Recently we showed that Lupeol, a dietary triterpene, activates apoptotic machinery, decreases serum- PSA levels and inhibits the tumorigenicity of androgen-sensitive prostate cancer (PCa) cells. Current study was conducted to understand the mechanistic basis of the effect of Lupeol in androgen-sensitive (LNCaP) PCa cells. Lupeol (30-50μM) treatment (for 21 days) was observed to inhibit the proliferative and clonogenic potential of LNCaP cells. Lupeol treated cells exhibited reduced expression level of PCNA protein, a marker of proliferation. To understand the mechanism of action of Lupeol, we first determined the effect of Lupeol in a focused microarray of 288-well characterized PCa-associated genes. We found that Lupeol (20 μM) significantly modulates the expression level of genes which are known to be associated with proliferation and survival. Lupeol treatment decreased the mRNA expression level of insulin growth factor receptor-1, myc, cyclin D1, ERBB2 and Jun, and caused an increase in the mRNA expression level of IGFBP6, STEAP1, TIMP3 and KLK10. Immunoblot analysis indicated that Lupeol-induced modulations in the mRNA expression levels of these genes corroborated well with the expression level of their products (proteins). Interestingly, genes which are modulated by Lupeol are also known to be associated directly or indirectly with β-catenin signaling which is highly aberrant in PCa patients. We therefore investigated the effect of Lupeol on the upstream regulators of β-catenin signaling including GSK3β/Axin complex. Treatment of cells with Lupeol (5-30 μM) significantly reduced levels of β-catenin in the cytoplasm as well as in nuclear fractions with an increase in the expression levels of GSK3β/Axin complex, thus rendering β-catenin prone to degradation. Next, we employed luciferase reporter assay to measure the transcriptional activity of TCF responsive element, a marker for β-catenin signaling, and observed that Lupeol caused a significant reduction in the transcriptional activation of TCF responsive element in pTK-TCF-Luc-transfected LNCaP cells. We further observed that Lupeol treatment significantly decreased the expression level of matrix metalloproteinase (MMP)-2 and MMP-7 proteins which are known as downstream targets of β-catenin signaling. Treatment of cells with Lupeol significantly reduced the enzymatic activity of MMP-2. Finally, we evaluated the effect of Lupeol treatment on the transcriptional activity of MMP-2. We observed that Lupeol treatment significantly reduced the transcriptional activity of MMP-2 gene in pGL2-MMP-2-Luc-transfected LNCaP cells. We conclude that Lupeol being a multi-target agent effectively targets IGFR1/β-catenin axis thus leading to the inhibition in the proliferation of PCa cells. We suggest that Lupeol could be developed as an agent for chemoprevention as well as chemotherapy of human PCa..

99th AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA