of pro-inflammatory cytokines. Receptor activator of nuclear factor κB ligand (RANKL) is a key cytokine involved in bone destruction and is highly expressed in synovial fluid B cells in patients with rheumatoid arthritis. In this study we sought to clarify the generation mechanism of RANKL(+) effector B cells and their impacts on osteoclast differentiation. METHODS: Peripheral blood and synovial fluid B cells from healthy controls and patients with rheumatoid arthritis were isolated using cell sorter. mRNA expression of RANKL, osteoprotegerin, tumor necrosis factor (TNF)-α, and Blimp-1 was analyzed by quantitative real-time polymerase chain reaction. Levels of RANKL, CD80, CD86, and CXCR3 were analyzed using flow cytometry. Functional analysis of osteoclastogenesis was carried out in the co-culture system using macrophage RAW264 reporter cells. RESULTS: RANKL expression was accentuated in CD80(+)CD86(+) B cells, a highly activated B-cell subset more abundantly observed in patients with rheumatoid arthritis. Upon activation via B-cell receptor and CD40, switched-memory B cells predominantly expressed RANKL, which was further augmented by interferon-γ (IFN-γ) but suppressed by interleukin-21. Strikingly, IFN-γ also enhanced TNF-α expression, while it strongly suppressed osteoprotegerin expression in B cells. IFN-γ increased the generation of CXCR3(+)RANKL(+) effector B cells, mimicking the synovial B cell phenotype in patients with rheumatoid arthritis. Finally, RANKL(+) effector B cells in concert with TNF-α facilitated osteoclast differentiation in vitro. CONCLUSIONS: Our current findings have shed light on the generation mechanism of pathogenic RANKL(+) effector B cells that would be an ideal therapeutic target for rheumatoid arthritis in the future.

Bi-cytopenia possibly induced by anti-PD-1 antibody for primary malignant melanoma of the esophagus: A case report.

BACKGROUND: Anti-programmed cell death 1 antibody nivolumab is a promising agent for various cancers. Immune-related adverse events are recognized; however, bi-cytopenia with nivolumab has not been reported. CASE PRESENTATION: A 73-year-old man was diagnosed with advanced primary malignant melanoma of the esophagus with liver, lung, and lymph node metastases. Previous therapies including dacarbazine and radiation of 39 Gy to the esophageal region were performed, but the liver metastases deteriorated. The patient was then administered nivolumab (2 mg/kg, every 3 weeks). After 3 cycles, the esophageal tumor and lymph nodes showed marked reductions in size, the lung metastases disappeared, and the liver metastases shrank partially. The treatment continued with 7 cycles for 4 months. However, severe anemia and thrombocytopenia appeared in the 6th cycle, and intermittent blood transfusions were required. The patient received high-dose intravenous methylprednisolone therapy for bi-cytopenia, but it was ineffective. Seven months after the initiation of nivolumab, the patient died of tumor. Although the mechanisms of bi-cytopenia were unclear, it could have been induced by nivolumab. CONCLUSION: The present case shows a rare but serious life-threatening bi-cytopenia possibly associated with nivolumab and suggests the importance of awareness of hematological adverse events during nivolumab therapy.

Clinicopathological significance of cervical adenocarcinoma associated with lobular endocervical glandular hyperplasia.

Lobular endocervical glandular hyperplasia (LEGH) is usually assumed to be a benign tumor-like lesion of the glands of the uterine cervix. However, LEGH has been associated with obvious cervical adenocarcinoma. The clinicopathological significance of coexistence of LEGH with adenocarcinoma remains unclear. We microscopically examined the presence or absence of LEGH components in 95 stage Ib cervical adenocarcinomas. Gastric mucin was detected with the use of clone HIK1083. Associations of the coexistence of LEGH components with clinicopathological variables were analyzed. LEGH components were present in 16 cases (16.8%). Gastric mucin was positive in all 16 LEGH components, as compared with only 6 of the 95 adenocarcinoma components. Of the 16 adenocarcinomas with LEGH components, 15 were well-differentiated mucinous adenocarcinomas, and one was poorly differentiated adenocarcinoma. The mortality rate of tumor recurrence was 25% (4 of 16) in patients whose tumors had LEGH components, and 21.5% (17 of 79) in those whose tumors had no LEGH components. There was no significant difference in survival. Early cervical adenocarcinoma was relatively frequently associated with LEGH components. LEGH may be one of the factors related to the development of cervical adenocarcinoma, but adenocarcinoma with LEGH components does not necessarily develop into a highly aggressive "adenoma malignum."