New derivatives of 11-methyl-6-[2-(dimethylamino)ethyl]-6H-indolo[2,3-b]quinoline as cytotoxic DNA topoisomerase II inhibitors

Novel indolo[2,3-b]quinoline derivatives substituted at N-6 and C-2 or C-9 positions with (dimethylamino)ethyl chains linked to heteroaromatic core by ether, amide or amine bonds, were manufactured and evaluated in vitro for their cytotoxic activity against several cell lines of different origin including multidrug resistant sublines and tested for their ability to influence the cell cycle and inhibit topoisomerase II activity. It was found, that all compounds show cytotoxic activity against cell lines tested, including multidrug resistant LoVo/DX, MES-SA/DX5 and HL-60 sublines. The tested compounds induce the G(2)M phase cell cycle arrest in Jurkat cells, and inhibit topoisomerase II activity.     

Synthesis, and cytotoxic activity of some novel indolo[2,3-b]quinoline derivatives: DNA topoisomerase II inhibitors

A series of new 5H-indolo[2,3-b]quinoline derivatives bearing methoxy and methyl groups at C-2 and C-9 was synthesized (according to the modified Graebe-Ullmann reaction). These compounds were evaluated for their antimicrobial and cytotoxic activity and tested as inhibitors of DNA topoisomerase II. Lipophilic and calf thymus DNA binding properties of these compounds were also established. In the SAR studies we used quantum-mechanical methodology to analyze the molecular properties of the drugs. All of the 5H-indolo[2,3-b]quinolines tested were found to inhibit the growth of gram-positive bacteria and pathogenic fungi at MIC ranging between 2.0 and 6.0 microM. They showed also cytotoxic activity in vitro against several human cancer cell lines of different origin (ID50 varied from 0.6 to 1.4 microM), and stimulated the formation of topoisomerase-II-mediated pSP65 DNA cleavage at concentration between 0.2 and 0.5 microM. The most active indolo[2,3-b]quinolines which had the greatest contribution to the increase in the Tm of DNA displayed also the highest DNA binding constants and the highest cytotoxic activity. The differences in DNA binding properties and cytotoxic activity seem to be more related to steric than electrostatic effects.     

Novel cyano- and amidino-substituted derivatives of thieno[2,3-b]- and thieno[3,2-b]thiophene-2-carboxanilides and thieno[3',2':4,5]thieno- and thieno[2',3':4,5]thieno [2,3-c]quinolones: synthesis, photochemical synthesis, DNA binding, and antitumor evaluation

A series of cyano- and amidino-substituted derivatives of thieno[2,3-b]- and thieno[3,2-b]thiophene-2-carboxanilides and their 'cyclic' derivatives (quinolones) were synthesized. 'Cyclic' compounds displayed a rather strong and differential antiproliferative effect on various cell lines, while the 'acyclic' amidino-substituted compounds were much more active, but showing mostly non-differential cytotoxicity, whereas cyano-substituted compounds (2a,b) produced a strikingly strong effect selectively on HeLa and Hep-2 cell lines. Antiproliferative activity of 'cyclic' derivatives is very likely caused by intercalation into DNA, while their 'acyclic' analogues use other target(s) and/or mechanisms of action.     

Synthesis and antidiabetic activity of 2,5-disubstituted-3-imidazol-2-yl-pyrrolo[2,3-b]pyridines and thieno[2,3-b]pyridines

In the present investigation, two series of 2,5-disubstituted-3-imidazol-2-yl-pyrrolo[2,3-b]pyridines (2a-l) and thieno[2,3-b]pyridines (3a-l) were designed as analogs of BL 11282 (1). The in vitro glucose dependent insulinotropic activity of all the test compounds was evaluated using RIN5F cell based assay and all the test compounds showed glucose and concentration dependent insulin secretion. The in vivo antidiabetic activities of most potent compounds from each series (2c and 3c) were assessed in C57BL/6J mice. Compounds 2c and 3c showed dose dependent insulin secretion and significant glucose reduction in vivo. In general, compounds 2c and 3c were found to be equipotent at all the three different doses selected and with respect to BL 11282, both the test compounds were found to be more potent, at all the time points.     

