Pd-Catalyzed Picolinamide-Directed C(sp2)-H Sulfonylation of Amino Acids/Peptides with Sodium Sulfinates.

This report describes a Pd-catalyzed picolinamide-directed site-selective C(sp2)-H sulfonylation of amino acids and peptides with sodium sulfinates in moderate to good yields. Sulfonylation of levodopa and dopamine drug molecules and late-stage directed peptide sulfonylation are studied for the first time. Broad substrate scope having various functionalities, late-stage drug modifications, and various post synthetic utilities such as chalcogenation, bromination, olefination, and arylation are potential advantages.

Ru(II)-Catalyzed C(sp2)-H Activation Annulation: Synthesis of Fluorescent Benzoisoquinolonyl Acetate/Peptides from N-Arylamides and Ethyne at Room Temperature.

Benzoisoquinolones are aryl ring extended isoquinolinone derivatives, which are constituents of alkaloid natural products. This report describes the synthesis of novel benzoisoquinolone amino acid/peptide derivatives from the respective N-aryl amino esters/peptides through Ru-catalyzed C(sp2)-H annulation at room temperature. The N-terminal amide acts as an intrinsic directing group and coordinates with the active Ru(II) catalyst for the C-H bond activation and annulation of the aryl ring to produce benzoisoqunolone derivatives. Importantly, these benzoisoquinolinones exhibit fluorescence (QY ∼35%) in protic polar solvents, possibly due to charge transfer, and exhibit cell internalization to the cell nucleus without any significant cytotoxicity to human cell lines (HEK293T). Hence, our results are exceptional to transform standard amino acids/peptides into fluorescent peptides at room temperature in the late stage, which could be applicable for tracking specific target peptides by fluorescence microscopy.

Synthesis of R-GABA Derivatives via Pd(II) Catalyzed Enantioselective C(sp3)-H Arylation and Virtual Validation with GABAB1 Receptor for Potential leads.

GABA (γ-amino butyric acid) analogues like baclofen, tolibut, phenibut, etc., are well-known GABAB1 inhibitors and pharmaceutically important drugs. However, there is a huge demand for more chiral GABA aryl analogues with promising pharmacological actions. Here, we demonstrate the chiral ligand acetyl-protected amino quinoline (APAQ) mediated enantioselective synthesis of GABAB1 inhibitor drug scaffolds from easily accessible GABA via Pd-catalyzed C(sp3)-H activation. The synthetic methodology shows moderate to good yields, up to 74% of ee. We have successfully demonstrated the deprotection and removal of the directing group to synthesize R-tolibut in 86% yield. Further, we employed computation to probe the binding of R-GABA analogues to the extracellular domain of the human GABAB1 receptor. Our Rosetta-based molecular docking calculations show better binding for four R-enantiomers of GABA analogues than R-baclofen and R-phenibut. In addition, we employed GROMACS MD simulations and MMPB(GB)SA calculations to identify per-residue contribution to binding free energy. Our computational results suggest analogues (3R)-4-amino-3-(3,4-dimethylphenyl) butanoic acid, (3R)-4-amino-3-(3-fluorophenyl) butanoic acid, (3R)-3-(4-acetylphenyl)-4-aminobutanoic acid, (3R)-4-amino-3-(4-methoxyphenyl) butanoic acid, and (3R)-4-amino-3-phenylbutanoic acid are potential leads which could be synthesized from our methodology reported here.

Pd-Catalyzed Picolinamide-Directed Late-Stage Chalcogenation of Tryptophan-Containing Peptides.

This report describes the Pd-catalyzed late-stage chalcogenation of tryptophan-containing peptides with disulfides/diselenides in moderate to good yields. It comprises broad substrate scope, functional group diversity, late-stage modification of drug molecules, and various valuable synthetic transformations, including room temperature easy removal of the picolinamide auxiliary, which could be applicable to tune the structure and function of peptides.

N-Salicyl-AAn-picolamide Foldameric Peptides Exhibit Quorum Sensing Inhibition of Pseudomonas aeruginosa (PA14).

