Isosteric Replacement Research Articles

Benzothiazepine CGP37157 and Its Isosteric 2′-Methyl Analogue Provide Neuroprotection and Block Cell Calcium Entry

Benzothiazepine CGP37157 is widely used as tool to explore the role of mitochondria in cell Ca(2+) handling, by its blocking effect of the mitochondria Na(+)/Ca(2+) exchanger. Recently, CGP37157 has shown to exhibit neuroprotective properties. In the trend to improve its neuroprotection profile, we have synthesized ITH12505, an isosteric analogue having a methyl instead of chlorine at C2' of the phenyl ring. ITH12505 has exerted neuroprotective properties similar to CGP37157 in chromaffin cells and hippocampal slices stressed with veratridine. Also, both compounds afforded neuroprotection in hippocampal slices stressed with glutamate. However, while ITH12505 elicited protection in SH-SY5Y cells stressed with oligomycin A/rotenone, CGP37157 was ineffective. In hippocampal slices subjected to oxygen/glucose deprivation plus reoxygenation, ITH12505 offered protection at 3-30 μM, while CGP37157 only protected at 30 μM. Both compounds caused blockade of Ca(2+) channels in high K(+)-depolarized SH-SY5Y cells. An in vitro experiment for assaying central nervous system penetration (PAMPA-BBB; parallel artificial membrane permeability assay for blood-brain barrier) revealed that both compounds could cross the blood-brain barrier, thus reaching their biological targets in the central nervous system. In conclusion, by causing a mild isosteric replacement in the benzothiazepine CGP37157, we have obtained ITH12505, with improved neuroprotective properties. These findings may inspire the design and synthesis of new benzothiazepines targeting mitochondrial Na(+)/Ca(2+) exchanger and L-type voltage-dependent Ca(2+) channels, having antioxidant properties.

Sulfonyl-morpholino-pyrimidines: SAR and development of a novel class of selective mTOR kinase inhibitor

High throughput screening to identify inhibitors of the mTOR kinase revealed sulfonyl-morpholino-pyrimidine 1 as an attractive start point. The compound displayed good physicochemical properties and selectivity over related kinases such as PI3Kα. Library preparation of related analogs allowed the establishment of additional SAR understanding and in particular the requirement for a key hydrogen bond donor motif at the 4-position of the phenyl ring in compounds such as indole 19. Isosteric replacement of the indole functionality led to the identification of urea compounds such as 32 that show good levels of mTOR inhibition in both enzyme and cellular assays.

Synthesis and alpha 1-adrenoceptor antagonistic activity of some 4-amino-5,7-dimethyl-2-(substituted) aminopyrido (2,3-d) pyrimidines.

A series of new 4-amino-5,7-dimethyl-2- (substituted)aminopyrido(2,3-d)pyrimidines (5) have been synthesized and tested for selective alpha 1-adrenoreceptor antagonistic activity. Some of the compounds were found to antagonize alpha 1-adrenoreceptor in a competitive and reversible manner. When screened on rat anococcygeus muscle some of the compounds exhibited significant alpha 1-adrenoreceptor antagonistic activity (pA2 values in the range of 5.2-7.8). The most potent compound (5j) was evaluated by an in vivo method and was found to reduce the systolic and diastolic blood pressure of spontaneously hypertensive rats. The percentage reduction in blood pressure by test compound 5j was found to be higher than that of the standard drug prazosin (CAS 19216-56-9) at the same dose level (1 mg/kg p.o.). Chemically, prazosin is a 4-aminoquinazolin derivative. Pyridopyrimidine is a known bioisostere of quinazoline. The study revealed that isosteric replacement of the benzene ring of prazosin by a pyridine ring increases the potency.

A new class of non-thiazolidinedione, non-carboxylic-acid-based highly selective peroxisome proliferator-activated receptor (PPAR) γ agonists: Design and synthesis of benzylpyrazole acylsulfonamides

Herein, we describe the design, synthesis, and structure–activity relationships of novel benzylpyrazole acylsulfonamides as non-thiazolidinedione (TZD), non-carboxylic-acid-based peroxisome proliferator-activated receptor (PPAR) γ agonists. Docking model analysis of in-house weak agonist 2 bound to the reported PPARγ ligand binding domain suggested that modification of the carboxylic acid of 2 would help strengthen the interaction of 2 with the TZD pocket and afford non-carboxylic-acid-based agonists. In this study, we used an acylsulfonamide group as the ring-opening analog of TZD as an isosteric replacement of carboxylic acid moiety of 2; further, preliminary modification of the terminal alkyl chain on the sulfonyl group gave the lead compound 3c. Subsequent optimization of the resulting compound gave the potent agonists 25c, 30b, and 30c with high metabolic stability and significant antidiabetic activity. Further, we have described the difference in binding mode of the carboxylic-acid-based agonist 1 and acylsulfonamide 3d.

