CFTR as an ABC Gene

CFTR is categorized as a member of the ABC or traffic ATPase family of genes. These ancient genes code for hundreds of polypeptides from both prokaryotes and eukaryotes, which transport nutrients, metabolites, toxins, and other small molecules across cellular membranes. All ABC proteins are characterized by two NBDs encoding canonical Walker A and B motifs (capable of binding and hydrolyzing ATP). ABC proteins also contain two membrane-spanning domains (MSDs) with multiple alpha helices that form highly selective passageways across lipid bilayers. Among ABC proteins, CFTR is atypical by virtue of a regulatory domain (R domain) with numerous sites for protein kinase A/cAMP-dependent protein kinase (cAMP/PKA) phosphorylation, as well as a regulatory insertion (RI) in nuclear binding domain 1 (NBD1) believed to participate in channel gating (see Fig. 47-1 ).

Structure/Activity and CFTR Gating Mechanism

The CFTR gene product is composed of approximately 1500 amino acids and functions as a transporter of Cl ⁻ and HCO ₃ ⁻ in numerous epithelial tissues. Although a high-resolution, three-dimensional structure of full-length CFTR is not yet available, site-directed mutagenesis indicates that gating is enabled by cAMP/PKA dependent phosphorylation of the R domain. The phosphorylation step has been shown to elicit a conformational change in the regulatory insertion, leading to NBD1/NBD2 heterodimerization and structural realignment in MSD1 and MSD2 that open the ion conductive pore. Key interactions between the intracellular loops of the transmembrane domains and the nucleotide binding domains are integral to protein stability and interdomain assembly, as well as to transmission of the forces required for channel opening (see Fig. 47-1 ). NBD dimerization is greatly augmented when two ATP binding sites (at the NBD1/2 interface) are occupied; closing of CFTR is attributable to ATP hydrolysis. It should be noted that ATP binding is not absolutely required for channel gating (so-called ATP-independent channels can also open, presumably on the basis of the tendency of NBDs to bind each other spontaneously). Transepithelial transport through CFTR is governed by the electrochemical driving force, and a CFTR bioelectric signature (by membrane patch clamp analysis or planar lipid bilayer reconstitution) describes an approximately 8 picosie­men channel with linear current/voltage relationship and characteristic opening bursts. CFTR therefore employs ATP binding to enable passive ion transport, rather than subserving the function typical of the ABC gene family (i.e., pumping of larger metabolites with a stoichiometric dependence on ATP hydrolysis).

Biogenesis and Processing of Native CFTR

Wild-type CFTR matures by a conventional pathway that includes cotranslational insertion in the endoplasmic reticulum (ER) membrane and post-translational modifications such as N-linked glycosylation and ubiquitination ( Fig. 47-3 ). Subdomain folding (e.g., within NBDs) and achievement of final tertiary structure requires complex domain binding interactions and represents a topic of intense interest due to the importance of misfolding as a mediator of clinical disease. As even wild-type CFTR processing is not completely efficient, a subset of CFTR molecules is targeted for proteosomal hydrolysis by ER-associated degradation (ERAD). For CFTR that advances from ER to Golgi, complex glycosylation takes place at two asparagine residues. CFTR reaching the Golgi is transported to the cell surface by vesicular traffic with subsequent recycling through sorting and recycling endosomes and reinsertion in the plasma membrane or routing to the lysosome. The cell surface apparatus that governs CFTR peripheral stability (i.e., plasma membrane) has been well characterized and includes ubiquitin conjugation to regulate internalization in response to environmental stress.

Scheme of CFTR biogenesis.

Biogenesis of CFTR in normal epithelial cell (right side of cell) and F508del cystic fibrosis (CF)-affected epithelial cell (left side). In the normal epithelial cell, CFTR is synthesized in the rough endoplasmic reticulum, glycosylated and folded in the Golgi apparatus, and chaperoned to the cell surface. In the F508del-affected epithelial cells, the CFTR polypeptide is misfolded and tagged for premature degradation via endoplasmic reticulum-associated degradation ( red arrow ) before reaching the cell surface (class II mutation).

(Adapted from Ameen N, Silvis M, Bradbury NA: Endocytic trafficking of CFTR in health and disease. J Cystic Fibrosis 6:1–14, Fig. 1, 2007.)

Cellular Defects Attributable to F508del CFTR

The vast majority of F508del CFTR is retained in the ER, heavily ubiquitinated, and routed to the proteasome by ERAD. As a consequence, F508del CFTR visualized by SDS PAGE is predominantly an ER localized, approximately 150KDa core glycosylated (“band B”) glycoform, whereas a substantial portion of post ER, wild-type protein migrates more slowly due to complex glycosylation (≈180 KDa “band C”). The F508del trafficking defect is temperature sensitive, and growth of epithelium at lower temperatures (e.g., 27° C) leads to measurable levels of band C with partial restoration of cell surface function.

The maturational processing defect mediated by F508del involves at least two distinct abnormalities. Omission of F508 (which is located on an externally exposed peptide loop of the first nucleotide binding domain) leads to pronounced loss of NBD1 stability as judged by thermal and isothermal calorimetry, protein aggregation, and protein yield measurements following recombinant overexpression. In addition, F508 facilitates binding of a cytosolic loop from MSD2, and disruption of the NBD1/MSD2 interface (independent of NBD1 stability) leads to further impairment of F508del maturation.

F508del CFTR molecules that have been rescued to the cell surface by low temperature display intrinsic channel gating abnormalities, diminished plasma membrane stability, and pronounced thermal unfolding with loss of function following incubation at 37° C. In addition, F508del CFTR mRNA may be misfolded and poorly utilized (leading to lower levels of CFTR translation) due to omission of the F508 codon. The observation of multiple distinct CFTR defects attributable to F508del points to the challenge of identifying a single agent capable of restoring mutant CFTR to therapeutically relevant levels. Intramolecular suppressor mutations that specifically ameliorate F508del NBD1 instability or the defective NBD1/MSD1 interface have been identified. CFTR constructs encoding these suppressors provide a means to characterize F508del corrective strategies and will enable compound library screening tailored to specific CFTR folding defects in the future.

Overview of Lung Pathophysiology

An improved understanding of CF pathogenesis has led to better diagnostic strategies, an improved understanding of the onset and progression of lung disease, and therapeutic approaches to target the underlying disease. An overview is provided in Figure 47-4 and described in further detail later. As with other monogenetic diseases, absent or dysfunctional CFTR results in diminished functional protein, which is necessary to transport chloride and bicarbonate across the airway epithelium. Several key manifestations include delayed mucociliary clearance because of depletion of airway surface liquid (ASL), abnormalities of the physical properties of CF mucus, and a predisposition to early infection because of abnormal mucosal defense; there may also be dysregulated inflammation. These processes initiate and perpetuate a cycle of destruction that ultimately results in irreversible lung injury, bronchiectasis, and respiratory failure.

CF pathophysiology.

CF lung disease results from consequences of genetic mutations in CFTR . Major operative pathways include reduced mucociliary clearance due to depleted airway surface liquid (ASL) hydration, abnormal mucus adhesion and viscosity, and defective bacterial killing that each contribute to the cycle of destruction.

