RESUMO
Viruses require host cells to replicate and proliferate, which indicates that viruses hijack the cellular machinery. Human immunodeficiency virus type 1 (HIV-1) primarily infects CD4-positive T cells, and efficiently uses cellular proteins to replicate. Cells already have proteins that inhibit the replication of the foreign HIV-1, but their function is suppressed by viral proteins. Intriguingly, HIV-1 infection also changes the cellular metabolism to aerobic glycolysis. This phenomenon has been interpreted as a cellular response to maintain homeostasis during viral infection, yet HIV-1 efficiently replicates even in this environment. In this review, we discuss the regulatory role of glycolytic enzymes in viral replication and the impact of aerobic glycolysis on viral infection by introducing various host proteins involved in viral replication. Furthermore, we would like to propose a "glyceraldehyde-3-phosphate dehydrogenase-induced shock (G-shock) and kill strategy" that maximizes the antiviral effect of the glycolytic enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH) to eliminate latently HIV-1-infected cells.
Assuntos
Glicólise , Infecções por HIV , HIV-1 , Replicação Viral , Humanos , HIV-1/fisiologia , Glicólise/fisiologia , Infecções por HIV/virologia , Infecções por HIV/metabolismo , Infecções por HIV/imunologia , Gliceraldeído-3-Fosfato Desidrogenases/metabolismoRESUMO
Tigliane diterpenoids possess exceptionally complex structures comprising common 5/7/6/3-membered ABCD-rings and disparate oxygen functionalities. While tiglianes display a wide range of biological activities, compounds with HIV latency-reversing activity can eliminate viral reservoirs, thereby serving as promising leads for new anti-HIV agents. Herein, we report collective total syntheses of phorbol (13) and 11 tiglianes 14-24 with various acylation patterns and oxidation states, and their evaluation as HIV latency-reversing agents. The syntheses were strategically divided into five stages to increase the structural complexity. First, our previously established sequence enabled the expeditious preparation of ABC-tricycle 9 in 15 steps. Second, hydroxylation of 9 and ring-contractive D-ring formation furnished phorbol (13). Third, site-selective attachment of two acyl groups to 13 produced four phorbol diesters 14-17. Fourth, the oxygen functionalities were regio- and stereoselectively installed to yield five tiglianes 18-22. Fifth, further oxidation to the most densely oxygenated acerifolin A (23) and tigilanol tiglate (24) was realized through organizing a 3D shape of the B-ring. Assessment of the HIV latency-reversing activities of the 12 tiglianes revealed seven tiglianes (14-17 and 22-24) with 20- to 300-fold improved efficacy compared with prostratin (12), a representative latency-reversing agent. Therefore, the robust synthetic routes to a variety of tiglianes with promising activities devised in this study provide opportunities for advancing HIV eradication strategies.
Assuntos
Diterpenos , Infecções por HIV , Forbóis , Humanos , Latência Viral , OxigênioRESUMO
The human immunodeficiency virus type 1 (HIV-1) Gag protein is responsible for facilitating HIV-1 virion assembly and budding. Our study demonstrates that cardiolipin (CL), a component found in the inner mitochondrial membrane, exhibits the highest binding affinity to the N-terminal MA domain of the HIV-1 Gag protein within the lipid group of host cells. To assess this binding interaction, we synthesized short acyl chain derivatives of CL and employed surface plasmon resonance (SPR) analysis to determine the dissociation constants (Kd) for CL and the MA domain. Simultaneously, we examined the Kd of D-myo-phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2 ) derivatives, known to play a crucial role in virion formation. Among all the derivatives, Tetra-C7 -CL exhibited the lowest Kd value (Kd = 30.8 ± 6.9 µM) for MA binding on the CL analog-immobilized sensorchip, indicating a higher affinity. Similarly, the Kd value of Di-C7 -PIP2 (Kd = 36.6 ± 4.7 µM) was the lowest on the PI(4,5)P2 analog-immobilized sensorchip. Thus, Tetra-C7 -CL binds to the MA domain using a distinct binding mode while displaying a comparable binding affinity to Di-C7 -PIP2. This discovery holds significant implications for comprehending the virological importance of CL-MA domain binding, such as its subcellular distribution, including mitochondrial translocation, and involvement in viral particle formation in concert with PI(4,5)P2 . Furthermore, this study has the potential to contribute to the development of drugs in the future.
