Early Stage Head And Neck Squamous Cell Carcinoma Research Articles

Abstract 1620: Multiregion spatial sequencing of early-stage head and neck squamous cell carcinoma revealed theranostic intratumor heterogeneity

Abstract Introduction: Head and neck squamous cell carcinoma (HNSCC) is characterized by substantial intratumor heterogeneity (ITH) at the molecular level that may foster tumor evolution, adaptation, and therapeutic failure. Personalized-medicine strategies mostly rely on single tumor-biopsy samples which might not capture the mutational burden of heterogeneous tumors. We investigated genetic ITH portrayed from multiregion spatial sequencing in early-stage HNSCC patients. Patients and Method: Seventy-eight early-stage HNSCC patients treated with upfront curative-intent surgery in the prospective biobanking SCANDARE study (NCT03017573) were included. We developed a pathological measure of ITH, based on the relative contribution of multiple poorly and well-differentiated intratumor regions (n = 199) in resected tumor samples. To characterize genetic ITH, we performed targeted next-generation sequencing on multiple spatially separated intratumor regions by using a custom panel of 571 genes (named DRAGON), in combination with 3’-Tag RNA sequencing and pathway enrichment analysis. Results: Fifty-nine patients displaying a minimum of two sequenced intratumor regions (total, n = 180) were analyzed. We observed genetic ITH in 22/59 patients (37%), of whom 6/22 (27%) based on theranostic molecular alterations. Three of the six patients exhibited molecular alterations in PIK3CA (n = 2) and TSC2 (n = 1) that have supporting clinical evidence of actionability in some cancer types (OncoKB level 1) and three patients exhibited actionable molecular alterations in PTEN (n = 2) and CDKN2A/B (n = 1) supported by substantial research evidence (OncoKB level 4). Two patients exhibited molecular alterations in SMARCA4 that is likely oncogenic, albeit not yet annotated by OncoKB. The pathological measure of ITH accounted for a significant source of variation in gene expression. Notably, we identified 346 differentially expressed genes imposing p-values ≤ 0.05 and |log fold-change| ≥ 1 involved in several Reactome’s signaling pathways, including the cell junction organization (R-HSA-446728) and the tight junction interaction (R-HSA-420029) pathways. Conclusions: We reported clinically relevant genetic ITH in early-stage HNSCC, which can lead to underestimation of the mutational landscape portrayed from single tumor biopsy and may present major challenges to precision medicine. Our findings uncovered the biology behind ITH and may have translational implications for biomarkers discovery in HNSCC. Citation Format: Grégoire Marret, Sophie Vacher, Constance Lamy, Ladidi Ahmanache, Laura Courtois, Choumouss Kamoun, Maral Halladjian, Zakhia El Beaino, Jerzy Klijanienko, Charlotte Martinat, Emmanuelle Jeannot, Joey Martin, Guillaume Rougier, Nathalie Badois, Maria Lesnik, Antoine Dubray-Vautrin, Olivier Choussy, Wahib Ghanem, Rabah Taouachi, Julien Masliah Planchon, Nicolas Servant, Maud Kamal, Ivan Bièche, Christophe Le Tourneau. Multiregion spatial sequencing of early-stage head and neck squamous cell carcinoma revealed theranostic intratumor heterogeneity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1620.

Identification of an immune-related signature as a prognostic classifier for patients with early-stage head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is the most common type and accounts for 90% of all head and neck cancer cases. Despite advances in early diagnosis and treatment strategies-chemotherapy, surgical resection, and radiotherapy-5-year survival remains grim. For patients with early-stage HNSCC, accurately predicting clinical outcomes is challenging. Considering the pivotal role of the immune system in HNSCC, we developed a reliable immune-related gene signature (IRGS) and explored its predictive accuracy in patients with early-stage HNSCC. We examined immune gene expression profiles and clinical information from 230 early-stage HNSCC specimens, including 100 cases from The Cancer Genome Atlas (TCGA), 49 cases from the Gene Expression Omnibus (GEO; GSE65858), and 81 cases from an independent clinical cohort. The prognostic signature was constructed using Kaplan-Meier analysis and the least absolute shrinkage and selection operator (LASSO) Cox algorithm. We also explored the IRGS-related biological pathways and immune landscape using bioinformatics analysis. A nine-immune-gene signature was generated to significantly stratify patients into high and low-risk groups. High risk patients exhibited shorter survival time [hazard ratio (HR) =13.795, 95% confidence interval (CI): 3.275-58.109, P<0.001]. The signature demonstrated robust prognostic ability in the training and validation sets and could independently predict overall survival (OS) and relapse-free survival (RFS). Subsequently, the receiver operating characteristic (ROC) curve and C-index confirmed the signature's predictive accuracy compared to clinical parameters. Additionally, cases classified as low risk showed more immune cell infiltration than high-risk cases. Our novel IRGS is a reliable and robust classifier for accurate patient stratification and prognostic evaluation. Future studies will attempt to affirm the signature's clinical application to early-stage HNSCC.