In the lungs, CF manifests as infected mucus secretions that compromise the airway lumen and contribute to obstructive pulmonary disease and reduced FEV ₁ ( Fig. 47-5 ). Airway disease is thought to begin in the small airways, resulting in airflow obstruction detected by spirometry midflows (i.e., FEF _25%-75% ). Submucosal gland hyperplasia and inspissated mucus secretions emanating from the glands are also prominent. Radiographic changes consistent with small airway obstruction (e.g., tree-in-bud opacities) followed by bronchiectasis become apparent over time. Development of bronchiectasis leads to irreversible changes that encourage continued infection and accelerate disease pathogenesis.

CF pathologic and radiologic correlation.

A, Gross pathologic specimen of explanted CF lung from a patient with end-stage CF lung disease. This specimen demonstrates heavy lungs characterized by bronchiectasis and numerous mucopurulent plugs. B, Large airway pathology of CF. A large mucus plug filled with inflammatory infiltrate is clearly seen, in addition to remodeling of the medium-sized airway (hematoxylin-eosin stain). C, Representative photomicrographs from a CF patient with end-stage lung disease is depicted. Note the thickening of the lamina propria (higher Reid index), the prominent arborized mucus plug extruding from the submucosal gland onto the airway lumen, and the hypertrophic surface epithelium (periodic acid–Schiff stain). D, Sagittal high-resolution CT image from a 16-year-old male with CF demonstrating mild to moderate multilobar bronchiectasis ( cyan arrows ) and tree-in-bud opacities representing small airways mucus plugs ( red arrow ).

( A–C, Images courtesy Dr. David Kelly, University of Alabama, Birmingham.)

Cellular Functions of the CFTR Gene Product

The CFTR gene is hundreds of millions of years old and utilized by diverse species including fish, amphibian, fowl, and mammalian. Although best described as a chloride and bicarbonate transporter, the protein appears to regulate numerous processes in addition to anion secretion. CFTR is situated within membrane complexes by virtue of a PDZ-type binding motif and configured in close proximity to a number of integral membrane proteins including other ion channels. In experimental systems, CFTR exerts a regulatory influence on the epithelial sodium channel (ENaC). Proteomic and transcriptome analyses demonstrate hundreds of cellular gene products that directly bind or are regulated by the gene product.

Regulation of Airway Surface Liquid Homeostasis

In a conventional model of CF respiratory pathogenesis, absence of apical CFTR leads to failure of chloride and bicarbonate secretion. Because release of water into the periciliary region is driven in large measure by a CFTR-dependent osmotic gradient, CFTR deficiency leads to failure of ASL hydration ( Fig. 47-6 ). In addition, a considerable body of evidence suggests that ENaC present on the airway surface may be down-regulated by wild-type CFTR, and that loss of the CF gene confers elevated ENaC-dependent sodium and water uptake, further exacerbating ASL desiccation. According to this prevailing model, depleted ASL leads to inadequate ciliary extension and impairment of mucociliary clearance.

CF airway surface and mucus transport defect.

Schematic of the mucociliary transport defect in CF. In the healthy state (A), adequate airway surface homeostasis ensures effective transport of mucus extruding from airway surface goblet cells and the submucosal glands. The airway surface liquid (ASL) is maintained by fluid secretion via the CFTR and fluid absorption via the epithelial sodium channel, ENaC (inset box on right) (CFTR, surface receptor in blue; ENaC, surface receptor in red). In CF (B), the ASL is depleted through the absence of CFTR-mediated fluid secretion accompanied by tonic fluid absorption via the ENaC. CFTR-dependent liquid dessication decreases the depth of the ASL including the periciliary layer causing abnormal clearance of mucus from the epithelial cell surface in the airways.

Recent findings indicate significant new levels of complexity with regard to the mechanisms that govern ASL depth and mucociliary clearance. For example, although the periciliary fluid layer has traditionally been viewed as primarily aqueous in composition, recent data indicate that a fluid layer containing tethered mucins is characterized by intrinsic viscosity that is sensitive to the osmolar forces of the overlying mucus, which are expected to strongly influence mucociliary clearance when the periciliary layer collapses due to mucus dehydration. Moreover, ENaC hyperactivity across the CF airway surface—thought to be a hallmark of CF pathophysiology in the past—has recently been questioned on the basis of (1) absence of sodium hyperabsorption from airway mucosa in newborn CF pigs (together with evidence of normal ASL depth) and (2) studies of primary human airway epithelium indicating that elevated sodium reabsorption in CF can be explained by an increased gradient for sodium uptake (rather than a direct biochemical influence on ENaC, per se).

Findings such as these pose a challenge to interpretation of experimental therapies intended to augment ASL depth by blocking respiratory ENaC or activating non-CFTR chloride secretory pathways. Such interventions represent an important approach for improving mucociliary clearance, and such strategies could improve mucus clearance regardless of their relationship to the fundamental CF defect. The significance of ASL expansion is underscored by therapeutic improvement among patients treated with hypertonic saline aerosols and dry powder mannitol to improve mucociliary clearance in CF lung disease by providing airway hydration.

Regulation of Sweat Chloride by CFTR

The concentration of chloride in human sweat is determined by the balance of secretion and reabsorption of sodium and chloride in the sweat gland and duct. Chloride is secreted by two parallel pathways. One involves CFTR; the other utilizes a CFTR-independent, calcium-activated chloride channel. Because CFTR is not the only pathway by which chloride exits the apical membrane, chloride is secreted into sweat even in the absence of functional CFTR. Normally, sodium is reabsorbed in the duct of the sweat gland because of ENaC in the apical membrane of the duct cells. Chloride follows sodium into the cell, through the CFTR chloride channel. In CF, with absent or defective CFTR, chloride reabsorption is significantly reduced and the chloride content of sweat is abnormally high.

Cystic Fibrosis Exocrine Glandular Epithelium

Submucosal glandular ducts filled with inspissated mucus are observed early in the course of CF pulmonary involvement. CFTR is heavily expressed within epithelial cells of submucosal glands, where it functions to activate fluid and electrolyte secretion and promote release of mucus onto the airway surface. The relative contributions of submucosal glands to overall CF respiratory pathophysiology are not known. Although absence of a lung phenotype in CF murine models has been attributed to non-CFTR Cl ⁻ secretory pathways that are highly active in mouse lungs, the paucity of murine airway glands has also been implicated (see Chapter 1 for differences in mouse and human lungs). The relative lack of murine airway glands is a likely explanation for the lack of a murine lung phenotype, particularly because other CF null animals with extensive submucosal gland formation (i.e., pig, ferret) exhibit a pulmonary phenotype similar to humans. The recent development of confocal imaging modalities that allow direct visualization of glandular activity in full-thickness airways and the ability to analyze biophysical properties of mucus extruded from CF pig, ferret, or rat submucosal glands will contribute new and important knowledge in this area.

Mucus Biogenesis, Adhesion, and Transport

Hyperviscous respiratory secretions obstruct small- and medium-sized airways in CF, leading to profound failure of mucociliary clearance that can be verified macroscopically by radioligand imaging. A primary biochemical defect in mucus composition has been extensively sought but not well established as a fundamental cause of the disease. Studies of CF pulmonary secretion are complicated by difficulty retrieving standardized mucus samples, the diversity of respiratory pathogens, pronounced lung inflammation, and other variables. Specific mucins expressed in CF respiratory secretions include the gel-forming mucins MUC5B and MUC5AC, in addition to a complex array of extracellular proteins. DNA strands released from dying bacteria, epithelial cells, and inflammatory cells represent an important contributor to excess mucus viscosity and provide rationale for use of recombinant DNAse as an aerosolized mucolytic.

