<p>Cardiac fibrosis is a critical pathological feature in nearly all forms of heart disease and contributes to heart failure. However, existing diagnostic and therapeutic approaches are insufficient for detecting fibrosis or targeting conventional pathways. Further molecular investigations are urgently needed to develop effective treatments. Circular RNAs (circRNAs), distinguished by their stability and regulatory roles, show significant potential. CircRNA_006640 was substantially upregulated in left ventricular tissues of male mice subjected to transverse aortic constriction, as well as in blood samples. Gain- and loss-of-function studies confirmed that circRNA_006640 promoted proliferation and phenotypic transformation in mouse cardiac fibroblasts. Real-time quantitative reverse transcription polymerase chain reaction and Western blotting demonstrated that circRNA_006640 suppressed miR-7648-3p and miR-185-3p, which in turn inhibited connective tissue growth factor (CTGF). Furthermore, there was a synergistic effect between miR-7648-3p and miR-185-3p to enhance the suppression of CTGF. In vivo, circRNA_006640 markedly exacerbated cardiac fibrosis. Knockdown of circRNA_006640 using small interfering RNA effectively mitigated cardiac fibrosis and preserved cardiac function, indicating therapeutic potential for antifibrotic strategies. CircRNA_006640 acts as a novel upstream regulator of CTGF, exacerbating cardiac fibrosis by sponging miR-7648-3p and miR-185-3p. The synergistic interaction between miR-7648-3p and miR-185-3p strengthens the profibrotic effect. Targeting CircRNA_006640 may hold promising therapeutic potential for cardiac fibrosis.</p> Graphical Abstract <p></p>