RESUMEN

Mycobacterium abscessus (Mab) is an opportunistic nontuberculous mycobacterium responsible of difficult-to-treat pulmonary infections in vulnerable patients, such as those suffering from Cystic Fibrosis (CF), where it represents a major cause of morbidity and mortality. Additionally, due to the intrinsic extensive antimicrobial resistance spectrum displayed by this species and the side effects reported for some available antibiotics, the therapeutic management of such infections remains extremely difficult. In the present study, we show that phosphatidylserine liposomes (PS-L) enhance intracellular mycobacterial killing of Mab infected human macrophages with functional or pharmacologically inhibited cystic fibrosis conductance regulator (CFTR), by a mechanism involving phagosome acidification and reactive oxygen species (ROS) production. Additionally, PS-L significantly reduce proinflammatory response of Mab infected macrophages in terms of NF-kB activation and TNF-α production, irrespective of CFTR inhibition. Altogether, these results represent the proof of concept for a possible future development of PS-L as a therapeutic strategy against difficult-to-treat Mab infection.

Asunto(s)

Liposomas , Macrófagos , Infecciones por Mycobacterium no Tuberculosas , Mycobacterium abscessus , Fagosomas , Fosfatidilserinas , Especies Reactivas de Oxígeno , Humanos , Mycobacterium abscessus/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Liposomas/metabolismo , Macrófagos/microbiología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Macrófagos/inmunología , Fagosomas/microbiología , Fagosomas/metabolismo , Fosfatidilserinas/metabolismo , Infecciones por Mycobacterium no Tuberculosas/microbiología , Factor de Necrosis Tumoral alfa/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , FN-kappa B/metabolismo , Fibrosis Quística/microbiología

RESUMEN

Two cystic fibrosis (CF) rat models, one carrying the common Phe508del mutation and the other a nonsense cystic fibrosis transmembrane conductance regulator (CFTR) mutation (knockout) were previously characterised. Although relevant CFTR mRNA reductions were present in the lung, no overt CF lung disease was observed. This study used flexiVent lung mechanic assessment and regional ventilation assessment via X-ray velocimetry (XV) functional imaging to assess the lung phenotype in both models. To determine the sensitivity of XV regional ventilation imaging, the effect of a localised physical obstruction (delivery of agar beads to part of the lungs) on lung ventilation was examined. At baseline, Phe508del and knockout CF rats had a lower inspiratory capacity, total respiratory system compliance, and static compliance than wildtype rats. Following agar bead delivery all XV ventilation parameters were altered, with substantial increases in poorly ventilated regions and ventilation heterogeneity. XV ventilation maps accurately identified locations of bead-induced airflow changes. Despite unremarkable lung histopathology, this study indicated that CF rats display altered respiratory mechanics, with CF rats needing to exert additional effort to expand and deflate their lungs due to increased stiffness. This study demonstrated the utility of XV imaging providing spatial lung ventilation information.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Modelos Animales de Enfermedad , Pulmón , Mecánica Respiratoria , Animales , Fibrosis Quística/diagnóstico por imagen , Fibrosis Quística/genética , Fibrosis Quística/fisiopatología , Ratas , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Pulmón/diagnóstico por imagen , Pulmón/metabolismo , Pulmón/patología , Pulmón/fisiopatología , Reología , Masculino , Ratas Sprague-Dawley

RESUMEN

BACKGROUND: Predictions based on patient-derived materials of CFTR modulators efficacy have been performed lately in patient-derived cells, extending FDA-approved drugs for CF patients harboring rare variants. Here we developed intestinal organoids from subjects carrying S737F- and T465N-CFTR in trans with null alleles to evaluate their functional impact on CFTR protein function and their restoration upon CFTR modulator treatment. The characterization of S737F-CFTR was performed in two subjects recently assessed in nasal epithelial cells but not in colonoids. RESULTS: Our functional analysis (Ussing chamber) confirmed that S737F-CFTR is a mild variant with residual function as investigated in colonoids of patients with S737F/Dele22-24 and S737F/W1282X genotypes. An increase of current upon Elexacaftor/Tezacaftor/Ivacaftor (ETI) treatment was recorded for the former genotype. T465N is a poorly characterized missense variant that strongly impacts CFTR function, as almost no CFTR-mediated anion secretion was registered for T465N/Q39X colonoids. ETI treatment substantially improved CFTR-mediated anion secretion and increased the rescue of mature CFTR expression compared to either untreated colonoids or to dual CFTR modulator therapies. CONCLUSIONS: Our study confirms the presence of a residual function of the S737F variant and its limited response to CFTR modulators while predicting for the first time the potential clinical benefit of Trikafta® for patients carrying the rare T465N variant.

