New potent and selective inhibitor of Pim-1/3 protein kinases sensitizes human colon carcinoma cells to doxorubicin.

International audienceThe Pim protein kinases (provirus insertion site of Moloney murine leukemia virus) have been identified as important actors involved in tumor cell survival, proliferation, migration and invasion. Therefore, inhibition of Pim activity by low molecular weight compounds is under investigation as a part of anticancer therapeutic strategies. We have synthesized a series of pyrrolo[2,3-a]carbazole derivatives that significantly inhibited Pim protein kinases at submicromolar concentrations. Particularly, benzodiazocine derivative 1 potently inhibited Pim-1 and -3 isoforms in in vitro kinase assays (IC50 8 nM and 13 nM, respectively), whereas Pim-2 activity was less affected (IC50 350 nM). We show here that no inhibitory effect of 1 was detectable at 1 µM against other 22 serine/threonine and tyrosine kinases. In addition, 1, possessing a planar pyrrolocarbazole scaffold, demonstrated no significant binding to DNA, nor was it a potent topoisomerase I inhibitor, suggesting that 1 is likely to be highly selective for Pim-1 and -3. Importantly, whereas 1 exerted a negligible cytotoxicity for human colon carcinoma HCT116 cell line at concentrations >10 µM within 72 h of cell exposure, it synergized at nontoxic concentrations with the antitumor drug doxorubicin (Dox) in killing HCT116 cells: IC50 of Dox alone and Dox+1 were ~200 nM and ~25 nM, respectively. These data strongly suggest that 1 emerges as a prospective antitumor drug candidate due to its selectivity to individual Pim protein kinases and the ability to potentiate the efficacy of conventional chemotherapeutics

The first series of 4,11-bis[(2-aminoethyl)amino]anthra[2,3-b]furan-5,10-diones: Synthesis and anti-proliferative characteristics

We developed the synthesis of a series of furan-fused tetracyclic analogues of the antitumor agent ametantrone. The reactions included nucleophilic substitution of propoxy groups in 4,11-dipropoxyanthra[2,3-b]furan-5,10-diones with ethylenediamines, producing the derivatives of 4,11-diaminoanthra[2,3-b]furan-5,10-dione in good yields. Studies of anti-proliferative activity on a panel of mammalian tumor cell lines demonstrated that anthra[2,3-b]furan-5,10-diones were the most potent derivatives among heteroarene-fused ametantrone analogues with one heteroatom. We identified several compounds that evoked a growth inhibitory effect at submicromolar concentrations. The anthra[2,3-b]furan-5,10-dione 9 with distal methylamino groups was markedly potent against drug-resistant cell lines with P-glycoprotein overexpression or p53 gene deletion. Furthermore, this derivative attenuated in vitro topoisomerase I-mediated DNA uncoiling at low micromolar concentrations. These results demonstrate that anthrafurandiones are a new class of heterocyclic anthraquinone derivatives with the properties potentially valuable for anticancer therapy.status: publishe

New antitumor anthra[2,3-b]furan-3-carboxamides: synthesis and structure-activity relationship

Chemical modifications of the anthraquinone scaffold are aimed at optimization of this exceptionally productive class of antitumor drugs. In particular, our previously reported anthra[2,3-b]furan-3-carboxamides demonstrated a high cytotoxic potency in cell culture and in vivo. In this study, we expanded our series of anthra[2,3-b]furan-3-carboxamides by modifying the key functional groups and dissected the structure-activity relationship within this chemotype. The majority of new compounds inhibited the growth of mammalian tumor cell lines at submicromolar to low micromolar concentrations. We found that 4,11-hydroxy groups as well as the carbonyl moiety in the carboxamide fragment were critical for cytotoxicity whereas the substituent at the 2-position of anthra[2,3-b]furan was not. Importantly, the new derivatives were similarly potent against wild type cells and their variants resistant to doxorubicin due to P-glycoprotein (Pgp) expression or p53 inactivation. The most cytotoxic derivatives 6 and 9 attenuated plasmid DNA relaxation by topoisomerase 1. Finally, we demonstrated that 6 and 9 at 1 μM induced intracellular oxidative stress, accumulation in G2/M phase of the cell cycle, and apoptosis in gastric carcinoma cell lines regardless of their p53 status. These results further substantiate the potential of anthra[2,3-b]furan-3-carboxamides as antitumor drug candidates.status: publishe

Synthesis and biological activity of pyrazole analogues of the staurosporine aglycon K252c.

