RESUMO
BACKGROUND: Tregs trafficking is controlled by CXCR4. In Renal Cell Carcinoma (RCC), the effect of the new CXCR4 antagonist, R54, was explored in peripheral blood (PB)-Tregs isolated from primary RCC patients. METHODS: PB-Tregs were isolated from 77 RCC patients and 38 healthy donors (HDs). CFSE-T effector-Tregs suppression assay, IL-35, IFN-γ, IL-10, TGF-ß1 secretion, and Nrp-1+Tregs frequency were evaluated. Tregs were characterised for CTLA-4, PD-1, CD40L, PTEN, CD25, TGF-ß1, FOXP3, DNMT1 transcriptional profile. PTEN-pAKT signalling was evaluated in the presence of R54 and/or triciribine (TCB), an AKT inhibitor. Methylation of TSDR (Treg-Specific-Demethylated-Region) was conducted. RESULTS: R54 impaired PB-RCC-Tregs function, reduced Nrp-1+Tregs frequency, the release of IL-35, IL-10, and TGF-ß1, while increased IFN-γ Teff-secretion. The CXCR4 ligand, CXCL12, recruited CD25+PTEN+Tregs in RCC while R54 significantly reduced it. IL-2/PMA activates Tregs reducing pAKT+Tregs while R54 increases it. The AKT inhibitor, TCB, prevented the increase in pAKT+Tregs R54-mediated. Moreover, R54 significantly reduced FOXP3-TSDR demethylation with DNMT1 and FOXP3 downregulation. CONCLUSION: R54 impairs Tregs function in primary RCC patients targeting PTEN/PI3K/AKT pathway, reducing TSDR demethylation and FOXP3 and DNMT1 expression. Thus, CXCR4 targeting is a strategy to inhibit Tregs activity in the RCC tumour microenvironment.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , PTEN Fosfo-Hidrolase , Receptores CXCR4 , Linfócitos T Reguladores , Humanos , Linfócitos T Reguladores/imunologia , PTEN Fosfo-Hidrolase/metabolismo , Receptores CXCR4/metabolismo , Neoplasias Renais/imunologia , Neoplasias Renais/patologia , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/metabolismo , Carcinoma de Células Renais/imunologia , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/metabolismo , Feminino , Masculino , Pessoa de Meia-Idade , Idoso , Adulto , Transdução de Sinais , Fatores de Transcrição Forkhead/metabolismoRESUMO
The present contribution introduces a novel computational protocol called PyRMD2Dock, which combines the Ligand-Based Virtual Screening (LBVS) tool PyRMD with the popular docking software AutoDock-GPU (AD4-GPU) to enhance the throughput of virtual screening campaigns for drug discovery. By implementing PyRMD2Dock, we demonstrate that it is possible to rapidly screen massive chemical databases and identify those with the highest predicted binding affinity to a target protein. Our benchmarking and screening experiments illustrate the predictive power and speed of PyRMD2Dock and highlight its potential to accelerate the discovery of novel drug candidates. Overall, this study showcases the value of combining AI-powered LBVS tools with docking software to enable effective and high-throughput virtual screening of ultralarge molecular databases in drug discovery. PyRMD and the PyRMD2Dock protocol are freely available on GitHub (https://github.com/cosconatilab/PyRMD) as an open-source tool.