Small extravesicular microRNA in head and neck squamous cell carcinoma and its potential as a liquid biopsy for early detection.

The objective was to assess secretion of small extracellular vesicular microRNA (exo-miRNA) in head and neck squamous cell carcinoma (HNSCC) according to human papillomavirus (HPV) status, and determine the translational potential as a liquid biopsy for early detection. This study employed a combination of cell culture and case-control study design using archival pretreatment serum. Small extracellular vesicles (sEV) were isolated from conditioned culture media and human serum samples via differential ultracentrifugation. miRNA-sequencing was performed on each sEV isolate. There were clear exo-miRNA profiles that distinguished HNSCC cell lines from nonpathologic oral epithelial control cells. While there was some overlap among profiles across all samples, there were apparent differences in exo-miRNA profiles according to HPV-status. Importantly, differential exo-miRNA profiles were also apparent in serum from early-stage HNSCC cases relative to cancer-free controls. Our findings indicate that exo-miRNA are highly dysregulated in HNSCC and support the potential of exo-miRNA as biomarkers for HNSCC.

GPD1L is negatively associated with HIF1α expression and predicts lymph node metastasis in oral and HPV- Oropharyngeal cancer.

The study is to determine if Glycerol-3-phosphate dehydrogenase 1-like (GPD1L) and hypoxia-inducible factor-1α (HIF1α) can identify high-risk patients with delayed lymph node metastasis in early-stage head and neck squamous cell carcinoma (HNSCC). The mRNA and protein expressions of markers were analyzed using fresh and paraffin embedded HNSCC specimens. The statistical analyses included chi-squared test, t test, correlation analysis, and univariate and multivariate analyses. GPD1L (downregulated) and HIF1α (upregulated) mRNA expression had a negative correlation (r=-.496, p=.001) in cT1-2N0 HNSCC. The low GPD1L+high HIF1α expression group (22.6%) showed a significant decrease in disease-free survival compared with the high GPD1L+low HIF1α expression group (71.4%) in the neck dissection group. The low GPD1L+high HIF1α expression (39.4%) resulted in a significantly higher delayed metastasis rate than the high GPD1L+low HIF1α expression (5.6%) for cT1-2N0 HNSCC in the neck observation group. GPD1L and HIF1α protein expression more accurately predicted lymph node metastasis than the WINTER hypoxia gene panel (false-negative rate in predicting metastasis: 8.1% versus 26.4%). Cox regression analyses found that the combined protein expression of GPD1L and HIF1α could predict delayed metastasis (HR:0.118, 95% CI:0.027-0.525). Low GPD1L+high HIF1α expression can serve as candidate biomarkers for high-risk populations with lymph node metastases in early-stage HNSCC.

Synchronous esophageal cancer impact on survival of head and neck squamous cell carcinoma (HNSCC) patients.

