Positive Oropharyngeal Squamous Cell Carcinoma Research Articles

Detection, Patterns, and Outcomes of Recurrent HPV-Positive Oropharyngeal Squamous Cell Carcinoma

Recurrent human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) is a relatively rare but serious disease with little empirical data. Previous works have studied patterns of recurrence in HPV-positive OPSCC, but only one has studied truly recurrent disease as opposed to persistent disease, and no work systematically analyzed posttreatment surveillance imaging strategies and how recurrences were detected. To refine the understanding of HPV-positive OPSCC recurrence and inform optimal imaging surveillance strategies. This retrospective cohort study involved electronic medical record review at a tertiary care hospital. Patients treated for OPSCC from 2012 to 2019 were extracted, and patients diagnosed with HPV-positive OPSCC were identified. Data were analyzed from December 2022 to May 2023. Percentage of patients with a true recurrence, location of recurrence, time of recurrence detection, and method of recurrence detection. Recurrence was demonstrated with a scan after an imaging-established disease-free state 3 to 6 months posttreatment. Of the 367 patients with HPV-positive OPSCC (mean [SD] age, 60.6 [9.2] years; 310 [84.5%] male), 37 (10.1%) experienced true disease recurrence. Median (IQR) follow-up time of the cohort was 3.6 years (8.5-88 months), defined as time from diagnosis to death or last contact. Within the true recurrence cohort, 21 patients (56.8%) experienced local, regional, or local and regional recurrence (LRR); 15 (40.5%) experienced distant metastasis (DM); and 1 (2.7%) experienced both LRR and DM. The mean (SD) time for detecting LRR was 2.46 (1.94) years and was considerably longer compared to the 1.89 (0.87) years for detecting DM (difference, 0.57 [95% CI, -0.29 to 1.02] years). The majority of patients identified their recurrence through symptom changes (31 [81.1%]) rather than through surveillance imaging (3 [8.1%]). In this cohort study, 10.1% of patients experienced true HPV-positive OPSCC disease recurrence, with most incidences of DM occurring in the lung and brain. Disease recurrence was identified primarily through symptomatic change, suggesting that further research may be needed to understand the optimal surveillance strategies after definitive treatment.

Immunoreactivity of LMO7 and other molecular markers as potential prognostic factors in oropharyngeal squamous cell carcinoma

BackgroundDespite the better prognosis associated with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC), some patients experience relapse and succumb to the disease; thus, there is a need for biomarkers identifying these patients for intensified treatment. Leucine-rich repeats and immunoglobulin-like domain (LRIG) protein 1 is a negative regulator of receptor tyrosine kinase signaling and a positive prognostic factor in OPSCC. Studies indicate that LRIG1 interacts with the LIM domain 7 protein (LMO7), a stabilizer of adherence junctions. Its role in OPSCC has not been studied before.MethodsA total of 145 patients diagnosed with OPSCC were enrolled. Immunohistochemical LMO7 expression and staining intensity were evaluated in the tumors and correlated with known clinical and pathological prognostic factors, such as HPV status and LRIG1, CD44, Ki67, and p53 expression.ResultsOur results show that high LMO7 expression is associated with significantly longer overall survival (OS) (p = 0.044). LMO7 was a positive prognostic factor for OS in univariate analysis (HR 0.515, 95% CI: 0.267–0.994, p = 0.048) but not in multivariate analysis. The LMO7 expression correlated with LRIG1 expression (p = 0.048), consistent with previous findings. Interestingly, strong LRIG1 staining intensity was an independent negative prognostic factor in the HPV-driven group of tumors (HR 2.847, 95% Cl: 1.036–7.825, p = 0.043).ConclusionsWe show for the first time that high LMO7 expression is a positive prognostic factor in OPSCC, and we propose that LMO7 should be further explored as a biomarker. In contrast to previous reports, LRIG1 expression was shown to be an independent negative prognostic factor in HPV-driven OPSCC.

