Combination of cytoplasmic and nuclear patterns on Hep-2 antinuclear antibody is useful as a screening test for anti-synthetase syndrome.

OBJECTIVE: This study aimed to establish a screening model for differentiating anti-synthetase syndrome (ASS) from other antinuclear antibody (ANA)-associated rheumatic diseases (AARD) using a combination of cytoplasmic and non-cytoplasmic ANA (ncANA) patterns. METHODS: This retrospective observational study included patients with AARDs such as systemic lupus erythematosus (SLE), systemic sclerosis (SSc), Sjögren's syndrome (SS), mixed connective tissue disease (MCTD), and polymyositis/dermatomyositis (PM/DM) who underwent ANA screening between April 2012 and December 2021. Variables included age, sex, ANA patterns (Cytoplasmic and ncANA), and titers. Logistic regression analysis of Cytoplasmic and ncANA patterns was performed to differentiate ASS from other AARDs. RESULT: The 981 diagnosed cases of AARDs consisted of SS (n = 451), SSc (n = 264), SLE (n = 201), PM/DM (n = 104), MCTD (n = 52), and ASS, including PM/DM (n = 64). Of these, 155 patients had ≥2 overlapping diseases; however, there was no overlap between AARDs and ASS. ASS is more likely to occur when the cytoplasmic titer is positive and the ncANA <320. Receiver operating characteristic (ROC) analysis of the Cytoplasmic and ncANA range revealed an area under the ROC curve (AUC) of 0.885 (95% CI: 0.844 to 0.927). CONCLUSION: It is important to detect cytoplasmic patterns as an ANA screening test for ASS diagnosis, even if the titer is low. Additionally, combining the cytoplasmic and ncANA patterns yields more accurate ASS screening results.

Genetic and clinical prediction models for the efficacy and hepatotoxicity of methotrexate in patients with rheumatoid arthritis: a multicenter cohort study.

The objective of the study is to develop genetic and clinical prediction models for the efficacy and hepatotoxicity of methotrexate (MTX) in patients with rheumatoid arthritis (RA). Among RA patients treated with MTX, 1966 polymorphisms of 246 enzymes/transporters relevant to pharmacokinetics and pharmacodynamics were measured by the Drug Metabolism Enzymes and Transporters (DMET) microarray and direct sequencing, and clinical variables at baseline were collected. For efficacy, response criteria of the European League Against Rheumatism were used to classify patients as responders or non-responders. Hepatotoxicity was defined as elevations of aspartate aminotransferase or alanine aminotransferase ≥1.5 times the reference range upper limit. Among 166 patients, a genetic prediction model for efficacy using seven polymorphisms showed the area under the receiver operating characteristic curve (AUC) was 0.822, with 74.3% sensitivity and 76.8% specificity. A combined genetic and clinical model indicated the AUC was 0.844, with 81.5% sensitivity and 76.9% specificity. By incorporating clinical variables into the genetic model, the overall category-free net reclassification improvement (NRI) was 0.663 (P < 0.0001) and the overall integrated discrimination improvement (IDI) was 0.083 (P = 0.0009). For hepatotoxicity, a genetic prediction model using seven polymorphisms showed the AUC was 0.783 with 70.0% sensitivity and 80.0% specificity, while the combined model indicated the AUC was 0.906 with 85.1% sensitivity and 87.8% specificity (overall category-free NRI: 1.002, P < 0.0001; overall IDI: 0.254, P < 0.0001). Our genetic and clinical models demonstrated moderate diagnostic accuracy for MTX efficacy and high accuracy for hepatotoxicity. These findings should, however, be validated and interpreted with a caution until external validation.

Predictive model of bosentan-induced liver toxicity in Japanese patients with pulmonary arterial hypertension.

Bosentan, an endothelin receptor antagonist, has been widely used as a first-line medication for the treatment of pulmonary arterial hypertension (PAH). It has been shown to improve symptoms of hypertension, exercise capacity, and hemodynamics and prolong time to clinical worsening. However, liver dysfunction is a major side effect of bosentan treatment that could hamper the optimal management of patients with PAH. Previously, we demonstrated, using drug metabolism enzymes and transporters analysis, that the carbohydrate sulfotransferase 3 (CHST3) and CHST13 alleles are significantly more frequent in patients with elevated aminotransferases during therapy with bosentan than they are in patients without liver toxicity. In addition, we constructed a pharmacogenomics model to predict bosentan-induced liver injury in patients with PAH using two single-nucleotide polymorphisms and two nongenetic factors. The purpose of the present study was to externally validate the predictive model of bosentan-induced liver toxicity in Japanese patients. We evaluated five cases of patients treated with bosentan, and one presented with liver dysfunction. We applied mutation alleles of CHST3 and CHST13, serum creatinine, and age to our model to predict liver dysfunction. The sensitivity and specificity were calculated as 100% and 50%, respectively. Considering that PAH is a rare disease, multicenter collaboration would be necessary to validate our model.

