137 THE ROLE OF LYSYL OXIDASE ON PROANGIOGENIC PHENOTYPES OF TUMOR ENDOTHELIAL CELLS

Abstract

INTRODUCTION AND OBJECTIVES: The molecules which are highly expressed in tumor endothelial cell (TEC) are important for targeting TEC specifically to inhibit tumor angiogenesis. However, there are still a few reports about TEC marker, because it is difficult to isolate TEC. We found LOX gene was upregulated in TEC compared to normal endothelial cell (NEC) by DNA microarray analysis in three different types of human tumor xenografts in nude mouse including renal cell carcinoma (RCC). LOX is an enzyme which is reported to play a critical role in the metastatic potential of tumor, possibly promoting tumor cell trafficking into the premetastatic niche in various types of malignancies. However, the effect of LOX on TEC has not been reported. To investigate whether LOX might be a TEC-specific target in cancer therapy, we analyzed its expression in clinical sample and examined the function of LOX on TEC. METHODS: We successfully purified human TEC (hTEC) and NEC (hNEC) from surgically resected RCC and normal parenchyma from six patients, respectively. Using cultured hTEC and hNEC, competitive reverse transcription-polymerase chain reaction (RT-PCR) was performed. LOX expression level was investigated using immunohistochemistry (IHC). To explore the role of LOX in TEC, mouse TEC (mTEC) and NEC (mNEC) were isolated from human tumor xenografts in nude mouse and the normal counterparts, respectively. To investigate the function of LOX, LOX was knock-downed using siRNA and TEC phenotypic changes were analyzed by migration assay and vascular tube formation assay. In addition, we investigated the effect of -aminopropionitrile (BAPN), an irreversible inhibitor of LOX, on tumor angiogenesis and metastatic colonization in vivo. RESULTS: LOX expression levels were higher in isolated TEC than NEC both in human (Tissues: clear cell carcinoma 6; Stage T1, 3; T3, 3; Grade 1, 1; Grade 2, 2; Grade 3, 3) and mouse by RT-PCR. In IHC, LOX was detected in blood vessels in human RCC, not in normal tissue. Cell migration and tube formation of mTEC were suppressed by LOX knock-down using siRNA with decrease in phosphorylation of focal adhesion kinase (Tyr397). Furthermore, BAPN specifically inhibited tumor angiogenesis and micrometastasis in vivo, via inhibition of LOX activity of TEC. CONCLUSIONS: Our findings suggest that LOX upregulated in TEC contributes in their proangiogenic phenotype and plays key roles in tumor metastasis. LOX might be a TEC-specific target and be useful for cancer therapeutic purposes.