e17513 Background: Optimal treatment of synchronous tumors (ST) of the aerodigestive tract is debatable and care is often individualized. Our goal was to characterize patients with HNSCC and esophageal cancer (EC) ST and to establish prognostic factors that could aid therapeutic decision. Methods: In this retrospective observational study, we evaluated data from 1650 consecutive patients diagnosed with HNSCC from 2008 to 2016. Patients with ST of HNSCC and esophagus with an interval of ≤ 6 months between both diagnoses were included. Patients ≥ 6 months between both diagnoses, incomplete treatment information and presence of another tumor site were excluded. Results: 52 patients were eligible. Median age was 57 years (39-91). Most were male (98%), with smoking and drinking habits (98%) and ECOG 0-1 (73%). HNSCCs were mainly in oropharynx (54%) and locally advanced disease (LA, III-IVB) (88%). In contrast, EC was early stage (I-II, 62%), located in the thorax (94%) and squamous histology (96%). 14 (27%) had LA in both primaries. Most LA HNSCCs (85%) were treated with radiotherapy (RT) with a median dose of 70Gy (5-70Gy). 50% received platinum and taxane induction chemotherapy. 81% of initial EC received at least surgery, mucosectomy or RT (median 50.4Gy). Hospitalization due to toxicity occurred in 12 (23%) and 7 (14%) of HNSCC and EC treatments, respectively. 16 patients (31%) had no definitive treatment directed to EC, without apparent impact on survival. Median time to progression was 13.8 months, being HNSCC the most frequent site of progression/relapse (40%). Median survival was 23.9 months (IC 95% 9.2-38.6). Early HNSCC survival was comparable to LA HNSCC (17.3 vs. 23.9 mo, p = 0.98). In LA HNSCC, LA vs. initial EC carried a worse prognosis (16 vs. 36.3 mo, p = 0.008). Anemia, BMI, tobacco exposure had no impact on survival. Conclusions: The occurrence of EC and HNSCC ST leads to a dismal survival, even in patients with early stage HNSCC. The presentation of LA in both sites is particularly challenging and associated with worse prognosis. Given the rate of treatment-related toxicity in this population, cautious efforts should be employed when planning definitive treatment in ST pts.

Tumor-associated B cells and humoral immune response in head and neck squamous cell carcinoma

ABSTRACTB lymphocytes are important players in immune responses to cancer. However, their composition and function in head and neck squamous cell carcinoma (HNSCC) has not been well described. Here, we analyzed B cell subsets in HNSCC (n = 38), non-cancerous mucosa (n = 14) and peripheral blood from HNSCC patients (n = 38) and healthy controls (n = 20) by flow cytometry. Intratumoral B cells contained high percentages of activated (CD86+), antigen-presenting (CD86+/CD21−) and memory B cells (IgD−/CD27+). T follicular helper cells (CD4+/CXCR5+/CD45RA−/CCR7−) as key components of tertiary lymphoid structures and plasma cells made up high percentages of the lymphocyte infiltrate. Percentages of regulatory B cell varied depending on the regulatory phenotype. Analysis of humoral immune responses against 23 tumor-associated antigens (TAA) showed reactivity against at least one antigen in 56% of HNSCC patients. Reactivity was less frequent in human papillomavirus associated (HPV+) patients and healthy controls compared to HPV negative (HPV−) HNSCC. Likewise, patients with early stage HNSCC or MHC-I loss on tumor cells had low TAA responses. Patients with TAA responses showed CD4+ dominated T cell infiltration compared to mainly CD8+ T cells in tumors without detected TAA response. To summarize, our data demonstrates different immune infiltration patterns in relation to serological TAA response detection and the presence of B cell subpopulations in HNSCC that can engage in tumor promoting and antitumor activity. In view of increasing use of immunotherapeutic approaches, it will be important to include B cells into comprehensive phenotypic and functional analyses of tumor-associated lymphocytes.

Stromal interleukin-33 promotes regulatory T cell-mediated immunosuppression in head and neck squamous cell carcinoma and correlates with poor prognosis.

Regulatory T cells (Tregs) mediate immunosuppressive signals that can contribute to the progression of head and neck squamous cell carcinoma (HNSCC). Interleukin-33 (IL-33) is defined as an 'alarmin', an endogenous factor that is expressed during tissue and cell damage, which has been shown to promote Treg proliferation in non-lymphoid organs. However, the interaction between IL-33 and Tregs in the HNSCC tumor microenvironment remains uncertain. In this study, we examined IL-33+ and Foxp3+ cells by immunohistochemistry in 68 laryngeal squamous cell cancer patients, followed by functional analysis of IL-33 in Tregs. In addition, the suppressive function of Tregs was assessed by cell proliferation assays. The level of stromal IL-33 was significantly upregulated in advanced versus early stage HNSCC patients and positively correlated with Foxp3+ Treg infiltration as well as a poor prognosis. ST2 is regarded as the only receptor of IL-33. Infiltrated ST2-expressing Tregs were responsive to IL-33, and the percentage of Tregs was increased upon IL-33 stimulation. Functional investigation demonstrated that IL-33 increased the proportion of Foxp3+GATA3+ Tregs and improved the suppressive functions of Tregs by inducing IL-10 and TGF-β1 as well as decreasing the proliferation of responder T cells. Blockade of ST2 abrogated the immunosuppression caused by IL-33. Our data demonstrate that stromal IL-33 both expands the Treg population and enhances their functions in the tumor microenvironment. Furthermore, stromal IL-33 has prognostic value for tumor progression. Thus, stromal IL-33 is a potential target for future HNSCC immunotherapy.