Asunto(s)

Aminofenoles , Benzodioxoles , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Organoides , Quinolonas , Humanos , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Organoides/metabolismo , Organoides/efectos de los fármacos , Benzodioxoles/farmacología , Fibrosis Quística/genética , Fibrosis Quística/tratamiento farmacológico , Fibrosis Quística/metabolismo , Fibrosis Quística/patología , Quinolonas/farmacología , Aminofenoles/farmacología , Indoles/farmacología , Combinación de Medicamentos , Pirazoles/farmacología , Masculino , Femenino , Quinolinas/farmacología , Piridinas , Pirrolidinas

RESUMEN

The correction of protein folding is fundamental for cellular functionality and its failure can lead to severe diseases. In this context, molecular chaperones are crucial players involved in the tricky process of assisting in protein folding, stabilization, and degradation. Chaperones, such as heat shock proteins (HSP) 90, 70, and 60, operate within complex systems, interacting with co-chaperones both to prevent protein misfolding and direct to the correct folding. Chaperone targeting drugs could represent a challenging approach for the treatment of cystic fibrosis (CF), an autosomal recessive genetic disease caused by mutations in the CFTR gene, encoding for the CFTR chloride channel. In this review, we discuss the potential role of molecular chaperones as proteostasis modulators affecting CFTR biogenesis. In particular, we focused on HSP90 and HSP70, for their key role in CFTR folding and trafficking, as well as on HSP60 for its involvement in the inflammation process.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Fibrosis Quística/tratamiento farmacológico , Fibrosis Quística/metabolismo , Fibrosis Quística/genética , Humanos , Chaperonas Moleculares/metabolismo , Pliegue de Proteína/efectos de los fármacos , Proteínas HSP90 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Animales , Chaperonina 60/metabolismo , Chaperonina 60/química , Chaperonina 60/antagonistas & inhibidores , Proteínas HSP70 de Choque Térmico/metabolismo

RESUMEN

Just over thirty years is the span of a generation. It is also the time that has passed since the discovery of the gene responsible for cystic fibrosis. Today, it is safe to say that this discovery has revolutionized our understanding, research perspectives, and management of this disease, which was, thirty years ago, a pediatric condition with a grim prognosis. The aim of this review is to present the advances that science and medicine have brought to our understanding of the pathophysiology of the disease and its management, which in many ways, epitomizes modern molecular genetic research. Since the discovery of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in 1989, modeling the CFTR protein, deciphering its function as an ion channel, and identifying its molecular partners have led to numerous therapeutic advances. The most significant advancement in this field has been the discovery of protein modulators that can target its membrane localization and chloride channel activity. However, further progress is needed to ensure that all patients can benefit from a therapy tailored to their mutations, with the primary challenge being the development of treatments for mutations leading to a complete absence of the protein. The present review delves into the history of the multifaceted world of CF, covering main historical facts, current landscape, clinical management, emerging therapies, patient perspectives, and the importance of ongoing research, bridging science and medicine in the fight against the disease.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Fibrosis Quística/terapia , Fibrosis Quística/genética , Fibrosis Quística/metabolismo , Humanos , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Mutación , Animales

Asunto(s)

Aminofenoles , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Quinolonas , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Aminofenoles/uso terapéutico , Benzodioxoles/uso terapéutico , Benzodioxoles/farmacología , Fibrosis Quística/tratamiento farmacológico , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Mutación , Pirazoles/uso terapéutico , Quinolinas/uso terapéutico , Quinolonas/uso terapéutico

RESUMEN

Cystic fibrosis (CF) is inherited by CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations. A variety of mutations have been identified in the CFTR gene that may be associated with cystic fibrosis, and these mutations demonstrate extensive molecular genetic heterogeneity in this disease. Little is known about the molecular mechanism by which mutations affect CFTR function, and only a minority of mutations have been characterized by functional studies. There has been an increase in the number of complex alleles. This may partly explain the difficulty in establishing genotype-phenotype correlations and complicate genetic counseling and diagnosis in some cases. Therefore, the identification of complex alleles has several important implications for recessive disorders. This will facilitate diagnosis; improve judgements concerning prognosis, and enable appropriate genetic counselling for affected families. This review describes the complex cystic fibrosis allele to better understand the contribution of this allele in the wide phenotypic variability of cystic fibrosis disease. It occurs in the complex allele that the second cis mutation can modulate the effects of the first mutation or vice versa. The phenotypic variability between CF or CFTR-RD (CFTR related disease) patients may be due to several factors, including different genetic and environmental backgrounds. It is important to determine the allele complex so that optimal treatment can be established.

