{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T18:28:08Z","timestamp":1786559288435,"version":"build-2736575974"},"reference-count":50,"publisher":"Public Library of Science (PLoS)","issue":"8","license":[{"start":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T00:00:00Z","timestamp":1786492800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    Glioblastoma (GBM) remains a lethal primary brain tumor, in part because therapeutic efficacy is limited by the blood\u2013brain barrier (BBB) and the complex tumor microenvironment (TME). Sonodynamic therapy (SDT), i.e., use of ultrasound to activate chemical sensitizers and generate cytotoxic stress, offers a non-invasive strategy for treating deep-seated intracranial disease, but progress is constrained by the scarcity of validated sonosensitizers and the inefficiency of conventional\n                    <jats:italic>in vitro<\/jats:italic>\n                    screening methods. Here, we introduce a New Approach Methodology (NAM) that couples a neural network-based positive-unlabeled (PU) learning framework with a high-throughput, magnetic field\u2013guided 3D bioprinting platform to accelerate identification and experimental validation of SDT-sensitizing agents. Using curated drug and small-molecule data and RDKit-derived molecular descriptors, the PU classifier identifies candidate ultrasound-responsive compounds without requiring reliable negative labels. We then validate the AI-based predictions in physiologically relevant U-87 MG glioblastoma spheroids that reproduce key TME features, including spatial heterogeneity and a hypoxic core. The NAM identifies two FDA-approved drugs, carboplatin (advanced ovarian cancer) and memantine hydrochloride (Alzheimer\u2019s disease), as effective ultrasound-responsive agents. In 3D spheroids, combining low-intensity pulsed ultrasound with either drug significantly reduces viability compared with drug-only controls, and both combinations outperform temozolomide (TMZ), the current standard chemotherapeutic. Time-resolved responses reveal distinct kinetics: memantine produces strong early cytotoxicity (24 h) enhanced by ultrasound, whereas carboplatin shows delayed but pronounced cytotoxicity (72 h), also improved by ultrasound. Together, these results establish an integrated computational\u2013experimental NAM that enables rapid repurposing of approved drugs as SDT sensitizers and provides a scalable framework for advancing GBM therapeutic discovery while reducing reliance on animal studies.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0354981","type":"journal-article","created":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T17:36:28Z","timestamp":1786556188000},"page":"e0354981","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":0,"title":["Repurposing Alzheimer\u2019s and ovarian cancer drugs as sonosensitizers for glioblastoma via a positive-unlabeled learning and 3D bioprinting-based new approach methodology (NAM)"],"prefix":"10.1371","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0009-0009-3361-5718","authenticated-orcid":true,"given":"Rudrajit","family":"Majumder","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2899-6714","authenticated-orcid":true,"given":"Priyankan","family":"Datta","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1046-447X","authenticated-orcid":true,"given":"Sreejesh","family":"Moolayadukkam","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8713-4188","authenticated-orcid":true,"given":"Ishwar K.","family":"Puri","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2026,8,12]]},"reference":[{"issue":"18","key":"pone.0354981.ref001","doi-asserted-by":"crossref","first-page":"18810","DOI":"10.1021\/acs.jmedchem.5c02019","article-title":"Temozolomide-derived therapeutic strategies to overcome resistance in glioblastoma","volume":"68","author":"H Zhang","year":"2025","journal-title":"J Med Chem"},{"key":"pone.0354981.ref002","doi-asserted-by":"crossref","DOI":"10.15586\/codon.glioblastoma.2017.ch11","volume-title":"Current standards of care in glioblastoma therapy","author":"C Fernandes","year":"2017"},{"issue":"8","key":"pone.0354981.ref003","doi-asserted-by":"crossref","first-page":"591","DOI":"10.1038\/nrn1728","article-title":"Drug resistance in brain diseases and the role of drug efflux transporters","volume":"6","author":"W L\u00f6scher","year":"2005","journal-title":"Nat Rev Neurosci"},{"issue":"32","key":"pone.0354981.ref004","doi-asserted-by":"crossref","first-page":"13184","DOI":"10.1021\/ja304986t","article-title":"Chemical molecule-induced light-activated system for anticancer and antifungal activities","volume":"134","author":"H Yuan","year":"2012","journal-title":"J Am Chem