Cystic Fibrosis Drug Research Articles

Could cystic fibrosis drugs be repurposed for treating alcohol‐induced pancreatitis?

Could cystic fibrosis drugs be repurposed for treating alcohol‐induced pancreatitis?

Allosteric inhibition of CFTR gating by CFTRinh-172 binding in the pore

Loss-of-function mutations of the CFTR gene cause the life-shortening genetic disease cystic fibrosis (CF), whereas overactivity of CFTR may lead to secretory diarrhea and polycystic kidney disease. While effective drugs targeting the CFTR protein have been developed for the treatment of CF, little progress has been made for diseases caused by hyper-activated CFTR. Here, we solve the cryo-EM structure of CFTR in complex with CFTRinh-172 (Inh-172), a CFTR gating inhibitor with promising potency and efficacy. We find that Inh-172 binds inside the pore of CFTR, interacting with amino acid residues from transmembrane segments (TMs) 1, 6, 8, 9, and 12 through mostly hydrophobic interactions and a salt bridge. Substitution of these residues lowers the apparent affinity of Inh-172. The inhibitor-bound structure reveals re-orientations of the extracellular segment of TMs 1, 8, and 12, supporting an allosteric modulation mechanism involving post-binding conformational changes. This allosteric inhibitory mechanism readily explains our observations that pig CFTR, which preserves all the amino acid residues involved in Inh-172 binding, exhibits a much-reduced sensitivity to Inh-172 and that the apparent affinity of Inh-172 is altered by the CF drug ivacaftor (i.e., VX-770) which enhances CFTR’s activity through binding to a site also comprising TM8.

Ribosomal Frameshifting Selectively Modulates the Assembly, Function, and Pharmacological Rescue of a Misfolded CFTR Variant.

The cotranslational misfolding of the cystic fibrosis transmembrane conductance regulator chloride channel (CFTR) plays a central role in the molecular basis of cystic fibrosis (CF). The misfolding of the most common CF variant (ΔF508) remodels both the translational regulation and quality control of CFTR. Nevertheless, it is unclear how the misassembly of the nascent polypeptide may directly influence the activity of the translation machinery. In this work, we identify a structural motif within the CFTR transcript that stimulates efficient -1 ribosomal frameshifting and triggers the premature termination of translation. Though this motif does not appear to impact the interactome of wild-type CFTR, silent mutations that disrupt this RNA structure alter the association of nascent ΔF508 CFTR with numerous translation and quality control proteins. Moreover, disrupting this RNA structure enhances the functional gating of the ΔF508 CFTR channel at the plasma membrane and its pharmacological rescue by the CFTR modulators contained in the CF drug Trikafta. The effects of the RNA structure on ΔF508 CFTR appear to be attenuated in the absence of the ER membrane protein complex (EMC), which was previously found to modulate ribosome collisions during "preemptive quality control" of a misfolded CFTR homolog. Together, our results reveal that ribosomal frameshifting selectively modulates the assembly, function, and pharmacological rescue of a misfolded CFTR variant. These findings suggest interactions between the nascent chain, quality control machinery, and ribosome may dynamically modulate ribosomal frameshifting in order to tune the processivity of translation in response to cotranslational misfolding.

Enhanced CFTR modulator efficacy in ΔF508 CFTR mouse organoids by ablation of RFFL ubiquitin ligase

The most common CFTR mutant in cystic fibrosis (CF), ΔF508 CFTR, is eliminated by ubiquitination even in the presence of CF drugs, reducing their therapeutic efficacy. RFFL is one of the ubiquitin ligases that remove ΔF508 CFTR from the cell surface despite treatment with the triple combination of CFTR modulators (TEZ/ELX/IVA) used clinically. Although RFFL knockdown has been shown to enhance the efficacy of TEZ/ELX/IVA in cell culture models, its impact in mouse models has not been evaluated. Here, we demonstrate that RFFL ablation significantly improves the effect of TEZ/ELX/IVA, resulting in enhanced function of ΔF508 CFTR in mouse organoids. Since RFFL knockout mice showed no significant abnormalities, our findings support RFFL inhibition as a promising strategy to improve CFtreatment.

