Risk of infection from glucocorticoid and methotrexate interaction in patients with rheumatoid arthritis using biologics: A retrospective cohort study.

AIMS: This retrospective cohort study aimed to evaluate the effect of the interaction between methotrexate and glucocorticoids on the risk of developing bacterial infections in patients with rheumatoid arthritis (RA) using biological disease-modifying antirheumatic drugs (bDMARDs). METHODS: We used the 2005-2018 JMDC claims database, a nationwide claims database in Japan. From the database of 7 175 048 patients, study patients were obtained by applying the following exclusion criteria: no use of bDMARDs; without information on the date of prescription; without RA as a disease; other than the new users of bDMARDs; and age <18 years. The exposures were glucocorticoids and methotrexate, and the outcome was bacterial infection. The interaction effects were examined using multivariate Cox regression analysis. Bacterial infections were identified according to antibiotic prescription and International Statistical Classification of Diseases and Related Health Problems, 10th revision codes. RESULTS: A total of 2837 RA patients were identified, with a median age of 50 years. The incidence of infection was 16.8% (95% confidence interval: 15.5-18.3). The interaction term for the doses of glucocorticoids and methotrexate was significant. Additionally, a higher dose of glucocorticoid was a significant risk factor for developing bacterial infections on the side of high doses of methotrexate. The incidence of bacterial infections tended to increase significantly with increasing methotrexate doses coprescribed with glucocorticoids ≥5 mg or glucocorticoid doses coprescribed with methotrexate ≥8 mg. CONCLUSION: Our results indicate a potential association between methotrexate dose and bacterial infections during bDMARDs administration with glucocorticoids in patients with RA.

A Novel Rac1-GSPT1 Signaling Pathway Controls Astrogliosis Following Central Nervous System Injury.

Astrogliosis (i.e. glial scar), which is comprised primarily of proliferated astrocytes at the lesion site and migrated astrocytes from neighboring regions, is one of the key reactions in determining outcomes after CNS injury. In an effort to identify potential molecules/pathways that regulate astrogliosis, we sought to determine whether Rac/Rac-mediated signaling in astrocytes represents a novel candidate for therapeutic intervention following CNS injury. For these studies, we generated mice with Rac1 deletion under the control of the GFAP (glial fibrillary acidic protein) promoter (GFAP-Cre;Rac1flox/flox). GFAP-Cre;Rac1flox/flox (Rac1-KO) mice exhibited better recovery after spinal cord injury and exhibited reduced astrogliosis at the lesion site relative to control. Reduced astrogliosis was also observed in Rac1-KO mice following microbeam irradiation-induced injury. Moreover, knockdown (KD) or KO of Rac1 in astrocytes (LN229 cells, primary astrocytes, or primary astrocytes from Rac1-KO mice) led to delayed cell cycle progression and reduced cell migration. Rac1-KD or Rac1-KO astrocytes additionally had decreased levels of GSPT1 (G1 to S phase transition 1) expression and reduced responses of IL-1ß and GSPT1 to LPS treatment, indicating that IL-1ß and GSPT1 are downstream molecules of Rac1 associated with inflammatory condition. Furthermore, GSPT1-KD astrocytes had cell cycle delay, with no effect on cell migration. The cell cycle delay induced by Rac1-KD was rescued by overexpression of GSPT1. Based on these results, we propose that Rac1-GSPT1 represents a novel signaling axis in astrocytes that accelerates proliferation in response to inflammation, which is one important factor in the development of astrogliosis/glial scar following CNS injury.

The extracellular A-loop of dual oxidases affects the specificity of reactive oxygen species release.