CHST3 and CHST13 polymorphisms as predictors of bosentan-induced liver toxicity in Japanese patients with pulmonary arterial hypertension.

Bosentan, an endothelin receptor antagonist, has been widely used as a first-line drug for the treatment of pulmonary arterial hypertension (PAH). In addition, bosentan is approved for patients with digital ulcers related to systemic sclerosis. Liver dysfunction is a major adverse effect of bosentan and may lead to discontinuation of therapy. The purpose of this study was to identify genomic biomarkers to predict bosentan-induced liver injury. A total of 69 PAH patients were recruited into the study. An exploratory analysis of 1936 single-nucleotide polymorphisms (SNPs) in 231 genes involved in absorption, distribution, metabolism, and elimination of multiple medications using Affimetrix DMET™ (Drug Metabolism Enzymes and Transporters) chips was performed. We extracted 16 SNPs (P < 0.05) using the Jonckheere-Terpstra trend test and multiplex logistic analysis; we identified two SNPs in two genes, CHST3 and CHST13, which are responsible for proteoglycan sulfation and were significantly associated with bosentan-induced liver injury. We constructed a predictive model for bosentan-induced liver injury (area under the curve [AUC]: 0.89, sensitivity: 82.61%, specificity: 86.05%) via receiver operating curve (ROC) analysis using 2 SNPs and 2 non-genetic factors. Two SNPs were identified as potential predictive markers for bosentan-induced liver injury in Japanese patients with pulmonary arterial hypertension. This is the first pharmacogenomics study linking proteoglycan sulfating genes to drug-induced liver dysfunction, a frequently observed clinical adverse effect of bosentan therapy. These results may provide a way to personalize PAH medicine as well as provide novel mechanistic insights to drug-induced liver dysfunction.

Clinical characteristics of and risk factors for serious infection in Japanese patients within six months of remission induction therapy for antineutrophil cytoplasmic antibody-associated vasculitis registered in a nationwide, prospective, inception cohort study.

OBJECTIVES: The purpose of this study was to identify the clinical characteristics and predictors of serious infections (SIs) in the RemIT-JAV, a nationwide, prospective, inception cohort study for Japanese patients with antineutrophil cytoplasmic antibody-associated vasculitis (AAV). METHODS: We analyzed SIs within six months of remission induction therapy in 156 AAV patients. Hazard ratios with 95% confidence intervals (CIs) for SIs were calculated using the COX proportional hazard model. RESULTS: Sixty-three SIs in 42 patients were identified. The incidence rate (IR) of SIs was 87.59/100 patient-years. The median length of time to the onset of first SIs was 54 days. Hazard ratios (95%CI) for SIs were 1.97 (0.99-3.95) for age >65 years, 0.47 (0.25-0.89) for female sex, 2.11 (1.05-4.27) for the severe form of AAV, and 2.88 (1.49-5.88) for initial PSL >0.8 mg/kg/day in the first model, and 2.64 (1.39-5.01) for smoking and 3.27 (1.66-6.45) for initial PSL >0.8 mg/kg/day in the second model. CONCLUSIONS: Lowering the IR of SIs in Japanese AAV patients is mandatory to improve the vital prognosis of these patients. For remission induction therapy of AAV patients with these risk factors, risk management of immunosuppressive treatment should be carefully considered.

[Evaluation of a Computer-Aided Microscope System and Its Anti-Nuclear Antibody Test Kit for Indirect Immunofluorescence Assay].