Assuntos
HIV-1 , Humanos , Membrana Celular/metabolismo , HIV-1/metabolismo , Cardiolipinas/análise , Cardiolipinas/metabolismo , Ligação Proteica , Produtos do Gene gag/análise , Produtos do Gene gag/metabolismoRESUMO
Nuclear localization signal (NLS) of HIV-1 integrase (IN) is implicated in nuclear import of HIV-1 preintegration complex (PIC). Here, we established a multiclass drug-resistant HIV-1 variant (HIVKGD) by consecutively exposing an HIV-1 variant to various antiretroviral agents including IN strand transfer inhibitors (INSTIs). HIVKGD was extremely susceptible to a previously reported HIV-1 protease inhibitor, GRL-142, with IC50 of 130 femtomolar. When cells were exposed to HIVKGD IN-containing recombinant HIV in the presence of GRL-142, significant decrease of unintegrated 2-LTR circular cDNA was observed, suggesting that nuclear import of PIC was severely compromised by GRL-142. X-ray crystallographic analyses revealed that GRL-142 interacts with NLS's putative sequence (DQAEHLK) and sterically blocks the nuclear transport of GRL-142-bound HIVKGD's PIC. Highly INSTI-resistant HIV-1 variants isolated from heavily INSTI-experienced patients proved to be susceptible to GRL-142, suggesting that NLS-targeting agents would serve as salvage therapy agents for highly INSTI-resistant variant-harboring individuals. The data should offer a new modality to block HIV-1 infectivity and replication and shed light on developing NLS inhibitors for AIDS therapy.
Assuntos
Integrase de HIV , HIV-1 , Humanos , Sinais de Localização Nuclear/genética , HIV-1/genética , Integrase de HIV/genética , AntiviraisRESUMO
Amamine (1), a new isoquinoline alkaloid, was isolated from the culture extract of an actinomycete Kitasatospora sp. HGTA304. The structure of 1 was determined by NMR and MS analyses in combination with UV data. Compound 1 displayed α-glucosidase inhibitory potential (IC50 value of 56 µM) compared with acarbose (IC50 value of 549 µM) as standard.
Assuntos
Alcaloides , Antineoplásicos , Streptomycetaceae , Inibidores de Glicosídeo Hidrolases/farmacologia , Inibidores de Glicosídeo Hidrolases/química , Alcaloides/farmacologia , Espectroscopia de Ressonância Magnética , Isoquinolinas/farmacologia , Estrutura MolecularRESUMO
Structurally simplified analogues of ansellone A, in which the decalin skeleton is replaced with a lipophilic chain, were prepared and their HIV latency-reversing activities biologically evaluated. In particular, two analogues bearing ether and alkenyl side chains, respectively, showed comparable activities to that of ansellone A. Each of the simplified compounds was easily synthesized using Prins cyclisation chemistry.
Assuntos
Infecções por HIV , Humanos , Relação Estrutura-AtividadeRESUMO
COVID-19 caused by SARS-CoV-2 has continually been serious threat to public health worldwide. While a few anti-SARS-CoV-2 therapeutics are currently available, their antiviral potency is not sufficient. Here, we identify two orally available 4-fluoro-benzothiazole-containing small molecules, TKB245 and TKB248, which specifically inhibit the enzymatic activity of main protease (Mpro) of SARS-CoV-2 and significantly more potently block the infectivity and replication of various SARS-CoV-2 strains than nirmatrelvir, molnupiravir, and ensitrelvir in cell-based assays employing various target cells. Both compounds also block the replication of Delta and Omicron variants in human-ACE2-knocked-in mice. Native mass spectrometric analysis reveals that both compounds bind to dimer Mpro, apparently promoting Mpro dimerization. X-ray crystallographic analysis shows that both compounds bind to Mpro's active-site cavity, forming a covalent bond with the catalytic amino acid Cys-145 with the 4-fluorine of the benzothiazole moiety pointed to solvent. The data suggest that TKB245 and TKB248 might serve as potential therapeutics for COVID-19 and shed light upon further optimization to develop more potent and safer anti-SARS-CoV-2 therapeutics.
