Cystic Fibrosis Transmembrane Regulator Correctors Research Articles

Cystic fibrosis transmembrane regulator correction attenuates heart failure-induced lung inflammation.

Heart failure (HF) affects 64 million people worldwide. Despite advancements in prevention and therapy, quality of life remains poor for many HF patients due to associated target organ damage. Pulmonary manifestations of HF are well-established. However, difficulties in the treatment of HF patients with chronic lung phenotypes remain as the underlying patho-mechanistic links are still incompletely understood. Here, we aim to investigate the cystic fibrosis transmembrane regulator (CFTR) involvement in lung inflammation during HF, a concept that may provide new mechanism-based therapies for HF patients with pulmonary complications. In a mouse model of HF, pharmacological CFTR corrector therapy (Lumacaftor (Lum)) was applied systemically or lung-specifically for 2 weeks, and the lungs were analyzed using histology, flow cytometry, western blotting, and qPCR. Experimental HF associated with an apparent lung phenotype characterized by vascular inflammation and remodeling, pronounced tissue inflammation as evidenced by infiltration of pro-inflammatory monocytes, and a reduction of pulmonary CFTR+ cells. Moreover, the elevation of a classically-activated phenotype of non-alveolar macrophages coincided with a cell-specific reduction of CFTR expression. Pharmacological correction of CFTR with Lum mitigated the HF-induced downregulation of pulmonary CFTR expression and increased the proportion of CFTR+ cells in the lung. Lum treatment diminished the HF-associated elevation of classically-activated non-alveolar macrophages, while promoting an alternatively-activated macrophage phenotype within the lungs. Collectively, our data suggest that downregulation of CFTR in the HF lung extends to non-alveolar macrophages with consequences for tissue inflammation and vascular structure. Pharmacological CFTR correction possesses the capacity to alleviate HF-associated lung inflammation.

The NSAID glafenine rescues class 2 CFTR mutants via cyclooxygenase 2 inhibition of the arachidonic acid pathway

Most cases of cystic fibrosis (CF) are caused by class 2 mutations in the cystic fibrosis transmembrane regulator (CFTR). These proteins preserve some channel function but are retained in the endoplasmic reticulum (ER). Partial rescue of the most common CFTR class 2 mutant, F508del-CFTR, has been achieved through the development of pharmacological chaperones (Tezacaftor and Elexacaftor) that bind CFTR directly. However, it is not clear whether these drugs will rescue all class 2 CFTR mutants to a medically relevant level. We have previously shown that the nonsteroidal anti-inflammatory drug (NSAID) ibuprofen can correct F508del-CFTR trafficking. Here, we utilized RNAi and pharmacological inhibitors to determine the mechanism of action of the NSAID glafenine. Using cellular thermal stability assays (CETSAs), we show that it is a proteostasis modulator. Using medicinal chemistry, we identified a derivative with a fourfold increase in CFTR corrector potency. Furthermore, we show that these novel arachidonic acid pathway inhibitors can rescue difficult-to-correct class 2 mutants, such as G85E-CFTR > 13%, that of non-CF cells in well-differentiated HBE cells. Thus, the results suggest that targeting the arachidonic acid pathway may be a profitable way of developing correctors of certain previously hard-to-correct class 2 CFTR mutations.

Therapeutic CFTR Correction Normalizes Systemic and Lung-Specific S1P Level Alterations Associated with Heart Failure.

Heart failure (HF) is among the main causes of death worldwide. Alterations of sphingosine-1-phosphate (S1P) signaling have been linked to HF as well as to target organ damage that is often associated with HF. S1P’s availability is controlled by the cystic fibrosis transmembrane regulator (CFTR), which acts as a critical bottleneck for intracellular S1P degradation. HF induces CFTR downregulation in cells, tissues and organs, including the lung. Whether CFTR alterations during HF also affect systemic and tissue-specific S1P concentrations has not been investigated. Here, we set out to study the relationship between S1P and CFTR expression in the HF lung. Mice with HF, induced by myocardial infarction, were treated with the CFTR corrector compound C18 starting ten weeks post-myocardial infarction for two consecutive weeks. CFTR expression, S1P concentrations, and immune cell frequencies were determined in vehicle- and C18-treated HF mice and sham controls using Western blotting, flow cytometry, mass spectrometry, and qPCR. HF led to decreased pulmonary CFTR expression, which was accompanied by elevated S1P concentrations and a pro-inflammatory state in the lungs. Systemically, HF associated with higher S1P plasma levels compared to sham-operated controls and presented with higher S1P receptor 1-positive immune cells in the spleen. CFTR correction with C18 attenuated the HF-associated alterations in pulmonary CFTR expression and, hence, led to lower pulmonary S1P levels, which was accompanied by reduced lung inflammation. Collectively, these data suggest an important role for the CFTR-S1P axis in HF-mediated systemic and pulmonary inflammation.

