{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,1]],"date-time":"2026-09-01T14:15:27Z","timestamp":1788272127668,"version":"build-2803163510"},"reference-count":52,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,9,1]],"date-time":"2026-09-01T00:00:00Z","timestamp":1788220800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003196","name":"Ministero della Salute","doi-asserted-by":"publisher","award":["RC 15\/23"],"award-info":[{"award-number":["RC 15\/23"]}],"id":[{"id":"10.13039\/501100003196","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003196","name":"Ministero della Salute","doi-asserted-by":"publisher","award":["RC 23\/23"],"award-info":[{"award-number":["RC 23\/23"]}],"id":[{"id":"10.13039\/501100003196","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Pharmacol."],"abstract":"<jats:sec>\n                    <jats:title>Introduction<\/jats:title>\n                    <jats:p>Mercaptopurine (MP) is used to treat pediatric acute lymphoblastic leukemia and inflammatory bowel disease (IBD). The Protein Kinase C and Casein Kinase Substrate in Neurons 2 (PACSIN2) rs2413739 polymorphism is an expression-quantitative trait locus associated with reduced PACSIN2 expression in gastrointestinal tissues and thiopurine-related gastrointestinal toxicity. PACSIN2 interacts with Rac1, a key thiopurine target protein involved in cytoskeletal regulation.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>LS180 intestinal cells with stable PACSIN2 knockdown (KD) or transient overexpression were used to assess the role of PACSIN2 in regulating Rac1 under basal conditions and after MP exposure. Rac1 levels and activity were measured by Western blot and G-LISA assay, respectively. Cytotoxicity was evaluated by MTT assay following exposure to MP and the Rac1 inhibitor NSC23766. Cell stiffness was assessed by atomic force microscopy, and cytoskeletal organization was examined by immunofluorescence.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>PACSIN2 KD cells exhibited elevated Rac1 expression and activity, and increased sensitivity to MP, which was further augmented by the Rac1 inhibitor NSC23766. Conversely, PACSIN2 overexpression reduced Rac1 levels and conferred MP resistance. PACSIN2 depletion decreased cell stiffness affecting the cytoskeletal architecture, particularly impairing tubulin levels and actin distribution. MP exposure reduced cell stiffness and decreased both Rac1 expression and activity across all tested cell lines, with more pronounced effects in LS180 KD cells. Altered cytoskeletal organization, especially actin redistribution, correlated with changes in cellular biomechanical properties and drug sensitivity.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>These findings suggest that PACSIN2 modulates MP response by regulating Rac1 levels and cytoskeletal integrity, providing a molecular basis for the clinical association between PACSIN2 and thiopurine-related gastrointestinal toxicity. These insights may contribute to the development of personalized approaches to optimize thiopurine therapy in pediatric patients.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.3389\/fphar.2026.1860702","type":"journal-article","created":{"date-parts":[[2026,9,1]],"date-time":"2026-09-01T13:30:41Z","timestamp":1788269441000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["PACSIN2 regulates mercaptopurine cytotoxicity and cellular biomechanics through Rac1 modulation in intestinal epithelial cells"],"prefix":"10.3389","volume":"17","author":[{"given":"Alessia","family":"Norbedo","sequence":"first","affiliation":[{"name":"Department of Medicine Surgery and Health Sciences, University of Trieste","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Giulia","family":"Zudeh","sequence":"additional","affiliation":[{"name":"Department of Translational and Advanced Diagnostics, Institute for Maternal and Child Health I.R.C.C.S. Burlo Garofolo","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Donatella","family":"Fiducioso","sequence":"additional","affiliation":[{"name":"Department of Life Sciences, University of Trieste","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Beatrice","family":"Senigagliesi","sequence":"additional","affiliation":[{"name":"Joint