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We integrated differential expression analysis, weighted gene co-expression network analysis (WGCNA), and protein\u2013protein interaction (PPI) network pharmacology to identify CRC hub genes and their ion channel connections, validated by dual single-cell perturbation approaches: variational graph autoencoder-based virtual knockout (VGAE-KO) and experimental HCT116 CRISPRi Perturb-seq (6 genes, 8445 cells). WGCNA identified 100 hub genes spanning three functional programs. Ribosomal proteins link to K+ channels (RPS21 \u2192 KCNQ2, targetable by EMA-approved ataluren, passed dual validation at 97.8th\u201398.7th percentile). RNA processing genes connect to Cl\u2212 channels (LSM7 \u2192 CLIC1, strongest signal at 99.8th\u201399.4th percentile). Immune checkpoint receptors (LAG3, CD27) connect via PPI intermediates to Ca2+ and K+ channels, targetable by relatlimab (FDA-approved) and varlilumab (Phase 2). This work maps previously unknown links between CRC driver genes and ion channel regulation, with the ataluren-RPS21-KCNQ2 axis ready for pharmacological testing.<\/jats:p>","DOI":"10.3390\/ijms27083412","type":"journal-article","created":{"date-parts":[[2026,4,10]],"date-time":"2026-04-10T12:01:52Z","timestamp":1775822512000},"page":"3412","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Ion-Channel-Mediated Drug Repurposing Opportunities Validated by Single-Cell Perturbation in Colorectal Cancer"],"prefix":"10.3390","volume":"27","author":[{"ORCID":"https:\/\/orcid.org\/0009-0006-4541-4524","authenticated-orcid":false,"given":"Zhongyuan","family":"Dong","sequence":"first","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Naval Medical University, Shanghai 200433, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9968-2069","authenticated-orcid":false,"given":"Xuanlin","family":"Meng","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Naval Medical University, Shanghai 200433, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8354-5895","authenticated-orcid":false,"given":"Lianghua","family":"Wang","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Naval Medical University, Shanghai 200433, China"},{"name":"Key Laboratory of Biosafety Defense, Naval Medical University, Ministry of Education, Shanghai 200433, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,10]]},"reference":[{"key":"ref_1","first-page":"209","article-title":"Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries","volume":"71","author":"Sung","year":"2021","journal-title":"CA Cancer J. 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