An organic acid, salicylic acid, and its derivatives are constituents of various natural products possessing remarkable bioactivity. O-Acetyl salicylate (aspirin) is a well-known life-saving drug. Its peptide derivative salicylamide has also been explored in the designing of peptide-based therapeutic drugs. An organic base, picolylamine has been recently explored for designing diagnostic probes. However, both the acid and base have common features as metal chelating with coordinating metals. Thus, these scaffolds could be used for designing inhibitors of various metalloenzymes. Their characteristic properties encourage us to design peptides containing both scaffolds (salicylic acid and picolylamine) at opposite terminals. So far there is no report available on such conjugated peptides. This report describes the synthesis, conformational analysis, and biochemical assessment of rationally designed N-salicyl-AAn-picolamide peptides. Pleasantly, we have obtained the crystal structures of representative peptides that confirm their roles in conformational changes. Our biological assessment as quorum sensing inhibitors has revealed that their di/tripeptides inhibit quorum sensing of the pathogenic bacterium PA14 strain. Hence, these peptides have promising foldameric and therapeutic values.

Synthesis of N-isoindolinonyl peptides via Pd-catalyzed C(sp2)-H olefination-activation and their conformational studies.

Isoindolinone is a constituent of several natural products that show a wide range of bioactivity, such as anticancer, antimicrobial, antiviral and anti-inflammatory properties. It would be interesting to explore the carbonyl group (H-bond acceptor) of isoindolinone and its structural and conformational changes. However, the synthesis of isoindolinone-comprising peptides in short steps is challenging. Herein, we have developed a synthetic methodology for introducing the isoindolinone residue to peptides via Pd-catalyzed C(sp2)-H activation/olefination, and demonstrated the conformational changes owing to the isoindolinone scaffold. Hence, isoindolinonyl peptides provide an avenue for the synthesis of novel foldamers and therapeutic agents.

A new transient directing group diethoxyethyl-L-proline facilitates ortho-arylation of aryl-amines/-amino acids via Pd-catalyzed C(sp2)-H activation.

Mono-ortho-arylated arylamines are constituents of various natural products but their syntheses are challenging. This report describes a new synthetic methodology for the ortho-arylation of arylamines and α-aromatic amino acids (phenylglycine and phenylalanine) through a Pd-catalyzed C(sp2)-H activation using the synthetic transient directing group diethoxyethyl-L-proline (DEP). A catalytic amount of diethoxyethyl-L-proline is sufficient to form mono-arylated arylamines as the major products using aryliodides. This method could be useful for the synthesis of various biphenyl amines and novel peptidomimetics.

Tropolone-Conjugated DNA: A Fluorescent Thymidine Analogue Exhibits Solvatochromism, Enzymatic Incorporation into DNA and HeLa Cell Internalization.

Tropolone is a non-benzenoid aromatic scaffold with unique photophysical and metal-chelating properties. Recently, it has been conjugated with DNA, and the photophysical properties of this conjugate have been explored. Tropolonyl-deoxyuridine (tr-dU) is a synthetic fluorescent DNA nucleoside analogue that exhibits pH-dependent emissions. However, its solvent-dependent fluorescence properties are unexplored owing to its poor solubility in most organic solvents. It would be interesting to incorporate it into DNA primer enzymatically. This report describes the solvent-dependent fluorescence properties of the silyl-derivative, and enzymatic incorporation of its triphosphate analogue. For practical use, its cell-internalization and cytotoxicity are also explored. tr-dU nucleoside was found to be a potential analogue to design DNA probes and can be explored for various therapeutic applications in the future.

Fluorescent DNA analog: 2-aminotroponyl-pyrrolyl-2'-deoxyuridinyl DNA oligo enhance fluorescence in DNA-duplex as compared to 2-aminotroponyl-ethynyl-2'-deoxyuridinyl DNA oligo.

