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1.
Mol Vis ; 23: 561-571, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28855795

RESUMO

PURPOSE: Age-related macular degeneration (AMD) is the leading cause of permanent vision loss among the elderly in many industrialized countries, and the complement system plays an important role in the pathogenesis of AMD. Inhibition of complement factor B, a key regulator of the alternative pathway, is implicated as a potential therapeutic intervention for AMD. Here we investigated the effect of liver factor B reduction on systemic and ocular factor B levels. METHODS: Second-generation antisense oligonucleotides (ASOs) targeting mouse and monkey factor B mRNA were administered by subcutaneous injection to healthy mice or monkeys, and the level of factor B mRNA was assessed in the liver and the eye. In addition, the factor B protein level was determined in plasma and whole eyes from the treated animals. RESULTS: Mice and monkeys treated with factor B ASOs demonstrated a robust reduction in liver factor B mRNA levels with no change in ocular factor B mRNA levels. Plasma factor B protein levels were significantly reduced in mice and monkeys treated with factor B ASOs, leading to a dramatic reduction in ocular factor B protein, below the assay detection levels. CONCLUSIONS: The results add to the increasing evidence that the liver is the main source of plasma and ocular factor B protein, and demonstrate that reduction of liver factor B mRNA by an ASO results in a significant reduction in plasma and ocular factor B protein levels. The results suggest that inhibition of liver factor B mRNA by factor B ASOs would reduce systemic alternative complement pathway activation and has potential to be used as a novel therapy for AMD.


Assuntos
Fator B do Complemento/genética , Fator B do Complemento/metabolismo , Olho/metabolismo , Fígado/metabolismo , Oligonucleotídeos Antissenso/administração & dosagem , RNA Mensageiro/metabolismo , Animais , Western Blotting , Eletroforese em Gel de Poliacrilamida , Injeções Subcutâneas , Macaca fascicularis , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase em Tempo Real
2.
J Hepatol ; 64(4): 781-9, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26658683

RESUMO

BACKGROUND & AIMS: Current treatment of chronic hepatitis B virus infection (CHB) includes interferon and nucleos(t)ide analogues, which generally do not reduce HBV surface antigen (HBsAg) production, a constellation that is associated with poor prognosis of CHB. Here we evaluated the efficacy of an antisense approach using antisense oligonucleotide (ASO) technology already in clinical use for liver targeted therapy to specifically inhibit HBsAg production and viremia in a preclinical setting. METHODS: A lead ASO was identified and characterized in vitro and subsequently tested for efficacy in vivo and in vitro using HBV transgenic and hydrodynamic transfection mouse and a cell culture HBV infection model, respectively. RESULTS: ASO treatment decreased serum HBsAg levels ⩾2 logs in a dose and time-dependent manner; HBsAg decreased 2 logs in a week and returned to baseline 4 weeks after a single ASO injection. ASO treatment effectively reduced HBsAg in combination with entecavir, while the nucleoside analogue alone did not. ASO treatment has pan-genotypic antiviral activity in the hydrodynamic transfection system. Finally, cccDNA-driven HBV gene expression is ASO sensitive in HBV infected cells in vitro. CONCLUSION: Our results demonstrate in a preclinical setting the efficacy of an antisense approach against HBV by efficiently reducing serum HBsAg (as well as viremia) across different genotypes alone or in combination with standard nucleoside therapy. Since the applied antisense technology is already in clinical use, a lead compound can be rapidly validated in a clinical setting and thus, constitutes a novel therapeutic approach targeting chronic HBV infection.


Assuntos
Antígenos de Superfície da Hepatite B/sangue , Hepatite B Crônica/tratamento farmacológico , Oligonucleotídeos Antissenso/uso terapêutico , Viremia/tratamento farmacológico , Animais , Células Hep G2 , Antígenos E da Hepatite B/sangue , Hepatite B Crônica/virologia , Humanos , Camundongos
3.
Adv Ther ; 40(4): 1317-1333, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36690839

RESUMO

TMPRSS6 is a serine protease highly expressed in the liver. Its role in iron regulation was first reported in 2008 when mutations in TMPRSS6 were shown to be the cause of iron-refractory iron deficiency anemia (IRIDA) in humans and in mouse models. TMPRSS6 functions as a negative regulator of the expression of the systemic iron-regulatory hormone hepcidin. Over the last decade and a half, growing understanding of TMPRSS6 biology and mechanism of action has enabled development of new therapeutic approaches for patients with diseases of erythropoiesis and iron homeostasis.ClinicalTrials.gov identifier NCT03165864.


