Cathepsin Levels and Atrial Fibrillation Risk: Insights From Bidirectional and Multivariable Mendelian Randomization Analyses.

Background: Atrial fibrillation (AF) is the most common cardiac arrhythmia, contributing to substantial morbidity, mortality, and healthcare burden. While genome-wide association studies (GWAS) have identified numerous genetic variants linked to AF risk, the causal roles of proteolytic enzymes such as cathepsins remain poorly defined. This study employed bidirectional and multivariable Mendelian randomization (MR) approaches to investigate the causal relationship between genetically determined cathepsin levels and AF risk. Methods: Genetic instruments for nine cathepsins were derived from the INTERVAL study (n = 3301, European ancestry), using a significance threshold of p < 5 × 10-6 and stringent LD pruning (r 2 < 0.001, 10,000 kb window). Only SNPs with F-statistics > 10 were retained. AF outcome data were obtained from a GWAS meta-analysis comprising 60,620 cases and 970,216 controls of European descent. Two-sample MR analyses were conducted using the inverse variance weighted (IVW) method, supported by MR-Egger and weighted median approaches. Multivariable MR was used to adjust for correlated cathepsins, and reverse MR assessed bidirectional causality. Results: Genetically elevated cathepsin O levels were significantly associated with increased AF risk (IVW: p = 0.0025, OR = 1.06, 95% CI 1.02-1.10), and this association remained robust in multivariable MR (IVW: p = 0.0265, OR = 1.0571, 95% CI 1.0065-1.1102). A suggestive association for cathepsin B was observed only in multivariable MR (IVW: p = 0.0356, OR = 1.0279, 95% CI 1.0018-1.0547), but did not survive multiple testing correction. Sensitivity analyses supported the validity of these findings, and reverse MR showed no evidence of reverse causation. Conclusion: This study provides genetic evidence that elevated cathepsin O levels-and conditionally cathepsin B-are causally linked to increased AF risk. These findings highlight the potential role of proteolytic enzymes in AF pathogenesis and suggest novel therapeutic targets. All analyses were conducted in European-ancestry populations; replication in diverse cohorts and mechanistic studies are warranted to validate and extend these insights.