Antinuclear antibody (ANA) testing is indispensable for diagnosing and understanding clinical conditions of autoimmune diseases. The indirect immunofluorescence assay (IFA) is the gold standard for ANA screening, and it can detect more than 100 different antibodies, such as anti-PCNA as well as anti-cytoplasmic antibodies. However, complicated procedures of conventional IFA and visual interpretation require highly skilled laboratory staff. This study evaluates the capability, characteristics, and applicability of the recently developed ANA detection system (EUROPattern Cosmic IFA System, EPA) using HEp20-10 cells and the automated pattern recognition microscope. Findings using EPA and conventional methods were compared in 282 sera obtained from connective tissue disease patients and 250 sera from healthy individuals. The concordance of the positivity rate, antibody titer (within +/- 1 tube difference), and the accurate recognition rate of ANA patterns between the automated EPA method and the microscopic judgement of the EPA image by eye was 98.9, 97.4, and 55.3%, respectively. The EPA method showed concordance of the positivity rate as high as 93.3% and concordance of the antibody titer as high as 94.0% (within +/- 1 titer) compared with the conventional method. Regarding the four typical patterns of ANA (homogeneous, speckled, nucleolar, and centromere), large differences between the EPA and conventional methods were not observed, and the rate of concordance between the final EPA result and the conventional method was from 94.1 to 100%. The positivity rate of ANA using the EPA and conventional methods showed marked agreement among the six connective tissue diseases (SLE, MCTD, SSc, PM/DM, and SS) and healthy individuals. Although the EPA system is not considered a complete system and laboratory staff should verify the results, it is a useful system for routine ANA analysis because it contributes to ANA standardization and an efficient workflow.

Granulomatosis with polyangiitis (Wegener's granulomatosis) diagnosed by transbronchial biopsy from stenotic portion of mainstem bronchus.

This report concerns a case of granulomatosis with polyangiitis (GPA) (Wegener's granulomatosis (WG)) with bronchus narrowing. Although nasal biopsy had been performed three times, no positive histology for GPA (WG) could be obtained. Flexible bronchoscopy revealed diffuse erythema, edema of the mucosa and stenosis of the right mainstem bronchus. Transbronchial biopsy identified granuloma with giant cells. These findings led to a diagnosis of GPA (WG). This case suggests that biopsy from the bronchus is useful for diagnosis of GPA (WG).

[Personalized Medicine in Rheumatoid Arthritis].

Medical strategy for rheumatoid arthritis (RA) has markedly advanced in recent years. The introductions of biologics and methotrexate as an anchor drug have made it possible to not only suppress pain and inflammation (clinical remission), but also to inhibit joint destruction (structural remission), leading to cure of the disease. In order to achieve this target, it is the most important to diagnose RA early and promote disease remission. However, since the condition and pathology are diverse among patients, optimal treatment for each patient is desired (personalized medicine). Treatment should be performed under consideration of the disease state such as activity, prognosis regarding joint destruction, and complications. It is also important to clarify the patient characteristics, such as responsiveness to the drugs and risk of adverse effects. Biomarkers, such as proteomics and pharmacogenomics (genetic polymorphism, etc.), are indispensable for personalized medicine. We have established a predictive model for methotrexate hepatotoxicity, consisting of 13 SNPs with a sensitivity of 100% and specificity of 89%, although the model should be validated with a larger-scale prospective study. RA is a multifactorial disorder with clinically heterogeneous features. Gene-environment interaction is closely involved in the production of anti-CCP antibodies (ACPA); thereafter, secondary stimuli of joints may lead to symptoms of RA. Joint injury, emotional stress, and infections often trigger the onset of RA. Cure can be achieved through complete remission by early aggressive treatment and returning to the pre-clinical state of RA with environmental improvement.

[Optimal Treatment for Rheumatoid Arthritis with Companion Diagnostics].

The medical strategy for rheumatoid arthritis (RA) has markedly advanced in recent years. The introduction of biologics in addition to methotrexate, an anchor drug, has made it possible to not only suppress pain and inflammation (clinical remission), but also inhibit joint destruction (structural remission), leading to cure from the disease. Since the condition and pathology are heterogeneous among individual patients, optimal treatment for each patient based on the use of companion diagnostics is desired (precision medicine). ACPA is important to diagnose RA, but also to assess the prognosis. ACPA is also a part of companion diagnostics for preclinical RA because it has been found to be positive before the onset. Treatment should be performed under consideration of the disease state such as activity, prognosis regarding joint destruction, and complications. It is also important to clarify the patient characteristics, such as responsiveness to the drugs and risk of adverse effects. Biomarkers, such as proteomics and pharmacogenomics, have been reported as companion diagnostics for optimal treatment of RA. RA is a multifactorial disorder with clinically heterogeneous features. Gene-environment interaction is closely involved in the production of ACPA, and then secondary stimuli to joints may lead to symptoms of RA. Joint injury, emotional stress, and infections often trigger the onset of RA. It is possible to cure RA, achieving complete remission, by early aggressive treatment and returning to the pre-clinical state with environmental improvement. [Review]

[MMP-3 as a Biomarker of Disease Activity of Rheumatoid Arthritis].

