Evaluation of pentaerythritol-based and trimethylolpropane-based cationic lipidic materials for gene delivery.

The chemical and physical structure of cationic liposomes pays an important effect on their gene transfection efficiency. Investigation on the structure-function relationship of cationic liposomes will guide the design of novel cationic liposomes with high transfection efficiency and biosafety. In this paper, two novel series of lipids based on the backbone of pentaerythritol and trimethylolpropane were discovered, and their gene transfection efficiencies were assayed in vitro. The four lipids 8c, 9c, 14b, and 15b, exhibited much better transfection efficiency in the HEK293 cell lines compared with Lipo2000, lipid 9c also showed good transfection efficiency in the SW480 cell lines. And the structure-efficiency relationship revealed that a hydroxyethyl polar head group boosted transfer potency in trimethylolpropane-type lipids, but reduced in pentaerythritol-type lipids.

Drug-Bearing Peptide-Based Nanospheres for the Inhibition of Metastasis and Growth of Cancer.

Employing a peptide-based nanoscale drug delivery system is an effective strategy to overcome the poor therapeutic outcomes of chemotherapeutic drugs. Here, we developed a self-assembling peptide-drug delivery system comprising a self-assembling anticancer peptide (R-lycosin-I), as revealed in our previous study, and 10-hydroxycamptothecin (HCPT) for cancer therapy. The results showed that peptide-drug conjugates (R-L-HCPT) could assemble into nanospheres of 40-60 nm in water. Compared with free HCPT, R-L-HCPT nanospheres not only inhibited tumor growth but also suppressed pulmonary metastatic nodules on B16-F10 cells in vivo. In summary, these results indicated that the self-assembling R-lycosin-I could provide a promising nanoscale platform for delivering small-molecule drugs. Moreover, our study might provide new opportunities for the development of a new class of functional peptide-drug-conjugated systems based on nanomaterials, which could synergistically enhance anticancer outcomes.

The Roles of Fatty-Acid Modification in the Activity of the Anticancer Peptide R-Lycosin-I.

We previously reported that R-lycosin-I, modified by amino acid substitution from lycosin-I, was a peptide with anticancer activity and a linear amphipathic α-helix conformation and that it can induce cancer cell apoptosis and inhibit cell proliferation. However, the anticancer activity of R-lycosin-I was not highly improved. In order to further improve the anticancer activity of R-lycosin-I, fatty acids with different chain lengths from 12 to 20 carbons were introduced to the N-terminal of R-lycosin-I to yield five lipopeptides (R-C12, R-C14, R-C16, R-C18, R-C20). The physicochemical properties of the five lipopeptides were determined by hydrodynamic size, ζ-potential, and circular dichroism spectroscopy, respectively. Then, the cytotoxic activity of these lipopeptides in A549 cells was evaluated with serum-containing and serum-free media, respectively, showing their anticancer activities were all increased through fatty-acid modification. This may be a result of the increased hydrophobicity and the enhanced interaction with the cancer cell membrane. The cytotoxic activity of R-C16 was 3-4-fold higher than that of the original R-lycosin-I and also was the strongest among all five lipopeptides, whether in serum or serum-free conditions. Compared with R-lycosin-I, the lactate dehydrogenase (LDH) leakage assay and scanning electron microscopy (SEM) indicated that R-C16 had a weakly destructive effect on the cancer cell membrane, but it might cause apoptosis to exert an anticancer activity. Finally, the impacts of fatty-acid length on the physicochemical properties and the anticancer potential of peptide were discussed. Our data consolidate work on fatty-acid-modified anticancer peptides.

Arginine modification of lycosin-I to improve inhibitory activity against cancer cells.

Lycosin-I is a linear amphipathic α-helical anticancer peptide (ACP) extracted from the spider Lycosa singoriensis, which can activate the mitochondrial death pathway to induce apoptosis in tumor cells and up-regulate p27 to inhibit cell proliferation. However, the applicability of lycosin-I as a novel anticancer drug is limited by its low cellular entry and efficacy in solid tumors. Amino acid substitution presents an effective and modest strategy to improve the anticancer activity and bioavailability of ACPs. Herein, an arginine-modified lycosin-I (named R-lycosin-I) was designed and synthesized by substituting lysine (Lys) with arginine (Arg). This peptide exhibited higher anticancer activity and penetrability against solid tumor cells than lycosin-I. They displayed noticeable differences in their physicochemical properties including the secondary structure, hydrodynamic size, and zeta potential. Fluorescence analyses have confirmed that R-lycosin-I exhibits increased cellular uptake and improved intracellular distribution. Due to its superior physical and chemical properties and high serum stability, R-lycosin-I could penetrate deeply into tumor spheroids and produce strong toxicity in the 3D tumor model. Overall, these findings suggest that arginine modification may provide an effective strategy for improving the anticancer activity of lycosin-I, and R-lycosin-I may be a useful lead for developing anticancer drugs.

