br Authorship contributions br Disclosure of conflicts of in

Authorship contributions
Disclosure of conflicts of interest
Acknowledgements
Introduction Melanomas are among the most commonly occurring cancers. Crude incidence rates in Australia (AIHW, 2014) and the USA (SEER, 2014) were approximately 50 cases (in 2010) and 20 cases (in 2011) per 100,000 respectively. With the number of new cases rising each year, melanoma is currently is listed as the 4th and 6th most common cancer in Australia and the USA respectively (AIHW, 2014; SEER, 2014). Current clinical staging criteria classify melanoma progression from a pre-invasive lesion, confined to the herpes simplex virus 2 (stage 0), a series of early stages of local invasion (I and II), a stage involving regional lymph nodes (stage III) and finally metastasis to distant sites (stage IV). The overall 5-year survival for melanoma is 91%, which is largely due to curative surgery for early stage disease. However, cure rates are <15% (Balch et al., 2009) if distant metastasis occurs (stage IV; AJCC 7th edition). We now have evidence that current therapeutic options for late stage disease are more effective if the disease is treated with a lower disease burden (Sosman et al., 2012; Hodi et al., 2010). Hence, melanoma must be treated in earlier stages to maximise the chances of patient survival. Therefore, the ability to identify signs of melanoma progression sooner would be a valuable clinical tool. Melanoma progression biomarkers have been studied intensively with varying levels of success. Serum lactate dehydrogenase (LDH) levels have been integrated into current staging regimens (Balch et al., 2009) and elevation of LDH levels increases in specificity as disease progresses (stages II (83%), III (87%), and IV (92%)). However the sensitivity of this marker is reduced during progression (stages II (95%), III (57%), and IV (79%)) (Brochez and Naeyaert, 2000; Finck et al., 1983; Karakousis et al., 1996; Sirott et al., 1993; Weide et al., 2012; Deichmann et al., 1999). S100B, a calcium binding protein, is raised in serum of stages III and IV melanoma patients (Guo et al., 1995; Smit et al., 2005). However, the proportion of patients with elevated S100B levels varies by stage: 0–9% in stages I/II, 5–98% in stage III, and 40–100% in stage IV (reviewed in Kruijff et al., 2009). As such, serum S100B is not routinely used in the clinic (Leiter et al., 2014), highlighting the fact that the current serological methods of progression detection, whilst relatively specific, are inadequate due to variability in sensitivity across all stages of disease. To date, there are no biomarkers that are sensitive or specific enough to be beneficial for early detection of melanoma (all stages). A blood test (‘circulating’ biomarkers) that detected melanoma with regional spread, prior to clinically evident distant metastasis, could improve treatment and outcomes for melanoma patients. For a circulating biomarker to be effective, not only must it be sufficiently sensitive and specific, but it must also be highly stable and resistant to degradation. In recent years, circulating microRNAs (miRNAs) have been studied for their utility as biomarkers in a wide range of malignancies and disorders (Allegra et al., 2012; De Guire et al., 2013). miRNAs are small (20–22nt) non-coding RNAs which function to regulate gene expression in the cell. Recently, tumour cells have been shown to release miRNAs into the circulation (Mitchell et al., 2008), contained primarily in micro-vesicles or exosomes (extracellular vesicles), or bound to AGO2 — a part of the miRNA-mediated silencing complex (Allegra et al., 2012; De Guire et al., 2013). Due to the ‘encapsulation’ of these miRNAs in serum or plasma they are highly resistant to degradation by RNases (highly concentrated in the blood), thus their potential usefulness as a ‘biomarker’ is relatively high. To date, circulating melanoma-related miRNAs have been rarely studied (Fleming et al., 2015; Friedman et al., 2012).