NADPH oxidase (Nox) family proteins produce superoxide (O2 (⨪)) directly by transferring an electron to molecular oxygen. Dual oxidases (Duoxes) also produce an O2 (⨪) intermediate, although the final species secreted by mature Duoxes is H2O2, suggesting that intramolecular O2 (⨪) dismutation or other mechanisms contribute to H2O2 release. We explored the structural determinants affecting reactive oxygen species formation by Duox enzymes. Duox2 showed O2 (⨪) leakage when mismatched with Duox activator 1 (DuoxA1). Duox2 released O2 (⨪) even in correctly matched combinations, including Duox2 + DuoxA2 and Duox2 + N-terminally tagged DuoxA2 regardless of the type or number of tags. Conversely, Duox1 did not release O2 (⨪) in any combination. Chimeric Duox2 possessing the A-loop of Duox1 showed no O2 (⨪) leakage; chimeric Duox1 possessing the A-loop of Duox2 released O2 (⨪). Moreover, Duox2 proteins possessing the A-loops of Nox1 or Nox5 co-expressed with DuoxA2 showed enhanced O2 (⨪) release, and Duox1 proteins possessing the A-loops of Nox1 or Nox5 co-expressed with DuoxA1 acquired O2 (⨪) leakage. Although we identified Duox1 A-loop residues (His(1071), His(1072), and Gly(1074)) important for reducing O2 (⨪) release, mutations of these residues to those of Duox2 failed to convert Duox1 to an O2 (⨪)-releasing enzyme. Using immunoprecipitation and endoglycosidase H sensitivity assays, we found that the A-loop of Duoxes binds to DuoxA N termini, creating more stable, mature Duox-DuoxA complexes. In conclusion, the A-loops of both Duoxes support H2O2 production through interaction with corresponding activators, but complex formation between the Duox1 A-loop and DuoxA1 results in tighter control of H2O2 release by the enzyme complex.

Negative charges in the flexible N-terminal domain of Rho GDP-dissociation inhibitors (RhoGDIs) regulate the targeting of the RhoGDI-Rac1 complex to membranes.

In its resting state, Rho GDP-dissociation inhibitor (RhoGDI) α forms a soluble cytoplasmic heterodimer with the GDP-bound form of Rac. Upon stimulation, the dissociation of RhoGDIα from the RhoGDIα-Rac complex is a mandatory step for Rac activation; however, this mechanism is poorly understood. In this study, we examined how the cytoplasm/membrane cycles of the RhoGDI-Rac complex are regulated, as well as where RhoGDI dissociates from the RhoGDI-Rac complex, during FcγR-mediated phagocytosis. The negatively charged and flexible N terminus (25 residues) of RhoGDIα, particularly its second negative amino acid cluster possessing five negatively charged amino acids, was a pivotal regulator in the cytoplasm/membrane cycles of the RhoGDI-Rac complex. We also found that RhoGDIα translocated to the phagosomes as a RhoGDIα-Rac1 complex, and this translocation was mediated by an interaction between the polybasic motif in the C terminus of Rac1 and anionic phospholipids produced on phagosomes, such as phosphatidic acid, that is, by a phagosome-targeting mechanism of Rac1. Thus, we demonstrated that the targeting/accumulation of the RhoGDIα-Rac1 complex to phagosomes is regulated by a balance between three factors: 1) the negatively charged and flexible N-terminal of RhoGDIα, 2) the binding affinity of RhoGDIα for Rac1, and 3) anionic phospholipids produced on phagosomes. Moreover, we demonstrated that the mechanism of targeting/accumulation of the RhoGDIα-Rac1 complex is also applicable for the RhoGDIß-Rac1 complex.

Potential of GSPT1 as a novel target for glioblastoma therapy.