Evaluation of Maltose-Based Cationic Liposomes with Different Hydrophobic Tails for Plasmid DNA Delivery.

In this paper, three cationic glycolipids with different hydrophobic chains Malt-DiC12MA (IX a), Malt-DiC14MA (IX b) and Malt-DiC16MA (IX c) were constructed by using maltose as starting material via peracetylation, selective 1-O-deacetylation, trichloroacetimidation, glycosylation, azidation, deacetylation, Staudinger reaction, tertiary amination and quaternization. Target compounds and some intermediates were characterized by ¹H-NMR, 13C-NMR, ¹H-¹H COSY and ¹H-13C HSQC. The results of gel electrophoresis assay, atomic force microscopy images (AFM) and dynamic light scattering (DLS) demonstrate that all the liposomes could efficiently bind and compact DNA (N/P ratio less than 2) into nanoparticles with proper size (88 nm-146 nm, PDI < 0.4) and zeta potential (+15 mV-+26 mV). The transfection efficiency and cellular uptake of glycolipids in HEK293 cell were evaluated through the enhanced green fluorescent protein (EGFP) expression and Cy3-labeled pEGFP-C1 (Enhanced Green Fluorescent Protein plasmid) images, respectively. Importantly, it indicated that Malt-DiC14MA exhibited high gene transfer efficiency and better uptake capability at N/P ratios of 8:1. Additionally, the result of cell viability showed glycolipids exhibited low biotoxicity and good biocompatibility by thiazolyl blue tetrazolium bromide (MTT) assay.

Design, synthesis and in vitro evaluation of d-glucose-based cationic glycolipids for gene delivery.

A cationic lipid consists of a hydrophilic headgroup, backbone and hydrophobic tails which have an immense influence on the transfection efficiency of the lipid. In this paper, two novel series of cationic cyclic glycolipids with a quaternary ammonium headgroup and different-length hydrophobic tails (dodecyl, tetradecyl, hexadecyl) have been designed and synthesized for gene delivery. One contains lipids 1-3 with two hydrophobic alkyl chains linked to the glucose ring directly via an ether link. The other contains lipids 4-6 with two hydrophobic chains on the positively charged nitrogen atoms. All of the lipids were characterized for their ability to bind to DNA, size, ζ-potential, and toxicity. Atomic force microscopy showed that the lipids and DNA-lipid complexes were sphere-like forms. The lipids were used to transfer enhanced green fluorescent protein (EGFP-C3) to HEK293 cells without a helper lipid, the results indicated that lipids 4-6 have better transfection efficiency, in particular lipids 5-6 have similar or better efficiency, compared with the commercial transfection reagent lipofectamine 2000.

A Lipid-Sensitive Spider Peptide Toxin Exhibits Selective Anti-Leukemia Efficacy through Multimodal Mechanisms.

Anti-cancer peptides (ACPs) represent a promising potential for cancer treatment, although their mechanisms need to be further elucidated to improve their application in cancer therapy. Lycosin-I, a linear amphipathic peptide isolated from the venom of Lycosa singorensis, shows significant anticancer potential. Herein, it is found that Lycosin-I, which can self-assemble into a nanosphere structure, has a multimodal mechanism of action involving lipid binding for the selective and effective treatment of leukemia. Mechanistically, Lycosin-I selectively binds to the K562 cell membrane, likely due to its preferential interaction with negatively charged phosphatidylserine, and rapidly triggers membrane lysis, particularly at high concentrations. In addition, Lycosin-I induces apoptosis, cell cycle arrest in the G1 phase and ferroptosis in K562 cells by suppressing the PI3K-AKT-mTOR signaling pathway and activating cell autophagy at low concentrations. Furthermore, intraperitoneal injection of Lycosin-I inhibits tumor growth of K562 cells in a nude mouse xenograft model without causing side effects. Collectively, the multimodal effect of Lycosin-I can provide new insights into the mechanism of ACPs, and Lycosin-I, which is characterized by high potency and specificity, can be a promising lead for the development of anti-leukemia drugs.

Peptide nanotube loaded with a STING agonist, c-di-GMP, enhance cancer immunotherapy against melanoma.

