William B Coman’s research while affiliated with Princess Alexandra Hospital (Queensland Health) and other places

With an increasing recognition of the link between oral and systemic disease, attention has turned to saliva as an alternative diagnostic medium for a diverse array of health conditions [1]. Compared with blood, saliva collection is non-invasive, easy sampling with multiple sampling opportunities, does not need pre-processing and is ideal for 3 rd world countries [2]. It is well established that tumour cells secrete biomolecules into the saliva [3]. Head and neck squamous cell carcinoma (HNSCC) encompasses a diverse group of aggressive tumours. HNSCC is the most distressing and disfiguring tumour for the patient to endure, for health workers to manage, and families to cope with. HNSCC patients, particularly those with a history of smoking, often develop secondary tumours. Currently, there are no diagnostic tests to detect these cancers at an early stage; as such, most patients present with metastatic disease at the time of diagnosis (regional nodal involvement in 43% and distant metastasis in 10%), leading to 5-year survival rates of less than 60% [4]. DNA methylation and microRNAs (miRNAs) are the most extensively studied epigenetic biomarkers in HNSCC [5]. We collected saliva (resting saliva and buccal swabs, DNA • SAL™) from HNSCC patients and healthy controls and interrogated CpG hypermethylation events in tumour suppressor genes using a sensitive methylation-specific PCR (MSP) assay. RASSF1a, DAPK1 and p16, showed an overall specificity of 87% and sensitivity of 80%. The test panel performed extremely well in the detection of the early stages of HNSCCs, with a sensitivity of 94% and specificity of 87%, and a high κ value of 0.8, indicating an excellent overall agreement between the presence of HNSCC and a positive MSP panel result. In addition, miR-9 and miR-191 provided a good discriminative ability with AUC values of 0.76 and 0.73 respectively (p<0.01) for discriminating HNSCC patients from healthy controls. In conclusion, we demonstrate that salivary DNA methylation and miRNA biomarkers are clinically useful in detecting HNSCC in a non-invasive manner.

Background MicroRNAs (miRNAs) are known to play an important role in cancer development by post-transcriptionally affecting the expression of critical genes. The aims of this study were two-fold: (i) to develop a robust method to isolate miRNAs from small volumes of saliva and (ii) to develop a panel of saliva-based diagnostic biomarkers for the detection of head and neck squamous cell carcinoma (HNSCC). Methods Five differentially expressed miRNAs were selected from miScript™ miRNA microarray data generated using saliva from five HNSCC patients and five healthy controls. Their differential expression was subsequently confirmed by RT-qPCR using saliva samples from healthy controls (n = 56) and HNSCC patients (n = 56). These samples were divided into two different cohorts, i.e., a first confirmatory cohort (n = 21) and a second independent validation cohort (n = 35), to narrow down the miRNA diagnostic panel to three miRNAs: miR-9, miR-134 and miR-191. This diagnostic panel was independently validated using HNSCC miRNA expression data from The Cancer Genome Atlas (TCGA), encompassing 334 tumours and 39 adjacent normal tissues. Receiver operating characteristic (ROC) curve analysis was performed to assess the diagnostic capacity of the panel. Results On average 60 ng/μL miRNA was isolated from 200 μL of saliva. Overall a good correlation was observed between the microarray data and the RT-qPCR data. We found that miR-9 (P

