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INTRODUCTION: Polymyxins, the cationic lipopeptide antibiotics, are the last line of therapeutics against the MDR Gram-negative bacterial (GNB) pathogens. Unfortunately, the rising cases of polymyxin-resistant strains from across the globe have adversely impacted their utility. While the molecular mechanisms responsible for developing polymyxin resistance (PolR) are largely understood, the prevalence of PolR strains in India has not been investigated systematically. The current study was undertaken to primarily determine the prevalence of PolR strains in India. Moreover, the extent of the spread of mobile colistin resistance (mcr) genes among the GNB strains in India was also determined. METHOD: A systematic search for articles using the relevant inclusion and exclusion criteria was performed in the applicable databases for the period January 2015 to December 2023. The included 41 studies were subjected to a meta-analysis using the Comprehensive Meta-Analysis software (V4.0). Publication biases were assessed using funnel plots and Egger's regression analysis. RESULT: Considering a total of 41 studies including 24â589 bacterial isolates the present meta-analysis found the rate of PolR bacteria in India to be at 15.0% (95% CI: 11.2 to 19.8). Among the Indian States, Tamil Nadu topped with the highest prevalence of PolR at 28.3%. Investigating the contribution of the mcr genes, it was observed that among the PolR strains, 8.4% (95% CI: 4.8 to 14.3) were mcr positive. CONCLUSION: The study determined the prevalence of PolR strains in India at 15.0%, which is higher than that of the global average at 10%. The study also determined that 8.4% of the PolR strains carried the mcr genes. The mcr-positive strains reported from India could be an underestimation of the actual numbers due to the non-inclusion of mcr screening in many previous studies. This study provides insight into the state of the PolR situation in India, which may be useful to develop a monitoring strategy to contain the spread of such strains and preserve the efficacy of the polymyxins.
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Antibacterianos , Infecções por Bactérias Gram-Negativas , Polimixinas , Índia/epidemiologia , Humanos , Prevalência , Antibacterianos/farmacologia , Polimixinas/farmacologia , Infecções por Bactérias Gram-Negativas/microbiologia , Infecções por Bactérias Gram-Negativas/epidemiologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Negativas/genética , Bactérias Gram-Negativas/classificação , Testes de Sensibilidade Microbiana , Farmacorresistência Bacteriana/genética , Farmacorresistência Bacteriana Múltipla/genética , Colistina/farmacologiaRESUMO
With the rising incidences of antimicrobial resistance (AMR) and the diminishing options of novel antimicrobial agents, it is paramount to decipher the molecular mechanisms of action and the emergence of resistance to the existing drugs. Polymyxin, a cationic antimicrobial lipopeptide, is used to treat infections by Gram-negative bacterial pathogens as a last option. Though polymyxins were identified almost seventy years back, their use has been restricted owing to toxicity issues in humans. However, their clinical use has been increasing in recent times resulting in the rise of polymyxin resistance. Moreover, the detection of "mobile colistin resistance (mcr)" genes in the environment and their spread across the globe have complicated the scenario. The mechanism of polymyxin action and the development of resistance is not thoroughly understood. Specifically, the polymyxin-bacterial lipopolysaccharide (LPS) interaction is a challenging area of investigation. The use of advanced biophysical techniques and improvement in molecular dynamics simulation approaches have furthered our understanding of this interaction, which will help develop polymyxin analogs with better bactericidal effects and lesser toxicity in the future. In this review, we have delved deeper into the mechanisms of polymyxin-LPS interactions, highlighting several models proposed, and the mechanisms of polymyxin resistance development in some of the most critical Gram-negative pathogens.
