Radiogenomics in head and neck cancer: correlation of radiomic heterogeneity and somatic mutations in TP53, FAT1 and KMT2D.

PURPOSE: Genetic tumour profiles and radiomic features can be used to complement clinical information in head and neck squamous cell carcinoma (HNSCC) patients. Radiogenomics imply the potential to investigate complementarity or interrelations of radiomic and genomic features, and prognostic factors might be determined. The aim of our study was to explore radiogenomics in HNSCC. METHODS: For 20 HNSCC patients treated with primary radiochemotherapy, next-generation sequencing (NGS) of tumour and corresponding normal tissue was performed. In total, 327 genes were investigated by panel sequencing. Radiomic features were extracted from computed tomography data. A hypothesis-driven approach was used for radiogenomic correlations of selected image-based heterogeneity features and well-known driver gene mutations in HNSCC. RESULTS: The most frequently mutated driver genes in our cohort were TP53 (involved in cell cycle control), FAT1 (Wnt signalling, cell-cell contacts, migration) and KMT2D (chromatin modification). Radiomic features of heterogeneity did not correlate significantly with somatic mutations in TP53 or KMT2D. However, somatic mutations in FAT1 and smaller primary tumour volumes were associated with reduced radiomic intra-tumour heterogeneity. CONCLUSION: The landscape of somatic variants in our cohort is well in line with previous reports. An association of somatic mutations in FAT1 with reduced radiomic tumour heterogeneity could potentially elucidate the previously described favourable outcomes of these patients. Larger studies are needed to validate this exploratory data in the future.

Prognostic value of CXCL12 and CXCR4 in inoperable head and neck squamous cell carcinoma.

OBJECTIVE: The chemokine CXCL12 and its receptor CXCR4 can affect tumor growth, recurrence, and metastasis. We tested the hypothesis that the CXCL12 and CXCR4 expression influences the prognosis of patients with inoperable head and neck cancer treated with definite radiotherapy or chemoradiotherapy. METHODS: Formalin-fixed paraffin-embedded pretreatment tumor tissue from 233 patients with known HPV/p16(INK4A) status was analyzed. CXCL12 and CXCR4 expressions were correlated with pretreatment parameters and survival data by univariate and multivariate Cox regression. RESULTS: CXCL12 was expressed in 43.3 % and CXCR4 in 66.1 % of the samples and both were correlated with HPV/p16(INK4A) positivity. A high CXCL12 expression was associated with increased overall survival (p = 0.036), while a high CXCR4 expression was associated with decreased metastasis-free survival (p = 0.034). CONCLUSION: A high CXCR4 expression could be regarded as a negative prognostic factor in head and neck cancer because it may foster metastatic spread. This may recommend CXCR4 as therapeutic target for combating head and neck cancer metastasis.

MAGE-A1-6 expression in patients with head and neck squamous cell carcinoma: impact on clinical patterns and oncologic outcomes.

BACKGROUND: Various subtypes of melanoma-associated antigens (MAGEs) are expressed in the tumor tissues of patients with head and neck squamous cell carcinoma (HNSCC). However, little data are currently available on how the gene expression of MAGEs impacts clinical patterns and oncologic outcomes. We have therefore evaluated the expression of MAGE-A1-6 (A1-6) subtypes in tumor tissues of patients with HNSCC and the clinical impact of this expression. METHODS: This was a retrospective review of 53 patients with histologically proven HNSCC of the oral cavity, oropharynx, larynx, or hypopharynx who underwent both treatment and analysis by reverse transcription (RT)-PCR assay with a common primer to identify the expression of MAGE-A1-6 subtypes in the tumor tissue. The clinicopathologic factors and oncologic outcomes of these patients and the correlations of both to MAGE-A1-6 gene expression were analyzed. RESULTS: MAGE-A1-6 subtypes were expressed in the tumor tissues of 37 patients (69.8 %). Patient age of ≥65 years [p = 0.031, hazard ratio (HR) 4.866] and advanced American Joint Committee on Cancer stage (p = 0.035, HR 4.291) were independent risk factors for expression of MAGE-A1-6 subtypes. Patients with MAGE-A1-6 expression had lower disease-free survival (p = 0.029), disease-specific survival (p = 0.070), and overall survival (p = 0.017) rates. Overall survival rate was independently associated to chemotherapy (p = 0.011, HR 2.859), while no surgery (p = 0.050, HR 2.400) and MAGE-A1-6 expression (p = 0.050, HR 2.527) showed borderline significance. CONCLUSION: In our patient group the expression of MAGE-A1-6 subtypes in tumor tissues of patients with HNSCC was correlated with advanced clinical stage of cancer and poor oncologic outcomes. We suggest that gene expression of MAGE-A1-6 subtypes may be considered to be a predictive factor to determine patient treatment or follow-up strategy.

