RESUMEN
Polyomaviruses are species-specific DNA viruses that can cause disease in immunocompromised individuals. Despite their role as the causative agents for several diseases, there are no currently approved antivirals for treating polyomavirus infection. Brincidofovir (BCV) is an antiviral approved for the treatment of poxvirus infections and has shown activity against other double-stranded DNA viruses. In this study, we tested the efficacy of BCV against polyomavirus infection in vitro and in vivo using mouse polyomavirus (MuPyV). BCV inhibited virus production in primary mouse kidney cells and brain cortical cells. BCV treatment of cells transfected with MuPyV genomic DNA resulted in a reduction in virus levels, indicating that viral inhibition occurs post-entry. Although in vitro BCV treatment had a limited effect on viral DNA and RNA levels, drug treatment was associated with a reduction in viral protein, raising the possibility that BCV acts post-transcriptionally to inhibit MuPyV infection. In mice, BCV treatment was well tolerated, and prophylactic treatment resulted in a reduction in viral DNA levels and a potent suppression of infectious virus production in the kidney and brain. In mice with chronic polyomavirus infection, therapeutic administration of BCV decreased viremia and reduced infection in the kidney. These data demonstrate that BCV exerts antiviral activity against polyomavirus infection in vivo, supporting further investigation into the use of BCV to treat clinical polyomavirus infections. IMPORTANCE: Widespread in the human population and able to persist asymptomatically for the life of an individual, polyomavirus infections cause a significant disease burden in the immunocompromised. Individuals undergoing immune suppression, such as kidney transplant patients or those treated for autoimmune diseases, are particularly at high risk for polyomavirus-associated diseases. Because no antiviral agent exists for treating polyomavirus infections, management of polyomavirus-associated diseases typically involves reducing or discontinuing immunomodulatory therapy. This can be perilous due to the risk of transplant rejection and the potential development of adverse immune reactions. Thus, there is a pressing need for the development of antivirals targeting polyomaviruses. Here, we investigate the effects of brincidofovir, an FDA-approved antiviral, on polyomavirus infection in vivo using mouse polyomavirus. We show that the drug is well-tolerated in mice, reduces infectious viral titers, and limits viral pathology, indicating the potential of brincidofovir as an anti-polyomavirus therapeutic.
Asunto(s)
Antivirales , Citosina , Organofosfonatos , Infecciones por Polyomavirus , Poliomavirus , Animales , Citosina/análogos & derivados , Citosina/farmacología , Citosina/uso terapéutico , Infecciones por Polyomavirus/tratamiento farmacológico , Infecciones por Polyomavirus/virología , Poliomavirus/efectos de los fármacos , Ratones , Antivirales/farmacología , Antivirales/uso terapéutico , Organofosfonatos/farmacología , Organofosfonatos/uso terapéutico , Replicación Viral/efectos de los fármacos , Riñón/virología , Riñón/efectos de los fármacos , Femenino , ADN Viral/genética , Células Cultivadas , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Encéfalo/virologíaRESUMEN
Human adenoviruses can cause serious, disseminated infections in immunocompromised patients. For pediatric allogeneic stem cell transplant patients, the case fatality rate can reach 80%. Still, there is no available antiviral drug that is specifically approved by the Food and Drug Administration for the treatment of adenovirus infections. To fill this pressing medical need, we have developed NPP-669, a prodrug of cidofovir with broad activity against double-stranded DNA viruses, including adenoviruses. Here, we report on the in vivo anti-adenoviral efficacy of NPP-669. Using the immunosuppressed Syrian hamster as the model, we show that NPP-669 is highly efficacious when dosed orally at 1 mg/kg and 3 mg/kg. In a delayed administration experiment, NPP-669 was more effective than brincidofovir, a similar compound that reached Phase III clinical trials. Furthermore, parenteral administration of NPP-669 increased its efficacy approximately 10-fold compared to oral dosing without apparent toxicity, suggesting that this route may be preferable in a hospital setting. Based on these findings, we believe that NPP-669 is a promising new compound that needs to be further investigated.
