RESUMO
PURPOSE OF REVIEW: The purpose of this literature review was to review the latest use of biologics in the management of anaphylaxis. The methodology was to highlight both the nonbiologic management of anaphylaxis and the biologic management and how they can be used in conjunction with each other. RECENT FINDINGS: As the phenotypes and endotypes of anaphylaxis are better portrayed, it furthers our understanding of the mechanisms of anaphylaxis. New applications of existing biologics to the prevention of anaphylaxis are described. SUMMARY: Anaphylaxis is a potentially life-threatening acute hypersensitivity reaction affecting up to 16.8% of the U.S. population. Acute management entails swift identification, removal of the causative agent, and the prevention of cardiovascular collapse, firstly with epinephrine. Adjunctive treatments such as antihistamines work to prevent anaphylaxis from recurring. Biologic management of anaphylaxis involves the use of large-molecule drugs such as monoclonal antibodies. Omalizumab, an IgG1 monoclonal antibody targeting unbound IgE, is the most prevalent and widely studied biologic in the prevention of anaphylaxis. Other monoclonal antibodies in development or approved for other indications, such as ligelizumab, quilizumab, MEDI4212, and dupilumab, may also have potential for preventing anaphylaxis through various mechanisms.
Assuntos
Anafilaxia , Produtos Biológicos , Humanos , Anafilaxia/tratamento farmacológico , Anafilaxia/prevenção & controle , Anafilaxia/etiologia , Omalizumab/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Epinefrina/uso terapêutico , Imunoglobulina E , Produtos Biológicos/uso terapêuticoRESUMO
BACKGROUND: Atopic dermatitis (AD) is characterized by TH2-dominated skin inflammation and systemic response to cutaneously encountered antigens. The TH2 cytokines IL-4 and IL-13 play a critical role in the pathogenesis of AD. The Q576->R576 polymorphism in the IL-4 receptor alpha (IL-4Rα) chain common to IL-4 and IL-13 receptors alters IL-4 signaling and is associated with asthma severity. OBJECTIVE: We sought to investigate whether the IL-4Rα R576 polymorphism is associated with AD severity and exaggerates allergic skin inflammation in mice. METHODS: Nighttime itching interfering with sleep, Rajka-Langeland, and Eczema Area and Severity Index scores were used to assess AD severity. Allergic skin inflammation following epicutaneous sensitization of mice 1 or 2 IL-4Rα R576 alleles (QR and RR) and IL-4Rα Q576 (QQ) controls was assessed by flow cytometric analysis of cells and quantitative RT-PCR analysis of cytokines in skin. RESULTS: The frequency of nighttime itching in 190 asthmatic inner-city children with AD, as well as Rajka-Langeland and Eczema Area and Severity Index scores in 1116 White patients with AD enrolled in the Atopic Dermatitis Research Network, was higher in subjects with the IL-4Rα R576 polymorphism compared with those without, with statistical significance for the Rajka-Langeland score. Following epicutaneous sensitization of mice with ovalbumin or house dust mite, skin infiltration by CD4+ cells and eosinophils, cutaneous expression of Il4 and Il13, transepidermal water loss, antigen-specific IgE antibody levels, and IL-13 secretion by antigen-stimulated splenocytes were significantly higher in RR and QR mice compared with QQ controls. Bone marrow radiation chimeras demonstrated that both hematopoietic cells and stromal cells contribute to the mutants' exaggerated allergic skin inflammation. CONCLUSIONS: The IL-4Rα R576 polymorphism predisposes to more severe AD and increases allergic skin inflammation in mice.
