Dissection of signaling pathways in fourteen breast cancer cell lines using reverse-phase protein lysate microarray.

Signal transduction pathways play a crucial role in breast cancer development, progression, and response to different therapies. A major problem in breast cancer therapy is the heterogeneity among different tumor types and cell lines commonly used in preclinical studies. To characterize the signaling pathways of some of the commonly used breast cancer cell lines and dissect the relationship among a number of pathways and some key genetic and molecular events in breast cancer development, such as p53 mutation, ErbB2 expression, and estrogen receptor (ER)/progesterone receptor (PR) status, we performed pathway profiling of 14 breast cancer cell lines by measuring the expression and phosphorylation status of 40 different cell signaling proteins with 53 specific antibodies using a protein lysate array. Cluster analysis of the expression data showed that there was close clustering of phosphatidylinositol 3-kinase, Akt, mammalian target of rapamycin (mTOR), Src, and platelet-derived growth factor receptor beta (PDGFRbeta) in all of the cell lines. The most differentially expressed proteins between ER- and PR-positive and ER- and PR-negative breast cells were mTOR, Akt (pThr308), PDGFRbeta, PDGFRbeta (pTyr751), panSrc, Akt (pSer473), insulin-like growth factor-binding protein 5 (IGFBP5), Src (pTyr418), mTOR (pSer2448), and IGFBP2. Many apoptotic proteins, such as apoptosis-inducing factor, IGFBP3, bad, bax, and cleaved caspase 9, were overexpressed in mutant p53-carrying breast cancer cells. Hexokinase isoenzyme 1, ND2, and c-kit were the most differentially expressed proteins in high and low ErbB2-expressing breast cancer cells. This study demonstrated that ER/PR status, ErbB2 expression, and p53 status are major molecules that impact downstream signaling pathways.

Effects of radiation on frozen lactate dehydrogenase.

Results concerning the influence of 6-MeV electron beam irradiation, of 2.45-GHz, 565-W microwaves, and of the combined electron and microwave irradiation, at -21 degrees C and -196 degrees C, on lactate dehydrogenase activity are presented. The microwave-irradiated samples exhibited a non-linear behaviour (successive activation and inactivation of the enzyme molecules), suggesting the major influence of the non-thermal component of microwave radiation. The combined electron and microwave irradiation led to a decrease of activity similar to the one caused by electron beam irradiation, which seemed to prove that microwave influence was insignificant in the dose, power and time ranges used. The radiation target analysis of the enzymatic decrease due to electron irradiation indicated a very large aggregation of the enzyme molecules. Our data suggest that radiation target analysis is not suitable to measure the molecular mass of lactate dehydrogenase, when frozen enzyme suspensions are irradiated. The D2O-protected enzyme, when exposed to electron irradiation, showed an even larger aggregation according to radiation target analysis, while the microwave irradiation of the protected enzyme led to a similar, though lesser, non-linear behaviour of the frozen enzyme molecules.