Human Uveal Melanoma Cells Express CXCR4 and a Synthetic Polypeptide CXCR4 Antagonist Decreases Hepatic Micrometastasis in Mouse Uveal Melanoma Model

Abstract

Purpose:: CXC chemokine receptor-4 (CXCR4) has been reported as the most frequently up-regulated chemokine receptor in hematopoietic and solid malignancies. The purpose of this study is to determine the level of CXCR4 expression in human uveal melanoma cells and mouse melanoma cell lines, to understand whether a synthetic polypeptide against CXCR4 inhibits hepatic micrometastases in a mouse uveal melanoma model, and to evaluate the effect of a synthetic polypeptide CXCR4 antagonist administrated before and after removal of the primary tumor.

Methods:: Total RNAs were prepared from cultured human uveal melanoma Mel290, Mel270, 02-1486 and OMM2.3 cells and mouse B16LS9, B16F10, Queens, metB16LS9 and metQueens cell lines with Trizol. Reverse transcription (RT)-PCR was performed to determine the expression level of CXCR4 mRNA. The synthetic polypeptide was designed as a CXCR4 antagonist containing a 14-mer peptide (TN14003). B16LS9 cells were inoculated into the posterior compartment of the 24 mice. The mice were divided into three groups: Group 1, received IP 1mM synthetic polypeptide (TN14003) three times a week starting at 3 days before inoculation; Group 2, received IP 1mM TN14003 three times a week starting at the day of enucleation; Group 3, received IP PBS three times a week. The inoculated eyes were enucleated at the 7^th day after inoculation, the mice were sacrificed and livers were collected at the 28^th day after inoculation. Immunohistochemical staining for TGF ß₂, Ki-67, MMP-2 was performed on the histology sections and the hepatic micrometastases were counted.

Results:: All human uveal melanoma cells and mouse melanoma cell lines expressed CXCR4 mRNA. The average number of hepatic micrometastases in each groups was: Group 1: 67.43±27.36; Group 2: 171.14±65.87; Group 3: 145.71±112.35. Differences between Group 1 and Group 2 (p<0.01), Group 1 and Group 3 (p<0.05) were significant. There was no significant difference between Group 2 and Group 3. Immunostaining demonstrated decreased expression of TGFß₂, Ki-67 and MMP-2 in Group 1.

Conclusions:: Human uveal melanoma expresses CXCR4. As a synthetic polypeptide CXCR4 antagonist effectively inhibits hepatic micrometastases in a mouse uveal melanoma model. Timing of administration of this anti-metastasic agent may be a crucial factor relative to efficacy.

Keywords: melanoma • uvea