American Association for Cancer Research
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Supplementary Figure from HPV Sequencing Facilitates Ultrasensitive Detection of HPV Circulating Tumor DNA

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journal contribution
posted on 2023-03-31, 22:42 authored by Eric Leung, Kathy Han, Jinfeng Zou, Zhen Zhao, Yangqiao Zheng, Ting Ting Wang, Ariana Rostami, Lillian L. Siu, Trevor J. Pugh, Scott V. Bratman
Supplementary Figure from HPV Sequencing Facilitates Ultrasensitive Detection of HPV Circulating Tumor DNA



Human papillomavirus (HPV) DNA offers a convenient circulating tumor DNA (ctDNA) marker for HPV-associated malignancies, but current methods, such as digital PCR (dPCR), provide insufficient accuracy for clinical applications in patients with low disease burden. We asked whether a next-generation sequencing approach, HPV sequencing (HPV-seq), could provide quantitative and qualitative assessment of HPV ctDNA in low disease burden settings. We conducted preclinical technical validation studies on HPV-seq and applied it retrospectively to a prospective multicenter cohort of patients with locally advanced cervix cancer (NCT02388698) and a cohort of patients with oropharynx cancer. HPV-seq results were compared with dPCR. The primary outcome was progression-free survival (PFS) according to end-of-treatment HPV ctDNA detectability. HPV-seq achieved reproducible detection of HPV DNA at levels less than 0.6 copies in cell line data. HPV-seq and dPCR results for patients were highly correlated (R2 = 0.95, P = 1.9 × 10–29) with HPV-seq detecting ctDNA at levels down to 0.03 copies/mL plasma in dPCR-negative posttreatment samples. Detectable HPV ctDNA at end-of-treatment was associated with inferior PFS with 100% sensitivity and 67% specificity for recurrence. Accurate HPV genotyping was successful from 100% of pretreatment samples. HPV ctDNA fragment sizes were consistently shorter than non–cancer-derived cell-free DNA (cfDNA) fragments, and stereotyped cfDNA fragmentomic patterns were observed across HPV genomes. HPV-seq is a quantitative method for ctDNA detection that outperforms dPCR and reveals qualitative information about ctDNA. Our findings in this proof-of-principle study could have implications for treatment monitoring of disease burden in HPV-related cancers. Future prospective studies are needed to confirm that patients with undetectable HPV ctDNA following chemoradiotherapy have exceptionally high cure rates.

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