Johnny Easmon

A small molecule modulates Jumonji histone demethylase activity and selectively inhibits cancer growth

The pharmacological inhibition of general transcriptional regulators has the potential to block growth through targeting multiple tumorigenic signaling pathways simultaneously. Here, using an innovative cell-based screen, we identify a structurally unique small molecule (named JIB-04) which specifically inhibits the activity of the Jumonji family of histone demethylases in vitro, in cancer cells, and in tumors in vivo. Unlike known inhibitors, JIB-04 is not a competitive inhibitor of α-ketoglutarate. In cancer but not in patient-matched normal cells, JIB-04 alters a subset of transcriptional pathways and blocks viability. In mice, JIB-04 reduces tumor burden and prolongs survival. Importantly, we find that patients with breast tumors that overexpress Jumonji demethylases have significantly lower survival. Thus JIB-04, a novel inhibitor of Jumonji demethylases in vitro and in vivo, constitutes a unique potential therapeutic and research tool against cancer, and validates the use of unbiased cellular screens to discover chemical modulators with disease relevance.

2-benzoxazolyl and 2-benzimidazolyl hydrazones derived from 2-acetylpyridine: A novel class of antitumor agents

Here we describe the effects of novel benzoxazol‐2‐yl and benzimidazol‐2‐yl hydrazones derived from 2‐pyridinecarbaldehyde and 2‐acetylpyridine. The IC50 values for inhibition of cell proliferation in KB‐3‐1, CCRF‐CEM, Burkitts lymphoma, HT‐29, HeLa, ZR‐75 and MEXF276L by most of the novel compounds are in the nanomolar range. In colony‐forming assays with human tumor xenografts the compounds 2‐actylpyridine benzoxazol‐2‐ylhydrazone (EPH52), 2‐acetylpyridine benzoimidazol‐2‐ylhydrazone (EPH61) and 2‐acetylpyridine 1‐methylbenzoimidazol‐2‐ylhydrazone (EPH116) exhibited above‐average inhibition of colon carcinoma (IC50 = 1.3–4.56 nM); EPH52 and EPH116 also exhibited above‐average inhibition of melanoma cells. As shown with human liver microsomes, EPH116 is only moderately metabolized. The compound inhibited the growth of human colon cancer xenografts in nude mice in a dose‐dependent manner. Thiosemicarbazones derived from 2‐formylpyridines have been shown to be inhibitors of ribonucleotide reductase (RR). The following results show that RR is not the target of the novel compounds: cells overexpressing the M2 subunit of RR and resistant to the RR inhibitor hydroxyurea are not cross‐resistant to the novel compounds; inhibition of RR occurs at 6‐ to 73‐fold higher drug concentrations than that of inhibition of cell proliferation; the pattern of cell cycle arrest in S phase induced by the RR inhibitor hydroxyurea is not observed after treatment with the novel compounds; and a COMPARE analysis with the related compounds 2‐acetylpyrazine benzothiazol‐2‐ylhydrazone (EPH95) and 3‐acetylisoquinoline benzoxazol‐2‐ylhydrazone (EPH136) showed that the pattern of these compounds is not related to any of the standard antitumor drugs. Therefore, these novel compounds show inhibition of colon cancers and exhibit a novel mechanism of action.

Investigations on the Mechanism of Action of the Novel Antitumor Agents 2‐Benzothiazolyl, 2‐Benzoxazolyl, and 2‐Benzimidazolyl Hydrazones Derived from 2‐Acetylpyridine

2‐Acetylpyridine hydrazone derivatives of benzothiazole, benzoxazole, and benzimidazole were found to exhibit potent cytotoxic activity against the growth of suspended leukemia and lymphomas. They were also active in a number of solid tumor screens, e.g. HeLa uterine carcinoma, SOS bone osteosarcoma, lung MB9812, lung A549, Mcf‐7 breast growth. In L1210 lymphoid leukemia cells the compounds preferentially inhibited RNA synthesis followed by DNA synthesis at 100 μM after 60 min. The reduction of de novo purine synthesis by the compounds at the regulatory sites PRPP‐amido transferase, IMP dehydrogenase and dihydrofolate reductase was responsible for the suppression of nucleic synthesis. Other minor sites where the agents have metabolic effects were thymidylate synthetase and thymidine kinase which would be additive with the overall inhibition of cell growth. The ct‐DNA studies suggest that the compounds also interacted with the DNA molecule itself, probably affecting template activity.

Comparative In Vitro Anti-Hepatitis C Virus Activities of a Selected Series of Polymerase, Protease, and Helicase Inhibitors

ABSTRACT We report here a comparative study of the anti-hepatitis C virus (HCV) activities of selected (i) nucleoside polymerase, (ii) nonnucleoside polymerase, (iii) α,γ-diketo acid polymerase, (iv) NS3 protease, and (v) helicase inhibitors, as well as (vi) cyclophilin binding molecules and (vii) alpha 2b interferon in four different HCV genotype 1b replicon systems.

Thiazolyl and benzothiazolyl hydrazones derived from α-(N)-acetylpyridines and diazines: synthesis, antiproliferative activity and CoMFA studies

Summary The synthesis of a series of thiazolyl and benzothiazolyl hydrazones derived from α-( N )-acylpyridines, -quinolines, -isoquinolines, -pyridazines, -pyrimidines, and -pyrazines is reported. The stereochemistry of these compounds was determined by NMR spectroscopic methods. The antiproliferative activity of the novel compounds was quantified in tissue culture (melanoma, breast carcinoma, colon adenocarcinoma, epitheloid cervix carcinoma, Burkitts lymphoma, leukemia, and hydroxyurea sensitive and resistant myelogenous leukemia sublines). All compounds exhibited profound antiproliferative activity, in particular against Burkitts lymphoma cells. Out of this series, compounds 6b, 7b, 7c, 8c and 8i were found to be 13–900 times more potent than hydroxyurea and no cross-resistance to hydroxyurea was observed. A predictive 3D-QSAR model using the CoMFA approach was established.