High-grade acute organ toxicity and p16(INK4A) expression as positive prognostic factors in primary radio(chemo)therapy for patients with head and neck squamous cell carcinoma.

BACKGROUND: Superior treatment response and survival for patients with human papilloma virus (HPV)-positive head and neck cancer (HNSCC) are documented in clinical studies. However, the relevance of high-grade acute organ toxicity (HGAOT), which has also been correlated with improved prognosis, has attracted scant attention in HPV-positive HNSCC patients. Hence we tested the hypothesis that both parameters, HPV and HGAOT, are positive prognostic factors in patients with HNSCC treated with definite radiotherapy (RT) or radiochemotherapy (RCT). PATIENTS AND METHODS: Pretreatment tumor tissue and clinical records were available from 233 patients receiving definite RT (62 patients) or RCT (171 patients). HPV infection was analysed by means of HPV DNA detection or p16(INK4A) expression; HGAOT was defined as the occurrence of acute organ toxicity >grade 2 according to the Common Toxicity Criteria. Both variables were correlated with overall survival (OS) using Cox proportional hazards regression. RESULTS: Positivity for HPV DNA (44 samples, 18.9 %) and p16(INK4A) expression (102 samples, 43.8 %) were significantly correlated (p < 0.01), and HGAOT occurred in 77 (33 %) patients. Overall, the 5-year OS was 23 %; stratified for p16(INK4A) expression and HGAOT, OS rates were 47 %, 42 %, 20 % and 10 % for patients with p16(INK4A) expression and HGAOT, patients with HGAOT only, patients with p16(INK4A) expression only, and patients without p16(INK4A) expression or HGAOT, respectively. After multivariate testing p16(INK4A) expression (p = 0.003) and HGAOT (p < 0.001) were significantly associated with OS. CONCLUSION: P16(INK4A) expression and HGAOT are independent prognostic factors for OS of patients with HNSCC, whereas p16(INK4A) expression is particularly important for patients without HGAOT.

[Epithelial neuroendocrine tumors of the upper respiratory tract: New entities, new perspectives]. / Neuroendokrine epitheliale Tumoren des oberen Respirationstrakts : Neue Entitäten, neue Blickwinkel.

Epithelial neuroendocrine tumors of the upper respiratory tract are rare and are classified as typical and atypical carcinoid versus small cell neuroendocrine carcinoma. Furthermore, a giant cell variant of neuroendocrine carcinoma is suggested corresponding to the bronchopulmonary system as well as a recently described subtype of oropharyngeal small cell neuroendocrine carcinoma associated with human papillomavirus. Many arguments relying on clinical as well as on molecular findings indicate that the distinction between carcinoid and poorly differentiated neuroendocrine carcinoma does not only reflect different degrees of differentiation of otherwise related tumors but indicates the existence of substantially different types of neoplasms.

Significance of p16 expression in head and neck cancer patients treated with radiotherapy and cetuximab.