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Lipopolissacarídeos , Polimixinas , Humanos , Polimixinas/farmacologia , Farmacorresistência Bacteriana/genética , Antibacterianos/farmacologia , Colistina/farmacologiaRESUMO
There is an unmet need to improve the efficacy of platinum-based cancer chemotherapy, which is used in primary and metastatic settings in many cancer types. In bladder cancer, platinum-based chemotherapy leads to better outcomes in a subset of patients when used in the neoadjuvant setting or in combination with immunotherapy for advanced disease. Despite such promising results, extending the benefits of platinum drugs to a greater number of patients is highly desirable. Using the multiomic assessment of cisplatin-responsive and -resistant human bladder cancer cell lines and whole-genome CRISPR screens, we identified puromycin-sensitive aminopeptidase (NPEPPS) as a driver of cisplatin resistance. NPEPPS depletion sensitized resistant bladder cancer cells to cisplatin in vitro and in vivo. Conversely, overexpression of NPEPPS in sensitive cells increased cisplatin resistance. NPEPPS affected treatment response by regulating intracellular cisplatin concentrations. Patient-derived organoids (PDO) generated from bladder cancer samples before and after cisplatin-based treatment, and from patients who did not receive cisplatin, were evaluated for sensitivity to cisplatin, which was concordant with clinical response. In the PDOs, depletion or pharmacologic inhibition of NPEPPS increased cisplatin sensitivity, while NPEPPS overexpression conferred resistance. Our data present NPEPPS as a druggable driver of cisplatin resistance by regulating intracellular cisplatin concentrations. SIGNIFICANCE: Targeting NPEPPS, which induces cisplatin resistance by controlling intracellular drug concentrations, is a potential strategy to improve patient responses to platinum-based therapies and lower treatment-associated toxicities.
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Cisplatino , Resistencia a Medicamentos Antineoplásicos , Neoplasias da Bexiga Urinária , Humanos , Cisplatino/farmacologia , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/metabolismo , Animais , Camundongos , Linhagem Celular Tumoral , Aminopeptidases/genética , Aminopeptidases/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Antineoplásicos/farmacologia , Organoides/efeitos dos fármacos , Organoides/metabolismoRESUMO
Invariant NKT (iNKT) cells comprise a heterogeneous group of non-circulating, tissue-resident T lymphocytes that recognize glycolipids, including alpha-galactosylceramide (αGalCer), in the context of CD1d, but whether peripheral iNKT cell subsets are terminally differentiated remains unclear. Here we show that mouse and human liver-resident αGalCer/CD1d-binding iNKTs largely correspond to a novel Zbtb16+Tbx21+Gata3+MaflowRorc- subset that exhibits profound transcriptional, phenotypic and functional plasticity. Repetitive in vivo encounters of these liver iNKT (LiNKT) cells with intravenously delivered αGalCer/CD1d-coated nanoparticles (NP) trigger their differentiation into immunoregulatory, IL-10+IL-21-producing Zbtb16highMafhighTbx21+Gata3+Rorc- cells, termed LiNKTR1, expressing a T regulatory type 1 (TR1)-like transcriptional signature. This response is LiNKT-specific, since neither lung nor splenic tissue-resident iNKT cells from αGalCer/CD1d-NP-treated mice produce IL-10 or IL-21. Additionally, these LiNKTR1 cells suppress autoantigen presentation, and recognize CD1d expressed on conventional B cells to induce IL-10+IL-35-producing regulatory B (Breg) cells, leading to the suppression of liver and pancreas autoimmunity. Our results thus suggest that LiNKT cells are plastic for further functional diversification, with such plasticity potentially targetable for suppressing tissue-specific inflammatory phenomena.
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Linfócitos B Reguladores , Células T Matadoras Naturais , Animais , Antígenos CD1d/metabolismo , Autoimunidade , Linfócitos B Reguladores/metabolismo , Galactosilceramidas , Interleucina-10/metabolismo , Fígado/metabolismo , CamundongosRESUMO
Cell proliferation is a crucial step that might promote cancer if deregulated. Therefore, this vital segment is critically controlled by a complicated cell-cycle process in normal cells that is regulated by some regulatory proteins. It has been observed that p16 protein, playing a crucial role in cell-cycle progression/regulation, remains inactivated in different cancer cells. This inactivity of p16 protein leads to the enhancement of cancer cell proliferation by allowing uncontrolled cancer cell division. Hence, the activity of p16 protein needs to be restored using new viral vectors, small molecules as well as peptides to control/suppress this type of abnormal cell proliferation. In this work, we have taken an interesting approach to increase the efficiency and bio-availability of p16 peptide (functional part of p16 protein) to be an aggressive anti-leukemia therapeutic agent by conjugating a nuclear-localized signal (NLS) sequence and a short peptide (AVPI) with it. Moreover, this newly designed NLS attached hybrid peptide greatly affects XIAP expressing but p16 lower expressing human chronic myelogenous leukemia (CML) cell proliferation by targeting both nuclear (CDK4/cyclin D) and cellular factors (XIAP) and promoting the caspase-3 dependent apoptosis pathway.
