Solid phase synthesis of structurally diverse pyrimido[4,5-d] pyrimidines for the potential use in combinatorial chemistry

An efficient solid phase synthesis of pyrimido[4,5-d]pyrimidine derivatives is described. Reaction of polymer-bound pyrimidine 1 with urea or thiourea followed by cleavage from the support provided 4-aminopyrimido[4,5-d]pyrimidines 4 and 5 while treatment of 6 with phenyl isocyanate or phenyl isothiocyanate followed by cleavage from resin afforded 3-phenylpyrimido[4,5-d]pyrimidines 9 and 10.     

Identification of 5,6-substituted 4-aminothieno[2,3-d]pyrimidines as LIMK1 inhibitors

4-Aminobenzothieno[3,2-d]pyrimidines were previously identified in a high throughput screening campaign as LIMK1 inhibitors. Scaffold reversal led to the identification of a series of simple 5,6-substituted 4-aminothieno[2,3-d]pyrimidines with low micromolar inhibition of LIMK1.     

Regiospecific one-pot synthesis of pyrimido[4,5-d]pyrimidine derivatives in the solid state under microwave irradiations

Electron rich 6-[(dimethylamino)methylene]amino uracil 1, undergoes [4+2] cycloaddition reactions with various in situ generated glyoxylate imine and imine oxides 6 to provide novel pyrimido[4,5-d]pyrimidine derivatives of biological significance, after elimination of dimethylamine from the (1:1) cycloadducts and oxidative aromatisation. This procedure provides a convenient method for the direct synthesis of pyrimido[4,5-d]pyrimidines in excellent yields when carried out in the solid state and under microwave irradiations.     

Synthesis and Biological Activity of Certain 4-Substituted Imidazo[4,5-d]Pyridazine Nucleosides

Abstract

Purine analogues and derivatives exhibit a broad range of pharmacological activities and are used in the chemotherapy of cancer, parasitic and viral infections, and for the suppression of immune responses. Undoubtedly, this wide range of biological activities reflect an equally wide number of biochemical sites of action, one of which is the purine de novo pathway. New agents which can either serve as inhibitors of enzymes involved in this pathway or as substrates are continually sought. The unique series of nucleosides described herein should meet these desired needs.

The synthesis of 1involved glycosylation of a suitably 4,5-disubstituted imidazole and subsequent cyclization of the imidazole nucleoside so formed to the imidazo[4,5-d]pyridazine nucleoside. Such methodology was successfully employed^1,2 in the preparation of certain 4,7-disubstituted imidazo[4,5-d]pyridazine nucleosides. Chlorination of 1furnished 4-chloro-1-(2,3,5-tri-O-acetyl-β-D-ribo-furanosyl)imidazo[4,5-dlpyridazine (2) in 80% yield. This versatile intermediate can now serve as a precursor to a variety of 4-substituted imidazo[4,5-d]pyridazine nucleosides.     

Novel 4-amino-furo[2,3-d]pyrimidines as Tie-2 and VEGFR2 dual inhibitors

A novel class of furo[2,3-d]pyrimidines has been discovered as potent dual inhibitors of Tie-2 and VEGFR2 receptor tyrosine kinases (TK) and a diarylurea moiety at 5-position shows remarkably enhanced activity against both enzymes. One of the most active compounds, 4-amino-3-(4-((2-fluoro-5-(trifluoromethyl)phenyl)amino-carbonylamino)phenyl)-2-(4-methoxyphenyl)furo[2,3-d]pyrimidine (7k) is <3 nM on both TK receptors and the activity is rationalized based on the X-ray crystal structure.     

Pyrazolo[3,4-d]pyrimidines as adenosine antagonists

A large number of nitrogen heterocycles structurally related to caffeine and theophylline have been tested for activity as adenosine antagonists. Preliminary screening, utilizing displacement of [3H]N6-phenylisopropyladenosine (PIA) binding to rat brain membranes, identified several pyrazolo[3,4-d]pyrimidines with potential antagonist activity. These were then tested for their ability to antagonize adenosine-stimulated adenylate cyclase of guinea-pig slices and to block adenosine receptors which mediate presynaptic inhibition of transmitter release from cholinergic nerves in guinea-pig ileum. Of several compounds found to have antagonist activity, one of these, 4,6-bis-alpha- carbamoylethylthio -1-phenylpyrazolo[3,4-d]pyrimidine ( DJB -KK) was approximately an order of magnitude more potent than theophylline in both tests. GTP greatly reduces the potency of purine agonists, but not antagonists, as inhibitors of [3H] PIA binding; the potency of the pyrazolo[3,4-d]pyrimidine compounds was not altered by GTP. The compounds have no significant activity against [3H]adenosine uptake or on the binding of ligands to muscarinic cholinergic, beta-adrenergic, GABA or L-glutamate receptors.     