Explainable prediction model for the human papillomavirus status in patients with oropharyngeal squamous cell carcinoma using CNN on CT images

Several studies have emphasised how positive and negative human papillomavirus (HPV+ and HPV−, respectively) oropharyngeal squamous cell carcinoma (OPSCC) has distinct molecular profiles, tumor characteristics, and disease outcomes. Different radiomics-based prediction models have been proposed, by also using innovative techniques such as Convolutional Neural Networks (CNNs). Although some of these models reached encouraging predictive performances, there evidence explaining the role of radiomic features in achieving a specific outcome is scarce. In this paper, we propose some preliminary results related to an explainable CNN-based model to predict HPV status in OPSCC patients. We extracted the Gross Tumor Volume (GTV) of pre-treatment CT images related to 499 patients (356 HPV+ and 143 HPV−) included into the OPC-Radiomics public dataset to train an end-to-end Inception-V3 CNN architecture. We also collected a multicentric dataset consisting of 92 patients (43 HPV+ , 49 HPV−), which was employed as an independent test set. Finally, we applied Gradient-weighted Class Activation Mapping (Grad-CAM) technique to highlight the most informative areas with respect to the predicted outcome. The proposed model reached an AUC value of 73.50% on the independent test. As a result of the Grad-CAM algorithm, the most informative areas related to the correctly classified HPV+ patients were located into the intratumoral area. Conversely, the most important areas referred to the tumor edges. Finally, since the proposed model provided additional information with respect to the accuracy of the classification given by the visualization of the areas of greatest interest for predictive purposes for each case examined, it could contribute to increase confidence in using computer-based predictive models in the actual clinical practice.

Systematic evaluation and meta-analysis of the prognosis of down-staging human papillomavirus (HPV) positive oropharyngeal squamous cell carcinoma using cetuximab combined with radiotherapy instead of cisplatin combined with radiotherapy.

To evaluate the efficacy and safety of cetuximab instead of cisplatin in combination with downstaging radiotherapy for papillomavirus (HPV) positive oropharyngeal squamous cell carcinoma (HPV+ OPSCC). Meta-analysis and systematic evaluation. The PubMed, Embase, Web of Science, and Cochrane library databases were searched up to June 8, 2023, as well as Clinicaltrials.gov Clinical Trials Registry, China Knowledge Network, Wanfang Data Knowledge Service Platform, and Wiprojournal.com. Randomized controlled trials reporting results of standard regimens of cetuximab + radiotherapy vs cisplatin + radiotherapy in treating HPV+ OPSCC were included. The primary outcomes of interest were overall survival (OS), progression-free survival (PFS), local regional failure rate (LRF), distant metastasis rate (DM), and adverse events (AE). Two reviewers independently extracted data and assessed the risk of bias of the included studies. The HR and its 95% CI were used as the effect analysis statistic for survival analysis, while the OR and its 95% CI were used as the effect analysis statistic for dichotomous variables. These statistics were extracted by the reviewers and aggregated using a fixed-effects model to synthesise the data. A total of 874 relevant papers were obtained from the initial search, and five papers that met the inclusion criteria were included; a total of 1,617 patients with HPV+ OPSCC were enrolled in these studies. Meta-analysis showed that OS and PFS were significantly shorter in the cetuximab + radiotherapy group of patients with HPV+ OPSCC compared with those in the conventional cisplatin + radiotherapy group (HR = 2.10, 95% CI [1.39-3.15], P = 0.0004; HR = 1.79, 95% CI [1.40-2.29], P < 0.0001); LRF and DM were significantly increased (HR = 2.22, 95% CI [1.58-3.11], P < 0.0001; HR = 1.66, 95% CI [1.07-2.58], P = 0.02), but there was no significant difference in overall grade 3 to 4, acute and late AE overall (OR = 0.86, 95% CI [0.65-1.13], P = 0.28). Cisplatin + radiotherapy remains the standard treatment for HPV+ OPSCC. According to the 7th edition AJCC/UICC criteria, low-risk HPV+ OPSCC patients with a smoking history of ≤ 10 packs/year and non-pharyngeal tumors not involved in lymphatic metastasis had similar survival outcomes with cetuximab/cisplatin + radiotherapy. However, further clinical trials are necessary to determine whether cetuximab + radiotherapy can replace cisplatin + radiotherapy for degraded treatment in individuals who meet the aforementioned characteristics, particularly those with platinum drug allergies. CRD42023445619.

Curcumin Alone and Combined With PI3K Inhibitors Elicits Positive Effects on Oropharyngeal Cancer Cell Lines Regardless of HPV Status.

