{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T13:01:11Z","timestamp":1791896471134,"version":"4.3.4"},"posted":{"date-parts":[[2025,11,4]]},"group-title":"Cancer Biology","reference-count":67,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,11,4]],"date-time":"2025-11-04T00:00:00Z","timestamp":1762214400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"funder":[{"award":["R15CA167476"],"award-info":[{"award-number":["R15CA167476"]}],"id":[{"id":"https:\/\/ror.org\/040gcmg81","id-type":"ROR","asserted-by":"publisher"}]},{"award":["8P20 GM103424-21"],"award-info":[{"award-number":["8P20 GM103424-21"]}],"id":[{"id":"https:\/\/ror.org\/04q48ey07","id-type":"ROR","asserted-by":"publisher"}]},{"award":["1038602"],"award-info":[{"award-number":["1038602"]}],"id":[{"id":"https:\/\/ror.org\/00sda2672","id-type":"ROR","asserted-by":"publisher"}]},{"award":["1032532"],"award-info":[{"award-number":["1032532"]}],"id":[{"id":"https:\/\/ror.org\/00sda2672","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,11,4]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Glucosylceramide synthase (GCS) catalyzes ceramide glycosylation in response to cell stress that produces glucosylceramide and other glycosphingolipids. GCS overexpression is a cause of drug resistance and enriches cancer stem cells (CSCs) during cancer chemotherapy. Previous studies showed that GCS modulates expression of p53 mutants and oncogenic gain-of-function (GOF) in heterozygous knock-in cell models (\n                  <jats:italic>TP53<\/jats:italic>\n                  R273H\n                  <jats:sup>-\/+<\/jats:sup>\n                  ). However, it is unclear whether GCS can modulate the effects of homozygous p53 mutations, which are common in many cancer cases. We report herewith that inhibition of GCS, via UGCG-knockout and using new inhibitor (Genz-161), effectively re-sensitizes drug resistance and diminishes CSCs in colon cancer cells carrying the homozygous p53 R273H mutation. In aggressive WiDr cells carrying\n                  <jats:italic>TP53<\/jats:italic>\n                  R273H mutation, knockout of\n                  <jats:italic>UGCG<\/jats:italic>\n                  gene using CRISPR\/Cas9 editing or inhibition of GCS with Genz-161 sensitized cancer cells to oxaliplatin, irinotecan and paclitaxel. With decreased ceramide glycosylation in lipidomic profiling, both UGCG-knockout and Genz-161 treatments substantially decreased wound healing, and diminished CSCs and tumor growth under chemotherapy. Interestingly, inhibition of RNA m\n                  <jats:sup>6<\/jats:sup>\n                  A methylation by neplanocin A reactivated p53 function and reversed drug resistance. Mechanistic investigation revealed that GCS inhibition downregulated METTL3 expression and repressed RNA-m\n                  <jats:sup>6<\/jats:sup>\n                  A modification on mutant p53 R273H effects. Altogether, our findings demonstrate that ceramide glycosylation promotes METTL3 expression and RNA m\n                  <jats:sup>6<\/jats:sup>\n                  A methylation in response to drug-induced stress, thereby promoting mutant p53 expression and associated GOF. Conversely, inhibition of GCS can diminish CSCs and drug resistance via reduction of m\n                  <jats:sup>6<\/jats:sup>\n                  A modification and reactivation of p53 function. GCS inhibition is an achievable approach for mutant cancer treatment.\n                <\/jats:p>","DOI":"10.1101\/2025.11.02.686136","type":"posted-content","created":{"date-parts":[[2025,11,4]],"date-time":"2025-11-04T14:15:24Z","timestamp":1762265724000},"source":"Crossref","is-referenced-by-count":1,"title":["Suppression of Glucosylceramide Synthase Reverses Drug Resistance in Cancer Cells Harbor Homozygous p53 Mutants"],"prefix":"10.64898","author":[{"given":"Md Saqline","family":"Mostaq","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mohammad N","family":"Amin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amanda","family":"Raphael","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Celine","family":"Asbury","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anish","family":"Gupta","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xin","family":"Gu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xianlin","family":"Han","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Davorka","family":"Sekulic","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pawel","family":"Michalak","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lin","family":"Kang","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7968-0162","authenticated-orcid":false,"given":"Yong-Yu","family":"Liu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2025110608300857000_2025.11.02.686136v1.1","doi-asserted-by":"publisher","DOI":"10.3389\/fonc.2021.807636"},{"key":"2025110608300857000_2025.11.02.686136v1.2","doi-asserted-by":"publisher","DOI":"10.1016\/B978-0-12-394274-6.00003-0"},{"key":"2025110608300857000_2025.11.02.686136v1.3","doi-asserted-by":"crossref","unstructured":"Russo D , Capolupo L , Loomba JS , Sticco L , D\u2019Angelo G: Glycosphingolipid metabolism in cell fate specification. 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