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Infections caused by multi-drug resistant (MDR) bacterial pathogens are a leading cause of mortality and morbidity across the world. Indiscriminate use of broad-spectrum antibiotics has seriously affected this situation. With the diminishing discovery of novel antibiotics, new treatment methods are urgently required to combat MDR pathogens. Polymyxins, the cationic lipopeptide antibiotics, discovered more than half a century ago, are considered to be the last-line of antibiotics available at the moment. This antibiotic shows a great bactericidal effect against Gram-negative bacteria. Polymyxins primarily target the bacterial membrane and disrupt them, causing lethality. Because of their membrane interacting mode of action, polymyxins cause nephrotoxicity and neurotoxicity in humans, limiting their usability. However, recent modifications in their chemical structure have been able to reduce the toxic effects. The development of better dosing regimens has also helped in getting better clinical outcomes in the infections caused by MDR pathogens. Since the mid1990s the use of polymyxins has increased manifold in clinical settings, resulting in the emergence of polymyxin-resistant strains. The risk posed by the polymyxin-resistant nosocomial pathogens such as the Enterobacteriaceae group, Pseudomonas aeruginosa, and Acinetobacter baumannii, etc. is very serious considering these pathogens are resistant to almost all available antibacterial drugs. In this review article, the mode of action of the polymyxins and the genetic regulatory mechanism responsible for the emergence of resistance are discussed. Specifically, this review aims to update our current understanding in the field and suggest possible solutions that can be pursued for future antibiotic development. As polymyxins primarily target the bacterial membranes, resistance to polymyxins arises primarily by the modification of the lipopolysaccharides (LPS) in the outer membrane (OM). The LPS modification pathways are largely regulated by the bacterial two-component signal transduction (TCS) systems. Therefore, targeting or modulating the TCS signalling mechanisms can be pursued as an alternative to treat the infections caused by polymyxin-resistant MDR pathogens. In this review article, this aspect is also highlighted.
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Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla , Polimixinas/farmacologia , HumanosRESUMO
We identified a new microtubule targeted small molecule, which showed significant anticancer activity and induced apoptotic death of cancer cells. Precisely the central bridged carbonyl group and trifluoro-acetophenone group of a bis-benzothiazole molecule (BBT) interacts with tubulin close to the curcumin site and perturbs microtubule dynamics as well as causes microtubule depolymerization. We observed a significant enhancement of fluorescence while BBT interacts with the tubulin through bridged carbonyl moiety, a similar phenomenon to colchicine. Further, BBT activates tumor-suppressing bim and p53-puma axes to inhibit cancer survival. It also shows promising results against a tumor spheroid model. BBT is also capable of tumor regression, which shows that this molecule can serve as a potential template for the design of next-generation microtubule targeted anticancer drugs.
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Acetofenonas/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzotiazóis/farmacologia , Microtúbulos/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Acetofenonas/química , Antineoplásicos/síntese química , Antineoplásicos/química , Benzotiazóis/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Teoria da Densidade Funcional , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Células MCF-7 , Microtúbulos/metabolismo , Estrutura Molecular , Polimerização/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Tubulina (Proteína)/metabolismo , Células Tumorais CultivadasRESUMO
Here, we demonstrate an interesting strategy of modulating mitochondrial reactive oxygen species (ROS) using the organic electron acceptor molecule carbonyl-bridged bithiazole attached with bis-trifluoroacetophenone (BBT). This molecule was found to affect complex I activity. It has the propensity to bind close to the flavin mononucleotide site of complex I of mitochondria where it traps electron released from nicotinamide adenine dinucleotide (NADH) and elevates intracellular ROS, which suggests that the bridged carbonyl in BBT plays a crucial role in the acceptance of electron from NADH. We understand that the potential of the NADH/NAD+ redox couple and low-lying LUMO energy level of BBT are compatible with each other, thus favoring its entrapment of released electrons in complex I. This effect of BBT in ROS generation activates JNK and p38 stress-dependent pathways and resulted in mitochondrial-dependent apoptotic cell death with the reduction in expression of several important cyto-protecting factors (Hsp27 and NFκB), indicating its potential in inhibition of cancer cell relapse. Intriguingly, we found that BBT is not a P-glycoprotein substrate, which further reveals its excellent anticancer potential. This study enlightens us on how the power of electron acceptor ability became an emerging strategy for modulation of intracellular function.
