Fluorinated indazoles as novel selective inhibitors of nitric oxide synthase (NOS): Synthesis and biological evaluation

In order to find new compounds with neuroprotective activity and NOS-I/NOS-II selectivity, we have designed, synthesized, and characterized 14 new NOS inhibitors with an indazole structure. The first group corresponds to 4,5,6,7-tetrahydroindazoles (4–8), the second to the N-methyl derivatives (9–12) of 7-nitro-1H-indazole (1) and 3-bromo-7-nitro-1H-indazole (2), and the latter to 4,5,6,7-tetrafluoroindazoles (13–17). Compound 13 (4,5,6,7-tetrafluoro-3-methyl-1H-indazole) inhibited NOS-I by 63% and NOS-II by 83%. Interestingly, compound 16 (4,5,6,7-tetrafluoro-3-perfluorophenyl-1H-indazole) inhibited NOS-II activity by 80%, but it did not affect to NOS-I activity. Structural comparison between these new indazoles further supports the importance of the aromatic indazole skeleton for NOS inhibition and indicate that bulky groups or N-methylation of 1 and 2 diminish their effect on NOS activity. The fluorination of the aromatic ring increased the inhibitory potency and NOS-II selectivity, suggesting that this is a promising strategy for NOS selective inhibitors.     

Biological properties and structural study of new aminoalkyl derivatives of benzimidazole and benzotriazole,dual inhibitors of CK2 and PIM1 kinases

The new aminoalkyl-substituted derivatives of known CK2 inhibitors 4,5,6,7-tetrabromo-1H-benzimidazole (TBBi) and 4,5,6,7-tetrabromo-1H-benzotriazole (TBBt) were synthesized, and their influence on the activity of recombinant human CK2 α, CK2 holoenzyme and PIM1 kinases was evaluated. All derivatives inhibited the activity of studied kinases and the most efficient were aminopropyl-derivatives 8b and 14b. These compounds also exerted inhibition of cancer cell lines – CCRF-CEM (acute lymphoblastoid leukemia), MCF-7 (human breast cancer), and PC-3 (prostate cancer) proliferation and their EC₅₀ is comparable with the value for clinically studied CK2 inhibitor CX-4945. Preliminary structure activity relationship analysis indicated that the spacer length affected antitumor potency, and two to three methylene units were more favorable. The complex of CK2 α^1-335/8b was crystallized, both under high-salt conditions and under low-salt conditions giving crystals which diffracted X-rays to about 2.4 Å resolution, what enabled the determination of the corresponding 3D-structures.     

Discovery and preliminary structure–activity relationship of 1H-indazoles with promising indoleamine-2,3-dioxygenase 1 (IDO1) inhibition properties

Indoleamine 2,3-dioxygenase 1 (IDO1)-mediated kynurenine pathway of tryptophan degradation is identified as an important immune effector pathway in the tumor cells to escape a potentially effective immune response. IDO1 is an attractive target for anticancer therapy and the discovery of IDO1 inhibitors has been intensely ongoing in both academic research laboratories and pharmaceutical organizations. Our study discovered that 1H-indazole was a novel key pharmacophore with potent IDO1 inhibitory activity. A series of new 1H-indazole derivatives were synthesized and determined the enzyme inhibitory activities, and the compound 2g exhibited the highest activity with an IC₅₀ value of 5.3 μM. The structure–activity relationships (SARs) analysis of the 1H-indazole derivatives as novel IDO1 inhibitors indicated that the 1H-indazole scaffold is necessary for IDO1 inhibition, and the substituent groups at the both 4-position and 6-position largely affect inhibitory activity. The docking model exhibited that the effective interactions of 1H-indazoles with ferrous ion of heme and key residues of hydrophobic Pocket A and B ensured the IDO1 inhibitory activities. The study suggested that the 1H-indazole was a novel interesting scaffold for IDO inhibition for further development.     

