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Best of Scientific Oral Presentations

Head and Neck Surgery

Session: Best of Scientific Orals 2

Nanopore Sequencing Detects Prevalence of HPV DNA in Pediatric and Young Adult Archival Tonsil Specimens

Sunday, October 12, 2025

2:30 PM - 2:36 PM EDT

Location: 120

Introduction: Human papillomavirus (HPV) accounts for the majority of oropharyngeal squamous cell carcinoma (OPSCC) in the US but understanding the rate of high-risk HPV infection in oropharyngeal tissue remains poor. Nanopore sequencing has emerged as a rapid and accurate long-read sequencing platform to detect genetic profiles, but it has not been tested on HPV-driven OPSCC with formalin-fixed paraffin-embedded (FFPE) tissue. Our objectives are to assess the feasibility of Nanopore sequencing on FFPE-derived tissue from known viral-associated OPSCC and to evaluate prevalence of HPV in pediatric and young adult tonsil tissue after routine tonsillectomy.

Methods: Archival specimens of HPV-positive and negative oropharyngeal cancers diagnosed at a single academic hospital from 2013-2023 underwent DNA extraction and polymerase chain reaction (PCR) amplification for sequencing with the Nanopore platform to detect HPV DNA. HPV detection was compared against p16 and HPV in situ hybridization status. FFPE specimens from routine non-oncologic tonsillectomies from 2016-2023 in patients ages 30 and younger were subsequently tested for HPV using Nanopore sequencing.

Results: Of the 54 OPSCC samples, Nanopore sequencing detected HPV DNA in 27 of 27 HPV-positive specimens and in 3 of 27 HPV-negative samples, resulting in 100% sensitivity, 89% specificity, 90% PPV, and 100% NPV. From 151 tonsillectomy cases for benign purposes (67% female, mean age 18.7 + 7.7 years old), there were 313 FFPE blocks subjected to Nanopore sequencing. Overall prevalence of HPV DNA was 27% of patients. In patients with ages ≤18 years (n=67) and >18 years old (n=84), prevalence of HPV was 30% and 25%, respectively. In patients < 10 years old (n=27), the prevalence was 37%. HPV 16 subtype was detected in all positive specimens.

Conclusions: Nanopore sequencing can reliably detect HPV DNA in FFPE-derived oropharyngeal tissue. Prevalence of high-risk HPV in pediatric and young adults appeared higher than previously studied, which warrants further studies on epidemiology and may impact vaccination policies.

Presenting Author(s)

Mikayla G. Hubbard, BS

Medical Student
Indiana University School of Medicine
Avon, IN, United States

Disclosure(s):
faculty for this accredited education activity has no relationship(s) with ineligible companies to disclose.

Co-Author(s)

TD

Thomas E. Davis, MD, PhD

Professor
Indiana University School of Medicine Department of Pathology and Laboratory Medicine
Indianapolis, IN, United States

Disclosure information not submitted.
GL

GUANG-SHENG LEI, PhD

Indiana University School of Medicine Department of Pathology and Laboratory Medicine
Carmel, Indiana, United States

Disclosure information not submitted.
RE

Robert E. Emerson, MD

Professor
Indiana University School of Medicine Department of Pathology and Laboratory Medicine
Indianapolis, IN, United States

Disclosure information not submitted.
DS

Danielle Schreiber, ASCP

Medical Laboratory Scientist
Indiana University School of Medicine Department of Pathology and Laboratory Medicine,
Indianapolis, Indiana, United States

Disclosure information not submitted.

Senior Author(s)

Diane W. Chen, MD

Assistant Professor
Indiana University
Indianapolis, IN, United States

Disclosure information not submitted.