Chlorotrimethylsilane-promoted one-pot synthesis of steroidal[17,16-d]pyrimidines

A novel and practical procedure was developed for the preparation of steroidal[17,16-d]pyrimidines by chlorotrimethylsilane (TMSCl)-promoted one-pot multicomponent Biginelli-like condensations of steroid-17-ones, urea and aromatic aldehydes. First, treatment of the steroid-17-ones with urea and aromatic aldehydes in dimethylformamide (DMF)/acetonitrile (ACN) gives the corresponding Biginelli products, following the aromatising reaction of the Biginelli products at the same time under air to yield the desired steroidal[17,16-d]pyrimidines (78-88%). Since steroidal[17,16-d]pyrimidines with hydroxyl group can be subsequently converted into steroidal[17,16-d]pyrimidine derivatives, this general method provides a highly efficient route to these biologically important compounds.     

Rational design of 4-amino-5,6-diaryl-furo[2,3-d]pyrimidines as potent glycogen synthase kinase-3 inhibitors

4-Amino-5,6-diaryl-furo[2,3-d]pyrimidines have been identified as inhibitors of glycogen synthase kinase-3beta (GSK-3beta). One representative derivative, 4-amino-3-(4-(benzenesulfonylamino)-phenyl)-2-(3-pyridyl)-furo[2,3-d]pyrimidine (12) exhibited potent GSK-3beta inhibitory activity in low nanomolar level of IC(50). The binding mode was proposed from a docking study.     

Synthesis of 7-Glycopyranosyl-5-oxq-pyrrolo[2,3-d]Pyrimidine and 4-Glyco Pyranosylamino-Furo[2,3-d]Pyrimidine Derivatives

Abstract

The synthesis of some glycosyl derivatives of furo[2,3-d] and pyrrolo[2,3-d]pyrimidines is described.     

Synthesis and evaluation of a classical 2,4-diamino-5-substituted-furo[2,3-d]pyrimidine and a 2-amino-4-oxo-6-substituted-pyrrolo[2,3-d]pyrimidine as antifolates

Two classical antifolates, a 2,4-diamino-5-substituted furo[2,3-d]pyrimidine and a 2-amino-4-oxo-6-substituted pyrrolo[2,3-d]pyrimidine, were synthesized as potential inhibitors of dihydrofolate reductase (DHFR) and thymidylate synthase (TS). The syntheses were accomplished by condensation of 2,6-diamino-3(H)-4-oxo-pyrimidine with alpha-chloro-ketone 21 to afford two key intermediates 23 and 24, followed by hydrolysis, coupling with l-glutamate diethyl ester and saponification of the diethyl ester to afford the classical antifolates 13 and 14. Compounds 13 and 14 with a single carbon atom bridge are both substrates for folylpoly-gamma-glutamate synthetase (FPGS), the enzyme responsible for forming critical poly-gamma-glutamate antifolate metabolites with increased potency and/or increased cell retention. Compound 14 is a highly efficient FPGS substrate demonstrating that 2,4-diamino-5-substituted furo[2,3-d]pyrimidines are important lead structures for the design of antifolates with FPGS substrate activity. It retains inhibitory potency for DHFR and TS compared to the two atom bridged analog 5. Compound 13 is a poor inhibitor of purified DHFR and TS, and both 13 and 14 are poor inhibitors of the growth of CCRF-CEM human leukemia cells in culture, indicating that single carbon bridged compounds in these series though conducive to FPGS substrate activity were not potent inhibitors.     

Synthesis of 4-substituted imidazo[4,5-d][1,2,3]triazine (2-azapurine)nucleosides

Several methods for functionalization of the 4-position of imidazo[4,5-d][1,2,3]triazin-4-one were investigated. These investigations were successful and led to the preparation of 4-amino, 4-triazol-1-yl, 4-methoxy, 4-methylthio, 4-methylamino, 4-thio, 4-nitrobenzyl, and 4-unsubstituted 9-(beta-D-ribofuranosyl)-imidazo-[4,5-d][1,2,3]triazine (2-azapurine ribosides). The 4-unsubstituted compound (19) was slightly active against HCMV in plaque and yield reduction experiments and was not cytotoxic at 100 microM. The methylamino (15), hydrazino (16), and p-nitrobenzylthio (20) were inactive against HCMV but slightly cytotoxic. The thiomethyl-substituted analog (21) was the most active with activity comparable to ganciclovir but with greater cytotoxicity. We conclude that even though none of the tested compounds had antiviral activity superior to ganciclovir, the new synthetic methods will provide a route to more interesting compounds.     