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Neutrophils with immunoregulatory properties, also referred to as type-2 neutrophils (N2), myeloid-derived suppressor cells (MDSCs), or tumor-associated neutrophils (TANs), comprise a heterogeneous subset of cells that arise from unknown precursors in response to poorly understood cues. Here, we find that, in several models of liver autoimmunity, pharmacologically induced, autoantigen-specific T regulatory type-1 (TR1) cells and TR1-cell-induced B regulatory (Breg) cells use five immunoregulatory cytokines to coordinately recruit neutrophils into the liver and program their transcriptome to generate regulatory neutrophils. The liver-associated neutrophils from the treated mice, unlike their circulating counterparts or the liver neutrophils of sick mice lacking antigen-specific TR1 cells, are proliferative, can transfer disease protection to immunocompromised hosts engrafted with pathogenic effectors, and blunt antigen-presentation and local autoimmune responses via cathelin-related anti-microbial peptide (CRAMP), a cathelicidin, in a CRAMP-receptor-dependent manner. These results, thus, identify antigen-specific regulatory T cells as drivers of tissue-restricted regulatory neutrophil formation and CRAMP as an effector of regulatory neutrophil-mediated immunoregulation.
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Autoimunidade , Catelicidinas/metabolismo , Fígado/imunologia , Linfócitos T Reguladores/imunologia , Animais , Antígenos/metabolismo , Linfócitos B Reguladores/imunologia , Polaridade Celular/genética , Citocinas/metabolismo , Regulação da Expressão Gênica , Inflamação/patologia , Células de Kupffer/metabolismo , Fígado/patologia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Mitose/genética , Células Supressoras Mieloides/imunologia , Infiltração de Neutrófilos , Neutrófilos , Especificidade de Órgãos , Fenótipo , Transcrição GênicaRESUMO
In Caulobacter crescentus the combined action of chromosome replication and the expression of DNA methyl-transferase CcrM at the end of S-phase maintains a cyclic alternation between a full- to hemi-methylated chromosome. This transition of the chromosomal methylation pattern affects the DNA-binding properties of the transcription factor GcrA that controls the several key cell cycle functions. However, the molecular mechanism by which GcrA and methylation are linked to transcription is not fully elucidated yet. Using a combination of cell biology, genetics, and in vitro analysis, we deciphered how GcrA integrates the methylation pattern of several S-phase expressed genes to their transcriptional output. We demonstrated in vitro that transcription of ctrA from the P1 promoter in its hemi-methylated state is activated by GcrA, while in its fully methylated state GcrA had no effect. Further, GcrA and methylation together influence a peculiar distribution of creS transcripts, encoding for crescentin, the protein responsible for the characteristic shape of Caulobacter cells. This gene is duplicated at the onset of chromosome replication and the two hemi-methylated copies are spatially segregated. Our results indicated that GcrA transcribed only the copy where coding strand is methylated. In vitro transcription assay further substantiated this finding. As several of the cell cycle-regulated genes are also under the influence of methylation and GcrA-dependent transcriptional regulation, this could be a mechanism responsible for maintaining the gene transcription dosage during the S-phase.