Cyclic Hydroxyamidines as Amide Isosteres: Discovery of Oxadiazolines and Oxadiazines as Potent and Highly Efficacious γ-Secretase Modulators in Vivo

Cyclic hydroxyamidines were designed and validated as isosteric replacements of the amide functionality. Compounds with these structural motifs were found to be metabolically stable and to possess highly desirable pharmacokinetic profiles. These designs were applied in the identification of γ-secretase modulators leading to highly efficacious agents for reduction of central nervous system Aβ(42) in various animal models.

Selenosemicarbazones as potent cruzipain inhibitors and their antiparasitic properties against Trypanosoma cruzi

The cysteine protease cruzipain is an essential T. cruzienzyme and one of the few validated drug targets for Chagas disease. Thiosemicarbazones have been described as cruzipain inhibitors. While searching for new antichagasic drugs, we synthesized a series of selenosemicarbazone analogs and demonstrated that the isosteric replacement of the sulfur atom with selenium resulted in an enhancement of the cysteine protease inhibitory effect. Three selenosemicarbazones were characterized enzymatically and proved to be reversible, slow-binding inhibitors for the Z-Phe-Arg-AMC substrate. Their KI values were in the low nM range (3.7 to 29.7 nM), suggesting a strong interaction with the enzyme, approaching a tight binding definition. All selenosemicarbazones tested showed better activities against epimastigotes than Benznidazole, the currently used drug, with IC50 values ranging from 1.2 to 5.9 μM (Bnz IC50 = 12.5 μM). Three of these compounds showed a better Selectivity Index (SI) than Benznidazole. These compounds also displayed activity against the infective intracellular amastigote form at low micromolar range. Overall, our results support the role of these novel organoselenium compounds as promising lead candidates for further drug development studies.

The design, synthesis and biological evaluation of novel URB602 analogues as potential monoacylglycerol lipase inhibitors

We have synthesised an extensive series of URB602 analogues as inhibitors of monoacylglycerol lipase (MAGL), which is the major enzyme responsible for metabolising the endocannabinoid 2-arachidonylglycerol. The recently identified crystal structure of MAGL was used in the design strategy and revealed three possible binding sites for URB602 and the proposed analogues. A test series of carbamate analogues were docked into the identified sites to predict the most favourable binding location. The synthesised analogues of URB602 explored the biological effects of isosteric replacement, ring size and substitution, para substitution of the biphenyl moiety and the incorporation of a bicyclic element. The compounds were tested for their ability to inhibit human MAGL. The carbamate analogue 16 displayed the most significant inhibitory activity, reducing MAGL activity to 26% of controls at 100 μM compared to 73% for the parent compound URB602.

Importance of Imidazolidinone Motif in 4-Phenyl-N-arylsulfonylimidazolidinone for their Anticancer Activity

To investigate the possible isosteric replacement of imidazolidinone moiety in 4-phenyl-N-arylsulfonylimidazolidinone for broad and potent anticancer agents, a series of 4-phenyl-l(N)-arylsulfonylimidazolidinones 6a-k, imidazolidinethione analogs 7a-i, and imidazolidine oxime analogs 8a-c were prepared and evaluated for their in vitro anticancer activity against four human cancer cell lines (human lung A549, human colon COLO205, human leukemia K562, human ovary SK-OV-3). Among all the derivatives of N-arylsulfonylimidazolidinone 6a-k, compounds 6f and 6g showed the best inhibition comparable to doxorubicin against all cancer cell lines. Increasing the carbon chain on alkyl moieties of carbamates as shown in 6c-g did not alter the activity. The imidazolidinethione analogs 7a-i and imidazolidin-2-one oxime derivatives 8a-c did not possess any good activity. Therefore, imidazolidinone moiety is the best pharmacophore among the 4-phenyl-Narylsulfonylimidazolidinone derivatives.

