Abstract 2319: The role of CDKN1A and CDKN2B in the reprogramming of AML cells toward cell cycle arrest

Abstract

Abstract AML is the most common acute leukemia in adults, showing rapid progression if left untreated. With current treatments, AML patients frequently achieve remission, but often experience relapse leading to the overall poor prognosis and high mortality of this cancer. We have employed human AML cell lines to examine the response of leukemia cells to phorbol esters. Phorbol esters mimic the action of diacyl glycerol (DAG), an activator of protein kinase C, which regulates several signal transduction pathways and cellular metabolic activities. In response to the phorbol ester PMA, AML cells undergo cell cycle arrest, differentiate into “normal” macrophage-like cells, and ultimately undergo programmed cell death. The transcriptome variations that accompany these changes have been explored by DNA microarray analysis and quantitative real-time PCR, revealing ~1250 genes that are differentially expressed as the cells transition from the cancer state to non-dividing, myeloid cells that initiate apoptosis. Two genes of great interest regarding cell cycle arrest are the cyclin-dependent kinase inhibitors CDKN1A and CDKN2B. We have shown that both of these genes are expressed at extremely low levels in AML cells, but are rapidly up-regulated during the PMA response, in keeping with their proposed function as tumor suppressor genes. We present here results from the transfection of expression plasmids containing the CDKN1A or CDKN2B genes into HL-60 cells, showing ~100 and ~1,000-fold overexpression, respectively. In addition, we have engineered a plasmid containing both genes and demonstrate their co-overexpression. RNA samples from cells transfected with CDKN1A, CDKN2B, and CDKN1A-CDKN2B have been subjected to RNA sequencing. RNA-seq analysis reveals changes in CDKN1A and CDKN2B-mediated gene expression, altering the AML transcriptome in ways that facilitate cell cycle arrest. Finally, we present results on the phenotypic and transcriptome changes of a stable HL-60 cell line containing an inducible CDKN2B gene after dox-induced overexpression, implicating CDKN2B as an important regulator of cell cycle progression in AML cells. Citation Format: Michael Roberts, Grace Crossland, Jeffrey Forrester. The role of CDKN1A and CDKN2B in the reprogramming of AML cells toward cell cycle arrest [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2319.