Novel synthetic derivatives of cinnamic and p-coumaric acids with antiproliferative effect on breast MCF-7 tumor cells.

p-Coumaric acid is derived from cinnamic acid and is one of the major compounds in the Brazilian green propolis extract. Studies have shown that both p-coumaric acid and cinnamic acid have promising antiproliferative effects. In this context, aiming to increase the complexity of these active natural products and their activities, we performed coupling reactions with propargylamine and benzylamine, as well as with threonine, phenylalanine and lysine amino acids, aiming to enhance their antiproliferative effects towards the hormone-dependent breast cancer MCF-7 cells. Overall, the p-coumaric acid coupling with L-threonine amino acid (compound 15) had the best selectivity index (SI = 5.1), with half-maximal inhibitory concentration of 39.6 ± 1 µM, showing a high selectivity against hormone-dependent breast cancer cell lines MCF-7 and low cytotoxicity against the normal breast cell lines MCF-10A. Thus, this new natural product derivative may represent a prototype for the future development of antiproliferative agents, especially against hormone-dependent breast cancer.

Synthesis, antitumor activity and in silico analyses of amino acid derivatives of artepillin C, drupanin and baccharin from green propolis.

Breast cancer has the highest incidence and mortality in females, while prostate cancer has the second-highest incidence in males. Studies have shown that compounds from Brazilian green propolis have antitumor activities and can selectively inhibit the AKR1C3 enzyme, overexpressed in hormone-dependent prostate and breast tumors. Thus, in an attempt to develop new cytotoxic inhibitors against these cancers, three prenylated compounds, artepillin C, drupanin and baccharin, were isolated from green propolis to synthesize new derivatives via coupling reactions with different amino acids. All obtained derivatives were submitted to antiproliferative assays against four cancer cells (MCF-7, MDA MB-231, PC-3, and DU145) and two normal cell lines (MCF-10A and PNT-2) to evaluate their cytotoxicity. In general, the best activity was observed for compound6e, derived from drupanin, which exhibited half-maximal inhibitory concentration (IC50) of 9.6 ± 3 µM and selectivity index (SI) of 5.5 against MCF-7 cells.In silicostudies demonstrated that these derivatives present coherent docking interactions and binding modes against AKR1C3, which might represent a possible mechanism of inhibition in MCF-7 cells.

Multifaceted antibodies development against synthetic α-dystroglycan mucin glycopeptide as promising tools for dystroglycanopathies diagnostic.

Dystroglycanopathies are diseases characterized by progressive muscular degeneration and impairment of patient's quality of life. They are associated with altered glycosylation of the dystrophin-glycoprotein (DGC) complex components, such as α-dystroglycan (α-DG), fundamental in the structural and functional stability of the muscle fiber. The diagnosis of dystroglycanopathies is currently based on the observation of clinical manifestations, muscle biopsies and enzymatic measures, and the available monoclonal antibodies are not specific for the dystrophic hypoglycosylated muscle condition. Thus, modified α-DG mucins have been considered potential targets for the development of new diagnostic strategies toward these diseases. In this context, this work describes the synthesis of the hypoglycosylated α-DG mimetic glycopeptide NHAc-Gly-Pro-Thr-Val-Thr[αMan]-Ile-Arg-Gly-BSA (1) as a potential tool for the development of novel antibodies applicable to dystroglycanopathies diagnosis. Glycopeptide 1 was used for the development of polyclonal antibodies and recombinant monoclonal antibodies by Phage Display technology. Accordingly, polyclonal antibodies were reactive to glycopeptide 1, which enables the application of anti-glycopeptide 1 antibodies in immune reactive assays targeting hypoglycosylated α-DG. Regarding monoclonal antibodies, for the first time variable heavy (VH) and variable light (VL) immunoglobulin domains were selected by Phage Display, identified by NGS and described by in silico analysis. The best-characterized VH and VL domains were cloned, expressed in E. coli Shuffle T7 cells, and used to construct a single chain fragment variable that recognized the Glycopeptide 1 (GpαDG1 scFv). Molecular modelling of glycopeptide 1 and GpαDG1 scFv suggested that their interaction occurs through hydrogen bonds and hydrophobic contacts involving amino acids from scFv (I51, Y33, S229, Y235, and P233) and R8 and α-mannose from Glycopeptide 1.

Synthesis of novel triazole-derived glycopeptides as analogs of α-dystroglycan mucins.