Assuntos
Caulobacter crescentus/genética , Metilação de DNA/genética , Regulação Bacteriana da Expressão Gênica/genética , Transcrição Gênica/genética , DNA (Citosina-5-)-Metiltransferases/biossíntese , DNA (Citosina-5-)-Metiltransferases/genética , Proteínas de Ligação a DNA/genética , RNA Polimerases Dirigidas por DNA/genética , Regiões Promotoras Genéticas/genética , Fator sigma/genéticaRESUMO
Peptide MHC class II-based (pMHCII-based) nanomedicines trigger the formation of multicellular regulatory networks by reprogramming autoantigen-experienced CD4+ T cells into autoimmune disease-suppressing T regulatory type 1 (TR1) cells. We have shown that pMHCII-based nanomedicines displaying liver autoimmune disease-relevant yet ubiquitously expressed antigens can blunt various liver autoimmune disorders in a non-disease-specific manner without suppressing local or systemic immunity against infectious agents or cancer. Here, we show that such ubiquitous autoantigen-specific T cells are also awakened by extrahepatic tissue damage and that the corresponding TR1 progeny can suppress experimental autoimmune encephalomyelitis (EAE) and pancreatic ß cell autoreactivity. In mice having EAE, nanomedicines displaying either ubiquitous or CNS-specific epitopes triggered the formation and expansion of cognate TR1 cells and their recruitment to the CNS-draining lymph nodes, sparing their liver-draining counterparts. Surprisingly, in mice having both liver autoimmunity and EAE, liver inflammation sequestered these ubiquitous or even CNS-specific TR1 cells away from the CNS, abrogating their antiencephalitogenic activity. In these mice, only the ubiquitous antigen-specific TR1 cells suppressed liver autoimmunity. Thus, the scope of antigen spreading in autoimmune disorders is larger than previously anticipated, involving specificities expected to be silenced by mechanisms of tolerance; the regulatory activity, but not the retention of autoreactive TR1 cells, requires local autoantigen expression.
Assuntos
Autoimunidade , Encefalomielite Autoimune Experimental/imunologia , Hepatite Autoimune/imunologia , Fígado/imunologia , Linfócitos T Reguladores/imunologia , Animais , Autoantígenos/imunologia , Encefalomielite Autoimune Experimental/patologia , Hepatite Autoimune/patologia , Antígenos de Histocompatibilidade Classe II/imunologia , Fígado/patologia , Camundongos , Camundongos Endogâmicos NOD , Linfócitos T Reguladores/patologiaRESUMO
The design and development of an efficacious tumor-specific drug-delivery system is a challenging task. In this study, we have synthesized target-specific small peptide substrates on an octaguanidine sorbitol scaffold, named small molecular targeted drug-delivery conjugate (SMTDDC). The SMTDDC fabrication, with dual targeting cRGD and Cathepsin B (Cath B)-specific tripeptide (Glu-Lys-Phe), altered the microtubule network of glioblastoma cells by the orchestrated release of the cytotoxic paclitaxel (PTX). Cath B assisted PTX delivery was monitored by high-performance liquid chromatography and Surface-Enhanced Raman Scattering (SERS) modalities. The time-dependent SERS fingerprinting and imaging revealed a fast and accurate PTX release profile and subsequent in vitro cytotoxicity as well as the apoptotic events and microtubule network alteration in U-87 MG glioblastoma cells. Furthermore, SMTDDC displayed adequate stability under physiological conditions and demonstrated biocompatibility toward red blood cells and lymphocytes. This study indicated a new insight on SERS-guided peptidomimetic sorbitol molecular transporter, enabling a greater promise with high potential for the further development of PTX delivery in glioblastoma treatment.
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Antineoplásicos Fitogênicos , Glioblastoma , Antineoplásicos Fitogênicos/uso terapêutico , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos , Glioblastoma/tratamento farmacológico , Humanos , Paclitaxel/uso terapêuticoRESUMO
Discovery of a nontoxic fluorescent molecular probe to "light up" specific cellular organelles is extremely essential to understand dynamics of intracellular components. Here, we report a new nontoxic mitochondria-targeted linear bithiazole compound, containing trifluoroacetyl terminal groups, which emits intense blue fluorescence and stained mitochondria of various cells. Interestingly, the power of fluorescence is completely off when the bithiazole unit is stapled by a carbonyl bridge.
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Reconstitution of a complex biological structure or system following a simple and facile strategy using minimum physiochemical cues is challenging for an in-depth understanding of the system. In particular, the brain is a highly sophisticated and complex network of trillions of neurons and glial cells that controls function of our body. Understanding this complex machinery requires an innovative and simple bottom-up approach. In this venture, we report an easy and efficient strategy to culture cortical and hippocampal primary neurons from the E14-E16 embryo of Sprague-Dawley rat. This generates spontaneous neurospheres within 6-7 days of primary neuron culture of E14-E16 embryo. It further proliferates and forms radial glia-like structures, which are known to be the primary neural progenitor cells that differentiate into neurons, astrocytes, and oligodendrocytes. Interestingly, neurospheres lead to the formation of large projection neurons and radial glia, which mimic the early stage of cortical development in an in vivo system. Overall, this new, facile, strategic mixed primary neuron culture method offers a potential platform for understanding the effect of neurochemical modulators, which has tremendous future implications in the screening of neurotherapeutics.