BACKGROUND: HPV-infection, p16 positivity, and EGFR expression have been correlated with favorable responses of head and neck cancer patients treated with radiotherapy (RT) with or without chemotherapy. However, a possible correlation of HPV/p16 and EGFR status on the effect of RT in combination with cetuximab has not been sufficiently investigated. MATERIALS AND METHODS: We analyzed tumor samples for p16 and EGFR expression and correlated these variables with treatment outcome. Cox-proportional-hazard regression models were applied to compare the risk of death among patients stratified according to risk factors. Survival was estimated by the Kaplan-Meier method. Results were compared with an institutional historical control group treated without cetuximab and with published data. RESULTS: Expression of p16 was predominantly found in oropharyngeal squamous cell cancer patients (OPSCC; 36.6% positivity; 92% of all cases), while EGFR was expressed at high levels in all tumor subsites (82%). p16 expression was associated with improved overall survival in irradiated OPSCC patients (2-year overall survival of 80% in p16-positive vs. 33% overall survival in p16-negative patients). In a multivariable analysis covering all tumor sites, nodal stage (> N2a vs. ≤ N2a) and tumor site (OPSSC vs. non-OPSCC) had an impact on overall survival. CONCLUSION: Our results show that p16 positivity is associated with a favorable outcome in OPSCC patients treated with RT and cetuximab.

[Role of miRNA in malignoma of the head and neck]. / Die Rolle von miRNA bei Kopf-Hals-Malignomen.

Despite optimized therapeutic strategies, the long-term survival of head and neck squamous cell carcinomas (HNSCC) has improved in recent years only slightly. Most studies on the tumor cell genome focus on protein-coding genes (exons). Data on changes within the non-coding sequences (introns) are limited. miRNAs (microRNA, miR) are small non-coding single-stranded RNAs that control gene expression at the posttranscriptional level by interacting with the mRNA. miRNA functions include many biological processes and control up to 50 % of human genes. They can have oncogenic or tumor suppressive functions. Altered expression patterns of miRNAs are involved in carcinogenesis and tumor progression even in HNSCC, or those processes (increased resistance to radiation or chemotherapy) that could be responsible for the poor long-term prognosis by forming metastases and inoperable local recurrences. Therefore, we here present miRNA groups, which are involved in these processes and may serve as new potential therapeutic treatment targets. miRNAs may also serve as biomarkers for early diagnosis, evaluation and monitoring of treatment and tumor recurrence. Alterations in miRNA expression before and after chemotherapy are therefore of high interest. In the long run, this knowledge could lead to more effective therapies with improved therapeutic outcome of HNSCC.

Prognostic value of HIF-1α expression during fractionated irradiation.

BACKGROUND AND PURPOSE: Hypoxia and reoxygenation are important determinants of outcome after radiotherapy. HIF-1α is a key molecule involved in cellular response to hypoxia. HIF-1α expression levels have been shown to change after irradiation. The objective of the present study was to explore the prognostic value of HIF-1α expression during fractionated irradiation. MATERIALS AND METHODS: Six human squamous cell carcinoma models xenografted in nude mice were analysed. Tumours were excised after 3, 5 and 10 fractions. HIF-1α expression was quantified by western blot. For comparative analysis, previously published data on local tumour control data and pimonidazole hypoxic fraction was used. RESULTS: HIF-1α expression in untreated tumours exhibited intertumoural heterogeneity and did not correlate with pimonidazole hypoxic fraction. During fractionated irradiation the majority of tumour models exhibited a decrease in HIF-1α expression, whereas in UT-SCC-5 no change was observed. Neither kinetics nor expression levels during fractionated irradiation correlated with local tumour control. CONCLUSION: Our data do not support the use of HIF-1α determined during treatment as a biomarker to predict outcome after fractionated irradiation.

Basal HIF-1α expression levels are not predictive for radiosensitivity of human cancer cell lines.

