HPV-negative Head And Neck Cancers Research Articles

Therapeutic Advances and Challenges for the Management of HPV-Associated Oropharyngeal Cancer.

The use of conventional chemotherapy in conjunction with targeted and immunotherapy drugs has emerged as an option to limit the severity of side effects in patients diagnosed with head and neck cancer (HNC), particularly oropharyngeal cancer (OPC). OPC prevalence has increased exponentially in the past 30 years due to the prevalence of human papillomavirus (HPV) infection. This study reports a comprehensive review of clinical trials registered in public databases and reported in the literature (PubMed/Medline, Scopus, and ISI web of science databases). Of the 55 clinical trials identified, the majority (83.3%) were conducted after 2015, of which 77.7% were performed in the United States alone. Eight drugs have been approved by the FDA for HNC, including both generic and commercial forms: bleomycin sulfate, cetuximab (Erbitux), docetaxel (Taxotere), hydroxyurea (Hydrea), pembrolizumab (Keytruda), loqtorzi (Toripalimab-tpzi), methotrexate sodium (Trexall), and nivolumab (Opdivo). The most common drugs to treat HPV-associated OPC under these clinical trials and implemented as well for HPV-negative HNC include cisplatin, nivolumab, cetuximab, paclitaxel, pembrolizumab, 5-fluorouracil, and docetaxel. Few studies have highlighted the necessity for new drugs specifically tailored to patients with HPV-associated OPC, where molecular mechanisms and clinical prognosis are distinct from HPV-negative tumors. In this context, we identified most mutated genes found in HPV-associated OPC that can represent potential targets for drug development. These include TP53, PIK3CA, PTEN, NOTCH1, RB1, FAT1, FBXW7, HRAS, KRAS, and CDKN2A.

Overview of head and neck cancer and the role of survivorship care

Head and neck cancer (HNC) accounted for 4% of all malignancies in 2022 and is often associated with tobacco and alcohol use, but the incidence of human papillomavirus (HPV)–associated HNC has steadily increased in recent decades. Unlike HPV-negative HNC, HPV-positive HNC has a favorable prognosis and survival is significantly better resulting in a large population of HNC survivors. Survivorship for HNC is of the utmost importance as there is a high burden of side effects that impact function and quality of life, and the patients with HPV HNC are living longer with the sequelae of their treatment.

Blood and saliva-derived ctDNA is a marker of residual disease after treatment and correlates with recurrence in human papillomavirus-associated head and neck cancer.

There is an alarming increase in human papillomavirus-associated head and neck cancer (HNC), reaching epidemic levels. While patient prognosis is generally good, off-target treatment effects are associated with decreased quality of life. Thus, non-invasive strategies to predict treatment response and risk of recurrence could help de-escalate treatment. In this study, we tested circulating tumor (ct)DNA in liquid biopsies (blood/saliva) of HPV-positive HNC patients to assess treatment response and disease progression. A total of 235 blood and saliva samples were collected from 60 HPV-positive and 17 HPV-negative HNC patients (control group) before and/or after treatment. Samples were analyzed using ddPCR for HPV16/18/31/33/35/45 and correlated with imaging and pathological examination. HPV-ctDNA detection was significantly higher prior to treatment (91%) than after treatment (8.0%) (χ2 p < 0.00001), with high concordance between saliva and blood (93%). In matched samples, all patients positive for ctDNA before treatment showed significant reductions in ctDNA levels post treatment (p < 0.0001). All but one patient with persistent ctDNA after treatment showed residual tumor and subsequent recurrence. Finally, fragmentomic analysis revealed shifts in cell-free DNA fragment size after treatment, suggesting a complementary biomarker for treatment response. Blood and saliva were found to be good sources of HPV-ctDNA. The presence of ctDNA strongly correlated with treatment response, demonstrating clinical utility as a non-invasive biomarker to monitor tumor progression in HPV-positive HNC. Liquid biopsy based ctDNA testing could be an effective approach to predict recurrence and stratify patients for de-escalation of treatment, thereby improving quality of life.

Examining the effect of Medicaid expansion on early detection of head and neck cancer of the oral cavity and pharynx by HPV-type and generosity of dental benefits.