Asunto(s)
Antivirales , Cidofovir , Citosina , Mesocricetus , Organofosfonatos , Profármacos , Animales , Antivirales/farmacología , Antivirales/uso terapéutico , Profármacos/farmacología , Profármacos/uso terapéutico , Humanos , Cidofovir/farmacología , Cidofovir/uso terapéutico , Organofosfonatos/farmacología , Organofosfonatos/uso terapéutico , Citosina/análogos & derivados , Citosina/farmacología , Citosina/uso terapéutico , Adenovirus Humanos/efectos de los fármacos , Infecciones por Adenovirus Humanos/tratamiento farmacológico , Infecciones por Adenovirus Humanos/virología , Modelos Animales de Enfermedad , Cricetinae , Administración OralRESUMEN
Mpox virus has wildly spread over 108 non-endemic regions in the world since May 2022. DNA replication of mpox is performed by DNA polymerase machinery F8-A22-E4, which is known as a great drug target. Brincidofovir and cidofovir are reported to have broad-spectrum antiviral activity against poxviruses, including mpox virus in animal models. However, the molecular mechanism is not understood. Here we report cryogenic electron microscopy structures of mpox viral F8-A22-E4 in complex with a DNA duplex, or dCTP and the DNA duplex, or cidofovir diphosphate and the DNA duplex at resolution of 3.22, 2.98 and 2.79 Å, respectively. Our structural work and DNA replication inhibition assays reveal that cidofovir diphosphate is located at the dCTP binding position with a different conformation to compete with dCTP to incorporate into the DNA and inhibit DNA synthesis. Conformation of both F8-A22-E4 and DNA is changed from the pre-dNTP binding state to DNA synthesizing state after dCTP or cidofovir diphosphate is bound, suggesting a coupling mechanism. This work provides the structural basis of DNA synthesis inhibition by brincidofovir and cidofovir, providing a rational strategy for new therapeutical development for mpox virus and other pox viruses.
Asunto(s)
Antivirales , Cidofovir , Citosina , Replicación del ADN , Organofosfonatos , Replicación Viral , Cidofovir/farmacología , Cidofovir/química , Organofosfonatos/farmacología , Organofosfonatos/química , Citosina/análogos & derivados , Citosina/farmacología , Citosina/química , Replicación del ADN/efectos de los fármacos , Humanos , Antivirales/farmacología , Antivirales/química , Replicación Viral/efectos de los fármacos , ADN Viral , Modelos MolecularesRESUMEN
Adenovirus infections of immunocompromised humans are a significant source of morbidity and mortality. Presently, there is no drug specifically approved for the treatment of adenovirus infections by the FDA. The state-of-the-art treatment of such infections is the off-label use of cidofovir, an acyclic nucleotide phosphonate. While cidofovir inhibits adenovirus replication, it has dose-limiting kidney toxicity. There is an apparent need for a better compound to treat adenovirus infections. To this end, we have been developing acyclic nucleotide phosphonate prodrugs that utilize an amino acid scaffold equipped with a lipophilic modifier. Here, we compare the antiviral potential of two prodrugs of HPMPA that differ only in the amino acid-based promoiety: USC-087, based on an N-hexadecyl tyrosinamide, and USC-093, based on an N-hexadecyl serinamide. Oral administration of both compounds was very efficacious against disseminated HAdV-C6 infection in immunosuppressed Syrian hamsters, suppressing virus replication and mitigating pathology even when treatment was withheld until 4 days after challenge. We saw only marginal efficacy after respiratory infection of hamsters, which may reflect suboptimal distribution to the lung. Importantly, neither compound induced intestinal toxicity, which was observed as the major adverse effect in clinical trials of brincidofovir, a prodrug of cidofovir which also contains a C-16 modifier. Notably, we found that there was a significant difference in the nephrotoxicity of the two compounds: USC-087 caused significant kidney toxicity while USC-093 did not, at effective doses. These findings will be valuable guidepoints in the future evolution of this new class of potential prodrugs to treat adenovirus infections.
Asunto(s)
Adenina/análogos & derivados , Infecciones por Adenoviridae , Infecciones por Adenovirus Humanos , Organofosfonatos , Profármacos , Tirosina/análogos & derivados , Cricetinae , Animales , Humanos , Infecciones por Adenovirus Humanos/tratamiento farmacológico , Cidofovir/farmacología , Cidofovir/uso terapéutico , Mesocricetus , Antivirales/uso terapéutico , Profármacos/farmacología , Profármacos/uso terapéutico , Adenoviridae , Replicación Viral , Organofosfonatos/farmacología , Organofosfonatos/uso terapéutico , Infecciones por Adenoviridae/tratamiento farmacológico , Citosina/farmacología , Citosina/uso terapéutico , Aminoácidos/farmacología , Nucleótidos/uso terapéuticoRESUMEN
Adenoviruses are the major cause of ocular viral infections worldwide. Currently, there is no approved antiviral treatment for these eye infections. Cyclopentenylcytosine (CPE-C) is an antiviral that has demonstrated activity against more than 20 viruses. The goals of the current study were to determine the in vitro and in vivo antiviral activity of CPE-C as well as its ocular toxicity. Antiviral activity was evaluated in vitro using standard plaque reduction assays to determine the 50% effective concentrations (EC50s) and in vivo in the Ad5/NZW rabbit ocular replication model. Ocular toxicity was determined in uninfected rabbit eyes following topical ocular application. The in vitro EC50s for CPE-C ranged from 0.03 to 0.059 µg/mL for nine adenovirus types that commonly infect the eye. Ocular toxicity testing determined CPE-C to be non-irritating or practically non-irritating by Draize scoring. In vivo, 3% CPE-C topically administered 4X or 2X daily for 7 days to adenovirus-infected eyes demonstrated effective antiviral activity compared with the negative control and comparable antiviral activity to the positive control, 0.5% cidofovir, topically administered twice daily for 7 days. We conclude CPE-C was relatively non-toxic to rabbit eyes and demonstrated potent anti-adenoviral activity in vitro and in vivo.