Synthesis and molecular modeling of some novel hexahydroindazole derivatives as potent monoamine oxidase inhibitors

A novel series of 2-thiocarbamoyl-2,3,4,5,6,7-hexahydro-1H-indazole and 2-substituted thiocarbamoyl-3,3a,4,5,6,7-hexahydro-2H-indazoles derivatives were synthesized and investigated for the ability to inhibit the activity of the A and B isoforms of monoamine oxidase (MAO). The target molecules were identified on the basis of satisfactory analytical and spectra data (IR, ¹H NMR, ¹³C NMR, ²D NMR, DEPT, EI-MASS techniques and elemental analysis). Synthesized compounds showed high activity against both the MAO-A (compounds 1d, 1e, 2c, 2d, 2e) and the MAO-B (compounds 1a, 1b, 1c, 2a, 2b) isoforms. In the discussion of the results, the influence of the structure on the biological activity of the prepared compounds was delineated. It was suggested that non-substituted and N-methyl/ethyl bearing compounds (except 2c) increased the inhibitory effect and selectivity toward MAO-B. The rest of the compounds, carrying N-allyl and N-phenyl, appeared to select the MAO-A isoform. The inhibition profile was found to be competitive and reversible for all compounds. A series of experimentally tested (1a–2e) compounds was docked computationally to the active site of the MAO-A and MAO-B isoenzyme. The autodock 4.01 program was employed to perform automated molecular docking. In order to see the detailed interactions of the docked poses of the model inhibitors compounds 1a, 1d, 1e and 2e were chosen because of their ability to reversibly inhibit the MAO-B and MAO-A and the availability of experimental inhibition data. The differences in the intermolecular hydrophobic and H-bonding of ligands to the active site of each MAO isoform were correlated to their biological data. Observation of the docked positions of these ligands revealed interactions with many residues previously reported to have an effect on the inhibition of the enzyme. Excellent to good correlations between the calculated and experimental K_i values were obtained. In the docking of the MAO-A complex, the trans configuration of compound 1e made various very close interactions with the residues lining the active site cavity these interactions were much better than those of the other compounds tested in this study. This tight binding observation may be responsible for the nanomolar inhibition of form of MAOA. However, it binds slightly weaker (experimental K_i = 1.23 μM) to MAO-B than to MAO-A (experimental K_i = 4.22 nM).     

Preparation of new C-nucleosides by intramolecular dehydration of 2-pentahydroxypentyl-4,5,6,7-tetrahydroindol-4-ones

Acid-catalysed dehydration of the polyhydroxyalkyl chain of 6,6-dimethyl-2-(d-gluco-pentitol-l-yl)-4,5,6,7-tetrahydroindol-4-one and of 6,6-dimethyl-2-(d-manno-pentitol-l-yl)-4,5,6,7-tetrahydroindol-4-one gave 2-α-d-arabinofuranosyl-6,6-dimethyl-4,5,6,7-tetrahydroindol-4-one (3). In a similar way, 2-β-d-lyxopyranosyl-6,6-dimethyl-4,5,6,7-tetrahydroindol-4-one (8) and 2-β-d-lyxopyranosyl-4,5,6,7-tetrahydroindol-4-one (9) were obtained by dehydration of 6,6-dimethyl-2-(d-galacto-pentitol-l-yl)-4,5,6,7-tetrahydroindol-4-one and 2-(d-galacto-pentitol-l-yl)-4,5,6,7-tetrahydroindol-4-one, respectively. The structures of the new C-nucleosides described (3, 8, and 9) were elucidated by chemical and physical methods.     

Development of novel tetrahydrothieno[2,3-c]pyridine-3-carboxamide based Mycobacterium tuberculosis pantothenate synthetase inhibitors: Molecular hybridization from known antimycobacterial leads

Twenty six 2,6-disubstituted 4,5,6,7-tetrahydrothieno[2,3-c]pyridine-3-carboxamide derivatives were designed by molecular hybridization approach using and synthesized from piperidin-4-one by five step synthesis. Compounds were evaluated for Mycobacterium tuberculosis (MTB) pantothenate synthetase (PS) inhibition study, in vitro activities against MTB, cytotoxicity against RAW 264.7 cell line. Among the compounds, 6-(4-nitrophenylsulfonyl)-2-(5-nitrothiophene-2-carboxamido)-4,5,6,7-tetrahydrothieno[2,3-c]pyridine-3-carboxamide (11) was found to be the most active compound with IC₅₀ of 5.87 ± 0.12 μM against MTB PS, inhibited MTB with MIC of 9.28 μM and it was non-cytotoxic at 50 μM. The binding affinity of the most potent inhibitor 11 was further confirmed biophysically through differential scanning fluorimetry.     