The nucleobase modified fluorescent DNA and RNA analogs are synthesized by the conjugation of aromatic scaffolds through linkers, comprising mostly ethyne/ethene or fused ring residues at the pyrimidine/purine ring. These scaffolds are mainly derived from the benzenoid aromatic molecules comprising electron withdrawing/donating characters. However, non-benzenoid aromatic scaffolds such as tropolone and related derivatives are constituents of various troponoid natural products. The conjugation of nucleobases with a troponyl moiety is underutilized for the synthesis of fluorescent DNA analogs. This report describes the synthesis and photophysical studies of 2-aminotroponyl conjugated deoxyuridine nucleosides and their DNA analogs. 2-Aminotropone derivatives are conjugated at the C-5 position of uridine through an ethynyl linker/pyrrolyl ring fusion and their DNA analogs. Their photophysical studies reveal that aminotroponyl deoxyuridine analogs exhibit solvent-dependent fluorescence properties. Moreover, pyrrolyl ring-fused aminotroponyl DNA oligonucleotides enhance the fluorescence after formation of duplexation with complementary sequences of native DNA oligonucleotides. Hence, these modified nucleosides and DNA are promising fluorescent analogs which could be useful to design the sequence-specific DNA probes.

Synthesis of Aminotroponyl-/Difluoroboronyl Aminotroponyl Deoxyuridine Phosphoramidites.

This report describes the chemical synthesis of aminotroponyl-conjugated deoxyuridine analog (at-dU) and its difluoroboron complex (dfbat-dU) and their phosphoramidites by using the versatile phosphorylating reagent 2-Cyanoethyl N,N-diisopropylchlorophosphoramidite. Tropolone is a non-benzenoid aromatic bioactive natural fluorescent molecule, possessing intramolecular charge transfer and metal chelating properties with transition metal ions such as Cu2+/ Zn2+/ Ni2+ . Its synthetic derivatives, 2-aminotropones also exhibit unique bioactivities and are considered potential therapeutic drug candidate. Recently, the fluorescence properties of aminotropone has improved by complexing with difluoroboron residue that generates aminotroponyl-BODIPY analog. These could be employed for the synthesis of at-dU/dfbat-dU containing DNA oligonucleotides for designing the 11 B/19 F-NMR/fluorescence-based DNA probes. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Synthesis of N-propargyl-2-aminotropone (2) and difluoroboronyl N-propargyl-2-aminotropone (3) molecules. Basic Protocol 2: Synthesis of N-propargyl-2-aminotroponyl deoxyuridinyl (at-dU) phosphoramidites (7). Basic Protocol 3: Synthesis of difluoroboronyl N-propargyl-2-aminotroponyl deoxyuridinyl (dfbat-dU) phosphoramidites (10).

A new amino acid, hybrid peptides and BODIPY analogs: synthesis and evaluation of 2-aminotroponyl-L-alanine (ATA) derivatives.

Natural aromatic α-amino acid residues play critical roles in the structural and functional organization of proteins owing to π-interactions. Their aromatic residues are derived from benzenoid scaffolds. Non-benzenoid aromatic scaffolds such as tropone and tropolone are also constituents of troponoid natural products. Tropolone has also the ability to exhibit π-interactions along with additional hydrogen bonding. Thus, amino acids comprising troponyl could be potential building blocks of novel peptidomimetics. This report describes the synthesis of the L-aminotroponylalanine amino acid (ATA) and its unusual activity with the peptide coupling agent EDC. Importantly, its di-peptides form ß-sheet/-turn type secondary structures in organic solvents owing to the troponyl residue. This amino acid is an excellent scaffold for the synthesis of fluorescent amino acids such as BODIPY amino acid analogs. Nevertheless, this amino acid and its BODIPY derivatives can enter HeLa cells without exhibiting significant cytotoxicity at low concentrations (∼50 µM). Hence, ATA and its BODIPY derivatives are promising aromatic amino acids for the construction of potential peptidomimetics and fluorescent labelling of target peptides.

