The detection of cell-free tumor DNA fragments shed into the circulation, reflecting the mutational landscape of a tumor type, has transformed the management of many cancers but not head and neck cancer. In contrast with other head and neck carcinomas, human papillomavirus (HPV)-associated oropharynx cancer has a unique and pervasive molecular signature that would make it an ideal candidate for detection and surveillance of HPV-mediated disease. Recent advances in the technology have enabled the sensitive and reliable identification and quantification of low copy number HPV gene expression in the blood. The clinical assay used in initial studies1 served as the platform for the commercially available NavDx test (Naveris), a blood test, or liquid biopsy, of circulating HPV tumor DNA.

In this issue of JAMA Otolaryngology–Head & Neck Surgery, Ferrandino et al2 assess the accuracy of tumor tissue–modified HPV DNA (TTMV-HPV DNA) using a commercially available test in the diagnosis and surveillance of HPV-associated oropharyngeal squamous cell carcinoma (SCC). The study has 2 parts: a diagnostic component and a surveillance component. The initial diagnostic component compared the frequency of positive circulating HPV tumor DNA tests in patients with biopsy-proven p16-positive SCC prior to treatment, yielding a test sensitivity and specificity of 91.5% and 100%, respectively. While the TTMV-HPV DNA is less invasive, performance characteristics are better than reported cytopathologic results using p16 and comparable with polymerase chain reaction–based HPV testing of cytologic specimens.3 The study raises the possibility that a blood test could supplant fine needle aspiration biopsies with p16 staining to diagnose some patients with oropharyngeal SCC.2 However, some limitations to this approach should be kept in mind.

Not all p16-positive SCCs are HPV related. Although p16-positive but HPV-negative cancers are grouped with HPV-associated cancers for staging and clinical trial purposes, these cancers are distinct and would not be expected to harbor HPV DNA. The diagnostic cohort in the study by Ferrandino et al2 included 163 patients with pathologically confirmed p16-positive disease in 98% but HPV-associated disease in only 85%. As expected, the TTMV-HPV DNA test detected HPV, but not all p16-positive disease, with high sensitivity and specificity. Historically, p16 immunostaining was used as a surrogate marker for HPV-mediated disease because of its widespread availability and low cost compared with in situ hybridization. As TTMV-HPV DNA testing becomes more widely available, the role of p16 immunostaining to define HPV-related disease may need to be reevaluated. However, TTMV-HPV testing appears to be highly effective in detecting DNA from HPV-related cancer. The test was designed to detect HPV DNA only from malignant tumors and not from HPV infection. Nevertheless, a positive test is not sufficient to establish a diagnosis; tumor tissue continues to be required to confirm the presence of SCC.

Ferrandino et al also found that TTMV-HPV DNA testing was useful in surveillance, predicting failure in advance of clinical recurrence.2 Of 290 patients in the surveillance cohort, 23 developed pathologically confirmed recurrences over a median follow up of 40.5 months. For 13 patients, a positive TTMV-HPV DNA test preceded the pathologically confirmed recurrence by a median of 47 days. One patient had a positive HPV DNA test more than 500 days prior to pathologically confirmed recurrence. In a preceding study of circulating HPV DNA, Chera et al also reported that abnormal HPV test results preceded the development of recurrence by 3.9 months in their study of 115 patients with p16-positive SCC.1 However, a greater number of patients had positive blood test results in the study by Chera et al (28 of 115 [24%]). Approximately one-half of those subsequently developed a recurrence, yielding a positive predictive value of only 54% compared with a positive predictive value of 100% for the study by Ferrandino et al,2 wherein every patient with a positive test result eventually had recurrence.

The reasons for these differences are not entirely clear, but a comparison of the 2 studies is instructive. In the study by Chera et al,1 patients had frequent blood draws during and after treatment—10 per patient over the course of the study. Testing was completed in conjunction with deintensification protocols in which imaging and laboratory surveillance was performed according to a prespecified schedule. All patients with persistently negative tests remained disease free. The authors determined that 2 positive tests were suggestive of recurrence, improving the positive predictive value to 94%.

In contrast, Ferrandino et al2 used data that were collected during the course of routine clinical care and at the discretion of the treating clinician. On average, patients had 1 to 2 TTMV-HPV DNA tests after treatment (591 tests in 290 patients), and it does not appear that testing was obtained according to a schedule. Furthermore, it is not clear if standardized surveillance imaging was used, which might have affected the time at which recurrences were clinically detected.

In both studies, negative predictive values were favorable. In the study by Chera et al,1 patients with durably negative tests (87 of 115 patients, or 76% of the cohort), never developed a recurrence (NPV of 100%). Patients in the current study were tested far less often, yet the negative predictive value of the test remained high (98.2%).2

The appropriate interval for surveillance testing remains unclear. Testing every 3 months in the first year, 4 in the next, and every 6 months subsequently—intervals frequently used for surveillance visits with or without imaging—seems reasonable. Baseline, pretreatment testing is preferred but not mandatory if HPV-related disease is confirmed. Ferrandino et al2 state that a prospective clinical validation study is needed, and I would agree with this sentiment. Nevertheless, the use of this technology shows remarkable promise to transform the ability to identify and follow patients with HPV-related disease. Testing is likely to be increasingly used in routine clinical care, as it is commercially available. It is incumbent on us to establish evidence for strong and detailed surveillance guidelines to share among the cancer community.

Corresponding Author: Miriam N. Lango, MD, Department of Head and Neck Surgery, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Blvd, Houston, TX 77030 (mnlango@mdanderson.org).

Published Online: July 9, 2023. doi:10.1001/jamaoto.2023.1938

Conflict of Interest Disclosures: None reported.

1.Chera  BS, Kumar  S, Shen  C,  et al.  Plasma circulating tumor HPV DNA for the surveillance of cancer recurrence in HPV-associated oropharyngeal cancer.   J Clin Oncol. 2020;38(10):1050-1058. doi:10.1200/JCO.19.02444PubMedGoogle ScholarCrossref 2.Ferrandino  RM, Chen  S, Kappauf  C,  et al.  Performance of liquid biopsy for diagnosis and surveillance of human papillomavirus–associated oropharyngeal cancer.   JAMA Otolaryngol Head Neck Surg. Published online July 9, 2023. doi:10.1001/jamaoto.2023.1937Google ScholarCrossref 3.El-Salem  F, Mansour  M, Gitman  M,  et al.  Real-time PCR HPV genotyping in fine needle aspirations of metastatic head and neck squamous cell carcinoma: exposing the limitations of conventional p16 immunostaining.   Oral Oncol. 2019;90:74-79. doi:10.1016/j.oraloncology.2019.02.006PubMedGoogle ScholarCrossref