Antiviral potential of natural compounds against influenza virus hemagglutinin

Influenza virus of different subtypes H1N1, H2N2, H3N2 and H5N1 cause many human pandemic deaths and threatening the people worldwide. The Hemagglutinin (HA) protein mediates viral attachment to host receptors act as an attractive target. The sixteen natural compounds have been chosen to target the HA protein. Molecular docking studies have been performed to find binding affinity of the compounds. Out of the sixteen, three compounds CI, CII and CIII found to posses a higher binding affinity. The molecular dynamics (MD) simulation has been performed to study the structural, dynamical properties for the nine different complexes CI, CII, CIII bound with H1, H2, H3 proteins and the results were compared. The molecular mechanics Poission-Boltzmann surface area (MM-PBSA) method is used to compare the binding free energy, its different energy components and per residue binding contribution. The H1 subtype shows higher binding preference for all the curcumin derivatives than H2 and H3. The binding capability of protein subtypes with curcumin derivatives and the binding affinity of curcumin compounds are in the order H1 > H2 > H3 and CI > CII > CIII respectively. The two -O-CH3- groups present in the CI compound help to have strong binding with HA protein than CII and CIII. The van der Waals interaction energy plays a significant role for binding in all the complexes. The hydrogen bonding interactions were monitored throughout the MD simulation. The conserved region (153–155) and the helix region (193–194) of H1, H2, H3 protein subtypes are found to possess higher binding susceptibility for binding of the curcumin derivatives.     

Mass spectrometric sequencing and acylation character analysis of C-terminal anchoring segment from Influenza A hemagglutinin

Influenza A virus hemagglutinin (HA) is a major envelope glycoprotein mediating viral and cell membrane fusion. HA is anchored in the viral envelope by a light HA(2) chain containing one transmembrane domain and a cytoplasmic tail. Three cysteine residues in the C-terminal region, one in the transmembrane domain and two in the cytoplasmic tail, are highly conserved and potentially palmitoylated in all HA subtypes. The HA(2) C- terminal anchoring segments were extracted to organic phase from the bromelain-digested viruses (subviral particles) of three strains: A/X-31 (H3 subtype), A/Puerto Rico/8/34 (H1 subtype) and A/FPV/Weybridge/34 (H7 subtype). Their primary structures were assessed by matrix-assisted laser desorption/ionization time-of-flight time-of- flight mass spectrometry (MALDI-ToF-ToF MS). Trypsin-type protease-cleaved peptides prevailed over bromelain- cleaved ones in the peptide mixtures. All of them included transmembrane domains. Several distinctive features of the C-terminal HA(2) peptides acylation character were discovered by MALDI-ToF MS: 1) the peptides isolated from the viruses, which were digested by bromelain in the absence of beta-mercaptoethanol, were predominantly triply acylated; 2) the peptides were acylated not only by palmitic, but also by stearic acid residues; 3) the palmitate/stearate ratio was different for the three strains studied; 4) the A/FPV/Weybridge/34 strain has a priority to stearate binding. This fatty acid residue was discovered at the first of three conservative cysteine residues located in the transmembrane domain. It was found that presence of thiol reagent during preparation of subviral particles led to the appearence of the C-terminal HA(2) peptides acylated to different degrees. Triply, doubly, mono- and even unacylated peptides were detected. It was demonstrated that the thioester bond in the isolated acylpeptides was extremely sensitive to thiol reagents.     

Synthesis and Antiviral Evaluation of Nucleoside Analogues Bearing One Pyrimidine Moiety and Two D-Ribofuranosyl Residues