Assuntos
Dermatite Atópica , Eczema , Camundongos , Animais , Interleucina-13/genética , Interleucina-13/metabolismo , Interleucina-4/genética , Interleucina-4/metabolismo , Células Th2 , Pele/metabolismo , Citocinas/metabolismo , Inflamação/metabolismo , Prurido/metabolismo , Eczema/metabolismoRESUMO
PURPOSE OF REVIEW: The purpose of this literature review was to review the latest advancements with biologics in rapid drug desensitization. Our methodology was to highlight both desensitization to biologics themselves and the use of biologics in desensitization to both biologic and nonbiologic drugs. RECENT FINDINGS: Biologics are a vast category of drugs that include monoclonal antibodies, nanobodies, modern vaccinations, and even hormones. Desensitization to biologics can be safely performed through standardized procedure. Biomarkers are used both in vitro and in vivo to help identify and classify hypersensitivity reactions. Hypersensitivity reactions to the mRNA vaccinations against SARS-CoV-2 present their own unique challenges to management. There are specific excipients in monoclonal antibodies that are thought to be responsible for many of their hypersensitivity reactions. Certain biologics can even be used to assist in desensitization to other drugs. Rapid drug desensitization is a standardized procedure that may be able to help many patients who have experienced hypersensitivity reactions to biologics and would best be treated with them to continue to receive them. Biologic drugs have opened a new era in medicine for the prevention and treatment of infectious diseases, cancer, and inflammatory diseases. Hypersensitivity reactions to biologics are quite common. This literature review presents the latest advancements in our understanding of hypersensitivity reactions to biologics, how rapid drug desensitization can be used to continue therapy despite history of hypersensitivity, and how biologics themselves can be used to aid in desensitization itself.
Assuntos
Anafilaxia , Produtos Biológicos , COVID-19 , Hipersensibilidade a Drogas , Humanos , Hipersensibilidade a Drogas/etiologia , Hipersensibilidade a Drogas/terapia , SARS-CoV-2 , Anticorpos Monoclonais/uso terapêutico , Dessensibilização Imunológica/métodos , Anafilaxia/etiologia , Produtos Biológicos/efeitos adversosRESUMO
Zinc Finger transcription factors are crucial in modulating various cellular processes, including differentiation. Chromatin reader Zinc Finger MYND (Myeloid, Nervy, and DEAF-1) type containing 8 (ZMYND8), an All-Trans Retinoic Acid (ATRA)-responsive gene, was previously shown to play a crucial role in promoting the expression of neuronal-lineage committed genes. Here, we report that ZMYND8 promotes neuronal differentiation by positively regulating canonical MAPT protein-coding gene isoform, a key player in the axonal development of neurons. Additionally, ZMYND8 modulates gene-isoform switching by epigenetically silencing key regulatory regions within the MAPT gene, thereby suppressing the expression of non-protein-coding isoforms such as MAPT213. Genetic deletion of ZMYND8 led to an increase in the MAPT213 that potentially suppressed the parental MAPT protein-coding transcript expression related to neuronal differentiation programs. In addition, ectopic expression of MAPT213 led to repression of MAPT protein-coding transcript. Similarly, ZMYND8-driven transcription regulation was also observed in other neuronal differentiation-promoting genes. Collectively our results elucidate a novel mechanism of ZMYND8-dependent transcription regulation of different neuronal lineage committing genes, including MAPT, to promote neural differentiation.
Assuntos
RNA Longo não Codificante , Diferenciação Celular/genética , Cromatina , Regulação da Expressão Gênica , RNA Longo não Codificante/genética , Tretinoína/farmacologia , Proteínas Supressoras de Tumor/metabolismoRESUMO
Hypersensitivity reactions to drugs have increased in the past 25 years due to increased exposures and availability of efficient, targeted, and personalized medications. Rapid drug desensitization is a clinical procedure that allows for the safe administration of a drug in patients with a history of such hypersensitivity reactions. Desensitization allows the continued use of first-line therapies, leading to higher efficacy of treatment, fewer side effects, cost-effectiveness, and increased quality of life and life expectancy of patients when compared with the use of second-line therapy. In this review, we discuss the who, what, where, when, why, and how of drug desensitization.
Assuntos
Anafilaxia , Hipersensibilidade a Drogas , Anafilaxia/etiologia , Dessensibilização Imunológica/métodos , Hipersensibilidade a Drogas/diagnóstico , Hipersensibilidade a Drogas/etiologia , Hipersensibilidade a Drogas/terapia , Humanos , Qualidade de VidaRESUMO
Long noncoding RNAs (lncRNAs) have emerged as critical regulators of biological processes. However, the aberrant expression of an isoform from the same lncRNA gene could lead to RNA with altered functions due to changes in their conformations, leading to diseases. Here, we describe a detailed characterization of the gene that encodes long intergenic non-protein-coding RNA 01016 (LINC01016, also known as LncRNA1195) with a focus on its structure, exon usage, and expression in human and macaque tissues. In this study we show that it is among the highly expressed lncRNAs in the testis, exclusively conserved among nonhuman primates, suggesting its recent evolution and is processed into 12 distinct RNAs in testis, cervix, and uterus tissues. Further, we integrate de novo annotation of expressed LINC01016 transcripts and isoform-dependent gene expression analyses to show that human LINC01016 is a multiexon gene, processed through differential exon usage with isoform-specific roles. Furthermore, in cervical, testicular, and uterine cancers, LINC01016 isoforms are differentially expressed, and their expression is predictive of survival in these cancers. This study has revealed an essential aspect of lncRNA biology, rarely associated with coding RNAs, that lncRNA genes are precisely processed to generate isoforms with distinct biological roles in specific tissues.