Over a decade of evidence supports the claim that increased access to insurance through Medicaid expansions improves early detection of cancer. Yet, evidence linking Medicaid expansions to early detection of head and neck cancers (HNC) of the oral cavity and pharynx, specifically, may be limited by the lack of attention to Human Papillomavirus (HPV) etiology, generosity of dental coverage, and valid inference analyzing state cancer registry data. This studyreexamined the effect of Medicaid expansion on early detection of HPV+/- HNC in states offering extensive dental benefits. Specialized data from the Surveillance, Epidemiology, and End Results (SEER) program was analyzed to account for, previously unmeasurable, differential detection patterns of HNCs associated with HPV. Then, to identify the effect of increasing Medicaid eligibility on staging patterns in states offering extensive benefits amidst potentially non-common trends between states, a "Triple Differences" design identifies the differential effect of Medicaid Expansion (with dental coverage) on HPV-negative HNCs relative to the change in HPV-positive HNCs. For valid inference analyzing a small number of state clusters (12) in cancer registry data, each regression model applies a Wild Cluster Bootstrap. Expanding Medicaid eligibility was found to be associated with a decrease in the proportion of distant-stage diagnoses of HPV(-) HNCs, but only among states which increased Medicaid dental generosity at the time of Medicaid expansion. These results suggest that adding extensive Medicaid dental benefits was the primary mechanism impacting HNC detection. This study highlights the potential positive spillover effects of policies which increase access to public dental coverage for low-income adults, while also showing the limitation of access to dental services for improving early detection of HPV+ HNCs.

Use of 3D Spheroid Models for the Assessment of RT Response in Head and Neck Cancer.

Radiotherapy (RT) is a key player in the treatment of head and neck cancer (HNC). The RT response, however, is variable and influenced by multiple tumoral and tumor microenvironmental factors, such as human papillomavirus (HPV) infections and hypoxia. To investigate the biological mechanisms behind these variable responses, preclinical models are crucial. Up till now, 2D clonogenic and in vivo assays have remained the gold standard, although the popularity of 3D models is rising. In this study, we investigate the use of 3D spheroid models as a preclinical tool for radiobiological research by comparing the RT response of two HPV-positive and two HPV-negative HNC spheroid models to the RT response of their corresponding 2D and in vivo models. We demonstrate that HPV-positive spheroids keep their higher intrinsic radiosensitivity when compared to HPV-negative spheroids. A good correlation is found in the RT response between HPV-positive SCC154 and HPV-negative CAL27 spheroids and their respective xenografts. In addition, 3D spheroids are able to capture the heterogeneity of RT responses within HPV-positive and HPV-negative models. Moreover, we demonstrate the potential use of 3D spheroids in the study of the mechanisms underlying these RT responses in a spatial manner by whole-mount Ki-67 and pimonidazole staining. Overall, our results show that 3D spheroids are a promising model to assess the RT response in HNC.

Impact of Secretion-Active Osteoblast-Specific Factor 2 in Promoting Progression and Metastasis of Head and Neck Cancer.

Simple SummaryHead and neck cancers (HNC) exhibit poor survival due to metastases. Our study identified osteoblast-specific factor 2 (OSF-2) as overexpressed in primary tumors, lymph node metastases, and the tumor microenvironment. High OSF-2 levels correlate with metastatic disease and reduced survival of HPV-negative HNC patients. OSF-2’s active secretion signal seems to promote metastases by supporting the tumor microenvironment via the ß1 integrin-induced PI3K and Akt/PKB signaling pathway. We suggest OSF-2 as a potential biomarker and drug target to control (HPV-negative) HNC metastasis and disease management.Treatment success of head and neck cancer (HNC) is still hampered by tumor relapse due to metastases. Our study aimed to identify biomarkers by exploiting transcriptomics profiles of patient-matched metastases, primary tumors, and normal tissue mucosa as well as the TCGA HNC cohort data sets. Analyses identified osteoblast-specific factor 2 (OSF-2) as significantly overexpressed in lymph node metastases and primary tumors compared to normal tissue. High OSF-2 levels correlate with metastatic disease and reduced overall survival of predominantly HPV-negative HNC patients. No significant correlation was observed with tumor localization or therapy response. These findings were supported by the fact that OSF-2 expression was not elevated in cisplatin-resistant HNC cell lines. OSF-2 was strongly expressed in tumor-associated fibroblasts, suggesting a tumor microenvironment-promoting function. Molecular cloning and expression studies of OSF-2 variants from patients identified an evolutionary conserved bona fide protein secretion signal (1MIPFLPMFSLLLLLIVNPINA21). OSF-2 enhanced cell migration and cellular survival under stress conditions, which could be mimicked by the extracellular administration of recombinant protein. Here, OSF-2 executes its functions via ß1 integrin, resulting in the phosphorylation of PI3K and activation of the Akt/PKB signaling pathway. Collectively, we suggest OSF-2 as a potential prognostic biomarker and drug target, promoting metastases by supporting the tumor microenvironment and lymph node metastases survival rather than by enhancing primary tumor proliferation or therapy resistance.