In extrapulmonary CF organs characterized by hyperviscous secretion (e.g., pancreas, liver), profound ductular obstruction is observed in the absence of polymicrobial infection, allowing more direct studies of the relationship between CFTR and mucus formation. An emerging notion implicates defective bicarbonate transport as a mediator of hyperviscosity and mucosal adhesion in CF secretions. In this model, exocrine mucus (highly negative in charge) is produced by acinar and other epithelial cells in compacted form due to bound calcium, which shields the negative repulsive force between sulfates and other anionic groups on constituent mucins. Bicarbonate secretion via CFTR chelates the calcium and permits mucinous expansion and a viscoelastic state compatible with physiologic clearance. Failure of bicarbonate release is hypothesized to result in a defective mucin expansion, leading to hyperviscous secretion with abnormally adherent properties (i.e., mucus that is tightly bound to the respiratory surface and difficult to mobilize). Evidence of excessive mucus adhesion that can be reversed by bicarbonate has been observed in the intestine of CF mice.

Host Defense and Infection

CF lungs are characterized by intense neutrophilic inflammation with polymorphonuclear cells densely infiltrating airway secretions. Whether robust lung inflammation is mediated directly by CFTR (as opposed to a consequence of polymicrobial infection) is a topic of considerable debate. Neutrophil chemoattractants such as interleukin-8, tumor necrosis factor (TNF)-α, and a collagen fragment, proline-glycine-proline, are present at high levels in airway secretions, and abnormalities of CFTR-dependent macrophage function have also been implicated. Although anti-inflammatory therapies can improve lung function and slow the rate of CF respiratory decline, immune blockade also can predispose individuals to worsening lung infection, and the proper clinical balance has been difficult to achieve.

Bacteria typically colonize the CF respiratory tract during infancy or early childhood. Longitudinal analysis of P. aeruginosa indicates sentinel infection due to a single genotype of the organism that often persists throughout the life of an individual patient. The explanation for early colonization is not well understood. Previous studies have suggested increased binding of CF pathogens to CFTR −/− airway epithelial cells or that antimicrobial peptides present in pulmonary secretions are inactivated by high salt concentrations bathing the respiratory surface. A recent study implicated acidic pH within CF ASL (attributable to absent CFTR bicarbonate release) as a likely cause of the increased susceptibility to bacteria. Findings such as these are most relevant to initial colonization events because airway pH may not be affected later in life.

Virulent respiratory pathogens such as P. aeruginosa evolve in a stereotypic fashion during the lifespan of individuals with CF and are characterized by rapid phenotype change and hypermutability. After several years of pulmonary infection, P. aeruginosa typically develops a mucoid phenotype in which considerable metabolic reserves are expended to synthesize and release the polyanionic protein, alginate. Appearance of mucoid P. aeruginosa in CF is a negative prognostic indicator. Selective advantage of alginate release has been attributed to the immunomodulating role of this exoproduct. Recent studies have demonstrated the complexity of the CF microbiome, which can be altered during exacerbation by the presence of a dominant pathogen.

Porcine and ferret models of CF pulmonary disease, as well as identification of very young CF patients by newborn screening, provide an important means to clarify relationships that exist among initial bacterial colonization, chronic infection, and inflammation in CF lungs. Longitudinal monitoring of CF respiratory infection in humans and transgenic animals will also facilitate a more sophisticated understanding of the complex CF microbiome and could provide new therapeutic opportunities in the future.

CFTR and Pulmonary Remodeling

Measurements in the CF porcine model indicate a change in tracheal diameter and density of submucosal glands before the advent of hyperviscous mucus obstruction. Similar findings have been reported in human CF lungs and in Cftr −/− rats (despite the absence of mucus plugging in the rodent model). In tissues such as porcine and human pancreas, fibrotic damage and fatty replacement can be profound, with extensive parenchymal scarring. Myofibroblast proliferation appears to mediate this effect, and TGF-β signal transduction (which engenders myofibroblast transformation) is markedly increased throughout the CF lung. TGF-β is also a well-established genetic modifier of F508del homozygous CF lung disease, and these findings suggest that changes in TGF-β–dependent profibrotic pathways govern lung tissue remodeling (in addition to inflammatory responsiveness elicited by the cytokine).

Overview

The diagnosis of CF is predicated on clinical suspicion for the disease, as indicated by clinical manifestations or family history, or alternatively through newborn screening programs. Early detection through newborn screening is leading to earlier recognition of illness before the clinical characteristics become apparent. The first organized CF newborn screening programs were developed in the 1980s and implemented in Colorado in 1987 and in the 1980s in Australia and Europe after retrospective trials demonstrated improved mortality and clinical outcomes. The number of programs in the United States and abroad increased in the past decade, and now all U.S. states have implemented newborn screening programs after a CDC/Cystic Fibrosis Foundation Consensus Statement in 2004 recommended this practice. The report invoked emerging prospective data that screening and early interventions in CF newborns diagnosed by newborn screening improved nutritional, gastrointestinal, respiratory, and cognitive functioning.

Several different screening algorithms are used and vary by state program, but serum detection of immunoreactive trypsinogen (IRT) is typically the primary screening method. IRT is quantified from blood spots taken in the perinatal period. IRT is a highly sensitive serum marker of obstructive pancreatic injury, although it is not specific for CF and thus additional testing is required to confirm the diagnosis. Elevated IRT in the perinatal period is typically followed by repeat IRT testing and/or a DNA panel for common CFTR mutations. If positive, refer to a diagnostic clinic for assessment of CFTR through mutation analysis and functional assays, such as sweat chloride, to confirm the diagnosis; this is frequently accomplished before the onset of clinical manifestations.

Diagnostic Algorithms

The diagnosis of CF is based on a positive newborn screening test or suspicious clinical characteristics in patients in infancy through adulthood ( Table 47-2 ). In 2008, the Cystic Fibrosis Foundation published comprehensive diagnostic guidelines for infants and adults with suspected CF ( Fig. 47-7A and B ); a similar approach was adopted in the European CF Society’s 2006 guidelines. The guidelines primarily differed only in the timing of CFTR mutation analysis and sweat chloride normative values. In addition, as compared with the European diagnostic guideline, the U.S. guideline deemphasizes measurement of nasal potential difference (NPD), a bioelectric measure of CFTR activity, mainly due to lack of standardization, although NPD is recommended as an alternative diagnostic modality. There is good concordance when comparing these methodologies for diagnostic accuracy.

Table 47-2

Diagnostic Criteria for CF by Clinical Phenotypic and CFTR Functional Abnormalities

The diagnosis of CF is confirmed by the presence of: Appropriate phenotypic clinical features (any of): Chronic sinopulmonary disease Chronic cough and sputum production Persistent infection with characteristic pathogens ( Staphylococcus aureus, Pseudomonas aeruginosa, other gram-negative organisms) Airflow obstruction Chronic chest radiographic abnormalities Sinus disease; nasal polyposis Gastrointestinal and nutritional abnormalities Exocrine pancreatic insufficiency Recurrent pancreatitis Fat-soluble vitamin deficiency Meconium ileus; DIOS Obstructive azoospermia in males Plus, Laboratory evidence of CFTR dysfunction (one or more of following): Elevated sweat chloride Disease-causing mutation in CFTR gene in both alleles Characteristic bioelectric abnormalities (potential difference) in nasal epithelium Abnormal ex vivo intestinal short-circuit current measurement

 

CF, cystic fibrosis; CFTR, cystic fibrosis transmembrane conductance regulator; DIOS, distal intestinal obstruction syndrome.