Preoperative anemia and perioperative blood transfusion in head and neck squamous cell carcinoma

ObjectivesTo evaluate the impact of preoperative anemia and perioperative blood transfusion (PBT) on disease free (DFS) and overall survival (OS) of patients with head and neck squamous cell carcinoma (HNSCC).MethodsRetrospective study of 354 patients primarily treated with surgery between 2006 and 2016. Cases were selected according to completeness and accuracy of available clinical data. Thus, a selection bias cannot be excluded. Patients who received PBT were identified by our controlling department and verified by our blood bank data base.ResultsBoth, preoperative anemia and PBT significantly decreased OS in univariate analysis. Although PBT was needed more frequently by older patients in worse physical conditions with more advanced HNSCC, subgroup analysis also demonstrate a profoundly negative effect of PBT on OS in younger patients and early stage HNSCC. According to a restrictive transfusion policy at our hospital the transfusion rate was comparably low. We could not verify increasing effects of PBT on cancer recurrence rates as it was previously shown.DiscussionPreoperative anemia is the most common paraneoplastic syndrome in HNSCC. Despite its devastating prognostic effect we suggest a restrictive transfusion policy whenever possible. Our data also show that anemia as an independent prognostic factor in head and neck surgical oncology is defined not only by low hemoglobin concentrations but low red blood cell counts as well.

Abstract 5332: Prognostic value of aberrant promoter hypermethylation of tumor-related genes in early-stage head and neck cancer

Abstract Staging and pathological grading are useful, but imperfect predictors of recurrence in head and neck squamous cell carcinoma (HNSCC). Accordingly, molecular biomarkers that predict the risk of recurrence are necessary to improve clinical outcomes. The ability to distinguish between low- and high-risk HNSCCs at an early stage may reduce follow-up costs. We hypothesized that the quantitative methylation specific PCR (Q-MSP) assay could be used to define patterns of DNA methylation that differentiate low- and high-risk HNSCCs. The methylation statuses of the promoters of 11 tumor-related genes (p16, RASSF1A, E-cadherin, H-cadherin, MGMT, DAPK, DCC, COL1A2, TAC1, SST, and GALR1) were analyzed in 133 HNSCC cases using quantitative methylation-specific PCR. Frequent methylation of p16 (44%), RASSF1A (18%), E-cadherin (53%), H-cadherin (35%), MGMT (35%), DAPK (53%), DCC (42%), COL1A2 (44%), TAC1 (61%), SST (64%), and GALR1 (44%) in HNSCC were detected. Disease-free survival was lower in patients with 6-11 methylated genes than in those with 0-5 methylated genes (log-rank test, P = 0.001). In a multivariate Cox proportional hazards analysis, the methylation of E-cadherin, COL1A2, TAC1, and GALR1 was associated with poor survival, with hazard ratios of 4.474 (95% CI, 1.241-16.124). In a joint analysis of these four genes, patients with 2-4 methylated genes had a significantly lower survival rate than those with 0-1 methylated genes in early-stage HNSCC. Importantly, the methylation of some genes was closely related to poor prognosis in early-stage HNSCC, providing strong evidence that these hypermethylated genes are valuable biomarkers for prognostic evaluation. In conclusion, the methylation profiles of E-cadherin, COL1A2, TAC1, and GALR1 were the most powerful combination for predicting early-stage HNSCC. This demonstrates that molecular stratification may predict cancer progression. These findings can benefit HNSCC screening and surveillance algorithms. Although our study was retrospective, was conducted at a single institution, and the number of patients was small, it serves as a platform to establish optimal therapeutic strategies for early-stage HNSCC. Citation Format: Atsushi Imai, Kiyoshi Misawa, Daiki Mochizuki, Hiroyuki Mineta, Takeharu Kanazawa. Prognostic value of aberrant promoter hypermethylation of tumor-related genes in early-stage head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5332.