Asunto(s)

Alelos , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Mutación , Fenotipo , Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Humanos , Mutación/genética

RESUMEN

A thin film of pulmonary surfactant lines the surface of the airways and alveoli, where it lowers the surface tension in the peripheral lungs, preventing collapse of the bronchioles and alveoli and reducing the work of breathing. It also possesses a barrier function for maintaining the blood-gas interface of the lungs and plays an important role in innate immunity. The surfactant film covers the epithelium lining both large and small airways, forming the first line of defense between toxic airborne particles/pathogens and the lungs. Furthermore, surfactant has been shown to relax airway smooth muscle (ASM) after exposure to ASM agonists, suggesting a more subtle function. Whether surfactant masks irritant sensory receptors or interacts with one of them is not known. The relaxant effect of surfactant on ASM is absent in bronchial tissues denuded of an epithelial layer. Blocking of prostanoid synthesis inhibits the relaxant function of surfactant, indicating that prostanoids might be involved. Another possibility for surfactant to be active, namely through ATP-dependent potassium channels and the cAMP-regulated epithelial chloride channels [cystic fibrosis transmembrane conductance regulators (CFTRs)], was tested but could not be confirmed. Hence, this review discusses the mechanisms of known and potential relaxant effects of pulmonary surfactant on ASM. This review summarizes what is known about the role of surfactant in smooth muscle physiology and explores the scientific questions and studies needed to fully understand how surfactant helps maintain the delicate balance between relaxant and constrictor needs.

Asunto(s)

Músculo Liso , Surfactantes Pulmonares , Humanos , Surfactantes Pulmonares/metabolismo , Músculo Liso/efectos de los fármacos , Músculo Liso/metabolismo , Animales , Tono Muscular/efectos de los fármacos , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo

Asunto(s)

Fibrosis Quística , Fibrosis Quística/fisiopatología , Fibrosis Quística/complicaciones , Humanos , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética

RESUMEN

BACKGROUND: Pancreatic fibrosis is an early diagnostic feature of the common inherited disorder cystic fibrosis (CF). Many people with CF (pwCF) are pancreatic insufficient from birth and the replacement of acinar tissue with cystic lesions and fibrosis is a progressive phenotype that may later lead to diabetes. Little is known about the initiating events in the fibrotic process though it may be a sequela of inflammation in the pancreatic ducts resulting from loss of CFTR impairing normal fluid secretion. Here we use a sheep model of CF (CFTR-/-) to examine the evolution of pancreatic disease through gestation. METHODS: Fetal pancreas was collected at six time points from 50-days of gestation through to term, which is equivalent to ~ 13 weeks to term in human. RNA was extracted from tissue for bulk RNA-seq and single cells were prepared from 80-day, 120-day and term samples for scRNA-seq. Data were validated by immunochemistry. RESULTS: Transcriptomic evidence from bulk RNA-seq showed alterations in the CFTR-/- pancreas by 65-days of gestation, which are accompanied by marked pathological changes by 80-days of gestation. These include a fibrotic response, confirmed by immunostaining for COL1A1, αSMA and SPARC, together with acinar loss. Moreover, using scRNA-seq we identify a unique cell population that is significantly overrepresented in the CFTR-/- animals at 80- and 120-days gestation, as are stellate cells at term. CONCLUSION: The transcriptomic changes and cellular imbalance that we observe likely have pivotal roles in the evolution of CF pancreatic disease and may provide therapeutic opportunities to delay or prevent pancreatic destruction in CF.