Structure-based discovery of CFTR potentiators and inhibitors

The cystic fibrosis transmembrane conductance regulator (CFTR) is a crucial ion channel whose loss of function leads to cystic fibrosis, whereas its hyperactivation leads to secretory diarrhea. Small molecules that improve CFTR folding (correctors) or function (potentiators) are clinically available. However, the only potentiator, ivacaftor, has suboptimal pharmacokinetics and inhibitors have yet to be clinically developed. Here, we combine molecular docking, electrophysiology, cryo-EM, and medicinal chemistry to identify CFTR modulators. We docked ∼155 million molecules into the potentiator site on CFTR, synthesized 53 test ligands, and used structure-based optimization to identify candidate modulators. This approach uncovered mid-nanomolar potentiators, as well as inhibitors, that bind to the same allosteric site. These molecules represent potential leads for the development of more effective drugs for cystic fibrosis and secretory diarrhea, demonstrating the feasibility of large-scale docking for ion channel drug discovery.

Cystic fibrosis rescued using a reprogrammed porosome secretory machinery

Cystic fibrosis (CF) is a genetic disorder resulting in the secretion of highly viscous mucus in the airways, leading to lung infection, respiratory failure, morbidity and mortality. CF is attributed to mutation in the CF Transmembrane Conductance Regulator (CFTR) gene that codes for a chloride transporting channel at the cell plasma membrane. More than 2,000 mutations in the CF gene have been identified; however, the ΔF508 CFTR is the most common, accounting for approximately 70% of all CFTR mutations. Our earlier studies demonstrate the CFTR protein to be among the nearly 30 proteins constituting the porosome secretory machinery at the plasma membrane in human airway epithelial mucous-secreting cells. Our recent studies show that stimulated human airway epithelial cells pre-exposed to CFTR inhibitors result in loss of mucus secretion, suggesting the involvement of CFTR in porosome-mediated mucus secretion. To further test the hypothesis that CFTR is involved in porosome-mediated mucus secretion in the human airways, and to develop a therapeutic approach to overcome this defect in ΔF508 CFTR human bronchial epithelial cells, the current study was undertaken. Mass spectrometry and Western Blot analysis of porosomes isolated from WT-CFTR Human Bronchial Epithelial (HBE) Cells and ΔF508-CFTR CF HBE cells, demonstrate a varying loss or gain of several porosome proteins, including undetectable levels of the Ras GTPase activating like-protein IQGAP1 in the ΔF508-CFTR CF cells. This suggested that mutation in porosome-associated CFTR protein additionally affects other proteins within the porosome secretory machinery, negatively impacting mucus secretion. Therefore, to ameliorate defects in mucus secretion in CF, the reconstitution of functional porosomes obtained from WT-CFTR HBE cells into the plasma membrane of ΔF508-CFTR mutant cells was performed. Results from the study demonstrate that porosome reconstitution rescues mucus secretion approximately two- to four-fold more effectively than the currently available CF drugs including TRIKAFTA. Acknowledgements: Work presented in this article was supported by the Viron Molecular Medicine Institute and Porosome Therapeutics, Inc., Boston, MA. Competing Financial Interest: This work is patent protected by Porosome Therapeutics, Inc. The authors hold shares in the company. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Contribution of the intestinal current measurement method to assessment of the efficacy of CFTR modulators in cystic fibrosis

Cystic fibrosis (CF) is a disease caused by pathogenic variants of the CFTR gene. In the last decade, the treatment algorithm has entered a new era as several drugs have become available that restore the function of the CFTR chloride channel and are called CFTR modulators. The efficacy and safety of targeted drugs in cystic fibrosis needs to be further investigated using additional assessment methods.The aim of this study was to investigate the role of intestinal current measurement (ICM) in assessing the efficacy of targeted therapy for cystic fibrosis.Methods. The efficacy of CFTR modulator therapy was evaluated in 15 patients, of which 10 were children and 5 were adults. In addition to the ICM method, patients’ clinical parameters, sweat test, and pulmonary function were also evaluated according to clinical guidelines.Results. Patients with genotypes 2143delT/7121G>T and G542X/R785X had no restoration of chloride channel function with elexacaftor + tezacaftor + ivacaftor therapy, and patients with the L467F;F508del genotype with lumacaftor + ivacaftor therapy. In patients with the F508del/F508del, N1303K/G461E, N1303K/3321delG genotype, improvements were noted in terms of the restoration of CFTR channel function during therapy with elexacaftor + tezacaftor + ivacaftor therapy, and in patients with the F508del/F508del genotype during therapy with tezacaftor + ivacaftor therapy and lumacaftor + ivacaftor.Conclusion. Restoring the function of the epithelial chloride channel (CFTR) is the basis for increasing life expectancy in CF. The crucial role of the ICM method in determining the efficacy of CFTR modulators is shown.