Proton Image-guided Radiation Assignment for Therapeutic Escalation via Selection of locally advanced head and neck cancer patients [PIRATES]: A Phase I safety and feasibility trial of MRI-guided adaptive particle radiotherapy

IntroductionRadiation dose-escalation for head and neck cancer (HNC) patients aiming to improve cure rates is challenging due to the increased risk of unacceptable treatment-induced toxicities. With “Proton Image-guided Radiation Assignment for Therapeutic Escalation via Selection of locally advanced head and neck cancer patients” (PIRATES), we present a novel treatment approach that is designed to facilitate dose-escalation while minimizing the risk of dose-limiting toxicities for locally advanced HPV-negative HNC patients. The aim of this Phase I trial is to assess the safety & feasibility of PIRATES approach. MethodsThe PIRATES protocol employs a multi-faceted dose-escalation approach to minimize the risk of dose-limiting toxicities (DLTs): 1) sparing surrounding normal tissue from extraneous dose with intensity-modulated proton therapy, 2) mid-treatment hybrid hyper-fractionation for radiobiologic normal tissue sparing; 3) Magnetic Resonance Imaging (MRI) guided mid-treatment boost volume adaptation, and 4) iso-effective restricted organ-at-risk dosing to mucosa and bone tissues.The time-to-event Bayesian optimal interval (TITE-BOIN) design is employed to address the challenge of the long DLT window of 6 months and find the maximum tolerated dose. The primary endpoint is unacceptable radiation-induced toxicities (Grade 4, mucositis, dermatitis, or Grade 3 myelopathy, osteoradionecrosis) occurring within 6 months following radiotherapy. The second endpoint is any grade 3 toxicity occurring in 3–6 months after radiation. DiscussionThe PIRATES dose-escalation approach is designed to provide a safe avenue to intensify local treatment for HNC patients for whom therapy with conventional radiation dose levels is likely to fail. PIRATES aims to minimize the radiation damage to the tissue surrounding the tumor volume with the combination of proton therapy and adaptive radiotherapy and within the high dose tumor volume with hybrid hyper-fractionation and not boosting mucosal and bone tissues. Ultimately, if successful, PIRATES has the potential to safety increase local control rates in HNC patients with high loco-regional failure risk.Trial registration: ClinicalTrials.gov ID: NCT04870840; Registration date: May 4, 2021.Netherlands Trial Register ID: NL9603; Registration date: July 15, 2021.

Association between human papillomavirus status and health-related quality of life in oropharyngeal and oral cavity cancer survivors

