PLoS ONE R2 | Published in PloSOne third February 2010 | Volume 5 | Issue 2 | T e9024 aggressiveness and metastasis. To this end, we have the use of HER2 overexpression and HER2 cell lines not of breast cancer. According to previous FAK cancer statements that the activity t of lapatinib correlated with the degree of HER2 expression that we SKBR3 and BT474 cells G1 arrest of cell cycle underwent place, and died sp Ter as a reaction to this drug. Conversely, in MCF7 and MDA-MB 231 cells lapatinib had no effect on the cell cycle and the Lebensf Conductivity at concentrations up to 1 mM. SKBR3, BT474, MCF7 and MDA-MB 231 cells were treated with lapatinib and gene expression profiles of cells treated lapatinib, compared to the vehicle were determined in the tables of low density.
Lapatinib has a gr Shown eren influence on the gene expression profile of SKBR3 and BT474 cells compared to non-HER2 overexpressing cell lines MCF7 and MDA-MB 231st In Figure 1B, we have arbitrarily weight hlt Defining 0.5-fold induction and induction FTY720 S1P Receptor inhibitor .1.5 times to the genes that were down and through medicine Se treatment regulated. If one of this criterion, in SKBR3 cells, studied 6% and 16% of the genes were inhibited by lapatinib and each. In BT474, 10% of the genes were upregulated, w While 15% were upregulated. In contrast, in MCF7 cells, a single gene of 82 modulated by lapatinib, w While in MDA-MB 231 UPOR gene was identified by the Regulation on the criteria defined above. It is important to some of the observed Ver Changes in gene expression were related to known or predicted the mode of action of lapatinib.
For example, CCND1, CCNE1 and CDC25, we found that downward adjusted by lapatinib in BT474 and SKBR3 cells are all involved in the transition G1 / S cell cycle. Figure 2 Lapatinib and the PI3K inhibitor LY294002 induce Grb7 upregulation. A, SKBR3 or BT474 were 26 105 cells per well seeded in 6-well plates t keep for 24 h and then End with 300 nM lapatinib or 20 mM LY294002 for the indicated times treated. Thereafter, total RNA was isolated, and mRNA levels were GRB7 that in vehicle-treated cells compared. B, C, SKBR3 or BT474 26 105 cells per well were seeded in 6-well plates t and keep for 24 h. Subsequently End cells were incubated with 300 nM lapatinib or 20 mM LY294002 treated for 24 h and then for protein lysate preparation, or incubated with 300 nM lapatinib for the indicated times before they lysed.
Grb7 was determined c tubulin, phosphorylated Akt, Akt, and the overall level of immunoblot. D, SKBR3 cells were cultured in 1.56103 bo covered Their 10 cm, has respect, and with 300 nM lapatinib for 36 hours. Subsequently End the cells were resuspended in lysis buffer and HER2 was immunpr Zipitiert using a specific antique Rpers fight against HER2. The immunpr Zipitierten proteins Was separated by SDS-PAGE, transferred to a PVDF membrane and Grb7 and HER2 were rpern using appropriate antibody. A are the results as mean values 6 SD of three separate experiments. BD, a repr Sentative experiment of three is shown. doi: 10.1371/journal.pone.0009024.g002 GRB7 level of HER2 PLoS ONE regulated | Published in PloSOne fourth February 2010 | Volume 5 | Issue 2 | e9024 Thus, these changes parallel Ver and probably represent the G1 cell cycle arrest observed in response to lapatinib. In addition, downregulation of CCND1 in primary Rtumoren of patients treated with this drug, documented. Other genes that are regulated by lapatinib in were consistently