Whole-genome sequencing (WGS)-based circulating tumor DNA (ctDNA) detection of human papillomavirus (HPV)-associated head and neck squamous cell carcinoma (HNSCC), termed HPV-DeepSeek by the study investigators, showed greater sensitivity, specificity, and diagnostic accuracy than any current single or combinatorial detection method. HPV-DeepSeek also had the highest positive predictive value of all detection methods, according to study findings published in Clinical Cancer Research. The high sensitivity and specificity rates with HPV-DeepSeek were also maintained when assessed in patients with early-stage disease, including asymptomatic patients.
“The goal of developing HPV-DeepSeek was to create a minimally invasive approach to detect HPV cancers that is significantly more sensitive than what is currently available for patients,” stated study author Daniel Faden, MD, FACS, Investigator, Mike Toth Head and Neck Cancer Research Center, Massachusetts Eye and Ear, and Assistant Professor of Otolaryngology–Head and Neck Surgery, Harvard Medical School. “Our findings demonstrate that we can use this approach to not only diagnose patients more accurately compared to what is currently available, but also provide the potential to screen for HPV cancers in the blood before patients ever develop symptoms, enabling us to catch and treat their cancers at the earliest stages.”
Study Methods and Results
The researchers sought to establish an early detection method for HPV-associated HNSCC with broad clinical utility. They developed HPV-DeepSeek, a multifeature HPV WGS liquid biopsy, to detect low-level ctHPVDNA.
Then, the researchers conducted a head-to-head comparison of WGS ctHPVDNA testing with standard detection methods, with a focus on other liquid biopsy approaches, to determine whether a single or combinatorial strategy could detect HPV-associated HNSCC early. The compared approaches included single-plex droplet digital polymerase chain reaction (ddPCR)–based ctHPVDNA detection, multiplex ddPCR-based ctHPVDNA detection, multiplex HPV protein antibody detection, and standard-of-care tissue biopsy.
Blood samples from 304 participants were tested with each liquid biopsy approach and compared with standard tissue biopsy. Of the patients, 152 had untreated HPV-associated HNSCC, the majority of which were stage I, and 152 population control patients.
HPV-DeepSeek yielded a sensitivity of 98.7% and a specificity of 98.7%, whereas ddPCR had rates of 94.2% and 98.6%, respectively. Multiplex ddPCR had a sensitivity of 90.6% and a specificity of 96.3%, whereas HPV protein antibody detection had rates of 87.4% and 98.8%, respectively.
In a head-to-head comparison, the WGS ctHPVDNA detection showed greater diagnostic accuracy than ddPCR (0.99 vs 0.90; P < .001), HPV antibody detection (0.99 vs 0.83; P < .001), and clinical workup (0.99 vs 0.82; P < .001). The WGS ctHPVDNA method maintained high diagnostic accuracy among patients with early-stage disease.
When the detection methods were modeled for screening biomarkers for HPV-associated HNSCC, and tested in men between the ages of 55 and 74, the WGS ctHPVDNA method had the lowest number of patients needed to screen (2,903) and the highest positive predictive value (2.6) of all approaches.
Further Studies
The study authors are now conducting other studies to potentially expand the utility of HPV-DeepSeek, especially for the early detection of cancers.
In another study currently in preprint, the study authors tested HPV-DeepSeek in 28 blood samples of patients who developed HPV-associated oropharyngeal cancer 1.3 to 10.8 years before their cancer diagnosis. The liquid biopsy was also tested in 28 gender-matched controls. Of the 28 cases and 0 controls, 22 of the cases showed signs of HPV-associated oropharyngeal cancer. The liquid biopsy achieved 100% detection within 4 years of diagnosis, and the longest lead time was 7.8 years. With a machine-learning model, 27 of the 28 cases were classified with 100% detection within 10 years of the cancer diagnosis.
“The natural history of these cancers is that they grow over a period of about 15 years, and as they grow, they release DNA from the HPV genome into the blood. If we can detect these cancers years earlier, at their earliest stages, it could drastically change how we treat patients; for example, being able to use less treatment leading to fewer side effects,” Dr. Faden stated. “While our results suggest great potential for clinical benefit, there are unanswered questions we need to address to understand how we could best utilize these approaches and what the follow-up regimens would be for patients who screen positive.”
In addition, the research team is also exploring a novel assay for evidence of measurable residual disease (MRD) after surgery in patients with HNSCC. The researchers developed a whole-genome, tumor-informed, mutation enrichment sequencing assay, named MAESTRO, which uses minor allele-enriched sequencing through recognition oligonucleotides. The assay was evaluated for early MRD detection after surgery in patients with HNSCC to guide adjuvant treatment decision-making.
According to the findings published in Clinical Cancer Research, the MAESTRO assay achieved a positive predictive value of 92.9% and a negative predictive value of 80% among 24 patients with HPV-independent HNSCC. Early MRD detection was associated with recurrence as well as worse overall survival and event-free survival irrespective of high-risk pathology.
“In this study, MAESTRO not only detected residual cancer in more patients who experienced future recurrence or death but also was highly predictive for it,” stated study author Viktor A. Adalsteinsson, PhD, Director of the Gerstner Center for Cancer Diagnostics at the Broad Institute.
“Whole-genome sequencing liquid biopsy approaches, like HPV-DeepSeek and MAESTRO used in these studies, are enormously powerful, allowing physicians to look for many hundreds or thousands of needles in haystacks as opposed to just a few, drastically increasing sensitivity,” Dr. Faden concluded. “For patients, this means significantly more accurate results and being one step closer to truly personalized care.”
Disclosure: For full disclosures of the study authors, visit aacrjournals.org.
