BackgroundPulmonary arterial hypertension (PAH) is one of the most severe complications and the leading cause of death in patients with systemic lupus erythematosus (SLE). Pathogenic mechanisms leading to SLE-PAH are still not fully understood. In idiopathic PAH, the dysfunction of the bone morphogenetic protein (BMP) pathway was found to be involved in pulmonary artery remodeling [1]. In SLE-PAH, our previous study identified serum autoantibodies targeting BMP receptors (BMPR) as the potential biomarker, including anti-BMPR2, BMPR1A, and Activin Receptor-Like Kinase type 1 (ALK1) antibodies [2]. However, the clinical and pathological roles of the above autoantibodies targeting BMP signaling in SLE-PAH remain uncertain.ObjectivesTo investigate the clinical and pathological roles of autoantibodies targeting BMPR2, BMPR1A, and ALK1 in SLE-PAH.MethodsPatients of SLE-PAH confirmed by right heart catheterization were enrolled and serum levels of autoantibodies targeting BMPR2, BMPR1A, and ALK1 were measured by ELISA. Patients with SLE-PAH were subtyped by cluster analysis. Survival and treatment goal achievement were further compared between different clusters. The Id1 and phospho-SMAD1/5 (p-SMAD1/5) levels were examined in human pulmonary artery endothelial cells (hPAEC) under the induction of BMP9 combined with anti-BMPR2, anti-ALK1, and anti-BMPR1A neutralizing antibodies (5 μg/mL). The effect of anti-ALK1 antibodies on the extent of monolayer permeability and apoptosis of hPAEC was further examined.Results60 SLE-PAH patients were enrolled and cluster analysis revealed two distinct clusters according to the positivity of autoantibodies targeting BMP signaling and clinical manifestations. Cluster 1 was a “low-antibody and high-disease activity” cluster while cluster 2 was a “high-antibody and low-disease activity” cluster. Patients in cluster 1 showed a higher proportion of nephropathy (76.9%) and SLE activity, however a low positivity rate of autoantibodies targeting BMP signaling. Patients in cluster 2 were characterized by a higher rate of anti-BMPR2 antibodies (82.4%), anti-ALK antibodies (70.6%), and lower SLE activity. Prognostic analysis showed that the proportion of patients who reached the treatment target was relatively higher in cluster 2. Mechanism study showed that the p-SMAD1/5 level and the Id1expression were decreased, indicating the suppression of BMP signaling in the presence of anti-ALK1 antibodies. Functional studies showed that anti-ALK1 antibodies increased the monolayer permeability of hPAEC. The late-stage apoptosis of hPAECs was also induced by anti-ALK1 antibodies.Table 1.The clinical features of the patients in cluster 1 and cluster 2.Cluster 1 (N=26)Cluster 2 (N=34)PAnti-BMPR2 antibodies5(19.2)28(82.4)<0.001Anti-BMPR1A antibodies8(30.2)4(1.8)0.068Anti-ALK antibodies5(19.2)24(70.6)<0.001Arthritis12(46.2)25(73.5)0.031Nephropathy20(76.9)1(2.9)<0.001SLEDAI5.46±4.282.62±2.100.001WHO III/IV17(65.4)14(41.2)0.063Figure 1.(A). Kaplan-Meier analysis of the prognosis of cluster1 and cluster2. (B) The level of Id1 expression under the induction of anti-BMPR2, anti-ALK1, and anti-BMPR1A antibodies in the presence or absence of BMP9. The ID1 (C) and p-Smad 1/5/8 (D) measured by western blotting under the induction of anti-ALK antibodies, BMP9, or anti-ALK antibodies+ BMP9. The fluorescence of the receiver plate well solution (E) and the percentage of late-stage apoptotic cells (E) under the induction of anti-ALK neutralizing antibodies.ConclusionThe serum positivity of autoantibodies targeting BMP signaling has clinical potential in dividing SLE-PAH patients into two distinct clusters. The anti-ALK1 antibodies can downregulate BMP signaling and mediate great permeability and apoptosis in hPAECs, which may be involved in the pathogenesis of SLE-PAH.References[1]Dewachter L, et.al. Eur Respir J 34 (5):1100-1110.[2]Xing Y, et.al. FASEB J 35 (12):e22044.AcknowledgementsWe thank CSTAR co-authors as following for assistance with cases collections.Disclosure of InterestsNone Declared.