Human papillomavirus positive (HPV+) oropharyngeal squamous cell carcinoma (OPSCC) is rising in incidence. Compared to HPV-negative (HPV-) OPSCC, HPV+ cases have a better 5-year survival. With its severe side-effects, today's chemoradiotherapy has not improved outcome compared to radiotherapy alone, so new therapies are needed. Mutations in phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), fibroblast growth factor receptor 3 (FGFR3) and cell division cycle 27 (CDC27) are found in HPV+ OPSCC, and in vitro targeted therapy combining PI3K and FGFR inhibitors showed synergistic effects. Here the effects of targeting CDC27 with curcumin with/without various inhibitors or cisplatin on OPSCC cell lines were examined. Curcumin was administered to HPV+ OPSCC cell lines CU-OP-2, CU-OP-3 and CU-OP-20, and HPV- CU-OP-17 with/without PI3K, cyclin-dependent kinase 4/6, FGFR, poly (ADP-ribose) polymerase or WEE1 inhibitors (BYL719, PD-0332991, JNJ-42756493, BMN-673 and MK-1775, respectively), or cisplatin. The cell lines were then assessed for 72 h after treatment for viability, proliferation and cytotoxicity. Curcumin led to dose-dependent responses with reduced viability and proliferation; upon combining it with BYL719, additional positive effects were found for most OPSCC lines grown as monolayers, and these effects were validated in CU-OP-2 cells grown as spheroids. Curcumin with MK-1775 or PD-0332991 also elicited some positive effects on CU-OP-2 and CU-OP-17 cells. Curcumin alone led to dose-dependent responses and when combined with BYL719, positive effects were revealed, as they were when it was combined with MK-1775 or PD-0332991, suggesting a potential use of some of these combinations for HPV+ OPSCC.

Relating pre-treatment non-Gaussian intravoxel incoherent motion diffusion-weighted imaging to human papillomavirus status and response in oropharyngeal carcinoma

Background and purposeDiffusion-weighted imaging (DWI) is a promising technique for response assessment in head-and-neck cancer. Recently, we optimized Non-Gaussian Intravoxel Incoherent Motion Imaging (NG-IVIM), an extension of the conventional apparent diffusion coefficient (ADC) model, for the head and neck. In the current study, we describe the first application in a group of patients with human papillomavirus (HPV)-positive and HPV-negative oropharyngeal squamous cell carcinoma. The aim of this study was to relate ADC and NG-IVIM DWI parameters to HPV status and clinical treatment response. Materials and methodsThirty-six patients (18 HPV-positive, 18 HPV-negative) were prospectively included. Presence of progressive disease was scored within one year. The mean pre-treatment ADC and NG-IVIM parameters in the gross tumor volume were compared between HPV-positive and HPV-negative patients. In HPV-negative patients, ADC and NG-IVIM parameters were compared between patients with and without progressive disease. ResultsADC, the NG-IVIM diffusion coefficient D, and perfusion fraction f were significantly higher, while pseudo-diffusion coefficient D* and kurtosis K were significantly lower in the HPV-negative compared to HPV-positive patients. In the HPV-negative group, a significantly lower D was found for patients with progressive disease compared to complete responders. No relation with ADC was observed. ConclusionThe results of our single-center study suggest that ADC is related to HPV status, but not an independent response predictor. The NG-IVIM parameter D, however, was independently associated to response in the HPV-negative group. Noteworthy in the opposite direction as previously thought based on ADC.

Treatment Delay and HPV Status on OPSCC With Upfront Surgery: Analysis of National Cancer Database.

Evaluate the effect of treatment delay on survival in human papillomavirus (HPV)-positive and HPV-negative oropharyngeal squamous cell carcinoma (OPSCC) patients undergoing primary surgical resection. Retrospective cohort study using the 2010-2017 National Cancer Database. Multicenter database study. Patients >18 years old with OPSCC and known HPV status, treated surgically with or without postoperative radiation/chemotherapy were included. Two cohorts based on HPV status were grouped by time to treatment initiation (TD-TI, ≤30, 31-60, ≥61 days) and surgery to radiotherapy (TS-RT, ≤42, 43-66, ≥67 days). Univariate, Kaplan-Meier, and multivariate analyses assessed correlations between demographic and clinical factors with overall survival in treatment delay groups. Included were 1643 HPV-positive OPSCC patients and 391 HPV-negative OPSCC patients. No associations between survival and gender, age, race, insurance, or radiotherapy length were observed. Regardless of HPV status, larger tumor size (>2 cm) and lymphovascular invasion predicted worse survival. HPV negative patients with >4 lymph nodes involved had 2.5×greater mortality risk (P = .039). Robotic surgery was associated with improved survival only in HPV positive patients (hazard ratio [HR]: 0.41, P < .001). In HPV positive patients, higher TD-TI related to lower mean survival, although this was not significant on multivariate analysis. HPV negative patients with >42 days of TS-RT had decreased survival (43-66 days, HR 1.63, P = .049; ≥67 days, HR 2.10, P = .032). Longer TS-RT was associated with lower overall survival in HPV negative patients. Treatment delay was not associated with survival in HPV positive OPSCC according to multivariate analysis. These findings enhance knowledge about treatment delay effects in OPSCC, aiding providers in decisions and patient communication.

