Ashraf H. Abadi

Sulindac sulfide selectively inhibits growth and induces apoptosis of human breast tumor cells by phosphodiesterase 5 inhibition, elevation of cyclic GMP, and activation of protein kinase G

Sulindac displays promising antineoplastic activity, but toxicities from cyclooxygenase inhibition limit its use for chemoprevention. Previous reports suggest that its anticancer properties may be attributed to a cyclooxygenase-independent mechanism, although alternative targets have not been well defined. Here, we show that sulindac sulfide (SS) induces apoptosis and inhibits the growth of human breast tumor cells with IC50 values of 60 to 85 μmol/L. Within the same concentration range, SS inhibited cyclic GMP (cGMP) hydrolysis in tumor cell lysates but did not affect cyclic AMP hydrolysis. SS did not induce apoptosis of normal human mammary epithelial cells (HMEC) nor did it inhibit phosphodiesterase (PDE) activity in HMEC lysates. SS increased intracellular cGMP levels and activated protein kinase G in breast tumor cells but not HMEC. The guanylyl cyclase (GC) activator, NOR-3, and cGMP PDE inhibitors, trequinsin and MY5445, displayed similar growth-inhibitory activity as SS, but the adenylyl cyclase activator, forskolin, and other PDE inhibitors had no effect. Moreover, GC activation increased the sensitivity of tumor cells to SS, whereas GC inhibition reduced sensitivity. By comparing PDE isozyme profiles in breast tumor cells with HMEC and determining the sensitivity of recombinant PDE isozymes to SS, PDE5 was found to be overexpressed in breast tumor cells and selectively inhibited by SS. The mechanism of SS binding to the catalytic domain of PDE5 was revealed by molecular modeling. These data suggest that PDE5 inhibition is responsible for the breast tumor cell growth–inhibitory and apoptosis-inducing activity of SS and may contribute to the chemopreventive properties of sulindac. [Mol Cancer Ther 2009;8(12):3331–40]

Design, synthesis and biological evaluation of novel pyridine derivatives as anticancer agents and phosphodiesterase 3 inhibitors

Two series of 4,6-diaryl-2-imino-1,2-dihydropyridine-3-carbonitriles and their isosteric 4,6-diaryl-2-oxo-1,2-dihydropyridine-3-carbonitriles were synthesized through a combinatorial approach. The prepared analogues were evaluated for their in vitro capacity to inhibit PDE3A and the growth of the human HT-29 colon adenocarcinoma tumor cell line. Compound 6-(4-bromophenyl)-4-(2-ethoxyphenyl)-2-imino-1,2-dihydropyridine-3-carbonitrile (Id) exhibited the strongest PDE3 inhibition when cGMP but not cAMP is the substrate with a IC(50)of 27microM, which indicates a highly selective mechanism of enzyme inhibition. On the other hand, compound 6-(1,3-benzodioxol-5-yl)-4-(2-ethoxyphenyl)-2-imino-1,2-dihydropyridine-3-carbonitrile (Ii) was the most active in inhibiting colon tumor cell growth with a IC(50) of 3microM. The electronic effects, steric effects and conformational aspects of Id seem to be the most crucial for the PDE3 inhibition. Meanwhile, steric factors and the H-bonding capability seem to be the most important factors for tumor cell growth inhibitory activity. Conversely, there is no direct correlation between PDE3 inhibition and anticancer activity for the prepared compounds. An in silico docking experiment indicates the potential involvement of other potential molecular targets such as PIM-1 kinase to explain its tumor cell growth inhibitory activity.

Sulindac selectively inhibits colon tumor cell growth by activating the cGMP/PKG pathway to suppress Wnt/β-catenin signaling