RESUMO
Recent technical and other advances in genomics provide unique opportunities to improve our understanding of human physiology and disease predisposition through a detailed analysis of gene structure and expression by examining data in public genome and gene-expression repositories. Yet, the vast majority of human genes remain understudied. This is particularly true of genes for long noncoding RNAs (lncRNAs). Here, we describe the detailed characterization of MIR503HG, a lncRNA gene found on the X chromosome in humans. Using information extracted from public databases, we show that human MIR503HG is a 5-exon gene, and that it is highly conserved among 5 non-human primates spanning over 85 million years ago of evolutionary diversification. MIR503HG is transcribed and processed into multiple distinct RNAs in each of these species through differential exon use and alternative RNA splicing, with a higher abundance of transcripts being found in reproductive tissues, especially during the early stages of ovary and testis development, indicating a possible role in reproductive biology. Furthermore, in select reproductive system cancers, MIR503HG transcripts are downregulated, with higher levels of RNA expression being associated with clinical outcomes. Collectively, these investigations show how the use of genomic, gene expression, and other genetic resources can lead to new insights about human biology and disease, and argue that MIR503HG is worthy of additional study.
Assuntos
Regulação da Expressão Gênica , RNA Longo não Codificante/química , RNA Longo não Codificante/genética , Animais , Genoma Humano , Humanos , Primatas , Prognóstico , Ligação Proteica , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
BACKGROUND: Parkinson's disease (PD) and multiple system atrophy (MSA) patients often suffer from gastrointestinal (GI) dysfunction and GI dysbiosis (microbial imbalance). GI dysfunction also occurs in mouse models of PD and MSA. OBJECTIVES: To assess gut dysfunction and dysbiosis in PD subjects as compared to controls, identify potential shared microbial taxa in humans and mouse models of PD and MSA, and to assess the effects of potential therapies on mouse GI microbiota. METHODS: In this human pilot study, GI function was assessed by fecal consistency/frequency measured using the Bristol Stool Form Scale and GI transit time assessed using Sitzmarks pills and abdominal radiology. Human and mouse microbiota were analyzed by extracting fecal genomic DNA followed by 16S rRNA sequencing. RESULTS: In our PD patients genera Akkermansia significantly increased while a trend toward increased Bifidobacterium and decreased Prevotella was observed. Families Bacteroidaceae and Lachnospiraceae and genera Prevotella and Bacteroides were detected in both humans and PD mice, suggesting potential shared biomarkers. In mice treated with the approved multiple sclerosis drug, FTY720, or with our FTY720-Mitoxy-derivative, we saw that FTY720 had little effect while FTY720-Mitoxy increased beneficial Ruminococcus and decreased Rickenellaceae family. CONCLUSION: Akkermansia and Prevotellaceae data reported by others were replicated in our human pilot study suggesting the use of those taxa as potential biomarkers for PD diagnosis. The effect of FTY720-Mitoxy on taxa Rikenellaceae and Ruminococcus and the relevance of S24-7 await further evaluation. It also remains to be determined if mouse microbiota have predictive power for human subjects.