Association of adjuvant radiation and survival in human papilloma virus-positive oropharynx squamous cell carcinoma with lymphovascular invasion as the sole adverse pathologic feature.

Postoperative radiotherapy radiation therapy (PORT) for early-stage human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) with positive lymphovascular invasion (LVI) has an unclear association with overall survival (OS). This retrospective cohort study queried the National Cancer Database for surgically treated, T1-2, N0-1 HPV+ OPSCC from 2010 to 2019. Primary exposures were LVI and PORT, and the main outcome was 5-year OS. Odds ratios and hazard ratios (HR) with 95% confidence intervals (CIs) were generated using multivariable models and Cox proportional hazard models, respectively. Of 2768 patients, average age was 59.3 years, 2207 (79.7%) were male, and 386 (13.9%) had LVI. Of patients with LVI as their sole adverse pathologic feature, 220 (57.0%) received PORT, which was not associated with 5-year OS (HR, 1.13; CI, 0.65-1.19). Patients with surgically treated, early-stage HPV+ OPSCC and positive LVI as their only pathologic adverse feature may not require PORT.

Human papillomavirus (HPV) load is higher in HPVDNA/p16 positive than in HPVDNA positive/p16 negative oropharyngeal squamous cell carcinoma but does not differ significantly between various subsites or correlate to survival

ObjectivePatients with human papillomavirus DNA positive (HPVDNA+) and p16ink4a overexpressing (p16+) oropharyngeal squamous cell carcinoma (OPSCC), especially those with cancer in the tonsillar and base of tongue subsites as compared to other OPSCC subsites have a better outcome than those with only HPVDNA+ or only p16+ cancer. Likewise having a high viral load has been suggested to be a positive prognostic factor. We therefore hypothesized, that HPV viral load could vary depending on OPSCC subsite, as well as with regard to whether the cancer was HPVDNA+ and p16+, or only HPVDNA+, or only p16+ and that this affected outcome. Material and methodsTo address these issues HPV viral load was determined by HPV digital droplet (dd) PCR in tumor biopsies with previously known HPVDNA/p16 status from 270 OPSCC patients diagnosed 2000–2016 in Stockholm, Sweden. More specifically, of these patients 235 had HPVDNA+/p16+, 10 had HPVDNA+/p16-, 13 had HPVDNA-/p16+ and 12 had HPVDNA-/p16- cancer. ResultsWe found that HPVDNA+/p16+ OPSCC had a significantly higher viral load than HPVDNA+/p16- OPSCC. Moreover, there was a tendency for a higher viral load in the tonsillar and base of tongue OPSCC subsites compared to the other subsites and for a low viral load to correlate to a better clinical outcome but none of these tendencies reached statistical significance. ConclusionTo conclude, the mean viral load in HPVDNA+/p16+ OPSCC was higher than in HPVDNA+/p16- OPSCC, but there was no statistically significant difference in viral load depending on OPSCC subsite or on clinical outcome.

Curative Radiotherapy for Oropharyngeal Squamous Cell Carcinoma Stratified by p16 Status, According to the Eighth Edition American Joint Committee on Cancer Staging Manual: A Retrospective Study.