Design,synthesis, cytotoxic evaluation,and QSAR study of some 6H-indolo[2,3-b]quinoxaline derivatives

In the pathway of anticancer drug development, we designed and synthesized some 6H-indolo[2,3-b]quinoxaline derivatives (which act as DNA intercalators) by structural modification. The structure of the 6H-indolo[2,3-b]quinoxaline derivatives was confirmed by IR, NMR, Mass and elemental analysis. The compounds (IDQ-5, IDQ-10, IDQ-11, IDQ-13, and IDQ-14) exhibited significant in vitro activity against a human leukemia (HL-60) cell line. The QSAR derived for modeling the cytotoxic activity of 6H-indolo[2,3-b]quinoxaline derivatives suggests that candidate structures for increased cytotoxic potency should incorporate cyclic substituents or substituents with primary carbon atoms.     

Synthesis and cytotoxicity evaluation of 6,11-dihydro-pyridazo- and 6,11-dihydro-pyrido[2,3-b]phenazine-6,11-diones

The 6,11-dihydro-pyridazo[2,3-b]phenazine-6,11-dione and 6,11-dihydro-pyrido[2,3-b]phenazine-6,11-dione derivatives were synthesized from 6,7-dichloro-5,8-phthalazinedione and 6,7-dichloro-5,8-quinolinedione, respectively, producing a series of new anticancer drugs. The cytotoxic activities of the prepared compounds were evaluated by a SRB (Sulforhodamine B) assay against the following tumor cell lines: A459 (human lung), SK-OV-3 (human ovarian), SK-MEL-2 (human melanoma), XF498 (human CNS), and HCT 15 (human colon). Almost all the derivatives of the 6,11-dihydro-pyridazo[2[,3-b]phenazine-6,11-dione and 6,11-dihydro-pyrido[2,3-b]phenazine-6,11-dione, tetracyclic heteroquinone analogues with four or three nitrogen atoms, exhibited excellent cytotoxicity on almost all the human tumor cell lines tested. Specifically, 6,11-dihydro-pyridazo[2,3-b]phenazine-6,11-dione (4a) exhibited potent activity against all the tumor cell lines, and in particular, its cytotoxic effect against HCT 15 (ED(50)=0.004 microg/mL) was 25 times greater than that of doxorubicin (ED(50)=0.093 microg/mL).     

Novel 1-(azacyclyl)-3-arylsulfonyl-1H-pyrrolo[2,3-b]pyridines as 5-HT6 agonists and antagonists

1-Aminoethyl-3-arylsulfonyl-1H-indoles 1 are 5-HT(6) receptor ligands with modest activity in a 5-HT(6) cyclase assay. Introduction of an additional nitrogen in the indole ring provides 1-aminoethyl-3-arylsulfonyl-1H-pyrrolo[2,3-b]pyridines 2 with both enhanced 5-HT(6) affinity and cyclase activity, many acting as 5-HT(6) agonists. We constrained the basic side chain as part of a ring to make 1-(azacyclyl)-3-arylsulfonyl-1H-pyrrolo[2,3-b]pyridines incorporating a pyrrolidinyl 3 or piperidinyl 4 ring system. Preparation of compounds 3 and 4 required synthesis of the key intermediates, 1-(pyrrolidin-3-yl)-1H-pyrrolo[2,3-b]pyridines 7 and 1-(piperidin-3-yl)-1H-pyrrolo[2,3-b]pyridines 8, respectively. Intermediates 7 were prepared through alkylation of 7-azaindole while the intermediates 8 required an alternate synthesis. The compounds of both series 3 and 4 were shown to have high binding affinities for the 5-HT(6) receptor. The in vitro functional activity at the 5-HT(6) receptor varied depending on various functionalities including the selection of the arylsulfonyl, the substitution on the arylsulfonyl group, the ring size, and the substitution on the basic amine moiety producing either 5-HT(6) receptor agonists or antagonists.     