The human papillomavirus (HPV) is a risk factor for a subgroup of head and neck cancers (HNC). HPV-positive and HPV-negative HNC patients encompass heterogeneous groups regarding risk factors, sociodemographic and clinical characteristics, which may influence health-related quality of life (HRQL) differently. Since this has been sparsely studied, our study investigated the association between HPV status and HRQL in HNC survivors in Denmark. This cross-sectional study included 179 recurrence-free oropharyngeal and oral cavity squamous cell carcinoma (OSCC) survivors. HRQL was assessed on the EORTC QLQ-C30 and QLQ-H&N35 questionnaires. Linear and logistic regression models were adjusted for sociodemographic, clinical and lifestyle factors. Most unadjusted results showed better HRQL among HPV-positive (n=119) compared to HPV-negative (n=60) OSCC survivors (average 18months since diagnosis). After adjustments, the HPV-positive survivors reported higher role functioning (mean difference [MD] 9.2, 95% confidence interval [CI] 0.1 to -18.4), and fewer problems with speech (MD -9.0, 95% CI -18.0 to -0.1), sexuality (MD -21.9, 95% CI -38.0 to -5.9) and opening mouth (MD -13.7, 95% CI -26.6 to -0.8) compared to HPV-negative survivors. Our findings support that HPV-positive OSCC survivors experience better HRQL than HPV-negative survivors. However, results indicate that sociodemographic, clinical and lifestyle factors explain most of the association between HPV status and HRQL. Findings suggest increased focus on the HPV-negative OSCC survivors with deteriorated HRQL in rehabilitation programs and future research to investigate the long-term effects of treatment among HPV-positive OSCC survivors who may develop symptoms later in survivorship.

Modulation of the Tumor Microenvironment by Abemaciclib in Operable HPV-Negative Head and Neck Cancer (HNC)

Modulation of the Tumor Microenvironment by Abemaciclib in Operable HPV-Negative Head and Neck Cancer (HNC)

Synergistic antitumour activity of HDAC inhibitor SAHA and EGFR inhibitor gefitinib in head and neck cancer: a key role for \u0394Np63\u03b1

BackgroundEpidermal growth factor receptor (EGFR) overexpression is associated with the development of head and neck cancer (HNC) and represents one of the main therapeutic targets for this disease. The use of EGFR inhibitors has limited efficacy due to their primary and acquired resistance, partially because of increased epithelial to mesenchymal transition (EMT). The HDAC inhibitor SAHA has been shown to revert EMT in different tumours, including HNC. In this study, we investigated the cooperative role of SAHA and the EGFR tyrosine kinase inhibitor gefitinib in both HPV-positive and HPV-negative HNC cell lines.MethodsA panel of 12 HPV-positive and HPV-negative HNC cell lines were screened for cell viability upon treatment with SAHA, gefitinib and the combination of the two. Epithelial/mesenchymal marker expression, as well as activation of signalling pathway, were assessed upon SAHA treatment. ΔNp63α silencing with shRNA lentiviral particles was used to determine its role in cell proliferation, migration and TGFβ pathway activation.ResultsWe found that both SAHA and gefitinib have antitumour activity in both HPV-positive and HPV-negative HNC cell lines and that their combination has a synergistic effect in inhibiting cell growth. SAHA treatment reverts EMT and inhibits the expression of the transcription factor ΔNp63α. Suppression of ΔNp63α reduces EGFR protein levels and decreases cell proliferation and TGFβ-dependent migration in both HPV-positive and HPV-negative HNC cell lines.ConclusionsOur results, by giving a clear molecular mechanism at the basis of the antitumour activity of SAHA in HNC cell lines, provide a rationale for the clinical evaluation of SAHA in combination with gefitinib in both HPV-positive and HPV-negative HNC patients. Further knowledge is key to devising additional lines of combinatorial treatment strategies for this disease.

The mutational landscape of recurrent versus nonrecurrent human papillomavirus-related oropharyngeal cancer.