Assuntos
Disbiose/microbiologia , Cloridrato de Fingolimode/farmacologia , Microbioma Gastrointestinal , Imunossupressores/farmacologia , Microbiota , Atrofia de Múltiplos Sistemas/microbiologia , Doença de Parkinson/microbiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Constipação Intestinal/fisiopatologia , Modelos Animais de Doenças , Feminino , Cloridrato de Fingolimode/administração & dosagem , Cloridrato de Fingolimode/análogos & derivados , Microbioma Gastrointestinal/efeitos dos fármacos , Motilidade Gastrointestinal/fisiologia , Humanos , Imunossupressores/administração & dosagem , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Projetos Piloto , RNA Ribossômico 16SRESUMO
Multiple system atrophy (MSA) is a fatal disorder with no effective treatment. MSA pathology is characterized by α-synuclein (aSyn) accumulation in oligodendrocytes, the myelinating glial cells of the central nervous system (CNS). aSyn accumulation in oligodendrocytes forms the pathognomonic glial cytoplasmic inclusions (GCIs) of MSA. MSA aSyn pathology is also associated with motor and autonomic dysfunction, including an impaired ability to sweat. MSA patients have abnormal CNS expression of glial-cell-line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF). Our prior studies using the parent compound FTY720, a food and drug administration (FDA) approved immunosuppressive for multiple sclerosis, reveal that FTY720 protects parkinsonian mice by increasing BDNF. Our FTY720-derivative, FTY720-Mitoxy, is known to increase expression of oligodendrocyte BDNF, GDNF, and nerve growth factor (NGF) but does not reduce levels of circulating lymphocytes as it is not phosphorylated so cannot modulate sphingosine 1 phosphate receptors (S1PRs). To preclinically assess FTY720-Mitoxy for MSA, we used mice expressing human aSyn in oligodendrocytes under a 2,' 3'-cyclic nucleotide 3'-phosphodiesterase (CNP) promoter. CNP-aSyn transgenic (Tg) mice develop motor dysfunction between 7 and 9 mo, and progressive GCI pathology. Using liquid chromatography-mass spectrometry (LC-MS/MS) and enzymatic assays, we confirmed that FTY720-Mitoxy was stable and active. Vehicle or FTY720-Mitoxy (1.1â¯mg/kg/day) was delivered to wild type (WT) or Tg littermates from 8.5-11.5 mo by osmotic pump. We behaviorally assessed their movement by rotarod and sweat production by starchiodine test. Postmortem tissues were evaluated by qPCR for BDNF, GDNF, NGF and GDNF-receptor RET mRNA and for aSyn, BDNF, GDNF, and Iba1 protein by immunoblot. MicroRNAs (miRNAs) were also assessed by qPCR. FTY720-Mitoxy normalized movement, sweat function and soleus muscle mass in 11.5 mo Tg MSA mice. FTY720-Mitoxy also increased levels of brain GDNF and reduced brain miR-96-5p, a miRNA that acts to decrease GDNF expression. Moreover, FTY720-Mitoxy blocked aSyn pathology measured by sequential protein extraction and immunoblot, and microglial activation assessed by immunohistochemistry and immunoblot. In the 3-nitropropionic acid (3NP) toxin model of MSA, FTY720-Mitoxy protected movement and mitochondria in WT and CNP-aSyn Tg littermates. Our data confirm potent in vivo protection by FTY720-Mitoxy, supporting its further evaluation as a potential therapy for MSA and related synucleinopathies.
Assuntos
Cloridrato de Fingolimode/análogos & derivados , Fator Neurotrófico Derivado de Linhagem de Célula Glial/biossíntese , Atrofia de Múltiplos Sistemas/patologia , Fármacos Neuroprotetores/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Cloridrato de Fingolimode/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Fator Neurotrófico Derivado de Linhagem de Célula Glial/efeitos dos fármacos , Humanos , Inflamação/metabolismo , Inflamação/patologia , Masculino , Camundongos , Camundongos Transgênicos , MicroRNAs/efeitos dos fármacos , MicroRNAs/metabolismo , Atrofia de Múltiplos Sistemas/metabolismo , Proteínas Proto-Oncogênicas c-ret/biossíntese , Proteínas Proto-Oncogênicas c-ret/efeitos dos fármacos , alfa-Sinucleína/genéticaRESUMO
Multiple system atrophy (MSA) is a fatal demyelinating disorder lacking any disease-modifying therapies. MSA pathology stems from aggregated α-synuclein (aSyn) accumulation in glial cytosolic inclusions of oligodendroglial cell (OLGs), the myelinating cells of brain. In MSA brains and in MSA animal models with aSyn accumulation in OLGs, aberrant expression of brain-derived neurotrophic factor (BDNF) and glial-cell-line-derived neurotrophic factor (GDNF) occur. Nerve growth factor (NGF) expression can also be altered in neurodegenerative diseases. It is unclear if oxidative stress impacts the viability of aSyn-accumulating OLG cells. Here, we show that OLN-93â¯cells stably expressing human wild type aSyn or the MSA-associated-aSyn-mutants G51D or A53E, are more vulnerable to oxidative stress. In dose response studies we found that OLN-93â¯cells treated 48â¯h with 160â¯nM FTY720 or our new non-immunosuppressive FTY720-C2 or FTY720-Mitoxy derivatives sustained normal viability. Also, FTY720, FTY720-C2, and FTY720-Mitoxy all stimulated NGF expression at 24â¯h. However only FTY720-Mitoxy also increased BDNF and GDNF mRNA at 24â¯h, an effect paralleled by increases in histone 3 acetylation and ERK1/2 phosphorylation. Myelin associated glycoprotein (MAG) levels were also increased in OLN-93â¯cells after 48â¯h treatment with FTY720-Mitoxy. FTY720, FTY720-C2, and FTY720-Mitoxy all prevented oxidative-stress-associated-cell-death of OLN-93â¯cells that lack any aSyn expression. However, only FTY720-Mitoxy protected MSA-like aSyn-expressing-OLN-93-cells against oxidative-cell-death. These data identify potent protective effects for FTY720-Mitoxy with regard to trophic factors as well as MAG expression by OLG cells. Testing of FTY720-Mitoxy in mice is thus a judicious next step for neuropharmacological preclinical development.