A series of 1,2,3-triazolyl nucleoside analogues in which 1,2,3-triazol-4-yl-β-d-ribofuranosyl fragments are attached via polymethylene linkers to both nitrogen atoms of the heterocycle moiety (uracil, 6-methyluracil, thymine, quinazoline-2,4-dione, alloxazine) or to the C-5 and N-3 atoms of the 6-methyluracil moiety was synthesized. All compounds synthesized were evaluated for antiviral activity against influenza virus A/PR/8/34/(H1N1) and coxsackievirus B3. Antiviral assays revealed three compounds, 2i, 5i, 11c, which showed moderate activity against influenza virus A H1N1 with IC₅₀ values of 57.5 µM, 24.3 µM, and 29.2 µM, respectively. In the first two nucleoside analogues, 1,2,3-triazol-4-yl-β-d-ribofuranosyl fragments are attached via butylene linkers to N-1 and N-3 atoms of the heterocycle moiety (6-methyluracil and alloxazine, respectively). In nucleoside analogue 11c, two 1,2,3-triazol-4-yl-2′,3′,5′-tri-O-acetyl-β-d-ribofuranose fragments are attached via propylene linkers to the C-5 and N-3 atoms of the 6-methyluracil moiety. Almost all synthesized 1,2,3-triazolyl nucleoside analogues showed no antiviral activity against the coxsackie B3 virus. Two exceptions are 1,2,3-triazolyl nucleoside analogs 2f and 5f, in which 1,2,3-triazol-4-yl-2′,3′,5′-tri-O-acetyl-β-d-ribofuranose fragments are attached to the C-5 and N-3 atoms of the heterocycle moiety (6-methyluracil and alloxazine respectively). These compounds exhibited high antiviral potency against the coxsackie B3 virus with IC₅₀ values of 12.4 and 11.3 µM, respectively, although both were inactive against influenza virus A H1N1. According to theoretical calculations, the antiviral activity of the 1,2,3-triazolyl nucleoside analogues 2i, 5i, and 11c against the H1N1 (A/PR/8/34) influenza virus can be explained by their influence on the functioning of the polymerase acidic protein (PA) of RNA-dependent RNA polymerase (RdRp). As to the antiviral activity of nucleoside analogs 2f and 5f against coxsackievirus B3, it can be explained by their interaction with the coat proteins VP1 and VP2.     

Overcoming the Lack of Oral Availability of Cyclic Hexapeptides: Design of a Selective and Orally Available Ligand for the Integrin αvβ3

A highly systematic approach for the development of both orally bioavailable and bioactive cyclic N‐methylated hexapeptides as high affinity ligands for the integrin αvβ3 is based on two concepts: a) screening of systematically designed libraries with spatial diversity and b) masking of the peptide charge with a lipophilic protecting group. The key steps of the method are 1) initial design of a combinatorial library of N‐methylated analogues of the stem peptide cyclo(d ‐Ala‐Ala₅); 2) selection of cyclic peptides with the highest intestinal permeability; 3) design of sublibraries with the bioactive RGD sequence in all possible positions; 4) selection of the best ligands for RGD‐recognizing integrin subtypes; 5) fine‐tuning of the affinity and selectivity by additional Ala to Xaa substitutions; 6) protection of the charged functional groups according to the prodrug concept to regain intestinal and oral permeability; 7) proof of biological effects in mice after oral administration.     

Exploring the Effect of Bioisosteric Replacement of Carboxamide by a Sulfonamide Moiety on N‐Glycosidic Torsions and Molecular Assembly: Synthesis and X‐ray Crystallographic Investigation of N‐(β‐D‐Glycosyl)sulfonamides as N‐Glycoprotein Linkage Region Analogues

N‐Glycoprotein linkage region constituents, 2‐acetamido‐2‐deoxy‐β‐D ‐glucopyranose (GlcNAc) and asparagine (Asn) are conserved among all the eukaryotes. To gain a better understanding for nature’s choice of GlcNAcβAsn as linkage region constituents and inter‐ and intramolecular carbohydrate–protein interactions, a detailed systemic structural study of the linkage region conformation is essential. Earlier crystallographic studies of several N‐(β‐glycopyranosyl)alkanamides showed that N‐glycosidic torsion, ?_N, is influenced to a larger extent by structural variation in the sugar part than that of the aglycon moiety. To explore the effect of the bioisosteric replacement of a carboxamide group by a sulfonamide moiety on the N‐glycosidic torsions as well as on molecular assembly, several glycosyl methanesulfonamides and glycosyl chloromethanesulfonamides were synthesized as analogues of the N‐glycoprotein linkage region, and crystal structures of seven of these compounds have been solved. A comparative analysis of this series of crystal structures as well as with those of the corresponding alkanamido derivatives revealed that N‐glycosidic torsion, ?_N, does not alter significantly. Methanesulfonamido and chloromethanesulfonamido derivatives of GlcNAc display a different aglycon conformation compared to other sulfonamido analogues. This may be due to the cumulative effect of the direct hydrogen bonding between N1 and O1′ and C? H???O interactions of the aglycon chain, revealing the uniqueness of the GlcNAc as the linkage sugar.     

