An effective method based on targeted deep sequencing to simultaneously estimate mosaic loss of Y chromosome and CHIP-related mutations

Clonal hematopoiesis (CH), prevalent in the elderly, is strongly associated with age-related diseases. Somatic mutations in driver genes and mosaic loss of Y chromosome (mLOY) represent the two major forms of CH in male populations. While capture-based deep sequencing has been widely used for detecting somatic mutations and droplet digital PCR (ddPCR) for mLOY, these methods suffer from limited sensitivity to low-level mLOY and lack the ability to concurrently assess mLOY and point mutations. Here, we present a novel strategy based on targeted deep sequencing to simultaneously estimate mLOY and CHIP-related mutations. First, we validated the reliability of mLOY estimation based on UTY and KDM5D coverage using both whole-genome sequencing and targeted sequencing data. In a cohort of 188 male patients with acute aortic syndrome (AAS), comparative analysis with ddPCR showed that our method exhibited exceptional reliability for detecting high-level mLOY (defined as > 20%), with an AUC of 0.999 (P<0.001). The combined detection of mLOY >20% achieved optimal performance: sensitivity of 87.5%, specificity of 100%, positive predictive value of 100%, and negative predictive value of 99.4%. Moreover, this strategy has been verified in an independent cohort of 240 male patients with acute coronary syndrome (ACS), which strengthens confidence in the method’s performance across different disease settings. We also explored the association between mLOY and CHIP-related mutations. The results revealed a significant trend: individuals with higher mLOY frequencies were more likely to harbor CHIP mutations. Notably, the co-occurrence of CHIP mutations and mLOY was more pronounced when detected with our method, highlighting that targeted deep sequencing is better suited for dissecting the complex interplay of genomic instabilities during aging. Collectively, this cost-effective strategy enables simultaneous detection of mLOY and CHIP mutations, providing a robust tool for investigating CH in age-related disease research.