BackgroundBaricitinib, an orally selective inhibitor of JAK1 and JAK2, was approved for adult patients with moderate-to-severe rheumatoid arthritis (RA) in China. The recommended dose is 2mg once daily and 4mg once daily in patients who have inadequately responded to baricitinib 2mg once daily (for 3 months) or TNF inhibitors. A single-arm, prospective, non-interventional post-marketing safety study (PMSS) was conducted in Chinese RA patients to describe the safety and effectiveness of baricitinib at 24 weeks.ObjectivesTo describe the safety and effectiveness of baricitinib in real-world setting of treating patients with moderate to severe active RA.MethodsThis PMSS (starting July 2020) included 667 patients with RA treated with baricitinib (2 mg or/and 4mg/day) and followed up for 24 weeks. Safety and effectiveness (disease activity) were assessed for 24 weeks. All statistical analyses are descriptive.ResultsSafety analyses included 667 patients (females=82.3%, mean age=53.3 years, mean RA duration 86.9 months). 106 (15.9%) were ≥65 and <75years and 19 (2.8%) were ≥75 years. 29 (4.3%) had previously received biologic therapy. Baricitinib dose regimen was as follows: 2 mg/day, n = 580 (87.0%); 4 mg/day, n = 53 (7.9%); 2/4 mg, n = 34 (5.1%). Concomitant use of MTX and leflunomide occurred in 54.3% and 35.5%, respectively. The overall exposure of baricitinib was 262.1 patient-years; 197 (29.5%) patients withdrew from the study, mostly for patient’s decision (n = 101). Adverse events (AEs) occurred in 250 (37.5%) patients [serious: 28 (4.2%)]. Two patients (0.3%) died: one of pneumonia and one with no cause reported. The incidence of serious infection, herpes zoster and hepatotoxicity was 0.6%, 1.0%, and 3.4%, respectively. No case met laboratory criteria for potential Hy’s Law (ALT/AST ≥3 x ULN and TBL ≥2 x ULN). Malignancy occurred in one patient (thyroid cancer). No venous thromboembolism (VTE) or major adverse cardiovascular event (MACE) were reported during the study observation period (Table 1).In the effectiveness analysis at Week 24, the proportions of patients achieving remission/ low disease activity were 66.6% (235/353) for DAS28-CRP, 64.6% (228/353) for SDAI, and 63.5% (242/381) for CDAI (Figure 1).ConclusionIn conclusion, the safety and effectiveness profile of baricitinib in this Chinese PMSS was generally similar to that in the global RA population with no VTEs or MACE reported and no new safety signals.ReferencesNIL.Table 1.Safety summary among patients with RA treated with baricitinibn (%) PYE [EAIR]Safety Population (N=667)12 weeks24 weeksAE214 (32.08)124.04 [172.52]250 (37.48)198.58 [125.89]AEs related to study treatmentas judged by the investigator95 (14.24)139.99 [67.86]120 (17.99)236.65 [50.71]Death/2 (0.30)261.30 [0.77]SAE22 (3.30)147.03 [14.96]28 (4.20)256.72 [10.91]SAEs related to study treatmentas judged by the investigator8 (1.20)148.32 [5.39]10 (1.50)260.30 [3.84]Treatment discontinuation due to AEs20 (3.00)148.18 [13.50]24 (3.60)260.79 [9.20]AESISerious infection3 (0.45)148.52 [2.02]4 (0.60)260.99 [1.53]Hepatotoxicity16 (2.40)147.13 [10.87]23 (3.45)256.76 [8.96]VTE0148.63 [0.00]0261.30 [0.00]Herpes zoster, n (%)3 (0.45)7 (1.05)Malignancy, n (%)1 (0.15)1 (0.15)MACE, n (%)00Abbreviations: AE= adverse event; AESI= adverse event with special interest; EAIR= exposure-adjusted incidence rate; MACE= major adverse cardiovascular events; N= number of patients in the safety analysis set; n= number of patients in the specified category; PYE= patient-years of exposure; RA= rheumatoid arthritis; SAE= serious adverse event; VTE= venous thromboembolismUsed EAIR per 100 PYE (patient exposure censored at the event).AESI was based on the judgement of investigator recorded in eCRF (electronic case report form)MACE included myocardial infarction, cardiovascular death, and stroke.Acknowledgements:NIL.Disclosure of InterestsChan-yuan Wu: None declared, Qian Wang: None declared, Jian Shi: None declared, XIUYING ZHANG: None declared, Rong Du: None declared, Jieruo Gu: None declared, Qi-huan Liu: None declared, Jiao Yu Employee of: Eli Lilly and Company, Jia-wei Xu Employee of: Eli Lilly and Company, Yan-jie Zhang Employee of: Eli Lilly and Company, Hao Zhu Employee of: Eli Lilly and Company, Mengtao Li: None declared, Xiaofeng Zeng: None declared.