The case of the missing mouse—developing cystic fibrosis drugs without using animals

Creating and developing new drugs can take decades, costs millions of dollars, requires untold human effort and usually, takes thousands of animal lives. Despite regulators professing confidence in non-animal approaches and guidance documents that permit submission of non-animal data, toxicity testing is routinely carried out in animals, employing rodents (invariably mice) and non-rodents. However, extensive preclinical testing in animals is still no guarantee that drugs will be safe and/or effective. In fact, more than nine out of every ten drugs that appear safe from animal trials will fail when tested in people, often due to unexplained toxicity or a lack of efficacy. This paper will describe recent advances in drug development where non-animal approaches have been used, to explore how and where these could be applied more widely to revolutionize the drug development pipeline and accelerate the creation of safe and effective medicines. As one case study, we look at the small molecule channel modifiers developed to address the consequences of the mutated chloride channel in the fatal genetic condition, cystic fibrosis. We then take a closer look at where drug development could be accelerated by focusing on innovative, human biology-based testing methods. Finally, we put forward recommendations, targeting all stakeholders, including the public, that will be needed to put this into practice and enable drug development to become more efficient - focusing on human-biology based testing and cutting out the middle-mouse.

NICE approves three cystic fibrosis drugs for NHS use

NICE approves three cystic fibrosis drugs for NHS use

Advances in the Cystic Fibrosis Drug Development Pipeline.

Cystic fibrosis is a genetic disease that results in progressive multi-organ manifestations with predominance in the respiratory and gastrointestinal systems. The significant morbidity and mortality seen in the CF population has been the driving force urging the CF research community to further advance treatments to slow disease progression and, in turn, prolong life expectancy. Enormous strides in medical advancements have translated to improvement in quality of life, symptom burden, and survival; however, there is still no cure. This review discusses the most current mainstay treatments and anticipated therapeutics in the CF drug development pipeline within the mechanisms of mucociliary clearance, anti-inflammatory and anti-infective therapies, restoration of the cystic fibrosis transmembrane conductance regulator (CFTR) protein (also known as highly effective modulator therapy (HEMT)), and genetic therapies. Ribonucleic acid (RNA) therapy, gene transfer, and gene editing are being explored in the hopes of developing a treatment and potential cure for people with CF, particularly for those not responsive to HEMT.

Effect of metabolite and temperature on artificial human sweat characteristics over a very wide frequency range (400 MHz–10.4 GHz) for wireless hydration diagnostic sensors

Sweat is an important biofluid that is excreted by the human body. It contains physiological biomarkers that provide vital information on the general health condition of the body. As a result, analysis of electrolytes in this biofluid is gaining traction in the development of non-invasive sweat diagnostics to determine level of dehydration, cystic fibrosis and illicit drugs. This paper presents a wireless measurement modality of permittivity properties of sweat of different metabolite concentration levels exemplifying the full spectrum of human sweat. The artificial sweat used in the study is an analogue of actual human sweat. It is synthesized from a composite mixture of metabolites and minerals including sodium chloride, potassium chloride, Urea and lactic acid. The effect of ambient temperature on the permittivity measurements of the various sweat solutions are studied across a frequency range of 400 MHz to 10.4 GHz. This information is important in the development of wireless radio frequency (RF) non-invasive biosensors.