Assuntos
Inteligência Artificial , Software , Simulação de Acoplamento Molecular , Proteínas/química , Descoberta de Drogas , Bibliotecas de Moléculas Pequenas , LigantesRESUMO
The present review focuses on synthetic peptide-based vaccine strategies in the context of anticancer intervention, paying attention to critical aspects such as peptide epitope selection, adjuvant integration, and nuanced classification of synthetic peptide cancer vaccines. Within this discussion, we delve into the diverse array of synthetic peptide-based anticancer vaccines, each derived from tumor-associated antigens (TAAs), including melanoma antigen recognized by T cells 1 (Melan-A or MART-1), mucin 1 (MUC1), human epidermal growth factor receptor 2 (HER-2), tumor protein 53 (p53), human telomerase reverse transcriptase (hTERT), survivin, folate receptor (FR), cancer-testis antigen 1 (NY-ESO-1), and prostate-specific antigen (PSA). We also describe the synthetic peptide-based vaccines developed for cancers triggered by oncovirus, such as human papillomavirus (HPV), and hepatitis C virus (HCV). Additionally, the potential synergy of peptide-based vaccines with common therapeutics in cancer was considered. The last part of our discussion deals with the realm of the peptide-based vaccines delivery, highlighting its role in translating the most promising candidates into effective clinical strategies. Although this discussion does not cover all the ongoing peptide vaccine investigations, it aims at offering valuable insights into the chemical modifications and the structural complexities of anticancer peptide-based vaccines.
Assuntos
Vacinas Anticâncer , Neoplasias , Humanos , Vacinas Anticâncer/química , Vacinas Anticâncer/imunologia , Neoplasias/imunologia , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/química , Vacinas de Subunidades Antigênicas/química , Vacinas de Subunidades Antigênicas/imunologia , Vacinas Sintéticas/química , Vacinas Sintéticas/imunologia , Peptídeos/química , Peptídeos/imunologia , Peptídeos/síntese químicaRESUMO
Antibiotic resistance is becoming a severe obstacle in the fight against acute and chronic infectious diseases that accompany most degenerative illnesses from neoplasia to osteo-arthritis and obesity. Currently, the race is on to identify pharmaceutical molecules or combinations of molecules able to prevent or reduce the insurgence and/or progression of infectivity. Attempts to substitute antibiotics with antimicrobial peptides have, thus far, met with little success against multidrug-resistant (MDR) bacterial strains. During the last decade, we designed and studied the activity and features of human ß-defensin analogs, which are salt-resistant, and hence active also under high salt concentrations as, for instance, in cystic fibrosis. Herein, we describe the design, synthesis, and major features of a new 21 aa long molecule, peptide γ2. The latter derives from the γ-core of the ß-defensin natural molecules, a small fragment of these molecules still bearing high antibacterial activity. We found that peptide γ2, which contains only one disulphide bond, recapitulates most of the biological properties of natural human ß-defensins and can also counteract both Gram-positive and Gram-negative MDR bacterial strains and biofilm formation. Moreover, it has great stability in human serum thereby enhancing its antibacterial presence and activity without cytotoxicity in human cells. In conclusion, peptide γ2 is a promising new weapon also in the battle against intractable infectious diseases.
Assuntos
Antibacterianos/farmacologia , Peptídeos Antimicrobianos/farmacologia , Bactérias/crescimento & desenvolvimento , Biofilmes/crescimento & desenvolvimento , beta-Defensinas/química , Bactérias/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla , Humanos , Testes de Sensibilidade MicrobianaRESUMO
Synthetic nucleic acid interactors represent an exciting research field due to their biotechnological and potential therapeutic applications. The translation of these molecules into drugs is a long and difficult process that justifies the continuous research of new chemotypes endowed with favorable binding, pharmacokinetic and pharmacodynamic properties. In this scenario, we describe the synthesis of two sets of homo-thymine nucleopeptides, in which nucleobases are inserted in a peptide structure, to investigate the role of the underivatized amino acid residue and the distance of the nucleobase from the peptide backbone on the nucleic acid recognition process. It is worth noting that the CD spectroscopy investigation showed that two of the reported nucleopeptides, consisting of alternation of thymine functionalized L-Orn and L-Dab and L-Arg as underivatized amino acids, were able to efficiently bind DNA and RNA targets and cross both cell and nuclear membranes.