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Técnicas de Cultura de Células/métodos , Córtex Cerebral/embriologia , Hipocampo/embriologia , Neurônios/citologia , Cultura Primária de Células , Animais , Córtex Cerebral/citologia , Embrião de Mamíferos , Células Ependimogliais/citologia , Hipocampo/citologia , Células-Tronco Neurais/citologia , Neurogênese , Ratos , Ratos Sprague-DawleyRESUMO
Streptococcus mutans, the primary aetiological agent of dental caries, is one of the major bacteria of the human oral cavity. The pathogenicity of this bacterium is attributed not only to the expression of virulence factors, but also to its ability to respond and adapt rapidly to the ever-changing conditions of the oral cavity. The two-component signal transduction system (TCS) CovR/S plays a crucial role in virulence and stress response in many streptococci. Surprisingly, in S. mutans the response regulator CovR appears to be an orphan, as the cognate sensor kinase, CovS, is absent in all the strains. We found that acetyl phosphate, an intracellular phosphodonor molecule known to act in signalling, might play a role in CovR phosphorylation in vivo. We also found that in vitro, upon phosphorylation by potassium phosphoramide (a high-energy phophodonor) CovR formed a dimer and showed altered electrophoretic mobility. As expected, we found that the conserved aspartic acid residue at position 53 (D53) was the site of phosphorylation, since neither phosphorylation nor dimerization was seen when an alanine-substituted CovR mutant (D53A) was used. Surprisingly, we found that the ability of CovR to act as a transcriptional regulator does not depend upon its phosphorylation status, since the D53A mutant behaved similarly to the wild-type protein in both in vivo and in vitro DNA-binding assays. This unique phosphorylation-mediated inhibition of CovR function in S. mutans sheds light on an unconventional mechanism of the signal transduction pathway.
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Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Streptococcus mutans/metabolismo , Fatores de Transcrição/metabolismo , Asparagina/genética , Asparagina/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Cárie Dentária/microbiologia , Mutação , Organofosfatos/metabolismo , Fosforilação , Ftalimidas/farmacologia , Regiões Promotoras Genéticas , Ligação Proteica , Multimerização Proteica/efeitos dos fármacos , Streptococcus mutans/genética , Fatores de Transcrição/química , Fatores de Transcrição/genéticaRESUMO
Microtubules regulate eukaryotic cell functions, which have tremendous implication in tumor progression. Thus, the design of novel approaches for controlling microtubule function is extremely important. In this manuscript, a novel tetrapeptide Ser-Leu-Arg-Pro (SLRP) has been designed and synthesized from a small peptide library consisting of 14 tetrapeptides, which perturbs microtubule function through interaction in the "anchor region". We have studied the role of peptides on microtubule function on a chemically functionalized 2D platform. Interestingly, we have found that SLRP binds with tubulin and inhibits the kinesin-driven microtubule motility on a kinesin-immobilized chemically functionalized 2D platform. Further, this peptide modulator interacts with intracellular tubulin/microtubule and depolymerizes the microtubule networks. These interesting findings of perturbation of microtubule function both on engineered platforms and inside the cell by this small peptide modulator inspired us to study the effect of this tetrapeptide on cancer cell proliferation. We found that the novel tetrapeptide modulator causes moderate cytotoxicity to the human breast cancer cell (MCF-7 cell), induces the apoptotic death of MCF-7 cell, and activates the tumor suppressor proteins p53 and cyclin-dependent kinase inhibitor 1 (p21). To the best of our knowledge, this is the shortest peptide discovered, which perturbs microtubule function both on an engineered 2D platform and inside the cell.