Diagnostic algorithm for CF.

A, Schematic depiction of the diagnostic algorithm for the diagnosis of CF in infants emphasizing newborn screening programs. B, Schematic depiction of a diagnostic algorithm for children and adults emphasizing sweat chloride as the cornerstone of the diagnosis. Note the differences in the lower limit of normal of sweat chloride for infants versus children and adults. Genetic testing for intermediate probability cases should trigger CFTR sequencing since analysis for only limited number of mutations can result in false negative evaluations in this disease category. CRMS, CFTR-related metabolic syndrome; IRT, immunoreactive trypsinogen; IRT/DNA, means confirmation with a DNA assay (see text); IRT/IRT, confirmation with a second IRT test; PCP, primary care physician.

( B, Adapted from Farrell PM, Rosenstein BJ, White TB, et al: Guidelines for diagnosis of cystic fibrosis in newborns through older adults: Cystic Fibrosis Foundation consensus report. J Pediatr 153(2):S4–S14, 2008.)

In the North American (CFF) algorithm (see Fig. 47-7A ), newborns are screened for CF using IRT, a byproduct of prenatal pancreatic inflammation. In most states this test is paired with a DNA probe panel for common CFTR mutations. IRT is not a highly specific measurement due to elevated levels observed in prematurity, traumatic delivery, and other neonatal gastrointestinal disorders. Because of declining IRT levels in the first few months after birth in CF and non-CF infants, this test is only useful for screening in the first few weeks after birth. Thereby, family history or clinical suspicion based on symptoms is the main impetus for diagnostic evaluation after the neonatal period has passed (see Fig. 47-7B ).

If the IRT is elevated or in situations where CF is suspected, the diagnosis should be confirmed by sweat chloride testing. Sweat chloride testing is available in many clinical laboratories and has been standardized to ensure accuracy. Because of its high sensitivity, sweat chloride testing has a central role in establishing the diagnosis in both the Cystic Fibrosis Foundation and European algorithms in the presence of a positive newborn screening or compatible clinical characteristics.

For infants screened at birth, a low sweat chloride value (<29 mmol/L) effectively rules out CF, whereas a high sweat chloride concentration (≥60 mmol/L) definitively establishes the diagnosis. In the case of clinical suspicion with an intermediate sweat chloride concentration (30 to 59 mmol/L), guidelines recommend performing CFTR sequencing or high-sensitivity DNA probe testing to establish the presence of CFTR mutations. If two CFTR mutations are found, the diagnosis of CF is established. If one or zero mutations are present, then repeat sweat chloride testing is recommended. It should be noted that standard genetic panels (i.e., panels that test fewer than 40 mutations) are not sufficient in patients with elevated sweat chloride and atypical manifestations because rare, partially functional mutations are common in these cases and may not be detected in limited panels. Some patients exhibit CF-like disease even in the absence of CFTR mutations, a finding probably caused by mutations in genes coding for other related proteins that can mimic mutations in CFTR (i.e., mutations in the ENaC channel).

If clinical suspicion remains elevated and sweat chloride is in the intermediate range, there are several alternatives to confirm CFTR functional deficits. NPD testing, available at select CF specialty centers, is viewed as an acceptable alternative in cases of inconclusive sweat chlorides values or inadequate genetic analysis. Testing of intestinal current measurements, another alternative. is only available at a few centers in the world and requires a biopsy of the rectal mucosa. Additionally, other measures of pancreatic function such as fecal elastase may help to support the diagnosis in this setting.

In analogous fashion, children and adults with clinical syndromes suspicious for CF are also first screened for CFTR dysfunction using sweat chloride testing (see Fig. 47-7B ). If the sweat chloride is low (≤39 mmol/L) CF is ruled out and other causes of the clinical syndrome should be sought. The lower limit of normal for sweat chloride values is higher in adults due to a relative, normal increase in sweat chloride with aging. In the presence of an elevated sweat chloride (>60 mmol/L), the diagnosis of CF is established in the patient and referral to a CF specialty center is recommended. In the case of an intermediate sweat chloride value (40 to 59 mmol/L), further CFTR genetic testing (i.e., sequencing) or alternative diagnostic testing as described earlier are appropriate. Additionally, in adult males, the diagnosis may be strongly suggested by the presence of a positive urologic evaluation for obstructive azoospermia.

It is increasingly recognized that CFTR-related disorders represent a spectrum of disease, with pancreatic-insufficient CF as the most severe form ( Fig. 47-8 ). It is notable that functional assessments of CFTR reflect this diversity, although the relationship between genotype and CFTR functional decrements can vary. CFTR biomarkers are generally able to distinguish between mild and severe phenotypes by demonstrating a continuum of CFTR decrement that correlates to clinical phenotype and is not apparent by genotype alone.

Spectrum of CF disorders.

This figure compares the findings in severe CF with the milder forms of the disease. Although manifestations are variable, severity in each organ system is generally consistent with degree of CFTR dysfunction conferred by genotype. CBAVD, congenital bilateral absence of the vas deferens; CUAVD, congenital unilateral absence of the vas deferens; DIOS, distal intestinal obstruction syndrome. *Refers to CFTR-related metabolic syndrome or CFTR-related disorders which are being defined currently.

Lower Respiratory Tract Disease

Progressive obstructive lung disease leading to bronchiectasis and respiratory failure causes the vast majority of mortality in individuals with CF. Even seemingly healthy infants with CF have significant subclinical lung disease; inflammation is often disproportionate to the degree of infection. The most common manifestation of lung disease is cough in association with bronchitis. Early in life, symptoms are often intermittent and exacerbated by episodes of acute respiratory tract infection that tend to exhibit a protracted course. With time, cough becomes a daily event. It is often worse at night and on arising in the morning. With progression or during exacerbations of lung disease, cough can become productive of tenacious, mucopurulent sputum secondary to chronic bacterial infection and resultant neutrophilic inflammation.

Hyperinflation of the lungs is common and often observed early in the progression of lung disease. Asthmatic or bronchiolitic-type wheezing is common during the first 2 years of life but may be encountered at any age. Wheezing is noted in up to one third of infants and may be present with or without evidence of atopy. Early in life, CF may often mimic the clinical manifestations of asthma and/or coexist with asthma syndrome, leading to delayed recognition.

Lung sounds are often unremarkable early in the disease process; the first detectable abnormalities may be a diminished intensity of breath sounds or subtle prolongation of the expiratory phase. Once CF lung disease becomes clinically apparent, adventitious lung sounds are usually first noted in the upper lobes. CF patients may have only mild bronchitic symptoms for long periods of time as lung homeostasis is maintained but typically manifest exacerbations of symptoms of increased cough intensity and sputum production. These exacerbations often present as tachypnea, shortness of breath, malaise, anorexia, and weight loss. Viral respiratory tract infection is a frequent trigger, as are other infectious agents, cigarette smoke, pollutants, allergens, and respiratory irritants that have been implicated in disturbing lower airway homeostasis. Initiation of broad-spectrum antibiotic therapy and aggressive maneuvers to facilitate clearance of mucus are usually required to improve symptoms and restore lung function. These pulmonary exacerbations are now recognized as a major driver of disease progression, and greater attempts to recognize and treat them before irreversible injury have become a priority. As lung disease progresses, exacerbations characteristically become more frequent and severe, often requiring extended courses of intravenous (IV) antibiotics and hospitalization. End-stage lung disease associated with impairment of daily activities heralds a sequence of terminal events in the absence of lung transplantation, including hypoxemia, pulmonary hypertension, cor pulmonale, respiratory failure, and death.

