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. 2022 Oct 15;20(1):469.
doi: 10.1186/s12967-022-03668-1.

A novel microRNA signature for the detection of melanoma by liquid biopsy

Claudia Sabato #  1 , Teresa Maria Rosaria Noviello #  2   3 , Alessia Covre  4   5 , Sandra Coral  4   6 , Francesca Pia Caruso  2   3 , Zein Mersini Besharat  1 , Elena Splendiani  7 , Laura Masuelli  1 , Cecilia Battistelli  7 , Alessandra Vacca  1 , Giuseppina Catanzaro  1 , Agnese Po  7 , Andrea Anichini  8 , Michele Maio  4   5 , Michele Ceccarelli  2   3 , Anna Maria Di Giacomo #  4   5 , Elisabetta Ferretti #  9
Affiliations

A novel microRNA signature for the detection of melanoma by liquid biopsy

Claudia Sabato et al. J Transl Med. .

Abstract

Background: Melanoma is the deadliest form of skin cancer and metastatic disease is associated with a significant survival rate drop. There is an urgent need for consistent tumor biomarkers to scale precision medicine and reduce cancer mortality. Here, we aimed to identify a melanoma-specific circulating microRNA signature and assess its value as a diagnostic tool.

Methods: The study consisted of a discovery phase and two validation phases. Circulating plasma extracellular vesicles (pEV) associated microRNA profiles were obtained from a discovery cohort of metastatic melanoma patients and normal subjects as controls. A pEV-microRNA signature was obtained using a LASSO penalized logistic regression model. The pEV-microRNA signature was subsequently validated both in a publicly available dataset and in an independent internal cohort.

Results: We identified and validated in three independent cohorts a panel of melanoma-specific circulating microRNAs that showed high accuracy in differentiating melanoma patients from healthy subjects with an area under the curve (AUC) of 1.00, 0.94 and 0.75 respectively. Investigation of the function of the pEV-microRNA signature evidenced their possible immune suppressive role in melanoma patients.

Conclusions: We demonstrate that a blood test based on circulating microRNAs can non-invasively detect melanoma, offering a novel diagnostic tool for improving standard care. Moreover, we revealed an immune suppressive role for melanoma pEV-microRNAs.

Keywords: Biomarkers signature; Diagnosis; Extracellular vesicles; Liquid biopsy; Melanoma; microRNAs.

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Conflict of interest statement

The authors declare that they have no competing interests.

The authors state no conflict of interest.

Figures

Fig. 1
Fig. 1
Characterization of pEVs in melanoma patients (Patient) and normal subjects (Ctrl). A Transmission electron microscopy visualization of EV isolated from human plasma samples. Isolated EV displayed multiple vesicles with a round-shaped morphology and a diameter of 100-400 nm. Scale bars correspond to 200 nm. B Western blot analysis of common exosomal markers (HSP70, TSG101, CD63 and CD81) and cell organelle (calnexin) in whole cell lysate (WCL) of WM793 melanoma cells and EV isolated from normal subject control (Ctrl) plasma sample. WCL was loaded as positive control. C, D Size distribution and concentration of isolated pEVs from healthy donor (C) and melanoma patient (D) using Tunable Resistive Pulse Sensing method
Fig. 2
Fig. 2
pEV-microRNA profiles in melanoma patients (Patient) and normal subjects (Ctrl). Heatmap of 65 differentially expressed pEV-microRNAs (21 down-regulated in Patients vs Ctrl, 44 up-regulated in Patients vs Ctrl), in melanoma patients (blue) and normal subject control (purple) with a statistically significance of p<0.05. Each row represents an individual microRNA, each column represents an individual sample
Fig. 3
Fig. 3
Evaluation of four microRNA signature predictive performance. A Parameter selection in LASSO regression. B, C ROC and PR curves for discovery cohort. D, E ROC and PR curves for external validation (GSE20994) cohort. F Comparison of pEV-microRNA signature model accuracy with an accuracy distribution of 1000 models with random microRNA signatures
Fig. 4
Fig. 4
Absolute quantification of 4 pEV-microRNA and evaluation of diagnostic performance in an independent internal cohort. A Absolute quantification of circulating levels of 4 pEV-microRNAs (miR-412-3p; miR-507; miR-1203 and miR-362-3p) in an independent internal cohort of normal control (Ctrl) and metastatic melanoma patients (Patients) using ddPCR. Significant differences are highlighted with a starlike symbol (* p value <0.05, **, p value < 0.005), whereas not statistically significant difference with the abbreviation of “ns”. B ROC curves and AUC of 4 pEV -microRNAs in an independent internal cohort (AUC=0.75, p-value=0.008)
Fig. 5
Fig. 5
MicroRNA target genes experimental validation. Relative Luciferase activity in the HEK293T cell line following co-transfection of the 3’UTR of TNFSF4 with miR-412-3p, miR-507, and miR-1203. Values represent the means ± S.D. of values from three experiments, each performed in triplicate
See this image and copyright information in PMC

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