Synthesis, antiplatelet and antithrombotic activities of new 2-substituted benzopyrano[4,3-d]pyrimidin-4-cycloamines and 4-amino/cycloamino-benzopyrano[4,3-d]pyrimidin-5-ones

Atherothrombotic coronary artery disease, associated with deep vein thrombosis, is one of the most common causes of death worldwide. Recently, antiplatelet combination therapy using agents with different mechanisms of action, such as aspirin, dipyridamole, and thienopyridines, seems to be an attractive preventive approach. Moreover, several large, randomized clinical trials support combination therapy with aspirin plus warfarin in high-risk patients with atherosclerotic heart disease. Our research on the benzopyrano[4,3-d]pyrimidine system gave rise to the synthesis of a large number of compounds endowed with in vitro anti-aggregating activity. Several SAR considerations suggest that the benzopyranopyrimidine system is an appropriate scaffold to obtain molecules that are able to act simultaneously in different pathways of aggregation. Now, we report the synthesis of new 2-substituted benzopyrano[4,3-d]pyrimidin-4-cycloamines and 4-amino/cycloamino-benzopyrano[4,3-d]pyrimidin-5-ones and the results of the pharmacological study on haemostasis. Some tested compounds showed a large-spectrum antiplatelet activity in vitro, and are more potent than aspirin as antithrombotics in vivo but, at variance with aspirin, they do not increase bleeding. This paper describes novel antithrombotic compounds with an interesting pharmacological profile and a potentially attractive benefit/risk ratio, with their mechanism of action generally, but not exclusively, dependent on antiplatelet activity, deserving further investigations.     

Synthesis and biological evaluation of novel steroidal[17,16-d][1,2,4]triazolo[1,5-a]pyrimidines

The preparation of steroidal[17,16-d][1,2,4]triazolo[1,5-a]pyrimidines and their biological evaluation as potential anticancer agents are herein reported. These novel heterosteroids (2, 4) were prepared through the condensation reaction of 3-amino-1,2,4-triazole with 16-arylidene-17-ketosteroids (1, 3). All the synthesized compounds were evaluated for their anticancer activity in vitro against PC-3 (human prostatic carcinoma), MCF-7 (human breast carcinoma) and EC9706 (human esophageal carcinoma) cell lines. Among the screened compounds, 2i, 2n and 4f showed significant inhibitory activity against all the three human cell lines.     

Antiangiogenic and antitumor agents. Design, synthesis, and evaluation of novel 2-amino-4-(3-bromoanilino)-6-benzylsubstituted pyrrolo[2,3-d]pyrimidines as inhibitors of receptor tyrosine kinases

Several different classes of growth factor receptors containing tyrosine kinases (RTK) are directly or indirectly involved in angiogenesis. Inhibition of these RTKs has provided a new paradigm in the treatment of tumors by restricting their growth and metastasis. We have designed, synthesized and evaluated eleven novel 2-amino-4-(3-bromoanilino)-6-substituted benzyl pyrrolo[2,3-d]pyrimidines as the first in a series of RTK inhibitors. These analogues were synthesized from appropriate alpha-bromomethylbenzyl ketones by cyclocondensation with 2,6-diamino-4-pyrimidone to afford the 2-amino-4-oxo-6-substituted benzyl pyrrolo[2,3-d]pyrimidines. Chlorination of the 4-position followed by displacement with 3-bromoaniline afforded the target compounds. In some instances, the 2-amino moiety of the pyrrolo[2,3-d]pyrimidines was protected prior to the chlorination and displacement followed by deprotection. The compounds were evaluated as inhibitors of vascular endothelial growth factor receptors VEGFR-2 (Flk-1, KDR) and VEGFR-1 (Flt-1); epidermal growth factor receptor (EGFR); and platelet-derived growth factor receptor-beta (PDGFR-beta). Selected compounds were also evaluated against the growth of A431 cells (which overexpress EGFR) in culture and as inhibitors of angiogenesis in the chicken embryo chorioallantonic membrane (CAM) assay. In each evaluation, a known standard compound was used as a comparison. Of the 11 analogues, five were more potent or equipotent as compared to standard compounds against the growth factor receptors. Two analogues showed superior inhibition of A431 cells in culture compared to the standard compounds. Three analogues were equipotent with the standard compound in the CAM assay and four of the analogues were dual inhibitors of RTKs. The structure-activity relationship for inhibition of different RTKs was quite distinct and different, and for VEGFR-2 and EGFR diametrically opposite. The inhibitory data against the RTKs in this study demonstrates that variation of the substituent(s) in the benzyl ring of these 2-amino-4-anilino 6-benzyl pyrrolo[2,3-d]pyrimidines does indeed control both the potency and specificity of inhibitory activity against RTKs.     