Asunto(s)

Biomarcadores , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Modelos Animales de Enfermedad , Células Estrelladas Pancreáticas , Fibrosis Quística/genética , Fibrosis Quística/metabolismo , Fibrosis Quística/patología , Animales , Células Estrelladas Pancreáticas/metabolismo , Células Estrelladas Pancreáticas/patología , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Femenino , Ovinos , Páncreas/metabolismo , Páncreas/patología , Embarazo , Enfermedades Pancreáticas/genética , Enfermedades Pancreáticas/metabolismo , Enfermedades Pancreáticas/patología , Transcriptoma , Humanos , Perfilación de la Expresión Génica

RESUMEN

Cystic fibrosis (CF) is the most common life-threatening genetic disease in the United States and among people of European descent. Despite the widespread distribution of the cystic fibrosis transmembrane conductance regulator (CFTR) along kidney tubules, specific renal phenotypes attributable to CF have not been well documented. Recent studies have demonstrated the downregulation of the apical Cl-/HCO3 - exchanger pendrin (Slc26a4) in kidney B-intercalated cells of CF mouse models. These studies have shown that kidneys of both mice and humans with CF have an impaired ability to excrete excess HCO3 -, thus developing metabolic alkalosis when subjected to excess HCO3 - intake. The purpose of this minireview is to discuss the latest advances on the role of pendrin as a molecule with dual critical roles in acid base regulation and systemic vascular volume homeostasis, specifically in CF. Given the immense prevalence of vascular volume depletion, which is primarily precipitated via enhanced chloride loss through perspiration, we suggest that the dominant presentation of metabolic alkalosis in CF is due to the impaired function of pendrin, which plays a critical role in systemic vascular volume and acid base homeostasis.

Asunto(s)

Alcalosis , Bicarbonatos , Fibrosis Quística , Transportadores de Sulfato , Humanos , Fibrosis Quística/metabolismo , Fibrosis Quística/patología , Fibrosis Quística/complicaciones , Alcalosis/metabolismo , Alcalosis/etiología , Bicarbonatos/metabolismo , Animales , Transportadores de Sulfato/metabolismo , Transportadores de Sulfato/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Ratones

RESUMEN

Our study probed the differences in ion channel gene expression in the endometrium of women with Recurrent Implantation Failure (RIF) compared to fertile women. We analyzed the relative expression of genes coding for T-type Ca2+, ENaC, CFTR, and KCNQ1 channels in endometrial samples from 20 RIF-affected and 10 control women, aged 22-35, via microarray analysis and quantitative real-time PCR. Additionally, we examined DNA methylation in the regulatory region of KCNQ1 using ChIP real-time PCR. The bioinformatics component of our research included Gene Ontology analysis, protein-protein interaction networks, and signaling pathway mapping to identify key biological processes and pathways implicated in RIF. This led to the discovery of significant alterations in the expression of ion channel genes in RIF women's endometrium, most notably an overexpression of CFTR and reduced expression of SCNN1A, SCNN1B, SCNN1G, CACNA1H, and KCNQ1. A higher DNA methylation level of KCNQ1's regulatory region was also observed in RIF patients. Gene-set enrichment analysis highlighted a significant presence of genes involved with ion transport and membrane potential regulation, particularly in sodium and calcium channel complexes, which are vital for cation movement across cell membranes. Genes were also enriched in broader ion channel and transmembrane transporter complexes, underscoring their potential extensive role in cellular ion homeostasis and signaling. These findings suggest a potential involvement of ion channels in the pathology of implantation failure, offering new insights into the mechanisms behind RIF and possible therapeutic targets.

Asunto(s)

Metilación de ADN , Implantación del Embrión , Endometrio , Humanos , Femenino , Endometrio/metabolismo , Adulto , Implantación del Embrión/genética , Canal de Potasio KCNQ1/genética , Canal de Potasio KCNQ1/metabolismo , Regulación de la Expresión Génica , Adulto Joven , Canales Iónicos/genética , Canales Iónicos/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Perfilación de la Expresión Génica , Infertilidad Femenina/genética , Infertilidad Femenina/metabolismo , Canales Epiteliales de Sodio/genética , Canales Epiteliales de Sodio/metabolismo