Glioblastoma is the most common malignant brain tumor in adults, the survival rate of which has not significantly improved over the past three decades. Therefore, there is an urgent need to develop novel treatment modalities. We previously reported that G1 to S phase transition 1 (GSPT1) depletion induces delayed cell cycle in primary astrocytes. Herein, we examined the potential of GSPT1 as a novel target for glioblastoma therapy. CC-885, a cereblon modulator that degrades GSPT1 by bridging GSPT1 to the CRL4 E3 ubiquitin ligase complex, was administered to nude mice with transplanted brain tumors of U87 glioblastoma cells. The survival period was significantly longer in CC-885 treated mice than in control mice. Furthermore, we generated GSPT1-knockout (KO) U87 cells and GSPT1-KO U87 cells with stable overexpression of FLAG-tagged GSPT1 (Rescued GSPT1-KO). Mice with transplanted GSPT1-KO U87 cells and Rescued GSPT1-KO U87 cells showed significantly longer and similar survival periods, respectively, as those with wild-type (WT) U87 cells. GSPT1-KO U87 cells showed enhanced apoptosis, detected by cleaved PARP1, compared to WT U87 cells. Brain tumors with transplantation of GSPT1-KO U87 cells also showed enhanced apoptosis compared to those with transplantation of WT and Rescued GSPT1-KO U87 cells. GSPT1 expression was confirmed in patients with glioblastoma. However, the clinical study using 87 glioblastoma samples showed that GSPT1 mRNA levels were not associated with overall survival. Taken together, we propose that GSPT1 is an essential protein for glioblastoma growth, but not its malignant characteristics, and that GSPT1 is a potential target for developing glioblastoma therapeutics.

Prescribing Trends of Biologic Disease-Modifying Anti-rheumatic Drugs Using a Claims Database from 6 Million People in Japan.

BACKGROUND AND OBJECTIVE: Biologic disease-modifying anti-rheumatic drugs (bDMARDs) are used either when conventional synthetic DMARDs are ineffective or when disease activity is high and with poor prognostic factors, based on various clinical guidelines. The purpose of this study was to investigate the prescribing trends of bDMARDs for patients with rheumatoid arthritis in Japan, and to clarify whether the pharmacological therapy of bDMARDs is administered based on guidelines. METHODS: We conducted a descriptive epidemiological study from 2012 to 2018 using the JMDC Claims Database, a nationwide claims database, and described the annual changes based on the number of patients prescribed bDMARDs. Anti-rheumatic drugs were identified based on the Anatomical Therapeutic Chemical codes, including methotrexate, glucocorticoids, non-steroidal anti-inflammatory drugs and bDMARDs. RESULTS: From the database including 6,862,244 people, the data of 6407 patients with rheumatoid arthritis were extracted. The present study demonstrated that the proportion of patients prescribed bDMARDs was 1.0 per 1000 people, with those aged ≥ 65 years being the most common age group. The proportion of patients with rheumatoid arthritis who were prescribed bDMARDs increased significantly over time (p < 0.0001). Additionally, the concomitant proportions of methotrexate (p < 0.0001), non-steroidal anti-inflammatory drugs (p < 0.0001) and glucocorticoids (p = 0.0001) prescribed with bDMARDs decreased significantly over time. CONCLUSIONS: The increase in bDMARD monotherapy may be attributed to the new bDMARDs that have been launched sequentially; furthermore, physicians have come to recognise monotherapy as the mainstay of treatment. Future studies must accumulate evidence on the long-term efficacy and safety of bDMARDs.

Difference in the effects of tandospirone on ataxia in various types of spinocerebellar degeneration: an open-label study.

The aim of this study was to investigate the effects of tandospirone on ataxia in various types of spinocerebellar degeneration (SCD). Fifteen milligram per day of tandospirone was administered to 39 patients with SCD (spinocerebellar atrophy (SCA) 1, five patients; SCA2, six patients; Machado-Joseph disease (MJD), 14 patient; SCA6, five patients; multiple system atrophy-cerebellar type (MSA-C), seven patients; and multiple system atrophy-Parkinson type (MSA-P), two patients). All patients were assessed before and 4 weeks after administration of the drug using the international cooperative ataxia rating scale total score (ARS), total length traveled (TLT) of body stabilometry, and a self-rating depression scale. Statistically, ARS showed a significant difference in MJD (p = 0.005) and SCA6 (p = 0.043). TLT also showed a significant difference in MJD (p = 0.002) and SCA6 (p = 0.043). Eight of 39 patients (SCA1, 1/5; SCA2, 0/6; MJD, 4/14; SCA6, 3/5; MSA-C, 0/7; and MSA-P, 0/2) showed more than a five point reduction in ARS, and 13 of 39 patients (SCA1, 0/5; SCA2, 1/6; MJD, 8/14; SCA6, 4/5; MSA-C, 0/7; and MSA-P, 0/2) showed a reduction of TLT. Our data indicate that the effects of tandospirone on ataxia are different between types of SCD. Therefore, tandospirone is useful for cerebellar ataxia in patients with MJD and SCA6.