The impact of time to treatment initiation on survival from head and neck cancer in north-eastern Italy

ObjectivesTo evaluate the impact of time to treatment initiation (TTI) on overall survival in patients with head-and-neck squamous cell carcinoma (HNSCC). Materials and methodsIn the period 2003–2009, 1616 HNSCC patients were diagnosed in Friuli Venezia Giulia Region, Northeastern Italy, including 462 oral, 346 oropharyngeal, 212 hypopharyngeal, and 596 laryngeal cancers. Clinical information, including date and type of first treatment, and follow-up were retrieved from the regional Cancer Registry and a population-based health database collecting comprehensive health information on people living in the Region. Multivariate hazard ratio (HR) and confidence intervals (CI) were calculated through Cox model. ResultsOverall, the median TTI was 28days, (Q1-Q3: 13–45days), but significant variations emerged according to anatomical site, cancer stage, treatment approach, and care transition to specialized centers. Five-year overall survival decreased with increasing treatment delay from 62% for TTI<30days to 39% for TTI≥90days (p<0.01). HR of death was 1.13 (95% CI: 0.92–1.39) for TTI between 45–89days, and 1.47 (1.05–2.05) for TTI≥90days. The association between TTI and poor prognosis was stronger for laryngeal cancers and early-stage HNSCCs. Further, care transition from community hospitals to specialized centers was associated to a better prognosis (HR=0.73; 95% CI: 0.60–0.88). ConclusionOur study findings suggest that HNSCC patients treated within 45days from diagnosis have increased survival probabilities and that early-stage patients suffered the most from treatment delay. Furthermore, care transition to specialized centers –though competitive to timely treatment– improves survival by providing the most innovative technologies and treatment approaches.

Endoscopic-assisted selective neck dissection via small lateral neck incision for early-stage (T1-2N0M0) head and neck squamous cell carcinoma: 3-year follow-up results.

Primary tumors of early-stage (T1-2N0M0) head and neck squamous cell carcinoma (HNSCC) can be treated by transoral minimally invasive surgery, but the cervical lymph node lacks corresponding minimally invasive treatment. Therefore, our study was to evaluate the safety and effectiveness of endoscopic-assisted selective neck dissection (EASND) via small lateral neck incision in treatment of early-stage HNSCC. From May 2008 to April 2012, 29 patients with early-stage HNSCC were enrolled in this retrospective study. EASND via small lateral neck incision was followed by primary tumors treated by transoral endoscopy or CO2 laser microsurgery in all patients. The clinical data and follow-up results were analyzed retrospectively. Twenty-nine patients (male 22, female 7) were successfully treated by EASND and no case conversed to open surgery. EASND included eight cases of level I-III and 21 cases of level II-IV. The operative time was 68.4±10.3min, and the operative bleeding was 40.8±12.4ml. The number of lymph nodes was 19.1±4.1. There were 19 cases of T1, 10 cases of T2, 22 cases of N0, 7 cases of N1, no cases with positive margin or extracapsular spread in lymph node. Temporary sensory changes in the postauricular area occurred in two cases, both recovered within 3months after the surgery, postoperative seromas occurred in one case. No patient showed postoperative bleeding, chylous fistula, incision infection, facial nerve injury, and accessory nerve injury. All patients were satisfied with the postoperative cosmetic results. The 3-year OS was 96.5%, while DFS, LRFS, and DMFS were 100%. EASND via small lateral neck incision is a feasible and safe technique with achieved short-term oncologic outcomes in early-stage (T1-2N0M0) HNSCC patients. But the long-term results of EASND on oncologic safety and functional outcomes are required to confirm by larger, randomised studies.

Prognostic value of aberrant promoter hypermethylation of tumor-related genes in early-stage head and neck cancer.