Human papillomavirus-related (HPV-related) oropharyngeal squamous cell carcinomas (OPSCCs) have an excellent response rate to platinum-based chemoradiotherapy. Genomic differences between primary HPV-related OPSCCs that do or do not recur are unknown. Furthermore, it is unclear if HPV-related OPSCCs that recur share a genomic landscape with HPV-negative head and neck cancers (HNCs). We utilized whole exome sequencing to analyze somatic nucleotide (SNVs) and copy number variants (CNVs) among a unique set of 51 primary HPV-related OPSCCs, including 35 that did not recur and 16 that recurred. We evaluated 12 metachronous recurrent OPSCCs (7 with paired primary OPSCCs) and 33 primary HPV-unrelated oral cavity and OPSCCs. KMT2D was the most frequently mutated gene among primary HPV-related OPSCCs (n = 51; 14%) and among metachronous recurrent OPSCCs (n = 12; 42%). Primary HPV-related OPSCCs that recurred shared a genomic landscape with primary HPV-related OPSCCs that did not recur. However, TSC2, BRIP1, NBN, and NFE2L2 mutations occurred in primary OPSCCs that recurred but not in those that did not recur. Moreover, primary HPV-related OPSCCs that recur harbor features of HPV-unrelated HNCs, notably including MAPK, JAK/STAT, and differentiation signaling pathway aberrations. Metachronous recurrent OPSCCs shared a genomic landscape with HPV-unrelated HNCs, including a high frequency of TP53, CASP8, FAT1, HLA-A, AJUBA, and NSD1 genomic alterations. Overall, primary HPV-related OPSCCs that recur share a genomic landscape with nonrecurrent OPSCCs. Metachronous recurrent OPSCCs share genomic features with HPV-negative HNCs. These data aim to guide future deescalation endeavors and functional experiments. This study is supported by the American Cancer Society (RSG TBG-123653), funding support for RAH (T32DC00018, Research Training in Otolaryngology, University of Washington), funds to EM from Seattle Translational Tumor Research (Fred Hutchinson Cancer Research Center), and center funds from the Fred Hutchinson Cancer Research Center to EM. UD is supported by the Department of Veterans Affairs, Biomedical Laboratory Research and Development (BLR&D), grant IO1-oo23456, and funds from the Pittsburgh Foundation and PNC Foundation.

Targeting Head and Neck Cancer by Vaccination.

Head and neck cancer (HNC) is a heterogeneous group of squamous cell cancers that affect the oral cavity, pharynx, and larynx. Worldwide, it is the sixth most common cancer but in parts of Southern and South-East Asia, HNC is one of the most common cancers. A significant proportion of HNC is driven by human papillomavirus (HPV) infection, whereas HPV-independent HNC is associated with alcohol, smoking, and smokeless tobacco consumption. Here, we review the past and present experience of targeting HNC with vaccination focusing on HPV-derived antigens as well as non-viral antigens for HPV-negative HNC. Novel therapeutic approaches for HNC will focus not only on effective vaccine platforms but will also target the stroma-rich immunosuppressive microenvironment found in those tumours.

Abstract 5898: The consomic xenograft model identifies genetic changes in the tumor microenvironment that alter the growth and metastasis of head and neck cancers

Abstract Background: The tumor microenvironment (TME) is known to impact tumor growth, metastatic potential, and treatment response. Nearly all studies of head and neck cancer (HNC) have focused on somatic mutations in the malignant cells. We hypothesized that genetic determinants limited to the microenvironment would influence HNC growth and metastatic potential. Approach: To demonstrate the impact of genetic differences in the TME on HNC cell line in vivo growth we utilized a novel tool, the consomic xenograft model (CXM). A consomic rat has an entire chromosome substituted into the isogenic background of another inbred strain by selective breeding. Use of immunodeficient (IL2Rγ-/-) consomic rats allows one to study the influence of stromal genetics on tumor biology without the confounding effect of differences in the immune system through the orthotopic implantation of cancer cells into different consomic rat strains. In this system, any differences in tumor growth or metastases are due to differences in the TME rather than cancer cells or immune response. We utilized SS and SS.BN3 consomic rat strains, previously shown to affect the growth of breast tumors, to study the effects of the TME on HNC tumor growth using two well-characterized HPV negative HNC cell lines, SCC-6 (base of tongue derived) and SCC-22b (derived from a hypopharyngeal cancer that had metastasized to lymph nodes). Both cell lines were modified to stably express luciferase. HNC cells were inoculated into the tongue of SS and SS.BN3 animals and tumor growth was monitored by biophotonic imaging after luciferin injection. Results: A significant difference in the tumor growth was seen between rat strains for both cell lines, with the SS.BN3 rats exhibiting less tumor growth and metastasis. Median luciferase activity from baseline increased by 4.1-fold vs. 1.1 fold in SCC-6 tumors in SS vs SS.BN3 rats, respectively (p&amp;lt;0.03). SCC-22B tumors demonstrated a significant difference in tumor size as well, with median luciferase activity from baseline 12.7 vs 4.4 fold on day 26, for SS vs SS.BN3, respectively (p&amp;lt;0.05). A significant differences in lung metastases was also seen between strains. Lung metastases were seen in 88% of SS and 0% of SS.BN3 rats (n=6 per group) inoculated with SCC-6 (p&amp;lt;0.02) and in 75% of SS and 30% of SS.BN3 rats (n=10 per group) inoculated with SCC-22b (p&amp;lt;0.08). Despite a non-statistically significant difference in the number of SCC-22b inoculated animals with lung metastases, there was a significantly higher metastatic burden as measured by luciferase signal, with the median signal 6.8 fold higher in SS as compared to SS.BN3 (p&amp;lt;0.03). Conclusions: The use of the CXM model demonstrates an important role for the TME in the growth and metastatic spread of HNC cell lines. This model allows for future congenic mapping to identify the causative genetic variants in the TME mediating the HNC changes in tumor growth and metastasis. Citation Format: Michael W. Straza, Amy Rymaszewski, Kwangok P. Nickel, Anne Frei, Anirban Chatterjee, Rachel Schlaak, Amit Joshi, Michael Flister, Randy J. Kimple, Carmen Bergom. The consomic xenograft model identifies genetic changes in the tumor microenvironment that alter the growth and metastasis of head and neck cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5898. doi:10.1158/1538-7445.AM2017-5898

