Expression and activation of matrix metalloproteinase-2 (MMP-2) and its co-localization with membrane-type 1 matrix metalloproteinase (MT1-MMP) correlate with melanoma progression

Matrix metalloproteinases (MMPs) and their specific tissue inhibitors (TIMP s) play an important role in cancer cell invasion and metastasis. Recently, it was shown that the presence of activated MMP-2 correlates with melanoma progression in vitro. This activation involves coordinated expression of M MP-2, membrane-type 1 MMP (MT1-MMP), and TIMP-2. To investigate the express ion profile of these enzymes in human melanoma, this study used tumour spec imens obtained from both a human melanoma xenograft model, consisting of ei ght melanoma cell lines with different metastatic capacity in nude mice, an d 60 fresh human cutaneous melanocytic Lesions comprising all stages of mel anocytic tumour progression. MT1-MMP and TIMP-2 mRNA and protein were prese nt in all cell lines. Cell surface expression level of MT1-MMP, as determin ed by flow cytometry, was similar on all cell lines. In addition, western b lot analysis revealed that both inactive and active MT1-MMP protein was exp ressed by all cell lines. MMP-2 mRNA and the pro-enzyme form of MMP-2 were expressed by all cell lines. Remarkably, the presence of functionally activ e MMP-2 was restricted to the most aggressive cell lines MV3 and BLM. Rever se transcription-polymerase chain reaction (RT-PCR) analysis of RNA isolate d from subcutaneous xenografts revealed MT1-MMP and TIMP-2 mRNA expression in all lesions, whereas MMP-2 mRNA could be detected only in xenografts der ived from the highly metastatic cell lines 1F6m, MV3, and BLM. Furthermore, immunohistochemistry demonstrated a marked increase of MMP-2 and MT1-MMP i n MV3 and BLM xenografts, whereas TIMP-2 expression showed no evident corre lation with metastatic capacity. In human cutaneous melanocytic lesions, MM P-2, MT1-MMP, and TIMP-2 mRNA were detectable by RT-PCR in all lesions. Exp ression of MMP-2 protein was not detectable, either in common and atypical naevi, or in melanoma in situ by immunohistochemistry. In these lesions, he terogeneous expression of MT1-MMP and TIMP-2 was present in melanocytic cel ls. In contrast, a large number of MMP-2 and MT1-MMP-positive tumour cells were observed in primary melanomas and melanoma metastases, Double staining experiments and immunohistochemistry on serial sections from the same lesi ons demonstrated that all tumour cells expressing MMP-2 also expressed MT1- MMP and TIMP-2. Finally, zymography of melanoma metastases revealed that MM P-2 was present in its functionally active form. This study demonstrates th at expression of MT1-MMP and TIMP-2 and activation of MMP-2 are correlated with tumour progression both in the xenograft model and in human melanocyti c lesions, strongly suggesting that these factors are required for melanoma invasion and metastasis formation. Copyright (C) 2000 John Wiley & Sons, L td.