Research Center","place":["Ispra, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pietro","family":"Parisse","sequence":"additional","affiliation":[{"name":"Istituto Officina dei Materiali, Consiglio Nazionale delle Ricerche (CNR - IOM)","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Loredana","family":"Casalis","sequence":"additional","affiliation":[{"name":"Elettra Sincrotrone Trieste","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Giuliana","family":"Decorti","sequence":"additional","affiliation":[{"name":"Department of Medicine Surgery and Health Sciences, University of Trieste","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marianna","family":"Lucaf\u00f2","sequence":"additional","affiliation":[{"name":"Department of Life Sciences, University of Trieste","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gabriele","family":"Stocco","sequence":"additional","affiliation":[{"name":"Department of Medicine Surgery and Health Sciences, University of Trieste","place":["Trieste, Italy"]},{"name":"Department of Translational and Advanced Diagnostics, Institute for Maternal and Child Health I.R.C.C.S. Burlo Garofolo","place":["Trieste, Italy"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,9,1]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"1165","DOI":"10.18632\/oncotarget.27974","article-title":"Dynamic cellular biomechanics in responses to chemotherapeutic drug in hypoxia probed by atomic force spectroscopy","volume":"12","author":"Alhalhooly","year":"2021","journal-title":"Oncotarget"},{"key":"B2","doi-asserted-by":"publisher","first-page":"3379","DOI":"10.3390\/cancers15133379","article-title":"The tumor suppressor DAB2IP is regulated by cell contact and contributes to YAP\/TAZ inhibition in confluent cells","volume":"15","author":"Apollonio","year":"2023","journal-title":"Cancers (Basel)"},{"key":"B3","doi-asserted-by":"publisher","first-page":"80","DOI":"10.1182\/blood-2014-07-58760","article-title":"FlnA binding to PACSIN2 F-BAR domain regulates membrane tubulation in megakaryocytes and platelets","volume":"126","author":"Begonja","year":"2015","journal-title":"Blood"},{"key":"B4","doi-asserted-by":"publisher","first-page":"1864","DOI":"10.1038\/s41467-021-22126-y","article-title":"Quantitative characterization of extracellular vesicle uptake and content delivery within Mammalian cells","volume":"12","author":"Bonsergent","year":"2021","journal-title":"Nat. Commun."},{"key":"B5","doi-asserted-by":"publisher","first-page":"e0166715","DOI":"10.1371\/journal.pone.0166715","article-title":"Characterization of novel molecular mechanisms favoring Rac1 membrane translocation","volume":"11","author":"Castro-Castro","year":"2016","journal-title":"PLoS One"},{"key":"B6","doi-asserted-by":"publisher","first-page":"2375","DOI":"10.1242\/jcs.08630","article-title":"The F-BAR domain protein PACSIN2 associates with Rac1 regulates cell spreading and migration","volume":"124","author":"de Kreuk","year":"2011","journal-title":"J. Cell Sci."},{"key":"B8","doi-asserted-by":"publisher","first-page":"45575","DOI":"10.1038\/srep45575","article-title":"On the determination of elastic moduli of cells by AFM based indentation","volume":"7","author":"Ding","year":"2017","journal-title":"Sci. Rep."},{"key":"B9","doi-asserted-by":"publisher","first-page":"e13783","DOI":"10.1111\/apha.13783","article-title":"PACSIN proteins in vivo: roles in development and physiology","volume":"234","author":"Dumont","year":"2022","journal-title":"Acta Physiol. (Oxf)"},{"key":"B10","doi-asserted-by":"publisher","first-page":"256","DOI":"10.20517\/cdr.2019.004","article-title":"Pharmacogenetics of thiopurines","volume":"2","author":"Franca","year":"2019","journal-title":"Cancer Drug Resist"},{"key":"B11","doi-asserted-by":"publisher","first-page":"415","DOI":"10.1038\/s41397-019-0130-0","article-title":"PACSIN2 rs2413739 influence on thiopurine pharmacokinetics: validation studies in pediatric patients","volume":"20","author":"Franca","year":"2020","journal-title":"Pharmacogenomics J."