Cu-Catalyzed Synthesis of Alkylaminotroponyl Sulfones as Pseudomonas Aeruginosa Quorum Sensing Inhibitors Targeting lasI/R QS Circuitry.

The scarcity of novel bioactive molecules against multidrug-resistant (MDR) bacterial strains like Pseudomonas aeruginosa is alarming. This bacterial virulence is regulated via Quorum sensing (QS), a cell-cell communication process. Disabling QS circuits (las, pqs, rhl) with small molecules has been proposed as a potential strategy to prevent bacterial pathogenicity. This strategy focuses on interruption of bacterial virulence, rather than killing them to tackle the drug resistance problem. Herein, we describe the synthesis of rationally designed Alklyamionotroponyl Sulfone (ATS) derivatives by Cu-catalyzed C(sp2 )-H functionalization at tropone ring and the screening of their anti-QS activity against P. aeruginosa. Importantly, two sulfones (∼20 µM) remarkably exhibit the down regulation of the lasI/R QS genes. These molecules also inhibit swarming motility, biofilm formation and pyocyanin production, which reduce P. aeruginosa virulence in cells. Hence, ATS derivatives could be considered as potential therapeutic candidates for the treatment of P. aeruginosa infections.

Nonbenzenoid BODIPY Analogues: Synthesis, Structural Organization, Photophysical Studies, and Cell Internalization of Biocompatible N-Alkyl-Aminotroponyl Difluoroboron (Alkyl-ATB) Complexes.

The alkyl-BODIPY derivatives are lipid types of fluorescent molecules that exhibit a unique structure and functions including sensing of hydrophobic microenvironments in living cells. Their synthesis involves multisteps from the core structure dipyrromethene scaffold. The alkyl-BODIPY analogues are sought to derivatize with minimal synthetic steps even by altering the core structures derived from benzenoid aromatic moiety. Recently, the nonbenzenoid scaffold (aminotropone) has been explored to synthesize troponyl-BODIPY analogues, which are fluorescent. In the repertoire of nonbenzenoid analogue, N-alkyl-aminotroponyl difluoroboron (alkyl-ATB) is rationally designed comprising long-chain hydrocarbons to explore the lipid type of fluorescent molecules. This report describes the synthesis, photophysical studies, structural organization, and biocompatibilities of ATB derivatives containing different lengths of alkyl chain at 2-aminotropone scaffold. The photophysical studies of ATB derivatives reveal their fluorescence behaviors in organic solvents (CH3OH/CH3CN) with a quantum yield of ∼10 to 15%. These ATB derivatives also exhibit fluorescence characters in the solid state though their quantum yield is relatively low. Cell permeability and cytotoxicity studies reveal that alkyl-ATB derivatives are permeable to HeLa/HEK293T cell lines and show negligible cytotoxicity. The biocompatibility of alkyl-ATB derivatives is studied and confirmed by cell viability (MTT) assay to the HeLa/HEK293T cell lines. Importantly, the cell internalization studies of the representative alkyl-ATB molecule by fluorescence microscopy show that octyl-ATB is efficiently detectable at the cytoplasmic membrane and cellular nucleus in HeLa cells. Hence, alkyl-ATB derivatives are potential fluorescent molecules for developing probes to visualize cellular components under a fluorescence microscope.

Novel cinnamic acid analogues: synthesis of aminotroponyl acrylates by Pd(II)-catalysed C(sp2)-H olefination.

Cinnamic acid, a benzenoid scaffold, is a building block of various natural products. This report describes the synthesis of new non-benzenoid cinnamate analogs, 3-(6-amino-7-oxocyclohepta-1,3,5-trien-1-yl)acrylates, obtained through Pd(II)-catalyzed C7-H olefination of 2-aminotropones in the presence of acrylates. In these site-selective couplings, the troponyl-carbonyl function acts as a directing group. This strategy has been employed for the synthesis of new pseudopeptides from prolyl/prolamide containing aminotropone derivatives. These novel troponyl cinnamate analogs are potential precursors of hairpin forming peptides.

