{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,28]],"date-time":"2026-07-28T20:36:11Z","timestamp":1785270971580,"version":"3.55.0"},"reference-count":66,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2023,2,17]],"date-time":"2023-02-17T00:00:00Z","timestamp":1676592000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Future Health Research and Innovation Fund scheme","award":["WANMA2021\/1"],"award-info":[{"award-number":["WANMA2021\/1"]}]},{"name":"Future Health Research and Innovation Fund scheme","award":["455-2019"],"award-info":[{"award-number":["455-2019"]}]},{"name":"Australian Dental Research Foundation","award":["WANMA2021\/1"],"award-info":[{"award-number":["WANMA2021\/1"]}]},{"name":"Australian Dental Research Foundation","award":["455-2019"],"award-info":[{"award-number":["455-2019"]}]},{"name":"University of Western Australia","award":["WANMA2021\/1"],"award-info":[{"award-number":["WANMA2021\/1"]}]},{"name":"University of Western Australia","award":["455-2019"],"award-info":[{"award-number":["455-2019"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>This study aimed to develop an in vitro three-dimensional (3D) cell culture model of oral carcinogenesis for the rapid, scalable testing of chemotherapeutic agents. Spheroids of normal (HOK) and dysplastic (DOK) human oral keratinocytes were cultured and treated with 4-nitroquinoline-1-oxide (4NQO). A 3D invasion assay using Matrigel was performed to validate the model. RNA was extracted and subjected to transcriptomic analysis to validate the model and assess carcinogen-induced changes. The VEGF inhibitors pazopanib and lenvatinib were tested in the model and were validated by a 3D invasion assay, which demonstrated that changes induced by the carcinogen in spheroids were consistent with a malignant phenotype. Further validation was obtained by bioinformatic analyses, which showed the enrichment of pathways associated with hallmarks of cancer and VEGF signalling. Overexpression of common genes associated with tobacco-induced oral squamous cell carcinoma (OSCC), such as MMP1, MMP3, MMP9, YAP1, CYP1A1, and CYP1B1, was also observed. Pazopanib and lenvatinib inhibited the invasion of transformed spheroids. In summary, we successfully established a 3D spheroid model of oral carcinogenesis for biomarker discovery and drug testing. This model is a validated preclinical model for OSCC development and would be suitable for testing a range of chemotherapeutic agents.<\/jats:p>","DOI":"10.3390\/ijms24044096","type":"journal-article","created":{"date-parts":[[2023,2,20]],"date-time":"2023-02-20T03:56:07Z","timestamp":1676865367000},"page":"4096","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":16,"title":["A Novel Preclinical In Vitro 3D Model of Oral Carcinogenesis for Biomarker Discovery and Drug Testing"],"prefix":"10.3390","volume":"24","author":[{"given":"Ravi Teja","family":"Chitturi Suryaprakash","sequence":"first","affiliation":[{"name":"UWA Dental School, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kate","family":"Shearston","sequence":"additional","affiliation":[{"name":"UWA Dental School, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1642-6204","authenticated-orcid":false,"given":"Camile S.","family":"Farah","sequence":"additional","affiliation":[{"name":"Australian Centre for Oral Oncology Research and Education, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2042-1809","authenticated-orcid":false,"given":"Simon A.","family":"Fox","sequence":"additional","affiliation":[{"name":"UWA Dental School, The University of Western Australia, Nedlands, WA 6009, Australia"},{"name":"Harry Perkins Institute of Medical Research, QEII Medical Centre and Centre for Medical Research, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8603-5348","authenticated-orcid":false,"given":"Muhammad Munir","family":"Iqbal","sequence":"additional","affiliation":[{"name":"Genomics WA, Harry Perkins Institute of Medical Research, Telethon Kids Institute, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ulrich","family":"Kadolsky","sequence":"additional","affiliation":[{"name":"Genomics WA, Harry Perkins Institute of Medical Research, Telethon Kids Institute, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiao","family":"Zhong","sequence":"additional","affiliation":[{"name":"Genomics WA, Harry Perkins Institute of Medical Research, Telethon Kids Institute, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5683-0618","authenticated-orcid":false,"given":"Alka","family":"Saxena","sequence":"additional","affiliation":[{"name":"Genomics WA, Harry Perkins Institute of Medical Research, Telethon Kids Institute, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5951-8280","authenticated-orcid":false,"given":"Omar","family":"Kujan","sequence":"additional","affiliation":[{"name":"UWA Dental School, The University of Western Australia, Nedlands, WA 6009, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,2,17]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"779","DOI":"10.1002\/hed.20573","article-title":"Current topics in the epidemiology of oral cavity and oropharyngeal cancers","volume":"29","author":"Gillison","year":"2007","journal-title":"Head Neck"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"209","DOI":"10.3322\/caac.21660","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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