Assuntos
Ceramidas/farmacologia , Cloridrato de Fingolimode/análogos & derivados , Atrofia de Múltiplos Sistemas/metabolismo , Oligodendroglia/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Moduladores do Receptor de Esfingosina 1 Fosfato/farmacologia , alfa-Sinucleína/efeitos dos fármacos , Animais , Fator Neurotrófico Derivado do Encéfalo/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Linhagem Celular , Cloridrato de Fingolimode/farmacologia , Fator Neurotrófico Derivado de Linhagem de Célula Glial/efeitos dos fármacos , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Glicoproteína Associada a Mielina/efeitos dos fármacos , Glicoproteína Associada a Mielina/metabolismo , Fator de Crescimento Neural/efeitos dos fármacos , Fator de Crescimento Neural/metabolismo , Oligodendroglia/metabolismo , Ratos , alfa-Sinucleína/metabolismoRESUMO
Parkinson's disease (PD) is a progressive aging disorder that affects millions worldwide, thus, disease-modifying-therapies are urgently needed. PD pathology includes α-synuclein (aSyn) accumulation as synucleinopathy. Loss of GM1 gangliosides occurs in PD brain, which is modeled in GM2 synthase transgenic mice. GM2+/- mice have low, not absent GM1 and develop age-onset motor deficits, making them an excellent PD drug testing model. FTY720 (fingolimod) reduces synucleinopathy in A53T aSyn mice and motor dysfunction in 6-OHDA and rotenone PD models, but no one has tested FTY720 in mice that develop age-onset PD-like motor problems. We confirmed that GM2+/-mice had equivalent rotarod, hindlimb reflexes, and adhesive removal functions at 9 mo. From 11 mo, GM2+/- mice received oral FTY720 or vehicle 3x/week to 16 mo. As bladder problems occur in PD, we also assessed GM2+/- bladder function. This allowed us to demonstrate improved motor and bladder function in GM2+/- mice treated with FTY720. By immunoblot, FTY720 reduced levels of proNGF, a biomarker of bladder dysfunction. In humans with PD, arm swing becomes abnormal, and brachial plexus modulates arm swing. Ultrastructure of brachial plexus in wild type and GM2 transgenic mice confirmed abnormal myelination and axons in GM2 transgenics. FTY720 treated GM2+/- brachial plexus sustained myelin associated protein levels and reduced aggregated aSyn and PSer129 aSyn levels. FTY720 increases brain derived neurotrophic factor (BDNF) and we noted increased BDNF in GM2+/- brachial plexus and cerebellum, which contribute to rotarod performance. These findings provide further support for testing low dose FTY720 in patients with PD.