Asunto(s)
Infecciones por Adenoviridae , Adenovirus Humanos , Infecciones del Ojo , Organofosfonatos , Animales , Conejos , Antivirales/uso terapéutico , Organofosfonatos/farmacología , Neuropatía Óptica Tóxica/tratamiento farmacológico , Citosina/farmacología , Infecciones por Adenoviridae/tratamiento farmacológico , Adenoviridae , Infecciones del Ojo/tratamiento farmacológico , Replicación ViralRESUMEN
The double-stranded DNA (dsDNA) viruses represented by adenovirus and monkeypox virus, have attracted widespread attention due to their high infectivity. In 2022, the global outbreak of mpox (or monkeypox) has led to the declaration of a Public Health Emergency of International Concern. However, to date therapeutics approved for dsDNA virus infections remain limited and there are still no available treatments for some of these diseases. The development of new therapies for treating dsDNA infection is in urgent need. In this study, we designed and synthesized a series of novel disulfide-incorporated lipid conjugates of cidofovir (CDV) as potential candidates against dsDNA viruses including vaccinia virus (VACV) and adenovirus (AdV) 5. The structure-activity relationship analyses revealed that the optimum linker moiety was C2H4 and the optimum aliphatic chain length was 18 or 20 atoms. Among the synthesized conjugates, 1c exhibited more potency against VACV (IC50 = 0.0960 µM in Vero cells; IC50 = 0.0790 µM in A549 cells) and AdV5 (IC50 = 0.1572 µM in A549 cells) than brincidofovir (BCV). The transmission electron microscopy (TEM) images revealed that the conjugates could form micelles in phosphate buffer. The stability studies in the GSH environment demonstrated that the formation of micelles in phosphate buffer might protect the disulfide bond from glutathione (GSH) reduction. The dominant means of the synthetic conjugates to liberate the parent drug CDV was by enzymatic hydrolysis. Furthermore, the synthetic conjugates remained sufficiently stable in simulated gastric fluid (SGF), simulated intestinal fluid (SIF), and pooled human plasma, which indicated the possibility for oral administration. These results indicated 1c may be a broad-spectrum antiviral candidate against dsDNA viruses with potential oral administration. Moreover, modification of the aliphatic chain attached to the nucleoside phosphonate group was involved as an efficient prodrug strategy for the development of potent antiviral candidates.
Asunto(s)
Antivirales , Profármacos , Animales , Chlorocebus aethiops , Humanos , Cidofovir/farmacología , Antivirales/química , Profármacos/farmacología , Células Vero , Micelas , Citosina/farmacología , Citosina/química , Virus Vaccinia , Lípidos , FosfatosRESUMEN
Mpox virus is an emergent human pathogen. While it is less lethal than smallpox, it can still cause significant morbidity and mortality. In this review, we explore 3 antiviral agents with activity against mpox and other orthopoxviruses: cidofovir, brincidofovir, and tecovirimat. Cidofovir, and its prodrug brincidofovir, are inhibitors of DNA replication with a broad spectrum of activity against multiple families of double-stranded DNA viruses. Tecovirimat has more specific activity against orthopoxviruses and inhibits the formation of the extracellular enveloped virus necessary for cell-to-cell transmission. For each agent, we review basic pharmacology, data from animal models, and reported experience in human patients.