Clonal repopulation dynamics in recurrent human papillomavirus-associated head and neck cancer.

e17517 Background: Despite the better prognosis for human papillomavirus (HPV)-associated head and neck cancer (HNC) relative to HPV-negative HNC, 10 to 25% of HPV-associated cases recur within 3 years of completing therapy. We sought to investigate clonal repopulation dynamics in recurrent HPV-related oropharyngeal squamous cell carcinoma (OPSCC). Methods: Deep sequencing ( &gt; 500X) using a CLIA-certified, targeted 262-gene multiplexed assay was performed on DNA extracted from 11 pairs of matched FFPE-preserved primary and subsequent recurrent p16-positive OPSCC samples including two pairs from the University of Washington and nine from the University of Pittsburgh. Results: 82% of patients were male with a median (IQR) age of 62 (15) years. Nine of eleven patients presented with advanced stage disease. Five were initially treated with surgery and concurrent chemoradiation, two with surgery, and four with concurrent chemoradiation. Median (IQR) time to recurrence was 10 (10.5) months. Four patients reported a history of tobacco use. The mean (SD) number of somatic nucleotide variants (SNVs) per tumor was 16.4 (12.2). 42% of SNVs occurred in the primary alone while 28% were unique to recurrences. 28% of SNVs were shared between primary-recurrent pairs. Three patients with distant or regional recurrences had different mutational profiles between the primary and recurrence suggesting repopulation by an early-appearing clone vs sampling error. One patient developed two local recurrences with conserved mutations in addition to de novo mutations in each subsequent recurrence suggesting repopulation by early and late evolving clones. Genes with a substantial functional impact that were frequently mutated in primary tumors include IDH2 and FBXW7. Recurrently mutated genes found in recurrent tumors include TET3, PIK3CA, STK11, and HDAC4. Conclusions: In thistargeted exome sequencing study of matched primary-recurrent OPSCC tumor pairs, we observed clonal repopulation with both transmitted and de novo mutations. Intriguingly, we noted a predilection for mutations involving genes regulating metabolism including IDH2 among primary tumors which has potential implications for precision medicine approaches.

Human Papillomavirus and Head and Neck Cancer: Psychosocial Impact in Patients and Knowledge of the Link – A Systematic Review