Rac-Dependent Signaling from Keratinocytes Promotes Differentiation of Intradermal White Adipocytes.

Rac signaling affects numerous downstream targets in vitro; however, few studies have established in vivo levels. We generated mice with a single knockout (KO) of Rac1 (Keratin5(K5)-Cre;Rac1flox/flox, Rac1-KO) and double KO of Rac1 and Rac3 (K5-Cre;Rac1flox/flox;Rac3-/-, Rac1/Rac3-DKO) in keratinocytes. The hairless phenotype in Rac1-KO mice was markedly exacerbated in Rac1/Rac3-DKO mice. Strikingly, Rac1-KO mice exhibited thinner dermal white adipose tissue, which was considerably further reduced in Rac1/Rac3-DKO mice. DNA microarray using primary keratinocytes from Rac1/Rac3-DKO mice exhibited decreased mRNA levels of Bmp2, Bmp5, Fgf20, Fgf21, Fgfbp1, and Pdgfα. Combinational treatment with bone morphogenetic protein (BMP) 2 and fibroblast growth factor (FGF) 21 in culture medium, but not individual purified recombinant proteins, could differentiate 3T3-L1 fibroblasts into adipocytes, as could culture media from primary keratinocytes. Conversely, addition of anti-BMP2 or anti-FGF21 antibodies into the culture medium inhibited fibroblast differentiation. In addition, BMP2 and FGF21 treatment promoted adipocyte differentiation only of rat primary white adipocyte precursors but not rat primary brown adipocyte precursors. Furthermore, BMP2 and FGF21 treatment enhanced adipogenesis of normal human dermal fibroblasts. Notably, brown adipogenesis promoted by FGF21 was inhibited by BMP2. Thus, we propose a complex paracrine pathway from keratinocytes to intradermal pre-adipocytes, which functions as a Rac-dependent modulator of both white and brown adipogenesis.

The 12-year trend report of antipsychotic usage in a nationwide claims database derived from four million people in Japan.

The current study aimed to describe the use of antipsychotics to clarify the gap between clinical guidelines and health care practice in Japan. We used data from the JMDC Claims Database (JMDC Inc., Tokyo, Japan), a nationwide claims database, from 2005 to 2016. Antipsychotics were defined as drugs coded as N05A with the Anatomical Therapeutic and Chemical (ATC) codes. We described the annual changes in proportions based on the number of patients prescribed any antipsychotics. From the database of 4,081,102 people, the data of 12,382 patients was extracted by applying the following exclusion criteria: no use of antipsychotics, missing the prescription date or dose, inpatients, prescribed antipsychotics only for use as needed, prescribed only injectable antipsychotics except for long-acting injections (LAIs), without schizophrenia as the primary disease, not exceeding 75 mg/day chlorpromazine equivalent, and less than 18 years old. The use of second-generation antipsychotics (SGA) has been expanding, while the use of first-generation antipsychotics has been decreasing. Aripiprazole accounted for the highest proportion of prescribed antipsychotics (31.9%) in 2016. Even though clozapine is categorized as a SGA, it accounted for a paltry 0.2%. The proportion of prescribed antipsychotics accounted for by LAIs was less than 5%. Although the use of antipsychotics for schizophrenia in Japan mostly corresponds to various clinical guidelines, limited use of clozapine and LAIs was identified. Further research focusing on the factors affecting the prescription of these underused antipsychotics may help advance the pharmacological therapy of schizophrenia.

What can predict and prevent the long-term use of benzodiazepines?