CFTR corrector C17 is effective in muscular dystrophy, in vivo proof of concept in LGMDR3.

Limb-girdle muscular dystrophy R3 (LGMDR3) is caused by mutations in the SGCA gene coding for α-sarcoglycan (SG). Together with β- γ- and δ-SG, α-SG forms a tetramer embedded in the dystrophin associated protein complex crucial for protecting the sarcolemma from mechanical stresses elicited by muscle contraction. Most LGMDR3 cases are due to missense mutations, which result in non-properly folded, even though potentially functional α-SG. These mutants are prematurely discarded by the cell quality control. Lacking one subunit, the SG-complex is disrupted. The resulting loss of function leads to sarcolemma instability, muscle fiber damage and progressive limb muscle weakness. LGMDR3 is severely disabling and, unfortunately, still incurable. Here, we propose the use of small molecules, belonging to the class of cystic fibrosis transmembrane regulator (CFTR) correctors, for recovering mutants of α-SG defective in folding and trafficking. Specifically, CFTR corrector C17 successfully rerouted the SG-complex containing the human R98H-α-SG to the sarcolemma of hind-limb muscles of a novel LGMDR3 murine model. Notably, the muscle force of the treated model animals was fully recovered. To our knowledge, this is the first time that a compound designated for cystic fibrosis is successfully tested in a muscular dystrophy and may represent a novel paradigm of treatment for LGMDR3 as well as different other indications in which a potentially functional protein is prematurely discarded as folding-defective. Furthermore, the use of small molecules for recovering the endogenous mutated SG has an evident advantage over complex procedures such as gene or cell transfer.

Targeting cerebrovascular sphingosine‐1‐phosphate signaling to treat heart failure‐induced brain dysfunction

Chronic cardiovascular diseases increasingly emerge as risk factors for neurodegeneration. Heart failure for instance, induces a chronic reduction in cerebral perfusion. Such chronic brain perfusion deficits are frequently accompanied by cognitive decline, which not only seriously impacts the patients’ quality of life but also reduces treatment compliance. Statistically, cognitive impairment is 5 times more prevalent in heart failure patients relative to the age‐matched general population. We previously demonstrated that heart failure associates to augmented signaling of a potent bioactive phospholipid called sphingosine‐1‐phosphate (S1P). During heart failure, such enhanced S1P signaling roots from dysfunctional cystic fibrosis transmembrane regulator (CFTR) expression and activity. CFTR controls the bioavailability of cerebrovascular S1P and hence, its pro‐constrictive effects. In heart failure, reduced microvascular CFTR expression correlates with enhanced myogenic tone and reduced cerebral blood flow.In this study, we investigated the therapeutic potential of CFTR corrector compounds to normalize cerebrovascular S1P signaling and thereby, improve neurological function during experimental heart failure in mice.Systemic CFTR corrector treatment stabilized cerebrovascular CFTR expression, improved myogenic tone and rectified deficient cerebral perfusion in heart failure, without altering the systemic parameters of cardiac output or total peripheral resistance. This normalization associated to recovery of neuronal integrity (i.e., dendrite morphology and dendritic spine density) and cognitive function. The heart failure‐associated dendrite atrophy (i.e., a 22% reduction in dendrite length) was not altered in mice treated with CFTR corrector compound. Longitudinal pre‐treatment (i.e., at 10 weeks post‐infarction) and post‐treatment analyses of the same vehicle‐ and CFTR corrector‐treated HF mice confirmed that CFTR corrector treatment reversed the impaired rhino‐cortical‐dependent, short‐term retention of object familiarity caused by heart failure.Our results put forward a new mechanistic understanding of heart failure‐associated brain complications, and CFTR correctors as potential treatment option for heart failure‐associated cognitive decline.Support or Funding InformationThis work is supported by the Knut & Alice Wallenberg Foundation and the Swedish Research Council.

Combined Use of CFTR Correctors in LGMD2D Myotubes Improves Sarcoglycan Complex Recovery.