Assuntos
Antivirais , COVID-19 , Proteases 3C de Coronavírus , Inibidores de Proteases , SARS-CoV-2 , Animais , Humanos , Camundongos , Antivirais/farmacologia , Benzotiazóis , Simulação de Acoplamento Molecular , Inibidores de Proteases/farmacologia , SARS-CoV-2/efeitos dos fármacos , Proteínas não Estruturais Virais/química , Proteases 3C de Coronavírus/antagonistas & inibidoresRESUMO
Supramolecular drug carriers are a promising approach for delivering anticancer drugs with high blood retention after administration. We previously synthesized folic acid-modified methyl-ß-cyclodextrin (FA-MßCD) as an anticancer drug. FA-MßCD has a selective autophagy-mediated antitumor effect on folic acid receptor (FR)-expressing cancer cells. Here, we enhanced the antitumor effect and safety of FA-MßCD by preparing a supramolecular nanoparticle formulation of FA-MßCD via host-guest interactions using an adamantane conjugate with human serum albumin (Ad-HSA). The Ad-HSA/FA-MßCD supramolecular complex prolonged the blood retention of FA-MßCD and improved its antitumor effect and safety after intravenous administration in tumor-bearing mice xenografted with FR-expressing cancer cells. These results suggest that the supramolecular technique using Ad-HSA is a promising approach for the delivery of CD-based anticancer drugs.
Assuntos
Adamantano , Antineoplásicos , Nanopartículas , Humanos , Animais , Camundongos , Ácido Fólico/farmacologia , Adamantano/farmacologia , AlbuminasRESUMO
The first total synthesis of marine sesterterpenoid ansellone G (2) was accomplished. This strategy utilizes the Prins cyclization reaction of a chloro-substituted homoallyl alcohol to synthesize the hydrobenzopyran skeleton. The preintroduction of the chloro groups facilitated the functional group transformation for 2 after constructing the carbon framework. Furthermore, we also successfully synthesized phorbadione (3) by dehydrating the tertiary alcohol. The HIV latency-reversing activity of the synthesized 2, 3, and deacetylated 2 was also evaluated.
Assuntos
Álcoois , CiclizaçãoRESUMO
Potent and biostable inhibitors of the main protease (Mpro) of SARS-CoV-2 were designed and synthesized based on an active hit compound 5h (2). Our strategy was based not only on the introduction of fluorine atoms into the inhibitor molecule for an increase of binding affinity for the pocket of Mpro and cell membrane permeability but also on the replacement of the digestible amide bond by a surrogate structure to increase the biostability of the compounds. Compound 3 is highly potent and blocks SARS-CoV-2 infection in vitro without a viral breakthrough. The derivatives, which contain a thioamide surrogate in the P2-P1 amide bond of these compounds (2 and 3), showed remarkably preferable pharmacokinetics in mice compared with the corresponding parent compounds. These data show that compounds 3 and its biostable derivative 4 are potential drugs for treating COVID-19 and that replacement of the digestible amide bond by its thioamide surrogate structure is an effective method.
RESUMO
Niemann-Pick disease type C (NPC) is characterized by the accumulation of glycolipids such as free cholesterol, sphingomyelin, and gangliosides in late endosomes/lysosomes (endolysosomes) due to abnormalities in the membrane proteins NPC1 or NPC2. The main symptoms of NPC caused by free cholesterol accumulation in various tissues vary depending on the time of onset, but hepatosplenomegaly and neurological symptoms accompanied by decreased motor, cognitive, and mental functions are observed in all age groups. However, the efficacy of NPC treatment remains limited. Herein, we have fabricated lactose-appended hydroxypropyl-ß-cyclodextrin (Lac-HPßCD) and evaluated its lowering effects on cholesterol accumulation in NPC model mice. We reveal that Lac-HPßCD lowers cholesterol accumulation in the liver and spleen by reducing the amount of free cholesterol. Moreover, Lac-HPßCD reduces the amount of free cholesterol in the cerebrum and slightly alleviates motor dysfunction. These results suggest that Lac-HPßCD has potential for the treatment of NPC.