α-Dystroglycan (α-DG) mucins are essential for maintenance of the structural and functional stability of the muscle fiber and, when hypoglycosylated, they are directly involved in pathological processes such as dystroglycanopathies. Thus, this work reports the synthesis of the novel 1,2,3-triazole-derived glycosyl amino acids αGlcNAc-1-O-triazol-2Manα-ThrOH (1) and Gal-ß1,4-αGlcNAc-1-O-triazol-2Manα-ThrOH (2), followed by solid-phase assembly to get the corresponding glycopeptides NHAcThrVal[αGlcNAc-1-triazol-2Manα]ThrIleArgGlyOH (3) and NHAcThrVal[Gal-ß1,4-αGlcNAc-1-triazol-2Manα]ThrIleArgGlyOH (4) as analogs of α-DG mucins. The glycosyl amino acids 1 (72%) and 2 (35%) were synthesized by Cu(I)-assisted 1,3-dipolar azide-alkyne cycloaddition reactions (CuAAC) between the azide-glycosyl amino acid αManN3-FmocThrOBn (5) and the corresponding alkyne-functionalyzed sugars 2'-propynyl-αGlcNAc (6) and 2'-propynyl-Gal-ß1,4-αGlcNAc (7), followed by hydrogenation reactions. Subsequently, glycopeptides 3 (23%) and 4 (12%) were obtained by solid phase synthesis, involving sequential couplings of Fmoc-protected amino acids or the glycosyl amino acids 1 and 2, followed by cleavage from resin, N-acetylation and O-deacetylation (NaOMe) reactions. Lastly, enzymatic galactosylation of glycopeptide 3 with bovine ß-1,4-GalT showed that it was not a substrate for this enzyme, which could be better elucidated by docking simulations with ß-1,4-GalT.

Insights into Anti-Trypanosomal Agents Based on Synthetic Glycoconjugates.

Chagas disease is still a worldwide threat, with estimated 6 to 7 million infected people, mainly in Latin America. Current treatments still rely only on benznidazol and nifurtimox, drugs with poor efficacy in chronic infection phase and recognized toxicity. Thus, there is an urgent need for new therapeutic agents against this disease. In this review we present an updated selection over the last decade of synthetic glycoconjugates as anti-trypanosomal agents, properly addressed as monosaccharideand disaccharide-based agents, and multivalent-based derivatives, disclosing relevant methods for their synthesis, along with their activities on T. cruzi and its trans-sialidase (TcTS). In addition, synthetic glycoconjugates as potential vaccines and diagnostic antigens against T. cruzi are also reported.

Binding of triazole-linked galactosyl arylsulfonamides to galectin-3 affects Trypanosoma cruzi cell invasion.

The synthesis of the O-3 triazole-linked galactosyl arylsulfonamides 1-7 as potential inhibitors of Trypanosoma cruzi cell invasion is described. These target compounds were synthesized by Cu(I)-catalysed azide-alkyne cycloaddition reaction ('click chemistry') between different azide arylsulfonamides and the alkyne-based sugar 3-O-propynyl-ßGalOMe. Inhibition assays of T. cruzi cell invasion with compounds 1-7 showed reduced values of infection index (∼20) for compounds 3 and 5, bearing the corresponding 5-methylisoxazole and 2,4-dimethoxypyrimidine groups, which also presented higher binding affinities to galectin-3 (EC50 17-18 µM) in Corning Epic label-free assays. In agreement with experimental results, the assessment of the theoretical binding of compounds 1-7 to galectin-3 by MM/PBSA method displayed higher affinities for compounds 3 (-9.7 kcal/mol) and 5 (-11.1 kcal/mol). Overall, these achievements highlight compounds 3 and 5 as potential T. cruzi cell invasion blockers by means of a galectin-3 binding-related mechanism, revealing galectin-3 as an important host target for design of novel anti-trypanosomal agents.

Synthetic glycoconjugates inhibitors of tumor-related galectin-3: an update.

Galectin-3 is associated with the development and malignancy of several types of tumor, mediating important tumor-related functions, such as tumorigenesis, neoplastic transformation, tumor cell survival, angiogenesis, tumor metastasis and regulation of apoptosis. Therefore, synthetic galectin-3 inhibitors are of utmost importance for development of new antitumor therapeutic strategies. In this review we present an updated selection of synthetic glycoconjugates inhibitors of tumor-related galectin-3, properly addressed as monosaccharide- and disaccharide-based inhibitors, and multivalent-based inhibitors, disclosuring relevant methods for their synthesis along with their inhibitory activities towards galectin-3. In general, Cu(I)-assisted 1,3-dipolar azide-alkyne cycloaddition (CuAAC) reactions were predominantly applied for the synthesis of the described inhibitors, which had their inhibitory activities against galectin-3 evaluated by fluorescence polarization, surface plasmon resonance (SPR), hemagglutination, ELISA and cell imaging assays. Overall, the presented synthetic glycoconjugates represent frontline galectin-3 inhibitors, finding important biomedical applications in cancer.

Trypanosoma cruzi Invasion into Host Cells: A Complex Molecular Targets Interplay.