Environmental Effects on a Prion's Helix II Domain: Copper(II) and Membrane Interactions with PrP180–193 and Its Analogues

An abnormal interaction between copper and the prion protein is believed to play a pivotal role in the pathogenesis of prion diseases. Copper binding has been mainly attributed to the N‐terminal domain of the prion protein, but this hypothesis has recently been challenged in some papers which suggest that the C‐terminal domain might also compete for metal anchoring. In particular, the segment corresponding to the helix II region of the prion protein, namely PrP180–193, has been shown both to bind copper and to exhibit a copper‐enhanced cytotoxicity, as well as to interact with artificial membranes. The present work is aimed at extending these results by choosing the most representative model of this domain and by determining its copper affinity. With this aim, the different role played by the electrostatic properties of the C‐ and N‐termini of PrP180–193 (VNITIKQHTVTTTT) in determining its conformational behaviour, copper coordination and ability to perturb model membranes was investigated. Owing to the low solubility of PrP180–193, its copper affinity was evaluated by using the shorter PrPAc184–188NH₂ (IKQHT) analogue as a model. ESI‐MS, ESR, UV/Vis, and CD measurements were carried out on the copper(II )/PrPAc184–188NH₂ and copper(II )/PrP180–193NH₂ systems, and showed that PrPAc184–188NH₂ is a reliable model for the metal interaction with the helix II domain. The affinity of copper(II ) for the helix II fragment is higher than that for the octarepeat and PrP106–126 peptides. Finally, the different ability of PrP180–193 analogues to perturb the DPPC model membrane was assessed by DSC measurements. The possible biological consequences of these findings are also discussed briefly.     

Chemoenzymatic Syntheses of Sialylated Oligosaccharides Containing C5‐Modified Neuraminic Acids for Dual Inhibition of Hemagglutinins and Neuraminidases

A fast chemoenzymatic synthesis of sialylated oligosaccharides containing C5‐modified neuraminic acids is reported. Analogues of GM₃ and GM₂ ganglioside saccharidic portions where the acetyl group of NeuNAc has been replaced by a phenylacetyl (PhAc) or a propanoyl (Prop) moiety have been efficiently prepared with metabolically engineered E. coli bacteria. GM₃ analogues were either obtained by chemoselective modification of biosynthetic N‐acetyl‐sialyllactoside (GM₃NAc) or by direct bacterial synthesis using C5‐modified neuraminic acid precursors. The latter strategy proved to be very versatile as it led to an efficient synthesis of GM₂ analogues. These glycomimetics were assessed against hemagglutinins and sialidases. In particular, the GM₃NPhAc displayed a binding affinity for Maackia amurensis agglutinin (MAA) similar to that of GM₃NAc, while being resistant to hydrolysis by Vibrio cholerae (VC) neuraminidase. A preliminary study with influenza viruses also confirmed a selective inhibition of N1 neuraminidase by GM₃NPhAc, suggesting potential developments for the detection of flu viruses and for fighting them.     

Epitope Structure of the Carbohydrate Recognition Domain of Asialoglycoprotein Receptor to a Monoclonal Antibody Revealed by High-Resolution Proteolytic Excision Mass Spectrometry

Recent studies suggest that the H1 subunit of the carbohydrate recognition domain (H1CRD) of the asialoglycoprotein receptor is used as an entry site into hepatocytes by hepatitis A and B viruses and Marburg virus. Thus, molecules binding specifically to the CRD might exert inhibition towards these diseases by blocking the virus entry site. We report here the identification of the epitope structure of H1CRD to a monoclonal antibody by proteolytic epitope excision of the immune complex and high-resolution MALDI-FTICR mass spectrometry. As a prerequisite of the epitope determination, the primary structure of the H1CRD antigen was characterised by ESI-FTICR-MS of the intact protein and by LC-MS/MS of tryptic digest mixtures. Molecular mass determination and proteolytic fragments provided the identification of two intramolecular disulfide bridges (seven Cys residues), and a Cys-mercaptoethanol adduct formed by treatment with β-mercaptoethanol during protein extraction. The H1CRD antigen binds to the monoclonal antibody in both native and Cys-alkylated form. For identification of the epitope, the antibody was immobilized on N-hydroxysuccinimide (NHS)-activated Sepharose. Epitope excision and epitope extraction with trypsin and FTICR-MS of affinity-bound peptides provided the identification of two specific epitope peptides (5–16) and (17–23) that showed high affinity to the antibody. Affinity studies of the synthetic epitope peptides revealed independent binding of each peptide to the antibody.     