Asunto(s)
Antivirales , Mpox , Organofosfonatos , Animales , Humanos , Antivirales/farmacología , Antivirales/uso terapéutico , Cidofovir , Citosina/farmacología , Organofosfonatos/farmacología , Mpox/tratamiento farmacológico , Monkeypox virus/efectos de los fármacosRESUMEN
African swine fever (ASF) is an acute, hemorrhagic, and devastating viral infectious disease that causes important economic losses to the swine industry. Currently, there are no effective vaccines or drugs available. Epigenetic mechanisms, especially cytosine methylation of cytosine- -phosphate-guanine (CpG) islands, have a significant impact on the life cycle of several viruses. Hence, drugs targeting DNA methylation may potentially be used for the treatment of ASF. Here, we selected the inner core, core shell, inner membrane, capsid, and external envelope membrane, to analyze the characteristics of CpG islands in the ASF virus (ASFV) genomes. Furthermore, we analyzed the promoters and CpG islands in the upstream regions of these genes. Results showed that the CpG islands of seven genes were conserved in the genomes of two genotype of ASFV strains, whereas the CpG islands of other genes were relatively conserved (ASFV strains differed mainly in the quantity of CpG islands). The different distribution of CpG islands in the genomes of different ASFV strains may affect their methylation status, which may in turn affect the regulation of viral gene expression, leading to different clinical outcomes. In addition, the predicted promoter regions based on the upstream sequences of most genes overlapped with CpG island positions. Methylation of the binding sites of the promoter regions inhibits the binding of the transcription factors to the promoters, thus inhibiting the activation of the promoters and limiting the synthesis of viral proteins. The results of this study provide a basis for exploring new antiviral therapeutic strategies from an epigenetic perspective.
Asunto(s)
Virus de la Fiebre Porcina Africana , Fiebre Porcina Africana , Enfermedades de los Porcinos , Virus de la Fiebre Porcina Africana/genética , Animales , Antivirales/farmacología , Islas de CpG , Citosina/metabolismo , Citosina/farmacología , Genotipo , Guanina/metabolismo , Guanina/farmacología , Fosfatos , Porcinos , Enfermedades de los Porcinos/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Factores de Transcripción/farmacología , Proteínas Virales/metabolismoRESUMEN
Selective inhibition of targeted protein kinases is an effective therapeutic approach for treatment of human malignancies, which interferes phosphorylation of cellular substrates. However, a drug-imposed selection creates pressures for tumor cells to acquire chemoresistance-conferring mutations or activating alternative pathways, which can bypass the inhibitory effects of kinase inhibitors. Thus, identifying downstream phospho-substrates conferring drug resistance is of great importance for developing poly-pharmacological and targeted therapies. To identify functional phosphorylation sites involved in 5-fluorouracil (5-FU) resistance during its treatment of colorectal cancer cells, CRISPR-mediated cytosine base editor (CBE) and adenine base editor (ABE) are utilized for functional screens by mutating phosphorylated amino acids with two libraries specifically targeting 7779 and 10 149 phosphorylation sites. Among the top enriched gRNAs-induced gain-of-function mutants, the target genes are involved in cell cycle and post-translational covalent modifications. Moreover, several substrates of RSK2 and PAK4 kinases are discovered as main effectors in responding to 5-FU chemotherapy, and combinational treatment of colorectal cancer cells with 5-FU and RSK2 inhibitor or PAK4 inhibitor can largely inhibit cell growth and enhance cell apoptosis through a RSK2/TP53BP1/γ-H2AX phosphorylation signaling axis. It is proposed that this screen approach can be used for functional phosphoproteomics in chemotherapy of various human diseases.
Asunto(s)
Neoplasias Colorrectales , Resistencia a Antineoplásicos , Humanos , Resistencia a Antineoplásicos/genética , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Fluorouracilo/farmacología , Fluorouracilo/uso terapéutico , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Proteínas Quinasas/farmacología , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Adenina/farmacología , Adenina/uso terapéutico , Aminoácidos/genética , Aminoácidos/farmacología , Aminoácidos/uso terapéutico , Citosina/farmacología , Citosina/uso terapéutico , Quinasas p21 Activadas/genética , Quinasas p21 Activadas/metabolismo , Quinasas p21 Activadas/farmacologíaRESUMEN
Intratumoural administration of unmethylated cytosine-phosphate-guanine motifs (CpG) to stimulate toll-like receptor (TLR)-9 has been shown to induce tumour regression in preclinical studies and some efficacy in the clinic. Because activated natural killer T (NKT) cells can cooperate with pattern-recognition via TLRs to improve adaptive immune responses, we assessed the impact of combining a repeated dosing regimen of intratumoural CpG with a single intratumoural dose of the NKT cell agonist α-galactosylceramide (α-GalCer). The combination was superior to CpG alone at inducing regression of established tumours in several murine tumour models, primarily mediated by CD8+ T cells. An antitumour effect on distant untreated tumours (abscopal effect) was reliant on sustained activity of NKT cells and was associated with infiltration of KLRG1+ NKT cells in tumours and draining lymph nodes at both injected and untreated distant sites. Cytometric analysis pointed to increased exposure to type I interferon (IFN) affecting many immune cell types in the tumour and lymphoid organs. Accordingly, antitumour activity was lost in animals in which dendritic cells (DCs) were incapable of signaling through the type I IFN receptor. Studies in conditional ablation models showed that conventional type 1 DCs and plasmacytoid DCs were required for the response. In tumour models where the combined treatment was less effective, the addition of tumour-antigen derived peptide, preferably conjugated to α-GalCer, significantly enhanced the antitumour response. The combination of TLR ligation, NKT cell agonism, and peptide delivery could therefore be adapted to induce responses to both known and unknown antigens.