The prevalence of oropharyngeal squamous cell carcinoma (OPSCC) is increasing worldwide. This retrospective study aimed to investigate the clinical outcomes of patients with OPSCC undergoing definitive radiotherapy, stratified according to their p16 status. A retrospective analysis was conducted on consecutive patients with OPSCC treated with curative external beam radiotherapy between May 2015 and September 2023. Clinical staging was determined by the eighth edition AJCC staging manual for p16 positive and negative OPSCC. All patients were treated with radiotherapy using a simultaneous integrated boost (SIB) with helical tomotherapy. The fractionation scheme, with or without chemotherapy, for the primary site and nodal lesions consisted of 2 Gy per fraction for a total dose of 70 Gy in 35 fractions over seven weeks. This study included 76 patients with a median age of 66 years. With a median follow-up time of 32.6 months, the 3-year progression-free survival rate was significantly higher in p16 positive patients compared to p16 negative patients (79.6% vs. 42.5%, p<0.001). Concerning 54 patients with p16-positive tumors, the overall survival rates indicated excellent clinical outcomes for stage I, II, and III with results of 100%, 100%, and 88.1%, respectively. This retrospective study revealed the clinical outcomes of patients with OPSCC treated with radical radiotherapy, emphasizing the significance of p16 status. While acknowledging the limitations of the retrospective nature of this study, future prospective studies with larger cohorts and extended follow-up periods are needed to enhance evidence quality.

INHBA is Enriched in HPV-negative Oropharyngeal Squamous Cell Carcinoma and Promotes Cancer Progression.

Patients with HPV-negative OPSCC have a poorer prognosis due to distinct molecular pathways. This study reveals significant transcriptomic differences between HPV-negative and HPV-positive OPSCC, identifying INHBA as a key upregulated gene in HPV-negative OPSCC's oncogenic pathways. INHBA is crucial in promoting EMT, cell proliferation, and an immunosuppressive tumor environment, suggesting its potential as a therapeutic target for HPV-negative OPSCC.

Could MMP3 and MMP9 Serve as Biomarkers in EBV-Related Oropharyngeal Cancer.

The high incidence of, and mortality from, head and neck cancers (HNCs), including those related to Epstein-Barr virus (EBV), constitute a major challenge for modern medicine, both in terms of diagnosis and treatment. Therefore, many researchers have made efforts to identify diagnostic and prognostic factors. The aim of this study was to evaluate the diagnostic usefulness of matrix metalloproteinase 3 (MMP 3) and matrix metalloproteinase 9 (MMP 9) in EBV positive oropharyngeal squamous cell carcinoma (OPSCC) patients. For this purpose, the level of these MMPs in the serum of patients with EBV-positive OPSCC was analyzed in relation to the degree of histological differentiation and TNM classification. Our research team's results indicate that the level of both MMPs is much higher in the EBV positive OPSCC patients compared to the EBV negative and control groups. Moreover, their levels were higher in more advanced clinical stages. Considering the possible correlation between the level of MMP 3, MMP 9 and anti-EBV antibodies, and also viral load, after statistical analysis using multiple linear regression, their high correlation was demonstrated. The obtained results confirm the diagnostic accuracy for MMP 3 and MMP 9. Both MMPs may be useful in the diagnosis of EBV positive OPSCC patients.

Clinical Predictors of Cisplatin Chemoradiation-Induced Ototoxicity in HPV-Positive Oropharyngeal Squamous Cell Carcinoma: A Case-Control Study.

Cisplatin-based chemoradiation is a standard treatment for many patients with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC), an etiologically distinct subset of head and neck cancer. Although associated with good long-term survival, clinical risk factors for ototoxicity have been understudied in this population. This study aimed to evaluate clinical predictors associated with ototoxicity in HPV-positive OPSCC patients treated with cisplatin chemoradiation. This retrospective case-control study included 201 adult patients (>18 years) with histologically confirmed HPV-positive OPSCC who received cisplatin chemoradiation as their primary treatment from 2001 and 2019 at a single tertiary cancer center. Ototoxicity was determined using baseline and follow-up audiometry and the Common Terminology Criteria for Adverse Events v5.0 grading criteria (Grade ≥2). Multivariable logistic regression [adjusted odds ratio (aOR)] identified significant predictors that increased the odds of ototoxicity. A total of 201 patients [165 males; median (IQR) age, 57 (11) years] were included in the study. The incidence of ototoxicity in the worst ear was 56.2%, with the greatest hearing loss occurring at high frequencies (4-8 kHz), resulting in a loss of 12.5 dB at 4 to 6 kHz and 20 dB at 6 to 8 kHz. High-dose cisplatin administration compared to weekly administration [aOR 4.93 (95% CI: 1.84-14.99), P = .003], a higher mean cochlear radiation dose [aOR 1.58 (95% CI: 1.12-2.30), P = .01], smoking history [aOR 2.89 (95% CI: 1.51-5.63), P = .001], and a 10 year increase in age [aOR 2.07 (95% CI: 1.25-3.52), P = .006] were each independently associated with increased odds of ototoxicity. Clinical predictors of ototoxicity in HPV-positive OPSCC patients treated with cisplatin-based chemoradiation include the use of a high-dose cisplatin regimen, higher cochlear radiation doses, a history of smoking, and older age. With the rising incidence of this malignancy in Western countries and overall improved survivorship, our research motivates future studies into risk stratification and earlier interventions to mitigate and reduce the risk of ototoxicity.