Staging and pathological grading are useful, but imperfect predictors of recurrence in head and neck squamous cell carcinoma (HNSCC). Accordingly, molecular biomarkers that predict the risk of recurrence are necessary to improve clinical outcomes. The methylation statuses of the promoters of 11 tumor-related genes (p16, RASSF1A, E-cadherin, H-cadherin, MGMT, DAPK, DCC, COL1A2, TAC1, SST, and GALR1) were analyzed in 133 HNSCC cases using quantitative methylation-specific PCR. We detected frequent methylation of p16 (44%), RASSF1A (18%), E-cadherin (53%), H-cadherin (35%), MGMT (35%), DAPK (53%), DCC (42%), COL1A2 (44%), TAC1 (61%), SST (64%), and GALR1 (44%) in HNSCC. Disease-free survival was lower in patients with 6–11 methylated genes than in those with 0–5 methylated genes (log-rank test, P = 0.001). In a multivariate Cox proportional hazards analysis, the methylation of E-cadherin, COL1A2, TAC1, and GALR1 was associated with poor survival, with hazard ratios of 4.474 (95% CI, 1.241–16.124). In a joint analysis of these four genes, patients with 2–4 methylated genes had a significantly lower survival rate than those with 0–1 methylated genes in early-stage HNSCC. Importantly, the methylation of some genes was closely related to poor prognosis in early-stage HNSCC, providing strong evidence that these hypermethylated genes are valuable biomarkers for prognostic evaluation.

Intraoperative optical assessment of photodynamic therapy response of superficial oral squamous cell carcinoma.

This study investigated whether diffuse optical spectroscopy (DOS) measurements could assess clinical response to photodynamic therapy (PDT) in patients with head and neck squamous cell carcinoma (HNSCC). In addition, the correlation between parameters measured with DOS and the crosslinking of signal transducer and activator of transcription 3 (STAT3), a molecular marker for PDT-induced photoreaction, was investigated. Thirteen patients with early stage HNSCC received the photosensitizer 2-[1-hexyloxyethyl]-2-devinylpyropheophorbide-a (HPPH) and DOS measurements were performed before and after PDT in the operating room (OR). In addition, biopsies were acquired after PDT to assess the STAT3 crosslinking. Parameters measured with DOS, including blood volume fraction, blood oxygen saturation (StO2), HPPH concentration (cHPPH), HPPH fluorescence, and blood flow index (BFI), were compared to the pathologic response and the STAT3 crosslinking. The best individual predictor of pathological response was a change in cHPPH (sensitivity=60%, specificity=100%), while discrimination analysis using a two-parameter classifier (change in cHPPH and change in StO2) classified pathological response with 100% sensitivity and 100% specificity. BFI showed the best correlation with the crosslinking of STAT3. These results indicate that DOS-derived parameters can assess the clinical response in the OR, allowing for earlier reintervention if needed.

Presentation and Treatment of a Patient with Three Primary Head and Neck Cancers: A Case Report

Triple primary head and neck carcinoma is a rare occurrence. In our case, a patient presented with early stage triple primary head and neck squamous cell carcinoma (HNSCC). Early stage HNSCC is usually treated with surgery and/or radiation. However, treatment of triple primary head and neck malignancies is not clearly defined in the literature, and although our patient presented with early stage lesions, our multidisciplinary team recommended concomitant chemoradiotherapy. Our patient subsequently had resolution of disease after completion of therapy but was lost to long-term follow-up.

When is radiation therapy in head and neck squamous cell carcinoma not indicated?

Head and neck squamous cell carcinoma (HNSCC) can be treated with surgery, radiotherapy (RT), and/or chemotherapy depending on the location and extent of the neoplasm. Treatment selection also depends on the goal of therapy (curative or palliative) and the expected efficacy weighed against the expected morbidity of the treatment. Moreover, it may be determined by the treatment modalities available, although all modalities will usually be available in most centers in developed countries. Earlystage HNSCCs can usually be treated with a single modality, whereas advanced-stage disease requires multimodality treatment. The choice of treatment modality will depend on factors including tumor site, existing and expected functionality, cosmetic outcome, age, performance status, the likelihood that adjuvant postoperative RT will be required, and the wishes of the patient. Following is a discussion of the curative-intent management of patients with mucosal HNSCC and situations where RT can be safely eliminated. Surgery alone is the preferred treatment for patients with HNSCC when the probability of a complete resection is high ([85–90 %) and the functional and cosmetic outcome is acceptable. An important caveat is that a functional/ cosmetic outcome that is acceptable to the physician may or may not be acceptable to the patient. Other factors such as patient age and medical co-morbidities also influence the decision. Patients with human papilloma virus (HPV)positive oropharyngeal squamous cell carcinomas (SCCs) have a better prognosis than those with HPV-negative SCCs. It could be reasoned that these patients should be treated differently. Depending on extent of disease, surgery alone, transoral robotic surgery (TORS) in particular, could be an attractive option as this group of patients is usually of younger age. However, the question if and how patients should be treated differently depending on HPV status is the subject of trials now and no firm recommendation can be made at this point [1, 2]. Although the adverse impacts The paper was written by members of the International Head and Neck Scientific Group (www.IHNSG.com).