(E)-Alkene and Ethylene Isosteres Substantially Alter the Hydrogen-Bonding Network in Class II MHC Aq/Glycopeptide Complexes and Affect T-Cell Recognition

The structural basis for antigen presentation by class II major histocompatibility complex (MHC) proteins to CD4(+) T-cells is important for understanding and possibly treating autoimmune diseases. In the work described in this paper, (E)-alkene and ethylene amide-bond isosteres were used to investigate the effect of removing hydrogen-bonding possibilities from the CII259-270 glycopeptide, which is bound by the arthritis-associated murine A(q) class II MHC protein. The isostere-modified glycopeptides showed varying and unexpectedly large losses of A(q) binding that could be linked to the dynamics of the system. Molecular dynamics (MD) simulations revealed that the backbone of CII259-270 and the A(q) protein are able to form up to 11 hydrogen bonds, but fewer than this number are present at any one time. Most of the strong hydrogen-bond interactions were formed by the N-terminal part of the glycopeptide, i.e., in the region where the isosteric replacements were made. The structural dynamics also revealed that hydrogen bonds were strongly coupled to each other; the loss of one hydrogen-bond interaction had a profound effect on the entire hydrogen-bonding network. The A(q) binding data revealed that an ethylene isostere glycopeptide unexpectedly bound more strongly to A(q) than the corresponding (E)-alkene, which is in contrast to the trend observed for the other isosteres. Analysis of the MD trajectories revealed that the complex conformation of this ethylene isostere was structurally different and had an altered molecular interaction pattern compared to the other A(q)/glycopeptide complexes. The introduced amide-bond isosteres also affected the interactions of the glycopeptide/A(q) complexes with T-cell receptors. The dynamic variation of the patterns and strengths of the hydrogen-bond interactions in the class II MHC system is of critical importance for the class II MHC/peptide/TCR signaling system.

Revisiting the Mechanism of the Triosephosphate Isomerase Reaction: The Role of the Fully Conserved Glutamic Acid 97 Residue

An analysis of 503 available triosephosphate isomerase sequences revealed nine fully conserved residues. Of these, four residues-K12, H95, E97 and E165-are capable of proton transfer and are all arrayed around the dihydroxyacetone phosphate substrate in the three-dimensional structure. Specific roles have been assigned to the residues K12, H95 and E165, but the nature of the involvement of E97 has not been established. Kinetic and structural characterization is reported for the E97Q and E97D mutants of Plasmodium falciparum triosephosphate isomerase (Pf TIM). A 4000-fold reduction in k(cat) is observed for E97Q, whereas the E97D mutant shows a 100-fold reduction. The control mutant, E165A, which lacks the key catalytic base, shows an approximately 9000-fold drop in activity. The integrity of the overall fold and stability of the dimeric structure have been demonstrated by biophysical studies. Crystal structures of E97Q and E97D mutants have been determined at 2.0 Å resolution. In the case of the isosteric replacement of glutamic acid by glutamine in the E97Q mutant a large conformational change for the critical K12 side chain is observed, corresponding to a trans-to-gauche transition about the Cγ-Cδ (χ(3)) bond. In the E97D mutant, the K12 side chain maintains the wild-type orientation, but the hydrogen bond between K12 and D97 is lost. The results are interpreted as a direct role for E97 in the catalytic proton transfer cycle. The proposed mechanism eliminates the need to invoke the formation of the energetically unfavourable imidazolate anion at H95, a key feature of the classical mechanism.

Cheminformatics and computational chemistry in lead optimisation

Lead Optimisation is the process by which a chemical lead series is improved with regard to multiple properties. These properties will vary from project to project but typically include affinity at the target protein, ADME properties and certain toxicological end-points. Techniques from computational chemistry and cheminformatics have a key role to play in many aspects of lead optimisation. A number of methodologies in use at GSK to assist our lead optimisation projects will be described in this talk. The integration of data from multiple data sources is an important starting point, leading to the use of novel computational tools for the analysis and visualisation of structure-activity relationships. The large number of lead optimisation programmes run at GSK over many years has resulted in many thousands of data points. A key question is, what knowledge can be extracted from such large data sets that can help in future optimisation efforts. Here we will summarise some of our work to derive medicinal chemistry rules of thumb and to probe for possible isosteric replacements. Finally, the use of experimental design techniques in array chemistry will be discussed.

Impact of the Nature of the Substituent at the 3-Position of 4H-1,2,4-Benzothiadiazine 1,1-Dioxides on Their Opening Activity toward ATP-Sensitive Potassium Channels

