Discovery of agents that eradicate leukemia stem cells using an in silico screen of public gene expression data

Abstract

Increasing evidence indicates that malignant stem cells are important for the pathogenesis of acute myelogenous leukemia (AML) and represent a reservoir of cells that drive the development of AML and relapse. Therefore, new treatment regimens are necessary to prevent relapse and improve therapeutic outcomes. Previous studies have shown that the sesquiterpene lactone, parthenolide (PTL), ablates bulk, progenitor, and stem AML cells while causing no appreciable toxicity to normal hematopoietic cells.Thus, PTL must evoke cellular responses capable of mediating AML selective cell death. Given recent advances in chemical genomics such as gene expression-based high-throughput screening (GE-HTS) and the Connectivity Map, we hypothesized that the gene expression signature resulting from treatment of primary AML with PTL could be used to search for similar signatures in publicly available gene expression profiles deposited into the Gene Expression Omnibus (GEO). We therefore devised a broad in silico screen of the GEO database using the PTL gene expression signature as a template and discovered 2 new agents, celastrol and 4-hydroxy-2-nonenal, that effectively eradicate AML at the bulk, progenitor, and stem cell level. These findings suggest the use of multicenter collections of highthroughput data to facilitate discovery of leukemia drugs and drug targets. IntroductionCancer stem cells (CSCs) have recently emerged as a potentially important consideration for studies of basic tumor biology and the development of improved therapies. Like normal stem cells, CSCs are thought to reside at the apex of a developmental hierarchy and are responsible for the continued growth and expansion of bulk tumor populations. 1 Consequently, the biological activity of CSCs may contribute to initiation, maintenance, and relapse of at least some forms of cancer. To date, CSCs are best characterized for the blood cancer, acute myelogenous leukemia (AML), where numerous studies have documented the phenotype, cell cycle status, and growth characteristics of malignant stem and progenitor cell types. [2][3][4] Notably, several studies have shown that AML stem cells (AML-SCs) are refractory to commonly used clinical agents such as cytarabine and anthracyclines, 5-8 thereby further supporting the hypothesis that malignant stem cells represent a probable reservoir from which disease relapse may occur. Given the central role of AML-SCs in leukemic disease, it is therefore important to identify therapeutic regimens that are capable of eradicating this subpopulation of malignant cells.To date, preclinical studies have demonstrated that selective ablation of AML-SCs using small molecule-based strategies is possible. For example, in vitro studies demonstrated that the combination of idarubicin and MG-132 (IDR/MG) can effectively eradicate leukemia stem cells via a mechanism involving concomitant inhibition of nuclear factor-B (NF-B)-mediated survival signals and induction of oxidative stress. 9 Subsequently, it was shown that parthenolide (PTL), 7 the bio...

A conceptual framework for the identification of candidate drugs and drug targets in acute promyelocytic leukemia

“…To test this strategy, we focused our efforts on the TF nuclear factor (NF)-kB, in part, because our promoter analysis revealed a significant enrichment of binding sites for NF-kB among genes dysregulated in APL, and in part, because NF-kB can be inhibited effectively by terpenoids such as Parthenolide. 32,38 Significantly, incubation of NB4 cells for 48 h with micromolar concentrations of Parthenolide resulted in a substantial inhibition of proliferation and induction of apoptosis ( Figure 5). The latter suggests that comprehensive promoter analysis of dysregulated genes in APL can identify TFs, which at least to some extent, might represent potential targets for therapeutic interventions.…”

“…25,26 In contrast, genes upregulated in APL cells versus PMs contained a significant enrichment of binding sites for TFs that are instrumental for malignant transformation in cell lines and AML (MZF1, ARNT), and most strikingly for proliferation, mobilization and self-renewal of HSCs and/or leukemic stem cells (LSCs) (ERG1, NFKB, NFKB1). [27][28][29][30][31][32] APL cells show a partial stemness signature Our promoter analysis revealed that genes upregulated in APL cells versus PMs were significantly enriched in binding sites for TFs that conduct HSC maintenance/self-renewal. This led us to the hypothesis that APL cells might differ form PMs by expression of genes associated with HSC maintenance/selfrenewal.…”

Gene expression signature-based approach identifies a pro-resolving mechanism of action for histone deacetylase inhibitors

“…This database can be queried with any gene-expression profile of interest (e.g., the expression profile of a compound, or a disease) and the results yield a list of the CMap drugs ranked, according to the similarity with the signature of interest. Hence, drugs with negative score (opposite profiles) might have the potential as new treatments for specific diseases, [12][13][14] while drugs with positive score (similar profiles) could be useful for identification of novel actions of existing drugs 15 or to unravel drug mechanisms of action. [16][17][18] Using the gene expression profile produced by the antiinflammatory pro-resolving molecule annexin A1 (AnxA1), 19 we found very strong positive connections with several members of the histone deacetylase inhibitors (HDACIs) class of drugs.…”

Towards novel paradigms for cancer therapy

“…It is important to emphasize that natural compounds have also been reported to target TICs selectively, with parthenolide (PTL) being the first such compound described (Guzman et al, 2005(Guzman et al, , 2007Steele et al, 2006;Hassane et al, 2008;Kawasaki et al, 2009). PTL is a sesquiterpene lactone found in feverfew (Chrysanthemum parthenium), which has been traditionally used to treat migraine and rheumatoid arthritis and exerts activity as nuclear factor-kB inhibitor, p53 activator (via ubiquitination of MDM2, mouse double minute 2-p53 binding protein homolog), DNMT1 (DNA (cytosine-5-)-methyltransferase 1) inhibitor and promoter of HDAC1 depletion by proteosome degradation (Gopal et al, 2007).…”

“…PTL is a sesquiterpene lactone found in feverfew (Chrysanthemum parthenium), which has been traditionally used to treat migraine and rheumatoid arthritis and exerts activity as nuclear factor-kB inhibitor, p53 activator (via ubiquitination of MDM2, mouse double minute 2-p53 binding protein homolog), DNMT1 (DNA (cytosine-5-)-methyltransferase 1) inhibitor and promoter of HDAC1 depletion by proteosome degradation (Gopal et al, 2007). Using the global transcription response to PTL, a collection of compounds that mimic the effect of PTL were discovered and shown to target acute myeloid leukemia stem cells (Hassane et al, 2008). Two of these, celastrol (a terpenoid) and 4-hydroxynonenal (a nonterpenoid lipid peroxidation product) are also natural products, whereas one additional hit, 15-deoxy-D12,14-PGJ2, is a prostaglandin derivative.…”