Microbiology

Patients with CF demonstrate colonization of the airways early in life with bacteria characteristic for the disease. Chronic infection of the lower airways, once established, is difficult to eradicate. Staphylococcus aureus and Haemophilus influenzae are usually the first organisms detected and often present with a benign clinical picture ( Fig. 47-9 ). Historically, acquisition of P. aeruginosa or Burkholderia cepacia was regarded as a particularly ominous clinical finding because these infections are associated with accelerated decline in lung function and increased mortality. With the increased prevalence of microbiologic screening and more aggressive antimicrobial treatment regimens, a more broad and diversified group of respiratory pathogens has emerged. These pathogens include methicillin-resistant S. aureus (MRSA), multidrug-resistant gram-negative rods, atypical mycobacteria, and fungal organisms. Often the clinical significance and factors contributing to the presence of these organisms in sputum culture are complex and multifactorial and thus require careful clinical consideration to determine the appropriate timing and intensity for treatment because some organisms may represent benign colonization.

Alterations in sputum microbiology with age.

This figure depicts increasing chronic infection with Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA) with advancing age based on data from CF centers in the National CF Patient Registry. This is contrasted with the higher prevalence of Haemophilus influenzae and methicillin-sensitive S. aureus in childhood and adolescence. MDR-PA, multidrug-resistant P. aeruginosa .

(Data from the Cystic Fibrosis Foundation Patient Registry; Bethesda, MD; 2012).

The prevalence of P. aeruginosa increases with age, infecting greater than two thirds of patients by the third decade of life. The frequency of detection earlier in life has increased in the era of newborn screening and may be as high as 20% in children younger than 2; detection in infants at the time of diagnosis is not uncommon. Acquisition of P. aeruginosa increases longitudinally (see Fig. 47-9 ) and has been associated with genotype severity—individuals homozygous for F508del demonstrate a higher prevalence of chronic infection.

Considerable concern has emerged surrounding the nosocomial acquisition of new infections and transmission of multidrug-resistant organisms. As the concept of eradication of initial infection has become standard practice, identification and earlier treatment of initial infection with P. aeruginosa using frequent respiratory sputum sampling to detect these inciting events is recommended in those not previously colonized. Sputum bacteriology correlates reasonably well with specimens obtained directly from the lower respiratory tract, although microbial sequencing is lending new insights into the validity of this conclusion. Oropharyngeal swab cultures that yield S. aureus or P. aeruginosa are modestly predictive of results from bronchoscopic specimens, but negative pharyngeal cultures do not rule out the presence of these organisms in lower airways. Nevertheless, recent studies supporting sputum or oropharyngeal swab culture surveillance and subsequent treatment with intent to eradicate infection have demonstrated similar efficacy to more invasive detection regimens. Other more sensitive means to detect P. aeruginosa, such as PCR, may further improve sensitivity of noninvasive techniques. Quantitative bacteriology may be particularly useful for determining the relative contributions of isolated organisms.

As lung disease progresses, P. aeruginosa often becomes the predominant organism recovered from sputum and may be present in multiple strains with different antibiotic sensitivity patterns. The emergence of a mucoid phenotype due to elaboration of large amounts of alginate is associated with worsened clinical outcome. Mucoid organisms are found as microcolonies of pseudomonads embedded and growing in biofilms of alginate. Biofilms inhibit phagocytosis and enhance bacterial adherence while limiting exposure to antibiotics and reactive intermediates produced by leukocytes. Although the presence of a mucoid phenotype is clearly associated with colonization, new mucoid Pseudomonas infection or conversion to mucoid phenotype may also be amenable to aggressive eradication strategies. Isolation of P. aeruginosa from the lower respiratory tract of a child or young adult with chronic lung symptoms is highly suggestive of CF but has been reported in patients with primary ciliary dyskinesia or other severe obstructive lung diseases.

Although P. aeruginosa has remained the dominant pathogen in adult CF lung disease, MRSA has emerged as a significant contributor to disease progression and mortality. In contrast, methicillin-susceptible strains are associated with improved outcomes. As MRSA has continued to emerge as a significant public health issue, its prevalence and severity in CF lung disease has become more evident. Two studies using the U.S. CF registry have demonstrated an association between MRSA and worsened lung function, whereas another has shown MRSA infection to be an independent risk factor for mortality. Considerable interest has emerged in the epidemiology and biologic significance of the staphylococcal chromosomal cassette (SCC) containing the methicillin-resistance gene (mecA) and the Panton-Valentine leukocidin (PVL) expressing strains, both of which have an effect on their virulence and impact on disease severity. Small-colony staphylococcus variants may also represent a distinct entity highly resistant to traditional therapy.

Many different gram-negative pathogens in addition to P. aeruginosa have demonstrated significant clinical impact on respiratory health, lending support to the consideration that CF lung infection represents a more complex microbiome than previously realized. Of these, B. cepacia has remained the most ominous due to its inherent multidrug antibiotic resistance and association with rapid decline in respiratory health. Infection can spread from patient to patient, leading to stringent infection control measures within CF care settings. Infection has been linked to the rapid demise of a small percentage of patients with what is referred to as “ cepacia syndrome.” Molecular analyses have shown that B. cepacia complex is composed of at least nine phenotypically indistinguishable but genetically distinct species known as genomovars. Genomovars II (Burkholderia multivorans) and III (B. cenocepacia) have been associated with the cepacia syndrome, and genomovar III includes the highly transmissible strain that may be linked to expression of the cable pilus. Other gram-negative rods present in sputum include mucoid Escherichia coli, Stenotrophomonas maltophilia, Achromobacter xylosoxidans, Klebsiella, and Proteus . Of these, S. maltophilia and A. xylosoxidans appear to have the strongest association with poor respiratory outcomes, but their clinical significance warrants further evaluation. Other gram-negative organisms such as Pandoraea and Ralstonia spp. have been isolated from CF sputum samples, but further clinical investigations are necessary to determine significant clinical associations. Obligate anaerobes have been identified from CF lung tissue; they are typically undetected by traditional clinical microbiologic samples but may be found in large abscess cavities on rare occasions.

Infection with nontuberculous mycobacteria is increasing in the CF population presumably due to concomitant antibacterial therapy resulting in a favorable ecologic niche, host susceptibility, improved detection, and environmental prevalence. Up to 20% of adult patients in some clinics are colonized by nontuberculous mycobacteria. Although infection is often transient, the clinical impact of infection with these organisms appears to be increasing. Mycobacterium avium complex infection is of variable clinical significance, yet rapidly growing mycobacteria such as Mycobacterium abscessus can exhibit a more virulent course, prompting aggressive and long treatment regimens. In contrast, Mycobacterium tuberculosis infection has only been seen in sporadic cases. Studies to standardize treatment for these emerging pathogens have been proposed.

Nearly 40% of individuals with CF will grow Aspergillus fumigatus in the sputum during their lifetime; patients who have severe lung disease appear to have an even higher incidence of sputum positivity. The pathogenic potential of Aspergillus in an otherwise immunocompetent host is not well established. However, Aspergillus is clearly associated with allergic bronchopulmonary aspergillosis (ABPA). ABPA is present in approximately 2% of CF patients and adds a considerable burden of care. See ABPA in Chapters 38 and 48 .

