Open Access Dissertation
Date of Award
Jeffrey W. Taub
Acute myeloid leukemia (AML) is a potentially devastating disease that can affect people of all ages. While there are certain patient groups that typically have favorable outcomes, in the pediatric population, overall survival (OS) is approximately 70%, leaving much progress to be made. Children with Down syndrome (DS) have a substantially elevated risk for developing AML, especially of the megakaryocytic phenotype (AMKL).
Patients with DS AMKL almost universally harbor a mutation in the gene encoding the GATA1 transcription factor, resulting in the translation of a short-form protein GATA1s, which has been shown to contribute to chemotherapy sensitivity. In contrast, in non-DS patients, overexpression of GATA1 has been correlated with poorer outcomes. In the first half of this work, we investigated the role that elevated GATA1 expression plays on the sensitivity of AMKL cells to chemotherapy. To begin, we confirmed that amongst non-DS patients, GATA1 transcripts were significantly higher in AMKL blasts compared to blasts from other AML subgroups. Further, GATA1 transcript levels significantly correlated with transcript levels for the anti-apoptotic protein Bcl-xL in our patient cohort. We demonstrated that Bcl-xL is a GATA1 target, and knockdown of GATA1 with shRNA sensitizes cells to chemotherapy treatment and decreases Bcl-xL expression. Treatment of Meg-01 cells with the histone deacetylase inhibitor valproic acid resulted in down-regulation of both GATA1 and Bcl-xL and significantly enhanced ara-C sensitivity. Furthermore, additional GATA1 target genes were identified by oligonucleotide microarray and ChIP-on-Chip analyses.
While OS for DS AML is generally very favorable (approximately 90%), DS patients with refractory or relapsed disease have dismal prognoses, with OS in that group only approximately 25%. Therefore, there is a need for the development of new therapeutic approaches for these patients. In the second half of this work, we investigated the response of 2 DS AML cell lines, CMK and CMY, to the combination of araC and one of either the aurora A kinase inhibitor MLN8237, the aurora B kinase inhibitor AZD1152-HQPA, the Plk1 inhibitor BI6237, or the wee1 inhibitor MK-1775. It was found that MK-1775, in contrast to the other three agents, synergized with araC in antiproliferative MTT assays in both cell lines as well as in ex vivo DS-AML primary patient samples. MK-1775 was able to decrease inhibitory CDK1(Y15) phosphorylation after only 4 hours, and was able to enhance araC-induced DNA damage in S-phase and partially abrogate araC-induced cell cycle arrest.
The work presented in this dissertation describes preclinical efforts at improving outcomes for pediatric AML patients in both the DS and the non-DS populations. The findings presented demonstrate a potential for using novel therapies, like HDAC or Bcl-2 family inhibitors to treat AMKL, as well as offering insight into the mechanisms of resistance in a difficult to treat disease. Furthermore, these findings lay the foundation for the use of MK-1775 to enhance the effects of araC in DS-AML. Though there is always more to be elucidated, the studies described herein set the stage for such further work, with the eventual goal of improving outcomes for pediatric AML.
Caldwell, John Timothy, "Identifying Mechanisms Of Resistance And Potential Therapeutic Targets For Pediatric Acute Myeloid Leukemia" (2016). Wayne State University Dissertations. 962.