The synthesis of diversely substituted 3-isopropoxy-, 3-isopropylsulfanyl-, 3-isopropylsulfinyl-, and 3-isobutyl-4H-1,2,4-benzothiadiazine 1,1-dioxides is described. Their activity on pancreatic β-cells (inhibitory effect on the insulin releasing process) and on vascular and uterine smooth muscle tissues (myorelaxant effects) was compared to that of previously reported K(ATP) channel openers belonging to 3-isopropylamino-4H-1,2,4-benzothiadiazine 1,1-dioxides. The present study aimed at evaluating the impact on biological activity of the isosteric replacement of the NH group of 3-alkylamino-4H-1,2,4-benzothiadiazine 1,1-dioxides by a O, S, S(═O), or CH(2) group. By comparing compounds bearing identical substituents, the following rank order of potency on pancreatic β-cells was observed: 3-isopropylamino > 3-isobutyl > 3-isopropoxy > 3-isopropylsulfanyl > 3-isopropylsulfinyl-substituted 4H-1,2,4-benzothiadiazine 1,1-dioxides (NH > CH(2) > O > S > S(═O)). A molecular modeling study revealed that 3-isopropoxy-, 3-isopropylsulfanyl-, and 3-isopropylamino-substituted compounds adopted a similar low-energy conformation (preferred orientation of the isopropyl chain). Moreover, no direct relationship was detected between the conformational freedom of the different classes of benzothiadiazines (from the most to the lowest conformationally constrained compounds: NH > O > S > CH(2)) and their biological activity on insulin-secreting cells. Therefore, the present study confirmed the critical role of the NH group at the 3-position for the establishment of a strong hydrogen bond responsible for optimal activity expressed by 3-alkylamino-4H-1,2,4-benzothiadiazine 1,1-dioxides on insulin-secreting cells. Radioisotopic and fluorimetric experiments conducted with 7-chloro-3-isopropoxy-4H-1,2,4-benzothiadiazine 1,1-dioxide 10c demonstrated that such a compound, bearing a short branched O-alkyl group instead of the NH-alkyl group at the 3-position, also behaved as a specific K(ATP) channel opener. Lastly, the present work further identified 3-(alkyl/aralkyl)sulfanyl-substituted 7-chloro-4H-1,2,4-benzothiadiazine 1,1-dioxides as a class of promising myorelaxant drugs acting on uterine smooth muscles, at least in part, through the activation of K(ATP) channels.

Synthesis of novel ceramide-like penetration enhancers

Ceramide-like penetration enhancers have been synthesized by isosteric replacements of the 1-hydroxymethyl and amide moieties of natural ceramides. Metabolically more stable oxime and five-membered heterocyclic (1,2,4-oxadiazole, 1,2,4-triazole, tetrazole) analogues are more polar and possess H-bond acceptor properties, which make them potentially useful in optimizing key intermolecular features in interactions between the modified ceramide structures and the lipid layers of stratum corneum.

Strategies and Chemical Design Approaches to Reduce the Potential for Formation of Reactive Metabolic Species

Metabolic activation of new chemical entities to reactive intermediates is routinely monitored in drug discovery and development. Reactive intermediates may bind to cellular macromolecules such as proteins, DNA and may eventually lead to cell death via necrosis, apoptosis or oxidative stress. The evidence that the ultimate outcome of metabolic activation is an adverse drug reaction manifested as in vivo toxicity, is at best circumstantial. However, understanding the process of bioactivation of structural alerts by trapping the reactive intermediates is critical to guide medicinal chemistry efforts in quest for safer and potent molecules. This commentary provides a brief introduction to adverse drug reactions and mechanisms of reactive intermediate formation for various functional groups, followed by a review of chemical design approaches, examples of such strategies, possible isosteric replacements for structural alerts and rationalization of laboratory approaches to determine reactive intermediates, as a guide to today's medicinal chemist.

Anthranilic acid-based inhibitors of phosphodiesterase: Design, synthesis, and bioactive evaluation

Our previous studies identified two 2-benzoylaminobenzoate derivatives 1, which potently inhibited superoxide (O(2)˙(-)) generation induced by formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP) in human neutrophils. In an attempt to improve their activities, a series of anthranilic acid derivatives were synthesized and their anti-inflammatory effects and underlying mechanisms were investigated in human neutrophils. Of these, compounds 17, 18, 46, 49, and 50 showed the most potent inhibitory effect on FMLP-induced release of O(2)˙(-) in human neutrophils with IC(50) values of 0.20, 0.16, 0.15, 0.06, and 0.29 μM, respectively. SAR analysis showed that the activities of most compounds were dependent on the ester chain length in the A ring. Conversely, a change in the linker between the A and B ring from amide to sulfonamide or N-methyl amide, as well as exchanges in the benzene rings (A or B rings) by isosteric replacements were unfavorable. Further studies indicated that inhibition of O(2)˙(-) production in human neutrophils by these anthranilic acids was associated with an elevation in cellular cAMP levels through the selective inhibition of phosphodiesterase 4. Compound 49 could be approved as a lead for the development of new drugs in the treatment of neutrophilic inflammatory diseases.