Imaging

The earliest radiographic change in CF lung disease is usually hyperinflation of the lungs, reflecting obstruction of small airways. Central airway thickening and linear opacities, often producing a “tram-track” appearance, representing bronchiectasis, are eventually seen ( eFig. 47-1 ) and often progress with age ( eFig. 47-2 ). Frequently these findings are most easily appreciated in the right upper lobe ( eFig. 47-3 ), and this lobe is often involved before other regions of lung are affected. The appearance of bronchiectasis at chest CT is characteristic ( eFigs. 47-1B-E and 47-2B-E ); see Chapter 48 ). Although chest radiography is appealing due to accessibility and relatively low doses of radiation exposure, its sensitivity for detecting acute or subtle chronic changes during a period of acute exacerbation remains limited. Despite its limitations, chest radiographs remain the standard first-line imaging in surveillance and during acute exacerbations, episodes of hemoptysis ( eFig. 47-4 ), or to identify complications such as pneumothorax ( eFig. 47-5 ); its use is recommended biannually in stable CF patients.

High-resolution cross-sectional imaging using computed tomography (CT) is clearly superior for detecting mild disease and quantifying the extent and pattern of bronchiectasis ( eFigs. 47-1B-E and 47-2B-E ). Mosaic perfusion, manifesting as areas of inhomogeneous lung opacity and resulting from air trapping producing disordered pulmonary parenchymal perfusion resulting in maldistribution of pulmonary blood flow, is common at chest CT in patients with CF ( eFig. 47-6 ). Small nodules resulting from bronchial and bronchiolar impaction are commonly seen at chest CT ( eFig. 47-7 ). CT studies have demonstrated significant early changes in young children, before obvious clinical disease, supporting the notion that CF lung disease begins early in life and its detection is limited by the sensitivity of routine clinical procedures including spirometry. Regardless of the imaging modality used, the degree of hyperinflation generally increases over time. As endobronchial infection and reactive inflammation emerge, peribronchial cuffing becomes increasingly evident. Evidence of bronchiectasis, such as enlarged ring shadows or airway dilation and cysts, is common even early in life (see eFig. 47-1 ); upwards of 50% of patients demonstrate bronchiectasis by 3 to 5 years of age.

During acute pulmonary exacerbations, a variety of radiographic findings may be noted such as peripheral round densities and mucoid impaction seen radiographically as branching opacities. These may resolve during treatment, replaced by the emergence of bronchiectasis or cystic changes. Additional findings including subpleural blebs often become evident during the second decade of life and are most prominent along the mediastinal border.

The indications for CT scanning in place of routine chest radiography are not established. One clear indication may be the evaluation of a patient with focal disease who may be considered for lobectomy, or patients that suffer acute deteriorations to help detect complications or evidence of new pathogens. Magnetic resonance (MR) imaging, which avoids radiation associated with CT scans, can also define lung morphology, particularly as techniques have improved, but the relative sensitivity and specificity of MR techniques have not yet been established. MR-based ventilation scans are also possible using hyperpolarized gas; although they sensitively demonstrate airway obstruction, the method is generally restricted to research use due to poor availability and stability of the noble gases required.

Lung Function

Lung function is believed to be normal at birth in infants with CF, though recent studies have demonstrated the cycle of inflammation and infection is present within weeks to months, and reduced cartilage size may impact early airway obstruction. Infants with CF can exhibit increased airway resistance, gas trapping, and diminished flow rates. Many centers use infant pulmonary function testing to detect early evidence of airway obstruction and guide institution of therapeutic strategies before permanent lung injury. Lung clearance index, a test performed by measuring the time for clearance of an inert gas through multiple breath washout technique, is also emerging as a research tool to detect early evidence of ventilation abnormality and is not confounded by patient effort. Reliable and consistent spirometry can be obtained routinely at 5 to 6 years of age when children are able to cooperate.

The earliest evidence of airway obstruction is typically limited to the small airways and thus is often first detectable by reduced forced midexpiratory flow rates, reduced flows at low lung volumes, and gas trapping (i.e., elevated residual volume to total lung capacity ratio [RV/TLC]). Although infrequently used in clinical applications, abnormalities such as increased alveolar-arterial oxygen gradient, frequency dependence of dynamic compliance, reduced response of flows to a helium-oxygen mixture, elevated slope of phase III of the single-breath nitrogen washout, and an elevated physiologic dead space are often present. Spirometry is the most clinically useful test to follow the course of respiratory disease progression and is typically measured at each clinic visit. Equally important is assessment of the flow volume loop, which may demonstrate obstruction via concavity toward the volume axis before either the FEV ₁ or FEF _25%-75% is affected.

Over time patients with active lung disease can experience a progressive deterioration of lung function; the annualized rate of decline in FEV ₁ is approximately 2% to 3% of the predicted FEV ₁ annually. Despite recent advances, this progressive loss of lung function often accelerates in young adulthood. As lung disease progresses, peripheral airway obstruction yields to more generalized obstruction and progressive air trapping late in the disease as airways cease to contribute to gas exchange, mimicking a restrictive pattern on spirometric measurements of lung function.

As the chronic cycle of inflammation and infection contributes to lung disease, many patients with CF begin to show mild decrements of arterial P o ₂ . Oxygenation declines slowly throughout life and this decline is often not clinically significant until late in the disease course. Patients who are able to sustain adequate oxygenation usually function well despite the presence of obstructive impairment. When arterial P o ₂ values are sustained below 55 mm Hg, patients are at high risk for symptomatic pulmonary hypertension. Clinically significant hypoxemia most commonly presents during sleep, particularly during REM-associated hypoventilation, and is a significant factor leading to pulmonary hypertension. Desaturation at rest or with exertion as measured during a 6-minute walk are strong predictors of mortality. Elevation of arterial P co ₂ and FEV ₁ less than 30% define end-stage disease. Patients with FEV ₁ less than 30% of predicted, arterial P co ₂ greater than 50 mm Hg, or arterial P o ₂ less than 55 mm have a predicted 2-year mortality of 50% and should be considered for lung trans­plantation. Other prediction tools can also facilitate appropriate patient selection for referral and outperform reliance on any single criterion.

Bronchial hyperresponsiveness is a consistent finding in asthma, but its presence and clinical significance in CF remain controversial. Bronchial hyperresponsiveness in CF has been demonstrated during exercise testing, bronchoprovocation testing, or response to bronchodilators, with two thirds of CF patients demonstrating decreased forced expiratory flows after a bronchoconstrictive challenge. In contrast to cross-sectional studies, repeated tests every 1 to 3 months for a year have demonstrated bronchodilator responsiveness at least once in 95% of subjects. Despite its frequency, the pathogenesis of bronchial hyperresponsiveness in CF is unclear. Hyperresponsiveness is unrelated to the severity of pulmonary disease or indices of atopy but seems to be more prevalent during winter months. Hyperresponsiveness diminishes with exacerbations of lung disease but returns as lung function improves after 2 weeks of intensive antibiotic therapy. Patients with CF may also have a lack of response to bronchodilators that may be related to loss of tone in bronchiectatic airways.

Exercise tolerance in CF is related to the severity of airway obstruction. Nearly 50% of patients with moderate to severe airway obstruction may experience oxygen desaturation to below 90% during peak exercise. Persons with CF have higher than expected ventilatory muscle endurance, and this endurance can be further improved with inspiratory muscle training. However, improved inspiratory muscle strength and endurance do not augment exercise performance.

