Abstract
BackgroundImmune checkpoint inhibition and in particular anti-PD-1 immunotherapy have revolutionized the treatment of advanced melanoma. In this regard, higher tumoral PD-L1 protein (gene name: CD274) expression is associated with better clinical response and increased survival to anti-PD-1 therapy. Moreover, there is increasing evidence that tumor suppressor proteins are involved in immune regulation and are capable of modulating the expression of immune checkpoint proteins. Here, we determined the role of p53 protein (gene name: TP53) in the regulation of PD-L1 expression in melanoma.MethodsWe analyzed publicly available mRNA and protein expression data from the cancer genome/proteome atlas and performed immunohistochemistry on tumors with known TP53 status. Constitutive and IFN-ɣ-induced PD-L1 expression upon p53 knockdown in wildtype, TP53-mutated or JAK2-overexpressing melanoma cells or in cells, in which p53 was rendered transcriptionally inactive by CRISPR/Cas9, was determined by immunoblot or flow cytometry. Similarly, PD-L1 expression was investigated after overexpression of a transcriptionally-impaired p53 (L22Q, W23S) in TP53-wt or a TP53-knockout melanoma cell line. Immunoblot was applied to analyze the IFN-ɣ signaling pathway.ResultsFor TP53-mutated tumors, an increased CD274 mRNA expression and a higher frequency of PD-L1 positivity was observed. Interestingly, positive correlations of IFNG mRNA and PD-L1 protein in both TP53-wt and -mutated samples and of p53 and PD-L1 protein suggest a non-transcriptional mode of action of p53. Indeed, cell line experiments revealed a diminished IFN-ɣ-induced PD-L1 expression upon p53 knockdown in both wildtype and TP53-mutated melanoma cells, which was not the case when p53 wildtype protein was rendered transcriptionally inactive or by ectopic expression of p53L22Q,W23S, a transcriptionally-impaired variant, in TP53-wt cells. Accordingly, expression of p53L22Q,W23S in a TP53-knockout melanoma cell line boosted IFN-ɣ-induced PD-L1 expression. The impaired PD-L1-inducibility after p53 knockdown was associated with a reduced JAK2 expression in the cells and was almost abrogated by JAK2 overexpression.ConclusionsWhile having only a small impact on basal PD-L1 expression, both wildtype and mutated p53 play an important positive role for IFN-ɣ-induced PD-L1 expression in melanoma cells by supporting JAK2 expression. Future studies should address, whether p53 expression levels might influence response to anti-PD-1 immunotherapy.
Highlights
Immune checkpoint inhibition and in particular anti-programmed cell death-1 (PD-1) immunotherapy have revolutionized the treatment of advanced melanoma
gene coding for p53 protein (TP53)-mutated melanoma present higher gene coding for programmed-death-ligand 1 (PD-L1) protein (CD274) mRNA expression levels To allow for an easier discrimination between mRNA and protein in our manuscript, we refer to the official gene names for mRNA (TP53 and CD274) and to the widely-used molecule names p53 and PD-L1 (CD274) for protein expression
For melanoma CD274 mRNA levels were significantly higher in TP53-mutated than in TP53wt samples (Fig. 1a; p = 0.0181; Mann-Whitney)
Summary
Immune checkpoint inhibition and in particular anti-PD-1 immunotherapy have revolutionized the treatment of advanced melanoma In this regard, higher tumoral PD-L1 protein (gene name: CD274) expression is associated with better clinical response and increased survival to anti-PD-1 therapy. Antibodies directed against the cytotoxic T-lymphocyteassociated antigen 4 (CTLA-4) receptor or programmed cell death-1 (PD-1) receptor have revolutionized the systemic therapy of advanced melanoma [1]. With these immunotherapeutic approaches durable responses in the treatment of metastatic melanoma have been achieved for the first time [2,3,4]. PD-L1 positivity (> 5% or > 1% of tumor cells positive for PD-L1 staining) is associated with a better overall response rate (ORR), progression free survival (PFS) and overall survival (OS) to anti-PD-1 immunotherapies [3, 4, 6, 8,9,10]
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