BackgroundAt present, specific immune targeted therapeutics including biologics and kinase inhibitors has made significant progress in Rheumatoid arthritis (RA). However, the irresponse to targeted therapies in some refractory RA patients suggests more attention should be paid on cell death pathways, such as pyroptosis. TNF-α is a proinflammatory cytokine that plays a fundamental role in the pathogenesis of RA, which could induce pyroptosis in monocytes and macrophages and cause the destruction of bone and cartilage. Iguratimod (IGU) has been confirmed as a highly efficacious and safe conventional synthetic disease-modifying anti-rheumatic drug for RA in Asia. Whether the combination of IGU and Tofacitinib (TOF), the Janus kinase inhibitor, would be better partner need to be elucidated.ObjectivesTo evaluate the therapeutic effect of IGU and TOF on active RA and secondary osteoporosis in collagen-induced arthritis (CIA)+TNF model.MethodsIn this study, hematoxylin and eosin (HE) staining were used to evaluate the pathological changes in ankle joints of CIA+TNF model. Immunohistochemistry (IHC) were used to evaluate the level of pyroptosis related proteins in synovial tissue. We performed Micro-computed tomography (Micro-CT), HE staining and IHC to analyze the trabecular bone changes in distal femoral metaphyses to investigate the destruction of knee joint.ResultsAfter 6 weeks treatment of IGU and/or TOF, the diameter of ankle joint and the level of interleukin (IL)-18, IL-1 of CIA+TNF model. Immunohistochemistry (IHC) were used to evaluate the level of pyroptosis related proteins in synovial tissue. We performed M group. HE staining showed that only a small amount of inflammatory cell infiltration and less pannus were seen in synovial tissue of both monotherapy and combination group, when compared with the CIA+TNF group. Of importance, the pyroptosis related proteins, such as gasdermin D (GSDMD), nucleotide-binding domain (NOD)-like receptor protein 3 (NLRP3), caspase-1, and IL-1β were significantly less expression in synovial tissue of combination group compared with the CIA+TNF group. Both the osteoblast bone formation and osteoclast bone absorption were sharply ruined in CIA+TNF model. However the bone destruction was significantly alleviated and bone turnover rate was remarkably increased in combination group, detected by Micro-CT, HE staining and IHC.ConclusionThe TOF-IGU combination synergistically alleviated the disease severity of the CIA model, including relieving joint inflammation and bone erosion, with suppressing the activation of the NLRP3 inflammasome and reducing GSDMD-mediated pyroptosis. The combined application of TOF and IGU may have a stronger therapeutic effect on RA and secondary osteoporosis bone remodeling.Figure 1.Therapeutic effects of TOF and IGU on the disease severity in CIA+TNF model.(A) The diameter of ankle joint; (B) HE staining in synovial tissue; (C) IHC staining of pyroptosis related proteins in synovial tissue; (D) Micro-CT analysis in distal femurs; (E) HE staining in knee joint; (F) IHC staining of ALP and TRAP staining in knee joint.Acknowledgementsthe ECCM Program of Clinical Research Center of Shandong University (No. 2021SDUCRCB010) and the Natural Science Foundation of Shandong Province (ZR2022MH177)Disclosure of InterestsNone Declared.

