The anticancer effects of tanshinone I in human non-small-cell lung cancer

Tanshinones are the major bioactive compounds of Salvia miltiorrhiza Bunge (Danshen), roots, which are used in many therapeutic remedies in Chinese traditional medicine. We investigated the anticancer effects of tanshinones on the highly invasive human lung adenocarcinoma cell line, CL1-5. Tanshinone I significantly inhibited migration, invasion, and gelatinase activity in macrophage-conditioned medium (CM)-stimulated CL1-5 cells in vitro and also reduced the tumorigenesis and metastasis in CL1-5-bearing severe combined immunodeficiency mice. Unlike tanshinone IIA, which induces cell apoptosis, tanshinone I had no significant cytotoxicity. Real-time quantitative polymerase chain reaction (RTQ-PCR), luciferase reporter assay, and an electrophoretic mobility shift assay revealed that tanshinone I reduces the transcriptional activity of interleukin-8 (IL-8), the angiogenic factor involved in cancer metastasis, by attenuating the DNA-binding activity of activator protein-1 and nuclear factor kappaB in CM-stimulated CL1-5 cells. Microarray and pathway analysis of tumor-related genes identified the differentially expressed genes responding to tanshinone I, and these results were validated by RTQ-PCR. The responsive genes included human platelet-derived growth factor beta chain, Shb, H-ras, N-Ras, mitogen-activated protein kinase kinase 3, phosphoinositide-3-kinase, CD44, Rac1, and collagen type IV; these genes may be associated with the Ras MAPK and Rac1 signaling pathways. These results suggest that tanshinone I exhibits anticancer effects both in vitro and in vivo, and that these effects are mediated at least partly through the IL-8, Ras MAPK, and Rac1 signaling pathways. Keywords: treatment with dose respone, cDNA array Five concentrations of tanshinone I in CM were used: 0, 0.01, 0.1, 1, 10 ug/mL; in addition to a control condition without CM. Thirty micrograms of total RNA derived from CL1-5 cells from each treatment described above were labeled with digoxigenin during reverse transcription. The averaged intensity for each gene (there were three copies on a chip for each gene) was used for the subsequent analyses. To reduce the variation arising from experimental results derived from different microarrays, the intensity values of the spots from each microarray were rescaled using a quantile normalization method and a housekeeping gene, GAPDH. To reduce background noise, any background intensity lower than 2,330 was assigned the value of 2,330. Genes were clustered into groups on the basis of expression profiles by the self-organizing maps (SOMs) algorithm using the Acuity 3.0 program (Axon Instruments, Union City, CA). After cluster analysis by the SOM method, genes whose expression profiles correlated with the concentration of tanshinone I were identified.