Effect of the isosteric replacement of the phenyl motif with furyl (or thienyl) of 4-phenyl-N-arylsulfonylimidazolones as broad and potent anticancer agents

To investigate the effect of isosteric replacement of phenyl with heteroaromatics of 4-phenyl-N-arylsulfonylimidazolones (1–4) as broad and potent anticancer agents, a series of 4-furyl (5a–e, 7a–c) or thienyl-N-arylsulfonylimidazolones (6a–e) were designed and synthesized to evaluate their in vitro anticancer activity against HCT116, A549 and NCI-H460 cancer cell lines. Replacement of phenyl with furyl or thienyl slightly reduced the activity, however, these analogs showed comparable activity to doxorubicin. The thienyl analogs exhibited more potent activity than furyl analogs due to their more isosteric similarity with the phenyl moiety. In addition, the carbamate derivatives (5a–e and 6a–e) of both the analogs had better activities than the carbamoyl derivatives (7a–c). Therefore, the size and planarity of the aryl motif at 4-position of N-arylsulfonylimidazolones are crucial for their anticancer activity.

Isosteric replacement of the Z-enone with haloethyl ketone and E-enone in a resorcylic acid lactone series and biological evaluation

The synthesis of a small library of resorcylic acid lactones and evaluation of their biological properties as kinase inhibitors is described. Within the series E-enones were found more active than corresponding Z-enones as inhibitors of a subset of kinases containing a conserved cysteine. Replacement of the enone moiety with a β-haloketone group led to compounds with an interesting kinase selectivity profile and also antiproliferative activity against Jurkat cells. An E-enone derivative also showed activity against capillary tube formation based on a co-culture of primary human umbilical cord endothelial cells (HUVECs) and vascular smooth muscle cells (vSMCs).

Identification of a novel pyrrole derivative endowed with antimycobacterial activity and protection index comparable to that of the current antitubercular drugs streptomycin and rifampin

A hit optimization procedure based on isosteric and bioisosteric replacement of decorating groups at both the N1 and the C5 phenyl rings of 1,5-diarylpyrroles led to identification of 4-((1-(4-fluorophenyl)-2-methyl-5-(4-(methylthio)phenyl)-1 H-pyrrol-3-yl)methyl)thiomorpholine that is characterized by a very high activity toward both Mycobacterium tuberculosis 103471 and H37Rv strains (MIC values of 0.125 μg/mL), and a safe profile in terms of cytotoxicity (CC 50 of >128 μg/mL) and protection index (>1000). Antitubercular activity and protection index of the new compound are comparable to those found for the current antitubercular drugs streptomycin and rifampin.

Docking of the alkaloid geissospermine into acetylcholinesterase: a natural scaffold targeting the treatment of Alzheimer’s disease

Pharmacological studies from our group [Lima et al. Pharmacol Biochem Behav 92:508, (2009)] revealed that geissospermine (GSP), the major alkaloid of the bark extract of Brazilian Geissospermum vellosii, inhibits acetylcholinesterases (AChEs) in the brains of rats and electric eels (Electrophorus electricus). However, the binding mode (i.e., conformation and orientation) of this indole-indoline alkaloid into the AChE active site is unknown. Therefore, in order to propose a plausible binding mode between GSP and AChE, which might explain the observed experimental inhibitory activity, we performed comparative automatic molecular docking simulations using the AutoDock and Molegro Virtual Docker (MVD) programs. A sample of ten crystal structures of the Pacific electric ray (Torpedo californica) TcAChE, in complex with ten diverse active site ligands, was selected as a robust re-docking validation test, and also for GSP docking. The MVD results indicate a preferential binding mode between GSP and AChE, in which GSP functional groups may perform specific interactions with residues in the enzyme active site, according to the ligand-protein contacts detected by the LPC/CSU server. Four hydrogen bonds were detected between GSP and Tyr121, Ser122, Ser200, and His440, in which the last two residues belong to the catalytic triad (Ser200···His440···Glu327). Hydrophobic and π-π stacking interactions were also detected between GSP and Phe330 and Trp84, respectively; these are involved in substrate stabilization at the active site. This study provides the basis to propose structural changes to the GSP structure, such as molecular simplification and isosteric replacement, in order to aid the design of new potential AChE inhibitors that are relevant to the treatment of Alzheimer's disease.

Synthesis and evaluation of amides surrogates of dopamine D3 receptor ligands

Isosteric replacement of the amide function and modulation of the arylpiperazine moiety of known dopamine D3 receptor ligands led to potent and selective compounds. Enhanced bioavailability and preferential brain distribution make compound 6c a good candidate for pharmacological and clinical evaluation.