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BACKGROUND: Guidelines remain unclear over whether patients with early stage oral cancer without overt neck disease benefit from upfront elective neck dissection (END), particularly those with the smallest tumours. METHODS: We conducted a randomised trial of patients with stage T1/T2 N0 disease, who had their mouth tumour resected either with or without END. Data were also collected from a concurrent cohort of patients who had their preferred surgery. Endpoints included overall survival (OS) and disease-free survival (DFS). We conducted a meta-analysis of all six randomised trials. RESULTS: Two hundred fifty randomised and 346 observational cohort patients were studied (27 hospitals). Occult neck disease was found in 19.1% (T1) and 34.7% (T2) patients respectively. Five-year intention-to-treat hazard ratios (HR) were: OS HR = 0.71 (p = 0.18), and DFS HR = 0.66 (p = 0.04). Corresponding per-protocol results were: OS HR = 0.59 (p = 0.054), and DFS HR = 0.56 (p = 0.007). END was effective for small tumours. END patients experienced more facial/neck nerve damage; QoL was largely unaffected. The observational cohort supported the randomised findings. The meta-analysis produced HR OS 0.64 and DFS 0.54 (p < 0.001). CONCLUSION: SEND and the cumulative evidence show that within a generalisable setting oral cancer patients who have an upfront END have a lower risk of death/recurrence, even with small tumours. CLINICAL TRIAL REGISTRATION: NIHR UK Clinical Research Network database ID number: UKCRN 2069 (registered on 17/02/2006), ISCRTN number: 65018995, ClinicalTrials.gov Identifier: NCT00571883.
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Carcinoma de Células Escamosas/cirugía , Procedimientos Quirúrgicos Electivos/métodos , Neoplasias de la Boca/cirugía , Disección del Cuello/métodos , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma de Células Escamosas/patología , Estudios de Cohortes , Supervivencia sin Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias de la Boca/epidemiología , Neoplasias de la Boca/patología , Cuello/inervación , Cuello/fisiopatología , Cuello/cirugía , Estadificación de Neoplasias , Ensayos Clínicos Controlados Aleatorios como Asunto , Resultado del TratamientoRESUMEN
BACKGROUND: Exosomes are extracellular vesicles released by almost all cell types, including cancer cells, into bodily fluids such as saliva, plasma, breast milk, semen, urine, cerebrospinal fluid, amniotic fluid, synovial fluid and sputum. Their key function being intercellular communication with both neighbouring as well as distant cells. Cancer exosomes have been shown to regulate organ-specific metastasis. However, little is known about the functional differences and molecular consequences of normal cells responding to exosomes derived from normal cells compared to those derived from cancer cells. METHODS: Here, we characterised and compared the transcriptome profiles of primary human normal oral keratinocytes (HNOK) in response to exosomes isolated from either primary HNOK or head and neck squamous cell carcinoma (HNSCC) cell lines. RESULTS: In recipient HNOK cells, we found that regardless of normal or cancer derived, exosomes altered molecular programmes involved in matrix modulation (MMP9), cytoskeletal remodelling (TUBB6, FEZ1, CCT6A), viral/dsRNA-induced interferon (OAS1, IFI6), anti-inflammatory (TSC22D3), deubiquitin (OTUD1), lipid metabolism and membrane trafficking (BBOX1, LRP11, RAB6A). Interestingly, cancer exosomes, but not normal exosomes, modulated expression of matrix remodelling (EFEMP1, DDK3, SPARC), cell cycle (EEF2K), membrane remodelling (LAMP2, SRPX), differentiation (SPRR2E), apoptosis (CTSC), transcription/translation (KLF6, PUS7). We have also identified CEP55 as a potential cancer exosomal marker. CONCLUSIONS: In conclusion, both normal and cancer exosomes modulated unique gene expression pathways in normal recipient cells. Cancer cells may exploit exosomes to confer transcriptome reprogramming that leads to cancer-associated pathologies such as angiogenesis, immune evasion/modulation, cell fate alteration and metastasis. Molecular pathways and biomarkers identified in this study may be clinically exploitable for developing novel liquid-biopsy based diagnostics and immunotherapies.
