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1.
Chembiochem ; 24(21): e202300418, 2023 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-37671979

RESUMO

Intramembrane serine proteases (rhomboid proteases) are involved in a variety of biological processes and are implicated in several diseases. Here, we report 4-oxo-ß-lactams as a novel scaffold for inhibition of rhomboids. We show that they covalently react with the active site and that the covalent bond is sufficiently stable for detection of the covalent rhomboid-lactam complex. 4-Oxo-ß-lactams may therefore find future use as both inhibitors and activity-based probes for rhomboid proteases.


Assuntos
Endopeptidases , beta-Lactamas , beta-Lactamas/farmacologia , Serina Proteases
2.
Mol Ther ; 30(10): 3284-3299, 2022 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-35765243

RESUMO

Existing evidence indicates that gut fungal dysbiosis might play a key role in the pathogenesis of colorectal cancer (CRC). We sought to explore whether reversing the fungal dysbiosis by terbinafine, an approved antifungal drug, might inhibit the development of CRC. A population-based study from Sweden identified a total of 185 patients who received terbinafine after their CRC diagnosis and found that they had a decreased risk of death (hazard ratio = 0.50) and metastasis (hazard ratio = 0.44) compared with patients without terbinafine administration. In multiple mouse models of CRC, administration of terbinafine decreased the fungal load, the fungus-induced myeloid-derived suppressor cell (MDSC) expansion, and the tumor burden. Fecal microbiota transplantation from mice without terbinafine treatment reversed MDSC infiltration and partially restored tumor proliferation. Mechanistically, terbinafine directly impaired tumor cell proliferation by reducing the ratio of nicotinamide adenine dinucleotide phosphate (NADP+) to reduced form of nicotinamide adenine dinucleotide phosphate (NADPH), suppressing the activity of glucose-6-phosphate dehydrogenase (G6PD), resulting in nucleotide synthesis disruption, deoxyribonucleotide (dNTP) starvation, and cell-cycle arrest. Collectively, terbinafine can inhibit CRC by reversing fungal dysbiosis, suppressing tumor cell proliferation, inhibiting fungus-induced MDSC infiltration, and restoring antitumor immune response.


Assuntos
Neoplasias Colorretais , Terbinafina , Animais , Antifúngicos , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Desoxirribonucleotídeos , Disbiose , Glucosefosfato Desidrogenase , Camundongos , NADP , Terbinafina/farmacologia
3.
Int J Mol Sci ; 24(3)2023 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-36768205

RESUMO

Aberrant miRNA expression has been associated with a large number of human diseases. Therefore, targeting miRNAs to regulate their expression levels has become an important therapy against diseases that stem from the dysfunction of pathways regulated by miRNAs. In recent years, small molecules have demonstrated enormous potential as drugs to regulate miRNA expression (i.e., SM-miR). A clear understanding of the mechanism of action of small molecules on the upregulation and downregulation of miRNA expression allows precise diagnosis and treatment of oncogenic pathways. However, outside of a slow and costly process of experimental determination, computational strategies to assist this on an ad hoc basis have yet to be formulated. In this work, we developed, to the best of our knowledge, the first cross-platform prediction tool, DeepsmirUD, to infer small-molecule-mediated regulatory effects on miRNA expression (i.e., upregulation or downregulation). This method is powered by 12 cutting-edge deep-learning frameworks and achieved AUC values of 0.843/0.984 and AUCPR values of 0.866/0.992 on two independent test datasets. With a complementarily constructed network inference approach based on similarity, we report a significantly improved accuracy of 0.813 in determining the regulatory effects of nearly 650 associated SM-miR relations, each formed with either novel small molecule or novel miRNA. By further integrating miRNA-cancer relationships, we established a database of potential pharmaceutical drugs from 1343 small molecules for 107 cancer diseases to understand the drug mechanisms of action and offer novel insight into drug repositioning. Furthermore, we have employed DeepsmirUD to predict the regulatory effects of a large number of high-confidence associated SM-miR relations. Taken together, our method shows promise to accelerate the development of potential miRNA targets and small molecule drugs.


Assuntos
Aprendizado Profundo , MicroRNAs , Neoplasias , Humanos , MicroRNAs/metabolismo , Neoplasias/metabolismo , Redes Reguladoras de Genes , Biologia Computacional
4.
ACS Chem Biol ; 18(4): 686-692, 2023 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-36920024

RESUMO

Aspartic proteases are a small class of proteases implicated in a wide variety of human diseases. Covalent chemical probes for photoaffinity labeling (PAL) of these proteases are underdeveloped. We here report a full on-resin synthesis of clickable PAL probes based on the natural product inhibitor pepstatin incorporating a minimal diazirine reactive group. The position of this group in the inhibitor determines the labeling efficiency. The most effective probes sensitively detect cathepsin D, a biomarker for breast cancer, in cell lysates. Moreover, through chemical proteomics experiments and deep learning algorithms, we identified sequestosome-1, an important player in autophagy, as a direct interaction partner and substrate of cathepsin D.