Head and neck cancers (HNCs) represent a significant and ever-growing burden to the modern society, mainly due to the lack of early diagnostic methods. A significant number of HNCs is often associated with drinking, smoking, chewing beetle nut, and human papilloma virus (HPV) infections. We have analyzed DNA methylation patterns in tumor and normal tissue samples collected from head and neck squamous cell carcinoma (HNSCC) patients who were smokers. We have identified novel methylation sites in the promoter of the mediator complex subunit 15 (MED15/PCQAP) gene (encoing a co-factor important for regulation of transcription initiation for promoters of many genes), hypermethylated specifically in tumor cells. Two clusters of CpG dinucleotides methylated in tumors, but not in normal tissue from the same patients, were identified. These CpG methylation events in saliva samples were further validated in a separate cohort of HNSCC patients (who developed cancer due to smoking or HPV infections) and healthy controls using methylation-specific PCR (MSP). We used saliva as a biological medium because of its non-invasive nature, close proximity to the tumors, easiness and it is an economically viable option for large-scale screening studies. The methylation levels for the two identified CpG clusters were significantly different between the saliva samples collected from healthy controls and HNSCC individuals (Welch's t-test returning P < 0.05 and Mann-Whitney test P < 0.01 for both). The developed MSP assays also provided a good discriminative ability with AUC values of 0.70 (P < 0.01) and 0.63 (P < 0.05). The identified novel CpG methylation sites may serve as potential non-invasive biomarkers for detecting HNSCC.

Conclusion: Alpha B-crystallin was found to be an independent prognostic marker for poor prognosis in oral cavity tumours. For oropharyngeal cancer, alpha B-crystallin had no prognostic value. Objective: The aim of this study was to see if earlier findings of alpha B-crystallin as an independent prognostic marker, and SPARC/osteonectin, PAI-1 and uPA as a prognostic combination for poor outcome in squamous cell carcinoma (SCC) of the head and neck could be confirmed in a new set of tumours. Methods: In a consecutive series of patients, assessed and primarily treated at a tertiary referral centre, histological sections from 55 patients with oral and SCC (OOPHSSC) with complete clinical data and follow-up were obtained. Oral and oropharyngeal tumours were studied separately. Immunohistochemical detection of alpha B-crystallin, SPARC/osteonectin, PAI-1 and uPA expression was performed. Results: Thirty-five patients had an oral tumour and 20 patients an oropharyngeal tumour. Twenty-five oral tumours stained negatively and 10 positively for alpha B-crystallin. For oropharyngeal tumours the figures were 15 negatively and 5 positively. Median disease-specific survival (DSS) for both sites was 33.8 and 11.9 months, for negative and positive alpha B-crystallin staining, respectively (p = 0.046). For the oral cavity, median DSS was 27.3 months for negative tumours and 7.5 months for positive tumours (p = 0.012). Corresponding figures for oropharyngeal tumours were 33.8 and 34.1 months (p = 0.95). Thus, significance in survival was only found in oral cavity tumours. In multivariate analyses there were no significant differences in DSS in the oropharyngeal group when adjusted for tumour size (T status) and presence of neck node metastasis (N status). In the oral cavity group, the significantly better DSS for negative tumours became even stronger when adjusted for T and N status. No statistical difference was found in DSS between positive and negative staining for SPARC/osteonectin, PAI-1 or uPA.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer in the world based on its mortality, and approximately 900,000 new cases are diagnosed each year with 300,000 deaths per annum. The known major risk factors for the development of SCC include smoking, drinking and human papilloma viral infections. Currently there are no biomarkers to detect HNSCC at an early stage and as a result 5-year survival is less than 50%. DNA promoter hypermethylation of tumour suppressor genes occurs in cancer initiation and progression. In this study, we collected saliva from HNSCC patients and healthy controls and using a sensitive methylation-specific PCR (MSP) assay based on detection of specific CpG hypermethylation events in the promoters of tumour suppressor genes (APC, p14, RASSF1a, DAPK1 and p16), for RASSF1a, DAPK1 and p16, we have demonstrated an overall specificity for this test panel of 73% (53.8-87.2%) for HNSCC patients (n=121) when compared with healthy controls (n=41). In addition, APC and p14 were methylated in a small subset of patients. The DNA methylation of DAPK1, p16 and RASSF1, in the saliva (drool and DNA • SAL™) enabled us to identify individuals with HNSCC at a sensitivity level of 60% (p=0.05 confidence interval 51-68%) and an accuracy of 63% (56-68%), with a Fisher's exact test p=0.002. In conclusion, we demonstrate that salivary DNA methylation biomarkers are clinically useful in detecting head and neck cancer in a non-invasive manner.