Asunto(s)
Células T Asesinas Naturales , Neoplasias , Animales , Linfocitos T CD8-positivos , Citosina/metabolismo , Citosina/farmacología , Guanina/metabolismo , Guanina/farmacología , Interferón gamma , Células Asesinas Naturales/metabolismo , Activación de Linfocitos , Ratones , Células T Asesinas Naturales/metabolismo , Neoplasias/tratamiento farmacológico , Fosfatos/metabolismo , Fosfatos/farmacologíaRESUMEN
A series of cytosine derivatives containing a sulfonamide moiety were designed and synthesized, and their antiviral activities against pepper mild mottle virus (PMMoV) were systematically evaluated. Then, a three-dimensional quantitative structure-activity relationship (3D-QSAR) model was constructed to study the structure-activity relationship according to the pEC50 of the compounds' protective activities. Next, compound A32 with preferable antiviral activity on PMMoV was obtained based on the CoMSIA and CoMFA models, with an EC50 of 19.5 µg/mL, which was superior to the template molecule A25 (21.3 µg/mL) and ningnanmycin (214.0 µg/mL). In addition, further studies showed that the antiviral activity of compound A32 against PMMoV was in accord with the up-regulation of proteins expressed in the defense response and carbon fixation in photosynthetic organisms. These results indicated that cytosine derivatives containing a sulfonamide moiety could be used as novel potential antiviral agents for further research and development.
Asunto(s)
Antivirales , Virus del Mosaico del Tabaco , Antivirales/farmacología , Citosina/farmacología , Diseño de Fármacos , Relación Estructura-Actividad Cuantitativa , Relación Estructura-Actividad , Sulfonamidas/farmacologíaRESUMEN
Cytosine-phosphate-guanine oligodeoxynucleotides (CpG-ODNs), which exist in vertebrate, bacterial, and viral genomes, are regarded as strong immune adjuvants. To date, the biological activities of CpG-ODNs in reproduction remain unknown. Here, we investigated the effects of CpG-ODNs on the cell cycle, apoptosis, and steroidogenesis in mouse granulosa cells (mGCs), in combination with inhibin alpha (1 ~ 32) fragments. mGCs were transfected with pEGFP (containing green fluorescent protein, as a control), pEGISI (containing inhibin alpha (1 ~ 32) fragments), or pEGISI-CpG-ODNs (containing inhibin alpha (1 ~ 32) fragments and CpG-ODNs motifs) plasmid for 48 h in vitro. Our results showed that the mRNA and protein expression levels of inhibin alpha were downregulated in mGCs transfected with pEGISI-CpG-ODNs, compared to those transfected with pEGISI. Flow cytometry demonstrated that pEGISI-CpG-ODNs transfection promoted cell proliferation (for example, increasing the number of cells in S and G2 phases) and decreased apoptosis, compared to pEGISI transfection. The present study also indicated that the expression of cell cycle-related genes (cyclin D2, cyclin D3, cyclin E1, Cdk2, and Cdk6) was increased, while the expression of apoptosis-related factors (Fas, FasL, caspase-8, and caspase-3) decreased after pEGISI-CpG-ODNs treatment. Additionally, pEGISI-CpG-ODNs reversed the effect of pEGISI on the secretion of estradiol in mGCs, which was further validated by upregulating the levels of its synthesis-related factors (StAR, Cyp11a1, and 17ß-HSD II). Nevertheless, pEGISI-CpG-ODNs or pEGISI did not affect the concentration of progesterone nor changed the expression levels of its synthesis-related factors (3ß-HSD I and Cyp19a1). In conclusion, this study demonstrated that CpG-ODNs may affect the cell cycle, apoptosis, and steroidogenesis by targeting the effects of inhibin alpha (1 ~ 32) fragments, supporting the potential role of CpG-ODNs in the development of granulosa cells.