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Proteínas de Ciclo Celular/genética , Exosomas/genética , Perfilación de la Expresión Génica/métodos , Neoplasias de Cabeza y Cuello/genética , Proteínas Nucleares/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Exosomas/patología , Proteína Forkhead Box M1/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias de Cabeza y Cuello/patología , Humanos , Queratinocitos/citología , Queratinocitos/metabolismo , Queratinocitos/patología , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Carcinoma de Células Escamosas de Cabeza y Cuello/patologíaRESUMEN
Histopathological discordance with molecular phenotype of many human cancers poses clinically challenging tasks for accurate cancer diagnosis, which impacts on treatment strategy and patient outcome. Hence, an objective, accurate and quantitative method is needed. A quantitative Malignancy Index Diagnostic System (qMIDS) was developed based on 14 FOXM1 (isoform B)-associated genes implicated in the regulation of the cell cycle, differentiation, ageing, genomic stability, epigenetic and stem cell renewal, and two reference genes. Their mRNA expression levels were translated via a prospectively designed algorithm, into a metric scoring system. Subjects from UK and Norway (n = 299) provided 359 head and neck tissue specimens. Diagnostic test performance was assessed using detection rate (DR) and false-positive rate (FPR). The median qMIDS scores were 1.3, 2.9 and 6.7 in healthy tissue, dysplasia and head and neck squamous cell carcinomas (HNSCC), respectively (UK prospective dataset, p<0.001); 1.4, 2.3 and 7.6 in unaffected, oral lichen planus, or HNSCC, respectively (Norwegian retrospective dataset with up to 19 years survival data, p<0.001). At a qMIDS cut-off of 4.0, DR was 94% and FPR was 3.2% (Norwegian dataset); and DR was 91% and FPR was 1.3% (UK dataset). We further demonstrated the transferability of qMIDS for diagnosing premalignant human vulva (n = 58) and skin (n = 21) SCCs, illustrating its potential clinical use for other cancer types. This study provided evidence that qMIDS was able to quantitatively diagnose and objectively stratify cancer aggressiveness. With further validation, qMIDS could enable early HNSCC detection and guide appropriate treatment. Early treatment intervention can lead to long-term reduction in healthcare costs and improve patient outcome.
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Biomarcadores de Tumor/genética , Carcinoma de Células Escamosas/diagnóstico , Factores de Transcripción Forkhead/genética , Neoplasias de Cabeza y Cuello/diagnóstico , Lesiones Precancerosas/diagnóstico , Neoplasias Cutáneas/diagnóstico , Neoplasias de la Vulva/diagnóstico , Algoritmos , Carcinoma de Células Escamosas/genética , Células Cultivadas , Diagnóstico Precoz , Femenino , Proteína Forkhead Box M1 , Perfilación de la Expresión Génica , Redes Reguladoras de Genes , Neoplasias de Cabeza y Cuello/genética , Humanos , Queratinocitos/citología , Queratinocitos/metabolismo , Metástasis Linfática , Masculino , Persona de Mediana Edad , Noruega , Lesiones Precancerosas/genética , Pronóstico , Estudios Prospectivos , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Neoplasias Cutáneas/genética , Neoplasias de la Vulva/genéticaRESUMEN
BACKGROUND: Epigenetic reprogramming of the methylome has been implicated in all stages of cancer evolution. It is now well accepted that cancer cells exploit epigenetic reprogramming, a mechanism that regulates stem/progenitor cell renewal and differentiation, to promote cancer initiation and progression. The oncogene FOXM1 has been implicated in all major forms of human cancer. METHODS: We have recently shown that aberrant upregulation of FOXM1 orchestrated a DNA methylation signature that mimics the cancer methylome landscape, from which we have identified a number of FOXM1-induced epigenetic markers. Differential promoter methylation and gene expression in clinical specimens were measured using commercially available bisulfite conversion kits and absolute quantitative PCR, respectively. RESULTS: Here, we investigated 8 FOXM1-induced differentially methylated genes, SPCS1, FLNA, CHPF, GLT8D1, C6orf136, MGAT1, NDUFA10, and PAFAH1B3, using human head and neck tissue specimens donated by 2 geographically independent patient cohorts from Norway and the United Kingdom. Two genes (GLT8D1 and C6orf136) were found to be differentially expressed in head and neck squamous cell carcinomas (HNSCCs). Using methylation-specific quantitative PCR, we confirmed that the promoters of GLT8D1 and C6orf136 were hypo- and hypermethylated, respectively, in HNSCC tissues. CONCLUSIONS: Given that epigenetic change precedes gene expression, methylation status of candidate genes identified from this study may represent a signature of premalignancy, rendering them potentially useful predictive biomarkers for precancer screening and/or therapeutic targets for cancer prevention.
