Methylation testing in plasma is a powerful diagnostic tool for molecular disease staging, risk stratification, and disease monitoring. Patients with initially high biomarker levels might benefit from intensified treatment and posttherapeutic surveillance. The early detection of a recurrent/metastatic disease or a second malignancy could lead to an earlier consecutive treatment, thereby improving patients' outcomes.

BackgroundSquamous cell carcinoma of the head and neck region (HNSCC) is a common malignant disease accompanied by a high risk of local or distant recurrence after curative-intent treatment. Biomarkers that allow for the prediction of disease outcome can guide clinicians with respect to treatment and surveillance strategies. Here, the methylation status of PITX2 and an adjacent lncRNA (PANCR) were evaluated for their ability to predict overall survival in HNSCC patients.ResultsPITX2 hypermethylation was associated with a better overall survival (hazard ratio, HR = 0.51, 95%CI: 0.35-0.74, p<0.001), while PANCR hypermethylation was significantly associated with an increased risk of death (HR = 1.64, 95%CI: 1.12-2.39, p=0.010).MethodsQuantitative, methylation-specific real-time PCR assays for PITX2 and PANCR were employed to measure bisulfite-converted DNA from formalin-fixed, paraffin-embedded (FFPE) tissues in a cohort of 399 patients with localized or locally advanced HNSCC who received curative-intent treatment (surgery with optional adjuvant radiochemotherapy or definite radiochemotherapy).ConclusionsPITX2 and PANCR methylation status were shown to be independent predictors for overall survival in HNSCC patients. Tissue-based methylation testing could therefore potentially be employed to identify patients with a high risk for death who might benefit from a more radical or alternative treatment.

Abstract. Background/Aim: Ectopic myoglobin (MB)expressionMyoglobin (MB) is a cytoplasmatic monomeric oxygenbinding hemoprotein known to be localized mainly in cardiac myocytes and mitochondrial-rich skeletal muscles. In contrast to its relative haemoglobin, MB is only found in the blood stream after muscle injury. The functional paradigm of MB is the storage and intracellular transport of oxygen (O 2 ) (1, 2) to the mitochondria for oxidative phosphorylation within the myocyte. However, MB also maintains homeostasis of the nitric oxide free radical (NO•) in muscle through scavenging or producing NO• molecules (3, 4) and appropriating excessive reactive oxygen species (ROS) (5). In addition, MB is able to bind fatty acids with physiological binding constants in vitro (6). On the other hand, MB-knockout mice are viable and exhibit normal exercise capacity but their survival depends on the activation of compensating mechanisms, i.e. increasing capillary densities and haematocrit (7,8).In the last years, the existence of non-muscle MB has been increasingly acknowledged, demonstrating MB, e.g., in the liver, brain or gills of carps (9) and in humans, 6235

BackgroundMolecular biomarkers assisting risk-group assignment and subsequent treatment stratification are urgently needed for patients with squamous cell cancer of the head and neck region (HNSCC). Aberrant methylation is a frequent event in cancer and, therefore, a promising source for potential biomarkers. Here, the methylation status of the paired-like homeodomain transcription factor 3 (PITX3) was evaluated in HNSCC.MethodsUsing a quantitative real-time PCR, PITX3 methylation was assessed in a cohort of 326 HNSCC patients treated for localized or locally advanced disease (training cohort). The results were validated with Infinium HumanMethylation450 BeadChip data from a 528 HNSCC patient cohort (validation cohort) generated by The Cancer Genome Atlas (TCGA) Research Network.Results PITX3 methylation was significantly higher methylated in tumor compared to normal adjacent tissue (NAT; training cohort: median methylation NAT 32.3%, tumor 71.8%, p < 0.001; validation cohort: median methylation NAT 16.9%, tumor 35.9%, p < 0.001). PITX3 methylation was also significantly correlated with lymph node status both in the training (p = 0.006) and validation (p < 0.001) cohort. PITX3 methylation was significantly higher in HPV-associated (p16-positive) tumors compared to p16-negative tumors (training cohort: 73.7 vs. 66.2%, p = 0.013; validation cohort: 40.0 vs. 33.1%, p = 0.015). Hypermethylation was significantly associated with the risk of death (training cohort: hazard ratio (HR) = 1.80, [95% confidence interval (CI) 1.20–2.69], p = 0.005; validation cohort: HR = 1.43, [95% CI 1.05–1.95], p = 0.022). In multivariate Cox analyses, PITX3 added independent prognostic information. Messenger RNA (mRNA) expression analysis revealed an inverse correlation with PITX3 methylation in the TCGA cohort.Conclusions PITX3 DNA methylation is an independent prognostic biomarker for overall survival in patients with HNSCC and might aid in the process of risk stratification for individualized treatment.

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