The aim of this study was to confirm the clinical significance of serum MMP-3 measurement in the evalua- tion of disease activity and effectiveness of treatment in patients with rheumatoid arthritis (RA). MMP-3 was measured for 206 outpatients with RA during a period of 4 months, and also serially measured for RA patients treated with methotrexate(MTX) alone or together with infliximab (IFX). Serum MMP-3 was significantly correlated with CRP, SAA, and ESR. Significant correlation of serum MMP-3 was found not only with DAS28 (CRP) in female and male patients (p <0.0001 and p < 0.0051, respectively) but also with the EULAR classification criteria for the disease activity of RA. Among the items of DAS28(CRP), the strongest association of MMP-3 was found with swollen joint counts. Furthermore, MMP-3 levels increased with advances in Stage and Class of RA. MMP-3 levels gradually decreased 12 and 24 weeks after successful treatment with MTX (p=0.0188 and p=0.0179, respectively). Extent of the decrease was more prominent in patients with better response to MTX than in those with poor response. MMP-3 levels significantly decreased 6 weeks after IFX treatment and continued to decrease until 48 weeks. Significant decrease of MMP-3 level from before treatment was shown only in the good response group to IFX after 48 weeks of treatment. MMP-3 level was shown to be useful as a disease activity marker in RA patients. In addition, serial measurement of MMP-3 maybe helpful to evaluate the effect of treatments with MTX and IFX.

CD45RA-Foxp3(low) non-regulatory T cells in the CCR7-CD45RA-CD27+CD28+ effector memory subset are increased in synovial fluid from patients with rheumatoid arthritis.

Increased numbers of regulatory T (Treg) cells are found in synovial fluid from patients with rheumatoid arthritis (RASF) compared with peripheral blood. However, Treg cells in RASF have been shown to have a decreased capacity to suppress T cells. Here we phenotypically classified CD4+ T cells in RASF into six subsets based on the expression of CD45RA, CCR7, CD27 and CD28, and demonstrated that the CCR7-CD45RA-CD27+CD28+ TEM subset was significantly increased in synovial fluid compared with peripheral blood. In addition, the proportion of Foxp3+ Treg cells in the CCR7-CD45RA-CD27+CD28+ TEM subset was significantly increased in RASF. Furthermore, most of the Foxp3+ Treg cells in RASF were non-suppressive CD45RA-Foxp3(low) non-Treg cells, and the frequency of the non-Treg cells in the CCR7-CD45RA-CD27+CD28+ TEM subset was significantly increased in RASF. Our findings suggest that the pro-inflammatory environment in RA joints may induce the increase of CD45RA-Foxp3(low) non-Treg cells in synovial fluid.

Cancer incidence in systemic sclerosis: meta-analysis of population-based cohort studies.

OBJECTIVE: To examine cancer incidence in patients with systemic sclerosis (SSc) through a meta-analysis of population-based cohort studies. METHODS: Five different databases (Medline, Scopus, CINAHL [Cumulative Index to Nursing and Allied Health Literature], Web of Science, and Cochrane Collaboration) were searched for articles published between January 1966 and May 2012; review articles and the reference lists from the articles that resulted from the search were also evaluated. Population-based cohort studies with data relevant to the determination of cancer risk in patients with SSc were included. All articles that met strict inclusion criteria were analyzed for data on population size, time of followup, and observed-to-expected cancer ratios, also known as standardized incidence ratios (SIRs). RESULTS: Six articles met criteria and were included in the meta-analysis. The pooled SIR for the incidence of cancer overall was 1.41 (95% confidence interval [95% CI] 1.18-1.68), and significant heterogeneity was observed as a consequence of variability in the participants, outcome, study design, and risk of bias among the studies. Men had a significantly higher pooled SIR (1.85 [95% CI 1.49-2.31]) than women (SIR 1.33 [95% CI 1.18-1.49]) (P < 0.01), and stratification for sex eliminated heterogeneity, which indicates that variability among the studies greatly contributed to differences between the sexes. There were no differences between limited cutaneous SSc and diffuse cutaneous SSc (P = 0.77). Significant increases were observed in the risk of cancer of the lung, liver, hematologic system, and bladder, as well as of non-Hodgkin's lymphoma and leukemia. CONCLUSION: SSc is associated with an increased risk of cancer, particularly lung, liver, hematologic, and bladder cancers, although absolute risk is relatively low. Men with SSc have a higher risk of developing cancer than women.