Molecular dynamics simulations of pro-apoptotic BH3 peptide helices in aqueous medium: relationship between helix stability and their binding affinities to the anti-apoptotic protein Bcl-X<Subscript>L</Subscript>

The B-cell lymphoma 2 (Bcl-2) family of proteins regulates the intrinsic pathway of apoptosis. Interactions between specific anti- and pro-apoptotic Bcl-2 proteins determine the fate of a cell. Anti-apoptotic Bcl-2 proteins have been shown to be over-expressed in certain cancers and they are attractive targets for developing anti-cancer drugs. Peptides from the BH3 region of pro-apoptotic proteins have been shown to interact with anti-apoptotic Bcl-2 proteins and induce biological activity similar to that observed in parent proteins. However, the specificity of BH3 peptides derived from different pro-apoptotic proteins differ for different anti-apoptotic Bcl-2 proteins. In this study, we have investigated the relationship between the stable helical nature of BH3 peptides and their affinities to Bcl-X_L, an anti-apoptotic Bcl-2 protein. We have carried out molecular dynamics simulations of six BH3 peptides derived from Bak, Bad and Bim pro-apoptotic proteins for a period of 50 ns each in aqueous medium. Due to the amphipathic nature of BH3 peptides, the hydrophobic residues on the hydrophobic face tend to cluster together in all BH3 peptides. While this process resulted in a complete loss of helical structure in 16-mer Bak and 16-mer Bad wild type peptides, stabilizing interactions in the hydrophilic face of the BH3 peptides and capping interactions helped to maintain partial helical character in 16-mer Bad mutant and 16-mer Bim peptides. The latter two 16-mer peptides exhibit higher affinity for Bcl-X_L. Similarly the longer BH3 peptides, 25-mer Bad and 33-mer Bim, also resulted in smaller and stable helical fragments and their helical conformation is stabilized by interactions between residues in the solvent-exposed hydrophilic half of the peptide. The stable nature of helical segment in a BH3 peptide can be directly correlated to its binding affinity and the helical region encompassed the highly conserved Leu residue. We propose that upon approaching the hydrophobic groove of anti-apoptotic proteins, a longer helix will be induced in high affinity BH3 peptides by extending the smaller stable helical segments around the conserved Leu residue in both N- and C-terminal regions. The results reported in this study will have implications in developing peptide-based inhibitors for anti-apoptotic Bcl-2 proteins.     

Halide‐Anion Binding by Singly and Doubly N‐Confused Porphyrins

The halide‐binding properties of N‐confused porphyrin (NCP, 1 ) and doubly N‐confused porphyrins (trans‐N₂CP ( 2 ), cis‐N₂CP ( 3 )) were examined in CH₂Cl₂. In the free‐base forms, cis‐N₂CP ( 3 ) showed the highest affinity to each anion (Cl^?, Br^?, I^?) with association constants K_a=7.8×10³, 1.9×10³, and 5.8×10² M ^?1, respectively. As metal complexes, on the other hand, trans‐N₂CP 2–Cu exhibited the highest affinity to Cl^?, Br^?, and I^? with K_a=9.0×10⁴, 2.7×10⁴, and 1.9×10³ M ^?1, respectively. The corresponding K_a values for cis‐N₂CP 3–Cu and NCP 1–Cu were about 1/10 and 1/2, respectively, of those of 2–Cu . With the help of density functional theory (DFT) calculations and complementary affinity measurements of a series of trisubstituted N‐confused porphyrins, the efficient anion binding of NCPs was attributed to strong hydrogen bonding at the highly polarized NH moieties owing to the electron‐deficient C₆F₅ groups at meso positions as well as the ideally oriented dipole moments and large molecular polarizability. The orientation and magnitude of the dipole moments in NCPs were suggested to be important factors in the differentiation of the affinity for anions.     

Diels-Alder adducts of 3-N-substituted derivatives of (−)-Cytisine as influenza A/H1N1 virus inhibitors; stereodifferentiation of antiviral properties and preliminary assessment of action mechanism