Although benzodiazepines (BZDs) are commonly prescribed for insomnia or anxiety, long-term use of BZDs causes serious adverse effects such as daytime drowsiness and cognitive decline. In the current study, we evaluated the predictors and preventers of long-term usage of BZDs from a retrospective survey by utilizing the 12-year prescription record of a university hospital. From the prescription data of 92,005 people, users of BZDs (n = 3,470, male = 39.2%, mean age = 60 ± 17.5) were analyzed. During this period, both the number of prescriptions (2722 in 2004 to 1019 in 2016) and the number of BZDs (1.73 in 2004 to 1.36 in 2016) gradually decreased, although more than half of the patients continued to take BZDs for over three years. High risk factors for long-term use of BZDs include elderly patients (>65 years old), high dosage (>5 mg diazepam per day), psychiatrist-prescribers, and users with polytherapy. Discontinuation is significantly found in users of hypnotic BZDs and alternative psychotropic medical drugs (including antipsychotics, serotonergic drugs, or newer types of sleep medicine). Future studies should focus on elucidating interventions that are more effective against long-term usage of BZDs.

CTLA-4 gene polymorphism is not associated with conventional multiple sclerosis in Japanese.

We investigated the polymorphisms of exon 1 (+49A/G) and promoter (-318C/T and -651C/T) regions of the CTLA-4 gene in 133 Japanese patients with conventional/classical multiple sclerosis (MS) and 156 healthy controls. Patients with optico-spinal MS (OSMS) or atypical clinical attacks were excluded from the study. There was no significant difference in the distribution of polymorphisms between patients and controls. Furthermore, there were no associations between polymorphisms and clinical characteristics, such as age at onset, disease prognosis, and HLA profiles. Our results suggest that CTLA-4 gene polymorphisms are neither conclusively related to susceptibility nor to the clinical characteristics of MS, especially in Japanese patients with conventional/classical form and clinical features identical to those of their counterparts in Western countries.

CSF pleocytosis and expansion of spinal lesions in Japanese multiple sclerosis with special reference to the new diagnostic criteria.

New diagnostic criteria for multiple sclerosis (MS) were recently proposed from the international panel on the diagnosis of MS, and they include exclusion criteria, such as lesions extending over more than two vertebral segments on spinal MRI and CSF pleocytosis of more than 50/mm3. We reviewed the clinical features of 158 patients who satisfied the diagnostic criteria for MS except for having the above atypical paraclinical findings. All patients exhibited two or more clinical attacks and objective clinical evidence of multiple lesions without any evidence of other disorders. Thirty-three (20.9%) patients had one or both atypical paraclinical findings. Twenty-one out of the 33 patients were classified as having optico-spinal MS (OSMS), and the other 12 as non-OSMS patients with atypical large expanding or destructive cerebral, cerebellar or brainstem lesions on MRI as well as one or both atypical paraclinical findings. Based on this heterogeneity in clinical findings in MS, there is an urgent need to develop a common general concept of the "MS" syndromes, and the ethnic-related heterogeneity should be considered in the revised criteria for the diagnosis of MS.

Estrogen receptor gene polymorphism and multiple sclerosis in Japanese patients: interaction with HLA-DRB1*1501 and disease modulation.

We investigated PvuII and XbaI polymorphism in the estrogen receptor gene (ERG) and HLA-DRB1*1501 positivity in 116 conventional multiple sclerosis (MS) patients and 101 healthy controls in a Japanese population. Logistic analysis revealed independent associations of [P] allele in the profiles for PvuII (p=0.0005, adjusted odds ratio (aOR)=3.17) and DRB1*1501 (p=0.0089, aOR=2.61) with conventional MS. Synergistic elevated risk of MS due to interaction between the [P] allele and HLA-DRB1*1501 allele was found among female patients (odds ratio=16.0; 95% CI=3.99-63.8, p<0.0001). The [P] allele-positive patients with disease duration of more than 5 years had a significantly higher progression index (PI) of disability (p=0.0230) and a worse ranked MS severity score (p=0.0152) than their non-[P] counterparts.

Positional vertigo and macroscopic downbeat positioning nystagmus in spinocerebellar ataxia type 6 (SCA6).