Assuntos
Doença de Niemann-Pick Tipo C , 2-Hidroxipropil-beta-Ciclodextrina/farmacologia , Animais , Colesterol/metabolismo , Endossomos/metabolismo , Lactose/metabolismo , Camundongos , Doença de Niemann-Pick Tipo C/tratamento farmacológicoRESUMO
Three new diterpenes, stellejasmins A (1) and B (2) and 12-O-benzoylphorbol-13-heptanoate (3), were isolated from the roots of Stellera chamaejasme L. The structures of 1-3 were elucidated by extensive NMR and mass spectroscopic analyses. Compounds 1 and 2 are the first derivatives containing a hydroxy group at C-2 in the family of daphnane and tigliane diterpenes. The presence of a chlorine atom in 1 is unique in the plant metabolite. Compound 3 has an odd-number acyl group, which is biosynthetically notable. Human immunodeficiency virus (HIV) LTR-driven transcription activity was tested with 1-3 and 17 known diterpenes isolated from S. chamaejasme L. and Wikstroemia retusa A.Gray. Among these, gnidimacrin (4), stelleralide A (5), and wikstroelide A (20) were highly potent, with EC50 values of 0.14, 0.33, and 0.39 nM, respectively. The structure-activity relationship (SAR) was investigated using 20 natural and eight synthetic diterpenes. This is the first SAR study on natural daphnane and tigliane diterpenes.
Assuntos
Fármacos Anti-HIV/síntese química , Fármacos Anti-HIV/farmacologia , Diterpenos/síntese química , Diterpenos/farmacologia , HIV/efeitos dos fármacos , Forbóis/química , Latência Viral/efeitos dos fármacos , Diterpenos/química , Modelos Moleculares , Simulação de Acoplamento Molecular , Forbóis/farmacologia , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Raízes de Plantas/química , Relação Estrutura-Atividade , Thymelaeaceae/química , Wikstroemia/químicaRESUMO
BACKGROUND: The genome of human immunodeficiency virus type 1 (HIV-1) is encapsulated in a core consisting of viral capsid proteins (CA). After viral entry, the HIV-1 core dissociates and releases the viral genome into the target cell, this process is called uncoating. Uncoating of HIV-1 core is one of the critical events in viral replication and several studies show that host proteins positively or negatively regulate this process by interacting directly with the HIV-1 CA. RESULTS: Here, we show that arginyl-tRNA-protein transferase 1 (ATE1) plays an important role in the uncoating process by governing the optimal core stability. Yeast two-hybrid screening of a human cDNA library identified ATE1 as an HIV-1-CA-interacting protein and direct interaction of ATE1 with Pr55gag and p160gag - pol via HIV-1 CA was observed by cell-based pull-down assay. ATE1 knockdown in HIV-1 producer cells resulted in the production of less infectious viruses, which have normal amounts of the early products of the reverse transcription reaction but reduced amounts of the late products of the reverse transcription. Interestingly, ATE1 overexpression in HIV-1 producer cells also resulted in the production of poor infectious viruses. Cell-based fate-of-capsid assay, a commonly used method for evaluating uncoating by measuring core stability, showed that the amounts of pelletable cores in cells infected with the virus produced from ATE1-knockdown cells increased compared with those detected in the cells infected with the control virus. In contrast, the amounts of pelletable cores in cells infected with the virus produced from ATE1-overexpressing cells decreased compared with those detected in the cells infected with the control virus. CONCLUSIONS: These results indicate that ATE1 expression levels in HIV-1 producer cells contribute to the adequate formation of a stable HIV-1 core. These findings provide insights into a novel mechanism of HIV-1 uncoating and revealed ATE1 as a new host factor regulating HIV-1 replication.