Assuntos
Ácidos Nucleicos Peptídicos , Timina , Aminoácidos/química , DNA/química , Ácidos Nucleicos Peptídicos/química , Peptídeos/química , RNA/genética , Timina/químicaRESUMO
During almost all 2020, coronavirus disease 2019 (COVID-19) pandemic has constituted the major risk for the worldwide health and economy, propelling unprecedented efforts to discover drugs for its prevention and cure. At the end of the year, these efforts have culminated with the approval of vaccines by the American Food and Drug Administration (FDA) and the European Medicines Agency (EMA) giving new hope for the future. On the other hand, clinical data underscore the urgent need for effective drugs to treat COVID-19 patients. In this work, we embarked on a virtual screening campaign against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Mpro chymotrypsin-like cysteine protease employing our in-house database of peptide and non-peptide ligands characterized by different types of warheads acting as Michael acceptors. To this end, we employed the AutoDock4 docking software customized to predict the formation of a covalent adduct with the target protein. In vitro verification of the inhibition properties of the most promising candidates allowed us to identify two new lead inhibitors that will deserve further optimization. From the computational point of view, this work demonstrates the predictive power of AutoDock4 and suggests its application for the in silico screening of large chemical libraries of potential covalent binders against the SARS-CoV-2 Mpro enzyme.
Assuntos
COVID-19 , Inibidores de Proteases , Antivirais/farmacologia , Humanos , Simulação de Acoplamento Molecular , Pandemias , Inibidores de Proteases/farmacologia , SARS-CoV-2RESUMO
αv ß6 Integrin is an epithelial transmembrane protein that recognizes latency-associated peptide (LAP) and primarily activates transforming growth factor beta (TGF-ß). It is overexpressed in carcinomas (most notably, pancreatic) and other conditions associated with αv ß6 integrin-dependent TGF-ß dysregulation, such as fibrosis. We have designed a trimeric Ga-68-labeled TRAP conjugate of the αv ß6 -specific cyclic pentapeptide SDM17 (cyclo[RGD-Chg-E]-CONH2 ) to enhance αv ß6 integrin affinity as well as target-specific in-vivo uptake. Ga-68-TRAP(SDM17)3 showed a 28-fold higher αv ß6 affinity than the corresponding monomer Ga-68-NOTA-SDM17 (IC50 of 0.26 vs. 7.4â nM, respectively), a 13-fold higher IC50 -based selectivity over the related integrin αv ß8 (factors of 662 vs. 49), and a threefold higher tumor uptake (2.1 vs. 0.66 %ID/g) in biodistribution experiments with H2009 tumor-bearing SCID mice. The remarkably high tumor/organ ratios (tumor-to-blood 11.2; -to-liver 8.7; -to-pancreas 29.7) enabled high-contrast tumor delineation in PET images. We conclude that Ga-68-TRAP(SDM17)3 holds promise for improved clinical PET diagnostics of carcinomas and fibrosis.
Assuntos
Adenocarcinoma de Pulmão/diagnóstico por imagem , Antígenos de Neoplasias/análise , Complexos de Coordenação/química , Integrinas/análise , Neoplasias Pulmonares/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos/química , Animais , Compostos Aza/química , Química Click , Complexos de Coordenação/síntese química , Feminino , Radioisótopos de Gálio , Humanos , Camundongos , Camundongos SCID , Estrutura Molecular , Neoplasias Experimentais/diagnóstico por imagem , Peptídeos Cíclicos/química , Ácidos Fosfínicos/química , Piperidinas/química , Compostos Radiofarmacêuticos/síntese química , Células Tumorais CultivadasRESUMO
Here we investigated the structural and biological effects ensuing from the disulfide bond replacement of a potent and selective C-X-C chemokine receptor type 4 (CXCR4) peptide antagonist, with 1,4- and 1,5- disubstituted 1,2,3-triazole moieties. Both strategies produced candidates that showed high affinity and selectivity against CXCR4. Notably, when assessed for their ability to modulate the CXCL12-mediated cell migration, the 1,4-triazole variant conserved the antagonistic effect in the low-mid nanomolar range, while the 1,5-triazole one displayed the ability to activate the migration, becoming the first in class low-molecular-weight CXCR4 peptide agonist. By combining NMR and computational studies, we provided a valuable model that highlighted differences in the interactions of the two peptidomimetics with the receptor that could account for their different functional profile. Finally, we envisage that our findings could be translated to different GPCR-interacting peptides for the pursuit of novel chemical probes that could assist in dissecting the complex puzzle of this fundamental class of transmembrane receptors.