Unnatural Amino Acid: 4-Aminopyrazolonyl Amino Acid Comprising Tri-Peptides Forms Organogel With Co-Solvent (EtOAc:Hexane).

Ampyrone is an amino-functionalized heterocyclic pyrazolone derivative that possesses therapeutic values such as analgesic, anti-inflammatory, and antipyretics. The chemical structure of ampyrone exhibits excellent hydrogen bonding sites and is considered as the potential scaffold of supramolecular self-assembly. Recently, this molecule has been derived into unnatural amino acids such as aminopyrazolone amino acid and its peptides. This report describes that one of its amino acids, O-alkylated ampyrone, containing hybrid (α/ß) peptides forms organogel after sonication at 50-55°C with 0.7-0.9% (w/v) in ethyl acetate: hexane (1:3). The formation/morphology of such organogels is studied by nuclear magnetic resonance Fourier-transform infrared (FT-IR), circular dichroism (CD), scanning electron microscope (SEM), transmission electron microscopy (TEM), powder X-ray diffraction (Powder-XRD), and thermogravimetric analysis (TGA). Energy-minimized conformation of APA-peptides reveals the possibility of intermolecular hydrogen bonding. Hence, APA-peptides are promising peptidomimetics for the organogel-peptides.

One-pot synthesis of cyclic-aminotropiminium carboxylate derivatives with DNA binding and anticancer properties.

Tropolone, a nonbenzenoid aromatic molecule, is a constituent of troponoid natural products possessing a wide range of bioactivities, including anticancer. This report describes the one-pot synthesis and mechanistic studies of fifteen fluorescent Caryl-Nalkyl-substituted cyclic-aminotroponiminium carboxylate (cATC) derivatives by unusual cycloaddition and rearrangement reactions. Herein, the biochemical studies of four cATC derivatives reveal a non-intercalative binding affinity with DNA duplex. In vitro/in vivo studies show strong anti-tumor activity in three cATC derivatives. These derivatives enter the cells and localize to the nucleus and cytoplasm, which are easily traceable due to their inherent fluorescence properties. These three cATC derivatives reduce the proliferation and migration of HeLa cells more than the non-cancer cell line. They induce p38-p53-mediated apoptosis and inhibit EMT. In xenograft-based mouse models, these cATC derivatives reduce tumor size. Overall, this study reports the synthesis of DNA binding fluorescent Caryl-Nalkyl-cyclic-aminotroponiminium derivatives which show anti-tumor activity with the minimum side effect.

Pd-Catalyzed C(sp2)-H olefination: synthesis of N-alkylated isoindolinone scaffolds from aryl amides of amino acid esters.

Isoindolinone is a constituent of various natural products and synthetic biologically active compounds. The classical multi-step synthetic methods used to prepare various indolinone derivatives are tedious and challenging. One-pot synthetic methods are attractive and economical. Transition-metal-catalyzed C-H activation is an emerging tool for synthesizing natural products and small organic molecules via reducing the number of synthetic steps necessary. This paper describes the synthesis of N-alkyl-3-methenyl chiral isoindolinone derivatives from aryl amides of L-amino acids and non-activated alkene via Pd-catalyzed C(sp2)-H olefination. Herein, the amino acid residue acts as a directing group for olefination at the aryl ring, and then cyclization occurs at the amide NH. Hence, this methodology could be helpful to transform standard amino acids into respective chiral isoindolinone derivatives.

Unusual Pseudopeptides: Syntheses and Structural Analyses of Ethylenediprolyl Peptides and Their Metal Complexes with Cu(II) Ion.