Assuntos
Aminoaciltransferases/metabolismo , Capsídeo/química , Infecções por HIV/enzimologia , HIV-1/metabolismo , Aminoaciltransferases/genética , Capsídeo/metabolismo , Proteínas do Capsídeo/química , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Infecções por HIV/genética , Infecções por HIV/fisiopatologia , Infecções por HIV/virologia , HIV-1/química , HIV-1/genética , Interações Hospedeiro-Patógeno , Humanos , Ligação Proteica , Estabilidade Proteica , Replicação Viral , Desenvelopamento do VírusRESUMO
Metformin is widely used for the treatment of type 2 diabetes, and increasing numbers of studies have shown that metformin also ameliorates tumor progression, inflammatory disease, and fibrosis. However, the ability of metformin to improve non-diabetic glomerular disease and chronic kidney disease (CKD) has not been explored. To investigate the effect of metformin on non-diabetic glomerular disease, we used a mouse model of Alport syndrome (Col4a5 G5X) which were treated with metformin or losartan, used as a control treatment. We also investigated the effect of metformin on adriamycin-induced glomerulosclerosis model. Pathological and biochemical analysis showed that metformin or losartan suppressed proteinuria, renal inflammation, fibrosis, and glomerular injury and extended the lifespan in Alport syndrome mice. Transcriptome analysis showed that metformin and losartan influenced molecular pathways-related to metabolism and inflammation. Metformin altered multiple genes including metabolic genes not affected by losartan. Metformin also suppressed proteinuria and glomerular injury in the adriamycin-induced glomerulosclerosis mouse model. Our results showed that metformin ameliorates the glomerular sclerosis and CKD phenotype in non-diabetic chronic glomerular diseases. Metformin may have therapeutic potential for not only diabetic nephropathy but also non-diabetic glomerular disease including Alport syndrome.
Assuntos
Hipoglicemiantes/uso terapêutico , Metformina/uso terapêutico , Nefrite Hereditária/tratamento farmacológico , Animais , Colágeno Tipo IV/genética , Nefropatias Diabéticas/fisiopatologia , Nefropatias Diabéticas/prevenção & controle , Modelos Animais de Doenças , Rim/metabolismo , Camundongos , Nefrite Hereditária/genética , Nefrite Hereditária/fisiopatologia , Fenótipo , Índice de Gravidade de Doença , Transdução de Sinais , TranscriptomaRESUMO
The cellular environment affects optimal viral replication because viruses cannot replicate without their host cells. In particular, metabolic resources such as carbohydrates, lipids, and ATP are crucial for viral replication, which is sensitive to cellular metabolism. Intriguingly, recent studies have demonstrated that human immunodeficiency virus type 1 (HIV-1) infection induces a metabolic shift from oxidative phosphorylation to aerobic glycolysis in CD4+ T cells to produce the virus efficiently. However, the importance of aerobic glycolysis in maintaining the quality of viral components and viral infectivity has not yet been fully investigated. Here, we show that aerobic glycolysis is necessary not only to override the inhibitory effect of virion-incorporated glycolytic enzymes, but also to maintain the enzymatic activity of reverse transcriptase and the adequate packaging of envelope proteins into HIV-1 particles. To investigate the effect of metabolic remodeling on the phenotypic properties of HIV-1 produced by infected cells, we replaced glucose with galactose in the culture medium because the cells grown in galactose-containing medium are forced to carry out oxidative metabolism instead of aerobic glycolysis. We found that the packaging levels of glyceraldehyde 3-phosphate dehydrogenase, alpha-enolase and pyruvate kinase muscle type 2, which decrease HIV-1 infectivity by packaging into viral particles, are increased in progeny viruses produced by the cells grown in galactose-containing medium. Furthermore, we found that the entry and reverse transcription efficiency of the progeny viruses were reduced, which was caused by a decrease in the enzymatic activity of reverse transcriptase in the viral particles and a decrease in the packaging levels of envelope proteins and reverse transcriptase. These results indicate that the aerobic glycolysis environment in HIV-1-infected cells may contribute to the quality control of viruses.