Soc"},{"issue":"36","key":"pone.0354981.ref005","doi-asserted-by":"crossref","first-page":"11522","DOI":"10.1002\/anie.201805138","article-title":"Innovative strategies for hypoxic-tumor photodynamic therapy","volume":"57","author":"X Li","year":"2018","journal-title":"Angew Chem Int Ed Engl"},{"key":"pone.0354981.ref006","doi-asserted-by":"crossref","first-page":"429","DOI":"10.1007\/978-3-319-22536-4_22","article-title":"Sonodynamic therapy: concept, mechanism and application to cancer treatment","author":"AP McHale","year":"2016","journal-title":"Therapeutic Ultrasound"},{"key":"pone.0354981.ref007","doi-asserted-by":"crossref","first-page":"0080","DOI":"10.34133\/bmef.0080","article-title":"Recent advances and future directions in sonodynamic therapy for cancer treatment","volume":"2024","author":"P Datta","year":"2024","journal-title":"BME Front"},{"issue":"9","key":"pone.0354981.ref008","doi-asserted-by":"crossref","first-page":"1553","DOI":"10.1016\/j.drudis.2016.06.024","article-title":"Cell spheroids: the new frontiers in in vitro models for cancer drug validation","volume":"21","author":"M Chatzinikolaidou","year":"2016","journal-title":"Drug Discov Today"},{"issue":"6","key":"pone.0354981.ref009","doi-asserted-by":"crossref","first-page":"1513","DOI":"10.1021\/acsptsci.5c00162","article-title":"Modernizing preclinical drug development: the role of new approach methodologies","volume":"8","author":"K Mehta","year":"2025","journal-title":"ACS Pharmacol Transl Sci"},{"issue":"13","key":"pone.0354981.ref010","doi-asserted-by":"crossref","first-page":"7519","DOI":"10.1021\/acs.jafc.5c02930","article-title":"Challenges and opportunities in the development and adoption of new approach methods (NAMs)","volume":"73","author":"SL Levine","year":"2025","journal-title":"J Agric Food Chem"},{"issue":"12","key":"pone.0354981.ref011","doi-asserted-by":"crossref","first-page":"796","DOI":"10.3390\/biomimetics10120796","article-title":"Integrating new approach methodologies (NAMs) into preclinical regulatory evaluation of oncology drugs","volume":"10","author":"MS Mirlohi","year":"2025","journal-title":"Biomimetics"},{"key":"pone.0354981.ref012","doi-asserted-by":"crossref","first-page":"1264","DOI":"10.1016\/j.proeng.2012.06.156","article-title":"A study on cheminformatics and its applications on modern drug discovery","volume":"38","author":"BF Begam","year":"2012","journal-title":"Procedia Eng"},{"issue":"32","key":"pone.0354981.ref013","doi-asserted-by":"crossref","first-page":"14455","DOI":"10.1039\/D5SC03648H","article-title":"Data-driven discovery of near-infrared type I photosensitizers for RNA-targeted tumor photodynamic therapy","volume":"16","author":"W Chen","year":"2025","journal-title":"Chem Sci"},{"issue":"6","key":"pone.0354981.ref014","doi-asserted-by":"crossref","first-page":"801","DOI":"10.1039\/c6pp00084c","article-title":"A quantitative structure-property relationship (QSPR) study of singlet oxygen generation by pteridines","volume":"15","author":"AA Buglak","year":"2016","journal-title":"Photochem Photobiol Sci"},{"key":"pone.0354981.ref015","doi-asserted-by":"crossref","first-page":"139850","DOI":"10.1016\/j.molstruc.2024.139850","article-title":"Accelerating the discovery of type \u2161 photosensitizer: Experimentally validated machine learning models for predicting the singlet oxygen quantum yield of photosensitive molecule","volume":"1321","author":"L He","year":"2025","journal-title":"J Mol Struct"},{"issue":"1","key":"pone.0354981.ref016","doi-asserted-by":"crossref","DOI":"10.1080\/23311916.2023.2301638","article-title":"Alternative cancer therapy through modeling pteridines photosensitizer quantum yield singlet oxygen production using swarm-based support vector regression and extreme learning machine","volume":"11","author":"N Aldhafferi","year":"2024","journal-title":"Cogent Eng"},{"issue":"47","key":"pone.0354981.ref017","doi-asserted-by":"crossref","first-page":"19769","DOI":"10.1021\/jacs.1c08211","article-title":"Self-improving photosensitizer discovery system via Bayesian search with first-principle simulations","volume":"143","author":"S Xu","year":"2021","journal-title":"J Am Chem Soc"},{"key":"pone.0354981.ref018","article-title":"Theoretical comparisons of positive-unlabeled learning against positive-negative learning","volume":"29","author":"G Niu","year":"2016","journal-title":"Adv Neural Inform Process Syst"},{"issue":"20","key":"pone.0354981.ref019","doi-asserted-by":"crossref","first-page":"2640","DOI":"10.1093\/bioinformatics\/bts504","article-title":"Positive-unlabeled learning for disease gene identification","volume":"28","author":"P Yang","year":"2012","journal-title":"Bioinformatics"},{"key":"pone.0354981.ref020","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1109\/IISA.2019.8900698","article-title":"Positive and unlabeled learning algorithms and applications: A survey.","volume-title":"2019 10th International conference on information, intelligence, systems and applications (IISA)","author":"K Jaskie","year":"2019"},{"issue":"32","key":"pone.0354981.ref021","doi-asserted-by":"crossref","DOI":"10.1002\/adhm.202402877","article-title":"In vitro sonodynamic therapy using a high throughput 3D glioblastoma spheroid model with 5-ALA and TMZ Sonosensitizers","volume":"13","author":"P Datta","year":"2024","journal-title":"Adv Healthc Mater"},{"key":"pone.0354981.ref022","volume-title":"Inventors; McMaster University, assignee. 