ng cells may account for the apparent disconnect between apoptosis seen in vitro versus the clinical NR. In order to understand whether there was a link between signaling by survival pathways and in vitro apoptotic responses, correlations were computed from the data depicted in August 2010 | Volume 5 | Issue 8 | e12405 Pathway Profiles in AML 11 August 2010 | Volume 5 | Issue 8 | e12405 Pathway Profiles in AML apoptosis profiles. Given the opportunity to apply SCNP assays to samples taken over time from the same patient it may be possible to determine which blast population confers refractoriness to chemotherapy. Further correlations to defined genetic abnormalities driving these signaling observations could underscore their potential roles in driving AML disease; such as analysis of intracellular signaling pathways in the context of FLT3 mutational status. The output from such studies could be to guide the choice of available investigational and approved agents to provide benefit for those AML patients refractory to current chemotherapy regimens. cytogenetics, FLT3-ITD positive or secondary AML. Criteria for sample inclusion in this study: diagnostic prior to initiation 18325633 of chemotherapy, AML classification by French American British criteria as M0 through M7 and data for clinical response to induction therapy. Found at: doi:10.1371/journal.pone.0012405.s003 Supporting Information MedChemExpress 193022-04-7 Definition of Metrics A) Summary of metrics used and the role each 12600694 has in determining a different measure of signaling biology. Median Fluorescence Intensities were calculated for leukemic blast cells under each condition and used to compute the metrics that represented protein expression, signaling and apoptosis, as described in Materials and Methods. Apoptosis was measured by levels of c-caspase 3, c-PARP as well as cells that were positive for the aqua viability dye. Measurements for cleaved caspase 3 were found to be correlated with cleaved PARP, but this was not the case for cleaved PARP and aqua. Therefore apoptosis metrics incorporated measurements for aqua and cleaved PARP to quantify cells that became positive for cPARP alone or for both aqua and c-PARP. Found at: doi:10.1371/journal.pone.0012405.s002 Acknowledgments The authors would like to thank David R. Parkinson for critical reading of the manuscript. Author Contributions Conceived and designed the experiments: DBR TC YWH GPN WF. Performed the experiments: DBR TC YWH. Analyzed the data: DBR SP GPN AC WF. Contributed reagents/materials/analysis tools: MM. Wrote the paper: DBR SP GPN AC MM WF. 25 NRs represent primary refractory AML. The ��other��values for race are black and Hispanic subgroups. Poor prognosis samples have one or more high risk features: age $60 years, unfavorable 12 August 2010 | Volume 5 | Issue 8 | e12405 Pathway Profiles in AML 20. Ravandi F, Burnett AK, Agura ED, Kantarjian HM Progress in the treatment of acute myeloid leukemia. Cancer 110: 1900910. 21. Tallman MS, Gilliland DG, Rowe JM Drug therapy for acute myeloid leukemia. Blood 106: 1154163. 22. Coffer PJ, Koenderman L, de Groot RP The role of STATs in myeloid differentiation and leukemia. Oncogene 19: 2511522. 23. Bromberg J, Darnell JE, Jr. The role 
of STATs in transcriptional control and their impact on cellular function. Oncogene 19: 2468473. 24. Yuan TL, Cantley LC PI3K pathway alterations in cancer: variations on a theme. Oncogene 27: 5497510. 25. Vogt PK, Gymnopoulos M, Hart JR PI 3-kinase and cancer: changing accents
