Purpose: This study was designed to identify mitochondrial (mt) DNA variations in primary and metastatic uveal melanoma (UM) cell lines and their relation with cell metabolism to gain insight into... Show morePurpose: This study was designed to identify mitochondrial (mt) DNA variations in primary and metastatic uveal melanoma (UM) cell lines and their relation with cell metabolism to gain insight into metastatic progression.Method: The entire mtDNA genomes were sequenced using Sanger sequencing from two primary UM cell lines (92.1 and MEL270) and two cell lines (OMM2.3 and OMM2.5) derived from liver metastases of the MEL270 patient. The mtDNA copy numbers determined by the ratio of nDNA versus mtDNA. qRT-PCR was used to evaluate expression levels of mitochondrial biogenesis genes.Results: Sequencing showed that cell line MEL270 and metastases-derived OMM2.3 and OMM2.5 cell lines had homoplasmic single nucleotide polymorphisms (SNPs) representing J1c7a haplogroup, whereas 92.1 cells had mtDNA H31a haplogroup. mtDNA copy numbers were significantly higher in primary cell lines. The metastatic UM cells showed down-regulation of POLG, TFAM, NRF-1 and SIRT1 compared to their primary MEL270 cells. PGC-1 alpha was downregulated in 92.1 and upregulated in MEL270, OMM2.3 and OMM2.5.Conclusions: Our finding suggests that within metastatic cells, the heteroplasmic SNPs, copy numbers and mitochondrial biogenesis genes are modulated differentially compared to their primary UM cells. Therefore, investigating pathogenic mtDNA variants associated with cancer metabolic susceptibility may provide future therapeutic strategies in metastatic UM. (C) 2021 Published by Elsevier Inc. Show less
BackgroundTo understand how to improve the effect of immune checkpoint inhibitors in uveal melanoma (UM), we need a better understanding of the expression of PD-1 and PD-L1, their relation with the... Show moreBackgroundTo understand how to improve the effect of immune checkpoint inhibitors in uveal melanoma (UM), we need a better understanding of the expression of PD-1 and PD-L1, their relation with the presence of tumor-infiltrating lymphocytes (TILs), and their prognostic relevance in UM patients. Materials and methodsExpression of PD-1 and PD-L1 was assessed in 71 UM tissue samples by immunohistochemistry and quantitative real-time PCR (qRT-PCR), and further validated by western blotting. The effect of interferon gamma (IFN-gamma) on PD-1/PD-L1 expression was determined on four UM cell lines. ResultsImmunoreactivity of PD-1 was found in 30/71 cases and of PD-L1 in 44/71 UM samples. Tumor-infiltrating lymphocytes were found in 46% of UM tissues. PD-1 was expressed on TILs while tumor cells expressed PD-L1. UM with and without TILs showed expression of PD-1 in 69% and 18% cases, respectively (p=0.001). Similarly, PD-L1 was found in 75% of UM with TILs and in 50% of cases without TILs, respectively (p=0.03). DFS rate were lower in patients with TILs with expression of PD-1 and PD-L1, but the rate of DFS was higher with expression of PD-L1 in patients without TILs. After treatment of UM cell lines with IFN-gamma, PD-1 expression was induced in all UM cell lines whereas PD-L1 expression was found at a lower level in untreated cells, while expression also increased following treatment with IFN-gamma .ConclusionOur study suggests that increased infiltration with TILs promotes the aggressive behavior and suppresses the immune response of UM cells, thereby inhibiting immunotherapy. Show less