Assuntos
Dissulfetos/química , Peptídeos/química , Peptídeos/farmacologia , Receptores CXCR4/química , Triazóis/química , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Quimiocina CXCL12/farmacologia , Humanos , Ligantes , Peptidomiméticos , Receptores CXCR4/agonistasRESUMO
Telomere repeat binding factor 2 (TRF2) has been increasingly recognized to be involved in telomere maintenance and DNA damage response. Here, we show that TRF2 directly binds SIRT6 in a DNA independent manner and that this interaction is increased upon replication stress. Knockdown of SIRT6 up-regulates TRF2 protein levels and counteracts its down-regulation during DNA damage response, leading to cell survival. Moreover, we report that SIRT6 deactetylates in vivo the TRFH domain of TRF2, which in turn, is ubiquitylated in vivo activating the ubiquitin-dependent proteolysis. Notably, overexpression of the TRF2cT mutant failed to be stabilized by SIRT6 depletion, demonstrating that the TRFH domain is required for its post-transcriptional modification. Finally, we report an inverse correlation between SIRT6 and TRF2 protein expression levels in a cohort of colon rectal cancer patients. Taken together our findings describe TRF2 as a novel SIRT6 substrate and demonstrate that acetylation of TRF2 plays a crucial role in the regulation of TRF2 protein stability, thus providing a new route for modulating its expression level during oncogenesis and damage response.
Assuntos
Dano ao DNA , Sirtuínas/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo , Acetilação , Antineoplásicos Fitogênicos/farmacologia , Camptotecina/farmacologia , Linhagem Celular , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Humanos , Imuno-Histoquímica , Modelos Moleculares , Poli(ADP-Ribose) Polimerases/metabolismo , Ligação Proteica , Conformação Proteica , Estabilidade Proteica , Proteólise/efeitos dos fármacos , Proteínas Recombinantes de Fusão/metabolismo , Sirtuínas/química , Especificidade por Substrato , Proteína 2 de Ligação a Repetições Teloméricas/química , Proteína 2 de Ligação a Repetições Teloméricas/genética , UbiquitinaçãoRESUMO
Cationic nucleopeptides belong to a family of synthetic oligomers composed by amino acids and nucleobases. Their capability to recognize nucleic acid targets and to cross cellular membranes provided the basis for considering them as novel non-covalent delivery agents for nucleic acid pharmaceuticals. Herein, starting from a 12-mer nucleopeptide model, the number of cationic residues was modulated in order to obtain new nucleopeptides endowed with high solubility in acqueous medium, acceptable bio-stability, low cytotoxicity and good capability to bind nucleic acid. Two candidates were selected to further investigate their potential as nucleic acid carriers, showing higher efficiency to deliver PNA in comparison with RNA. Noteworthy, this study encourages the development of nucleopeptides as new carriers to extend the known strategies for those nucleic acid analogues, especially PNA, that still remain difficult to drive into the cells.