Sarcoglycanopathies are rare limb girdle muscular dystrophies, still incurable, even though symptomatic treatments may slow down the disease progression. Most of the disease-causing defects are missense mutations leading to a folding defective protein, promptly removed by the cell’s quality control, even if possibly functional. Recently, we repurposed small molecules screened for cystic fibrosis as potential therapeutics in sarcoglycanopathy. Indeed, cystic fibrosis transmembrane regulator (CFTR) correctors successfully recovered the defective sarcoglycan-complex in vitro. Our aim was to test the combined administration of some CFTR correctors with C17, the most effective on sarcoglycans identified so far, and evaluate the stability of the rescued sarcoglycan-complex. We treated differentiated myogenic cells from both sarcoglycanopathy and healthy donors, evaluating the global rescue and the sarcolemma localization of the mutated protein, by biotinylation assays and western blot analyses. We observed the additive/synergistic action of some compounds, gathering the first ideas on possible mechanism/s of action. Our data also suggest that a defective α-sarcoglycan is competent for assembly into the complex that, if helped in cell traffic, can successfully reach the sarcolemma. In conclusion, our results strengthen the idea that CFTR correctors, acting probably as proteostasis modulators, have the potential to progress as therapeutics for sarcoglycanopathies caused by missense mutations.

Respiratory System Disease.

Respiratory system involvement in cystic fibrosis is the leading cause of morbidity and mortality. Defects in the cystic fibrosis transmembrane regulator (CFTR) gene throughout the sinopulmonary tract result in recurrent infections with a variety of organisms including Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, and nontuberculous mycobacteria. Lung disease occurs earlier in life than once thought and ideal methods of monitoring lung function, decline, or improvement with therapy are debated. Treatment of sinopulmonary disease may include physiotherapy, mucus-modifying and antiinflammatory agents, antimicrobials, and surgery. In the new era of personalized medicine, CFTR correctors and potentiators may change the course of disease.

Targeting protein–protein interactions to rescue Δf508‐cftr: a novel corrector approach to treat cystic fibrosis

Cystic fibrosis (CF) is one of the most prevalent life‐threatening autosomal recessive disorders in the Western World, with an estimated incidence of 1 in 2000–3000 Caucasian newborns and a median age of survival of 40 years. The first report of CF as a distinct pathology was published in 1938 by Dorothy Andersen, a physician at the Babies & Children's Hospital of Columbia University, who differentiated its diagnosis from celiac disease in the pancreas (Andersen, 1938). This seminal description was followed by other contributions that linked CF to altered Cl− permeability in epithelial cells in the gastrointestinal and hepato‐billiary systems, respiratory tract, reproductive system and sweat glands (Kreindler, 2010); and culminated with the discovery of the gene responsible of the disease, referred to as the cystic fibrosis transmembrane regulator (CFTR) (Riordan et al, 1989). The CFTR is a cAMP‐regulated chloride and bicarbonate channel that controls anion conductance in epithelial cells located in mucosal surfaces. Functional alteration of CFTR leads to the accumulation of mucous secretions that cannot be cleared favouring inflammation and infection. Cumulative evidence shows a strong correlation between the severity of the CF phenotype and the degree of CFTR dysfunction, being stronger for those patients where the channel function is absent (Kreindler, 2010). The CF phenotype is caused by more than 1000 mutations of the CFTR gene including missense, such as R117H or G551D that significantly reduce channel activity, and nonsense like G542X, R553X or W1282X, which abrogate protein expression (Kreindler, 2010). However, the most prevalent mutation in CF patients (70%) is the in‐frame deletion of Phenylalanine at position 508 (ΔF508‐CFTR) (Kreindler, 2010; Rowe & Verkman, 2013). This deletion produces a partially unfolded CFTR protein that is retained in the endoplasmic reticulum and diverted to degradation by the proteosome, resulting in a virtually absent CFTR activity …

Cystic Fibrosis Transmembrane Regulator Correctors and Potentiators

Cystic fibrosis (CF) is caused by loss-of-function mutations in the CF transmembrane conductance regulator (CFTR) protein, a cAMP-regulated anion channel expressed primarily at the apical plasma membrane of secretory epithelia. Nearly 2000 mutations in the CFTR gene have been identified that cause disease by impairing its translation, cellular processing, and/or chloride channel gating. The fundamental premise of CFTR corrector and potentiator therapy for CF is that addressing the underlying defects in the cellular processing and chloride channel function of CF-causing mutant CFTR alleles will result in clinical benefit by addressing the basic defect underlying CF. Correctors are principally targeted at F508del cellular misprocessing, whereas potentiators are intended to restore cAMP-dependent chloride channel activity to mutant CFTRs at the cell surface. This article reviews the discovery of CFTR potentiators and correctors, what is known regarding their mechanistic basis, and encouraging results achieved in clinical testing.