},{"key":"B12","doi-asserted-by":"publisher","first-page":"301","DOI":"10.1136\/flgastro-2015-100681","article-title":"Monitoring thiopurine metabolites in inflammatory bowel disease","volume":"7","author":"Gonz\u00e1lez-Lama","year":"2016","journal-title":"Frontline Gastroenterol"},{"key":"B13","doi-asserted-by":"publisher","first-page":"1991","DOI":"10.1016\/j.yexcr.2008.03.015","article-title":"PACSIN proteins bind tubulin and promote microtubule assembly","volume":"314","author":"Grimm-G\u00fcnter","year":"2008","journal-title":"Exp. Cell Res."},{"key":"B14","doi-asserted-by":"publisher","first-page":"580","DOI":"10.1038\/ng.2653","article-title":"The genotype-tissue expression (GTEx) project","volume":"45","year":"2013","journal-title":"Nat. Genet."},{"key":"B15","doi-asserted-by":"publisher","first-page":"2024.03.04.583337","DOI":"10.1101\/2024.03.04.583337","article-title":"Quantitative superresolution imaging of F-Actin in the cell body and cytoskeletal protrusions using phalloidin-based single-molecule labeling and localization microscopy","volume":"2024","author":"Gunasekara","year":"2024","journal-title":"bioRxiv"},{"key":"B16","doi-asserted-by":"publisher","first-page":"220607","DOI":"10.1098\/rsos.220607","article-title":"Changes in the mechanical properties of human mesenchymal stem cells during differentiation","volume":"10","author":"He","year":"2023","journal-title":"R. Soc. Open Sci."},{"key":"B17","doi-asserted-by":"publisher","first-page":"G1137","DOI":"10.1152\/ajpgi.00031.2006","article-title":"Rac1 mediates intestinal epithelial cell apoptosis via JNK","volume":"291","author":"Jin","year":"2006","journal-title":"Am. J. Physiol. Gastrointest. Liver Physiol."},{"key":"B18","doi-asserted-by":"publisher","first-page":"5","DOI":"10.1186\/s40580-017-0099-9","article-title":"Mechanical properties of paraformaldehyde-treated individual cells investigated by atomic force microscopy and scanning ion conductance microscopy","volume":"4","author":"Kim","year":"2017","journal-title":"Nano Converg."},{"key":"B19","doi-asserted-by":"publisher","first-page":"1154","DOI":"10.15252\/embr.201439267","article-title":"Direct interaction of actin filaments with F-BAR protein pacsin2","volume":"15","author":"Kostan","year":"2014","journal-title":"EMBO Rep."},{"key":"B20","doi-asserted-by":"publisher","first-page":"8786","DOI":"10.3390\/ijms21228786","article-title":"Nanomechanics in monitoring the effectiveness of drugs targeting the cancer cell cytoskeleton","volume":"21","author":"Kubiak","year":"2020","journal-title":"Int. J. Mol. Sci."},{"key":"B21","doi-asserted-by":"publisher","first-page":"7675","DOI":"10.1038\/s41598-019-43975-0","article-title":"The small GTPase Rac1 increases cell surface stiffness and enhances 3D migration into extracellular matrices","volume":"9","author":"Kunschmann","year":"2019","journal-title":"Sci. Rep."},{"key":"B22","first-page":"1326","article-title":"Atomic force microscopy-based single-cell indentation: experimentation and finite element simulation","volume-title":"IEEE\/RSJ International Conference on Intelligent Robots and Systems","author":"Ladjal","year":"2009"},{"key":"B23","doi-asserted-by":"publisher","first-page":"447","DOI":"10.1083\/jcb.201612069","article-title":"Rho GTPase signaling complexes in cell migration and invasion","volume":"217","author":"Lawson","year":"2018","journal-title":"J. Cell Biol."},{"key":"B24","doi-asserted-by":"publisher","first-page":"72","DOI":"10.1186\/s12915-021-00997-3","article-title":"Rac1 activation can generate untemplated, lamellar membrane ruffles","volume":"19","author":"Leyden","year":"2021","journal-title":"BMC Biol."},{"key":"B25","first-page":"551","volume-title":"Effects of Methotrexate on the Viscoelastic Properties of Single Cells Probed by Atomic Force Microscopy. J. Biol. Phys.","author":"Li","year":"2016"},{"key":"B26","doi-asserted-by":"publisher","first-page":"2979","DOI":"10.1007\/s00204-019-02549-9","article-title":"Biophysical and biomechanical properties of neural progenitor cells as indicators of developmental neurotoxicity","volume":"93","author":"Mahajan","year":"2019","journal-title":"Arch. Toxicol."