Assuntos
Anemia Ferropriva , Eritropoese , Camundongos , Animais , Humanos , Eritropoese/genética , Anemia Ferropriva/tratamento farmacológico , Ferro/metabolismo , Fígado/metabolismo , Homeostase , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo
4.
EBioMedicine ; 29: 92-103, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29463471

RESUMO

Chronic hepatitis leads to liver fibrosis and cirrhosis. Cirrhosis is a major cause of worldwide morbidity and mortality. Macrophages play a key role in fibrosis progression and reversal. However, the signals that determine fibrogenic vs fibrolytic macrophage function remain ill defined. We studied the role of interleukin-4 receptor α (IL-4Rα), a potential central switch of macrophage polarization, in liver fibrosis progression and reversal. We demonstrate that inflammatory monocyte infiltration and liver fibrogenesis were suppressed in general IL-4Rα-/- as well as in macrophage-specific IL-4Rα-/- (IL-4RαΔLysM) mice. However, with deletion of IL-4RαΔLysM spontaneous fibrosis reversal was retarded. Results were replicated by pharmacological intervention using IL-4Rα-specific antisense oligonucleotides. Retarded resolution was linked to the loss of M2-type resident macrophages, which secreted MMP-12 through IL-4 and IL-13-mediated phospho-STAT6 activation. We conclude that IL-4Rα signaling regulates macrophage functional polarization in a context-dependent manner. Pharmacological targeting of macrophage polarization therefore requires disease stage-specific treatment strategies. RESEARCH IN CONTEXT: Alternative (M2-type) macrophage activation through IL-4Rα promotes liver inflammation and fibrosis progression but speeds up fibrosis reversal. This demonstrates context dependent, opposing roles of M2-type macrophages. During reversal IL-4Rα induces fibrolytic MMPs, especially MMP-12, through STAT6. Liver-specific antisense oligonucleotides efficiently block IL-4Rα expression and attenuate fibrosis progression.


Assuntos
Subunidade alfa de Receptor de Interleucina-4/metabolismo , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Macrófagos/metabolismo , Transdução de Sinais , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Expressão Gênica , Subunidade alfa de Receptor de Interleucina-4/genética , Cirrose Hepática/tratamento farmacológico , Cirrose Hepática/etiologia , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Metaloproteinases da Matriz/genética , Metaloproteinases da Matriz/metabolismo , Camundongos , Camundongos Knockout , Células Mieloides/imunologia , Células Mieloides/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Células RAW 264.7 , Fator de Transcrição STAT6/metabolismo , Baço/imunologia , Baço/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo
5.
Immunobiology ; 221(6): 701-8, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-26307001

RESUMO

Systemic lupus erythematosus is an autoimmune disease that manifests in widespread complement activation and deposition of complement fragments in the kidney. The complement pathway is believed to play a significant role in the pathogenesis and in the development of lupus nephritis. Complement factor B is an important activator of the alternative complement pathway and increasing evidence supports reducing factor B as a potential novel therapy to lupus nephritis. Here we investigated whether pharmacological reduction of factor B expression using antisense oligonucleotides could be an effective approach for the treatment of lupus nephritis. We identified potent and well tolerated factor B antisense oligonucleotides that resulted in significant reductions in hepatic and plasma factor B levels when administered to normal mice. To test the effects of factor B antisense oligonucleotides on lupus nephritis, we used two different mouse models, NZB/W F1 and MRL/lpr mice, that exhibit lupus nephritis like renal pathology. Antisense oligonucleotides mediated reductions in circulating factor B levels were associated with significant improvements in renal pathology, reduced glomerular C3 deposition and proteinuria, and improved survival. These data support the strategy of using factor B antisense oligonucleotides for treatment of lupus nephritis in humans.


Assuntos
Complexo Antígeno-Anticorpo/metabolismo , Fator B do Complemento/genética , Hepatócitos/fisiologia , Rim/metabolismo , Lúpus Eritematoso Sistêmico/terapia , Nefrite Lúpica/terapia , Oligonucleotídeos Antissenso/genética , Animais , Células Cultivadas , Complemento C3/metabolismo , Fator B do Complemento/metabolismo , Via Alternativa do Complemento/genética , Modelos Animais de Doenças , Humanos , Rim/patologia , Lúpus Eritematoso Sistêmico/imunologia , Nefrite Lúpica/imunologia , Camundongos , Camundongos Endogâmicos MRL lpr , Camundongos Endogâmicos NZB , Proteinúria
6.
Invest Ophthalmol Vis Sci ; 56(11): 6362-75, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26436889