Assuntos
Portadores de Fármacos/metabolismo , Ácidos Nucleicos Peptídicos/metabolismo , Polilisina/metabolismo , RNA/metabolismo , Timina/análogos & derivados , Timina/metabolismo , Cátions/síntese química , Cátions/química , Cátions/metabolismo , Cátions/toxicidade , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Dicroísmo Circular , Portadores de Fármacos/síntese química , Portadores de Fármacos/química , Portadores de Fármacos/toxicidade , Humanos , Conformação de Ácido Nucleico , Hibridização de Ácido Nucleico , Ácidos Nucleicos Peptídicos/química , Ácidos Nucleicos Peptídicos/genética , Polilisina/síntese química , Polilisina/química , Polilisina/toxicidade , RNA/química , RNA/genética , Solubilidade , Temperatura , Timina/síntese química , Timina/toxicidade , Transfecção/métodosRESUMO
The RGD-recognizing αvß6 integrin has only recently emerged as a major target for cancer diagnosis and therapy. Thus, the development of selective, low-molecular-weight ligands of this receptor is still in great demand. Here, a metadynamics-driven design strategy allowed us to successfully convert a helical nonapeptide into a cyclic pentapeptide (6) showing remarkable potency and αvß6 specificity. NMR and docking studies elucidated the reasons for the high affinity and selectivity of this compound, setting the ground for the rational design of new αvß6-specific small peptides or even peptidomimetics. In vivo PET imaging studies demonstrated the potential use of 6 for medical applications.
Assuntos
Antígenos de Neoplasias/química , Integrinas/química , Humanos , Simulação de Acoplamento Molecular , Ressonância Magnética Nuclear BiomolecularRESUMO
Upon binding, ligands can chaperone their protein targets by preventing them from misfolding and aggregating. Thus, an organic molecule that works as folding chaperone for a protein might be its specific ligand, and, similarly, the chaperone potential could represent an alternative readout in a molecular screening campaign toward the identification of new hits. Here we show that small molecules selected for acting as pharmacological chaperones on a misfolded mutant of the Frizzled4 (Fz4) receptor bind and modulate wild-type Fz4, representing what are to our knowledge the first organic ligands of this until-now-undruggable GPCR. The novelty and the advantages of the screening platform, the allosteric binding site addressed by these new ligands and the mechanism they use to modulate Fz4 suggest new avenues for development of inhibitors of the Wnt-ß-catenin pathway and for drug discovery.
Assuntos
Receptores Frizzled/química , Chaperonas Moleculares/química , Sítio Alostérico , Motivos de Aminoácidos , Sequência de Bases , Linhagem Celular Tumoral , Química Farmacêutica/métodos , Relação Dose-Resposta a Droga , Desenho de Fármacos , Glicerol/química , Células HEK293 , Células HeLa , Humanos , Ligantes , Microscopia de Fluorescência , Dados de Sequência Molecular , Mutagênese , Ligação Proteica , Dobramento de Proteína , Receptores Acoplados a Proteínas G/químicaRESUMO
Nucleopeptides are promising nucleic acid mimetics in which the peptide backbone bears nucleobases. They can recognize DNA and RNA targets modulating their biological functions. To date, the lack of an effective strategy for the synthesis of nucleopeptides prevents their evaluation for biological and biomedical applications. Herein, we describe an unprecedented approach that enables the synthesis of cationic both homo and heterosequence nucleopeptides wholly on solid support with high yield and purity. Spectroscopic studies indicate advantageous properties of the nucleopeptides in terms of binding, thermodynamic stability and sequence specific recognition. Biostability assay and laser scanning confocal microscopy analyses reveal that the nucleopeptides feature acceptable serum stability and ability to cross the cell membrane.
Assuntos
DNA/química , Proteínas Nucleares/síntese química , Peptídeos/síntese química , RNA/química , Técnicas de Síntese em Fase Sólida/métodos , Sequência de Aminoácidos , Linhagem Celular Tumoral , Dicroísmo Circular , Humanos , Proteínas Nucleares/química , Peptídeos/químicaRESUMO
The αvß6 integrin binds the RGD-containing peptide of the foot and mouth disease virus with high selectivity. In this study, the long binding helix of this ligand was downsized to an enzymatically stable cyclic peptide endowed with sub-nanomolar binding affinity toward the αvß6 receptor and remarkable selectivity against other integrins. Computational studies were performed to disclose the molecular bases underlying the high binding affinity and receptor subtype selectivity of this peptide. Finally, the utility of the ligand for use in biomedical studies was also demonstrated here.