Asunto(s)
Citosina , Oligodesoxirribonucleótidos , Animales , Apoptosis/fisiología , Ciclo Celular , Citosina/metabolismo , Citosina/farmacología , Femenino , Células de la Granulosa/metabolismo , Guanina/metabolismo , Guanina/farmacología , Inhibinas/genética , Inhibinas/metabolismo , Inhibinas/farmacología , Ratones , Oligodesoxirribonucleótidos/metabolismo , Fosfatos/metabolismo , Fosfatos/farmacologíaRESUMEN
Human mastadenovirus (HAdV), a linear double-stranded DNA (dsDNA) virus, is the causal agent of several diseases, including pharyngoconjunctival fever, epidemic keratoconjunctivitis, and hemorrhagic cystitis, in immunocompromised individuals. There are more than 100 reported types of adenoviruses, but the pathogenicity of many HAdVs remains unknown. Brincidofovir (BCV) is a hexadecyloxypropyl lipid conjugate of cidofovir (CDV) that is active against dsDNA viruses. Clinical effectiveness of BCV against certain HAdV species has been reported; however, its activity against novel HAdV types remains unknown. We investigated the half-maximal inhibitory concentration (IC50) values of BCV for novel HAdV types and found that the epidemic keratoconjunctivitis-associated HAdV-D54 prevalent in the Asian region was the most susceptible. The mean overall IC50 value of BCV was lower than that of CDV, indicating that BCV is effective against HAdVs, including the novel types. IMPORTANCE We investigated the IC50 values of BCV for novel HAdV types and found that the epidemic keratoconjunctivitis-associated HAdV-D54 prevalent in the Asian region was the most susceptible. In addition, the mean overall IC50 value of BCV was lower than that of CDV, indicating that BCV is effective against HAdVs.
Asunto(s)
Infecciones por Adenoviridae/virología , Infecciones por Adenovirus Humanos/virología , Citosina/análogos & derivados , Queratoconjuntivitis/virología , Mastadenovirus/efectos de los fármacos , Organofosfonatos/farmacología , Infecciones por Adenoviridae/inmunología , Infecciones por Adenovirus Humanos/inmunología , Cistitis , Citosina/farmacología , Humanos , Huésped Inmunocomprometido , Queratoconjuntivitis/inmunología , Mastadenovirus/clasificación , Mastadenovirus/genética , Mastadenovirus/fisiologíaAsunto(s)
Infecciones por Adenoviridae , Trasplante de Células Madre Hematopoyéticas , Infecciones por Adenoviridae/tratamiento farmacológico , Antivirales/uso terapéutico , Citosina/análogos & derivados , Citosina/farmacología , Citosina/uso terapéutico , Humanos , Organofosfonatos , Estudios RetrospectivosRESUMEN
Smallpox, caused by Variola virus (VARV), was eradicated in 1980; however, VARV bioterrorist threats still exist, necessitating readily available therapeutics. Current preparedness activities recognize the importance of oral antivirals and recommend therapeutics with different mechanisms of action. Monkeypox virus (MPXV) is closely related to VARV, causing a highly similar clinical human disease, and can be used as a surrogate for smallpox antiviral testing. The prairie dog MPXV model has been characterized and used to study the efficacy of antipoxvirus therapeutics, including recently approved TPOXX (tecovirimat). Brincidofovir (BCV; CMX001) has shown antiviral activity against double-stranded DNA viruses, including poxviruses. To determine the exposure of BCV following oral administration to prairie dogs, a pharmacokinetics (PK) study was performed. Analysis of BCV plasma concentrations indicated variability, conceivably due to the outbred nature of the animals. To determine BCV efficacy in the MPXV prairie dog model, groups of animals were intranasally challenged with 9 × 105 plaque-forming units (PFU; 90% lethal dose [LD90]) of MPXV on inoculation day 0 (ID0). Animals were divided into groups based on the first day of BCV treatment relative to inoculation day (ID-1, ID0, or ID1). A trend in efficacy was noted dependent upon treatment initiation (57% on ID-1, 43% on ID0, and 29% on ID1) but was lower than demonstrated in other animal models. Analysis of the PK data indicated that BCV plasma exposure (maximum concentration [Cmax]) and the time of the last quantifiable concentration (AUClast) were lower than in other animal models administered the same doses, indicating that suboptimal BCV exposure may explain the lower protective effect on survival.IMPORTANCE Preparedness activities against highly transmissible viruses with high mortality rates have been highlighted during the ongoing coronavirus disease 2019 (COVID-19) pandemic. Smallpox, caused by variola virus (VARV) infection, is highly transmissible, with an estimated 30% mortality. Through an intensive vaccination campaign, smallpox was declared eradicated in 1980, and routine smallpox vaccination of individuals ceased. Today's current population has little/no immunity against VARV. If smallpox were to reemerge, the worldwide results would be devastating. Recent FDA approval of one smallpox antiviral (tecovirimat) was a successful step in biothreat preparedness; however, orthopoxviruses can become resistant to treatment, suggesting the need for multiple therapeutics. Our paper details the efficacy of the investigational smallpox drug brincidofovir in a monkeypox virus (MPXV) animal model. Since brincidofovir has not been tested in vivo against smallpox, studies with the related virus MPXV are critical in understanding whether it would be protective in the event of a smallpox outbreak.