The activation of the stimulating factor of the interferon gene (STING) pathway can enhance the immune response within the tumor. Cyclic diguanylate monophosphate (c-di-GMP) is a negatively charged, hydrophilic STING agonist, however, its effectiveness is limited due to the poor membrane permeability and low bioavailability. Herein, we introduced KL-7 peptide derived from Aß amyloid fibrils that can self-assemble to form nanotubes to load and deliver c-di-GMP, which significantly enhanced c-di-GMP's effectiveness and then exhibited a robust "in situ immunity" to kill melanoma cells. KL-7 peptide nanotube, also called PNT, was loaded with negatively charged c-di-GMP via electrostatic interaction, which prepared a nanocomposite named c-di-GMP-PNT. Treatment of RAW 264.7 cells (leukemia cells in mouse macrophage) with c-di-GMP-PNT markedly stimulated the secretion of IL-6 and INF-ß along with phospho-STING (Ser365) protein expression, indicating the activation of the STING pathway. In the unilateral flank B16-F10 (murine melanoma cells) tumor-bearing mouse model, compared to PNT and c-di-GMP, c-di-GMP-PNT can promote the expression of INF-ß, TNF-α, IL-6, and IL-1ß. At the same time, up-regulated CD4 and CD8 active T cells kill tumors and enhance the immune response in tumor tissues, resulting in significant inhibition of tumor growth in tumor-bearing mice. More importantly, in a bilateral flank B16-F10 tumor model, both primary and distant tumor growth can also be significantly inhibited by c-di-GMP-PNT. Moreover, c-di-GMP-PNT demonstrated no obvious biological toxicity on the main organs (heart, liver, spleen, lung, and kidney) and biochemical indexes of mice. In summary, our study provides a strategy to overcome the barriers of free c-di-GMP in the tumor microenvironment and c-di-GMP-PNT may be an attractive nanomaterial for anti-tumor immunity. Electronic Supplementary Material: Supplementary material (synthesis and characterization of KL-7 peptide; the encapsulation rate and cumulative release rate of c-di-GMP-PNT; cytotoxicity of PNT, c-di-GMP, and c-di-GMP-PNT; anti-tumor effect of c-di-GMP-PNT (equivalent to 1 and 5 µg c-di-GMP per mouse); representative immunofluorescence images; and biosafety analysis) is available in the online version of this article at 10.1007/s12274-022-5102-z.

Structure-based discovery of potent inhibitors of Axl: design, synthesis, and biological evaluation.

Commonly overexpressed in many cancers and associated with tumor growth, metastasis, drug resistance, and poor overall survival, Axl has emerged as a promising target for cancer therapy. However, the availability of new chemical forms for Axl inhibition is limited. Herein, we present the development and characterization of novel Axl inhibitors, including the design, synthesis, and structure-activity relationships (SARs) of a series of diphenylpyrimidine-diamine derivatives. Most of these compounds exhibited remarkable activity against the Axl kinase. In particular, the promising compound m16 showed the highest enzymatic inhibitory potency (IC50 = 5 nM) and blocked multiple tumor cells' proliferation potencies (the CC50 of 4 out of 42 cancer cell lines <100 nM). Furthermore, compound m16 also possessed preferable pharmacokinetic profiles and liver microsome stability. All these favorable results make m16 a good leading therapeutic candidate for further development.

Improvement of anticancer effect of berberine by salt formation modifications.

BACKGROUND: Berberine is a quaternary isoquinoline alkaloid that possesses a significant therapeutic effect on a variety of cancers. However, due to poor bioavailability, an increased dose is often required to achieve therapeutic goals. To improve the activities of natural berberine, most modifications were focused on the positive isoquinoline unit by grafting long aliphatic chains or heterocycles. However, the negative part is ignored. At this point, the strategy of salt formation modifications with short- and medium-chain fatty acids was proposed in this article. PURPOSE: Using salt modification to enhance the antitumor activity of berberine and explore the mechanism. METHODS: Four short- and medium-chain fatty acid salts of berberine were prepared from berberine hydrochloride by salt formation modification with the sodium salt of butyric, caproic, octanoic, and decanoic acid, respectively. The cytotoxicity of four berberine salts on B16-F10, A549, HepG2, and U373 cancer cell lines was explored. Through cell localization, Mitochondrial membrane potential assay, and Western blotting analysis explored the mechanism of berberine salt-induced apoptosis. Its anticancer activity in vivo was demonstrated by the mouse xenograft model. RESULTS: The four berberine fatty acid salts exhibited an enhanced inhibitory effect on B16-F10, A549, HepG2, and U373 cancer cell lines, particularly on B16-F10 cells. Meanwhile, the four berberine fatty acid salts can inhibit the migration of B16-F10 cells. The four berberine fatty acid salts induce cancer cell apoptosis through the mitochondrial pathway, which was confirmed by the mitochondrial colocalization, the decreased mitochondrial membrane potential as well as activation of caspase-3, cytochrome C (Cyt-C), and down-regulated expression of B-cell lymphoma 2 (Bcl-2). Most importantly, the four berberine fatty acid salts inhibited tumor growth in the in vivo B16-F10 melanoma model without generating side effects intraperitoneally. CONCLUSIONS: This study revealed that salt formation modification may be an effective strategy to optimize the anticancer property of berberine hydrochloride and demonstrated the four berberine fatty acid salts induced apoptosis through the mitochondrial apoptotic pathway.

