Prognostic value and cost benefit of HPV testing for oropharyngeal cancer patients.

OBJECTIVES: High-risk human papillomavirus (HR-HPV) can cause oropharyngeal squamous cell carcinoma (OpSCC). The revised 8th edition of the AJCC Staging Manual now stages OpSCC by incorporating p16 immunohistochemistry (IHC), the surrogate marker for HPV status. This study assessed the prognostic values of p16 and HPV markers. METHODS: We identified 244 OpSCC patients diagnosed between 2000 and 2008 from the British Columbia Cancer Registry with enough tissue to conduct experiments. Formalin-fixed, paraffin-embedded tissue sections were stained for p16 IHC, RNA in situ hybridization (ISH) HPV 16 and 18, and DNA ISH HR-HPV. Electronic charts were reviewed to collect clinical and outcome data. Combined positive RNA and/or DNA ISH was used to denote HPV status. RESULTS: Human papillomavirus was positive among 77.9% of samples. Using HPV as the benchmark, p16 IHC had high sensitivity (90.5%), but low specificity (68.5%). Distinct subgroups of patients were identified by sequential separation of p16 then HPV status. Among both p16-positive and p16-negative groups, HPV-positive patients were younger, more males, and had better clinical outcomes, especially 5-year overall survival. We further evaluated the technical costs associated with HPV testing. CONCLUSION: Human papillomavirus is more prognostic than p16 for OpSCC. Clinical laboratories can adopt HPV RNA ISH for routine analysis.

Quantitative nuclear phenotype signatures predict nodal disease in oral squamous cell carcinoma.

BACKGROUND: Early-stage oral squamous cell carcinoma (OSCC) patients have a one-in-four risk of regional metastasis (LN+), which is also the most significant prognostic factor for survival. As there are no validated biomarkers for predicting LN+ in early-stage OSCC, elective neck dissection often leads to over-treatment and under-treatment. We present a machine-learning-based model using the quantitative nuclear phenotype of cancer cells from the primary tumor to predict the risk of nodal disease. METHODS AND FINDINGS: Tumor specimens were obtained from 35 patients diagnosed with primary OSCC and received surgery with curative intent. Of the 35 patients, 29 had well (G1) or moderately (G2) differentiated tumors, and six had poorly differentiated tumors. From each, two consecutive sections were stained for hematoxylin & eosin and Feulgen-thionin staining. The slides were scanned, and images were processed to curate nuclear morphometric features for each nucleus, measuring nuclear morphology, DNA amount, and chromatin texture/organization. The nuclei (n = 384,041) from 15 G1 and 14 G2 tumors were randomly split into 80% training and 20% test set to build the predictive model by using Random Forest (RF) analysis which give each tumor cell a score, NRS. The area under ROC curve (AUC) was 99.6% and 90.7% for the training and test sets, respectively. At the cutoff score of 0.5 as the median NRS of each region of interest (n = 481), the AUC was 95.1%. We then developed a patient-level model based on the percentage of cells with an NRS ≥ 0.5. The prediction performance showed AUC of 97.7% among the 80% (n = 23 patient) training set and with the cutoff of 61% positive cells achieved 100% sensitivity and 91.7% specificity. When applying the 61% cutoff to the 20% test set patients, the model achieved 100% accuracy. CONCLUSIONS: Our findings may have a clinical impact with an easy, accurate, and objective biomarker from routine pathology tissue, providing an unprecedented opportunity to improve neck management decisions in early-stage OSCC patients.

Tumor microRNA profile and prognostic value for lymph node metastasis in oral squamous cell carcinoma patients.

Neck lymph node metastasis (LN+) is one of the most significant prognostic factors affecting 1-in-2 patients diagnosed with oral squamous cell carcinoma (OSCC). The different LN outcomes between clinico-pathologically similar primary tumors suggest underlying molecular signatures that could be associated with the risk of nodal disease development. MicroRNAs (miRNAs)are short non-coding molecules that regulate the expression of their target genes to maintain the balance of cellular processes. A plethora of evidence has indicated that aberrantly expressed miRNAs are involved in cancers with either an antitumor or oncogenic role. In this study, we characterized miRNA expression among OSCC fresh-frozen tumors with known outcomes of nodal disease (82 LN+, 76 LN0). We identified 49 differentially expressed miRNAs in tumors of the LN+ group. Using penalized lasso Cox regression, we identified a group of 10 miRNAs of which expression levels were highly associated with nodal-disease free survival. We further reported a 4-miRNA panel (miR-21-5p, miR-107, miR-1247-3p, and miR-181b-3p) with high accuracy in discriminating LN status, suggesting their potential application as prognostic biomarkers for nodal disease.

Plasma-Derived Inflammatory Proteins Predict Oral Squamous Cell Carcinoma.