The synthesis and anti-influenza activity study of Diels-Alder adducts of 3-N-substituted derivatives of (−)-cytisine with N-substituted maleimides are described. Synthesized compounds were studied for antiviral activity against influenza virus A/California/07/09 (H1N1)pdm09 in MDCK. The values of CC₅₀, IC₅₀ and selectivity indexes (SI) of obtained derivatives were determined. It was shown that anti-influenza activity of ‘α-endo’ adducts is higher (SI of three samples is 79 and higher) than activity of ‘β-endo’ adducts. By means of ‘time-of-addition’ experiment it was established that the leading compound (3aS,4R,8S,12R,12aR,12bS)-10-benzyl-2-phenyloctahydro-1H-4,12a-etheno-8,12-methanopyrrolo[3′,4':3,4]pyrido[1,2-a][1,5]diazocine-1,3,5(4H)-trione (16a) demonstrates anti-influenza activity at the middle and late stages of the virus life cycle. The possibility of interaction of synthesized derivatives with the active sites of the PA_N and PB2 was estimated via in silico approach. The difference in the locations of ‘α-endo’ and ‘β-endo’ adducts in PB2 active site (5JUN) is offered as an explanation of the dependence of their virus-inhibiting properties on stereochemistry.     

Paradigms and paradoxes: why is the electron affinity of the azide radical, N3, so large?

The consensus value for the electron affinity of azide radical is 261 kJ mol⁻¹, anomalously higher than many species that are also made of highly electronegative elements. N₃ ⁻ has two equivalent resonance structures analogous to NO₂ ⁻ that has a lower electron affinity. Electronegativity trends rationalize why the electron affinity of N₃ is higher than that of P₃; however, those of N and N₂ are lower than those of P and P₂. We suggest the reason for the observed high electron affinity of azide radical is Coulombic stabilization in the ionic triplet resonance structure, ⁻N=N⁺=N⁻.     

Aladan scanning: The structure-activity relationship of dynorphin A

An unnatural amino acid, β-[6′-(N, N-dimethyl)amino-2′-naphthoyl]alanine (Ald) showing polarity-sen sitive fluorescence characteristics, was synthesized. A thorough Ald-scan of dynorphin A (Dyn A), the putative endogenous ligand for κ opioid receptors, was then performed. Replacement of the amino acid residues in positions 5, 8, 10, 12 or 14 of Dyn A(1-13)-NH2 with Ald resulted in compounds that had almost equal κ binding affinity compared with that of the parent compound; on the other hand, substi-tution o...     

99mTc-labeling and in vitro characterization of N4- and N3S-RGDS-derivative peptides

The aim of this work was to characterize the in vitro behavior of N4- and N3S-RGDS-derivative peptides labeled with ^99mTc. Peptides AGGG-Abu-GRGDSPK-NH₂ (F22) and C(acm)-GGG-Abu-GRGDSPK-NH₂ (SMA1) were synthesized by solid phase. The stability of ^99mTc-labeled peptides was assessed in a 30-fold molar excess of cysteine and in plasma. The affinity for plasma proteins was also evaluated. Labeling yield was >95% for both peptides. ^99mTc-F22 was not stable in presence of cysteine, but 63% of ^99mTc remained chelated to SMA1 up to 24 hours. Both peptides showed low affinity to plasma proteins. N3S-RGDS-derivative peptide (SMA1) showed more stable coordination binding with ^99mTc and a higher stability in plasma with regard to N4-RGDS-derivative peptide (F22).     

Interdigitated array microelectrode based impedance immunosensor for detection of avian influenza virus H5N1

Continuous outbreaks of avian influenza (AI) in recent years with increasing threat to animals and human health have warranted the urgent need for rapid detection of pathogenic AI viruses. In this study, an impedance immunosensor based on an interdigitated array (IDA) microelectrode was developed as a new application for sensitive, specific and rapid detection of avian influenza virus H5N1. Polyclonal antibodies against AI virus H5N1 surface antigen HA (Hemagglutinin) were oriented on the gold microelectrode surface through protein A. Target H5N1 viruses were then captured by the immobilized antibody, resulting in a change in the impedance of the IDA microelectrode surface. Red blood cells (RBCs) were used as biolabels for further amplification of the binding reaction of the antibody-antigen (virus). The binding of target AI H5N1 onto the antibody-modified IDA microelectrode surface was further confirmed by atomic force microscopy. The impedance immunosensor could detect the target AI H5N1 virus at a titer higher than 10³ EID₅₀/ml (EID₅₀: 50% Egg Infective Dose) within 2 h. The response of the antibody-antigen (virus) interaction was shown to be virus titer-dependent, and a linear range for the titer of H5N1 virus was found between 10³ and 10⁷ EID₅₀/ml. Equivalent circuit analysis indicated that the electron transfer resistance of the redox probe [Fe(CN)₆]^3−/4− and the double layer capacitance were responsible for the impedance change due to the protein A modification, antibody immobilization, BSA (bovine serum albumin) blocking, H5N1 viruses binding and RBCs amplification. No significant interference was observed from non-target RNA viruses such as Newcastle disease virus and Infectious Bronchitis disease virus. (The H5N1 used in the study was inactivated virus.)     