Nonsteroidal anti-inflammatory drugs (NSAID) display promising antineoplastic activity for colorectal and other cancers, but toxicity from COX inhibition limits their long-term use for chemoprevention. Previous studies have concluded that the basis for their tumor cell growth inhibitory activity does not require COX inhibition, although the underlying mechanism is poorly understood. Here, we report that the NSAID sulindac sulfide inhibits cyclic guanosine 3′,5′-monophosphate phosphodiesterase (cGMP PDE) activity to increase intracellular cGMP levels and activate cGMP-dependent protein kinase (PKG) at concentrations that inhibit proliferation and induce apoptosis of colon tumor cells. Sulindac sulfide did not activate the cGMP/PKG pathway, nor affect proliferation or apoptosis in normal colonocytes. Knockdown of the cGMP-specific PDE5 isozyme by siRNA and PDE5-specific inhibitors tadalafil and sildenafil also selectively inhibited the growth of colon tumor cells that expressed high levels of PDE5 compared with colonocytes. The mechanism by which sulindac sulfide and the cGMP/PKG pathway inhibits colon tumor cell growth involves the transcriptional suppression of β-catenin to inhibit Wnt/β-catenin T-cell factor transcriptional activity, leading to downregulation of cyclin D1 and survivin. These observations suggest that safer and more efficacious sulindac derivatives can be developed for colorectal cancer chemoprevention by targeting PDE5 and possibly other cGMP-degrading isozymes. Mol Cancer Ther; 12(9); 1848–59. ©2013 AACR.

Synthesis of 4-alkyl (aryl)-6-aryl-3-cyano-2(1H)-pyridinones and their 2-imino isosteres as nonsteroidal cardiotonic agents

Thirty new 1,2-dihydropyridine derivatives of the general formula 4-alkyl (aryl)-6-aryl-3-cyano-2(1H)-pyridinones (1-15) and 4-alkyl (aryl)-6-aryl-3-cyano-2(1H)-iminopyridines (16-30) were synthesized using one-pot multicomponent reactions of the properly substituted acetophenone, appropriate aldehyde, ammonium acetate and ethyl cyanoacetate (1-15) or malononitrile (16-30) in ethanol. These target compounds (1-30) were evaluated for their cardiotonic activity using the spontaneously beating atria model, from reserpine-treated guinea pigs. The best pharmacological profile was obtained with 3-cyano-6-(3,4-dimethoxyphenyl)-4-(4-hydroxyphenyl)-2(1H)-pyridinone (9) which displaced selectivity for increasing the force of contraction (108.7 +/- 6.7,% change over control) rather than the frequency rate (40.8 +/- 5.3,% change over control) at a 5 x 10(-4) M concentration. The effects of structural changes upon activity are reported.

A Novel Sulindac Derivative that Potently Suppresses Colon Tumor Cell Growth by Inhibiting cGMP Phosphodiesterase and β-Catenin Transcriptional Activity

Nonsteroidal anti-inflammatory drugs (NSAIDs) have been widely reported to inhibit tumor growth by a COX-independent mechanism, although alternative targets have not been well defined or used to develop improved drugs for cancer chemoprevention. Here, we characterize a novel sulindac derivative referred to as sulindac benzylamine (SBA) that does not inhibit COX-1 or COX-2, yet potently inhibits the growth and induces the apoptosis of human colon tumor cells. The basis for this activity appears to involve cyclic guanosine 3′,5′,-monophosphate phosphodiesterase (cGMP PDE) inhibition as evident by its ability to inhibit cGMP hydrolysis in colon tumor cell lysates and purified cGMP-specific PDE5, increase intracellular cGMP levels, and activate cGMP-dependent protein kinase G at concentrations that suppress tumor cell growth. PDE5 was found to be essential for colon tumor cell growth as determined by siRNA knockdown studies, elevated in colon tumor cells as compared with normal colonocytes, and associated with the tumor selectivity of SBA. SBA activation of PKG may suppress the oncogenic activity of β-catenin as evident by its ability to reduce β-catenin nuclear levels, Tcf (T-cell factor) transcriptional activity, and survivin levels. These events preceded apoptosis induction and appear to result from a rapid elevation of intracellular cGMP levels following cGMP PDE inhibition. We conclude that PDE5 and possibly other cGMP degrading isozymes can be targeted to develop safer and more efficacious NSAID derivatives for colorectal cancer chemoprevention. Cancer Prev Res; 5(6); 822–33. ©2012 AACR.

Synthesis and antitumor activity of some new substituted quinolin-4-one and 1,7-naphthyridin-4-one analogs.

The synthesis of some new analogs of quinolin‐4‐one and 1,7‐naphthyridin‐4‐one is described. The prepared compounds were tested for their in vitro antitumor and cdc2 kinase or cdc25 phosphatase inhibitory activity. Compound ethyl 7‐oxo‐2,3‐dihydro‐7H‐pyrido [1,2,3‐de][2,3‐b]pyrido‐1,4‐thiazine‐6‐carboxylate (6b) showed antitumor activity against CNS SNB‐75, breast T‐47D, and lung NCI‐H522 cancer cell lines with GI50 values of 8.3, 17.6, and 22.7 μM, respectively. Meanwhile, the compounds ethyl 4‐oxo‐8‐phenylthio‐1H,4H‐quinoline‐3‐carboxylate (11a) and 4‐oxo‐8‐phenylthio‐1H,4H‐1,7‐naphthyridine‐3‐carboxylic acid (12b) have proved to be cdc25 phosphatase inhibitors at IC50 values of 11 and 5 μM, respectively.