BackgroundPulmonary arterial hypertension (PAH) is one of the most important complications that seriously threatens the prognosis of patients with systemic lupus erythematosus (SLE), with complicated and unclear pathogenesis.ObjectivesBased on genomic studies and functional experiments, we aim to investigate candidate biomarkers and targeted therapy for the early diagnosis and timely treatment of SLE-PAH patients.Methods:1) In order to screen susceptible genes of SLE-PAH, a number of 150 peripheral blood from SLE-PAH patients were subject to whole-exome sequencing (WES), and genome-wide association study (GWAS) was performed by comparing with 934 healthy controls.2) The transcriptional expression levels on peripheral blood of SLE-PAH patients were examined by RT-qPCR to further evaluate the possible pathogenesis of the above screened genes.3) Intervention experiments on human pulmonary artery endothelial cells (hPAEC) were performed to figure out the potential pathogenesis of the selected gene in vitro. RNA-seq and gene ontology were applied to identify the downstream pathways.4) Established by pristane injection and hypoxia induction, SLE-PAH mice model was used to evaluate the pathogenicity and therapeutic value of selected gene. Pulmonary arterial pressure (PAP) was measured by right heart catheterization after tail-intravenous injection of therapeutic vectors.Results:1) The tumor necrosis factor receptor-associated factor 5 (TRAF5) was identified as a susceptible gene of SLE-PAH based on WES and GWAS.2) The significant reductions of TRAF5 on transcriptional level in peripheral blood of SLE-PAH patients were identified, indicating clinical diagnosis values.3) Knockdown of TRAF5 significantly increased early apoptosis of hPAEC and triggered the pathogenesis of PAH through distinct pathways.4) SLE-PAH mouse model was successfully established since they showed lupus phenotype and the mean PAPs were measured as over 40mmHg. Tail-intravenous injection of TRAF5-overexpression vector attenuated PAH.ConclusionLack of TRAF5 triggers the pathogenesis of PAH in SLE patients through inducing hPAEC abnormality. It is a susceptible gene of SLE-PAH and could be a candidate marker for diagnosis and therapy for SLE-PAH patients.Figure 1.A) Genomic and protein simulation structure TRAF5 (a susceptible gene of SLE-PAH). Red dots represent mutation sites that screened from SLE-PAH patients. P.G468R mutation causes dysfunction of protein. B) TRAF5 mRNA expression levels in PBMC of healthy-controls, SLE-PAH and SLE-nPAH patients. *p<0.05. C) shTRAF5 transfected human PAEC. The transfection efficiency reaches 80% (The percentage of EGFP positive cells in all cells of bright field) when MOI=20. All three knockdown vectors of shTRAF5 showed significantly down regulation of TRAF5. *p<0.05, **p<0.01. D) FACS was performed to detect early apoptotic cells labeled with Annexin V-APC (Apoptosis Detection Kit). The group of shTRAF5 showed significantly increased apoptosis compared with groups of control/shScramble. *p<0.05 E) Wound healing experiments were performed in different groups, and the distances of scratches at the same area in each group were measured at the time point of 0h, 6h, 24h, 48h. Abnormal migration was observed in shTRAF5 transfected PAECs.REFERENCES:NIL.Acknowledgements:NIL.Disclosure of InterestsNone Declared.