Combined Analysis of Valproic Acid Induced MicroRNA and Gene Expression Changes in Acute Myeloid Leukemia.
Authors: Russ Annika C. , Dugas Martin , Schmid-Fetzer Sonja C. , Schlenk Richard F. , Pollack Jonathan R. , Dohner Konstanze , Schmid Mathias , Dohner Hartmut , Bullinger Lars 
Published: October 2019

Abstract
Inhibitors of histone deacetylases (HDACs) like valproic acid (VPA) display activity in murine leukemia models and induce apoptosis and myeloid differentiation of acute myeloid leukemia (AML) blasts. While recently several studies examined the underlying VPA-mediated mechanisms, until today not many genes have been identified whose expression is altered by VPA treatment. Recently, microRNAs (miRs), a novel abundant class of negative gene regulators, have been shown to control a wide range of biological functions such as proliferation, differentiation and apoptosis by either translational repression or by mRNA cleavage or miR-mediated decay of the respective target mRNA. Furthermore, deregulated miR expression has been associated with various human cancers including leukemia. This led us to investigate whether VPA treatment of AML cells affects miR-expression which in turn might influence the level of miR target genes involved in VPA effects. First, we identified an in vitro miR VPA-response signature by profiling miR expression in 4 different myeloid leukemia cell lines following 48 hours of VPA treatment (Ambion microarray platform comprising 281 human miRs). In parallel, we profiled gene expression by using both cDNA microarrays and Affymetrix U133Aplus2.0 GeneChips. 13 miRs were found to be differentially expressed, 10 miRs were down-regulated and 3 miRs were up-regulated by VPA. Gene expression profiling revealed several hundred differentially regulated genes containing some known VPA influenced targets like e.g. cyclin-dependent kinase inhibitor CDKN1A coding for p21. To correlate miR and gene expression, we next searched for an enrichment of putative miR target genes of the VPA-regulated miRs in the VPA-induced gene expression pattern. Interestingly, there were several candidates for which miR expression in response to VPA inversely correlated with gene expression of the respective targets. These included genes involved in DNA damage checkpoint like e.g. CHEK1 which was found to be down-regulated in response to VPA and which is a predicted target of miR-15a and miR-16, both found to be up-regulated by VPA treatment. In addition, potential miR-targets included genes known to be regulated by HDAC inhibitors in cancer cells like e.g. the homeobox gene HOXA1 found to be up-regulated in response to VPA and being a putative target of miR-99a, found to be down-regulated by VPA. Our study is the first to show that VPA treatment significantly affects expression levels of several miRs in myeloid cell lines, and based on the correlation of VPA-induced miR and gene expression patterns we could identify putative miR-targets that included genes with tumorigenic relevance. While it remains to be determined whether VPA-induced miR-mediated mRNA cleavage or decay of the respective target mRNAs is involved in leukemogenesis, our data nevertheless provide new insights into VPA-induced mechanisms of myeloid differentiation and into deregulated miR expression in leukemia.