sgi-1027 and Neoplasms

DNA methylation is a mammalian epigenetic mark that is involved in defining where and when genes are expressed, both in normal cells and in the context of diseases. Like other epigenetic marks, it is reversible and can be modulated by chemical agents. Because it plays an important role in cancer by silencing certain genes, such as tumor suppressor genes, and by reactivating other regions, such as repeated elements, it is a promising therapeutic target. Two compounds are already approved to treat hematological cancers. Many efforts have been carried out to discover new molecules that are able to efficiently inhibit DNA methylation in cancer cells. We will briefly overview the foremost of these efforts by focusing on what we have learned to this point on non-nucleoside inhibitors and on what we consider to be the features of an ideal inhibitor.

Topics: Animals; Antineoplastic Agents; DNA Methylation; DNA Modification Methylases; Drug Discovery; Epigenesis, Genetic; Gene Expression Regulation, Neoplastic; Humans; Models, Molecular; Molecular Targeted Therapy; Neoplasms; Nucleosides

The transcription master regulator MYC plays an essential role in regulating major cellular programs and is a well-established therapeutic target in cancer. However, MYC targeting for drug discovery is challenging. New therapeutic approaches to control MYC-dependent malignancy are urgently needed. The mitogen-activated protein kinase kinase 3 (MKK3) binds and activates MYC in different cell types, and disruption of MKK3-MYC protein-protein interaction may provide a new strategy to target MYC-driven programs. However, there is no perturbagen available to interrogate and control this signaling arm. In this study, we assessed the drugability of the MKK3-MYC complex and discovered the first chemical tool to regulate MKK3-mediated MYC activation. We have designed a short 44-residue inhibitory peptide and developed a cell lysate-based time-resolved fluorescence resonance energy transfer (TR-FRET) assay to discover the first small molecule MKK3-MYC PPI inhibitor. We have optimized and miniaturized the assay into an ultra-high-throughput screening (uHTS) 1536-well plate format. The pilot screen of ~6,000 compounds of a bioactive chemical library followed by multiple secondary and orthogonal assays revealed a quinoline derivative SGI-1027 as a potent inhibitor of MKK3-MYC PPI. We have shown that SGI-1027 disrupts the MKK3-MYC complex in cells and in vitro and inhibits MYC transcriptional activity in colon and breast cancer cells. In contrast, SGI-1027 does not inhibit MKK3 kinase activity and does not interfere with well-known MKK3-p38 and MYC-MAX complexes. Together, our studies demonstrate the drugability of MKK3-MYC PPI, provide the first chemical tool to interrogate its biological functions, and establish a new uHTS assay to enable future discovery of potent and selective inhibitors to regulate this oncogenic complex.

Topics: Antineoplastic Agents; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Drug Discovery; Humans; MAP Kinase Kinase 3; Molecular Docking Simulation; Molecular Structure; Neoplasms; Protein Binding; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-myc; Small Molecule Libraries; Structure-Activity Relationship

DNA methyltransferases (DNMTs) are epigenetic enzymes involved in embryonic development, cell differentiation, epithelial to mesenchymal transition, and control of gene expression, whose overexpression or enhanced catalytic activity has been widely reported in cancer initiation and progression. To date, two DNMT inhibitors (DNMTi), 5-azacytidine (5-AZA) and 5-aza-2'-deoxycytidine (DAC), are approved for the treatment of myelodysplastic syndromes and acute myeloid leukemia. Nevertheless, they are chemically instable and quite toxic for healthy cells; thus, the discovery of novel DNMTi is urgent.. Here, we report the identification of a new quinoline-based molecule, MC3353, as a non-nucleoside inhibitor and downregulator of DNMT. This compound was able, in promoter demethylating assays, to induce enhanced green fluorescence protein (EGFP) gene expression in HCT116 cells and transcription in a cytomegalovirus (CMV) promoter-driven luciferase reporter system in KG-1 cells. Moreover, MC3353 displayed a strong antiproliferative activity when tested on HCT116 colon cancer cells after 48 h of treatment at 0.5 μM. At higher doses, this compound provided a cytotoxic effect in double DNMT knockout HCT116 cells. MC3353 was also screened on a different panel of cancer cells (KG-1 and U-937 acute myeloid leukemia, RAJI Burkitt's lymphoma, PC-3 prostate cancer, and MDA-MB-231 breast cancer), where it arrested cell proliferation and reduced viability after 48 h of treatment with IC. The present work describes MC3353 as a novel DNMTi displaying a stronger in cell demethylating ability than both 5-AZA and DAC, providing re-activation of the silenced ubiquitin C-terminal hydrolase L1 (UCHL1) gene. MC3353 displayed dose- and time-dependent antiproliferative activity in several cancer cell types, inducing cell death and affecting EMT through E-cadherin and MMP2 modulation. In addition, this compound proved efficacy even in primary osteosarcoma cell models, through the modulation of genes involved in osteoblast differentiation.