Head and neck cancer (HNC) currently affects approximately 11 200 people in the UK, with an increasing proportion known to be caused by the human papillomavirus (HPV). We undertook a systematic review of studies measuring the psychosocial impact of HPV-related HNC and also studies measuring knowledge about the link between HPV and HNC among different populations. Searches were conducted on MEDLINE, Embase, PsycINFO, CINAHL Plus and Web of Science, with reference and forward citation searches also carried out on included studies. Studies were selected if they (i) were original peer-reviewed research (qualitative or quantitative), (ii) mentioned HPV and HNC, (iii) measured an aspect of the psychosocial impact of the diagnosis of HPV-related HNC as the dependent variable and/or (iv) measured knowledge of the association between HPV and HNC. In total, 51 papers met the inclusion criteria; 10 measuring psychosocial aspects and 41 measuring knowledge of the link between HPV and HNC. Quality of life in those with HPV-positive HNC was found to be higher, lower or equivalent to those with HPV-negative HNC. Longitudinal studies found quality of life in patients was at its lowest 2–3 months after diagnosis and some studies found quality of life almost returned to baseline levels after 12 months. Knowledge of the link between HPV and HNC was measured among different populations, with the lowest knowledge in the general population and highest in medical and dental professionals. Due to the limited studies carried out with patients measuring the psychosocial impact of a diagnosis of HPV-positive HNC, future work is needed with the partners of HPV-positive HNC patients and health professionals caring for these patients. The limited knowledge of the association between HPV and HNC among the general population also indicates the need for research to explore the information that these populations are receiving.

Abstract A1-63: Characterizing the DNA damage repair defect in HPV-positive oropharyngeal squamous cell carcinoma

Abstract Although tobacco-associated head and neck cancers (HNCs) are declining in incidence, overall HNC rates are escalating due to increasing prevalence of human papillomavirus (HPV)-associated tumors, especially oropharyngeal squamous cell carcinomas (OPSCCs). Clinically, patients with HPV-associated OPSCCs have improved overall survival and increased response to therapy, especially agents which act by damaging DNA. Based on these observations, we hypothesized that HPV-positive OPSCCs harbor a defect in DNA repair activity and are sensitive to DNA repair-targeted therapy. We evaluated DNA repair activity by immunofluorescent staining for DNA damage-induced formation of DNA repair protein foci. Baseline expression of DNA repair proteins was determined by NanoString nCounter analysis of transcript levels and immunoblotting for protein levels. Therapeutic sensitivity was assessed in vitro using the colony formation assay and in vivo using both cell line-derived xenografts and a patient-derived xenograft. Consistent with our hypothesis, in vitro disease models demonstrated delayed resolution of radiation-induced DNA double strand breaks (DSBs) as assessed by γH2AX foci staining and neutral comet assay. Investigation of the two main DSB repair mechanisms, non-homologous end joining (NHEJ) and homologous recombination (HR), indicated intact activation of both pathways but strikingly diminished recruitment of downstream repair factors DNA-Pk (NHEJ) and BRCA2 (HR) to sites of damage. In addition, protein expression of both DNA-PK and BRCA2 was decreased in HPV-positive compared to HPV-negative HNC cell lines. We next studied susceptibility of HPV-positive OPSCCs to PARP inhibition, a class of anticancer agents which block DNA repair signaling pathways and have proven effective clinically in DNA repair-deficient cancers. In vitro colony formation assays revealed a negative effect on cell survival in HPV-positive but not HPV-negative HNCs treated with the PARP inhibitor veliparib. Importantly, these results were confirmed in vivo in both cell line-derived xenografts and a patient-derived xenograft. In summary, our findings demonstrate for the first time the presence of a significant DNA DSB repair defect in HPV-positive OPSCCs encompassing both NHEJ and HR repair, and suggest therapies targeting DNA repair pathways may help improve therapeutic ratio in this disease. Citation Format: Alice N. Weaver, Tiffiny S. Cooper, Hoa Q. Trummell, James A. Bonner, Eben L. Rosenthal, Eddy S. Yang. Characterizing the DNA damage repair defect in HPV-positive oropharyngeal squamous cell carcinoma. [abstract]. In: Proceedings of the AACR Special Conference on Translation of the Cancer Genome; Feb 7-9, 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 1):Abstract nr A1-63.