RESUMEN

Cystic fibrosis (CF) is an autosomal recessive inherited disease caused by variants of cystic fibrosis transmembrane conductance regulation (CFTR) gene. This report presents a case of a Chinese boy diagnosed with CF, attributed to the presence of two specific CFTR gene variations: 4056G > C (NM_000492.4) (p.Gln1352His, legacy: Q1352H) and c.1210-34TG[13]T[5] (NM_000492.4)(legacy: 5T; TG13). A ten-year-old boy was admitted to the hospital due to recurrent pneumonia, cough, and intermittent fever for seven years. Lung auscultation revealed rales, and a lung CT scan indicated parenchymal transformation with infection in both lungs. Whole Exome Sequencing (WES) identified two CFTR gene variants, Q1352H and 5T; TG13, which were significantly associated with clinical phenotype. Following a two-year course of azithromycin combined with inhalation therapy with budesonide, the patient experienced no further episodes of respiratory infections. Moreover, significant improvements were observed in pulmonary function, pulmonary infection, and bronchiectasis. The occurrence of combined variations, Q1352H and 5T; TG13, in the CFTR gene is rare and specific to Chinese populations. WES proves to be a valuable diagnostic tool for detecting CFTR gene variants.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Mutación , Humanos , Masculino , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Fibrosis Quística/genética , Fibrosis Quística/diagnóstico , Niño , Pueblo Asiatico/genética , Heterocigoto , Fenotipo , Secuenciación del Exoma , Antibacterianos/uso terapéutico , Pueblos del Este de Asia

RESUMEN

Cystic fibrosis is caused by biallelic pathogenic variants in the CFTR gene, which contains a polymorphic (TG)mTn sequence (the "poly-T/TG tract") in intron 9. While T9 and T7 alleles are benign, T5 alleles with longer TG repeats, e.g., (TG)12T5 and (TG)13T5, are clinically significant. Thus, professional medical societies currently recommend reporting the TG repeat size when T5 is detected. Sanger sequencing is a cost-effective method of genotyping the (TG)mTn tract; however, its polymorphic length substantially complicates data analysis. We developed CFTR-TIPS, a freely available web-based software tool that infers the (TG)mTn genotype from Sanger sequencing data. This tool detects the (TG)mTn tract in the chromatograms, quantifies goodness of fit with expected patterns, and visualizes the results in a graphical user interface. It is broadly compatible with any Sanger chromatogram that contains the (TG)mTn tract ± 15 bp. We evaluated CFTR-TIPS using 835 clinical samples previously analyzed in a CLIA-certified, CAP-accredited laboratory. When operated fully automatically, CFTR-TIPS achieved 99.8% concordance with our clinically validated manual workflow, while generally taking less than 10 s per sample. There were two discordant samples: one due to a co-occurring heterozygous duplication that confounded the tool and the other due to incomplete (TG)mTn tract detection in the reverse chromatogram. No clinically significant misclassifications were observed. CFTR-TIPS is a free, accurate, and rapid tool for CFTR (TG)mTn tract genotyping using cost-effective Sanger sequencing. This tool is suitable both for automated use and as an aid to manual review to enhance accuracy and reduce analysis time.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Genotipo , Técnicas de Genotipaje , Programas Informáticos , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Humanos , Fibrosis Quística/genética , Técnicas de Genotipaje/métodos , Alelos , Análisis de Secuencia de ADN/métodos

RESUMEN

Although substantial advances have been obtained in the pharmacological treatment of cystic fibrosis (CF) with the approval of Kaftrio, a combination of two correctors (VX-661, VX-445) and one potentiator (VX-770), new modulators are still needed to rescue F508del and other CFTR mutants with trafficking defects. We have previously identified PP compounds based on a tricyclic core as correctors with high efficacy in the rescue of F508del-CFTR on native epithelial cells of CF patients, particularly in combination with class 1 correctors (VX-809, VX-661). Compound PP028 was found as a lead candidate for the high rescue of F508del-CFTR and used for mechanistic insight indicating that PP028 behaves as a class 3 corrector, similarly to VX-445. From the exploration of the chemical space around the hit structure, based on iterative cycles of chemical synthesis and functional testing, the class of 6,9-dihydro-5H-pyrrolo [3,2-h]quinazolines with corrector activity was discovered. Within a series of 38 analogues, two derivatives emerged as promising candidates and used for further insight to assess the mechanism of action. Both compounds, decorated with a benzensulfonylamino group at the pyrimidine moiety, were able to generate a dose-dependent increase in CFTR function, particularly in the presence of VX-809. Half-effective concentrations (EC50) were in the single digit micromolar range and decreased in the presence of VX-809 thus indicating a synergistic interaction with class 1 correctors. Synergy was also observed with corr-4a (class 2 corrector) but not with VX-445 and PP028 (class 3 correctors) indicating that the new compounds behave as class 3 correctors. These results suggest that tricyclic pyrrolo-quinazolines interact with CFTR at a site different from that of VX-809 and represent a novel class of CFTR correctors suitable for combinatorial pharmacological treatments for the basic defect in CF.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Quinazolinas , Fibrosis Quística/tratamiento farmacológico , Fibrosis Quística/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Humanos , Quinazolinas/farmacología , Quinazolinas/química , Quinazolinas/síntesis química , Relación Estructura-Actividad , Relación Dosis-Respuesta a Droga , Estructura Molecular , Pirroles/farmacología , Pirroles/química , Pirroles/síntesis química , Mutación

