Brush swab as a noninvasive surrogate for tissue biopsies in epigenomic profiling of oral cancer

Abstract

Background Oral squamous cell carcinoma (OSCC) has poor survival rates. There is a pressing need to develop more precise risk assessment methods to tailor clinical treatment. Epigenome-wide association studies in OSCC have not produced a viable biomarker. These studies have relied on methylation array platforms, which are limited in their ability to profile the methylome. In this study, we use MethylCap-Seq (MC-Seq), a comprehensive methylation quantification technique, and brush swab samples, to develop a noninvasive, readily translatable approach to profile the methylome in OSCC patients. Methods Three OSCC patients underwent collection of cancer and contralateral normal tissue and brush swab biopsies, totaling 4 samples for each patient. Epigenome-wide DNA methylation quantification was performed using the SureSelectXT Methyl-Seq platform. DNA quality and methylation site resolution were compared between brush swab and tissue samples. Correlation and methylation value difference were determined for brush swabs vs. tissues for each respective patient and site (i.e., cancer or normal). Correlations were calculated between cancer and normal tissues and brush swab samples for each patient to determine the robustness of DNA methylation marks using brush swabs in clinical biomarker studies. Results There were no significant differences in DNA yield between tissue and brush swab samples. Mapping efficiency exceeded 90% across all samples, with no differences between tissue and brush swabs. The average number of CpG sites with at least 10x depth of coverage was 2,716,674 for brush swabs and 2,903,261 for tissues. Matched tissue and brush swabs had excellent correlation (r = 0.913 for cancer samples and r = 0.951 for normal samples). The methylation profile of the top 1000 CpGs was significantly different between cancer and normal samples (mean p-value = 0.00021) but not different between tissues and brush swabs (mean p-value = 0.11). Conclusions Our results demonstrate that MC-Seq is an efficient platform for epigenome profiling in cancer biomarker studies, with broader methylome coverage than array-based platforms. Brush swab biopsy provides adequate DNA yield for MC-Seq, and taken together, our findings set the stage for development of a non-invasive methylome quantification technique for oral cancer with high translational potential.

Feasibility of a novel non-invasive swab technique for serial whole-exome sequencing of cervical tumors during chemoradiation therapy

“…A previous study examining epigenome-wide DNA methylation compared sample acquisition by biopsy and isohelix brush swab for cancers in the oral cavity found no significant difference in DNA yield between tissue and brush samples and matched tissue. Isohelix brush swabs had an excellent correlation in the oral cavity 10 . In this prior study, mapping efficacy was over 90% for swab-acquired DNA.…”

“…In this prior study, mapping efficacy was over 90% for swab-acquired DNA. Investigators successfully identified potential prognostic and predictive biomarkers for malignant lesions in the oral cavity with high sensitivity and specificity using a comparable isohelix swab design as was used here 10 . While isohelix brushes have been designed and marketed for DNA acquisition of the buccal mucosa, we were able to acquire sufficient quality and quantity DNA for WES in all but one tumor sample with collected samples by isohelix brushings from cervical mucosa.…”

“…Nevertheless, cervical tumors are an ideal setting for the serial study of treatment response because tumors can be readily monitored by physical exam and are accessible for sampling through the course of chemoradiotherapy. Encouragingly, successful non-invasive swab-based sample acquisition of DNA has been demonstrated in the screening and diagnosis of precancerous and cancerous lesions in the oral cavity and detection of HPV-associated cutaneous lesions by PCR [10][11][12][13] . This study hypothesizes that non-invasive swab-based sampling of cervical tumors can serve as a robust, noninvasive method to acquire tumor specimens for WES.…”

Feasibility of a novel non-invasive swab technique for serial whole-exome sequencing of cervical tumors during chemoradiation therapy

“…An additional study comparing sample acquisition by biopsy and isohelix brush swab for cancers in the oral cavity for epigenome-wide DNA methylation found no significant difference in DNA yield between tissue and brush samples and matched tissue. Isohelix brush swabs had an excellent correlation in the oral cavity [ 10 ]. Mean DNA yield from frozen tissue was 0.39ug (range 0.19–0.66ug), and 0.53ug (range 0.51–2.00ug) from swab-acquired DNA.…”

“…Nevertheless, cervical tumors are an ideal setting for the serial study of treatment response because tumors can be readily monitored by physical exam and are accessible for sampling through the course of chemoradiotherapy. Encouragingly, successful non-invasive swab-based sample acquisition of DNA has been demonstrated in the screening and diagnosis of precancerous and cancerous lesions in the oral cavity and detection of HPV-associated cutaneous lesions by PCR [ 10 – 13 ].…”