Traits, trends and hits of orphan drug designations in cystic fibrosis

BackgroundIn the United States (US) and in Europe, cystic fibrosis (CF) qualifies as a rare disease, thus positioning the field to benefit from regulatory incentives provided by orphan drug designation (ODD) to boost pharmaceutical research and development.In this study, we analyzed the pool of products for the treatment of CF that received such incentives from the US Food and Drug Administration (FDA) and/or the European Medicines Agency (EMA) over the past two decades. We describe the characteristics and trends in ODDs over time and explore factors that might be determinants of successful drug development. MethodsWe collected the products that received the ODD from the registries of the FDA and the EMA from 2000 to 2021, characterizing their nature, development stage, and type of sponsor. We categorized the study drugs according to the therapeutic target addressed and described trends of drug development over the study period. A logistic regression analysis was done to assess how ODD characteristics were associated with the approval for market authorization. ResultsFrom 2000–2021, 107 ODDs were collectively granted by the FDA and the EMA for products developed for the treatment of CF. Although the trends of the number of ODDs granted remained stable over time, those targeting the CF basic protein defect increased from 6 out of 54 (11.1%) in the first half of the study period up to 20 out of 54 (37.7%) in the second half, while those treating symptoms decreased from 48/54 (88.9%) to 33/53 (62.3%).Overall, 10 products obtained marketing approval: 7 in both the US and Europe, 3 only in Europe. All the approved ODDs were chemical products for chronic use. No statistically significant difference was found across the examinated variables, but we observed possible drivers of successful drug development for ODDs targeting CFTR, as well as for those with active substances previously marketed, and for those developed by large companies and companies with experience in developing orphan drugs. By contrast, our findings suggest that financial issues most hamper the development of ODDs sponsored by small-medium enterprises. ConclusionsAlthough ODDs for treating infection and other CF sequelae accounted for the majority, we observed a shift of ODDs toward mechanism-based products over the study period.In line with other rare diseases, we found that approximately 1/10 ODDs for CF reached the status of marketing approval.Advances in disease genetics paved the way for a shift in CF drug development; however, we described how the convergence of pharmaceutical technology, the financial environment, and the regulatory ecosystem played a crucial role in successful marketing authorization in CF.

Innovative Strategy toward Mutant CFTR Rescue in Cystic Fibrosis: Design and Synthesis of Thiadiazole Inhibitors of the E3 Ligase RNF5.

In cystic fibrosis (CF), deletion of phenylalanine 508 (F508del) in the CF transmembrane conductance regulator (CFTR) is associated to misfolding and defective gating of the mutant channel. One of the most promising CF drug targets is the ubiquitin ligase RNF5, which promotes F508del-CFTR degradation. Recently, the first ever reported inhibitor of RNF5 was discovered, i.e., the 1,2,4-thiadiazol-5-ylidene inh-2. Here, we designed and synthesized a series of new analogues to explore the structure-activity relationships (SAR) of this class of compounds. SAR efforts ultimately led to compound 16, which showed a greater F508del-CFTR corrector activity than inh-2, good tolerability, and no toxic side effects. Analogue 16 increased the basal level of autophagy similar to what has been described with RNF5 silencing. Furthermore, co-treatment with 16 significantly improved the F508del-CFTR rescue induced by the triple combination elexacaftor/tezacaftor/ivacaftor in CFBE41o- cells. These findings validate the 1,2,4-thiadiazolylidene scaffold for the discovery of novel RNF5 inhibitors and provide evidence to pursue this unprecedented strategy for the treatment of CF.

WS02.04 Triple HEMT in Pregnancy and Lactation: effects on the developing lung, gut and pancreas

Introduction/aim: The development of new cystic fibrosis (CF) drugs, including the highly effective modulator therapies (HEMT) such as elexacaftor-tezacaftor-ivacaftor (ETI), has enabled more women with CF to reach child-bearing age. However, safety of ongoing use of ETI during pregnancy and breastfeeding, especially for the baby’s developing organs, is unknown. We aim to investigate placental and milk transfer of maternally administered ETI in pre- and postnatal wildtype and F508del-CFTR rats and their bioaccumulation into the developing lungs, gut and pancreas.

P190 Nanostructured formulation of a novel hybrid iminosugar/steroid agent for application in cystic fibrosis lung disease

Chronic bacterial infections and exaggerated inflammation of airways are hallmarks of Cystic Fibrosis (CF) and are significant causes of morbidity and mortality in CF patients. Accordingly, development of novel anti-inflammatory and anti-bacterial agents, to combine with CFTR modulators, is a key goal in CF drug discovery. Recent studies, granted by Italian Cystic Fibrosis Research Foundation, revealed the ability of two compounds, the iminosugar L-Miglustat and beta-sitosterol (BSS), to reduce both the levels of inflammation and the bacterial load in murine models of CF infection.

The Unbearable Cost of Drug Development

The Unbearable Cost of Drug Development

Simultaneous Quantification of Ivacaftor, Tezacaftor, and Elexacaftor in Cystic Fibrosis Patients’ Plasma by a Novel LC–MS/MS Method

