The transcription factor C/EBPa is necessary and sufficient for neutrophil differentiation. Expression and/or function of C/EBPalpha were found to be perturbed by various mechanisms in many cases of acute myeloid leukemia (AML). Consequently, conditional expression of C/EBPa in leukemic cells re-established their neutrophilic differentiation. This proposal offers novel C/EBPalpha-mediated transcriptional therapies for AML by two approaches. In one, noninvasive delivery of functional C/EBPa protein into nuclei of diseased cells will be utilized. Cell permeable peptides will be fused in frame with C/EBPa protein and expressed in eukaryotic cells. Intracellular protein transport will be achieved through co-culture of the producer cell line with the recipient cells, or by supplying the purified cell permeable C/EBPa proteins into culture media of the recipient cells. Optimized procedure will be applied to primary leukemic bone marrow cells. The expected effects of C/EBPa protein delivery, such as induction of differentiation and apoptosis, will be monitored. In the second approach, a rapid and reproducible cell-based high throughput screen of small molecule chemicals was developed. Briefly, a stable indicator line was made, which harbors luciferase gene under the control of C/EBP-responsive element. Increase in luciferase activity will likely result from augmentation of C/EBPa expression and/or function. Active compounds will be identified and studied to determine the mechanisms of their action. Although this proposal is focused on development of new therapies for AML, successful C/EBPa targeting therapies might be also used for treatment of CML refractory to interferon` or ST1571. The applicant has considerable skills necessary to perform these experiments. Nevertheless, completion of the proposed career development program will further expand her research experience and will enable her to pursue an independent research career.