Mechanisms of crosstalk between the oropharyngeal microbiome and human papillomavirus in oropharyngeal carcinogenesis: a mini review.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer globally. Notably, human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) is on the rise, accounting for 70% of all OPSCC cases. Persistent high-risk HPV infection is linked to various cancers, but HPV infection alone is not sufficient to cause cancer. Advances in next-generation sequencing have improved our understanding of changes in the human microbiome of cancerous environments. Yet, there remains a dearth of knowledge on the impact of HPV-microbiome crosstalk in HPV-positive OPSCC. In this review, we examine what is known about the oropharyngeal microbiome and the compositional shifts in this microbiome in HPV-positive OPSCC. We also review potential mechanisms of crosstalk between HPV and specific microorganisms. Additional research is needed to understand these interactions and their roles on cancer development and progression.

Analysis of cells of epithelial, connective tissue and immune differentiation in HPV-positive-, HPV-negative oropharyngeal carcinoma and normal oropharyngeal tissue by immunofluorescence multiplex image cytometry: a preliminary report

BackgroundEpithelial, connective tissue and immune cells contribute in various ways to the pathophysiology of HPV positive (HPV+) and HPV negative (HPV-) oropharyngeal squamous cell carcinoma (OPSCC). We aimed to investigate the abundance of these cell lineages and their coexpression patterns in patients with HPV + and HPV- OPSCC.MethodsWe used a 4-channel immunofluorescence-microscopy technique for the simultaneous detection of three direct-conjugated antibodies (pancytokeratin, vimentin and CD45/CD18) and DAPI (4’,6-Diamidin-2-phenylindole) in formalin fixed paraffin-embedded tissue samples (FFPE) of patients with HPV + and HPV- OPSCC, and of control patients. Image acquisition and analysis were performed with TissueFAXS and StrataQuest (TissueGnostics, Vienna, Austria), respectively, in tumor cell clusters/stroma in OPSCC specimens and epithelial layer/lamina propria in control specimens. Cell populations were created based on antibodies’ coexpression patterns. Isotype and positive controls were examined for plausibility.ResultsThe proportion of cells of epithelial differentiation in tumor cell clusters was higher in HPV + OPSCC (55%) than in HPV- OPSCC samples (44%). The proportion of connective tissue cells in tumor cell cluster was lower in HPV + OPSCC patients (18%) than in HPV- OPSCC patients (26%). The proportion of immune cells in tumor cell clusters was higher in HPV + OPSCC patients (25%) than in HPV- OPSCC patients (18%). The percentage of anaplastic, potentially de-differentiated cells, was 2% in control patients, and it was higher in HPV- OPSCC (21%) than in HPV + OPSCC samples (6%).ConclusionsThis study provided the first quantitative data for the abundance of cells of epithelial, connective tissue and immune differentiation, in patients with OPSCC and control patients. The abundance of these different crucial cell populations was consistently originating from the same tissue sample. De-differentiation of tumor cells was higher in HPV- OPSCC than in HPV + OPSCC. In tumor cells clusters, the antitumoral host immune response was higher in HPV + OPSCC than in HPV- OPSCC, whereas the fibroblast response was higher in HPV- OPSCC than in HPV + OPSCC. This study contributed to the understanding of histopathologic differences between HPV + OPSCC and HPV- OPSCC patients.

Circulating tumor-tissue modified HPV DNA testing in the clinical evaluation of patients at risk for HPV-positive oropharynx cancer: The IDEA-HPV study