Assuntos
Comportamento Animal/efeitos dos fármacos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Cloridrato de Fingolimode/farmacologia , Doença de Parkinson Secundária/tratamento farmacológico , Moduladores do Receptor de Esfingosina 1 Fosfato/farmacologia , alfa-Sinucleína/metabolismo , Animais , Encéfalo/metabolismo , Cloridrato de Fingolimode/uso terapêutico , Camundongos , Camundongos Transgênicos , Destreza Motora/efeitos dos fármacos , N-Acetilgalactosaminiltransferases/genética , N-Acetilgalactosaminiltransferases/metabolismo , Doença de Parkinson Secundária/metabolismo , Teste de Desempenho do Rota-Rod , Moduladores do Receptor de Esfingosina 1 Fosfato/uso terapêuticoRESUMO
BACKGROUND: Patch testing is the "gold standard" to identify culprit allergen(s) causing allergic contact dermatitis (ACD), but there are limited studies of patch testing from allergy practice settings. OBJECTIVE: We sought to explore patch test findings in a large academic allergy practice, including patch testing results, history of atopy, location of dermatitis, and referral source. We also wanted to determine whether patch testing using an extended panel, such as the North American screening series, compared with a limited series, such as the Thin-Layer Rapid-Use Epicutaneous (T.R.U.E.) Test, increased the sensitivity. METHODS: A retrospective chart review was conducted of patients referred for patch testing over a 6-year period. RESULTS: A total of 585 patients (mean age 48.7 years, 71.6 % female) underwent patch testing over the 6-year period, of which 369 (63%) had a positive test. Of those who tested positive, 202 (55%) reported a history of atopy. The extremities were the most commonly involved site, followed by the head/neck and trunk. The 5 most common positive allergens were nickel sulfate, gold sodium thiosulfate, methylchloroisothiazolinone, thimerosal, and bacitracin. Three hundred fourteen (53.6%) patients were positive to at least 1 allergen on TRUE testing. Extended screening series identified an additional 10.8% of patients with positive tests who were negative to T.R.U.E. test allergens. CONCLUSION: Patch testing is a valuable diagnostic tool for the practicing allergist and provides early identification of culprit allergens in ACD. Performing an extended screening series such as the North American Contact Dermatitis Group (NACDG) or supplemental panel of allergens increased sensitivity when compared with a limited series.
Assuntos
Alérgenos/administração & dosagem , Dermatite Alérgica de Contato/diagnóstico , Testes do Emplastro , Pele/efeitos dos fármacos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Bacitracina/administração & dosagem , Criança , Pré-Escolar , Dermatite Alérgica de Contato/imunologia , Dermatite Alérgica de Contato/fisiopatologia , Feminino , Tiossulfato Sódico de Ouro/administração & dosagem , Humanos , Masculino , Pessoa de Meia-Idade , Níquel/administração & dosagem , Estudos Retrospectivos , Pele/imunologia , Pele/fisiopatologia , Tiazóis/administração & dosagem , Timerosal/administração & dosagemAssuntos
Doenças do Tecido Conjuntivo/tratamento farmacológico , Hipersensibilidade a Drogas/tratamento farmacológico , Neoplasias/tratamento farmacológico , Rituximab/efeitos adversos , Rituximab/uso terapêutico , Dessensibilização Imunológica/métodos , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
In searching for Parkinson's disease (PD) pharmacotherapies we began studying FTY720, a food and drug administration (FDA) approved drug. We also created derivatives, FTY720-C2 and FTY720-Mitoxy, and began assessing them. Here we treated dopaminergic MN9D cells with FTY720s then measured microRNA (miRNA) levels by PCR arrays. We discovered that all three FTY720s increased miR376b-3p, while FTY720-C2 also increased miR-128-3p, miR-146b-5p, miR-7a-5p, and miR-9-5p, and FTY720-Mitoxy also increased miR-30d-5p. Investigations revealed that some miRNAs downregulate alpha-synuclein, while others reduce apoptosis, suggesting that FTY720s may act to reduce synucleinopathy and dopaminergic neuron loss in PD and related disorders.