Proteasome assembly defect due to a proteasome subunit beta type 8 (PSMB8) mutation causes the autoinflammatory disorder, Nakajo-Nishimura syndrome.

Nakajo-Nishimura syndrome (NNS) is a disorder that segregates in an autosomal recessive fashion. Symptoms include periodic fever, skin rash, partial lipomuscular atrophy, and joint contracture. Here, we report a mutation in the human proteasome subunit beta type 8 gene (PSMB8) that encodes the immunoproteasome subunit ß5i in patients with NNS. This G201V mutation disrupts the ß-sheet structure, protrudes from the loop that interfaces with the ß4 subunit, and is in close proximity to the catalytic threonine residue. The ß5i mutant is not efficiently incorporated during immunoproteasome biogenesis, resulting in reduced proteasome activity and accumulation of ubiquitinated and oxidized proteins within cells expressing immunoproteasomes. As a result, the level of interleukin (IL)-6 and IFN-γ inducible protein (IP)-10 in patient sera is markedly increased. Nuclear phosphorylated p38 and the secretion of IL-6 are increased in patient cells both in vitro and in vivo, which may account for the inflammatory response and periodic fever observed in these patients. These results show that a mutation within a proteasome subunit is the direct cause of a human disease and suggest that decreased proteasome activity can cause inflammation.

[Evaluation of cut-off value for autoantibodies against double-stranded DNA-complexed nucleosomes based on enzyme linked immunosorbent assay].

The new classification criteria for systemic lupus erythematosus (SLE) by Systemic Lupus International Collaborating Clinics (SLICC) published in 2012 have defined positive level titer of antibody against double stranded (ds) DNA as more than double the reference range if tested by enzyme linked immunosorbent assay (ELISA). The aim of this study was to evaluate optimal cut-off value and diagnostic performance of the anti-dsDNA nucleosome-complexed (anti-dsDNA-NcX) ELISA, which uses salmon testis DNA complexed with purified nucleosomes as antigens, for diagnosis of SLE. Titers of antibodies against dsDNA-complexed nucleosome were measured in sera of 76 patients with SLE, 148 with other connective tissue diseases, and 323 healthy volunteers. Sensitivity, specificity, correlation and concordance rate were compared among anti-dsDNA-NcX, conventional ELISA methods (EIA) and radioimmunoassay (RIA). As a results, concordance rates and Spearman's coefficient of rank correlation (r(s)) of anti-dsDNA-NcX with EIA and RIA were 59.2%, r(s) = 0.40 and 53.9%, r(s) = 0.21, respectively. Receiver operating characteristic curve analysis showed that the titer of 44 IU/mL calculated as 99 percentile of 323 healthy volunteers is a better cut-off value for anti-dsDNA-NcX than the 100 IU/mL recommended by the manufacturer. By setting the cut-off value at 44 IU/mL, anti-dsDNA-NcX showed the highest sensitivity (75.0%) and specificity (90.5%) of the 3 assays. With SLICC criterion, positivity rates of anti-dsDNA-NcX, EIA and RIA for SLE patients were 50.0%, 30.3% and 50.0%, respectively. In conclusion, anti-dsDNA-NcX has a good diagnostic performance for SLE with our proposed cut-off value of 44 IU/mL.

CD45RA-Foxp3(high) activated/effector regulatory T cells in the CCR7 + CD45RA-CD27 + CD28+central memory subset are decreased in peripheral blood from patients with rheumatoid arthritis.