ObjectivesWhile survival outcomes are favorable for Human Papillomavirus (HPV)-positive oropharyngeal squamous cell carcinomas (OPSCCs), early diagnosis may minimize treatment-related morbidity and mortality. This study evaluated circulating tumor tissue-modified viral (TTMV)-HPV DNA plasma testing to facilitate early diagnosis of HPV-positive OPSCCs. MethodsIn this prospective exploratory cohort study, patients presenting to an Otolaryngology-Head and Neck Surgery clinic with unexplained signs or symptoms considered high-risk for HPV-positive OPSCC were recruited between March 2021-October 2022. Circulating TTMV-HPV DNA testing was performed, and results were shared with subjects and treating clinicians. Clinicians were surveyed regarding the perceived clinical utility of the test. ResultsThirty-nine subjects were included. Most subjects were women (N = 23, 59 %), white (N = 32, 82 %) and never-smokers (N = 20, 51 %) with median age 60 years. Circulating TTMV-HPV DNA was detected in 2/39 subjects, both subsequently diagnosed with HPV-positive OPSCC. Both were white men aged 70–80 years with a neck mass. One subject with undetectable TTMV-HPV DNA was also diagnosed with HPV-positive OPSCC through excisional neck mass biopsy. Other eventual diagnoses included 3 HPV-negative head and neck squamous cell carcinomas and 4 other malignancies. Testing was perceived as helpful in clinical decision-making for 26/38 (68 %) subjects, and useful for similar future patients for 32/37 (86 %) subjects. ConclusionCirculating TTMV-HPV DNA testing is feasible and holds potential as a diagnostic aid for HPV-positive OPSCC alongside standard clinical workup. Clinicians should be cognizant of its limitations, as a negative test does not necessarily indicate the absence of disease. Further studies to evaluate its utility are warranted.

Clarification of the margin status by the multidisciplinary tumor board following transoral robotic surgery for p16 positive oropharyngeal squamous cell carcinoma.

Margin status interpretation following transoral robotic surgery (TORS) for oropharyngeal squamous cell carcinoma (OPSCC) is challenging. This study aims to assess the discrepancy between status of margins as reported by the pathologist versus as determined by multi-disciplinary team review (MDTB). A retrospective study of 57 patients with OPSCC who underwent TORS from January 2010 to December 2016 was conducted. Our primary outcome measure was the discrepancy between the surgical specimen margins as described in the pathology report versus final margin status that was determined after the multi-disciplinary team discussion. Fisher's exact test was used. Based on the pathologist-report, 29 subjects (51%) had positive margins, compared to 2 (4%) after multi-disciplinary team discussion. Receipt of chemotherapy correlated with final margin status as determined by MDTB, not with initial main specimen margins (p = .02 and p = .08, respectively). With a median follow up of 28.4 months, two subjects (4%) had loco-regional recurrence. Following TORS, there was a significant discrepancy between status of margins as reported by the pathologist versus as determined by MDTB review. Chemotherapy was avoided in 93.1% of cases that were originally reported as positive margins by the pathologist with an acceptably low recurrence rate. 4.

Loss of LRP1B expression drives acquired chemo and radio-resistance in HPV-positive head and neck cancer

ObjectivesAlthough human papillomavirus positive (HPV+) oropharyngeal squamous cell carcinoma (OPSCC) patients typically experience excellent survival, 15–20 % of patients recur after treatment with chemotherapy and radiation. Therefore, there is a need for biomarkers of treatment failure to guide treatment intensity. Materials and methodsWhole genome sequencing was carried out on HPV+OPSCC patients who were primarily treated with concurrent chemotherapy (cisplatin) and radiation. We then explored whether the loss of LRP1Bwas sufficient to drive an aggressive phenotype, and promote a resistance to cisplatin and radiation therapy both in vitro using HPV+ cell lines (93VU147T, UMSCC47, UWO37 and UWO23) and in vivo. ResultsThrough integrative genomic analysis of three HPV+OPSCC tumour datasets, we identified that deletion of LRP1B was enriched in samples that recurred following chemo-radiation. Knockdown using siRNA in four HPV+ cell lines (UWO23, UWO37, UMSCC47 and 93VU147T) resulted in increased proliferation of all cases. CRISPR/Cas9 deletion of LRP1B in the same cell line panel demonstrated increased proliferation, clonogenic growth and migration, as well as resistance to both cisplatin and radiation in LRP1B deleted cells compared to their respective non-targeting control cells. Cell line derived xenograft studies indicated that the LRP1B knockout tumours were more resistant to cisplatin and radiation therapy compared to their controls invivo. ConclusionTaken together, our work implicates LRP1B deletion as a potential biomarker for identifying treatment resistant HPV+ OPSCC cases.