Pyridazines. IIIL: The configuration of novel thiosemicarbazone derivatives of pyridazinecarbaldehydes and alkyl pyridazinyl ketones

Determination de la structure et de la configuration des derives de thiosemicarbazones contenant un groupement pyridazinyl-3, pyridazinyl-4 ou pyridyl-2 par spectrometrie RMN 1 H et 13 C

N-benzoxazol-2-yl-N′-1-(isoquinolin-3-yl-ethylidene)-hydrazine, a novel compound with antitumor activity, induces radicals and dissipation of mitochondrial membrane potential

SummaryThe novel compound N-benzoxazol-2-yl-N′-1-(isoquinolin-3-yl-ethylidene)-hydrazine (EPH136) has been shown to exhibit antitumor activity in vitro and in vivo. A COMPARE analysis showed that the patterns of cellular effects of EPH136 are not related to any of 175 standard antitumor agents with a known mechanism of action. In order to help identify the mechanism of action we employed a bioinformatics approach called partial least squares modelling in latent variables in which the expression levels of ~8,000 genes in each of 56 untreated NCI panel cell lines were correlated with the respective IC50 values of each cell line following treatment with EPH136. The 60 genes found to be most important for the antiproliferative effect of EPH136 are involved in nucleoside, nucleotide, nucleic acid binding and metabolism, developmental processes, protein modification and metabolism. In addition, using a DNA microarray we measured the expression of ~5,000 known genes following treatment of HT-29 colon carcinoma cells with a two-fold IC50 concentration of EPH136. The genes that were up-regulated more than two-fold compared to untreated controls belong to the same classes as found by the bioinformatic approach. Many of these proteins are regulated by oxidation/reduction and so we concluded that formation of radicals may be involved in the mechanism of action. We show here that EPH136 leads to generation of oxygen radicals, swelling of mitochondria and dissipation of the mitochondrial membrane potential. The antiproliferative activity of EPH136 was prevented by the radical scavenger N-acetylcysteine. Cells with elevated glutathione exhibited resistance to EPH136. In summary, the mechanism of the novel experimental anticancer drug EPH136 is generation of radicals and dissipation of the mitochondrial membrane potential.

2-Benzazolyl-4-Piperazin-1-Ylsulfonylbenzenecarbohydroxamic Acids as Novel Selective Histone Deacetylase-6 Inhibitors with Antiproliferative Activity.

We have screened our compound collection in an established cell based assay that measures the derepression of an epigenetically silenced transgene, the locus derepression assay. The screen led to the identification of 4-[4-(1-methylbenzimidazol-2-yl)piperazin-1-yl]sulfonylbenzenecarbohydroxamic acid (9b) as an active which was found to inhibit HDAC1. In initial structure activity relationships study, the 1-methylbenzimidazole ring was replaced by the isosteric heterocycles benzimidazole, benzoxazole, and benzothiazole and the position of the hydroxamic acid substituent on the phenyl ring was varied. Whereas compounds bearing a para substituted hydroxamic acid (9a-d) were active HDAC inhibitors, the meta substituted analogues (8a-d) were appreciably inactive. Compounds 9a-d selectively inhibited HDAC6 (IC50 = 0.1–1.0μM) over HDAC1 (IC50 = 0.9–6μM) and moreover, also selectively inhibited the growth of lung cancer cells vs. patient matched normal cells. The compounds induce a cell cycle arrest in the S-phase while induction of apoptosis is neglible as compared to controls. Molecular modeling studies uncovered that the MM-GBSA energy for interaction of 9a-d with HDAC6 was higher than for HDAC1 providing structural rationale for the HDAC6 selectivity.

Synthesis, Cytotoxic, and Antitumor Activities of 2-Pyridylhydrazones Derived from 3-Benzoylpyridazines

A series of 2‐pyridylhydrazones derived from phenyl‐pyridazin‐3‐yl‐methanones were prepared in search for potential novel antitumor agents. The stereochemistry of these compounds was established by means of NMR spectroscopy. Whereas hydrazones derived from 3‐benzoylpyridazines (IC50 = 0.99–8.74 µM) inhibited the proliferation of the tumor cell lines tested, the non‐fully aromatic 3‐benzoylpyridazinone hydrazones (IC50 > 10 µM) turned out to be inactive. Compounds E‐1b (IC50 = 0.12 µM) and E‐1d (IC50 = 0.18 µM) exert high cytotoxic activities in clonogenic assays involving human tumor cells of different tissue origins. In vivo application of compound E‐1b (300 mg/kg/day) resulted in a 66% reduction in tumor burden.

Novel diazine derived hydrazones as potential ribonucleotide reductase inhibitors

Thiosemicarbazones derived from pyridine aldehydes aud ketoues (A) represent an interesti lg c ass of r bonucleotide reductase inhibilors I Previously we have showu, that replacement of the azine system by a 1,2-diaziue uucleus not only leads to improved solubility, but also to a reduction of cytotoxicily ia this class of compmmds without impairiug autiviral activity. Moreover, moderate synergism with AZT and acyclovir has been observed with such diazine derivatives 2 ~ R 1 I l e L ~ R1