ET 1 in combination with SDF 1 promotes 6 10B

The U metric is useful for comparing data on a normalized scale of zero to one and was used here to compare DDR readouts in AML samples and to assess the cell cycle selectivity of various drugs by comparing the proportion of responding buy 17-AAG cells within CyclinA2 or CyclinA2 populations for each agent. Popu lation frequency metrics for the percentage of healthy non apoptotic cell line or leukemic cells were calculated. For both cell lines and AML samples, per centages of CyclinA2 or CyclinA2 cells of the parent population were calculated. For both cell lines and AML samples DDR metrics were calculated for all healthy non apoptotic cells as well as CyclinA2 and CyclinA2 subsets where possible. Statistics Individual DDR readouts were correlated against each other using Pearsons linear correlation.<br><br> Data on drug induced cytotoxicity and DNA damage were correlated with proliferation levels using Pearsons linear correlation. Comparison of CyclinA2 vs CyclinA2 cells for magnitude and CV of signaling was performed with using a paired students t test. Students t test オーダー 17-DMAG was performed to compare BRCA1vs. BRCA1cell line responses. Results Development of SCNP assays to quanitfy induction of DNA damage and ATM mediated activation of DNA damage repair pathways To quantitatively assess induction of DNA damage and activation of DDR pathways at the single cell level, re agents and experimental conditions were first optimized to detect the induction of activation motifs within the two major double strand break repair pathwaysNHEJ and HRR.<br><br> Using the topoisomerase II inhibitor etoposide to induce DDR pathway activation and DNA damage, re agents recognizing the induced phosphorylation オーダー A66 sites on multiple proteins within the NHEJ and HRR pathways were examined. In a panel of cell lines con taining defined mutations including ATM. ATM, and ATMgenotypes, etoposide in duced phosphorylation and activation of all proteins tested. DDR response data for each ATM genotype and an illustration of the gating scheme used are shown in Figure 3. Consistent with the functional role of ATM in mediating signaling through both HRR and NHEJ path ways, etopside treatment induced lower activation of all readouts in the ATM cell lines compared to ATMand ATMcell lines and showed slightly lower activa tion of the ATMcell line vs. ATM cell lines for two readouts.<br><br> Of note, no significant differences were seen comparing the percen tages of live, healthy cells or CyclinA2 cells between ATM. ATMand ATMgenotypes, suggesting that DDR pathway measurements may be more informative than general measures of cell death cell cycle status. This supports the utility of the evaluated DDR readouts to quantify ATM mediated activa tion of DNA damage response pathways. Quantification of PARPi TMZ induced p H2AX levels in CyclinA2 cells robustly identifies alterations in HRR pathway activity The ability of SCNP to measure DDR responses in cellular subpopulations was next exploited as a method to quan tify the activity of cell cycle specific DDR pathways such as HRR.