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Droplet Digital PCR Analysis of Liquid Biopsy Samples Unveils the Diagnostic Role of hsa-miR-133a-3p and hsa-miR-375-3p in Oral Cancer

1
Department of General Surgery, Section of Maxillo Facial Surgery, Policlinico San Marco, University of Catania, 95123 Catania, Italy
2
Epidemiology Unit, IRCCS Istituto Nazionale Tumori “Fondazione G. Pascale”, 80131 Naples, Italy
3
Department of Biomedical and Biotechnological Sciences, University of Catania, 95123 Catania, Italy
4
Otolaryngology Unit, Department of Medical Sciences, Surgical and Advanced Technologies, University of Catania, 95123 Catania, Italy
5
Research Center for Prevention, Diagnosis and Treatment of Cancer, University of Catania, 95123 Catania, Italy
*
Author to whom correspondence should be addressed.
These authors contributed equally to this work.
These authors share senior authorship.
Biology 2020, 9(11), 379; https://doi.org/10.3390/biology9110379
Received: 12 October 2020 / Revised: 29 October 2020 / Accepted: 4 November 2020 / Published: 6 November 2020
Despite the availability of screening programs, oral cancer is often diagnosed due to the lack of effective biomarkers. Therefore, the identification of new effective diagnostic and late prognostic biomarkers is of fundamental importance for the management of this tumor type. In our previous computational study, we have identified a set of microRNAs (miRNAs) significantly dysregulated in oral cancer and with a potential diagnostic and prognostic significance for oral cancer patients. Starting from our preliminary bioinformatics results, the aim of the present study was to validate the diagnostic potential of four selected miRNAs, hsa-miR-133a-3p, hsa-miR-375-3p, hsa-miR-503-5p and hsa-miR-196a-5p, in liquid biopsy samples obtained from oral cancer patients and healthy donors. For this purpose, the expression levels of the selected miRNAs were determined in plasma samples by using specific miRNA probes and droplet digital PCR (ddPCR). The ddPCR results showed that the hsa-miR-133a-3p and hsa-miR-375-3p were significantly down-regulated in oral cancer and their evaluation in liquid biopsy samples can predict the risk of oral cancer development with high sensitivity and specificity. Finally, the computational analysis of miRNA expression and clinical-pathological features of patients allowed us to establish the functional role and prognostic significance of the two validated miRNAs.
Despite the availability of screening programs, oral cancer deaths are increasing due to the lack of diagnostic biomarkers leading to late diagnosis and a poor prognosis. Therefore, there is an urgent need to discover novel effective biomarkers for this tumor. On these bases, the aim of this study was to validate the diagnostic potential of microRNAs (miRNAs) through the analysis of liquid biopsy samples obtained from ten oral cancer patients and ten healthy controls. The expression of four selected miRNAs was evaluated by using droplet digital PCR (ddPCR) in a pilot cohort of ten oral cancer patients and ten healthy donors. Bioinformatics analyses were performed to assess the functional role of these miRNAs. The expression levels of the predicted down-regulated hsa-miR-133a-3p and hsa-miR-375-3p were significantly reduced in oral cancer patients compared to normal individuals while no significant results were obtained for the up-regulated hsa-miR-503-5p and hsa-miR-196a-5p. ROC analysis confirmed the high sensitivity and specificity of hsa-miR-375-3p and hsa-miR-133a-3p. Therefore, both miRNAs are significantly down-regulated in cancer patients and can be used as biomarkers for the early diagnosis of oral cancer. The analysis of circulating miRNAs in a larger series of patients is mandatory to confirm the results obtained in this pilot study. View Full-Text
Keywords: oral cancers; microRNAs; ddPCR; diagnosis; biomarkers; cancer; epigenetics oral cancers; microRNAs; ddPCR; diagnosis; biomarkers; cancer; epigenetics
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MDPI and ACS Style

Crimi, S.; Falzone, L.; Gattuso, G.; Grillo, C.M.; Candido, S.; Bianchi, A.; Libra, M. Droplet Digital PCR Analysis of Liquid Biopsy Samples Unveils the Diagnostic Role of hsa-miR-133a-3p and hsa-miR-375-3p in Oral Cancer. Biology 2020, 9, 379. https://doi.org/10.3390/biology9110379

AMA Style

Crimi S, Falzone L, Gattuso G, Grillo CM, Candido S, Bianchi A, Libra M. Droplet Digital PCR Analysis of Liquid Biopsy Samples Unveils the Diagnostic Role of hsa-miR-133a-3p and hsa-miR-375-3p in Oral Cancer. Biology. 2020; 9(11):379. https://doi.org/10.3390/biology9110379

Chicago/Turabian Style

Crimi, Salvatore, Luca Falzone, Giuseppe Gattuso, Caterina M. Grillo, Saverio Candido, Alberto Bianchi, and Massimo Libra. 2020. "Droplet Digital PCR Analysis of Liquid Biopsy Samples Unveils the Diagnostic Role of hsa-miR-133a-3p and hsa-miR-375-3p in Oral Cancer" Biology 9, no. 11: 379. https://doi.org/10.3390/biology9110379

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