BACKGROUND AND PURPOSE: High levels of hypoxia inducible factor (HIF)-1α in tumors are reported to be associated with tumor progression and resistance to therapy. To examine the impact of HIF-1α on radioresistance under normoxia, the sensitivity towards irradiation was measured in human tumor cell lines that differ significantly in their basal HIF-1α levels. MATERIAL AND METHODS: HIF-1α levels were quantified in lysates of H1339, EPLC-272H, A549, SAS, XF354, FaDu, BHY, and CX- tumor cell lines by ELISA. Protein levels of HIF-1α, HIF-2α, carbonic anhydrase IX (CA IX), and GAPDH were assessed by Western blot analysis. Knock-down experiments were performed using HIF-1α siRNA. Clonogenic survival after irradiation was determined by the colony forming assay. RESULTS: According to their basal HIF-1α status, the tumor cell lines were divided into low (SAS, XF354, FaDu, A549, CX-), intermediate (EPLC-272H, BHY), and high (H1339) HIF-1α expressors. The functionality of the high basal HIF-1α expression in H1339 cells was proven by reduced CA IX expression after knocking-down HIF-1α. Linear regression analysis revealed no correlation between basal HIF-1α levels and the survival fraction at either 2 or 4 Gy in all tumor cell lines investigated. CONCLUSION: Our data suggest that basal HIF-1α levels in human tumor cell lines do not predict their radiosensitivity under normoxia.

Squamous cell carcinomas of the head and neck in Fanconi anemia: risk, prevention, therapy, and the need for guidelines.

Fanconi anemia (FA) is a rare recessive DNA repair disorder that is clinically characterized by congenital malformations, progressive bone marrow failure, and increased incidence of malignancies, especially acute myeloid leukemia and squamous cell carcinomas of the head and neck (HNSCCs) and the anogenital regions. On a cellular level, typical features of the disorder are a high degree of genomic instability and an increased sensitivity to bi-functionally alkylating agents. So far, germ-line defects in 15 different FA genes have been identified. Some of these FA genes are also established as tumor susceptibility genes for familiar cancers.In recent years, the prevention and therapy of HNSCCs in FA patients has become more important as the percentage of patients surviving into adulthood is rising. HNSCCs appear in very young FA patients without common risk factors. Since cisplatin-based chemotherapy in combination with radiotherapy, essential parts of the standard treatment approach for sporadic HNSCCs, cannot be used in FA patients due to therapy-associated toxicities and mortalities even with reduced dosing, surgery is the most important treatment option for HNSCCs, in FA patients and requires an early and efficient detection of malignant lesions. So far, no uniform treatment protocol for the management of HNSCCs in FA patients exists. Therefore, we propose that the information on affected FA patients should be collected worldwide, practical therapeutic guidelines developed and national treatment centers established.

[Translational approaches in cancer stem cell research]. / Translationsansätze in der Tumorstammzellforschung.

There has been a growing body of evidence in recent years to indicate that the presence of cancer stem cells may be responsible for tumour development and early recurrence after conventional therapy strategies such as surgery, radiation, or chemotherapy. Although this concept of a small subpopulation of cancer cells with stem cell properties is not new as such and was already discussed by Virchow decades ago, the identification of cells of this kind in human malignancies was first successful in 1997 in acute myeloid leukemia. The recent identification of cancer stem cells and the detection of their fundamental signalling pathways (e.g. Hedgehog, Notch) may offer new therapeutic options in the future and become part of a therapeutic concept. In this article, we introduce the cancer stem cell model, provide an overview of current cancer stem cell markers in different human malignancies as well as head and neck squamous cell carcinoma, and discuss studies on the first targeted therapies against cancer stem cells and their signalling pathways.

[HPV-associated head and neck cancer. The basics of molecular and translational research]. / Humane Papillomaviren bei Kopf-Hals-Karzinomen. Molekulare und translationale Grundlagen.

Translational research refers to the interfaces between preclinical research and targeted short- and medium-term developments through to clinical standards. There are two distinct groups of oropharyngeal malignancies: those caused by tobacco and alcohol abuse and those caused by HPV infection. Although the prognosis of patients in the latter group is significantly better, this is not taken into consideration in the choice of treatment. However, less intensive use of radiotherapy, chemotherapy, or surgery, as well as targeted multimodal therapeutic approaches, is under research. This article summarizes the main events in the HPV life cycle, with emphasis on carcinogenic mechanisms and potential new molecular targets. Identifying distinct tumor entities of the oropharynx enables the design and development of new preventive and therapeutic strategies to reduce the incidence and mortality of HPV-associated oropharyngeal cancers in the near future.