},{"key":"B27","doi-asserted-by":"publisher","first-page":"275","DOI":"10.1136\/gutjnl-2021-325520","article-title":"Epithelial RAC1-dependent cytoskeleton dynamics controls cell mechanics, cell shedding and barrier integrity in intestinal inflammation","volume":"72","author":"Mart\u00ednez-S\u00e1nchez","year":"2023","journal-title":"Gut"},{"key":"B28","doi-asserted-by":"publisher","first-page":"475","DOI":"10.3389\/fcell.2020.00475","article-title":"Editorial: biomechanical properties of cells and tissues and their impact on cellular adhesion and motility","volume":"8","author":"Mierke","year":"2020","journal-title":"Front. Cell Dev. Biol."},{"key":"B29","doi-asserted-by":"publisher","first-page":"14","DOI":"10.1016\/j.bbrc.2021.05.083","article-title":"Chemical inhibitor anticancer drugs regulate mechanical properties and cytoskeletal structure of non-invasive and invasive breast cancer cell lines: study of effects of letrozole, exemestane, and everolimus","volume":"565","author":"Mohammadi","year":"2021","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"B30","doi-asserted-by":"publisher","first-page":"458","DOI":"10.1016\/j.carbon.2021.01.142","article-title":"Shedding plasma membrane vesicles induced by graphene oxide nanoflakes in brain cultured astrocytes","volume":"176","author":"Musto","year":"2021","journal-title":"Carbon"},{"key":"B31","doi-asserted-by":"publisher","first-page":"316","DOI":"10.1080\/21541248.2016.1224399","article-title":"Rac1 and RhoA: networks, loops and bistability","volume":"9","author":"Nguyen","year":"2016","journal-title":"Small GTPases"},{"key":"B32","doi-asserted-by":"publisher","first-page":"189","DOI":"10.1016\/j.cbi.2017.08.009","article-title":"Pharmacokinetics and pharmacodynamics of thiopurines in an in vitro model of human hepatocytes: insights from an innovative mass spectrometry assay","volume":"275","author":"Pelin","year":"2017","journal-title":"Chem. Biol. Interact."},{"key":"B33","doi-asserted-by":"publisher","first-page":"640","DOI":"10.4049\/jimmunol.176.1.640","article-title":"Azathioprine suppresses ezrin-radixin-moesin-dependent T Cell-APC conjugation through inhibition of vav guanosine exchange activity on rac proteins","volume":"176","author":"Poppe","year":"2006","journal-title":"J. Immunology Baltim. Md"},{"key":"B34","doi-asserted-by":"publisher","first-page":"2515","DOI":"10.1091\/mbc.E19-06-0352","article-title":"PACSIN2-dependent apical endocytosis regulates the morphology of epithelial microvilli","volume":"30","author":"Postema","year":"2019","journal-title":"Mol. Biol. Cell"},{"key":"B35","doi-asserted-by":"publisher","first-page":"5198","DOI":"10.1111\/febs.12343","article-title":"Syndapin--a membrane remodelling and endocytic F-BAR protein","volume":"280","author":"Quan","year":"2013","journal-title":"FEBS J."},{"key":"B36","doi-asserted-by":"publisher","first-page":"13039","DOI":"10.1074\/jbc.M208741200","article-title":"The requirement for polyamines for intestinal epithelial cell migration is mediated through Rac1","volume":"278","author":"Ray","year":"2003","journal-title":"J. Biol. Chem."},{"key":"B37","doi-asserted-by":"publisher","first-page":"573","DOI":"10.1016\/j.devcel.2013.02.010","article-title":"Dynamin 2 potentiates invasive migration of pancreatic tumor cells through stabilization of the Rac1 GEF Vav1","volume":"24","author":"Razidlo","year":"2013","journal-title":"Dev. Cell"},{"key":"B38","doi-asserted-by":"publisher","first-page":"103","DOI":"10.1016\/j.ceb.2015.08.005","article-title":"Rho GTPase signalling in cell migration","volume":"36","author":"Ridley","year":"2015","journal-title":"Curr. Opin. Cell Biol."},{"key":"B39","doi-asserted-by":"publisher","first-page":"621","DOI":"10.1097\/FTD.0000000000000326","article-title":"Rac1 as a potential pharmacodynamic biomarker for thiopurine therapy in inflammatory bowel disease","volume":"38","author":"Seinen","year":"2016","journal-title":"Ther. Drug Monit."},{"key":"B40","doi-asserted-by":"publisher","first-page":"102582","DOI":"10.1016\/j.nano.2022.102582","article-title":"Triple negative breast cancer-derived small extracellular vesicles as modulator of biomechanics in target cells","volume":"44","author":"Senigagliesi","year":"2022","journal-title":"Nanomedicine"},{"key":"B41","doi-asserted-by":"publisher","first-page":"273103","DOI":"10.1088\/0953-8984\/27\/27\/273103","article-title":"Membrane tension and cytoskeleton organization in cell motility","volume":"27","author":"Sens","year":"2015","journal-title":"J. Phys. Condens. Matter"},{"key":"B42","doi-asserted-by":"publisher","first-page":"13699","DOI":"10.1074\/jbc.M115.694422","article-title":"Thiopurine prodrugs mediate immunosuppressive effects by interfering with Rac1 protein function","volume":"291","author":"Shin","year":"2016","journal-title":"J. Biol. Chem."