Topics: Aminoquinolines; Cell Line, Tumor; Cell Proliferation; Cell Survival; DNA Methylation; DNA-Cytosine Methylases; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Enzyme Inhibitors; Epigenesis, Genetic; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; HCT116 Cells; Humans; Neoplasms; Pyrimidines

DNA methyltransferases (DNMTs) and histone deacetylases (HDACs) are important epigenetic targets during anticancer drug development. Recent study indicates that DNMT inhibitors and HDAC inhibitors display synergistic effects in certain cancers, therefore, development of molecules targeting both DNMT and HDAC is of therapeutic advantage against these cancers. Based on the structure of DNMT inhibitor NSC-319745 and the pharmacophore characteristics of HDAC inhibitors, a series of hydroxamic acid derivatives of NSC-319745 were designed and synthesized as DNMT and HDAC multifunctional inhibitors. Most compounds displayed potential DNMT inhibitory potency and potent HDAC inhibitory activity, especially compound 15a showed much better DNMT1 inhibitory potency than NSC-319745, and inhibited HDAC1, HDAC6 with IC

Topics: Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; DNA (Cytosine-5-)-Methyltransferases; DNA Methylation; Drug Design; Drug Screening Assays, Antitumor; Gene Expression Regulation, Neoplastic; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Molecular Docking Simulation; Neoplasms

Aberrant DNA hypermethylation of promoter of tumor suppressor genes is commonly observed in cancer, and its inhibition by small molecules is promising for their reactivation. Here we designed bisubstrate analogues-based inhibitors, by mimicking each substrate, the S-adenosyl-l-methionine and the deoxycytidine, and linking them together. This approach resulted in quinazoline-quinoline derivatives as potent inhibitors of DNMT3A and DNMT1, some showing certain isoform selectivity. We highlighted the importance of (i) the nature and rigidity of the linker between the two moieties for inhibition, as (ii) the presence of the nitrogen on the quinoline group, and (iii) of a hydrophobic group on the quinazoline. The most potent inhibitors induced demethylation of CDKN2A promoter in colon carcinoma HCT116 cells and its reactivation after 7 days of treatment. Furthermore, in a leukemia cell model system, we found a correlation between demethylation of the promoter induced by the treatment, chromatin opening at the promoter, and the reactivation of a reporter gene.

Topics: Cell Line, Tumor; DNA (Cytosine-5-)-Methyltransferase 1; DNA (Cytosine-5-)-Methyltransferases; DNA Methylation; DNA Methyltransferase 3A; Drug Design; Enzyme Inhibitors; Genes, Tumor Suppressor; Humans; Neoplasms; Substrate Specificity

Among the epigenetic marks, DNA methylation is one of the most studied. It is highly deregulated in numerous diseases, including cancer. Indeed, it has been shown that hypermethylation of tumor suppressor genes promoters is a common feature of cancer cells. Because DNA methylation is reversible, the DNA methyltransferases (DNMTs), responsible for this epigenetic mark, are considered promising therapeutic targets. Several molecules have been identified as DNMT inhibitors and, among the non-nucleoside inhibitors, 4-aminoquinoline-based inhibitors, such as SGI-1027 and its analogs, showed potent inhibitory activity. Here we characterized the in vitro mechanism of action of SGI-1027 and two analogs. Enzymatic competition studies with the DNA substrate and the methyl donor cofactor, S-adenosyl-l-methionine (AdoMet), displayed AdoMet non-competitive and DNA competitive behavior. In addition, deviations from the Michaelis-Menten model in DNA competition experiments suggested an interaction with DNA. Thus their ability to interact with DNA was established; although SGI-1027 was a weak DNA ligand, analog 5, the most potent inhibitor, strongly interacted with DNA. Finally, as 5 interacted with DNMT only when the DNA duplex was present, we hypothesize that this class of chemical compounds inhibit DNMTs by interacting with the DNA substrate.

Topics: Aminoquinolines; DNA; DNA (Cytosine-5-)-Methyltransferases; DNA Methylation; Enzyme Inhibitors; Epigenomics; Humans; Neoplasms; Pyrimidines