The synthetic unnatural amino acids and their peptides as peptidomimetics have shown remarkable structural and functional properties. In the repertoire of synthetic peptides, pseudopeptides have emerged as attractive small peptidomimetics that are capable of forming the characteristic secondary structures in the solid/solution phase, as in natural peptides. This report describes the synthesis and structural analyses of novel pseudopeptides as ethylenediprolyl (etpro) tetra/hexapeptides, comprising a chiral diaminedicarboxylate scaffold. Their NMR and CD spectral analyses strongly support the formation of the ß-turn-type structures in organic solvents (ACN/MeOH). Further, the single-crystal X-ray studies of tetrapseudopeptide confirm the formation of a unique self-assembly structure as ß-strand type in the solid state through hydrogen bonding. Importantly, their diamine moiety influences the formation of Cu-complexes with Cu(II) ions. A tetrapseudopeptide monocarboxylate-Cu(II) complex forms the single crystal that is studied by the single-crystal X-ray diffractometer. The crystal structure of the tetrapseudopeptide-Cu(II) complex confirms the formation of the distorted square planar geometry structure, almost like the amyloid ß(Aß)-peptide-Cu(II) complex structural geometry. Hence, these etpro-pseudopeptides are emerging peptidomimatics that form ß-turn types of structures and metal complexes mainly with Cu(II) ions. These molecules could be considered for the development of peptide-based catalysts and peptide-based therapeutic drug candidates.

The 2'-caged-tethered-siRNA shows light-dependent temporal controlled RNAi activity for GFP gene into HEK293T cells.

Small interfering RNA (siRNA) exhibits gene-specific RNAi activity by the formation of RISC complex with mRNA of gene. The structural modification of siRNA at appropriate positions affects the structure of RISC complex and then RNAi activity. The modified siRNA are mostly prepared from the incorporation of sugar ring modified, and nucleobase modified RNA nucleotides. It is learned that the introduction of the sterically hindered nucleoside at the specific position of siRNA, severely affects siRNA-RISC complex formation. This report describes the syntheses of bulkier siRNA from 2'-caged-tethered-siRNAs, containing bulkier photolabile protecting group (o-nitrobenzyl) at 2'-position of ribose nucleoside. Importantly, these 2'-caged-siRNAs exhibit the light-dependent RNA interference (RNAi) activity into HEK293T cells for the GFP gene expression. The 2'-caged-siRNAs are synthesized by caging the sense and antisense strand of siRNA. The biochemical evaluations of these caged-siRNAs show that antisense-strand caged-siRNAs decrease RNAi activity temporarily in dark while enhancing RNAi activity, almost like control, after exposure withUV- light. However, 2'-caged sense strand siRNA increase RNAi activity temporarily while decreasing RNAi activity after exposure with light. These caged-siRNAs are also stable in the serum (fetal bovine serum) as like native siRNA. Hence these results strongly support that 2'-caged-tethered-siRNAs are promising analogues to control RNAi activity by UV-light.

Instability of Amide Bond with Trifluoroacetic Acid (20%): Synthesis, Conformational Analysis, and Mechanistic Insights into Cleavable Amide Bond Comprising ß-Troponylhydrazino Acid.

The instability of an amide bond with dilute trifluoroacetic acid (TFA) is a rare chemical event. The native amide bonds are stable even in the neat TFA, which is one of the reagents that releases the peptides from the solid support in the solid-supported peptide synthesis method. In the repertoire of unnatural peptidomics, α-/ß-hydrazino acids and their peptides are explored for the synthesis of N-amino peptide derivatives, and their amide bonds are stable in TFA (∼100%) as natural amide bonds. This report describes the synthesis of a ß-hydrazino acid analogue as ß-troponylhydrazino acid, containing a nonbenzenoid natural troponyl scaffold. The structural and conformational studies of their hybrid di-/tripeptides with the natural amino acid show that the 2-aminotroponyl residue is involved in hydrogen bonding. Surprisingly, the amide bond of ß-troponylhydrazino peptides is cleavable with TFA (∼20%) through the formation of a new heterocyclic molecule N-troponylpyrazolidinone or troponylpyrazolidinone. Tropolone and related compounds are excellent biocompatible chromophores. Hence, ß-troponylhydrazino acid could be employed for tuning the peptide structure and considered a promising chromophoric acid-sensitive protecting group of a free amine of amino acids/peptides. It could be applied for the estimation of the free amine group functionality by a UV-vis spectrophotometer.