To investigate the frequency of positioning nystagmus in degenerative ataxic disorders, we examined downbeat positioning nystagmus (DPN) in 25 patients with spinocerebellar ataxia type 6 (SCA6) and 58 patients with other types of degenerative ataxia. DPN was observed in 21 of the 25 patients with SCA6 (84 %) versus only 3 of the 58 patients (5.2 %) with other types of degenerative ataxia, including multiple system atrophy, SCA1, SCA2, SCA3/Machado-Joseph disease, and non-SCA6 late-onset pure cerebellar ataxia. Our findings indicated that DPN is a distinct part of the clinical presentation of SCA6, showing that vestibular cerebellum is more affected in SCA6 than other types of degenerative ataxia.

Attack-related severity: a key factor in understanding the spectrum of idiopathic inflammatory demyelinating disorders.

Understanding the spectrum of idiopathic inflammatory demyelinating disorders (IIDD) is a fundamental issue for the diagnosis and treatment of these disorders as well as for the approach to their pathogenesis. The spectrum of IIDD is usually classified according to clinical course and lesion distribution. We compared the demographic features, clinical characteristics, laboratory findings, and genetic backgrounds between 193 Japanese patients with and without clinically or radiographically fulminant attacks who all satisfied the diagnostic criteria for multiple sclerosis (MS). "Fulminant attacks" in the current study represent attack-related clinically or radiologically severe relapses but do not necessarily mean severe disability. Patients with fulminant attacks were clinically and immunogenetically distinct from those free of such attacks, and the previously described characteristics of the opticospinal form of MS (OSMS) or neuromyelitis optica (NMO) were mostly shared by patients with fulminant attacks. HLA profiles were similar among patients with fulminant attacks irrespective of the lesion distributions. The GG homozygous and G alleles of the CTLA4 gene A/G coding SNP at position 49 in exon 1 were significantly more common in patients with fulminant attacks than in those without. Attack-related severity may be an important factor if validated by prospective studies defining criteria and establishing relationships to disease course and treatment regimens.

Effects of tandospirone on "5-HT1A receptor-associated symptoms" in patients with Machado-Josephe disease: an open-label study.

BACKGROUND: We investigated the frequencies of the symptoms such as "ataxia, depression, insomnia, anorexia, and pain," that have been reported to be associated with 5-HT1A receptor, and the effect of tandospirone citrate (tandospirone: 5-HT1A agonist) in patients with Machado-Joseph disease (MJD). METHODS: Ten MJD patients received tandospirone (15-30 mg/d) for seven weeks. During that time, they were evaluated weekly using the Ataxia Rating Scale (ARS) and Total Length Traveled (TLT) by Stabilimetry tests, the Self-rating Depression Scale (SDS), which in addition to evaluating their level of depression, also evaluated their degree of insomnia and anorexia, and a pain questionnaire. RESULTS: Before tandospirone therapy, all patients displayed cerebellar ataxia, while insomnia, and leg pain was observed in 7 patients, depression in 6 patients, and anorexia was observed in 2 patients. In response to treatment, 7 of the 10 patients who were ataxic showed a reduction in their ARS, while 3 of 6 patients showed a reduction in their SDS, and 5 of 7 patients showed an alleviation of their insomnia and leg pain. Both of the affected patients showed a marked improvement in their anorexia. A stabilimetry test could be performed in 7 patients, 5 of whom showed a reduction in TLT. CONCLUSIONS: Our data indicate that the patients with MJD are prone to manifest 5-HT1A receptor-associated symptoms, and tandospirone is a useful drug for these symptoms in patients with MJD, though a double-blind study is needed.

[Susceptibility genes for multiple sclerosis].

Multiple lines of evidence support the role of genetic factors for susceptibility to multiple sclerosis (MS), although unknown environmental factors must play an important role in developing MS. In this paper, we review the reports studied for the possible associations between Japanese MS and candidate genes by using case-control method. As for HLA alleles, HLA-DRB1*1501 and -DPB1*0501 allele have been confirmed to be associated with conventional MS and opticospinal MS, respectively. Some polymorphisms of vitamin D receptor gene, estrogen receptor gene, CTLA-4 gene, and osteopontin gene were reported to be associated with conventional MS. Appropriate case ascertainment and collection as well as proper statistical method are needed to isolate the susceptibility genes for MS.