Assuntos
Ceramidas/farmacologia , Neurônios Dopaminérgicos/metabolismo , Cloridrato de Fingolimode/análogos & derivados , Cloridrato de Fingolimode/farmacologia , MicroRNAs/metabolismo , Fármacos Neuroprotetores/metabolismo , Regulação para Cima/efeitos dos fármacos , Animais , Células Cultivadas , CamundongosAssuntos
Dermatite Alérgica de Contato/diagnóstico , Ouro/análise , Testes do Emplastro/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alérgenos/imunologia , Criança , Pré-Escolar , Cobalto/imunologia , Feminino , Ouro/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Níquel/imunologia , Estudos Retrospectivos , Adulto JovemRESUMO
Characterizing the normal function(s) of the protein α-Synuclein (aSyn) has the potential to illuminate links between Parkinson's disease (PD) and diabetes and also point the way toward new therapies for these disorders. Here we provide a perspective for consideration based on our discovery that aSyn normally acts to inhibit insulin secretion from pancreatic ß-cells by interacting with the Kir6.2 subunit of the ATP-sensitive potassium channel (K-ATP). It is also known that K-ATP channels act to inhibit brain dopamine secretion, and we have also shown that aSyn is a normal inhibitor of dopamine synthesis. The finding, that aSyn modulates Kir6.2 and other proteins involved in dopamine and insulin secretion, suggests that aSyn interacting proteins may be negatively impacted when aSyn aggregates inside cells, whether in brain or pancreas. Furthermore, identifying therapies for PD that can counteract dysfunction found in diabetes, would be highly beneficial. One such compound may be the multiple sclerosis drug, FTY720, which like aSyn can stimulate the activity of the catalytic subunit of protein phosphatase 2A (PP2Ac) as well as insulin secretion. In aging aSyn transgenic mice given long term oral FTY720, the mice had reduced aSyn pathology and increased levels of the protective molecule, brain derived neurotrophic factor (BDNF) (Vidal-Martinez et al., 2016). In collaboration with medicinal chemists, we made two non-immunosuppressive FTY720s that also enhance PP2Ac activity, and BDNF expression (Vargas-Medrano et al., 2014; Enoru et al., 2016; Segura-Ulate et al., 2017a). FTY720 and our novel FTY720-based-derivatives, may thus have therapeutic potential for both diabetes and PD.
RESUMO
BACKGROUND: An array of glycoside hydrolases with multiple substrate specificities are required to digest plant cell wall polysaccharides. Cel5E from Clostridium thermocellum and Cel5A from Thermotoga maritima are two glycoside hydrolase family 5 (GH5) enzymes with high sequence and structural similarity, but notably possess different substrate specificities; the former is a bifunctional cellulase/xylanase and the latter is a cellulase/mannanase. A specific loop in TmCel5A, Tmloop, is one of the most structurally divergent regions compared to CtCel5E and interacts with substrates, suggesting the importance for mannan recognition. METHOD: A Tmloop inserted CtCel5E and its related mutants were produced to investigate the role of Tmloop in catalysis. Crystal structure of CtCel5E-TmloopF267A followed by site-direct mutagenesis reveals the mechanism. RtCelB, a homolog with Tmloop was identified to have mannanase activity. RESULT: Tmloop incorporation enables CtCel5E to gain mannanase activity. Tyr270, His277, and Trp282 in the Tmloop are indispensable for CtCel5E-Tmloop catalysis, and weakening hydrophobic environment near the Tmloop enhances enzyme kcat. Using our newly identified loop motif to search for structurally conserved homologs in other subfamilies of GH5, we identified RtCelB. This homolog, originally annotated as a cellulase also possesses mannanase and xylanase activities. CONCLUSION: Our studies show that Tmloop enhances GH5 enzyme promiscuity and plays a role in catalysis. GENERAL SIGNIFICANCE: The study identified a loop of GH5 for mannan recognition and catalysis. Weakening the hydrophobic environment near the loop can also enhance the enzyme catalytic rate. Our findings provide a new insight on mannan recognition and activity enhancement of GH5.