Assuntos
Antígenos de Neoplasias/química , Integrinas/química , Peptídeos/química , Sequência de Aminoácidos , Linhagem Celular Tumoral , Humanos , Ligantes , Espectroscopia de Ressonância Magnética , Simulação de Dinâmica MolecularRESUMO
The shelterin protein TRF2 has come to the limelight for its role in telomere maintenance and tumorigenesis. Herein, the application of rational design and synthesis allowed identifying the first TRF2TRFH binder able to elicit a marked DNA damage response in cancer cells. This work paves the way for the unprecedented employment of a chemical tool to finely tune specific mechanisms underlying telomere maintenance.
Assuntos
Desenho de Fármacos , Peptídeos Cíclicos/farmacologia , Proteína 2 de Ligação a Repetições Teloméricas/antagonistas & inibidores , Humanos , Modelos Moleculares , Peptídeos Cíclicos/síntese química , Peptídeos Cíclicos/química , Proteína 2 de Ligação a Repetições Teloméricas/metabolismoRESUMO
(Z)-Arylchlorooximes and α-isocyanoacetamides undergo a smooth reaction to produce 1,3-oxazol-2-oxime derivatives in good yields. Opening of the oxazole ring and deoximation reaction give a facile access to aryl-α-ketoamide amides, a class of privileged scaffolds in medicinal chemistry and important synthetic intermediates in organic chemistry.
Assuntos
Amidas/síntese química , Nitrilas/química , Oxazóis/química , Oximas/síntese química , Amidas/química , Catálise , Estrutura Molecular , Oxazóis/síntese química , Oximas/químicaRESUMO
BACKGROUND AND PURPOSE: While HCC is an inflammation-associated cancer, CRLM develops on permissive healthy liver microenvironment. To evaluate the immune aspects of these two different environments, peripheral blood-(PB), peritumoral-(PT) and tumoral tissues-(TT) from HCC and CRLM patients were evaluated. METHODS: 40 HCC and 34 CRLM were enrolled and freshly TT, PT and PB were collected at the surgery. PB-, PT- and TT-derived CD4+CD25+ Tregs, M/PMN-MDSC and PB-derived CD4+CD25- T-effector cells (Teffs) were isolated and characterized. Tregs' function was also evaluated in the presence of the CXCR4 inhibitor, peptide-R29, AMD3100 or anti-PD1. RNA was extracted from PB/PT/TT tissues and tested for FOXP3, CXCL12, CXCR4, CCL5, IL-15, CXCL5, Arg-1, N-cad, Vim, CXCL8, TGFß and VEGF-A expression. RESULTS: In HCC/CRLM-PB, higher number of functional Tregs, CD4+CD25hiFOXP3+ was detected, although PB-HCC Tregs exert a more suppressive function as compared to CRLM Tregs. In HCC/CRLM-TT, Tregs were highly represented with activated/ENTPD-1+Tregs prevalent in HCC. As compared to CRLM, HCC overexpressed CXCR4 and N-cadherin/vimentin in a contest rich in arginase and CCL5. Monocytic MDSCs were highly represented in HCC/CRLM, while high polymorphonuclear MDSCs were detected only in HCC. Interestingly, the function of CXCR4-PB-Tregs was impaired in HCC/CRLM by the CXCR4 inhibitor R29. CONCLUSION: In HCC and CRLM, peripheral blood, peritumoral and tumoral tissues Tregs are highly represented and functional. Nevertheless, HCC displays a more immunosuppressive TME due to Tregs, MDSCs, intrinsic tumor features (CXCR4, CCL5, arginase) and the contest in which it develops. As CXCR4 is overexpressed in HCC/CRLM tumor/TME cells, CXCR4 inhibitors may be considered for double hit therapy in liver cancer patients.