The effectiveness of a dual-component regimen of induction chemotherapy in the treatment of HPV-positive oropharyngeal squamous cell carcinoma

Given the favorable prognosis associated with HPV-positive oropharyngeal cancer, cancer care professionals are actively discussing the feasibility of de-escalating treatment strategy for this cohort of patients.The purpose of the study was to improve the treatment outcomes in patients with locally advanced HPV- positive oropharyngeal squamous cell carcinoma by identifying the optimal induction chemotherapy (ICH) regimen, in particular, by using a two-drug ICH.Material and Methods. The study included 27 patients with locally advanced (T3–4N0–1, or T1–4N2–3) P16-positive oropharyngeal squamous cell carcinoma. All patients received 3 cycles of ICT according to the TP (docetaxel + cisplatin) regimen. To assess the effect and toxicity of two-drug ICH, a control group of patients, who received ICH according to the standard three-drug TPF (docetaxel, cisplatin and 5FU) regimen, was formed.Results. In the TP group, complete response (CR) was achieved in 3 (11 %) patients, partial response (PR) in 17 (63 %), stable disease (SD) in 7 (26 %) patients. In the control group: CR 4 (20 %) patients, PR – 13 (60 %), SD – 4 (20 %) patients. The median follow-up time in the TP group was 9 months (range: 2 to 22 months). The 1-year progression-free (PFS) and overall survival (OS) rates were 88.2 % and 100 %, respectively. In the TPF group, the PFS and OS rates were 84.4 % and 100 %, respectively.Conclusion. The results revealed that TP ICH regimen was non-inferior to the standard TPF regimen in the rates of the objective response, 1-year OS and PFS.

Abstract LB252: HPV integration events are heterogenous, clonally selected and associated with spatially distinct transcriptomic profiles in aggressive HPV positive oropharyngeal squamous cell carcinoma

Abstract HPV positive Oropharyngeal Squamous Cell Carcinoma (HPV positive OPSCC) is rising in incidence and, for patients that recur, survival is extremely poor. Recent technological advances have led to a better understanding of patterns of HPV integration in primary tumors but have not yet tested the evolution of these events in deadly forms of the disease. Therefore, we performed multi-region sequencing analysis on a cohort of 34 recurrent and metastatic patients with HPV positive OPSCC (29% survival rate) plus one recurrent HPV positive model pair, UPCI-90 and UPCI-152. We then used our recently developed capture-based sequencing HPV integration caller and assembler pipeline, SearcHPV, to identify patterns of viral integration into the host genome and Oxford Nanopore long-read whole genome sequencing to validate the integration events in selected cases. In total, 1591 HPV integrations were called and assembled. Unexpectedly, our analysis revealed that HPV integration events are heterogenous as multiple independent integration events were observed in different spatial regions of the same tumor. Further, when comparing independent and anatomically distinct regions of recurrence from the same patient, independent integration events were also observed. Heterogeneity of integration structures was found to include variation of local HPV copy number, distribution of the specific HPV integration sites within the structures as well as distinct characteristics of rearrangement breakpoints (human-HPV, HPV-HPV, human-human). Given this observation, we then evaluated the hypothesis that HPV integration events could be clonally selected by comparing the integration profiles from a cohort of matched primary and recurrent tumors. In this cohort, 64 integration events (11 exact same; 53 clustered in the same genes) were maintained throughout tumor progression to recurrence, while other integration events were lost, supporting a model of clonal evolution with driver and passenger integrations. We then explored this further using one recurrent HPV positive model pair, UPCI-90 and UPCI-152, using long-read whole genome sequencing which validated the heterogeneity and clonal selection of HPV integration events. Finally, to test if differences in HPV integration content in spatially distinct regions could drive spatially distinct phenotypic differences, we performed multi-region RNA-seq analysis on 8 HPV16 positive OPSCCs, which demonstrated spatial differences in HPV16 E7 expression, that were validated with RNAscope assays, as well as statistically significant spatial enrichment of independent oncogenic gene sets. Collectively, we report evidence showing that HPV integration events are heterogenous, clonally selected during progression and associated with distinct transcriptomic profiles. Based on this data, we now propose a new working model of disease pathogenesis in which heterogeneity and clonal evolution of HPV integration events serve as a critical driver of the disease process. Citation Format: Wenjin Gu, Apurva Bhangale, Collin V. Brummel, Elizabeth Gensterblum-Miller, Matthew E. Spector, Ryan E. Mills, Chad Brenner. HPV integration events are heterogenous, clonally selected and associated with spatially distinct transcriptomic profiles in aggressive HPV positive oropharyngeal squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB252.