Studies on substituted benzo[h]quinazolines,benzo[g]indazoles,pyrazoles, 2,6-diarylpyridines as anti-tubercular agents

Various substituted 5,6-dihydro-8-methoxybenzo[h]quinazolin-2-amine, 1-(3-(4-alkoxyphenyl)-7-methoxy-3,3a,4,5-tetrahydro-2H benzo[g]indazol-2-yl)ethanone, pyrazole and 2,6-diarylpyridine derivatives have been synthesized in good yields by an efficient methodology. The synthesized compounds (4–23) were evaluated for their in vitro anti-tubercular activity against Mycobacterium tuberculosis H37Rv strain. Compounds 6a, 6c, 8a, 19a and 19e exhibited significant anti-tubercular activity at MIC values 50, 100, 50, 25 and 100 μM concentration. In vitro cytotoxicity data using THP-1 cells indicated that most active compound 19a is safe as its MIC value is much lower than the cytotoxic value.     

Synthesis of novel 2-amino-4-(5′-substituted 2′-phenyl-1H-indol-3′-yl)-6-aryl-4H-pyran-3-carbonitrile derivatives as antimicrobial and antioxidant agents

As a part of systematic investigation of synthesis and biological activities of indole analogues linked to various heterocyclic systems, we have synthesized new compounds viz., 2-amino-4-(5′-substituted 2′-phenyl-1H-indol-3′-yl)-6-aryl-4H-pyran-3-carbonitriles (2a–i), 4,5-diamino-6-(5′-substituted 2′-phenyl-1H-indol-3′-yl)-8-aryl-2-oxo-2,6-dihydrodipyrano [2,3-b:3,2-e]pyridine-3-carbonitriles (3a–i), 4-amino-5-(5′-substituted 2′-phenyl-1H-indol-3-yl)-7-aryl-1H-pyrano[2,3-d]pyrimidin-2(5H)-ones (4a–i), 4-amino-5-(5′-substituted 2′-phenyl-1H-indol-3′-yl)-7-aryl-1H-pyrano[2,3-d]pyrimidin-2(5H)-thiones (5a–i), 4-(5′-subtituted 2′-phenyl-1H-indol-3′-yl)-6-aryl-1,4-dihydropyrano[2,3-c]pyrazol-3-amines (6a–i) and 5-(5′-substituted 2′-phenyl-1H-indol-3′-yl)-7-aryl-3H-pyrano[2,3-d]pyrimidin-4(5H)-ones (7a–i). Antibacterial activity results revealed that, compound 6a showed promising activity versus Escherichia coli, Staphylococcus aureus and Klebsiella pneumoniae. Compound 6d exhibited good activity against S. aureus, K. pneumoniae and Pseudomonas aeruginosa. Antifungal activity results indicated that, compound 4d exhibited maximum zone of inhibition against Aspergillus oryzae and Aspergillus flavus. In case of antioxidant activity, compound 4a showed promising radical scavenging activity, ferric ions (Fe³⁺) reducing antioxidant power (FRAP) and metal chelating activity.     

Amine-free melanin-concentrating hormone receptor 1 antagonists: Novel non-basic 1-(2H-indazole-5-yl)pyridin-2(1H)-one derivatives and mitigation of mutagenicity in Ames test

To develop non-basic melanin-concentrating hormone receptor 1 (MCHR1) antagonists with a high probability of target selectivity and therapeutic window, we explored neutral bicyclic motifs that could replace the previously reported imidazo[1,2-a]pyridine or 1H-benzimidazole motif. The results indicated that the binding affinity of a chemically neutral 2H-indazole derivative 8a with MCHR1 (hMCHR1: IC₅₀ = 35 nM) was comparable to that of the imidazopyridine and benzimidazole derivatives (1 and 2, respectively) reported so far. However, 8a was positive in the Ames test using TA1537 in S9− condition. Based on a putative intercalation of 8a with DNA, we introduced a sterically-hindering cyclopropyl group on the indazole ring to decrease planarity, which led to the discovery of 1-(2-cyclopropyl-3-methyl-2H-indazol-5-yl)-4-{[5-(trifluoromethyl)thiophen-3-yl]methoxy}pyridin-2(1H)-one 8l without mutagenicity in TA1537. Compound 8l exerted significant antiobesity effects in diet-induced obese F344 rats and exhibited promising safety profile.     