RESUMEN

The relationship between dental fluorosis and alterations in the salivary proteome remains inadequately elucidated. This study aimed to investigate the salivary proteome and fluoride concentrations in urine and drinking water among Thai individuals afflicted with severe dental fluorosis. Thirty-seven Thai schoolchildren, aged 6-16, were stratified based on Thylstrup and Fejerskov fluorosis index scores: 10 with scores ranging from 5 to 9 (SF) and 27 with a score of 0 (NF). Urinary and water fluoride levels were determined using an ion-selective fluoride electrode. Salivary proteomic profiling was conducted via LC-MS/MS, followed by comprehensive bioinformatic analysis. Results revealed significantly elevated urinary fluoride levels in the SF group (p = 0.007), whereas water fluoride levels did not significantly differ between the two cohorts. Both groups exhibited 104 detectable salivary proteins. The NF group demonstrated notable upregulation of LENG9, whereas the SF group displayed upregulation of LDHA, UBA1, S100A9, H4C3, and LCP1, all associated with the CFTR ion channel. Moreover, the NF group uniquely expressed 36 proteins, and Gene Ontology and pathway analyses suggested a link with various aspects of immune defense. In summary, the study hypothesized that the CFTR ion channel might play a predominant role in severe fluorosis and highlighted the depletion of immune-related salivary proteins, suggesting compromised immune defense in severe fluorosis. The utility of urinary fluoride might be a reliable indicator for assessing excessive fluoride exposure.

Asunto(s)

Fluoruros , Fluorosis Dental , Proteómica , Saliva , Fluorosis Dental/metabolismo , Humanos , Niño , Masculino , Saliva/metabolismo , Saliva/química , Femenino , Fluoruros/orina , Fluoruros/análisis , Adolescente , Proteómica/métodos , Proteoma/análisis , Tailandia , Proteínas y Péptidos Salivales/metabolismo , Proteínas y Péptidos Salivales/análisis , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Espectrometría de Masas en Tándem , Agua Potable

RESUMEN

The cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-dependent Cl- channel, is closely associated with multiple pathogen infections, such as SARS-CoV-2. However, whether the function of the CFTR is involved in herpes simplex virus (HSV) infection has not been reported. To evaluate the association of CFTR activity with HSV infection, the antiviral effect of CFTR inhibitors in epithelial cells and HSV-infected mice was tested in this study. The data showed that treatment with CFTR inhibitors in different concentrations, Glyh-101 (5-20 µM), CFTRi-172 (5-20 µM) and IOWH-032 (5-20 µM), or the gene silence of the CFTR could suppress herpes simplex virus 1 (HSV-1) and herpes simplex virus 2 (HSV-2) replication in human HaCaT keratinocytes cells, and that a CFTR inhibitor, Glyh-101 (10-20 µM), protected mice from HSV-1 and HSV-2 infection. Intracellular Cl- concentration ([Cl-]i) was decreased after HSV infection via the activation of adenylyl cyclase (AC)-cAMP signaling pathways. CFTR inhibitors (20 µM) increased the reduced [Cl-]i caused by HSV infection in host epithelial cells. Additionally, CFTR inhibitors reduced the activity and phosphorylation of SGK1 in infected cells and tissues (from the eye and vagina). Our study found that CFTR inhibitors can effectively suppress HSV-1 and HSV-2 infection, revealing a previously unknown role of CFTR inhibitors in HSV infection and suggesting new perspectives on the mechanisms governing HSV infection in host epithelial cells, as well as leading to potential novel treatments.