Exercise therapy has many physiologic and biologic benefits but has not been convincingly demonstrated to improve standard spirometry. Nevertheless, standardized aerobic and resistive exercise does improve cardiorespiratory fitness and quality of life (QOL) and has been associated with a reduced risk of hospitalization. Maximum oxygen consumption during exercise may be a better predictor of survival than routine pulmonary function testing, but the test is time consuming and not available in all clinical settings. Several more clinically efficient measures of physical fitness show strong correlation with peak oxygen consumption and disease prognosis, further reinforcing that improvement of physical health through exercise may positively affect overall health in CF.

Upper Respiratory Tract Disease

Chronic rhinosinusitis is present in virtually all patients with CF. CF sinus disease manifests as chronic, relapsing symptoms of increased upper airway secretions, moderate airflow obstruction, and widening of the nasal bridge. This is seen at imaging ( eFig. 47-8 ) as opacification of the paranasal sinuses in more than 90% of patients in the first year of life. Nasal polyps are seen in an additional 15% to 20% of patients. Nasal polyps usually present toward the end of the first or during the second decade of life and may be the clinical finding that triggers diagnostic evaluation. The presence of acute and chronic nasal obstruction can diminish olfactory function and may contribute to diminished dietary intake and subsequent nutritional decline. Despite the presence of radiographic abnormalities, symptoms can be surprisingly well tolerated. Acute or chronic symptoms of sinusitis manifest in less than 10% of children and in approximately 24% of adults. Nevertheless, colonization of the upper airway may contribute to lower airway disease, necessitating attention to its presence and severity.

Complications of Respiratory Tract Disease

Atelectasis is present in approximately 5% of CF patients during the first 5 years of life with diminishing frequency with advanced age. Atelectasis may be lobar or subsegmental, with the right lung being the most commonly affected. Furthermore, it may develop concurrently with pulmonary exacerbations or in the absence of clinical symptoms. Occasionally, atelectasis may result from endobronchial aspergillosis presenting as mucoid impaction with volume loss. However, in most instances, a discrete mucus plug is not evident on bronchoscopy.

Pneumothorax (see eFig. 47-5 ) is a well-recognized CF complication due to air trapping and subsequent rupture of subpleural blebs. Although the overall incidence is equal among sexes and fairly low (about 1% per year), this increases sharply with age and disease severity, with 20% of CF adults experiencing at least one pneumothorax during their lifetime. Typical presentations include acute onset of chest pain, dyspnea, respiratory distress, or hemoptysis. Tension pneumothorax, which is more common in CF than in other obstructive lung diseases, represents an emergent situation because the rapid accumulation of pleural air may become life-threatening. Similarly, simultaneous bilateral pneumothoraces have been described and constitute an emergency. Small asymptomatic pneumothoraces may also be discovered in patients following routine surveillance chest radiography. Recurrent pneumothoraces are common.

Hemoptysis is a relatively common event in CF (see eFig. 47-4 ) and is believed to result from mucosal erosions and bronchial artery hypertrophy, which are a consequence of chronic inflammation. The presence of hemoptysis correlates with disease severity and is more common in the setting of chronic MRSA infection. Blood streaking in the sputum is a frequent finding and may be chronic. Massive hemoptysis (>240 mL blood in 24 hours) presents in approximately 5% of patients during their lifetime. Due to the strong correlation between hemoptysis and exacerbations of lung infection, initial treatment should include antibiotic therapy and rest from chest physiotherapy. Typically, aerosol therapies are temporarily held and measures to promote clot stabilization are considered. There are anecdotal reports of the use of the antifibrinolytic, tranexamic acid. Beta blockade has also been reported to be helpful in acute and chronic hemoptysis presumably by reducing blood pressure and blunting the sympathetic response to coughing spells. Although bronchoscopy can help to localize the site of bleeding, emergent bronchoscopy or radiographic imaging are of limited clinical utility. Often, emergent bronchial artery embolization (see eFig. 47-4F and G ) should be attempted without waiting for these measures. Advances in invasive vascular interventions and improved clinical management strategies have resulted in significantly reduced mortality, which was historically as high as 10% after massive hemoptysis.

ABPA has a lifetime incidence of 2% to 8% in CF patients, although a few small cohorts report an incidence rate as high as 20%, suggesting an environmental association. Aspergillus is a common environmental mold and is frequently recovered in sputum culture. Although up to 50% of patients with CF may demonstrate precipitating antibodies to A. fumigatus in their serum, an IgE-mediated allergic hypersensitization must develop to manifest ABPA clinically. Clinical features include increased cough, dyspnea, wheezing, and the expectoration of rusty brown plugs containing many eosinophils. Radiographic findings can be present, including characteristic finger-in-glove pattern. Atelectasis and volume loss may also result from hyphae-laden mucoid impaction in segmental bronchi. A diagnosis of ABPA is made by fulfilling major and minor criteria that include characteristic clinical findings plus skin test hypersensitivity, elevated total IgE, and elevated levels of IgG and IgE antibodies against A. fumigatus or other fungi. (See Chapters 38 and 48 .)

Staphylococcal and pseudomonal empyemas have been described in patients with CF, but respiratory tract infections usually spare the pleural space, making complications such as pleural effusions and empyema uncommon. Nonetheless, pulmonary exacerbations may often be accompanied by pleuritic-type pain.

Digital clubbing, which is caused by hyperplasia and hypertrophy of connective tissue and increased vascularity of the distal phalanges, appears in virtually all patients with advanced CF and is often present early in individuals with active lung disease. The cause of clubbing is unknown, but it is known to resolve following lung transplantation. Its severity generally correlates with the severity of lung disease. Hypertrophic pulmonary osteoarthropathy is a common clinical entity presenting with advanced CF lung disease in up to 15% of older adolescents and adults. Radiographic evidence for periostitis may be present in up to 8% of subjects. The distal aspects of the tibia, fibula, radius, and ulna are the most commonly affected sites. Signs and symptoms include pain, bone tenderness, swelling, and warmth over the involved areas; however, some patients may not manifest clinical symptoms. Effusions are uncommon but may arise in adjacent joints. Pain with ambulation or following strenuous physical exercise is common. Hypertrophic pulmonary osteoarthropathy is exacerbated during periods of poor respiratory health and tends to subside with the resolution of pulmonary exacerbations. Its presence with end-stage lung disease may yield persistent clinical symptoms requiring chronic analgesic therapy. There are also rare instances of cutaneous vasculitis causing self-limited, painless, palpable purpura, typically involving the lower extremities.

Respiratory failure is the greatest contributor to mortality and is the cause of death in 90% of CF patients. Hypoxemia is first seen during exertion or sleep and progresses with lung disease. Hypercapnia is a late finding reflecting advanced lung disease, which typically progresses with disease severity and worsens during pulmonary exacerbations. Pulmonary hypertension and cor pulmonale may develop late in the disease process, resulting in hepatic congestion and peripheral edema. The role of pulmonary hypertension in CF is currently being studied to prioritize treatment regimens. Hypoxemia, if not recognized or adequately treated, will contribute to worsening pulmonary hypertension and cardiac failure. Pneumothorax, hemoptysis, and infections such as respiratory syncytial virus or influenza can cause acute respiratory failure that is reversible with aggressive treatment.

