Evaluation of the G-quadruplex Binding Drug Telomestatin as an Inhibitor of c-myb in Chronic Myelogenous Leukemia.

摘要

Concept: We propose to investigate MYB as a potential molecular target of telomestatin in CML, and we will seek to answer two major questions about telomestatin in CML. How does telomestatin prevent MYB expression in CML, and is telomestatin active against CML in vivo. We hypothesize that the antileukemia activity of telomestatin in CML is in part due to inhibition of MYB expression by the binding of telomestatin to a G-quadruplex in the MYB promoter. Aims: The specific objectives of this proposal are: 1) to evaluate the ability and mechanism of telomestatin to selectively suppress c-MYB expression in K562 and K562R CML cells in culture; and 2) to investigate the anti-leukemia activity of telomestatin in murine K562 and K562R xenografts. In aim 1, we will treat imatinib sensitive and resistant CML cells with telomestatin alone or in combination with imatinib and measure the expression of C-MYB, housekeeping control genes, a panel of other genes that we have identified as containing potential G-quadruplex forming units in their promoters, and telomerase (hTERT) expression. We will show that telomestatin can prevent transcription factor binding to the MYB promoter in solution by EMSAs and footprinting, and in CML cells by ChIP assays. In aim 2, we will form K562 and K562R xenografts in immunodeficient (scid) mice, administer telomestatin to these mice by tail vein injection, and measure the activity of telomestatin on the growth of these CML xenografts. To determine the in vivo mechanism of action of telomestatin, tumors will be harvested and analyzed by standard histopathology, immunohistochemistry for MYB, and analysis of telomerase activity.