Asunto(s)

Antivirales , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Herpes Simple , Herpesvirus Humano 1 , Herpesvirus Humano 2 , Replicación Viral , Animales , Ratones , Antivirales/farmacología , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/antagonistas & inhibidores , Humanos , Herpes Simple/tratamiento farmacológico , Herpes Simple/virología , Herpesvirus Humano 1/efectos de los fármacos , Herpesvirus Humano 1/fisiología , Replicación Viral/efectos de los fármacos , Herpesvirus Humano 2/efectos de los fármacos , Herpesvirus Humano 2/fisiología , Femenino , Línea Celular , Células Epiteliales/virología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células HaCaT , Queratinocitos/virología , Queratinocitos/efectos de los fármacos , Ratones Endogámicos BALB C , Chlorocebus aethiops , Simplexvirus/efectos de los fármacos , Simplexvirus/fisiología

RESUMEN

Cystic fibrosis (CF) is caused by the functional expression defect of the cystic fibrosis transmembrane conductance regulator (CFTR) protein. Despite the recent success in CFTR modulator development, the available correctors only partially restore the F508del-CFTR channel function, and several rare CF mutations show resistance to available drugs. We previously identified compound 4172 that synergistically rescued the F508del-CFTR folding defect in combination with the existing corrector drugs VX-809 and VX-661. Here, novel CFTR correctors were designed by applying a classical medicinal chemistry approach on the 4172 scaffold. Molecular docking and three-dimensional quantitative structure-activity relationship (3D-QSAR) studies were conducted to propose a plausible binding site and design more potent and effective analogs. We identified three optimized compounds, which, in combination with VX-809 and the investigational corrector 3151, increased the plasma membrane density and function of F508del-CFTR and other rare CFTR mutants resistant to the currently approved therapies.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Pirazoles , Pirimidinonas , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Pirazoles/farmacología , Pirazoles/síntesis química , Pirazoles/química , Humanos , Pirimidinonas/farmacología , Pirimidinonas/síntesis química , Pirimidinonas/química , Fibrosis Quística/tratamiento farmacológico , Fibrosis Quística/metabolismo , Relación Estructura-Actividad Cuantitativa , Simulación del Acoplamiento Molecular , Benzodioxoles/farmacología , Benzodioxoles/síntesis química , Benzodioxoles/química , Mutación , Aminopiridinas/farmacología , Aminopiridinas/síntesis química , Aminopiridinas/química

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Pancreatitis , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Humanos , Pancreatitis/genética , Pancreatitis/metabolismo , Fibrosis Quística/genética , Fibrosis Quística/metabolismo , Mutación

RESUMEN

Cystic fibrosis (CF) is a serious genetic disease caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Approved small molecule therapies benefit the majority of people with CF (pwCF), but unfortunately not all. Gene addition offers a mutation agnostic treatment option for all pwCF. SP-101 is an adeno-associated virus gene therapy vector (AAV2.5T) that has been optimized for efficient human airway cell transduction, and that contains a functional and regulated shortened human CFTR minigene (hCFTRΔR) with a small synthetic promoter/enhancer. To understand SP-101 airway distribution, activity, and the associated immune response, in vivo studies were performed in wild-type and CF ferrets. After single dose inhaled delivery of SP-101, followed by single dose inhaled doxorubicin (an AAV transduction augmenter) or saline, SP-101 vector genomes were detected throughout the respiratory tract. hCFTRΔR mRNA expression was highest in ferrets also receiving doxorubicin and persisted for the duration of the study (13 weeks). Pre-existing mucus in the CF ferrets did not present a barrier to effective transduction. Binding and neutralizing antibodies to the AAV2.5T capsid were observed regardless of doxorubicin exposure. Only a portion of ferrets exhibited a weak T-cell response to AAV2.5T and no T-cell response was seen against hCFTRΔR. These data strongly support the continued development of inhaled SP-101, followed by inhaled doxorubicin, for the treatment of CF.

Asunto(s)

Regulador de Conductancia de Transmembrana de Fibrosis Quística , Fibrosis Quística , Dependovirus , Doxorrubicina , Hurones , Terapia Genética , Vectores Genéticos , Transgenes , Animales , Fibrosis Quística/terapia , Fibrosis Quística/genética , Doxorrubicina/farmacología , Doxorrubicina/administración & dosificación , Doxorrubicina/uso terapéutico , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Dependovirus/genética , Administración por Inhalación , Terapia Genética/métodos , Vectores Genéticos/administración & dosificación , Vectores Genéticos/genética , Humanos , Modelos Animales de Enfermedad , Expresión Génica