Abstract 726: Copy number amplification of the PIK3CA gene is associated with poor prognosis in early-stage head and neck squamous cell carcinoma

Abstract Background: Deregulation of the EGFR signaling pathway is one of the most frequently observed genetic abnormalities that drive cancer development. Mutations in downstream components of the EGFR signaling pathway, including KRAS, BRAF and PIK3CA, have been reported in numerous cancers. However, extensive mutation analysis of these genes in clinical samples has not been performed for head and neck squamous cell carcinoma (HNSCC). Patient and methods: We examined mutations and copy number alterations of KRAS, BRAF and PIK3CA in 115 clinical specimens of HNSCC obtained from surgically treated patients. Results: We identified three mutations in K-RAS and two mutations in PIK3CA. Copy number amplification was found in 42 cases (36.4%) for PIK3CA, 12 cases (10.4%) for K-RAS and nine cases (7.8%) for BRAF. Tumor recurrence and death occurred in 49 (42.6%) and in 22 (19.3 %) patients during the median follow-up of 723 days, respectively. Kaplan-Meier survival analysis revealed that in patients with early-stage carcinoma (Stages I-III), copy-number amplification of PIK3CA was drastically associated with cancer relapse. Conclusion: Copy number amplification of the PIK3CA is associated with poor prognosis in early-stage HNSCC. The PIK3CA copy number status will serve as a marker of poor prognosis in patients with HNSCC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 726. doi:1538-7445.AM2012-726

Genetic variations in regulator of G-protein signaling genes as susceptibility loci for second primary tumor/recurrence in head and neck squamous cell carcinoma

Curatively treated patients with early-stage head and neck squamous cell carcinoma (HNSCC) are at high risks for second primary tumor (SPT) and recurrence. The regulator of G-protein signaling (RGS) is important in essential signaling transduction and cellular activities. We hypothesize that genetic variations of RGS may modulate the risk of SPT/recurrence in patients with early-stage HNSCC. In a nested case-control study, we evaluated 98 single-nucleotide polymorphisms (SNPs) in 17 RGS genes for the risk of SPT/recurrence among 450 HNSCC patients. Eight SNPs showed significant associations with the risk of SPT/recurrence, with the most significant one of rs2179653, which is located in the 5'-flanking region of RGS2 gene. Under a recessive genetic model, the homozygous variant genotype of this SNP was associated with 2.95-fold [95% confidence interval (CI): 1.52-5.74] increased risk of SPT/recurrence. This association remained significant after the adjustment for multiple comparisons. Cumulative effects analysis revealed that the risk increased significantly with the increasing numbers of unfavorable genotypes. Compared with subjects carrying 0-2 unfavorable genotypes, the hazard ratios (95% CIs) for those carrying 3 or 4+ were 1.73 (1.10-2.70) and 3.05 (1.92-4.83), respectively. Furthermore, survival tree analysis revealed potential higher order gene-gene interactions and indicated different outcomes based on distinct genotype profiles. Genetic variations of RGS genes may modulate the susceptibility to SPT/recurrence in early-stage HNSCC patients individually and cumulatively. Our results stressed the importance of taking a polygenic approach to evaluate the cumulative and interaction effects of genetic variations in the prediction of cancer risk and prognosis.

Genetic variants in the telomere maintenance pathway as predictors of second primary tumors and recurrence of early-stage head and neck cancer.