Human CD4+ T cells can be classified as either naïve, central memory (TCM), or effector memory (TEM) cells. To identify the CD4+ T cell subsets most important in the pathogenesis of rheumatoid arthritis (RA), we phenotypically defined human CD4+ T cells as functionally distinct subsets, and analyzed the distribution and characteristics of each subset in the peripheral blood. We classified CD4+ T cells into six novel subsets based on the expression of CD45RA, CCR7, CD27, and CD28. The CCR7 + CD45RA-CD27 + CD28+ TCM subset comprised a significantly smaller proportion of CD4+ T cells in RA patients compared to healthy controls. The frequency of TNF-α-producing cells in the CCR7-CD45RA-CD27 + CD28+ TEM subset was significantly increased in RA. Furthermore, within the CCR7 + CD45RA-CD27 + CD28+ TCM subset, which was decreased in periperal blood from RA, the proportions of total Foxp3+ Treg cells and CD45RA-Foxp3(high) activated/effector Treg cells were significantly lower in RA patients. Our findings suggest that the increased proportion of TNF-α-producing cells and the decreased proportion of CD45RA-Foxp3(high) activated/effector Treg cells in particular subsets may have critical roles in the pathogenesis of RA.

Mycophenolate mofetil versus intravenous cyclophosphamide for induction treatment of proliferative lupus nephritis in a Japanese population: a retrospective study.

OBJECTIVES: Recent studies have shown that mycophenolate mofetil (MMF) is similar to intravenous cyclophosphamide (IVC) for the treatment of lupus nephritis (LN), but that treatment response may vary according to location and race/ethnicity. Moreover, no studies have been conducted to compare the efficacy of MMF with that of IVC for a Japanese population. We therefore conducted a retrospective study to clarify the efficacy and safety of MMF compared with that of IVC for induction therapy for active LN, classes III and IV, in a Japanese population of 21 patients, 11 of whom received MMF and 10 IVC. METHODS: The primary endpoint was expressed as the percentage of responders, who in turn were defined as the patients who met complete or partial response criteria according to the European consensus statement. The secondary endpoints comprised the renal activity component and serological activity. RESULTS: The primary endpoint was achieved in nine (81.8 %) patients receiving MMF and in four (40.0 %) receiving IVC, with no significant difference between the two groups (p = 0.081), while there was also no significant difference between them in terms of secondary endpoints. However, the MMF group suffered significantly fewer hematologic toxic effects than the IVC group. CONCLUSIONS: MMF may be used as an alternative to IVC for inducing renal remission of LN in Japanese patients.

[Introductory remarks by chairpersons].

Personalized medicine (PM) is expected to change the healthcare system due to great progress in areas such as pharmacogenomics (PGx) and molecular diagnostics; however, PM is more popular in oncology than in clinic practice. Oncological therapies are easily personalized because accurate diagnostic tests are available to identify patients who can benefit from targeted therapies. Nevertheless, personalized medicine has become a more imminent reality for several reasons, such as the increased awareness of drugs with an adequate benefit/risk ratio to patients, better methods to optimize drug selection and dosing and so on. We invited four speakers to discuss the importance of PGx in the practice of PM from the viewpoint of basic and clinical research.

Combination of echocardiography and pulmonary function tests could predict no complication of pulmonary hypertension during 5 years in patients with systemic sclerosis.

OBJECTIVE: To determine whether complications of pulmonary hypertension (PH) can be predicted by noninvasive screening tests in systemic sclerosis (SSc). METHODS: Forty-seven of 113 SSc patients underwent right heart catheterization (RHC) during 2011-2014. Clinical data, hemodynamic features, echocardiography, and pulmonary function tests had been followed up from the first RHC until 5 years later. RESULTS: At the first RHC, out of 44 patients, 8 were diagnosed with pre-capillary PH (mean pulmonary arterial pressure [mPAP] > 20 mm Hg), and 36 patients were defined as no-PH (mPAP ≤ 20 mm Hg). Three patients with >15 mm Hg of pulmonary artery wedge pressure were excluded. Receiver operating characteristic analyses for pre-capillary PH using estimated systolic PAP (esPAP) revealed an area under the curve (AUC) of 0.736, with a sensitivity and specificity of 62.5% and 86.1%, respectively, at a cutoff level of 35.0 mm Hg. The predicted percentage diffusing lung capacity for carbon monoxide (DLCO%) revealed an AUC of 0.840, with a sensitivity and specificity of 85.7% and 80.0%, respectively, at a cutoff level of 70.0%. Six pre-capillary PH patients, including one who died from PH 14 months after the first RHC, indicated exacerbations of mPAP or esPAP within 5 years. When esPAP < 35.0 mm Hg and DLCO% > 70% were met as the cutoff, none had been newly diagnosed with PH over 5 years. CONCLUSIONS: The conventional screening tests may be useful for detecting pre-capillary PH with SSc, and both esPAP < 35.0 mm Hg and DLCO% > 70% indicated a lower risk of developing PH for at least 5 years.