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Desenho de Fármacos , Microtúbulos/metabolismo , Oligopeptídeos/metabolismo , Tubulina (Proteína)/metabolismo , Apoptose/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Simulação de Acoplamento Molecular , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Ligação Proteica , Conformação Proteica , Proteína Supressora de Tumor p53/metabolismoRESUMO
Identification of key amino acids is required for development of efficient cell-penetrating peptides (CPPs) and has tremendous implications in medicine. Extensive research work has enlightened us about the importance of two amino acids, arginine and tryptophan, in cell penetration. Here, we present a top-down approach to show how spatial positions of two tryptophans regulate the cellular entry and nuclear localization. This enables us to develop short, non-toxic tetrapeptides with excellent potential for cell penetration and nuclear localization. Among them, Glu-Thr-Trp-Trp (ETWW) emerges as the most promising. Results suggest that it enters into cancer cells following an endocytic pathway and binds at the major groove of nuclear DNA, where successive tryptophan plays major role. We subsequently show that it is not a P-glycoprotein substrate and is non-toxic to PC12-derived neurons, suggesting its excellent potential as a CPP. Furthermore, its potential as a CPP is validated in multi-cellular 3D cell culture (spheroid) and in in vivo mice model. This study provides major fundamental insights about the positional importance of tryptophan and opens new avenues toward the development of next-generation CPPs and major-groove-specific anticancer drugs.
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Núcleo Celular/metabolismo , Peptídeos Penetradores de Células/metabolismo , Triptofano/metabolismo , Animais , Núcleo Celular/química , Peptídeos Penetradores de Células/química , Células Cultivadas , Humanos , Células MCF-7 , Camundongos , Células PC12 , Ratos , Triptofano/químicaRESUMO
Nitric oxide (NO), an endogenously produced free radical species, is an extremely important signalling molecule in several biochemical processes related to neurotransmission, neuronal communication, and vasodilation, to name a few. Other than relying on endogenous synthesis, intracellular NO delivery presents an interesting challenge to fully exploit the therapeutic potential of this gaseous molecule. We have applied a self-assembling peptide conjugate strategy to devise a construct carrying a NO-release arm, which can be activated under standard redox conditions. Consequently, a tryptophan-based peptide carrier was designed, which self-assembled in the solution phase to afford soft nanospherical structures, and released NO in Neuro2a cell line, resulting in neurite outgrowth.
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A novel neuro-compatible peptide-based hydrogel has been designed and developed, which contains microtubule stabilizing and neuroprotective short peptide. This hydrogel shows strong three-dimensional cross-linked fibrillary networks, which can capture water molecules. Interestingly, this hydrogel serves as excellent biocompatible soft material for 2D and 3D (neurosphere) neuron cell culture and provides stability of key cytoskeleton filaments such as microtubule and actin. Remarkably, it was observed that this hydrogel slowly enzymatically degrades and releases neuroprotective peptide, which promotes neurite outgrowth of neuron cell as well as exhibits excellent neuroprotection against anti-NGF-induced toxicity in neuron cells. Further, it can encapsulate anti-Alzheimer and anticancer hydrophobic drug curcumin, releases slowly, and inhibits significantly the growth of a 3D spheroid of neuron cancer cells. Thus, this novel neuroprotective hydrogel can be used for both neuronal cell transplantation for repairing brain damage as well as a delivery vehicle for neuroprotective agents, anti-Alzheimer, and anticancer molecules.
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Hidrogéis/química , Doença de Alzheimer , Humanos , Neuritos , Crescimento Neuronal , Neuroproteção , PeptídeosRESUMO
Microtubule dynamics play a crucial role in cancer cell division. Various drugs are developed to target microtubule. Although a few of them show potential in treatment of cancer, but success rate is limited due to their poor bioavailability and lack of specificity. Thus, development of highly bioavailable and target specific anticancer drug is extremely necessary. To address these key issues, here, a combination of approaches such as development of a dodecapeptide-docetaxel nanoassembly targeted to tubulin and MUC1 (mucin 1, cell surface associated glycoprotein) targeting oligonucleotide aptamer conjugated liposome for delivering peptide-docetaxel nanoassembly into the breast cancer cell have been demonstrated. These studies reveal that the peptide forms nanoassembly and entraps docetaxel drug. Further, the liposomal formulation of peptide-docetaxel exerts synergistic anticancer effect, activates key mitotic check point proteins, and inhibits bipolar spindle formation, metastatic cancer cell migration, and growth of tumor mimicking 3D multicellular spheroid.