[Translational research in head and neck cancer. Biological characteristics and general aspects]. / Translationale Forschung bei Kopf-Hals-Tumoren. Biologische Besonderheiten und generelle Problematik.

Translational research in head and neck oncology is subject to the same laws as all other solid tumors. It is based on the one hand on a solid framework of well prepared clinical studies and / or workflows according to consensus criteria with comparable documentation of clinical outcomes, while on the other on methodolgically solid and reproducible laboratory research within an effeciently interacting network. Translationally applicable single molecular markers from basic research [with the exception of p16(INK4a) as a surrogate marker for human papillomavirus (HPV)] have not found their way into clinical routine in head and neck squamous cell carcinoma (HNSCC). "Correlated gene sets" and "metagenes", including genetic profiling (omics) within clinically characterized patient groups, play an increasing role in the translational research of HNSCC. Although methodological problems currently hinder clinical oncological research, increasing focus on translational research can be observed.

[Paragangliomas and paraganglioma syndromes]. / Paragangliome und Paragangliomsyndrome.

Paragangliomas are rare tumors of neural crest origin. They are benign in the majority of cases and are characterized by a strong vascularisation. In the head and neck region they most commonly occur as carotid body tumors. Jugulotympanic and especially vagal paragangliomas are seen less frequently. Complete surgical resection represents the only curative treatment option even though resection of locally advanced tumors regularly results in lesions of the lower cranial nerves and major vessels. Approximately 30% of all head and neck paragangliomas (HNPs) are hereditary and associated with different tumor syndromes. The paraganglioma syndromes 1, 3 and 4 (PGL 1, 3 and 4) make up the majority of those familial cases. PGL 1 is associated with mutations of the succinate dehydrogenase subunit D (SDHD) gene, PGL 3 is caused by SDHC and PGL 4 by SDHB gene mutations. Multiple HNPs and the occurrence of HNPs together with pheochromocytomas are seen in SDHD as well as SDHB mutation carriers. In patients with SDHB mutations the risk for the development of malignant paraganglial tumors is significantly higher compared to SDHC and SDHD patients as well as patients with sporadic tumors. SDHC mutation carriers almost exclusively present with benign HNPs that are unifocal in the majority of cases. The role of transmission is autosomal dominant for all 3 symptoms. Interestingly, there is a "parent-of-origin-dependent-inheritance" in subjects with SDHD gene mutations. This means that the disease phenotype may only become present if the mutation is inherited through the paternal line. We recommend screening for mutations of the genes SDHB, SDHC and SDHD in all patients with HNPs. Certain clinical parameters can help to set up the order in which the 3 genes should be tested.

[Innovations in biomarker development: a pathway towards individualized cancer therapy?]. / Moderne Methoden der Biomarkerentwicklung: eine Chance für die individualisierte Tumortherapie?

Despite multiple medical and scientific achievements, cancer remains a leading cause of death worldwide. Next to imaging technologies, molecular methods for early detection and for monitoring of the course of disease are of increasing interest. Thus, over the past years numerous studies have focused on the identification of biomarkers for cancer diagnosis, prognosis and response to therapy. The study of biomarkers seems to pose a high degree of complexity because many different types of molecules may, in principle, serve as potential biomarkers. In addition, these molecules can be produced either by the tumor or by the tumor-host in response to the presence of cancer. In this review the authors will address several major topics encompassed by the field of biomarker research. They will discuss the primary sources from which biomarker candidates can be 'mined' as well as the technological or methodological challenges associated with identification of biomarkers. Furthermore, the review will focus on current biomarker candidates for head and neck squamous cell carcinoma (HNSCC), with particular interest on several molecules yielding potential relevance for detection and prognosis of this type of cancer. Finally, several biomarker candidates with predictive potential for the response to therapy of HNSCC patients will be discussed, since identifying such molecules is crucial for developing individually-tailored and improved therapeutic strategies.

Current potential and limitations of molecular diagnostic methods in head and neck cancer.