Synthesis of cyclic and acyclic Nα-methyl-Nω-alkyl-l-arginine analogues

A series of Nα-methyl-alkyl-l-arginine (Arg) analogues have been synthesized from inexpensive, commercially available starting materials. These analogues, once incorporated into pharmaceutically relevant peptides, are expected to increase binding affinity, receptor selectivity, lipophilicity, and stability as demonstrated with analogues of similar design and structure.     

Screening and Identification of Peptides that Bind Specifically to the X gene Promoter of Hepatitis B Virus Using a Combinatorial Peptide Library Approach

Hepatitis B virus (HBV) X protein (HBx) plays a key role in the development of hepatocellular carcinoma (HCC) in HBV carriers. A drug that can bind to the promoter region of HBV may shut down the expression of HBx and subsequently prevent the development of HCC in the HBV carrier. We have constructed a seven amino acid residue peptide library on a TentaGel resin using a combinatorial one‐bead one‐sequence peptide synthesis method. The fluorescently labeled eicosanucleotide (5′‐(6‐FAM) CTTTTGGGCT TTGCTGCCCC‐3′) of the HBx promoter region was used as a monitor to screen for peptides that have high binding affinity to the HBx promoter. Two heptapeptides, KAPLFSI and SRVRMTW, were identified, and synthesized. The binding affinities of the peptides to the HBx promoter oligonucleotide were determined using Surface Plasmon Resonance (SPR). The peptide KAPLFSI had a greater binding affinity constant (k_a) and equilibrium constant (K_D) than SRVRMTW. The k_a and K_D values with the full X‐promoter sequence were found to be 1.425 E+5 (1/Ms) and 1.186 E‐8 (M), respectively. The peptide may open a new route for tumor suppression in HBV carriers.     

Identification of Novel Influenza Polymerase PB2 Inhibitors Using a Cascade Docking Virtual Screening Approach

To discover novel inhibitors that target the influenza polymerase basic protein 2 (PB2) cap-binding domain (CBD), commercial ChemBridge compound libraries containing 384,796 compounds were screened using a cascade docking of LibDock–LigandFit–GOLD, and 60 compounds were selected for testing with cytopathic effect (CPE) inhibition assays and surface plasmon resonance (SPR) assay. Ten compounds were identified to rescue cells from H1N1 virus-mediated death at non-cytotoxic concentrations with EC₅₀ values ranging from 0.30 to 67.65 μM and could bind to the PB2 CBD of H1N1 with Kd values ranging from 0.21 to 6.77 μM. Among these, four compounds (11D4, 12C5, 21A5, and 21B1) showed inhibition of a broad spectrum of influenza virus strains, including oseltamivir-resistant ones, the PR/8-R292K mutant (H1N1, recombinant oseltamivir-resistant strain), the PR/8-I38T mutant (H1N1, recombinant baloxavir-resistant strain), and the influenza B/Lee/40 virus strain. These compounds have novel chemical scaffolds and relatively small molecular weights and are suitable for optimization as lead compounds. Based on sequence and structure comparisons of PB2 CBDs of various influenza virus subtypes, we propose that the Phe323/Gln325, Asn429/Ser431, and Arg355/Gly357 mutations, particularly the Arg355/Gly357 mutation, have a marked impact on the selectivities of PB2 CBD-targeted inhibitors of influenza A and influenza B.     

Functionalization of Piperidine Derivatives for the Site-Selective and Stereoselective Synthesis of Positional Analogues of Methylphenidate

Rhodium-catalyzed C−H insertions and cyclopropanations of donor/acceptor carbenes have been used for the synthesis of positional analogues of methylphenidate. The site selectivity is controlled by the catalyst and the amine protecting group. C−H functionalization of N-Boc-piperidine using Rh₂(R-TCPTAD)₄, or N-brosyl-piperidine using Rh₂(R-TPPTTL)₄ generated 2-substitited analogues. In contrast, when N-α-oxoarylacetyl-piperidines were used in combination with Rh₂(S-2-Cl-5-BrTPCP)₄, the C−H functionalization produced 4-susbstiuted analogues. Finally, the 3-substituted analogues were prepared indirectly by cyclopropanation of N-Boc-tetrahydropyridine followed by reductive regio- and stereoselective ring-opening of the cyclopropanes.