Integrated fragmentomic profile and 5-Hydroxymethylcytosine of capture-based low-pass sequencing data enables pan-cancer detection via cfDNA

Using epigenetic markers and fragmentomics of cell-free DNA for cancer detection has been proven applicable. We further combine the two features and explore the diagnostic potential of the features on pan-cancer detection. We extracted cfDNA fragmentomic features from 191 whole-genome sequencing data and investigated them in 396 low-pass 5hmC sequencing data from four common cancer types and controls. We identified aberrant ultra-long fragments (220-500bp) of cancer samples in 5hmC sequencing data, both in size and coverage profile, and showed its dominant role in cancer prediction. Since cfDNA hydroxymethylation and fragmentomic markers can be detected simultaneously in low-pass 5hmC sequencing data, we built an integrated model including 63 features of both fragmentomic features and hydroxymethylation signatures for pan-cancer detection with high sensitivity and specificity (88.52% and 82.35%, respectively). We showed that fragmentomic information in 5hmC sequencing data is an ideal marker for cancer detection and that it shows high performance in low-pass sequencing data. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This study was supported by the 1·3·5project for disciplines of excellence, West China Hospital, Sichuan University (ZYYC20006) to D. Xie; Thousand Talents Program of the West China Hospital (0040205401F58) to D. Xie; Sichuan Provincial Foundation of Science and Technology (2020YFS0051) to D. Xie, (2017SZ0006) to Y. Liu; Clinical Research Innovation Project, West China Hospital, Sichuan University (19HXCX009) to Y. Liu; the San Hang Program of the Second Military Medical University, Medical basic research project of the First Affiliated Hospital, the Second Military Medical University (2021JCMS16) to W. Zhang; the Science and technology project of Sichuan Province (2021YFS0109) to A. Li; Post–Doctor Research Project, West China Hospital, Sichuan University (20HXBH035) to S. Zhang; the high quality development of Guang’an People’s Hospital (21FZ003) to A. Wei. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: This study was approved by the Ethics Committee of Sichuan Cancer Hospital (SCCHEC-02-2016-005). The written informed consent was obtained from all participants. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes The 5hmC sequencing data for controls, LUAD, HCC and PDAC were publicly available as described in the Method section. The 5hmC sequencing data for GBM can be accessed at