Assuntos
Proteínas de Bactérias/química , Celulase/química , Glucanos/metabolismo , Mananas/metabolismo , Thermotoga maritima/enzimologia , Xilanos/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Catálise , Celulase/genética , Celulase/metabolismo , Clostridium thermocellum/enzimologia , Cristalografia por Raios X , Ativação Enzimática , Modelos Moleculares , Família Multigênica , Mutagênese Sítio-Dirigida , Polissacarídeos/metabolismo , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade por Substrato , Thermotoga maritima/genéticaRESUMO
Multiple system atrophy (MSA) is a demyelinating neurodegenerative disorder characterized by accumulation of aggregated α-synuclein (aSyn) inside oligodendrocyte precursors, mature oligodendroglia, and neurons. MSA dysfunction is associated with loss of trophic factor production by glial and neuronal cells. Here, we report that recombinant wild type human aSyn uptake by OLN-93, an oligodendroglia cell-line, reduced brain-derived neurotrophic factor (BDNF) expression. Furthermore, OLN-93 cells stably transfected with human wild type or an MSA-associated mutant aSyn, A53E that produces neuronal and glial inclusions, reduced BDNF mRNA to nearly unmeasurable qPCR levels. Curiously, another MSA-associated aSyn mutant, G51D that also produces neuronal and glial inclusions, caused only a trend toward BDNF mRNA reduction in transfected OLN-93 cells. This suggests that oligodendrocyte-associated BDNF loss occurs in response to specific aSyn types. Treating OLN-93 cells with 160 nM FTY720 (Fingolimod, Gilenya®), a Food and Drug Administration (FDA) approved therapeutic for multiple sclerosis, counteracted BDNF downregulation in all aSyn OLN-93 cells. FTY720 also restored BDNF mRNA in OLN-93 cells treated with recombinant aSyn, as measured by qPCR or semiquantitatively on agarose gels. Immunoblots confirmed that FTY720 increased histone 3 acetylation in OLN-93, and chromatin immunoprecipitation assays showed increased acetylated histone 3 at BDNF promoter 1 after FTY720. Moreover, OLN-93 cells treated with valproic acid, a classic histone deacetylase inhibitor, confirmed that increasing acetylated histone 3 levels increases BDNF expression. Cumulatively, the data suggest that FTY720-associated histone deacetylase inhibition stimulates BDNF expression in oligodendroglial cells, raising the possibility that MSA patients may also benefit by treatment with FTY720.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/biossíntese , Cloridrato de Fingolimode/farmacologia , alfa-Sinucleína/antagonistas & inibidores , alfa-Sinucleína/metabolismo , Acetilação/efeitos dos fármacos , Animais , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Histonas/metabolismo , Humanos , Mutação , Neuroglia/metabolismo , Neurônios/metabolismo , Oligodendroglia/metabolismo , Ratos , Ácido Valproico/farmacologia , alfa-Sinucleína/genética , alfa-Sinucleína/farmacologia , beta-Sinucleína/metabolismoRESUMO
Parkinson's disease (PD) is a neurodegenerative aging disorder in which postmortem PD brain exhibits neuroinflammation, as well as synucleinopathy-associated protein phosphatase 2A (PP2A) enzymatic activity loss. Based on our translational research, we began evaluating the PD-repurposing-potential of an anti-inflammatory, neuroprotective, and PP2A stimulatory oral drug that is FDA-approved for multiple sclerosis, FTY720 (fingolimod, Gilenya®). We also designed two new FTY720 analogues, FTY720-C2 and FTY720-Mitoxy, with modifications that affect drug potency and mitochondrial localization, respectively. Herein, we describe the metabolic stability and metabolic profiling of FTY720-C2 and FTY720-Mitoxy in liver microsomes and hepatocytes. Using mouse, rat, dog, monkey, and human liver microsomes the intrinsic clearance of FTY720-C2 was 22.5, 79.5, 6.0, 20.2 and 18.3 µL/min/mg; and for FTY720-Mitoxy was 1.8, 7.8, 1.4, 135.0 and 17.5 µL/min/mg, respectively. In hepatocytes, both FTY720-C2 and FTY720-Mitoxy were metabolized from the octyl side chain, generating a series of carboxylic acids similar to the parent FTY720, but without phosphorylated metabolites. To assess absorption and distribution, we gave equivalent single intravenous (IV) or oral doses of FTY720-C2 or FTY720-Mitoxy to C57BL/6 mice, with two mice per time point evaluated. After IV delivery, both FTY720-C2 and FTY720-Mitoxy were rapidly detected in plasma and brain; and reached peak concentrations at the first sampling time points. After oral dosing, FTY720-C2 was present in plasma and brain, although FTY720-Mitoxy was not orally bioavailable. Brain-to-plasma ratio of both compounds increased time-dependently, suggesting a preferential partitioning to the brain. PP2A activity in mouse adrenal gland increased ~2-fold after FTY720-C2 or FTY720-Mitoxy, as compared to untreated controls. In summary, FTY720-C2 and FTY720-Mitoxy both (i) crossed the blood-brain-barrier; (ii) produced metabolites similar to FTY720, except without phosphorylated species that cause S1P1-mediated-immunosuppression; and (iii) stimulated in vivo PP2A activity, all of which encourage additional preclinical assessment.