International audienceA derivative of the staurosporine aglycon (K252c), in which the lactam ring was replaced by a pyrazole moiety, was synthesized. The resulting indolopyrazolocarbazole (3) inhibited Pim isoforms 1–3 whereas it did not impair the activity of two known targets of K252c, protein kinase C isoforms α and γ. Compound 3 exhibited moderate cytotoxic activity toward human leukemia and colon carcinoma cell lines (K562 and HCT116), strongly suggesting that this new scaffold deserves further investigations for treatment of malignancies associated with Pim activity

Fluorescence of free and bound to DNA of compounds 1–3.

<p>Comparison of fluorescence intensity of compounds <b>1</b> (circles)<b>, 2</b> (squares) and <b>3 (</b>triangles) in the presence (A) or absence (B) of magnesium cations. Open symbols, fluorescence of free (unbound) <b>1–</b>3; filled symbols, fluorescence of <b>1–3</b> in complexes with pUC19 DNA (10 μM; bp). Excitation: 440 nm, fluorescence detection: 480 nm.</p

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<p><b>Dependence of DNA binding affinity of compounds 1 (A) and 3 (B) on the ionic strength of solution.</b> DNA binding constants of <b>1</b> and <b>3</b> obtained in solutions that contained 5mM MgCl₂ (filled circles) and no MgCl₂ (open circles). Linear approximations shown by solid lines with the slope SK = ∂logK/∂log[KCl].</p

New antitumor anthra[2,3-b]furan-3-carboxamides: Synthesis and structure-activity relationship

Chemical modifications of the anthraquinone scaffold are aimed at optimization of this exceptionally productive class of antitumor drugs. In particular, our previously reported anthra[2,3-b]furan-3-carboxamides demonstrated a high cytotoxic potency in cell culture and in vivo. In this study, we expanded our series of anthra[2,3-b]furan-3-carboxamides by modifying the key functional groups and dissected the structure-activity relationship within this chemotype. The majority of new compounds inhibited the growth of mammalian tumor cell lines at submicromolar to low micromolar concentrations. We found that 4,11-hydroxy groups as well as the carbonyl moiety in the carboxamide fragment were critical for cytotoxicity whereas the substituent at the 2-position of anthra[2,3-b]furan was not. Importantly, the new derivatives were similarly potent against wild type cells and their variants resistant to doxorubicin due to P-glycoprotein (Pgp) expression or p53 inactivation. The most cytotoxic derivatives 6 and 9 attenuated plasmid DNA relaxation by topoisomerase 1. Finally, we demonstrated that 6 and 9 at 1 μM induced intracellular oxidative stress, accumulation in G2/M phase of the cell cycle, and apoptosis in gastric carcinoma cell lines regardless of their p53 status. These results further substantiate the potential of anthra[2,3-b]furan-3-carboxamides as antitumor drug candidates

Salt dependent thermodynamic parameters of binding of 1 and 3 to DNA calculated on the basis of fluorescence measurements.

<p>Salt dependent thermodynamic parameters of binding of 1 and 3 to DNA calculated on the basis of fluorescence measurements.</p

Binding of compounds 1–3 to the pUC19 plasmid DNA monitored by electrophoretic mobility in 1% agarose gel.

<p>The plasmid was incubated with the compounds at indicated concentrations (μM) in BB-Mg buffer containing 5 mM MgCl₂ (A) or BB buffer (same buffer with no MgCl₂) (B). Migration of the free compound is shown at the highest concentration (25 μM) for each drug (arrows). Bottom panels in A and B: electrophoresis with EtBr in the gel and in the running buffer.</p

Changes of CD spectra of compound 3 upon binding to DNA.

<p>CD spectra of unbound compound <b>3</b> in the absence of DNA (open circles) and in complexes with DNA (filled circles) in BB-Mg (A) or BB (B). DNA concentration increased in the range 5–50 μM (bp), the concentration of <b>3</b> was 20 μM.</p