Chagas' disease is still a worldwide threat, with estimated from 6 to 7 million infected people, mainly in Latin America. Despite all efforts, especially from international consortia (DNDi, NMTrypI), to develop an innovative therapeutic strategy against this disease, no candidate has achieved full requirements for clinical use yet. In this review, we point out the general molecular and cellular mechanisms involved in T. cruzi cell invasion and elucidate the roles of specific parasite and host targets in the progress of Chagas' disease. Among these molecular targets are Gp85/transsialidase, mucins, cruzipain and oligopeptidase B, found in parasite cell surface, and Galectin-3 and Toll-like receptors present in host cells. Thus, the deep understanding of their interplay and involvement on T. cruzi host cell adhesion, invasion and evasion from host immune may expand the chances for discovering new therapeutic agents against this neglected disease. Additionally, these targets may represent a remarkable strategy to block parasite invasion in the early stages of infection.

Synthetic 1,2,3-triazole-linked glycoconjugates bind with high affinity to human galectin-3.

This work describes the synthesis of the 1,2,3-triazole amino acid-derived-3-O-galactosides 1-6 and the 1,2,3-triazole di-lactose-derived glycoconjugate 7 as potential galectin-3 inhibitors. The target compounds were synthesized by Cu(I)-catalyzed azide-alkyne cycloaddition reaction ('click chemistry') between the azido-derived amino acids N3-ThrOBn, N3-PheOBn, N3-N-Boc-TrpOBn, N3-N-Boc-LysOBn, N3-O-tBu-AspOBn and N3-l-TyrOH, and the corresponding alkyne-based sugar 3-O-propynyl-GalOMe, as well as by click chemistry reaction between the azido-lactose and 2-propynyl lactose. Surface plasmon resonance (SPR) assays showed that all synthetic glycoconjugates 1-7 bound to galectin-3 with high affinity, but the highest binders were the amino acids-derived glycoconjugates 2 (KD 7.96µM) and 4 (KD 4.56µM), and the divalent lactoside 7 (KD1 0.15µM/KD2 19µM). Molecular modeling results were in agreement with SPR assays, since more stable interactions with galectin-3 were identified for glycoconjugates 2, 4 and 7. Regarding compounds 2 and 4, they established specific cation-π (Arg144) and ionic (Asp148) interactions, whereas glycoconjugate 7 was capable to bridge two independent galectin-3 CRDs, creating a non-covalent cross-link between two monomers and, thus, reaching a submicromolar affinity towards galectin-3.

Chemical and chemoenzymatic synthesis of glycosyl-amino acids and glycopeptides related to Trypanosoma cruzi mucins.

This study describes the synthesis of the alpha- and beta-linked N-acetyllactosamine (Galp-beta-1,4-GlcNAc; LacNAc) glycosides of threonine (LacNAc-Thr). LacNAc-a-Thr was prepared by direct chemical coupling of a 2-azido-2-deoxy-lactose disaccharide donor to a suitable partially protected threonine unit. In contrast, stepwise chemical generation of beta-linked N-acetylglucosamine followed by enzymatic galactosylation to give LacNAc-beta-Thr proved effective, whereas use of a 2-azido-2-deoxy-lactose donor in acetonitrile failed to give the desired beta-linked disaccharyl glycoside. This study illustrates that it is possible to overcome the inherent stereoselection for 1,2-trans chemical glycosylation with a GlcNAc donor, and that the well-established preference of bovine beta-1,4-galactosyltransferase for beta-linked acceptor substrates can also be overcome. Using this knowledge, short glycopeptide fragments based on T. cruzi mucin sequences, Thr-Thr-[LacNAcThr]-Thr-Thr-Gly, were synthesised. All LacNAc-based compounds outlined were shown to serve as acceptor substrates for sialylation by T. cruzi trans-sialidase.

Molecular modeling, docking and ADMET studies applied to the design of a novel hybrid for treatment of Alzheimer's disease.

Alzheimer's disease (AD) is the most common form of dementia in adults, which is characterized by senile plaquets and cholinergic deficit as the disease progresses. Improvement of cholinergic neurotransmission is the basis of some drugs currently used in the treatment of AD. It is achieved by acetylcholinesterase (AChE) inhibition, the enzyme responsible for acetylcholine hydrolysis. Molecular modeling techniques were of utmost importance to design a new pharmaceutical against Alzheimer's disease, with potential inhibitory activity over AChE, since the inhibition of human plasma butyrylcholinesterase (BChE) may cause side effects. Some of the drugs currently used in the treatment of AD are capable of increasing the cholinergic transmission through the AChE inhibition. In this work we proposed molecular hybrids of tacrine with donepezil (fusion of these structures), in order to suggest new proposals of AChE inhibitors for future treatment of AD. We have analyzed all the structures by docking, density functional studies and drug like properties.