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Biomarcadores de Tumor/genética , Carcinoma de Células Escamosas/genética , Factores de Transcripción Forkhead/genética , Neoplasias de Cabeza y Cuello/genética , Metilación de ADN , Epigénesis Genética , Femenino , Proteína Forkhead Box M1 , Regulación Neoplásica de la Expresión Génica , Glicosiltransferasas/genética , Humanos , Masculino , Persona de Mediana Edad , Noruega , Regiones Promotoras Genéticas , Estudios Retrospectivos , Carcinoma de Células Escamosas de Cabeza y Cuello , Reino Unido , Regulación hacia ArribaRESUMEN
BACKGROUND: Heterogeneity in oral potentially malignant disorder (OPMD) poses a problem for accurate prognosis that impacts on treatment strategy and patient outcome. A holistic assessment based on gene expression signatures from both the tumour cells and their microenvironment is necessary to provide a more precise prognostic assessment than just tumour cell signatures alone. METHODS: We reformulated our previously established multigene qPCR test, quantitative Malignancy Index Diagnostic System (qMIDS) with new genes involved in matrix/stroma and immune modulation of the tumour microenvironment. An algorithm calculates and converts a panel of 16 gene mRNA expression levels into a qMIDS index to quantify risk of malignancy for each sample. RESULTS: The new qMIDSV2 assay was validated in a UK oral squamous cell carcinoma (OSCC) cohort (n = 282) of margin and tumour core samples demonstrating significantly better diagnostic performance (AUC = 0.945) compared to previous qMIDSV1 (AUC = 0.759). Performance of qMIDSV2 were independently validated in Chinese (n = 35; AUC = 0.928) and Indian (n = 95; AUC = 0.932) OSCC cohorts. Further, 5-year retrospective analysis on an Indian dysplastic lesion cohort (n = 30) showed that qMIDSV2 was able to significantly differentiate between lesions without transformation and those with malignant transformation. CONCLUSIONS: This study validated a novel multi-gene qPCR test on a total of 535 tissue specimens from UK, China and India, demonstrating a rapid minimally invasive method that has a potential application for dysplasia risk stratification. Further study is required to establish if qMIDSV2 could be used to improve OPMD patient management, guide treatment strategy and reduce oral cancer burden.
RESUMEN
Metastasis of oral squamous cell carcinoma (OSCC) to the cervical lymph nodes has a significant impact on prognosis. Accurate staging of the neck is important in order to deliver appropriate treatment for locoregional control of the disease and for prognosis. The management of the neck in early, low volume disease (clinically T1/T2 oral cavity tumours) has long been debated. The risk of occult nodal involvement in cT1/T2 OSCC is estimated around 20-30%. We describe the natural evolutionary history of OSCC and its patterns of spread and metastasis to the local lymphatic basins. We discuss most published literature and studies on management of the clinically negative neck (cN0). Particular focus is given to prospective randomized trials comparing the outcomes of upfront elective neck dissection against the observational stance, and we summarize the results of the sentinel node biopsy studies. The paper discusses the significance of the primary tumour histological characteristics and specifically the tumour's depth of invasion (DOI) and its impact on predicting nodal metastasis. The DOI has been incorporated in the TNM staging highlighting its significance in aiding the treatment decision making and this is reflected in world-wide oncological guidelines. The critical analysis of all available literature amalgamates the existing evidence in early OSCC and provides recommendations in the management of the clinically N0 neck.
