CRISPR-Cas9 induced formation of Myc-containing ecDNA drives carcinogenesis in mouse liver and cancer phenotypes in human cell models

CRISPR-Cas9 induced formation of Myc-containing ecDNA drives carcinogenesis in mouse liver and cancer phenotypes in human cell models | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article CRISPR-Cas9 induced formation of Myc-containing ecDNA drives carcinogenesis in mouse liver and cancer phenotypes in human cell models Birgitte Regenberg, Weijia Feng, Yiyuan Niu, Egija Zole, Annette Snejbjerg Bartels, and 11 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9185034/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Over 40% of human cancers harbor extrachromosomal DNA (ecDNA), which correlates with therapy resistance and poor prognosis. Yet research is limited by a lack of robust de novo ecDNA models. We present a CRISPR-Cas9 based approach for generation of ecDNA in cell lines and mouse liver. In HEK293 cells, CRISPR-induced MYC-bearing ecDNA ([MYCcircle]) promoted cancer hallmarks, including enhanced proliferation, increased migration, and reduced apoptosis. To model ecDNA in mice, we combined Cas9 knock in mice with lipid-nanoparticle delivery of sgRNAs in the liver targeting the Myc–Pvt1 locus. When combined with Trp53 or Pten loss, the resultant [Myccircle] caused hepatocellular carcinomas with amplified levels of [Myccircle] and elevated, heterogenous Myc expression, supporting evidence of ecDNA involvement in tumorigenesis. Our approach offers a fast and versatile tool, where target-sequence flexibility allows rapid generation of animal and cell models with any investigator-specified ecDNA, thereby enabling study of the direct role of ecDNA in cancer progression. Biological sciences/Genetics/Cancer genetics Biological sciences/Genetics/Genome Biological sciences/Biological techniques/Genetic engineering Biological sciences/Cancer/Tumour heterogeneity Full Text Additional Declarations Yes there is potential Competing Interest. Birgitte Regenberg is co-founder of CARE-DNA IP Aps (CVR number: 45303667, https://care-dna.eu/ ). Egija Zole is employed by CARE-DNA IP Aps. Morten Frödin is co-founder of BioPhenyx Aps. Supplementary Files SupplementaryTable1.xlsx Primers and sgRNA sequences SupplementaryTable2.xlsx Treatment groups SupplementaryTable3CharacteristicsofsgRNALNPs.docx Physicochemical properties of sgRNA-LNPs dispersed in D-PBS, pH 7.4. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9185034","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":610793177,"identity":"e60d0fef-2970-4164-b129-55bedcd13cd7","order_by":0,"name":"Birgitte 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