Assuntos
Glucose/metabolismo , Glicólise , HIV-1/patogenicidade , Vírion/metabolismo , Aerobiose/efeitos dos fármacos , Biomarcadores Tumorais/metabolismo , Proteínas de Transporte/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Meios de Cultura , Proteínas de Ligação a DNA/metabolismo , Galactose/farmacologia , Produtos do Gene env/metabolismo , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/metabolismo , Glicólise/efeitos dos fármacos , HIV-1/efeitos dos fármacos , HIV-1/genética , Humanos , Proteínas de Membrana/metabolismo , Fosfopiruvato Hidratase/metabolismo , Transcrição Reversa/efeitos dos fármacos , Transcrição Reversa/genética , Hormônios Tireóideos/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Empacotamento do Genoma Viral/efeitos dos fármacos , Proteínas de Ligação a Hormônio da TireoideRESUMO
The total synthesis and biological evaluation of the marine sesterterpenoid ansellone A (1), an HIV latency-reversing agent, and its analogues are reported. The key to the success of this synthetic route is a Prins cyclization reaction enabled by the strategic use of the TfO group for stabilization of the acid-labile tertiary allylic alcohol. The SAR study found the alcohol analogue to exhibit more potent activity than 1.
Assuntos
Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Sesterterpenos/síntese química , Ciclização , HIV-1/química , HIV-1/fisiologia , Estrutura Molecular , Sesterterpenos/químicaRESUMO
Except remdesivir, no specific antivirals for SARS-CoV-2 infection are currently available. Here, we characterize two small-molecule-compounds, named GRL-1720 and 5h, containing an indoline and indole moiety, respectively, which target the SARS-CoV-2 main protease (Mpro). We use VeroE6 cell-based assays with RNA-qPCR, cytopathic assays, and immunocytochemistry and show both compounds to block the infectivity of SARS-CoV-2 with EC50 values of 15 ± 4 and 4.2 ± 0.7 µM for GRL-1720 and 5h, respectively. Remdesivir permitted viral breakthrough at high concentrations; however, compound 5h completely blocks SARS-CoV-2 infection in vitro without viral breakthrough or detectable cytotoxicity. Combination of 5h and remdesivir exhibits synergism against SARS-CoV-2. Additional X-ray structural analysis show that 5h forms a covalent bond with Mpro and makes polar interactions with multiple active site amino acid residues. The present data suggest that 5h might serve as a lead Mpro inhibitor for the development of therapeutics for SARS-CoV-2 infection.
Assuntos
Tratamento Farmacológico da COVID-19 , Inibidores de Protease de Coronavírus/farmacologia , SARS-CoV-2/efeitos dos fármacos , Proteases Virais/efeitos dos fármacos , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/farmacologia , Alanina/análogos & derivados , Alanina/farmacologia , Animais , Antivirais/farmacologia , Linhagem Celular , Chlorocebus aethiops , Humanos , Indóis/farmacologia , Piridinas/farmacologia , Células Vero , Proteases Virais/metabolismoRESUMO
One compound sometimes shows two biological functions, becoming important aspect of recent drug discovery. This study began with an attempt to confirm the previously reported molecular mechanism of the anti-human immunodeficiency virus (HIV) heterocyclic compound BMMP [2-(benzothiazol-2-ylmethylthio)-4-methylpyrimidine], i.e., induction of abnormal uncoating of the viral core at the post-entry step. Our mechanistic study gave results consistent with this mechanism. We further attempted to find out the molecular target of BMMP by a pulldown approach using previously synthesized biotinylated BMMP (Biotin-BMMP) and successfully identified heterogenous nuclear ribonucleoprotein M (hnRNP M) as a BMMP-binding protein. This protein was found not to be accountable for the anti-HIV activity of BMMP. As hnRNP M has been reported to promote cancer metastasis, we tested this mechanism and found that BMMP suppressed migration of the human lung carcinoma cell line A549 stimulated with transforming growth factor-ß (TGF-ß). Mechanistic study showed that BMMP suppressed the expression of CD44 mRNA via the regulation of hnRNP M. Furthermore, six new derivatives of BMMP were synthesized, and the patterns of their activities against HIV-1 and cell migration were not uniform, suggesting that the anti-HIV mechanism and the anti-cell migration mechanism of BMMP are independent. Taken together, the anti-cell migration activity of the anti-HIV heterocyclic compound BMMP was newly discovered by identification of its binding protein hnRNP M using a chemical biology approach.