Synthesis of some novel pyrazolo[3,4-d]pyrimidine derivatives as potential antimicrobial agents

The reaction of 4-hydrazino-8-(trifluoromethyl)quinoline (2) with ethoxymethylenecyanoacetate afforded ethyl 5-amino-1-[8-(trifluoromethyl)quinolin-4-yl]-1H-pyrazole-4-carboxylate (3) and that with ethoxymethylenemalononitrile afforded 5-amino-1-[8-(trifluoromethyl)quinolin-4-yl]-1H-pyrazole-4-carbonitrile (5). Compounds 3 and 5 were hydrolyzed to get 5-amino-1-[8-(trifluoromethyl)quinolin-4-yl]-1H-pyrazole-4-carboxylic acid and then reacted with acetic anhydride to afford 6-methyl-1-[8-(trifluoromethyl)quinolin-4-yl]pyrazolo[3,4-d]oxazin-4-one (6), which was condensed with different aromatic amines to give a series of 5-substituted 6-methyl-1-[8-(trifluoromethyl)quinolin-4-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidin-4-ones (7). Compounds 3 and 5 also reacted with formamide, urea, and thiourea affording the corresponding pyrazolo[3,4-d]pyrimidines (8-13), respectively. Structures of the products have been determined by chemical reactions and spectral studies. All compounds of the series have been screened for their antibacterial and antifungal activity studies. The results are summarized in Tables 1 and 2.     

Discovery and in vitro evaluation of potent kinase inhibitors: Pyrido[1',2':1,5]pyrazolo[3,4-d]pyrimidines

The discovery, synthesis, potential binding mode, and in vitro kinase profile of several pyrido[1',2':1,5]pyrazolo[3,4-d]pyrimidines as potent kinase inhibitors are discussed.     

Pyrrolo[2,3-d]pyrimidines containing an extended 5-substituent as potent and selective inhibitors of lck II

Pyrrolo[2,3-d]pyrimidines containing a 5-(4-phenoxyphenyl) substituent are novel, potent and selective inhibitors of lck in vitro. Exploration of C-6 position of the pyrrolo[2,3-d]pyrimidine and the terminal phenyl group structure-activity relationship (SAR) is detailed. Compound 1 is orally active in animal models.     

Pyrazolo[4,3-d]pyrimidines as new generation of cyclin-dependent kinase inhibitors

A search among analogues of anti-CDK purines led to the identification of substituted pyrazolo[4,3-d]pyrimidines as novel inhibitors of CDK1/cyclin B. Some of these derivatives also show antiproliferative activity on cancer cell line K-562, thus may find an application as anticancer agents.     

Discovery of 4-anilino-N-methylthieno[3,2-d]pyrimidines and 4-anilino-N-methylthieno[2,3-d]pyrimidines as potent apoptosis inducers

We report the discovery of N-((benzo[d][1,3]dioxol-5-yl)methyl)-6-phenylthieno[3,2-d]pyrimidin-4-amine (2a) as an apoptosis inducer using our proprietary cell- and caspase-based ASAP HTS assay, and SAR study of HTS hit 2a which led to the discovery of 4-anilino-N-methylthieno[3,2-d]pyrimidines and 4-anilino-N-methylthieno[2,3-d]pyrimidines as potent apoptosis inducers. Compounds 5d and 5e were the most potent with EC₅₀ values of 0.008 and 0.004 μM in T47D human breast cancer cells, respectively. Compound 5d was found to be highly active in the MX-1 breast cancer model. Functionally, compounds 5d and 5e both induced apoptosis through inhibition of tubulin polymerization.     

Synthesis and biological evaluation of 6,7-disubstituted 4-aminopyrido[2,3-d]pyrimidines as adenosine kinase inhibitors

The synthesis and structure-activity relationship of a series of 6,7-disubstituted 4-aminopyrido[2,3-d]pyrimidines as novel non-nucleoside adenosine kinase inhibitors is described. A variety of substituents, primarily aryl, at the C6 and C7 positions of the pyridopyrimidine core were found to yield analogues that are potent inhibitors of adenosine kinase. In contrast to the 5,7-disubstituted and 5,6,7-trisubstituted pyridopyrimidine series, these analogues exhibited only modest potency to inhibit AK in intact cells.