PO-149 Expression profiling of TP53, TP73, NME and GLI families of genes/proteins in metastatic melanoma

Abstract

IntroductionMalignant melanoma is the most aggressive form of skin cancer and resistant to available therapies, therefore new molecular approaches for better understanding of disease are needed. Although TP53 is rarely mutated in melanoma, it fails to function as a tumour suppressor. This may result from alterations in p53 family members, including the diverse isoforms of p53 and its homologue p73. Moreover, we assume that p53 functions in melanoma might be altered through interactions with small molecular weight variants of p53 and p73 isoforms, NME and GLI families of proteins. In this study, we conducted a gene/protein expression profiling for p53 and its potential interaction partners (p73/NME/GLI) in metastatic melanoma tissue.Material and methodsMetastatic melanoma and adjacent healthy skin tissues were obtained from 38 patients during surgery in the Sestre milosrdnice University Hospital Centre, Zagreb. Expression of 9 TP53 isoforms, both N- (full-length, Δ40 and Δ133) and C-terminal (α, β and γ), 2 TP73 isoforms (TAp73 and ΔNN’p73), NME1, NME2, GLI1, GLI2, GLI3 and PTCH1, was analysed by RT-qPCR. Expression of p53 (p53α, p53β, Δ40p53α, Δ133p53α, Δ133p53β and Δ160p53α isoforms), p73 (TAp53α, TAp53β, ΔNp73α and ΔNp73β), NME1, NME2, GLI1 (130 and 160 kDa isoforms), GLI2 (133 and 250 kDa) and GLI3 (activator/repressor forms) was analysed by western blot.Results and discussionsRelative expression of ‘long’ TP53 isoforms in tumour tissue was as follows: p53α>p53β > Δ40α>p53γ > Δ40β > Δ40γ. Expression of ‘short’ TP53 isoforms was: Δ133α > Δ133β > Δ133γ. Only Δ40β and Δ40γ were significantly downregulated in tumours. Expression of full length TAp73 was higher than ΔNp73, and both were significantly downregulated in tumours. Significant downregulation in tumours was also observed for PTCH1, GLI1 and GLI2; while NME1 and NME2 were generally the most expressed genes but without significant difference between healthy and tumour tissue. In addition, in metastatic melanomas the most expressed proteins were p53α and NME1, while ΔNp73β, GLI2 (250 kDa) and Δ133p53α showed lowest expression. Eight proteins showed significantly higher expression in tumours compared with healthy skin: 2 GLI1 isoforms (130 and 160 kDa), 133 kDa GLI2 isoform, NME1 and NME2, ΔNp73Δ and 2 p53 isoforms with shortest N- and longest C-terminus.ConclusionWe have shown that TP53/TP73/NME/GLI genes are generally downregulated in metastatic melanoma tissue compared with healthy skin, while, on the contrary, their protein products seem to be upregulated in tumours.