RESUMO

PURPOSE: To preserve photoreceptor cell structure and function in a rodent model of retinitis pigmentosa with P23H rhodopsin by selective inhibition of the mutant rhodopsin allele using a second generation antisense oligonucleotide (ASO). METHODS: Wild-type mice and rats were treated with ASO by intravitreal (IVT) injection and rhodopsin mRNA and protein expression were measured. Transgenic rats expressing the murine P23H rhodopsin gene (P23H transgenic rat Line 1) were administered either a mouse-specific P23H ASO or a control ASO. The contralateral eye was injected with PBS and used as a comparator control. Electroretinography (ERG) measurements and analyses of the retinal outer nuclear layer were conducted and correlated with rhodopsin mRNA levels. RESULTS: Rhodopsin mRNA and protein expression was reduced after a single ASO injection in wild-type mice with a rhodopsin-specific ASO. Transgenic rat eyes that express a murine P23H rhodopsin gene injected with a murine P23H ASO had a 181 ± 39% better maximum amplitude response (scotopic a-wave) as compared with contralateral PBS-injected eyes; the response in control ASO eyes was not significantly different from comparator contralateral eyes. Morphometric analysis of the outer nuclear layer showed a significantly thicker nuclear layer in eyes injected with murine P23H ASO (18%) versus contralateral PBS-injected eyes. CONCLUSIONS: Allele-specific ASO-mediated knockdown of mutant P23H rhodopsin expression slowed the rate of photoreceptor degeneration and preserved the function of photoreceptor cells in eyes of the P23H rhodopsin transgenic rat. Our data indicate that ASO treatment is a potentially effective therapy for the treatment of retinitis pigmentosa.


Assuntos
Regulação da Expressão Gênica , Degeneração Macular/prevenção & controle , Oligonucleotídeos Antissenso/genética , RNA Mensageiro/genética , Rodopsina/genética , Alelos , Animais , Western Blotting , Modelos Animais de Doenças , Eletrorretinografia , Degeneração Macular/genética , Degeneração Macular/metabolismo , Masculino , Camundongos , Ratos , Ratos Transgênicos , Reação em Cadeia da Polimerase em Tempo Real , Rodopsina/biossíntese
7.
Chronic Obstr Pulm Dis ; 2(2): 177-190, 2015 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-28848840

RESUMO

Clinical research in rare diseases, including alpha-1 antitrypsin deficiency (AATD), faces challenges not shared by common disease research. These challenges may include the limited number of patient volunteers available for research, lack of natural history studies on which to base many clinical trial interventions, an urgency for the development of drug therapies given the often poor prognosis of rare diseases and uncertainties about appropriate biomarkers and clinical outcomes critical to clinical trial design. To address these challenges and initiate formal discussions among key stakeholders-patients, researchers, industry, federal regulators-the Alpha-1 Foundation hosted the Clinical Trial Design for Alpha-1 Antitrypsin Deficiency: A Model for Rare Diseases conference February 3-4, 2014 in Bethesda, Maryland. Discussions at the conference led to the conclusions that 1) adaptive designs should be considered for rare disease clinical trials yet more dialogue and study is needed to make these designs feasible for smaller trials and to address current limitations; 2) natural history studies, including the identification of appropriate biomarkers are critically needed and precompetitive collaborations may offer a means of creating these costly studies; and 3) patient registries and databases within the rare disease community need to be more publicly available and integrated, particularly for AATD. This report summarizes the discussions leading to these conclusions.

8.
Rare Dis ; 2: e28511, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25054094

RESUMO

Alpha-1 antitrypsin (AAT) is a serum protease inhibitor that belongs to the serpin superfamily. Mutations in AAT are associated with α-1 antitrypsin deficiency (AATD), a rare genetic disease with two distinct manifestations: AATD lung disease and AATD liver disease. AATD lung disease is caused by loss-of-function of AAT and can be treated with plasma-derived AAT. AATD liver disease is due to the aggregation and retention of mutant AAT protein in the liver; the only treatment available for AATD liver disease is liver transplantation. Here we demonstrate that antisense oligonucleotides (ASOs) targeting human AAT efficiently reduce levels of both short and long human AAT transcript in vitro and in transgenic mice, providing a novel therapy for AATD liver disease. In addition, ASO-mediated depletion of mouse AAT may offer a useful animal model for the investigation of AATD lung disease.