Synthesis and Antitumor Activity of Ethyl 2-Substituted-aminothiazole-4-carboxylate Analogs

The synthesis of several new ethyl 2‐substituted‐aminothiazole‐4‐ carboxylate analogs is described. The prepared compounds were tested for their in vitro antitumor activity against 60 human tumor cell lines by the National Cancer Institute (NCI) and showed potential anticancer activity. Ethyl 2‐[3‐(diethylamino)‐propanamido]‐thiazole‐4‐carboxylate (14) exhibited remarkable activity against RPMI‐8226 leukemia cell line with GI50 value of 0.08 μM, and a broad spectrum activity against all the tumor cell lines used with GI50 (MG‐MID) value of 38.3 μM. The detailed synthesis and antitumor screening data are reported.

Discovery of colon tumor cell growth inhibitory agents through a combinatorial approach

Two series with the general formula of 4,6-diaryl-2-oxo-1,2 dihydropyridine-3-carbonitriles and their isosteric 4,6-diaryl-2-imino-1,2-dihydropyridine-3-carbonitrile were synthesized through one pot reaction of the appropriate acetophenone, aldehyde, ammonium acetate with ethyl cyanoacetate or malononitrile, respectively. The synthesized compounds were evaluated for their tumor cell growth inhibitory activity against the human HT-29 colon tumor cell line, as well as their PDE3 inhibitory activity. Compound 4-(2-Ethoxyphenyl)-2-oxo-6-thiophen-3-yl-1,2-dihydropyridine-3 carbonitrile (21) showed tumor cell growth inhibitory activity with an IC50 value of 1.25 microM. Meanwhile, 4-(4-Ethoxyphenyl)-2-imino-6-(thiophen-3-yl)-1,2-dihydropyridine-3-carbonitrile (26) showed inhibitory effect upon PDE3 using cAMP or cGMP as substrate. No correlation exists between PDE3 inhibition and the tumor cell growth inhibitory activity. Docking compound 21 to other possible molecular targets showed the potential to bind PIM1 Kinase.

The voltammetric study and determination of ramipril in dosage forms and biological fluids

The voltammetric behavior of ramipril was studied using cyclic voltammetry, direct current polarography (DCt), differential pulse polarography (DPP) and alternating current polarography (ACt). Ramipril developed well-defined cathodic waves in Britton-Robinson buffers over the pH range 6-12. The waves were characterized as being diffusion-controlled, irreversible and partially affected by adsorption phenomenon. The diffusion-current constant (Id) was 1.24 +/- 0.02. The current-concentration plots were rectilinear over the range 10-50, 4-40 and 0.16-12 micrograms/ml in the DCt, DPP and ACt modes, respectively, with a minimum detectability (S/N = 2) of 0.02 microgram/ml (4.8 x 10(-8) M) using the latter mode. The proposed method was successfully applied to the determination of ramipril in commercial tablets. Hydrochlorothiazide, which is frequently co-formulated with ramipril, did not interfere with the assay. Furthermore, the proposed method was applied to the determination of ramipril in urine and plasma adopting the ACt technique. The percentage recoveries were 97.12 +/- 0.56 and 94.97 +/- 0.62%, respectively. A pathway for the electrode reaction was proposed.

Synthesis and in vitro antiproliferative effect of novel quinoline-based potential anticancer agents.

Several derivatives with a quinoline scaffold and a flexible, semi-flexible or rigid side chains at position 8 of the quinoline ring were synthesized and assessed for their in vitro activity versus the human colon cancer cell line HT29 and the human breast cancer cell line MDA-MB231. The HT29 cell line was more refractory to the cytotoxic activity of some compounds, meanwhile all the quinoline derivatives except one displayed high to moderate activity against MDA-MB231 with IC50 values ranging between 4.6 and 48.2 μM. The most active derivative in this study against both tested cell lines was the Schiffs base 4e with IC50 of 4.7 and 4.6 μM against HT29 and MDA-MB231, respectively.