Reaction of 2-amino-2-deoxyheptoses with cyclic β-dicarbonyl compounds

The reaction between 2-amino-2-deoxyaldoses and β-dicarbonyl compounds yields polyhydroxyalkylpyrroles. Thus, 6,6-dimethyl-2-(D-galacto-pentitol-1-yl)-4,5,6,7-tetrahydroindol-4-one (4a), 6,6-dimethyl-2-(D-gluco-pentitol-1-yl)-4,5,6,7-tetrahydroindol-4-one (4b), and 6,6-dimethyl-2-(D-manno-pentitol-1-yl)-4,5,6,7-tetrahydroindol-4-one (4c) have been obtained from 5,5-dimethylcyclohexane-1,3-dione (2) and 2-amino-2-deoxyheptoses having D-glycero-L-gluco (1a), D-glycero-D-ido (1b), and D-glycero-D-talo (1c) configurations, respectively. 2-Amino-2-deoxy-D-glycero-L-manno-heptose (1d), the epimer of 1a, also reacts with 2, to yield 4a. In a similar way, 1a, 1b, and 1c react with cyclohexane-1,3-dione (3), to give 2-(D-galacto-pentitol-1-yl)-4,5,6,7-tetrahydroindol-4-one (5a), 2-D-gluco-pentitol-1-yl)-4,5,6,7-tetrahydroindol-4-one (5b), and 2-(D-manno-pentitol-1-yl)-4,5,6,7-tetrahydroindol-4-one (5c), respectively.     

Facile synthesis of SSR180575 and discovery of 7-chloro-N,N,5-trimethyl-4-oxo-3(6-[18F]fluoropyridin-2-yl)-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide,a potent pyridazinoindole ligand for PET imaging of TSPO in cancer

A novel synthesis of the translocator protein (TSPO) ligand 7-chloro-N,N,5-trimethyl-4-oxo-3-phenyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide (SSR180575, 3) was achieved in four steps from commercially available starting materials. Focused structure–activity relationship development about the pyridazinoindole ring at the N3 position led to the discovery of 7-chloro-N,N,5-trimethyl-4-oxo-3(6-fluoropyridin-2-yl)-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide (14), a novel ligand of comparable affinity. Radiolabeling with fluorine-18 (¹⁸F) yielded 7-chloro-N,N,5-trimethyl-4-oxo-3(6-[¹⁸F]fluoropyridin-2-yl)-3,5-dihydro-4H-pyridazino[4,5-b]indole-1-acetamide ([¹⁸F]-14) in high radiochemical yield and specific activity. In vivo studies of [¹⁸F]-14 revealed this agent as a promising probe for molecular imaging of glioma.     

Synthesis of pyrazole encompassing 2-pyridone derivatives as antibacterial agents

A series of novel compounds 6-amino-1-((1,3-diphenyl-1H-pyrazole-4-yl)methyleneamino)-4-(aryl)-2-oxo-1,2-dihydropyridine-3,5-dicarbonitriles (4a–t) were synthesized and characterized by IR, ¹H NMR, ¹³C NMR and mass spectral data. These compounds were screened for their in vitro antibacterial activity against Staphylococcus aureus, Streptococcus pyogenes (Gram positive), Escherichia coli, Pseudomonas aeruginosa (Gram negative) by serial broth dilution and cytotoxic activity (NIH 3T3 & HeLa) by MTT assay. The results indicated that compounds 4g, 4i, 4m, 4o, 4r and 4t exhibit potent antibacterial activity against bacterial strains at non-cytotoxic concentrations.     

Synthesis and structures of aryloxo- and binaphthoxogermanium(IV) alkyl iodide complexes

The germanium(II) aryloxide complexes (S)-[Ge{O₂C₂₀H₁₀-(SiMe₂Ph)₂-3,3′}{NH₃}] (1) and [Ge(OC₆H₃Ph₂-2,6)₂] (2) react with either Bu^tI or MeI to yield the corresponding germanium(IV) compounds (S)-[Ge{O₂C₂₀H₁₀-(SiMe₂Ph)₂-3,3′}{Bu^t}{I}] (3), (S)-[Ge{O₂C₂₀H₁₀-(SiMe₂Ph)₂-3,3′}{Me}{I}] (4), [Ge(OC₆H₃Ph₂-2,6)₂(Bu^t)(I)] (5), and [Ge(OC₆H₃Ph₂-2,6)₂(Me)(I)] (6). Compound 6 reacts with 2,6-diphenylphenol to yield [Ge(OC₆H₃Ph₂-2,6)₃(Me)] (7), while 3-5 do not. The X-ray crystal structures of 3-5 and 7 were determined, and 3-5 represent the first structurally characterized germanium(IV) species having germanium bound to both oxygen and iodine.     