Assuntos
Ácido Aspártico Endopeptidases , Catepsina D , Pepstatinas , Marcadores de Fotoafinidade , Humanos , Ácido Aspártico Endopeptidases/química , Catepsina D/química , Diazometano , Pepstatinas/química , Pepstatinas/farmacologia , Marcadores de Fotoafinidade/química , Proteína Sequestossoma-1/química
5.
ACS Omega ; 8(28): 25487-25495, 2023 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-37483247

RESUMO

Photoaffinity labeling followed by tandem mass spectrometry is an often used strategy to identify protein targets of small-molecule drugs or drug candidates, which, under ideal conditions, enables the identification of the actual drug binding site. In the case of bioactive peptides, however, identifying the distinct binding site is hampered because of complex fragmentation patterns during tandem mass spectrometry. We here report the development and use of small cleavable photoaffinity reagents that allow functionalization of bioactive peptides for light-induced covalent binding to their protein targets. Upon cleavage of the covalently linked peptide drug, a chemical remnant of a defined mass remains on the bound amino acid, which is then used to unambiguously identify the drug binding site. Applying our approach to known peptide-drug/protein pairs with reported crystal structures, such as the calmodulin-melittin interaction, we were able to validate the identified binding sites based on structural models. Overall, our cleavable photoaffinity labeling strategy represents a powerful tool to enable the identification of protein targets and specific binding sites of a wide variety of bioactive peptides in the future.

6.
Cell Oncol (Dordr) ; 46(4): 1049-1067, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37012514

RESUMO

PURPOSE: Gastric cancer (GC) is a malignant tumour with high mortality, and liver metastasis is one of the main causes of poor prognosis. SLIT- and NTRK-like family member 4 (SLITRK4) plays an important role in the nervous system, such as synapse formation. Our study aimed to explore the functional role of SLITRK4 in GC and liver metastasis. METHODS: The mRNA level of SLITRK4 was evaluated using publicly available transcriptome GEO datasets and Renji cohort. The protein level of SLITRK4 in the tissue microarray of GC was observed using immunohistochemistry. Cell Counting Kit-8, colony formation, transwell migration assays in vitro and mouse model of liver metastasis in vivo was performed to investigate the functional roles of SLITRK4 in GC. Bioinformatics predictions and Co-IP experiments were applied to screen and identify SLITRK4-binding proteins. Western blot was performed to detect Tyrosine Kinase receptor B (TrkB)-related signaling molecules. RESULTS: By comparing primary and liver metastases from GC, SLITRK4 was found to be upregulated in tissues of GC with liver metastasis and to be closely related to poor clinical prognosis. SLITRK4 knockdown significantly abrogated the growth, invasion, and metastasis of GC in vitro and in vivo. Further study revealed that SLITRK4 could interact with Canopy FGF Signalling Regulator 3 (CNPY3), thus enhancing TrkB- related signaling by promoting the endocytosis and recycling of the TrkB receptor. CONCLUSION: In conclusion, the CNPY3-SLITRK4 axis contributes to liver metastasis of GC according to the TrkB-related signaling pathway. which may be a therapeutic target for the treatment of GC with liver metastasis.


Assuntos
Neoplasias Hepáticas , Neoplasias Gástricas , Animais , Camundongos , Neoplasias Gástricas/genética , Linhagem Celular Tumoral , Transdução de Sinais , Neoplasias Hepáticas/patologia , Endocitose , Proliferação de Células/genética
7.
Mol Omics ; 17(2): 197-209, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33507200

RESUMO

Covalent chemical probes are important tools in chemical biology. They range from post-translational modification (PTM)-derived metabolic probes, to activity-based probes and photoaffinity labels. Identification of the probe targets is often performed by tandem mass spectrometry-based proteomics methods. In the past fifteen years, cleavable linker technologies have been implemented in these workflows in order to identify probe targets with lower background and higher confidence. In addition, the linkers have enabled identification of modification sites. Overall, this has led to an increased knowledge of PTMs, enzyme function and drug action. This review gives an overview of the different types of cleavable linkers, and their benefits and limitations. Their applicability in target identification is also illustrated by several specific examples.


Assuntos
Marcadores de Fotoafinidade , Processamento de Proteína Pós-Traducional/genética , Proteoma/genética , Proteômica/tendências , Humanos , Espectrometria de Massas em Tandem
8.
J Mater Chem B ; 9(40): 8512-8517, 2021 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-34554170

RESUMO

Fluorescence imaging with high temporal and spatial resolution has emerged as one of the most promising techniques to monitor biomolecules and biological processes in living systems. Among many kinds of small molecular fluorescent dyes, 2,1,3-benzoxadiazole (BD) derivatives have been widely applied in many chemical and biological applications due to their excellent photophysical properties. However, only a limited number of BD dyes with long emission wavelengths were reported. Herein, we have reported a new class of red-to near-infrared-emitting small molecular dyes 2a-3a based on benzodioxazole scaffolds, which are named VBDfluors. To bathochromically shift both absorption and emission, the conjugation system was extended by introducing electron-withdrawing group-substituted vinyl groups at position 7 via a Knoevenagel condensation reaction. The basic photophysical properties of VBDfluors were detected and summarized. The VBDfluors display excellent photophysical properties, including emission in the red-to-NIR region, large Stokes shifts, good stability/photostability and cell permeability. The geometry of the molecules was optimized by density functional theory (DFT) and time-dependent DFT (TDDFT) methods. Bioimaging results indicated that 2a and 3a exhibited excellent cell permeability and could be utilized for visualization of lipid droplets in living cells.


Assuntos
Benzodioxóis/química , Corantes Fluorescentes/química , Corantes Fluorescentes/síntese química , Imagem Óptica , Teoria da Densidade Funcional , Células HeLa , Humanos , Concentração de Íons de Hidrogênio , Estrutura Molecular
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