Assuntos
Carcinoma Hepatocelular , Neoplasias Colorretais , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Microambiente Tumoral , Arginase/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismoRESUMO
Despite progress in the prevention and diagnosis of cancer, current technologies for tumor detection present several limitations including invasiveness, toxicity, inaccuracy, lengthy testing duration and high cost. Therefore, innovative diagnostic techniques that integrate knowledge from biology, oncology, medicinal and analytical chemistry are now quickly emerging in the attempt to address these issues. Following this approach, here we developed a paper-based electrochemical device for detecting cancer-derived Small Extracellular Vesicles (S-EVs) in fluids. S-EVs were obtained from cancer cell lines known to express, at a different level, the αvß6 integrin receptor, a well-established hallmark of numerous epithelial cancer types. The resulting biosensor turned out to recognize αvß6-containing S-EVs down to a limit of 0.7*103 S-EVs/mL with a linear range up to 105 S-EVs /mL, and a relative standard deviation of 11%, thus it may represent a novel opportunity for αvß6 expressing cancers detection.
RESUMO
The present study describes a small library of peptides derived from a potent and selective CXCR4 antagonist (3), wherein the native disulfide bond is replaced using a side-chain to tail macrolactamization technique to vary ring size and amino acid composition. The peptides were preliminary assessed for their ability to interfere with the interaction between the receptor and anti-CXCR4 PE-conjugated antibody clone 12G5. Two promising candidates (13 and 17) were identified and further evaluated in a125I-CXCL12 competition binding assay, exhibiting IC50 in the low-nanomolar range. Furthermore, both candidates displayed high selectivity towards CXCR4 with respect to the cognate receptor CXCR7, ability to block CXCL12-dependent cancer cell migration, and receptor internalization, albeit at a higher concentration compared to 3. Molecular modeling studies on 13 and 17 produced a theoretical model that may serve as a guide for future modifications, aiding in the development of analogs with improved affinity. Finally, the study provides valuable insights into developing therapeutic agents targeting CXCR4-mediated processes, demonstrating the adaptability of our lead peptide 3 to alternative cyclization approaches and offering prospects for comprehensive investigations into the receptor region's interaction with its C-terminal region.
Assuntos
Dissulfetos , Peptídeos , Receptores CXCR4 , Receptores CXCR4/antagonistas & inibidores , Receptores CXCR4/metabolismo , Humanos , Sítios de Ligação/efeitos dos fármacos , Peptídeos/química , Peptídeos/farmacologia , Peptídeos/síntese química , Dissulfetos/química , Dissulfetos/farmacologia , Relação Estrutura-Atividade , Estrutura Molecular , Relação Dose-Resposta a Droga , Lactamas/química , Lactamas/farmacologia , Lactamas/síntese química , Movimento Celular/efeitos dos fármacos , Modelos Moleculares , Linhagem Celular TumoralRESUMO
Despite several major achievements in the development of vaccines and antivirals, the fight against SARS-CoV-2 and the health problems accompanying COVID-19 are still ongoing. SARS-CoV-2 main protease (Mpro), an essential viral cysteine protease, is a crucial target for the development of antiviral agents. A virtual screening analysis of in-house cysteine protease inhibitors against SARS-CoV-2 Mpro allowed us to identify two hits (i.e., 1 and 2) bearing a methyl vinyl ketone warhead. Starting from these compounds, we herein report the development of Michael acceptors targeting SARS-CoV-2 Mpro, which differ from each other for the warhead and for the amino acids at the P2 site. The most promising vinyl methyl ketone-containing analogs showed sub-micromolar activity against the viral protease. SPR38, SPR39, and SPR41 were fully characterized, and additional inhibitory properties towards hCatL, which plays a key role in the virus entry into host cells, were observed. SPR39 and SPR41 exhibited single-digit micromolar EC50 values in a SARS-CoV-2 infection model in cell culture.