Cu(BF4)2/AC-Catalyzed Synthesis of N-Substituted Anilines, N-Substituted 1,6-Naphthyridin-5(6H)-one, and Isoquinolin-1(2H)-one.

Herein, we report practical Cu(BF4)2/activated carbon-catalyzed amination of various anilines, isoquinolinone, and naphthyridinone with aryl boronic acids. The ultrasonic and rotary evaporation treatment of the mixture of aq. Cu(BF4)2 and activated carbon in methanol afforded a novel Cu(II)-catalyst, which is air-stable and can be effectively applied in the Chan-Lam coupling reaction. The products of N-arylation were isolated in good to excellent yields at low catalytic loading. And Cu(BF4)2/AC also showed good reusability.

Small-molecule albumin ligand modification to enhance the anti-diabetic ability of GLP-1 derivatives.

Glucagon-like peptide-1 (GLP-1) receptor agonists modified with albumin ligands which can specificity bind to the human serum albumin (HSA) was an efficient strategy to prolong the half-time of GLP-1. Herein, we investigated the effect of small-molecule albumin ligand modification on the hypoglycemic activities of GLP-1 derivatives. Two GLP-1 derivatives MPA-C12-GLP-1 and Rhein-C12-GLP-1 were achieved by modification of the side chain amino of lysine in position 26 of the Arg34-GLP-1(7-37)-OH with Rhein and 3-Maleimidopropionic acid respectively using 12-aminolauric acid as a linker, and its specific albumin-conjugating characteristics, pharmaceutical characterization, and the antidiabetic effects were investigated. In vitro level, two GLP-1 derivatives demonstrated a higher binding capacity to GLP-1 receptor than that of Arg34-GLP-1(7-37)-OH. Interestingly, although the binding ability of MPA-C12-GLP-1 was equal to liraglutide, the binding ability of Rhein-C12-GLP-1 was 10-fold higher than liraglutide. In vivo level, the two GLP-1 derivatives can significantly increase their glucose tolerance and prolong their half-life in ICR mice, and they were also superior to GLP-1 in controlling glucose homeostasis and suppression of food intake and water consumption in db/db mice. Importantly, the two GLP-1 derivatives showed comparable efficacy to liraglutide for the therapy of type 2 diabetes mellitus. The in vitro INS-1 cells toxicity and the in vivo hepatotoxicity indicated that the Rhein-C12-GLP-1 was a safe candidate for the therapy of type 2 diabetes, and the serum biomarkers determination results showed that the Rhein-modified GLP-1 could significantly improve the HbA1c and blood lipids, and the H&E stain exhibited that the Rhein-C12-GLP-1 can effectively promote ß-cell proliferation and differentiation. In conclusion, the 3-Maleimidopropionic acid or Rhein-modified GLP-1derivatives have great potential for development as a Type 2 diabetes mellitus therapeutic drug.

Dual-sensitive antibacterial peptide nanoparticles prevent dental caries.