Rebeccamycin analogues from indolo[2,3-c]carbazole

Glycosylated indolocarbazoles related to the antibiotic rebeccamycin represent an important series of antitumor drugs. In the course of structure-activity relationship studies, we report the synthesis of two new derivatives containing an indolo[2,3-c]carbazole chromophore instead of the conventional indolo[2,3-a]carbazole unit found in the natural metabolites. The N-methylated compound 8 containing one glucose residue behaves as a typical DNA intercalating agent, as judged from circular and electric linear dichroism measurements with purified DNA. In contrast, the bis-glycosylated derivative 7 containing a glucose residue on each indole nitrogen has lost its capacity to form stable complexes with DNA. DNA relaxation experiments reveal that the two drugs 7 and 8 have weak effects on human DNA topoisomerase I. The modified conformation of the indolocarbazole chromophore is detrimental to the stabilization of topoisomerase I-DNA complexes. The lack of potent topoisomerase I inhibition leads to decreased cytotoxicity but, however, we observed that the DNA-intercalating mono-glycosyl derivative 8 is about 5 times more cytoxic than the bis-glycosyl analogue 7. The study suggests that the naturally-occurring indolo[2,3-a]carbazole skeleton should be preserved to maintain the topoisomerase I inhibitory and cytotoxic activities.     

Interaction of ellipticine and an indolo[2,3b]-quinoxaline derivative with DNA and synthetic polynucleotides

The non-covalent DNA interaction of the anticancer drug ellipticine (Scheme I, 1a) as well as an indolo[2,3-b]-quinoxaline derivative (Scheme I, 3b) with a dimethylaminoethyl side chain has been studied by light absorption, linear dichroism (LD) and fluorescence. Compound 3b (Scheme I) has antitumorigenic as well as antiviral activity. Both compounds bind to DNA or synthetic polynucleotides such as poly(dA-dT).(dA-dT) and poly(dG-dC).(dG-dC) by intercalation. In contrast to ellipticine, compound 3b (Scheme I) exhibits a significant binding specificity for alternating AT sequences. Its fluorescence is strongly enhanced in AT sequences and quenched in GC sequences. Fluorescence titrations evaluated as Scatchard plots show that both ellipticine and compound 3b (Scheme I) bind to the nucleic acids according to a non-cooperative neighbor exclusion model.     

Cytotoxic activity toward KB cells of 2-substituted naphtho[2,3-b]furan-4,9-diones and their related compounds

We investigated the cytotoxic activity of 2-substituted naphtho[2,3-b]furan-4,9-diones. We have previously synthesized 33 types of 2-substituted and related compounds, and the cytotoxic activity of these compounds was then examined by a KB cell culture assay. 2-(3-Furanoyl)benzoic acids and 1,4-naphthoquinones had no activity. 2-Acetyl-4,9-dimethoxynaphtho[2,3-b]furan 4 showed low activity. However, parent naphtho[2,3-b]furan-4,9-dione 2 and most 2-substituted derivatives exhibited cytotoxic activity. The parent structure was therefore for cytotoxicity. 2-Formylnaphtho[2,3-b]furan-4,9-dione 11 had particularly potent activity (ED50=0.09 microg/ml).     

2-Anilinonicotinyl linked 2-aminobenzothiazoles and [1,2,4]triazolo[1,5-b] [1,2,4]benzothiadiazine conjugates as potential mitochondrial apoptotic inducers

