Ultrasensitive detection of tumor‐specific mutations in saliva of patients with oral cavity squamous cell carcinoma

Abstract

BACKGROUND: Oral cavity squamous cell carcinoma (OCSCC) is the most common head and neck malignancy. Although the survival rate of patients with advanced-stage disease remains approximately 20% to 60%, when detected at an early stage, the survival rate approaches 80%, posing a pressing need for a well validated profiling method to assess patients who have a high risk of developing OCSCC. Tumor DNA detection in saliva may provide a robust biomarker platform that overcomes the limitations of current diagnostic tests. However, there is no routine saliva-based screening method for patients with OCSCC. METHODS: The authors designed a custom nextgeneration sequencing panel with unique molecular identifiers that covers coding regions of 7 frequently mutated genes in OCSCC and applied it on DNA extracted from 121 treatment-naive OCSCC tumors and matched preoperative saliva specimens. RESULTS: By using stringent variant-calling criteria, mutations were detected in 106 tumors, consistent with a predicted detection rate ≥88%. Moreover, mutations identified in primary malignancies were also detected in 93% of saliva samples. To ensure that variants are not errors resulting in false-positive calls, a multistep analytical validation of this approach was performed: 1) re-sequencing of 46 saliva samples confirmed 88% of somatic variants; 2) no functionally relevant mutations were detected in saliva samples from 11 healthy individuals without a history of tobacco or alcohol; and 3) using a panel of 7 synthetic loci across 8 sequencing runs, it was confirmed that the platform developed is reproducible and provides sensitivity on par with droplet digital polymerase chain reaction. CONCLUSIONS: The current data highlight the feasibility of somatic mutation identification in driver genes in saliva collected at the time of OCSCC diagnosis.

Next generation sequencing-based salivary biomarkers in oral squamous cell carcinoma

“…( Tables 3 , 4 ) Most of these studies started from Gene Expression Omnibus datasets ( https://www.ncbi.nlm.nih.gov/gds ) in combination with differentially expressed genes, the protein-protein interaction network, gene ontology and the Kyoto Encyclopedia of Genes and Genomes pathway. Shanmugam et al 70 have reported a customized NGS analysis of unique coding regions of seven mutated genes from OSCC saliva, and similar data from whole-exome sequencing were also discovered in tumors from The Cancer Genome Atlas 71 , the International Cancer Genome Consortium gingiva-buccal cohort 69 , and MD Anderson Cancer Center OSCC cohort 72 .…”

“…Compared with known genomic salivary biomarkers, such as p53, FAT1, CASP8, PIK3CA, HRAS, NOTCH1, and CDKN2A, NGS-based genomics consider mutation of p53 and promoter hypermethylation of DAPK, TIMP3, p16, and MGMT, cyclone D1 and mammary serine protease inhibitors. Also, up-regulated oncogenic tissue miRNAs and down-regulated tissue suppressive marker miRNAs, including miR 31, miR 125, and miR 3928, are recommended as subjects for further research 70 , 74 , 76 - 79 , 81 . ( Table 5 )…”

Liquid Biopsy as a Tool for the Characterisation and Early Detection of the Field Cancerization Effect in Patients with Oral Cavity Carcinoma

“…However, the concordance in saliva was low; indeed, the analysis of TP53 mutations in healthy individuals revealed the presence of pathogenic mutations, which should be taken into account in any subsequent design of ctDNA assays for early diagnosis. Finally, a recent study by Shanmugam et al [33] used massive sequencing techniques to analyse tissue and saliva samples from 121 pre-surgical patients with OSCC. Alterations were identified in more than 95% of these patients, and in more than 97% the alteration was present in saliva, although this concordance was lower in initial disease stages.…”

Epidemiological, Clinical, and Genomic Profile in Head and Neck Cancer Patients and Their Families

“…Most cases (90%) are histologically classified as squamous cell carcinoma. Despite several therapeutic advances in cancer treatment, patients with advanced-stage disease presented a 5-year survival rate varying from ~20 to 60% [ 2 ]. The most significant improvements in survival were described only among patients with nasopharynx and oropharynx cancer [ 3 , 4 ].…”