Background: Dental caries is the most prevalent bacterial biofilm-induced disease. Current clinical prevention and treatment agents often suffer from adverse effects on oral microbiota diversity and normal tissues, predominately arising from the poor biofilm-targeting property of the agents. Methods: To address this concern, we herein report dual-sensitive antibacterial peptide nanoparticles pHly-1 NPs upon acid and lipid-binding for treatment of dental caries. Amino acid substitutions were performed to design the peptide pHly-1. The potential, morphology and secondary structure of pHly-1 were characterized to elucidate the mechanisms of its pH and lipid sensitivity. Bacterial membrane integrity assay and RNA-seq were applied to uncover the antimicrobial mechanism of peptides under acidic condition. The in vitro and ex vivo antibiofilm assays were used to determine the antibiofilm performance of pHly-1 NPs. We also carried out the in vivo anti-caries treatment by pHly-1 NPs on dental caries animal model. Oral microbiome and histopathological analyses were performed to assess the in vivo safety of pHly-1 NPs. Results: The pHly-1 peptide underwent the coil-helix conformational transition upon binding to bacterial membranes in the acidic cariogenic biofilm microenvironment, thereby killing cariogenic bacteria. Under normal physiological conditions, pHly-1 adopted a ß-sheet conformation and formed nanofibers, resulting in negligible cytotoxicity towards oral microbes. However, in acidic solution, pHly-1 NPs displayed reliable antibacterial activity against Streptococcus mutans, including standard and clinically isolated strains, mainly via cell membrane disruption, and also suppressed in vitro and human-derived ex vivo biofilm development. Compared to the clinical agent chlorhexidine, in vivo topical treatment with pHly-1 NPs showed an advanced effect on inhibiting rat dental caries development without adverse effects on oral microbiota diversity and normal oral or gastric tissues. Conclusion: Our results demonstrated the high efficacy of dual-sensitive antimicrobial peptides for the selective damage of bacterial biofilms, providing an efficient strategy for preventing and treating dental caries.

Glucose-Lipopeptide Conjugates Reveal the Role of Glucose Modification Position in Complexation and the Potential of Malignant Melanoma Therapy.

Glycosylation and fatty acid modification are promising strategies to improve peptide performance. We previously studied glycosylation and fatty acid modification of the anticancer peptide R-lycosin-I. In this study, we further investigated the co-modification of fatty acids and monosaccharides in R-lycosin-I. A glucose derivative was covalently coupled to the ε-amino group of the Lys residues of the lipopeptide R-C12, which was derived from R-lycosin-I modified with dodecanoic acid, and obtained seven glycolipid peptides. They exhibited different cytotoxicity profiles, which may be related to the changes in physicochemical properties and binding ability to glucose transporter 1 (GLUT1). Among them, R-C12-4 exhibited the highest cytotoxicity and improved selectivity. A further study demonstrated that R-C12-4 showed significant cytotoxicity and antimetastasis activity in murine melanoma cells, melanoma spheroids, and animal models. Our results indicated that the glucose derivative modification position plays important roles in glucose-lipopeptide conjugates, and R-C12-4 might be a promising lead for developing anticancer drugs.

Fatty Acid Modification of the Anticancer Peptide LVTX-9 to Enhance Its Cytotoxicity against Malignant Melanoma Cells.

Spider venom is a valuable resource for the development of novel anticancer drugs. In this study, we focused on novel linear amphipathic α-helical anticancer peptide LVTX-9, which was derived from the cDNA library of the venom gland of the spider Lycosa vittata. The cytotoxicity of LVTX-9 against murine melanoma cells in the range of 1.56-200 µM was tested and found to be significantly lower than those of most anticancer peptides reported. Its IC50 was determined to be 59.2 ± 19.8 µM in a serum or 76.3 ± 12.7 µM in serum-free medium. Fatty acid modification is a promising strategy for improving peptide performance. Therefore, to enhance the cytotoxic activity of LVTX-9, fatty acid modification of this peptide was performed, and five different carbon chain length lipopeptides named LVTX-9-C12-C20 were produced. Among them, the lipopeptide LVTX-9-C18 showed the highest cytotoxic activity in relation to B16-F10 cells, whether in a serum or serum-free medium. Most importantly, the cytotoxic activity of LVTX-9-C18 was improved by about 12.9 times in a serum medium or 19.3 times in a serum-free medium compared to that of LVTX-9. Subsequently, assays including scanning electron microscopy, trypan blue staining, lactate dehydrogenase leakage assay, and hemolytic activity could indicate that the potential direct cell membrane disruption is the main mechanism of LVTX-9-C18 to induce cancer cell death. Furthermore, the LVTX-9-C18 also showed strong cytotoxicity in relation to 3D B16-F10 spheroids, which indicates it might be a promising lead for developing anticancer drugs.

[One-stage phallic reconstruction with extended pudendal-thigh flap: a desirable surgical option].