A series of N-(2-anilino-pyridyl) linked 2-amino benzothiazoles (4a-n) and [1,2,4]triazolo [1,5-b]benzothiadiazine conjugates (5a-j) have been designed, synthesized and evaluated for their antiproliferative activity. Some of these compounds (4h-k, 4n, and 5e) have exhibited potent cytotoxicity specifically against human leukemia HL-60 cell lines with IC(50) values in the range of 0.08-0.70 μM. All these compounds were tested for their effects on the cell cycle perturbations and induction of apoptosis. Morphological evidences of apoptosis, including fragmentation of nuclei and inter nucleosomal DNA laddering formation were clearly observed after 24h exposure to compound 4i. Flow cytometry analysis revealed that compound 4i showed drastic cell cycle perturbations due to concentration dependant increase in the sub-G0 region which comprises of both the apoptotic and debris fraction, thus implying the extent of cell death. These compounds trigger the mitochondrial apoptotic pathway that results in the loss of mitochondrial membrane potential through activation of multiple caspases followed by activation of caspase-3, and finally cleavage of PARP. Further the mechanism of cell death was analysed by fluorescent microscopic analysis and also by scanning electron microscopy. The cytotoxicity of 4i correlated with induction of apoptosis, caspases activation and DNA damage and thus indicating the apoptotic pathway of anticancer effect of these compounds.     

Synthesis and evaluation of the antiproliferative activity of novel isoindolo[2,1-a]quinoxaline and indolo[1,2-a]quinoxaline derivatives

A novel series of isoindolo[2,1-a]quinoxaline and indolo[1,2-a]quinoxaline derivatives was synthesized and evaluated in vitro against various human cancer cell lines for antiproliferative activity. These new compounds displayed activity against leukemia and breast cancer cell lines in the 3- to 18-μM concentration range.     

UDP-glucuronosyltransferase-mediated metabolic activation of the tobacco carcinogen 2-amino-9H-pyrido[2,3-b]indole

2-Amino-9H-pyrido[2,3-b]indole (AαC) is a carcinogenic heterocyclic aromatic amine (HAA) that arises in tobacco smoke. UDP-glucuronosyltransferases (UGTs) are important enzymes that detoxicate many procarcinogens, including HAAs. UGTs compete with P450 enzymes, which bioactivate HAAs by N-hydroxylation of the exocyclic amine group; the resultant N-hydroxy-HAA metabolites form covalent adducts with DNA. We have characterized the UGT-catalyzed metabolic products of AαC and the genotoxic metabolite 2-hydroxyamino-9H-pyrido[2,3-b]indole (HONH-AαC) formed with human liver microsomes, recombinant human UGT isoforms, and human hepatocytes. The structures of the metabolites were elucidated by (1)H NMR and mass spectrometry. AαC and HONH-AαC underwent glucuronidation by UGTs to form, respectively, N(2)-(β-D-glucosidurony1)-2-amino-9H-pyrido[2,3-b]indole (AαC-N(2)-Gl) and N(2)-(β-D-glucosidurony1)-2-hydroxyamino-9H-pyrido[2,3-b]indole (AαC-HON(2)-Gl). HONH-AαC also underwent glucuronidation to form a novel O-linked glucuronide conjugate, O-(β-D-glucosidurony1)-2-hydroxyamino-9H-pyrido[2,3-b]indole (AαC-HN(2)-O-Gl). AαC-HN(2)-O-Gl is a biologically reactive metabolite and binds to calf thymus DNA (pH 5.0 or 7.0) to form the N-(deoxyguanosin-8-yl)-AαC adduct at 20-50-fold higher levels than the adduct levels formed with HONH-AαC. Major UGT isoforms were examined for their capacity to metabolize AαC and HONH-AαC. UGT1A4 was the most catalytically efficient enzyme (V(max)/K(m)) at forming AαC-N(2)-Gl (0.67 μl·min(-1)·mg of protein(-1)), and UGT1A9 was most catalytically efficient at forming AαC-HN-O-Gl (77.1 μl·min(-1)·mg of protein(-1)), whereas UGT1A1 was most efficient at forming AαC-HON(2)-Gl (5.0 μl·min(-1)·mg of protein(-1)). Human hepatocytes produced AαC-N(2)-Gl and AαC-HN(2)-O-Gl in abundant quantities, but AαC-HON(2)-Gl was a minor product. Thus, UGTs, usually important enzymes in the detoxication of many procarcinogens, serve as a mechanism of bioactivation of HONH-AαC.     