},{"key":"B43","doi-asserted-by":"publisher","first-page":"491","DOI":"10.1097\/CJI.0b013e318176fae4","article-title":"Low-dose chemotherapeutic agents regulate small rho GTPase activity in dendritic cells","volume":"31","author":"Shurin","year":"2008","journal-title":"J. Immunother."},{"key":"B44","doi-asserted-by":"publisher","first-page":"30244","DOI":"10.1038\/srep30244","article-title":"PACSIN2 polymorphism is associated with thiopurine-induced hematological toxicity in children with acute lymphoblastic leukaemia undergoing maintenance therapy","volume":"6","author":"Smid","year":"2016","journal-title":"Sci. Rep."},{"key":"B45","doi-asserted-by":"publisher","first-page":"942","DOI":"10.3390\/cells12060942","article-title":"Caveolae mechanotransduction at the interface between cytoskeleton and extracellular matrix","volume":"12","author":"Sotodosos-Alonso","year":"2023","journal-title":"Cells"},{"key":"B46","doi-asserted-by":"publisher","first-page":"1133","DOI":"10.1172\/JCI200316432","article-title":"CD28-dependent Rac1 activation is the molecular target of azathioprine in primary human CD4+ T lymphocytes","volume":"111","author":"Tiede","year":"2003","journal-title":"J. Clin. Invest"},{"key":"B47","doi-asserted-by":"publisher","first-page":"658","DOI":"10.2174\/138920007782109742","article-title":"In vitro methods to study intestinal drug metabolism","volume":"8","author":"van de Kerkhof","year":"2007","journal-title":"Curr. Drug Metab."},{"key":"B48","doi-asserted-by":"publisher","first-page":"449","DOI":"10.1007\/s40272-020-00411-5","article-title":"Thiopurines in pediatric inflammatory bowel disease: current and future place","volume":"22","author":"van Hoeve","year":"2020","journal-title":"Paediatr. Drugs"},{"key":"B49","doi-asserted-by":"publisher","first-page":"815","DOI":"10.1053\/j.gastro.2018.05.028","article-title":"Abnormal small intestinal epithelial microvilli in patients with crohn\u2019s disease","volume":"155","author":"VanDussen","year":"2018","journal-title":"Gastroenterology"},{"key":"B50","doi-asserted-by":"publisher","first-page":"jcs260827","DOI":"10.1242\/jcs.260827","article-title":"Pacsin 2-dependent N-cadherin internalization regulates the migration behaviour of malignant cancer cells","volume":"136","author":"Wint","year":"2023","journal-title":"J. Cell Sci."},{"key":"B51","doi-asserted-by":"publisher","first-page":"2793","DOI":"10.3390\/ma13122793","article-title":"Insights into In Vitro wound closure on two biopolyesters-polylactide and polyhydroxyoctanoate","volume":"13","author":"Witko","year":"2020","journal-title":"Mater. (Basel)"},{"key":"B52","doi-asserted-by":"publisher","first-page":"6348","DOI":"10.3748\/wjg.v27.i38.6348","article-title":"Biomarkers for gastrointestinal adverse events related to thiopurine therapy","volume":"27","author":"Zudeh","year":"2021","journal-title":"World J. Gastroenterology"},{"key":"B53","doi-asserted-by":"publisher","first-page":"e202201610","DOI":"10.26508\/lsa.202201610","article-title":"PACSIN2 as a modulator of autophagy and mercaptopurine cytotoxicity: mechanisms in lymphoid and intestinal cells","volume":"6","author":"Zudeh","year":"2023","journal-title":"Life Sci. Alliance"}],"container-title":["Frontiers in Pharmacology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fphar.2026.1860702\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,1]],"date-time":"2026-09-01T13:30:43Z","timestamp":1788269443000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fphar.2026.1860702\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,9,1]]},"references-count":52,"alternative-id":["10.3389\/fphar.2026.1860702"],"URL":"https:\/\/doi.org\/10.3389\/fphar.2026.1860702","relation":{},"ISSN":["1663-9812"],"issn-type":[{"value":"1663-9812","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,9,1]]},"article-number":"1860702"}}