The new breakthrough cystic fibrosis (CF) drug combination of ivacaftor (IVA), tezacaftor (TEZ), and elexacaftor (ELX), namely "caftor" drugs, directly modulates the activity and trafficking of the defective CF transmembrane conductance regulator protein (CFTR) underlying the CF disease. The role of therapeutic drug monitoring (TDM) of caftor drugs in clinical settings has recently been established. The availability of reliable and robust analytical methods for the quantification of IVA, TEZ, and ELX is essential to support dose-concentration-effect studies. We have developed and validated a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the rapid and simultaneous quantification of IVA, TEZ, and ELX from the plasma of CF patients. The method was based on a rapid extraction protocol from 50 μL human plasma and separation on a reversed-phase C-18 HPLC column after the addition of deuterated internal standards. Accurate analyte quantification using multiple reaction monitoring (MRM) detection was then obtained using a Thermofisher Quantiva triple-quadrupole MS coupled to an Ultimate 3000 UHPLC. The method has been validated following international (EMA) guidelines for bioanalytical method validation and has been tested on plasma samples from 62 CF patients treated with the three-drug combination IVA/TEZ/ELX, marketed as Kaftrio® or Trikafta®, in steady-state condition. The assay was linear over wide concentration ranges (0.008-12 mg/L) in plasma for IVA, TEZ, and ELX, suitable for a broad range of plasma concentrations, and accurate and reproducible in the absence of matrix effects. The stability of analytes for at least 30 days at room temperature could allow for cost-effective shipment and storage. On the same day of sample collection, a sweat test was evaluated for 26 associated patients' samples, FEV1 (%) for 58, and BMI was calculated for 62. However, Spearman correlation showed no correlation between Cthrough plasma concentrations of analytes (IVA, TEZ, ELX) and sweat test, FEV1 (%), or BMI. Our method proved to be suitable for TDM and could be helpful in assessing dose-concentration-response correlations in larger studies.

NICE does not recommend three cystic fibrosis drugs in draft guidance

NICE does not recommend three cystic fibrosis drugs in draft guidance

Organoid Technology and Its Role for Theratyping Applications in Cystic Fibrosis.

Cystic fibrosis (CF) is a autosomal recessive, multisystemic disease caused by different mutations in the CFTR gene encoding CF transmembrane conductance regulator. Although symptom management is important to avoid complications, the approval of CFTR modulator drugs in the clinic has demonstrated significant improvements by targeting the primary molecular defect of CF and thereby preventing problems related to CFTR deficiency or dysfunction. CFTR modulator therapies have positively changed the patients' quality of life, especially for those who start their use at the onset of the disease. Due to early diagnosis with the implementation of newborn screening programs and considerable progress in the treatment options, nowadays pediatric mortality was dramatically reduced. In any case, the main obstacle to treat CF is to predict the drug response of patients due to genetic complexity and heterogeneity. Advances in 3D culture systems have led to the extrapolation of disease modeling and individual drug response in vitro by producing mini organs called "organoids" easily obtained from nasal and rectal mucosa biopsies. In this review, we focus primarily on patient-derived intestinal organoids used as in vitro model for CF disease. Organoids combine high-validity of outcomes with a high throughput, thus enabling CF disease classification, drug development and treatment optimization in a personalized manner.

Dynamic Perfluorinated Gas MRI Shows Improved Lung Ventilation in People with Cystic Fibrosis after Elexacaftor/Tezacaftor/Ivacaftor: An Observational Study.

The availability of highly effective CFTR modulators is revolutionizing the treatment of cystic fibrosis (CF) and drastically improving outcomes. MRI-based imaging modalities are now emerging as highly sensitive endpoints, particularly in the setting of mild lung disease. Adult CF patients were recruited from a single center prior to starting treatment with E/T/I. The following studies were obtained before and after one month on treatment: spirometry, multiple breath nitrogen washout (MBW), 1H UTE MRI (structural images) and 19F MRI (ventilation images). Changes between visits were calculated, as were correlations between FEV1, lung clearance index (LCI), MRI structural scores, and MRI-based ventilation descriptors. Eight subjects had complete datasets for evaluation. Consistent with prior clinical trials, FEV1 and LCI improved after 28 days of E/T/I use. 1H UTE MRI detected improvements in bronchiectasis/airway wall thickening score and mucus plugging score after 28 days of therapy. 19F MRI demonstrated improvements in fractional lung volume with slow gas washout time (FLV↑tau2) and ventilation defect percentage (VDP). Improvements in FLV↑tau2 and VDP correlated with improvement in FEV1 (r = 0.81 and 0.86, respectively, p < 0.05). This observational study establishes the ability of 19F MRI and 1H UTE MRI to detect improvements in lung structure and function after E/T/I treatment. This study supports further development of 19F MRI and 1H UTE MRI as outcome measures for cystic fibrosis research and drug development.