Antibacterial profile against drug-resistant Staphylococcus epidermidis clinical strain and structure-activity relationship studies of 1H-pyrazolo[3,4-b]pyridine and thieno[2,3-b]pyridine derivatives

Antibacterial resistance is a complex problem that contributes to health and economic losses worldwide. The Staphylococcus epidermidis is an important nosocomial pathogen that affects immunocompromised patients or those with indwelling devices. Currently, there are several resistant strains including S. epidermidis that became an important medical issue mainly in hospital environment. In this work, we report the biological and theoretical evaluations of a 4-(arylamino)-1-phenyl-1H-pyrazolo[3,4-b]pyridine-5-carboxylic acids series (1, 1a-m) and the comparison with a new isosteric ring nucleus series, 4-(arylamino)thieno[2,3-b]pyridine-5-carboxylic acids derivatives (2, 2a-m). Our results revealed the 1H-pyrazolo[3,4-b]pyridine derivatives significant antibacterial activity against a drug-resistant S. epidermidis clinical strain in contrast to the thieno[2,3-b]pyridine series. The minimal inhibitory concentration (MIC) of the most active derivatives (1a, 1c, 1e, and 1f) against S. epidermidis was similar to that of oxacillin and twofold better than chloramphenicol. Interestingly, the position of the functional groups has a great impact on the activity as observed in our structure-activity relationship (SAR) study. The SAR of 1H-pyrazolo[3,4-b]pyridine derivatives shows that the highest inhibitory activity is observed when the meta position is occupied by electronegative substituents. The molecular modeling analysis of frontier molecular orbitals revealed that the LUMO density is less intense in meta than in ortho and para positions for both series (1 and 2), whereas HOMO density is overconcentrated in 1H-pyrazolo[3,4-b]pyridine ring nucleus compared to the thieno[2,3-b]pyridine system. The most active derivatives of series 1 were submitted to in silico ADMET screening, which confirmed these compounds as potential antibacterial candidates.     

Design, synthesis, and biological evaluation of substituted-N-(thieno[2,3-b]pyridin-3-yl)-guanidines, N-(1H-pyrrolo[2,3-b]pyridin-3-yl)-guanidines, and N-(1H-indol-3-yl)-guanidines

Sulfonylureas stimulate insulin secretion independent of the blood glucose concentration and therefore cause hypoglycemia in type 2 diabetic patients. Over the last years, a number of aryl-imidazoline derivatives have been identified that stimulate insulin secretion in a glucose-dependent manner. In the present study, we have developed three series of substituted N-(thieno[2,3-b]pyridin-3-yl)-guanidine (2a-l), N-(1H-pyrrolo[2,3-b]pyridin-3-yl)-guanidine (3a-l), and N-(1H-indol-3-yl)-guanidine (4a-l) as new class of antidiabetic agents. In vitro glucose-dependent insulinotropic activity of test compounds 2a-l, 3a-l, and 4a-l was evaluated using RIN5F (Rat Insulinoma cell) based assay. All the test compounds showed concentration-dependent insulin secretion, only in presence of glucose load (16.7mmol). Some of the test compounds (2c, 3c, and 4c) from each series were found to be equipotent to BL 11282 (standard aryl-imidazoline), which indicated that the guanidine group acts as a bioisostere of imidazoline ring system.     

3-(4-Piperidinyl)indoles and 3-(4-piperidinyl)pyrrolo-[2,3-b]pyridines as ligands for the ORL-1 receptor

A novel series of indoles and 1H-pyrrolo[2,3-b]pyridines having a piperidine ring at the 3-position were synthesized and found to bind with high affinity to the ORL-1 receptor. Structure-activity relationships at the piperidine nitrogen were investigated in each series. Substitution on the phenyl ring and nitrogen atom of the indole and 1H-pyrrolo[2,3-b]pyridine cores generated several selective high-affinity ligands that were agonists of the ORL-1 receptor.     