Traditional diagnostic methods such as clinical assessment, histopathological examination and imaging techniques are limited in their capacity to provide information on prognosis and treatment choice of head and neck cancer. In recent years, molecular techniques have been developed that enabled us to get more insight into the molecular biological cellular pathways underlying tumor progression and metastasis. Correlation of these molecular changes with clinical events has been explored. However, consistently useful markers have not been identified yet, although many promising developments are in progress. It may be expected that in the near future, molecular markers will be useful for clinical purposes. In this paper, an overview will be given of the several molecular techniques that may have potential to be introduced in clinical practice in the management of head and neck squamous cell carcinoma.

Human papillomavirus in head and neck tumors: epidemiological, molecular and clinical aspects.

High-risk types of human papillomaviruses (HR HPV) play an important role in the etiology of a group of head and neck squamous cell cancers (HNSCC). This review is focused on epidemiological, molecular, and clinical aspects of HPV infection in head and neck cancer. High risk HPV DNA is being detected in a very different proportion of HNSCC with the highest prevalence in oropharynx. Patients with HPV-associated tumors are characterized by moderate tobacco and alcohol consumption. Some aspects of sexual behavior may represent a risk factor. Recently, it has been shown that HPV infection is spreading and the rising prevalence of HPV-positive tumors can probably be attributed to this epidemic. On molecular level the viral oncoproteins E6 and E7 were shown to be involved in oncogenesis. HPV-positive cancers have better prognosis and HPV status should be considered in clinical decision-making. The rising proportion of HPV-positive tumors underlines the importance of HPV vaccination also for the prevention of HNSCC.

Oral squamous cell carcinoma: overview of current understanding of aetiopathogenesis and clinical implications.

MEDLINE contains well over 14,000 papers revealed by a search using keywords 'oral squamous cell cancer' or 'oral squamous cell carcinoma', over 27,000 using 'oral carcinoma' and over 48,000 using the keywords 'oral cancer'. It is difficult to see how clinicians could keep abreast of such a subject. This paper attempts to help by providing an overview of the aetiopathogenesis of oral squamous cell carcinoma (OSCC), discussing changes in epidemiology and increasing awareness of the wide range of risk factors, emphasising the genetic background to cancer susceptibility and the genetic changes associated with progression to OSCC and highlighting clinical implications.

Associations between variants of the 8q24 chromosome and nine smoking-related cancer sites.

Recent genome-wide association studies identified key single nucleotide polymorphisms (SNPs) in the 8q24 region to be associated with prostate cancer. 8q24 SNPs have also been associated with colorectal cancer, suggesting that this region may not be specifically associated to just prostate cancer. To date, the association between these polymorphisms and tobacco smoking-related cancer sites remains unknown. Using epidemiologic data and biological samples previously collected in three case-control studies from U.S. and Chinese populations, we selected and genotyped one SNP from each of the three previously determined "regions" within the 8q24 loci, rs1447295 (region 1), rs16901979 (region 2), and rs6983267 (region 3), and examined their association with cancers of the lung, oropharynx, nasopharynx, larynx, esophagus, stomach, liver, bladder, and kidney. We observed noteworthy associations between rs6983267 and upper aerodigestive tract cancers [adjusted odds ratio (ORadj), 1.69; 95% confidence interval (95% CI), 1.28-2.24], particularly in oropharynx (ORadj, 1.80; 95% CI, 1.30-2.49) and larynx (ORadj, 2.04; 95% CI, 1.12-3.72). We also observed a suggestive association between rs6983267 and liver cancer (ORadj, 1.51; 95% CI, 0.99-2.31). When we stratified our analysis by smoking status, rs6983267 was positively associated with lung cancer among ever-smokers (ORadj, 1.45; 95% CI, 1.05-2.00) and inversely associated with bladder cancer among ever-smokers (ORadj, 0.35; 95% CI, 0.14-0.83). Associations were observed between rs16901979 and upper aerodigestive tract cancer among never-smokers and between rs1447295 and liver cancer among ever-smokers. Our results suggest variants of the 8q24 chromosome may play an important role in smoking-related cancer development. Functional and large epidemiologic studies should be conducted to further investigate the association of 8q24 SNPs with smoking-related cancers.