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Carcinoma de Células Escamosas/cirugía , Neoplasias de la Boca , Humanos , Disección del Cuello , Estadificación de Neoplasias , Estudios Prospectivos , Biopsia del Ganglio Linfático CentinelaRESUMEN
BACKGROUND: Major ablative surgery in the head and neck region may create composite defects involving the oral mucosa, bone and the overlying facial skin. The large surface area and the three-dimensional nature of these defects pose a difficult reconstructive challenge requiring adequate bone and large, positionally versatile skin flaps. PATIENTS AND METHODS: From September 1993 to May 2000, 19 patients with through-and-through osteocutaneous defects of the mouth and face were reconstructed with composite subscapular artery system flaps. The evaluated parameters included: (i) site and dimensions of the tissue defect; (ii) specific flap properties; and (iii) review of the recipient and donor site morbidity. RESULTS: 10 variants of scapular osteocutaneous flaps, eight latissimus dorsi with serratus anterior and rib osteo-myocutaneous flaps, and one combination of an osteocutaneous scapular and myocutaneous latissimus dorsi flap were used to reconstruct composite facial defects with mean dimensions of: skin 54.4 cm(2), mucosa 56.2 cm(2) and bone of 8.2 cm. Ischaemic complications occurred in three patients including one total flap failure and one failure of the bony component in previously irradiated patients. The third flap was successfully salvaged. No significant long-term donor site morbidity was noted. CONCLUSION: Composite flaps based on the subscapular artery system are a versatile reconstructive modality for large through-and-through defects of the mouth and face.
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Cara/cirugía , Huesos Faciales/cirugía , Boca/cirugía , Procedimientos de Cirugía Plástica/métodos , Colgajos Quirúrgicos , Carcinoma de Células Escamosas/cirugía , Huesos Faciales/lesiones , Femenino , Humanos , Masculino , Neoplasias Mandibulares/cirugía , Neoplasias de la Boca/cirugía , Invasividad Neoplásica , Heridas por Arma de Fuego/cirugíaRESUMEN
OBJECTIVE: To compare anxieties of general dental practitioners (GDPs) across the UK in communicating with patients about oral cancer and confidence in clinical skills required to perform soft tissue screening for oral cancer. DESIGN: A questionnaire was sent to 2200 randomly selected GDPs from across the UK. Responses to the questionnaires were analysed using 95% confidence intervals. SETTING: Dental practitioners in general practice within England, Northern Ireland, Scotland and Wales. RESULTS: The response rate varied between 57% in England and 65% in Northern Ireland. A high percentage of dental practitioners across all UK regions reported performing soft tissue examinations (range 78% to 88%). The number of soft tissue examinations per month varied between 129 (95% CI 109, 148) and 162 (95% CI 154, 170) indicating criteria when selecting patients for screening. Using a nine-point rating scale (1 = not at all, 9 = extremely), confidence in the clinical skills required during oral cancer screening was generally good (ratings varying between 5.4 and 6.7). With the exception of reporting positive findings to patients (rating 4.5 to 5.2), anxiety in communication skills used during oral cancer screening was generally low (ratings varying between 1.8 and 3.9). CONCLUSION: While concerns over generalizing the results exist, the situation with respect to the clinical and communication skills required by GDPs during oral cancer screening is generally encouraging. An area of concern is discussing positive findings with patients. This may be overcome by developing specialist courses on breaking bad news within undergraduate dental curricula and programmes of continuing professional development.