Asunto(s)
Citosina/análogos & derivados , Monkeypox virus/efectos de los fármacos , Organofosfonatos/farmacología , Organofosfonatos/farmacocinética , Viruela/tratamiento farmacológico , Animales , Antivirales/farmacocinética , Antivirales/farmacología , Benzamidas/farmacocinética , Benzamidas/farmacología , Citosina/farmacocinética , Citosina/farmacología , Modelos Animales de Enfermedad , Perros , Femenino , Isoindoles/farmacocinética , Isoindoles/farmacología , Masculino , Virus de la Viruela/efectos de los fármacosRESUMEN
Thiouracil and thiocytosine are important heterocyclic pharmacophores having pharmacological diversity. Antitumor and antiviral activity is commonly associated with thiouracil and thiocytosine derivatives, which are well known fragments for adenosine receptor affinity with many associated pharmacological properties. In this respect, 33 novel compounds have been synthesized in two groups: 24 thiouracil derivatives (4a-x) and 9 thiocytosine derivatives (5a-i). Antitumor activity of all the compounds was determined in the U87 MG glioblastoma cell line. Compound 5e showed an anti-proliferative IC50 of 1.56 µM, which is slightly higher activity than cisplatin (1.67 µM). The 11 most active compounds showed no signficant binding to adenosine A1, A2A or A2B receptors at 1 µM. Brain tumors express high amounts of phosphodiesterases. Compounds were tested for PDE4 inhibition, and 5e and 5f showed the best potency (5e: 3.42 µM; 5f: 0.97 µM). Remakably, those compounds were also the most active against U87MG. However, the compounds lacked a cytotoxic effect on the HEK293 healthy cell line, which encourages further investigation.
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Antineoplásicos/farmacología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Citosina/farmacología , Glioblastoma/tratamiento farmacológico , Inhibidores de Fosfodiesterasa 4/farmacología , Receptores Purinérgicos P1/metabolismo , Tiouracilo/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/química , Sitios de Unión/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Citosina/análogos & derivados , Citosina/química , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Glioblastoma/metabolismo , Glioblastoma/patología , Humanos , Estructura Molecular , Inhibidores de Fosfodiesterasa 4/síntesis química , Inhibidores de Fosfodiesterasa 4/química , Relación Estructura-Actividad , Tiouracilo/síntesis química , Tiouracilo/químicaRESUMEN
OBJECTIVES: Human adenovirus (HAdV) infections are associated with a high morbidity and mortality in transplant patients requiring the use of antiviral treatments. Brincidofovir (BCV), a cytidine analog, inhibits HAdV replication through viral DNA elongation termination and likely through other mechanisms. To elucidate if BCV regulates cellular antiviral pathways, we analyzed its impact on HAdV-infected and non-HAdV-infected lung epithelial cells. METHODS: We assessed the cellular and viral transcriptome of A549 cells infected and non-infected with HAdV C5 and treated or non-treated with BCV by RNAseq after 72 h. RESULTS: BCV treatment of HAdV infected cells resulted in a profound decrease of viral transcription associated with a relative overexpression of the early genes E1A and E4 and of the late gene L1. BCV had also a profound impact on A549 cells' transcriptome. Ontologic analysis revealed an effect of BCV on several pathways known to interact with adenovirus replication as mTor signalling and Wnt pathways. A549 cells treated with BCV demonstrated a significant inhibition of the biological function of "viral replication" including 25 dysregulated genes involved in inflammation pathways. CONCLUSION: We demonstrated that BCV alters viral gene expression and promotes the expression of antiviral cellular pathways in A549 cells. These results provide new insights how to interfere with cellular pathways to control HAdV infections.