Novel pyrimidine-pyridine hybrids: Synthesis,cyclooxygenase inhibition,anti-inflammatory activity and ulcerogenic liability

Some derivatives containing pyrido[2,3-d:6,5d′]dipyrimidine-4,5-diones (9a-f), tetrahydropyrido[2,3-d]pyrimidine-6-carbonitriles (11a-c) and 6-(4-acetylphenyl)-2-thioxo-2,3,5,6,7,8-hexahydro-1H-pyrimido[4,5-d]pyrimidin-4-one (12) were synthesized from 6-amino-2-thioxo-2,3-dihydro-1H-pyrimidin-4-one (8). The anti-inflammatory effect of these candidates was determined and the ulcer indices were calculated for active compounds. 7-Amino-5-(3,4,5-trimethoxyphenyl)-4-oxo-2-thioxo-1,2,3,4-tetrahydropyrido[2,3-d] pyrimidine-6-carbonitrile (11c) exhibited better edema inhibition than celecoxib. Moreover, compounds 9b, 9d and 11c revealed better COX-2 inhibitory activity in a range (IC₅₀ = 0.25–0.89 µM) than celecoxib (IC₅₀ = 1.11 µM). Regarding ulcerogenic liability, all of the compounds under the study were less ulcerogenic than indomethacin. Molecular docking studies had been carried on active candidates 9d and 11c to explore action mode of these candidates as leads for discovering other anti-inflammatory agents.     

Short hybrid peptides incorporating β- and γ-amino acids as antimicrobial agents

The peptides containing β- and γ-amino acids, LA-Lys(Z)-PEA, P1; LA-Lys(Z)-β^3,3-Ac₆c-PEA, P2; LA-Orn(Z)-β^3,3-Ac₆c-PEA, P3; LA-Lys(Z)-Gpn-PEA, P4; LA-Orn(Z)-Gpn-PEA, P5; LA-Lys(Z)-γ⁴-Phe-PEA, P6, LA-γ⁴-Leu-Lys(Z)-PEA, P7 and LA-β^3,3-Pip(Ac)-Lys(Z)-PEA, P8 were synthesized, characterized and evaluated against Gram-positive and Gram-negative bacteria. Among all, peptides P2, P3, P4 and P5 exhibited potent activity (MIC 6.25 μM) against S. aureus MTCC 737 and P. aeruginosa MTCC 424. In order to understand the efficacy of peptides and mechanism of action, time kill kinetics and fluorescence microscopic studies were performed against S. aureus and P. aeruginosa for the peptides P2, P3, P4 and P5. P4 took half time to show the bactericidal effect on P. aeruginosa and S. aureus in comparison to P2 at their 2x MICs. Fluorescence microscopic studies suggested that peptides P2 and P4 both killed the bacteria via membrane disruption. Further, P4 exhibited lowest haemolytic activity among active peptides and negligible cytotoxic activity against human cancer cell lines A-549, PC-3 and HCT-116 at its MIC.     

Synthesis of indazole based diarylurea derivatives and their antiproliferative activity against tumor cell lines

New series of indazole based diarylureas were synthesized and their anticancer activity against cancer cells H460, A549, OS-RC-2, HT-29, Lovo, HepG2, Bel-7402, SGC-7901 and MDA-MB-231 were examined. These derivatives of diarylureas, except azaindazole based diarylureas 5f, 5l and 5m, showed superior or similar activity against most of these selected cancer cell lines to the reference compound sorafenib. The effect of substituents on the indazole ring was also investigated. Derivatives with trifluoromenthy or halogen substituent on the indazole ring showed higher activity against the selected cancer cell lines than sorafenib. The acute toxicity assay showed that compounds 5a, 5b and 5i possessed lower toxicity than sorafenib. Compound 5i with 4-(trifluoromenthy)-1H-indazole and 4-(trifluoromenthy) benzene moieties exhibited the most potent anticancer activity.     