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Odontología General , Neoplasias de la Boca/diagnóstico , Adulto , Ansiedad , Competencia Clínica , Comunicación , Femenino , Humanos , Masculino , Autoimagen , Encuestas y Cuestionarios , Reino UnidoRESUMEN
Diseases of the thyroid are common and surgical treatment is often the preferred option. Thyroid surgery is becoming subspecialised and falls within the repertoire of maxillofacial, and head and neck surgeons. Multidisciplinary management of most patients with diseases of the thyroid is key to providing the best care particularly for those with malignancies and retrosternal extension. To reduce postoperative complications a meticulous search for, and protection of the recurrent laryngeal nerve and parathyroid glands, with an incision along the skin crease in the lower neck, which can be extended for neck dissection, are paramount. Recent advances in thyroid surgery include ultrasound-guided cervical plexus block, use of the Harmonic Scalpel(®) (Ethicon Endo-Surgery, Inc., USA), intraoperative nerve stimulation to monitor the recurrent laryngeal nerve, use of TissuePatch™ 3 (Tissuemed Ltd., Leeds, UK) adhesive sealant, and minimal access surgery.
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Disección del Cuello/métodos , Glándulas Paratiroides/cirugía , Nervio Laríngeo Recurrente/cirugía , Enfermedades de la Tiroides/cirugía , Humanos , Disección del Cuello/instrumentación , Complicaciones Posoperatorias/prevención & control , Neoplasias de la Tiroides/cirugíaRESUMEN
The oncogene FOXM1 has been implicated in all major types of human cancer. We recently showed that aberrant FOXM1 expression causes stem cell compartment expansion resulting in the initiation of hyperplasia. We have previously shown that FOXM1 regulates HELLS, a SNF2/helicase involved in DNA methylation, implicating FOXM1 in epigenetic regulation. Here, we have demonstrated using primary normal human oral keratinocytes (NOK) that upregulation of FOXM1 suppressed the tumour suppressor gene p16(INK4A) (CDKN2A) through promoter hypermethylation. Knockdown of HELLS using siRNA re-activated the mRNA expression of p16(INK4A) and concomitant downregulation of two DNA methyltransferases DNMT1 and DNMT3B. The dose-dependent upregulation of endogenous FOXM1 (isoform B) expression during tumour progression across a panel of normal primary NOK strains (nâ=â8), dysplasias (nâ=â5) and head and neck squamous cell carcinoma (HNSCC) cell lines (nâ=â11) correlated positively with endogenous expressions of HELLS, BMI1, DNMT1 and DNMT3B and negatively with p16(INK4A) and involucrin. Bisulfite modification and methylation-specific promoter analysis using absolute quantitative PCR (MS-qPCR) showed that upregulation of FOXM1 significantly induced p16(INK4A) promoter hypermethylation (10-fold, P<0.05) in primary NOK cells. Using a non-bias genome-wide promoter methylation microarray profiling method, we revealed that aberrant FOXM1 expression in primary NOK induced a global hypomethylation pattern similar to that found in an HNSCC (SCC15) cell line. Following validation experiments using absolute qPCR, we have identified a set of differentially methylated genes, found to be inversely correlated with in vivo mRNA expression levels of clinical HNSCC tumour biopsy samples. This study provided the first evidence, using primary normal human cells and tumour tissues, that aberrant upregulation of FOXM1 orchestrated a DNA methylation signature that mimics the cancer methylome landscape, from which we have identified a unique FOXM1-induced epigenetic signature which may have clinical translational potentials as biomarkers for early cancer screening, diagnostic and/or therapeutic interventions.