9.
J Clin Invest ; 124(1): 251-61, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24355919

RESUMO

Alpha-1 antitrypsin deficiency (AATD) is a rare genetic disease that results from mutations in the alpha-1 antitrypsin (AAT) gene. The mutant AAT protein aggregates and accumulates in the liver leading to AATD liver disease, which is only treatable by liver transplant. The PiZ transgenic mouse strain expresses a human AAT (hAAT) transgene that contains the AATD-associated Glu342Lys mutation. PiZ mice exhibit many AATD symptoms, including AAT protein aggregates, increased hepatocyte death, and liver fibrosis. In the present study, we systemically treated PiZ mice with an antisense oligonucleotide targeted against hAAT (AAT-ASO) and found reductions in circulating levels of AAT and both soluble and aggregated AAT protein in the liver. Furthermore, AAT-ASO administration in these animals stopped liver disease progression after short-term treatment, reversed liver disease after long-term treatment, and prevented liver disease in young animals. Additionally, antisense oligonucleotide treatment markedly decreased liver fibrosis in this mouse model. Administration of AAT-ASO in nonhuman primates led to an approximately 80% reduction in levels of circulating normal AAT, demonstrating potential for this approach in higher species. Antisense oligonucleotides thus represent a promising therapy for AATD liver disease.


Assuntos
Oligonucleotídeos Antissenso/genética , Deficiência de alfa 1-Antitripsina/terapia , Animais , Feminino , Técnicas de Silenciamento de Genes , Células Hep G2 , Hepatócitos/enzimologia , Humanos , Fígado/enzimologia , Fígado/patologia , Cirrose Hepática/enzimologia , Cirrose Hepática/genética , Cirrose Hepática/terapia , Macaca fascicularis , Masculino , Camundongos , Camundongos Transgênicos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , alfa 1-Antitripsina/genética , alfa 1-Antitripsina/metabolismo , Deficiência de alfa 1-Antitripsina/enzimologia , Deficiência de alfa 1-Antitripsina/genética
10.
PLoS One ; 8(7): e66923, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23922646

RESUMO

Obesity is a primary risk factor for multiple metabolic disorders. Many drugs for the treatment of obesity, which mainly act through CNS as appetite suppressants, have failed during development or been removed from the market due to unacceptable adverse effects. Thus, there are very few efficacious drugs available and remains a great unmet medical need for anti-obesity drugs that increase energy expenditure by acting on peripheral tissues without severe side effects. Here, we report a novel approach involving antisense inhibition of fibroblast growth factor receptor 4 (FGFR4) in peripheral tissues. Treatment of diet-induce obese (DIO) mice with FGFR4 antisense oligonucleotides (ASO) specifically reduced liver FGFR4 expression that not only resulted in decrease in body weight (BW) and adiposity in free-feeding conditions, but also lowered BW and adiposity under caloric restriction. In addition, combination treatment with FGFR4 ASO and rimonabant showed additive reduction in BW and adiposity. FGFR4 ASO treatment increased basal metabolic rate during free-feeding conditions and, more importantly, prevented adaptive decreases of metabolic rate induced by caloric restriction. The treatment increased fatty acid oxidation while decreased lipogenesis in both liver and fat. Mechanistic studies indicated that anti-obesity effect of FGFR4 ASO was mediated at least in part through an induction of plasma FGF15 level resulted from reduction of hepatic FGFR4 expression. The anti-obesity effect was accompanied by improvement in plasma glycemia, whole body insulin sensitivity, plasma lipid levels and liver steatosis. Therefore, FGFR4 could be a potential novel target and antisense reduction of hepatic FGFR4 expression could be an efficacious therapy as an adjunct to diet restriction or to an appetite suppressant for the treatment of obesity and related metabolic disorders.


Assuntos
Adiposidade/efeitos dos fármacos , Metabolismo Basal/efeitos dos fármacos , Obesidade/metabolismo , Oligonucleotídeos Antissenso/farmacologia , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/metabolismo , Animais , Ácidos e Sais Biliares/metabolismo , Peso Corporal/efeitos dos fármacos , Restrição Calórica , Dieta , Quimioterapia Combinada , Ácidos Graxos/metabolismo , Fígado Gorduroso/sangue , Fígado Gorduroso/genética , Fígado Gorduroso/patologia , Comportamento Alimentar/efeitos dos fármacos , Fatores de Crescimento de Fibroblastos/sangue , Regulação da Expressão Gênica/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Insulina/farmacologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Obesidade/sangue , Obesidade/tratamento farmacológico , Obesidade/genética , Oligonucleotídeos Antissenso/uso terapêutico , Oxirredução/efeitos dos fármacos , Piperidinas/farmacologia , Piperidinas/uso terapêutico , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptor CB1 de Canabinoide/metabolismo , Receptor Tipo 4 de Fator de Crescimento de Fibroblastos/genética , Rimonabanto
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