3D label-free contactless formation of cellular structures and co-cultures through diamagnetophoresis","author":"AR Fattah","year":"2023"},{"key":"pone.0354981.ref023","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gkr777","article-title":"ChEMBL: a large-scale bioactivity database for drug discovery","volume":"40","author":"A Gaulton","year":"2012","journal-title":"Nucleic Acids Res"},{"key":"pone.0354981.ref024","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gkp456","article-title":"PubChem: a public information system for analyzing bioactivities of small molecules","volume":"37","author":"Y Wang","year":"2009","journal-title":"Nucleic Acids Res"},{"key":"pone.0354981.ref025","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gkm958","article-title":"DrugBank: a knowledgebase for drugs, drug actions and drug targets","volume":"36","author":"DS Wishart","year":"2008","journal-title":"Nucleic Acids Res"},{"key":"pone.0354981.ref026","first-page":"5281","article-title":"RDKit: A software suite for cheminformatics, computational chemistry, and predictive modeling","volume":"8","author":"G Landrum","year":"2013","journal-title":"J Cheminform"},{"issue":"5","key":"pone.0354981.ref027","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1016\/0893-6080(89)90020-8","article-title":"Multilayer feedforward networks are universal approximators","volume":"2","author":"K Hornik","year":"1989","journal-title":"Neural Netw"},{"issue":"4","key":"pone.0354981.ref028","doi-asserted-by":"crossref","first-page":"455","DOI":"10.1007\/BF02134016","article-title":"Approximation by superpositions of a sigmoidal function","volume":"5","author":"G Cybenko","year":"1992","journal-title":"Math Control Signal Syst"},{"issue":"1","key":"pone.0354981.ref029","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1007\/s10822-024-00578-w","article-title":"MolGraph: a Python package for the implementation of molecular graphs and graph neural networks with TensorFlow and Keras","volume":"39","author":"A Kensert","year":"2024","journal-title":"J Comput Aided Mol Des"},{"key":"pone.0354981.ref030","article-title":"An overview of gradient descent optimization algorithms","author":"S Ruder","year":"2016","journal-title":"arXiv preprint arXiv:1609.04747"},{"key":"pone.0354981.ref031","volume-title":"Neural networks: a systematic introduction","author":"R Rojas","year":"2013"},{"key":"pone.0354981.ref032","article-title":"Positive-unlabeled learning with non-negative risk estimator","volume":"30","author":"R Kiryo","year":"2017","journal-title":"Adv Neural Inform Process Syst"},{"key":"pone.0354981.ref033","doi-asserted-by":"crossref","first-page":"213","DOI":"10.1145\/1401890.1401920","article-title":"Learning classifiers from only positive and unlabeled data.","volume-title":"Proceedings of the 14th ACM SIGKDD international conference on Knowledge discovery and data mining","author":"C Elkan","year":"2008"},{"issue":"4","key":"pone.0354981.ref034","doi-asserted-by":"crossref","first-page":"719","DOI":"10.1007\/s10994-020-05877-5","article-title":"Learning from positive and unlabeled data: a survey","volume":"109","author":"J Bekker","year":"2020","journal-title":"Mach Learn"},{"key":"pone.0354981.ref035","article-title":"Tensorflow: large-scale machine learning on heterogeneous distributed systems","author":"M Abadi","year":"2016","journal-title":"arXiv preprint"},{"issue":"2003","key":"pone.0354981.ref036","first-page":"448","article-title":"Learning with positive and unlabeled examples using weighted logistic regression","author":"WS Lee","year":"2003","journal-title":"ICML"},{"issue":"11","key":"pone.0354981.ref037","doi-asserted-by":"crossref","first-page":"1153","DOI":"10.1038\/s41592-019-0575-8","article-title":"Positive-unlabeled convolutional neural networks for particle picking in cryo-electron micrographs","volume":"16","author":"T Bepler","year":"2019","journal-title":"Nat Methods"},{"issue":"7","key":"pone.0354981.ref038","doi-asserted-by":"crossref","DOI":"10.1093\/bioinformatics\/btae416","article-title":"ADMET-AI: a machine learning