OBJECTIVE: To investigate the feasibility of one-stage phallic reconstruction with the extended pudendal-thigh flap (EPTF) to achieve a higher survival rate, shorter surgical cycle, and more experimental evidence for the clinical application of the technique. METHODS: We equally randomized 36 healthy adult New Zealand rabbits to an experimental and a control group to receive one-stage phallic reconstruction, the former with EPTF (pudendal-thigh flap + thigh cutaneous branches of the superficial external pudendal artery, PTF + TCB), the urethra reconstructed with the TCB flap, and the latter with bilateral pudendal-thigh flaps. RESULTS: The survival rate of the reconstructed phallus was 83.33% (15/18) and the urine leakage was 55.56% (10/18) in the experimental group, as compared with 38.89% (7/18) and 72.22% (13/18) in the controls. CONCLUSION: One-stage phallic reconstruction with EPTF, with its higher survival rate and shorter surgical cycle, is a desirable substitute for phallic reconstruction with bilateral pudendal-thigh flaps.

Monosaccharide Analogues of Anticancer Peptide R-Lycosin-I: Role of Monosaccharide Conjugation in Complexation and the Potential of Lung Cancer Targeting and Therapy.

Glycoconjugation is a promising modification strategy for the optimization of peptide drugs. In this study, five different monosaccharide derivatives (7a-e) were covalently linked to the N-terminal of R-lycosin-I, which yielded five glycopeptides (8a-e). They demonstrated increased or reduced cytotoxicity depending on monosaccharide types, which might be explained by the changes of physicochemical properties. Among all synthesized glycopeptides, only 8a exhibited increased cytotoxicity (IC50 = 9.6 ± 0.3 µM) and selectivity (IC50 = 37.4 ± 5.9 µM). The glucose transporter 1 (GLUT1) with high expression in cancer cells was approved to be involved in the cytotoxicity and selectivity enhancement of 8a. Furthermore, 8a but not R-lycosin-I inhibited tumor growth in the nude mice xenograft model without generating side effects intraperitoneally. Taken together, this study reveals the different monosaccharide roles in peptide modification and also provides an optimized anticancer peptide with high activity and selectivity, that is, 8a might be a promising lead for developing anticancer drugs.

Position Effect of Fatty Acid Modification on the Cytotoxicity and Antimetastasis Potential of the Cytotoxic Peptide Lycosin-I.

Peptide modification with fatty acids is an effective method to improve peptide performance. We previously investigated the fatty acid modification of R-lycosin-I, a cytotoxic peptide derived from lycosin-I from the venom of the spider Lycosa singoriensis. In this study, we further investigated the position effects of fatty acid modification of lycosin-I. Dodecanoic acid was covalently coupled to the α/ε-amino group of one of the seven Lys residues of lycosin-I, generating eight different lipopeptides. Although all the lipopeptides had significantly improved cytotoxicity compared with lycosin-I, they displayed different cytotoxic potencies and profiles, which might be explained by multifactors including charge, size, helicity, hydrophobicity, and so forth. Of the eight lipopeptides, L-C12 demonstrated highest cytotoxicity and antimetastasis activity in two-dimensional cells, tumor spheroids, subcutaneous transplantation mouse models, and experimental melanoma metastasis mouse models. Collectively, our finding indicated that fatty acid modification position plays important roles in physiochemical parameters and biological activities of cytotoxic peptides.

Installation of electron-donating protective groups, a strategy for glycosylating unreactive thioglycosyl acceptors using the preactivation-based glycosylation method.

Preactivation-based chemoselective glycosylation is a powerful strategy for oligosaccharide synthesis with its successful application in assemblies of many complex oligosaccharides. However, difficulties were encountered in reactions where glycosyl donors bearing multiple electron-withdrawing groups failed to glycosylate hindered unreactive acceptors. In order to overcome this problem, it was discovered that the introduction of electron-donating protective groups onto the glycosyl donors can considerably enhance their glycosylating power, leading to productive glycosylations even with unreactive acceptors. This observation is quite general and can be extended to a wide range of glycosylation reactions, including one-pot syntheses of chondroitin and heparin trisaccharides. The structures of the reactive intermediates formed upon preactivation were determined through low-temperature NMR studies. It was found that for a donor with multiple electron-withdrawing groups, the glycosyl triflate was formed following preactivation, while the dioxalenium ion was the major intermediate with a donor bearing electron-donating protective groups. As donors were all cleanly preactivated prior to the addition of the acceptors, the observed reactivity difference between these donors was not due to selective activation encountered in the traditional armed-disarmed strategy. Rather, it was rationalized by the inherent internal energy difference between the reactive intermediates and associated oxacarbenium ion like transition states during nucleophilic attack by the acceptor.