Oral squamous cell carcinoma (OSCC) is a major concern with high morbidity and mortality worldwide, even with the current knowledge and the advancement in treatment. OSCCs diagnosed at late-stage often require wide-excision with or without neck dissection, radiotherapy, or chemotherapy. When deemed successful, treatment often results in diminished quality of life, impaired function, and disfigurement. Strategies for early detection are urgently needed for patients afflicted with this disease. Inflammatory protein plasma biomarkers have shown to be potential tests for early detection and disease monitoring in several cancers. There has been no study on inflammation-related plasma biomarkers in OSCC. The objectives of the study were to use a multiplex approach to screen plasma-derived biomarkers and to examine the association of measurable proteins with OSCC. A total of 260 plasma samples (210 OSCC and 50 normal controls) were collected to measure for concentration of inflammatory related biomarkers using electrochemiluminescence multiplex assay. After screening of 82 potential biomarkers of the first 160 OSCC, 16 cytokines, chemokines, and growth factors were identified and verified in the second set of samples containing 50 OSCC and 50 normal. After adjustment of age and batch effects, the adjusted differential expression analysis showed that the OSCCs were markedly lower in 14 biomarkers and significantly higher level of interleukin 1 receptor antagonist (IL1Ra). By performing unsupervised clustering analysis, we observed distinctive groups of normal and two subgroups of OSCC. Linear regression of IL2, IL1Ra, and macrophage inhibitory factor (MIF) showed high accuracy in classifying OSCC with sensitivity of 0.96 and specificity of 0.92. In conclusion, this is the first paper to identify potential inflammatory plasma protein biomarkers of patients with OSCC. With further validation, the set of biomarkers can potentially be used to assist in early detection of OSCC when the disease is localized and in more treatable stage.

Tumor-associated immune aggregates in oral cancer: Their cellular composition and potential prognostic significance.

There is growing evidence supporting the importance of immune microenvironment in cancer development and progression, especially with the rapid development of immunotherapy. Presence of immune cell aggregates in solid tumors has been associated with clinical outcomes, but little is known about the immune microenvironment in oral squamous cell carcinoma (OSCC), which has high morbidity and mortality. Based on our preliminary observation, we hypothesize that there is the presence of tumor-associated immune aggregates (TaIAs) during oral cancer development. Adapting to the dynamic change of the composition of cellular membership and co-evolving with the tumor at invasion fronts, these TaIAs, either pro-inflammatory or immune suppressive, are associated with clinical consequences. With the unique access to a set of prospectively collected, highly annotated OSCC surgical samples and the use of multi-color immunostaining of key immune cells, the confirmation of our hypothesis may shed light of the underlying biology related to OSCC and the knowledge learned can potentially be used to identify prognostic markers, response predictive markers for immunotherapies, as well as novel therapeutic targets.

Nodal Disease Burden for Early-Stage Oral Cancer.

Importance: Nodal disease has a significant effect on survival of patients with oral squamous cell carcinoma (OSCC). The decision for elective neck dissection for clinically node-negative (cN0) disease remains elusive. Objectives: To determine the efficacy of prophylactic neck treatment and to assess the value of commonly used clinicopathologic factors associated with nodal disease for early-stage OSCC. Design, Setting, and Participants: This retrospective study from a population-based cancer registry included patients diagnosed as having OSCC from January 11, 2001, to December 24, 2007, who were identified from the British Columbia Cancer Agency Registry. Comprehensive clinicopathologic data, treatment information, and time to outcome were collected. Five-year overall survival, disease-specific survival, and cumulative incidence of regional failure (RF) were analyzed. Receiver operating characteristic curve analysis with sensitivity and specificity was used to determine the association of these covariates with RF during follow-up. Data were analyzed from January 16 to June 30, 2015. Interventions: Follow-up of patients with cN0 OSCC with or without prophylactic neck treatment (elective neck dissection [END] and or radiotherapy). Main Outcomes and Measures: Patient demographic characteristics, clinicopathologic data, treatment data, and time from the initial surgery to last follow-up, the development of RF, or death due to oral cancer or other causes. Results: Of the 469 patients with cN0 primary OSCC who underwent intent-to-cure surgery for the intraoral lesion, 447 received local excision (LE) for the primary tumor (256 men [57.3%] and 191 women [42.7%]; mean [SD] age, 63.3 [14.7] years). Patients who received prophylactic treatment of the neck (n = 125) compared with LE only (n = 322) had no survival advantage. The estimated 5-year overall and disease-specific survival rates were 61.9% (95% CI, 56.5%-67.8%) and 80.8% (95% CI, 76.1%-85.6%), respectively, for the LE-only group; 54.4% (95% CI, 45.9%-64.5%) and 73.1% (95% CI, 65%-82.3%), respectively, for the LE + END ± radiotherapy group; and 61.7% (95% CI, 52.3%-72.8%) and 80.3% (95% CI, 72%-89.4%), respectively, for the LE + END group. Among the patients with cN0 disease receiving LE only, 89 (27.6%; 95% CI, 23%-33%) developed RF at a median time of 10.8 months, and 71 of the RFs (79.8%) developed within 30 months. Tumor depth of invasion of at least 4 mm and tumor grade of 2 or 3 showed an association with RF but had poor sensitivity and specificity. Conclusions and Relevance: Commonly used pathologic factors to decide neck dissection for cN0 OSCC are not effective and can cause overtreatment or undertreatment. The need for identification of new objective approaches for risk assessment of RF is urgent.