2-Styrylindolium based fluorescent probes visualize neurofibrillary tangles in Alzheimer’s disease

We evaluated 2-styrylindolium derivatives (6–11) as novel and selective probes for neurofibrillary tangles (NFTs) on brain sections of AD patients. The staining experiments indicated that these compounds may bind selectively to NFTs in the presence of ß-amyloid (Aß) plaques. Cell free binding assays confirmed that 2-[2-[4-(1-pyrrolidinyl)phenyl]ethenyl]-1,3,3-trimethyl-3H-indolium iodide (9) and 2-[2-[4-(diethylamino)phenyl]ethenyl]-1-butyl-3,3-dimethyl-3H-indolium iodide (11) display excellent affinities to Tau-aggregates (IC₅₀ values of 5.1 and 1.4 nM, respectively) in the displacement of Thiazin Red R. These probes have good solubility in distilled water and low or no cytotoxicity in zebrafish embryo and liver hepatocellular carcinoma cell assays.     

Synthesis and activity of substituted heteroaromatics as positive allosteric modulators for α4β2α5 nicotinic acetylcholine receptors

The design and synthesis of a series of substituted heteroaromatic α4β2α5 positive allosteric modulators is reported. The optimization and development of the heteroaromatic series was carried out from NS9283, and several potent analogues, such as 3-(5-(pyridin-3-yl)-2H-tetrazol-2-yl)benzonitrile (5k) and 3,3′-(2H-tetrazole-2,5-diyl)dipyridine (12h) with good in vitro efficacy were discovered.     

Synthesis of carbon-11-labeled 5-HT6R antagonists as new candidate PET radioligands for imaging of Alzheimer’s disease

Carbon-11-labeled serotonin (5-hydroxytryptamine) 6 receptor (5-HT₆R) antagonists, 1-[(2-bromophenyl)sulfonyl]-5-[¹¹C]methoxy-3-[(4-methyl-1-piperazinyl)methyl]-1H-indole (O-[¹¹C]2a) and 1-[(2-bromophenyl)sulfonyl]-5-methoxy-3-[(4-[¹¹C]methyl-1-piperazinyl)methyl]-1H-indole (N-[¹¹C]2a), 5-[¹¹C]methoxy-3-((4-methylpiperazin-1-yl)methyl)-1-(phenylsulfonyl)-1H-indole (O-[¹¹C]2b) and 5-methoxy-3-((4-[¹¹C]methylpiperazin-1-yl)methyl)-1-(phenylsulfonyl)-1H-indole (N-[¹¹C]2b), 1-((4-isopropylphenyl)sulfonyl)-5-[¹¹C]methoxy-3-((4-methylpiperazin-1-yl)methyl)-1H-indole (O-[¹¹C]2c) and 1-((4-isopropylphenyl)sulfonyl)-5-methoxy-3-((4-[¹¹C]methylpiperazin-1-yl)methyl)-1H-indole (N-[¹¹C]2c), 1-((4-fluorophenyl)sulfonyl)-5-[¹¹C]methoxy-3-((4-methylpiperazin-1-yl)methyl)-1H-indole (O-[¹¹C]2d) and 1-((4-fluorophenyl)sulfonyl)-5-methoxy-3-((4-[¹¹C]methylpiperazin-1-yl)methyl)-1H-indole (N-[¹¹C]2d), were prepared from their O- or N-desmethylated precursors with [¹¹C]CH₃OTf through O- or N-[¹¹C]methylation and isolated by HPLC combined with SPE in 40–50% radiochemical yield, based on [¹¹C]CO₂ and decay corrected to end of bombardment (EOB). The radiochemical purity was >99%, and the molar activity (MA) at EOB was 370–740?GBq/μmol with a total synthesis time of ～40-min from EOB.     

The alkenic unreactivity of mono- and bi-cyclic derivatives of 3,6-dihydro-2-(methylthio)-2H-thiopyran S,S,S′,S′-tetraoxide

The inertness of the alkenic bond towards electrophilic additions in 3-exocyano-3-(methylthio)-2-thiabicyclo[2.2.1]hept-5-ene S,S,S′,S′-tetraoxide (5), 3,6-dihydro-2-(methylthio)-2H-thiopyran-2-carbonitrile S,S,S′,S′-tetraoxide (3), and 2-(acetamidomethyl)-3,6-dihydro-2-(methylthio)-2H-thiopyran S,S,S′,S′-tetraoxide (4) is attributed to the “supra-annular effect” and field effects. Conformational analysis of a pentadeuterated derivative of 4 (10) is reported. On the basis of the 220-MHz ¹H n.m.r.-spectral data of 10, the compound was concluded to adopt the ⁰H₂ conformation in chloroform solution.