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Adenovirus Humanos/efectos de los fármacos , Antivirales/farmacología , Citosina/análogos & derivados , Organofosfonatos/farmacología , Transcriptoma , Células A549 , Citosina/farmacología , Interacciones Microbiota-Huesped , Humanos , Replicación Viral/efectos de los fármacosRESUMEN
5-Formylcytosine (5fC) is a chemically edited, naturally occurring nucleobase which appears in the context of modified DNA strands. The understanding of the impact of 5fC on dsDNA physical properties is to date limited. In this work, we applied temperature-dependent 1H Chemical Exchange Saturation Transfer (CEST) NMR experiments to non-invasively and site-specifically measure the thermodynamic and kinetic influence of formylated cytosine nucleobase on the melting process involving dsDNA. Incorporation of 5fC within symmetrically positioned CpG sites destabilizes the whole dsDNA structure-as witnessed from the â¼2°C decrease in the melting temperature and 5-10 kJ mol-1 decrease in ΔG°-and affects the kinetic rates of association and dissociation. We observed an up to â¼5-fold enhancement of the dsDNA dissociation and an up to â¼3-fold reduction in ssDNA association rate constants, over multiple temperatures and for several proton reporters. Eyring and van't Hoff analysis proved that the destabilization is not localized, instead all base-pairs are affected and the transition states resembles the single-stranded conformation. These results advance our knowledge about the role of 5fC as a semi-permanent epigenetic modification and assist in the understanding of its interactions with reader proteins.
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Citosina/análogos & derivados , ADN/efectos de los fármacos , Conformación Molecular/efectos de los fármacos , Termodinámica , Emparejamiento Base/genética , Islas de CpG/genética , Citosina/química , Citosina/farmacología , ADN/química , ADN/genética , ADN de Cadena Simple/efectos de los fármacos , ADN de Cadena Simple/genética , Cinética , Espectroscopía de Resonancia Magnética , Temperatura de TransiciónRESUMEN
We synthesized affinity-based chemical probes of cytosine-adenosine bisubstrate analogs and identified several potential targets by proteomic analysis. The validation of the proteomic analysis identified the chemical probe as a specific inhibitor of glucose-regulated protein 94 (GRP94), a potential drug target for several types of cancers. Therefore, as a result of the use of bisubstrate-type chemical probes and a chemical-biology methodology, this work opens the way to the development of a new family of GRP94 inhibitors that could potentially be of therapeutic interest.
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Adenosina/análogos & derivados , Adenosina/farmacología , Marcadores de Afinidad/farmacología , Citosina/análogos & derivados , Citosina/farmacología , Glicoproteínas de Membrana/antagonistas & inhibidores , Adenosina/efectos de la radiación , Marcadores de Afinidad/síntesis química , Marcadores de Afinidad/efectos de la radiación , Línea Celular Tumoral , Química Clic , Citosina/efectos de la radiación , Humanos , Glicoproteínas de Membrana/química , Proteoma/química , Proteómica , Rayos UltravioletaRESUMEN
OBJECTIVE: To evaluate the anti-cancer effect of unmethylated cytosine-phosphorothioate-guanine (CpG)-containing oligodeoxynucleotides (ODNs) on human bladder cancer UM-UC-3 cells, our study was carried out. METHODS: The viability of cells (UM-UC-3, T24 and SV-HUC-1) with CpG ODN treatments was examined by cell counting kit-8 (CCK-8) assay. Apoptosis and cell cycle phase were determined by flow cytometry analysis. Pre-apoptosis factors of caspase-3, p53, B-cell lymphoma 2 associated X protein (Bax) and anti-apoptosis factor of B-cell lymphoma 2 (Bcl-2) were detected by western blot. RESULTS: Experimental results showed that the viability of human bladder cancer cells (UM-UC-3 and T24) with CpG ODN treatment was decreased and the viability of human normal urothelial cells (SV-HUC-1) with CpG ODN treatment was increased with time-dependance manner. Moreover, CpG ODN increased the apoptosis rate of UM-UC-3 cells and arrested more cells in G0G1 phase. Furthermore, the expression of caspase-3, p53 and Bax were increased and the expression of Bcl-2 was decreased with CpG ODN treatment on UM-UC-3 cells. CONCLUSION: CpG ODN promoted the proliferation of normal urinary transitional epithelial cells (SV-HUC-1) and inhibited the cell viability of human bladder cancer cells (UM-UC-3 and T24) in vitro. CpG ODN induced the apoptosis of human bladder cancer (UM-UC-3) cells in a cascade progress via enhancing the expression of caspase-3, p53 and Bax, and inhibiting the expression of Bcl-2 with significant time-dependancy. CpG ODN inhibited cell cycle distribution of human bladder cancer (UM-UC-3) cells with more cells were arrested in G0G1 phase. This study suggested that the CpG ODN is the potential candidate on human bladder cancer.