1504 Background: Telomere is critical for chromosome stability. Telomere shortening has been associated with cancer development. In this study, we evaluated genetic variants in the telomere maintenance pathway as predictors for the risk of second primary tumor (SPT) or recurrence in early-stage head and neck squamous cell carcinoma (HNSCC) patients. Methods: We genotyped 427 single nucleotide polymorphisms (SNP) from 41 telomere maintenance pathway genes in a nested 1:2 case-control study of 150 early stage HNSCC patients with SPT/recurrence and 300 patients without. Individual and cumulative effects and interactions of these SNPs on SPT/recurrence were assessed. Results: In the Cox proportional hazards model, patients carrying the variant genotypes of rs9562605, located in the BRCA2 gene, were associated with a 0.5-fold (95% CI 0.4- 0.8) reduced risk of SPT/recurrence, and longer median SPT/recurrence-free survival time (MST) (> 93 months) than those with wild-type genotype (81.6 months) (p = 2.4 × 10-3). A cumulative effect analysis was performed by combining the unfavorable genotypes of the 53 significant SNPs with p < 0.05. A progressively increasing SPT/recurrence risk was observed as the number of unfavorable genotypes increased (p for trend < 0.0001). Specifically, compared to the subjects with fewer than 19 unfavorable genotypes, subjects with more than 30 unfavorable genotypes had 19.7-fold (95% CI 9.6-40.4) increased risk and significantly shorter MST (28.2 months vs. > 93 months, log-rank p = 2.0 × 10-24). From survival tree analysis, rs10849605 was the first split with the smallest p-value (2.5 × 10-3). Node 18 ranked the highest risk group (HR > 3.7, 95% CI 9.6-568.4, MST = 13.4 months) as compared to the reference node, node 1 (MST > 93 months) (log-rank p = 1.1 × 10-23). Conclusions: The study identified several promising SNPs and further demonstrated that the combination of multiple SNPs and their interactions may be useful to predict clinical outcome of early-stage head and neck cancer. No significant financial relationships to disclose.

Novel susceptibility loci for second primary tumors/recurrence in head and neck cancer patients: large-scale evaluation of genetic variants.

This study was aimed to identify novel susceptibility variants for second primary tumor (SPT) or recurrence in curatively treated early-stage head and neck squamous cell carcinoma (HNSCC) patients. We constructed a custom chip containing a comprehensive panel of 9,645 chromosomal and mitochondrial single nucleotide polymorphisms (SNP) representing 998 cancer-related genes selected by a systematic prioritization schema. Using this chip, we genotyped 150 early-stage HNSCC patients with and 300 matched patients without SPT/recurrence from a prospectively conducted randomized trial and assessed the association of these SNPs with risk of SPT/recurrence. Individually, six chromosomal SNPs and seven mitochondrial SNPs were significantly associated with risk of SPT/recurrence after adjustment for multiple comparisons. A strong gene-dosage effect was observed when these SNPs were combined, as evidenced by a progressively increasing SPT/recurrence risk as the number of unfavorable genotypes increased (P for trend < 1.00 x 10(-20)). Several polygenic analyses suggest an important role of interconnected functional network and gene-gene interaction in modulating SPT/recurrence. Furthermore, incorporation of these genetic markers into a multivariate model improved significantly the discriminatory ability over the models containing only clinical and epidemiologic variables. This is the first large-scale systematic evaluation of germ-line genetic variants for their roles in HNSCC SPT/recurrence. The study identified several promising susceptibility loci and showed the cumulative effect of multiple risk loci in HNSCC SPT/recurrence. Furthermore, this study underscores the importance of incorporating germ-line genetic variation data with clinical and risk factor data in constructing prediction models for clinical outcomes.

Autoantibody Approach for Serum-Based Detection of Head and Neck Cancer

Currently, no effective tool exists for screening or early diagnosis of head and neck squamous cell carcinoma (HNSCC). Here, we describe an approach for cancer detection based on analysis of patterns of serum immunoreactivity against a panel of biomarkers selected using microarray-based serologic profiling and specialized bioinformatics. We biopanned phage display libraries derived from three different HNSCC tissues to generate 5,133 selectively cloned tumor antigens. Based on their differential immunoreactivity on protein microarrays against serum immunoglobulins from 39 cancer and 41 control patients, we reduced the number of clones to 1,021. The performance of a neural network model (Multilayer Perceptron) for cancer classification on a data set of 80 HNSCC and 78 control samples was assessed using 10-fold cross-validation repeated 100 times. A panel of 130 clones was found to be adequate for building a classifier with sufficient sensitivity and specificity. Using these 130 markers on a completely new and independent set of 80 samples, an accuracy of 84.9% with sensitivity of 79.8% and specificity of 90.1% was achieved. Similar performance was achieved by reshuffling of the data set and by using other classification models. The performance of this classification approach represents a significant improvement over current diagnostic accuracy (sensitivity of 37% to 46% and specificity of 24%) in the primary care setting. The results shown here are promising and show the potential use of this approach toward eventual development of diagnostic assay with sufficient sensitivity and specificity suitable for detection of early-stage HNSCC in high-risk populations.