TY - JOUR
T1 - A Novel Preclinical In Vitro 3D Model of Oral Carcinogenesis for Biomarker Discovery and Drug Testing
T2 - International Journal of Molecular Sciences
AU - Chitturi Suryaprakash, Ravi T.
AU - Shearston, Kate
AU - Farah, Camile S.
AU - Fox, Simon A.
AU - Iqbal, Muhammad M.
AU - Kadolsky, Ulrich
AU - Zhong, Xiao
AU - Saxena, Alka
AU - Kujan, Omar
N1 - Funding Information:
This project was funded by the Future Health Research and Innovation Fund scheme through the WA Near-miss Award Program (Grant No. WANMA2021/1), the Australian Dental Research Foundation (ADRF Grant No. 455-2019) and The University of Western Australia. RNAseq library preparation, next-generation sequencing, and bioinformatics data analysis were conducted in the Genomics WA Laboratory in Perth, Australia. This facility is supported by BioPlatforms Australia, the State Government of Western Australia, the Australian Cancer Research Foundation, the Cancer Research Trust, the Harry Perkins Institute of Medical Research, the Telethon Kids Institute and the University of Western Australia. We gratefully acknowledge the Australian Cancer Research Foundation and the Centre for Advanced Cancer Genomics for making available Illumina Sequencers for the use of Genomics WA.
Publisher Copyright:
© 2023 by the authors.
PY - 2023/2/17
Y1 - 2023/2/17
N2 - 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.
AB - 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.
KW - oral squamous cell carcinoma
KW - oral epithelial dysplasia
KW - spheroid
KW - chemoprevention
KW - tobacco
KW - alcohol
KW - VEGF
KW - pazopanib
KW - lenvatinib
UR - http://www.scopus.com/inward/record.url?scp=85148947835&partnerID=8YFLogxK
U2 - 10.3390/ijms24044096
DO - 10.3390/ijms24044096
M3 - Article
C2 - 36835505
SN - 1422-0067
VL - 24
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
IS - 4
M1 - 4096
ER -