ADMET platform for evaluation of large-scale chemical libraries","volume":"40","author":"K Swanson","year":"2024","journal-title":"Bioinformatics"},{"key":"pone.0354981.ref039","article-title":"ADMETlab 3.0: an updated comprehensive online ADMET prediction platform enhanced with broader coverage, improved performance, API functionality and decision support","volume":"52","author":"L Fu","year":"2024","journal-title":"Nucleic Acids Res"},{"issue":"12","key":"pone.0354981.ref040","doi-asserted-by":"crossref","first-page":"4491","DOI":"10.1021\/mp400316e","article-title":"Memantine transport across the mouse blood-brain barrier is mediated by a cationic influx H+ antiporter","volume":"10","author":"DC Mehta","year":"2013","journal-title":"Mol Pharm"},{"issue":"343","key":"pone.0354981.ref041","doi-asserted-by":"crossref","DOI":"10.1126\/scitranslmed.aaf6086","article-title":"Clinical trial of blood-brain barrier disruption by pulsed ultrasound","volume":"8","author":"A Carpentier","year":"2016","journal-title":"Sci Transl Med"},{"issue":"13","key":"pone.0354981.ref042","doi-asserted-by":"crossref","first-page":"3793","DOI":"10.1158\/1078-0432.CCR-18-3643","article-title":"Safety and feasibility of repeated and transient blood\u2013brain barrier disruption by pulsed ultrasound in patients with recurrent glioblastoma","volume":"25","author":"A Idbaih","year":"2019","journal-title":"Clin Cancer Res"},{"issue":"5","key":"pone.0354981.ref043","doi-asserted-by":"crossref","first-page":"509","DOI":"10.1016\/S1470-2045(23)00112-2","article-title":"Repeated blood\u2013brain barrier opening with an implantable ultrasound device for delivery of albumin-bound paclitaxel in patients with recurrent glioblastoma: a phase 1 trial","volume":"24","author":"AM Sonabend","year":"2023","journal-title":"The Lancet Oncology"},{"issue":"4","key":"pone.0354981.ref044","doi-asserted-by":"crossref","first-page":"681","DOI":"10.1007\/s00018-018-2954-1","article-title":"Comprehensive review on how platinum- and taxane-based chemotherapy of ovarian cancer affects biology of normal cells","volume":"76","author":"J Miku\u0142a-Pietrasik","year":"2019","journal-title":"Cell Mol Life Sci"},{"issue":"1","key":"pone.0354981.ref045","doi-asserted-by":"crossref","first-page":"16161","DOI":"10.1038\/s41598-020-73204-y","article-title":"Evaluation of low-intensity pulsed ultrasound on doxorubicin delivery in 2D and 3D cancer cell cultures","volume":"10","author":"M Pa\u0161kevi\u010di\u016bt\u0117","year":"2020","journal-title":"Sci Rep"},{"issue":"4","key":"pone.0354981.ref046","doi-asserted-by":"crossref","first-page":"503","DOI":"10.3390\/cancers11040503","article-title":"NMDA receptor-mediated signaling pathways enhance radiation resistance, survival and migration in glioblastoma cells-a potential target for adjuvant radiotherapy","volume":"11","author":"A M\u00fcller-L\u00e4ngle","year":"2019","journal-title":"Cancers (Basel)"},{"issue":"9","key":"pone.0354981.ref047","doi-asserted-by":"crossref","first-page":"1010","DOI":"10.1038\/nm0901-1010","article-title":"Glutamate release promotes growth of malignant gliomas","volume":"7","author":"T Takano","year":"2001","journal-title":"Nat Med"},{"issue":"19","key":"pone.0354981.ref048","doi-asserted-by":"crossref","first-page":"9463","DOI":"10.1158\/0008-5472.CAN-07-2034","article-title":"Autocrine glutamate signaling promotes glioma cell invasion","volume":"67","author":"SA Lyons","year":"2007","journal-title":"Cancer Res"},{"issue":"4","key":"pone.0354981.ref049","first-page":"513","article-title":"Temozolomide: an updated overview of resistance mechanisms, nanotechnology advances and clinical applications","volume":"19","author":"R Ortiz","year":"2021","journal-title":"Curr Neuropharmacol"},{"issue":"3","key":"pone.0354981.ref050","doi-asserted-by":"crossref","first-page":"463","DOI":"10.1038\/sj.bjc.6600711","article-title":"Temozolomide induces apoptosis and senescence in glioma cells cultured as multicellular spheroids","volume":"88","author":"W G\u00fcnther","year":"2003","journal-title":"Br J Cancer"}],"container-title":["PLOS One"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0354981","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,12]],"date-time":"2026-08-12T17:36:37Z","timestamp":1786556197000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0354981"}},"subtitle":[],"editor":[{"given":"Michael C","family":"Burger","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2026,8,12]]},"references-count":50,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2026,8,12]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0354981","relation":{},"ISSN":["1932-6203"],"issn-type":[{"value":"1932-6203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,8,12]]}}}