Developing alternative targets and drugs for rheumatoid arthritis (RA) treatment is currently an urgent issue. The relationship between TGM2 and the abnormal immune microenvironment in synovium tissues, as well as the specific role of TGM2 in RA are yet to be elucidated. Sarsasapogenin (Sar) is a sapogenin extracted from the Chinese medical herb Anemarrhena asphodeloides Bunge. and served as a representative anti-inflammatory drug capable of ameliorating inflammatory responses in several human diseases. However, the therapeutic effect of Sar on RA remains unknown.This investigation aims to elucidate the role of TGM2 in RA and investigate whether Sar is a candidate drug to target TGM2 of fibroblast-like synoviocytes (FLS).Bioinformatics analyses were applied for elucidating the role of N(6)-methyladenine (m6A) RNA methylation in RA and identifying the specific target regulated by m6A methylation in RA-FLS. Methylated RNA immunoprecipitation, CCK8 assay, Edu assay, flow cytometry, RT-qPCR and Western blot were utilized to investigate the function of Sar and TGM2 in RA-FLS.Bioinformatics analyses emphasized the importance of m6A RNA methylation in RA and identified an m6A methylation-mediated gene TGM2. Interestingly, both m6A RNA methylation and TGM2 expression in RA synovium tissues correlated with activated immuno-inflammatory phenotype and associated with clinical characteristics and therapy response of RA patients. TGM2 served as a promoter of RA-FLS proliferation by inducing DNA replication and cell cycle transition and inhibiting apoptosis through activating NF-κB signaling. Intriguingly, Sar could impair m6A methylation of TGM2 mRNA and downregulate TGM2 expression. Downregulated TGM2 contributed to the suppressive role of Sar in DNA replication and the stimulatory role of Sar in cell cycle arrest and apoptosis of RA-FLS. Mechanically, Sar inhibited the expression of key regulators in DNA replication, cell cycle, and apoptosis by impairing NF-κB signaling, thus abolishing FLS proliferation to ameliorate RA progression.This cross-validated work based on three independent datasets is detailedly delineated using cell lines and clinical samples, recognizing that TGM2 can be an attractive target and Sar might be a novel anti-RA drug.Copyright © 2021 Elsevier GmbH. All rights reserved.