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Carcinoma de Células Escamosas/genética , Metilación de ADN , Epigenómica , Factores de Transcripción Forkhead/genética , Neoplasias de Cabeza y Cuello/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Células Cultivadas , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , ADN (Citosina-5-)-Metiltransferasa 1 , ADN (Citosina-5-)-Metiltransferasas/genética , ADN (Citosina-5-)-Metiltransferasas/metabolismo , ADN Helicasas/genética , ADN Helicasas/metabolismo , ADN Metiltransferasa 3A , Proteína Forkhead Box M1 , Factores de Transcripción Forkhead/metabolismo , Perfilación de la Expresión Génica , Neoplasias de Cabeza y Cuello/metabolismo , Neoplasias de Cabeza y Cuello/patología , Humanos , Immunoblotting , Regiones Promotoras Genéticas/genética , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
BACKGROUND: Keratin 15 (K15) is a type I keratin that is used as a marker of stem cells. Its expression is restricted to the basal layer of stratified epithelia, and the bulge in hair follicles. However, in certain clinical situations including oral lichen planus, K15 is induced in suprabasal layers, which is inconsistent with the role of a stem cell marker. This study provides insights into the mechanisms of K15 expression in the basal and differentiating keratinocytes. METHODOLOGY/PRINCIPAL FINDINGS: Human keratinocytes were differentiated by three different methods; suspension in methylcellulose, high cell density and treatment with phorbol ester. The expression of mRNA was determined by quantitative PCR and protein by western blotting and immunostaining. Keratinocytes in suspension suppressed ß1-integrin expression, induced differentiation-specific markers and K15, whereas FOXM1 (a cell cycle regulated protein) and K14 were downregulated. Rescuing ß1-integrin by either fibronectin or the arginine-glycine-aspartate peptide suppressed K15 but induced K14 and FOXM1 expression. Specific inhibition of PKCδ, by siRNA, and AP-1 transcription factor, by TAM67 (dominant negative c-Jun), suppressed K15 expression, suggesting that PKC/AP-1 pathway plays a role in the differentiation-specific expression of K15. The basal cell-specific K15 expression may involve FOXM1 because ectopic expression of the latter is known to induce K15. Using chromatin immunoprecipitation, we have identified a single FOXM1 binding motif in the K15 promoter. CONCLUSIONS/SIGNIFICANCE: The data suggests that K15 is induced during terminal differentiation mediated by the down regulation of ß1-integrin. However, this cannot be the mechanism of basal/stem cell-specific K15 expression in stratified epithelia, because basal keratinocytes do not undergo terminal differentiation. We propose that there are two mechanisms regulating K15 expression in stratified epithelia; differentiation-specific involving PKC/AP-1 pathway, and basal-specific mediated by FOXM1, and therefore the use of K15 expression as a marker of stem cells must be viewed with caution.
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Factores de Transcripción Forkhead/metabolismo , Queratina-15/metabolismo , Queratinocitos/metabolismo , Proteína Quinasa C-delta/metabolismo , Transducción de Señal , Factor de Transcripción AP-1/metabolismo , Western Blotting , Diferenciación Celular , Células Cultivadas , Células Epidérmicas , Epidermis/metabolismo , Fibronectinas/metabolismo , Técnica del Anticuerpo Fluorescente , Proteína Forkhead Box M1 , Perfilación de la Expresión Génica , Humanos , Queratinocitos/citología , Oligopéptidos/metabolismo , Proteína Quinasa C-delta/antagonistas & inhibidores , Proteína Quinasa C-delta/genética , ARN Interferente Pequeño/genética , Piel/citología , Piel/metabolismo , Células Madre/citología , Células Madre/metabolismoRESUMEN
Malignant tumors of deep head and neck structures can invade skin, but the tumor periphery is difficult to assess clinically. The surgeon's dilemma is achieving tumor clearance with safe margins while at the same time minimizing skin loss on the face. We show, in 2 cases involving the face, that high-resolution diagnostic ultrasound was superior to CT scan in demonstrating the periphery of the tumor. The tumor was distinguished from surrounding edema by its lower echogenicity and homogeneous echotexture. The maximum contour of the tumor was marked on the skin surface with ink under ultrasound guidance. The ink marking aided excision and reconstruction planning. Subsequent histology showed the surgical margins were clear of tumor. The patients remained tumor-free for more than 3 years. Ultrasound imaging therefore shows good potential for planning surgical resection with a safe margin and for aiding decisions on donor site and type of flap for reconstruction.