Background: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cause of cancer mortality in the world and the 5th most commonly occurring cancer. Tobacco smoking, alcohol consumption and human papilloma virus (HPV) infections have been associated with the occurrence of HNSCC. Despite advances that have been made in HNSCC treatment, smoking-associated HNSCC patients still exhibit a poor 5 year survival rate (30-50 %) and a concomitant poor quality of life. The major clinical challenge to date lies in the early detection of dysplastic lesions,which can progress to malignancy. In addition, there are currently no tools available to monitor HNSCC patients for early stages of local recurrences or distant metastases. In the recent past, micro-RNAs (miRNA) have been assessed for their role in cancer initiation and progression, including HNSCC. It is now well-established that deregulation of these single stranded, small non-coding, 19-25 nt RNAs can e.g. enhance the expression of oncogenes or subdue the expression of tumor suppressor genes. The aims of this review are three-fold: first to retrieve from the literature miRNAs that have specifically been associated with HNSCC, second to group these miRNAs into those regulating tumor initiation, progression and metastasis, and third to discern miRNAs related to smoking-associated HNSCC versus HPV-associated HNSCC development. Conclusions: This review gives an overview on the miRNAs regulating the development of head and neck cancers. The ultimate establishment of miRNA expression profiles that are HNSCC specific, and miRNAs that orchestrate altered gene and protein expression levels in HNSCC, could pave the way for a better understanding of the mechanism underlying its pathogenesis and the development of novel, targeted therapies.

Head and neck squamous cell carcinoma (HNSCC) accounts for a bulk of the oral and laryngeal cancers, the majority (70%) of which are associated with smoking and excessive drinking, major known risk factors for the development of HNSCC. In contrast to reports that suggest an inverse relationship between smoking and global DNA CpG methylation, hypermethylation of promoters of a number of genes was detected in saliva collected from patients with HNSCC. Using a sensitive methylation-specific polymerase chain reaction (MSP) assay to determine specific methylation events in the promoters of RASSF1A, DAPK1, and p16 genes, we demonstrate that we can detect tumor presence with an overall accuracy of 81% in the DNA isolated from saliva of patients with HNSCC (n = 143) when compared with the DNA isolated from the saliva of healthy nonsmoker controls (n = 31). The specificity for this MSP panel was 87% and the sensitivity was 80% (with a Fisher exact test P < .0001). In addition, the test panel performed extremely well in the detection of the early stages of HNSCCs, with a sensitivity of 94% and a specificity of 87%, and a high κ concordance value of 0.8, indicating an excellent overall agreement between the presence of HNSCC and a positive MSP panel result. In conclusion, we demonstrate that the promoter methylation of RASSF1A, DAPK1, and p16 MSP panel is useful in detecting hypermethylation events in a noninvasive manner in patients with HNSCC.

Malignancies of the nasal septum are rare diseases and fewer than 400 cases were reported. The understanding of the disease is limited due to its rarity. We present a series of patients with nasal septum malignancies, who were referred to the Princess Alexandra Hospital, Ear, Nose and Throat Department from 2007 to 2010. Seventeen patients were found to have nasal septum malignancies. The average age was 59.5 years old (range: 36 to 83 years old). The commonest initial symptom on presentation was nasal obstruction (nine out of 17, 53%), seconded by epistaxis (eight out of 17, 47%). The average time from the initial onset of symptoms to presentation averaged 18.8 months (range: 1 to 48 months). The commonest physical finding on presentation was nasal masses (11 out of 17, 65%), followed by nasal septum ulcers (four out of 17, 24%). The histology of the lesions was predominantly squamous cell carcinoma. The mean duration of follow-up was 24.7 months. The overall 3-year survival was 81.9% with the relapse free survival 66.7%. Nasal septum malignancies are highly treatable with good prognoses when in early stages. They required high degree of suspicion to be detected early. Treatment options include surgical resection and radiotherapy and they offered similar 3-year survival rate. Combined therapy is adopted in larger tumours; however, it is not verified with randomized trials. Vigilant follow-up is vital to detect early recurrence, which is common in advanced stage lesions.