Saliva collection methods for DNA biomarker analysis in oral cancer patients.
Patients with head and neck cancers are predisposed to local recurrence and second primaries because of the phenomenon of field cancerisation, and clinical detection of recurrence remains challenging. DNA biomarkers in saliva may prove to be an adjunct to current diagnostic methods, but irradiation of the primary site often leads to xerostomia. We assessed 3 methods of collecting saliva for their ability to generate DNA of sufficient quantity and quality to use in biomarker assays. Paired saliva samples were collected from 2 groups of patients with oral squamous cell carcinoma (SCC). In the first group saliva was collected in Oragene(®) vials and as saline mouthwash from non-irradiated patients (n=21) (4 had had radiotherapy before collection); in the second group it was collected using Oragene(®) sponge kits and as mouthwash from irradiated patients (n=24). Quantitative polymerase chain reaction (qPCR) showed that Oragene(®) vials contained DNA in significantly greater amounts (median 122 μg, range 4-379) than mouthwash (median 17 μg, range 2-194) (p=0.0001) in the non-irradiated patients, while Oragene(®) sponge kits (median 4 μg, range 0.1-61) and mouthwash (median 5.5 μg, range 0.1-75) generated comparable concentrations of DNA from the irradiated group. All 90 samples contained DNA of sufficient quantity and quality for p16 promoter quantitative methylation-specific PCR (qMSP). While Oragene(®) vials contained the most DNA, all 3 methods yielded enough to detect DNA biomarkers using qMSP. The method of collection should depend on the compliance of the patient and oral competency.
School of Dentistry, University of Liverpool and Regional Maxillofacial, Unit, Aintree University Hospitals NHS Foundation Trust, Liverpool, UK.
April M
AM
Matthews
Harpal
H
Kaur
Michael
M
Dodd
Jacob
J
D'Souza
Triantafillos
T
Liloglou
Richard J
RJ
Shaw
Janet M
JM
Risk
D000818:Animals; D014408:Biomarkers, Tumor / Q000032:analysis*; D002294:Carcinoma, Squamous Cell / Q000378:metabolism* / Q000532:radiotherapy / Q000601:surgery; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000032:analysis; D019175:DNA Methylation / Q000235:genetics; D004273:DNA, Neoplasm / Q000032:analysis*; D006801:Humans; D009062:Mouth Neoplasms / Q000378:metabolism* / Q000532:radiotherapy / Q000601:surgery; D009067:Mouthwashes / Q000008:administration & dosage; D016133:Polymerase Chain Reaction; D011161:Porifera; D012463:Saliva / Q000737:chemistry*; D012965:Sodium Chloride / Q000008:administration & dosage; D013048:Specimen Handling / Q000295:instrumentation / Q000379:methods*
D014408:Biomarkers, Tumor; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004273:DNA, Neoplasm; D009067:Mouthwashes; D012965:Sodium Chloride