Mass spectrometry imaging-based proteomic analysis to differentiate melanocytic nevi and malignant melanoma

Question: Histological criteria for the diagnosis of melanocytic skin lesions such as benign nevus and malignant melanoma may be equivocal. We used mass spectrometry imaging (MSI) technology to determine proteomic differences between these lesions. These proteomic patterns may assist in the differential diagnosis. Methods: Formalin-fixed paraffin-embedded tissue from cutaneous melanomas (n = 27) and melanocytic nevi (n = 12) were cut at 3 μm. One section per case was mounted onto a conductive glass slide for IMS analysis, while the adjacent serial section was used for hematoxylin and eosin (HE) staining and annotation of the tumor area. Unstained sections were processed for deparaffination, heat-induced epitope retrieval, in-situ trypsin digestion and matrix application. From these sections, mass spectral data were acquired at a spatial resolution of 50 μm using a rapifleX MALDI Tissuetyper mass spectrometer. The histology-annotated image was merged to an image of the MSI section and spectra were collected from each tumor annotation from each section. The MSI data were then imported into SCiLS Lab software and R for processing, generation of peptide profiles and statistical analysis. Results: Spectra were extracted from both benign and malignant lesions and compared. Principal Component Analysis highlighted spectra patterns correlating with melanoma and nevus tissue. Comparison of the mass spectra average profiles revealed several significant peptide peaks (P< 0.001, AUC >0.7) between the two tissue types. Specifically, 137 and 89 peptides were overexpressed in melanoma and nevus, respectively. These peaks were used to generate a Linear Discriminant Analysis classification model, which could discriminate melanoma from nevus with an accuracy of 100%. Some significant discriminant peptides such as m/z 976.5, 1287.5, 1410.6, 1428.7, were identified as actin, macrophage migration inhibitory factor, CK5, and vimentin. Conclusion: A proteomic signature discerning melanoma from benign nevus was established using MSI and might be used as a supplement to standard histology.