Specific stabilization of DNA triple helices by indolo[2,1-b]quinazolin-6,12-dione derivatives

Derivatives of indolo[2,1-b]quinazolinone containing aminoalkylamino side chains were synthesized as specific DNA triplex stabilizing agents. The aminoalkylamino side chains are essential for triplex stabilization. The position-8 fluorine atom or a methyl group to the nitrogen adjacent to the planar core can enhance triplex stability by 6 degrees C and the effect is additive. Conformational analysis reveals that the orientation of the side chain underlies the ability of this compound to stabilize a DNA triplex.     

One-pot solvent free synthesis and DNA binding studies of thieno[2,3-b]-1,8-naphthyridines

With the aim of evaluating interaction between double-stranded calf thymus (ds)DNA and sulphur containing fused planar rings, the derivatives of 1,8-naphthyridine containing thiono groups were synthesized by the condensation of 2-mercapto-3-formyl[1,8]naphthyridines using 1-chloroacetone, 2-chloroacetamide, chloroaceticacid, and 2-chloro-1-phenylethanone in the presence of anhydrous potassium carbonate as s catalyst under solvent free microwave irradiation. The structures of the compounds were elucidated on the basis of elemental analysis, IR, (1)H NMR, and mass spectra. The interaction of thieno[2,3-b]-1,8-naphthyridine-2-carboxylic acid (TNC) (3a) with ct-DNA was studied by UV-Vis spectrophotometry, viscosity, thermal denaturation, as well as cyclic voltammetry experiments. On binding to DNA, the absorption spectrum underwent bathochromic and hypochromic shifts. Binding parameters, determined from spectrophotometric measurements indicated a binding constant of Kb=2.1 x 10(6) M(-1). The thieno[2,3-b]-1,8-naphthyridine-2-carboxylic acid (3a) increases the viscosity of sonicated rod-like DNA fragments. The binding of TNC to DNA increased the melting temperature by about 4 degrees C. The decrease in peak current heights and shifts of peak potential values are observed by the addition of calf thymus DNA in cyclic voltammetry studies.     

Synthesis and cytotoxicity of 6,11-dihydro-pyrido- and 6,11-dihydro-benzo[2,3-b]phenazine-6,11-dione derivatives

6,11-Dihydro-pyrido[2,3-b]phenazine-6,11-diones and 6,11-dihydro-benzo[2,3-b]phenazine-6,11-diones were synthesized from 6,7-dichloro-5,8-quinolinedione and 2,3-dichloro-1,4-naphthoquinone. The study on the cytotoxicity on these products revealed that the pyridophenazinediones, tetracyclic heteroquinone analogues with three nitrogen atoms exhibited a high cytotoxicity on several human tumor cell lines. Compound 9c and 9e showed in vitro antitumor activity comparable or superior to doxorubicin against the human ovarian tumor cells (SK-OV-3) and the human CNS cells (XF 498). The IC(50) value for compound 9e was 0.06 microM against the human CNS cells (XF 498), which was 2.6 times higher than that (0.16 microM) of doxorubicin. In addition, the X-ray crystallographic analysis of two phenazinedione derivatives (9b,c) showed clearly the exact position of the nucleophilic substitution of 6,7-dichloro-5,8-quinolinedione.     

Synthesis and antiproliferative evaluation of 4-anilino-n-methoxyfuro[2,3-b]quinoline derivatives (n=6, 7). Part 5

A series of 27 differently substituted 4-anilinofuro[2,3-b]quinolines were synthesized and evaluated for their antiproliferative activities against the HeLa, SKHep1, SAS, AGS, A549, and CE81T cell lines, cancers commonly found in Asian countries. Among the compounds tested, 1-{4-[(3-chloro-7-methoxyfuro[2,3-b]quinolin-4-yl)amino]phenyl}ethanone (1) was the most potent, with IC(50) values of 3.1, 3.0, and 4.2 microM, resp., against the growth of HeLa, SKHep, and CE81T cells. Compound 1 was, thus, further evaluated by flow cytometry to evaluate its effect on the cell-cycle distribution of HeLa cells. Our results indicated that 1 readily induces cell-cycle arrest in the G2/M phase, followed by DNA fragmentation and, ultimately, cell death.