Abstract 4899: Small proline rich protein 3 (SPRR3) is a potential mediator of radiation resistance in HPV negative head and neck squamous cell carcinoma

Abstract Purpose: Head and neck cancers (HNCs) are caused by two major etiologies that have significant prognostic significance: tobacco and human papillomavirus (HPV). Patients with HPV-positive HNC have significantly better outcomes than those with HPV-negative HNC. We have previously shown that this difference may be due, in part, to increased sensitivity to radiation in HPV-positive HNCs. In order to identify additional factors that may underlie differences in radiation sensitivity, we sought to understand radiation-induced changes in global gene expression between HPV-positive and HPV-negative HNCs. Methods: A panel of eight cell lines, four HPV positive (UD-SCC2, UM-SCC47, UPCI-SCC90, 93-VU-147T) and four HPV negative (SCC1, SCC6, SCC22B, SCC1483 were treated with radiation (4 Gy) or mock and total RNA isolated 4, 24, and 48 hrs after treatment. Global gene expression was analyzed using Affymetrix Gene 1.1 ST array chips. Comparisons between untreated, baseline control, and post-radiation expression were made using ArrayStar. Alterations in differentially expressed genes were validated by Real-Time quantitative PCR (RT-qPCR). Results: Out of the top 15 hits, several members of the small proline rich repeat (SPRR) family were identified. SPRR1A, SPRR1B, and SPRR3 were significantly upregulated in HPV negative as compared to HPV-positive cell lines. Differences in expression were validated using RT-qPCR in a panel of HNC cell lines. Conclusions: We identified changes in expression of SPRR3 in HPV negative cell lines. This gene is a member of a protein family known to scavenge reactive oxygen species. Its increased expression in HPV negative HNC may mediate resistance to radiation and contribute to the poor outcomes of patients in this cohort. SPRR3 is a potential novel molecular target that could improve the outcomes of patients with HPV negative HNC. Citation Format: Joshua A. Martin, Dana Gunderson, Randall J. Kimple. Small proline rich protein 3 (SPRR3) is a potential mediator of radiation resistance in HPV negative head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4899. doi:10.1158/1538-7445.AM2014-4899

Abstract 73: Preclinical evaluation of chemoradiation with cetuximab HPV-positive head and neck cancer suggests significant radiosensitization.

Abstract PURPOSE: Whether cetuximab, a monoclonal antibody to the epidermal growth factor receptor, should be combined with radiation in patients with human papillomavirus (HPV) positive locally advanced head and neck cancer (HNC) is not known. Currently available data are inadequate to address this important therapeutic question. We sought to examine the ability of cetuximab to act as a radiosensitizer and to examine the mechanism of action of this combination in a model system of HPV+ HNC. METHODS: Four cell lines (UD-SCC2, UM-SCC47, UPCI-SCC90, 93-VU-147T) validated for HPV expression were used for all assays. Subcutaneous flank xenografts were performed in Hsd:athymic Nude-Foxn1nu female mice treated with intraperitoneal cetuximab (0.2mg/mouse) or flank radiation (2 Gy/fraction) delivered twice weekly for 2 weeks. Time to tumor quadrupling from baseline was assessed by Kaplan-Meier method. Flow cytometry was used to assess apoptosis by Annexin V staining and cell cycle distribution by propidium iodide staining. RESULTS: While there was minimal in vitro radiosensitization after treatment with cetuximab (similar to that seen in HPV-negative HNC), cetuximab + radiation resulted in a significant increase in time to tumor quadrupling compared to cetuximab alone (p=0.001) or radiation alone (p&amp;lt;0.0001) in HPV+ HNC xenografts. Increased apoptosis was seen in cetuximab and radiation treated cells. Analysis of cell cycle distribution 24 hours after drug treatment showed that cetuximab induced a modest G1 cell cycle arrest when given prior to radiation, but that 24 hours after combined therapy, a significant G2 arrest was present. CONCLUSIONS: Using HPV+ HNC cell lines, radiation and cetuximab appear to be beneficial compared to either treatment alone. This combination, currently undergoing study in randomized clinical trial is being further evaluated in a direct from patient tumorgraft model of HPV+ HNC to further understand predictors of response. Citation Format: Molly A. Smith, Grace C. Blitzer, Alexandra D. Torres, Eric A. Armstrong, Paul F. Lambert, Paul M. Harari, Randall J. Kimple. Preclinical evaluation of chemoradiation with cetuximab HPV-positive head and neck cancer suggests significant radiosensitization. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 73. doi:10.1158/1538-7445.AM2013-73