Gastrointestinal Manifestations

Meconium ileus is seen in about 15% of newborns and is pathognomonic for CF. Failure to pass thick inspissated meconium in the first 48 hours of life is associated with abdominal distention and rapidly advances to bilious emesis. Affected infants are at risk for intestinal perforation and peritonitis accompanied by shock. Radiographic features are typical of high-grade bowel obstruction revealing multiple dilated loops of intestine and air-fluid levels ( eFig. 47-9 ). A granular appearance of the lower abdomen may be noted due to accumulated meconium containing small air bubbles. The colon is characteristically small when visualized with contrast imaging ( eFig. 47-10 ). Scrotal and peritoneal calcification can be seen following ileal perforation in utero. Meconium obstruction in the colon may only delay passage of stool. This has been termed the meconium plug syndrome and is much less specific for CF.

Beyond the newborn period, 20% of patients may develop the distal intestinal obstruction syndrome (DIOS). This is characterized by obstruction in the cecum, proximal colon, or terminal ileum associated with voluminous, viscous, and incompletely digested intestinal contents and is similar to meconium ileus ( eFig. 47-11 ). Partial obstruction may manifest as a chronic or recurrent entity with intermittent crampy abdominal pain as the only symptom. Fulminant complete obstruction is associated with failure to pass stool, resulting in abdominal distention and vomiting that may be bilious or fecal if allowed to progress. A mobile right lower quadrant mass may be palpable. Risk factors for DIOS include previous episodes, dehydration, dietary change, immobilization, bacterial overgrowth, treatment with antibiotics, and constipating medications. Other causes of acute abdominal pain with obstruction should also be considered, including intussusception, intestinal adhesions from previous abdominal surgery, and appendicitis, which may be partially suppressed due to concurrent antibiotic therapy. Appendicitis is thought to be uncommon in CF, with a 2% lifetime risk compared with a 7% to 8% risk in the general population. Nonfilling of the appendix with contrast enema is seen in patients with CF; radiographic and histologic findings of a dilated, mucus-filled appendix are typical features, although the uninflamed appendix is frequently enlarged at CT in CF patients ( eFig. 47-12 ). The loss of pancreatic and bowel secretion of bicarbonate to buffer gastric acid may result in duodenal irritation and recurrent epigastric pain.

Gastroesophageal reflux is common in CF; increased abdominal pressure associated with obstructive lung disease contributes to its high prevalence. Consideration of gastroesophageal reflux is important because it can impair nutritional status and may contribute to microaspiration, accelerating lung disease. The presence of gastroesophageal reflux has been associated with worsened outcome after lung transplantation, and therefore patients with CF considered for lung transplantation should be evaluated thoroughly; some centers advocate surgical therapy for ongoing reflux during the peritransplant period.

Rectal prolapse develops in nearly 20% of children with CF but is an infrequent event for adults. Rectal prolapse in a child should raise consideration of CF even if it is the only obvious clinical symptom. Rectal prolapse is precipitated by the presence of bulky, sticky stools that adhere to rectal mucosa, poor nutritional status with loss of the perirectal fat that normally supports the rectum, and the presence of high intra-abdominal pressure due to frequent paroxysmal coughing.

Pancreatic Disease

Exocrine pancreatic insufficiency (PI), due to intraluminal obstruction with thickened, dehydrated secretions, is present from birth in 85% of patients with CF. Adequate exocrine pancreatic secretion is present in 10% to 15% of patients and has defined genotypic associations. Insufficient release of pancreatic enzymes into the gut results in impaired fat and protein digestion and impaired absorption in the small bowel. Pancreatic insufficiency results in frequent bulky, greasy, foul-smelling stools and protuberance of the abdomen due to increased intraluminal bacterial gas production. Assessment of pancreatic function by ELISA measurement of stool fecal elastase-1 may enhance clinical assessment of PI.

Untreated malabsorption results in nutritional failure and ultimately failure of linear growth, which has been linked to worsened outcomes. Patients with CF also grow slowly because of factors beyond nutritional intake and intestinal absorption. For example, increased expenditure of energy to accomplish the work of breathing may be an important contributor; systemic inflammation may also play a role. Fat-soluble vitamin deficiency was historically a common association at diagnosis in young children presenting with nutritional failure but is much less common in the era of newborn screening.

Symptomatic pancreatitis develops in less than 1% of adolescent or adult CF patients, usually in patients who have at least some residual exocrine pancreatic function. However, recurrent pancreatitis has been reported in association with CFTR dysfunction and can be the presenting feature of the disorder. Complete fatty replacement of the pancreas is common at CT ( eFig. 47-13 ), and pancreatic lipomatosis and fibrosis are characteristic of pediatric CF. Pancreatic ductal dilation and calcifications may also be seen in CF patients. Rarely, there may be cystic replacement of the pancreas, referred to as pancreatic cystosis .

Hepatobiliary Disease

Although relatively common, there is not yet consensus on the definition of CF-associated liver disease. Histologically, liver disease manifests as focal biliary cirrhosis and produces clinically significant liver disease in a greater proportion of CF patients as longevity increases. One description of clinically significant disease includes the constellation of an abnormal physical exam, abnormal liver function testing, and imaging findings. Liver disease may present as hepatosplenomegaly or as persistent elevation of hepatic enzymes (AST, ALT, bilirubin, GGT). Current guidelines recommend evaluation for liver disease when enzymes are elevated more than 3 times normal or remain 1.5 to 3 times normal for 3 months. Patients with CF-associated liver disease rarely develop acute fulminant hepatic failure but with advanced cirrhosis can develop portal hypertension and clinically significant esophageal varices. Steatosis is a common clinical finding and has been associated with malnutrition, essential fatty acid deficiency, and/or oxidative stress. Ultrasound is generally used to monitor severity and is currently being studied in longitudinal studies.

Biliary tract disease is common in CF. Cholelithiasis develops in approximately 10% of patients but causes significant clinical symptoms in less than 4% of cases. Formation of stones is encouraged by the abnormal ion transport environment resident in the gallbladder. A small or microgallbladder is seen in 20% to 30% of patients and is of unknown clinical significance.

Genitourinary Tract Abnormalities

The vas deferens is congenitally absent in almost all males with CF. Semen analysis is typically required to identify the 1% of male CF patients who are fertile. The volume of ejaculate is usually one third to one half of normal; complete absence of spermatozoa in addition to a number of chemical abnormalities that reflect absence of secretions from the seminal vesicles is usually apparent. An increased incidence of inguinal hernia and hydrocele has also been reported.

Although significantly less common, female infertility in CF may be as high as 20%. Many women with CF and advanced lung disease and/or malnutrition are anovulatory. Another obstacle to conception is the presence of thick tenacious cervical mucus and an increased presence of endocervicitis. This dehydrated mucus also has abnormal electrolyte concentrations, preventing the usual ferning at midcycle. As a result, this mucosal abnormality is thought to impede normal sperm migration. Urinary incontinence can be seen beginning in adolescence and does not clearly correlate with disease severity; rather, incontinence probably reflects chronic cough and increased abdominal pressure due to airflow obstruction.

Pregnancy in females with CF appears to be increasing as clinical outcomes have improved. A longitudinal study of 325 pregnant women with CF demonstrated 258 live births (79%) and 67% therapeutic abortions. Compared with 1142 age- and severity-matched controls, pregnancy in a woman with CF did not have an independent negative effect on pulmonary status or mortality over 2 years. Successful pregnancy and delivery is possible and may be carried out safely, but it is essential that women with CF consider implications to their health as they consider family planning. Successful breast feeding has been reported in women with CF.

Sweat Gland Dysfunction

Sweat chloride is elevated in most CF patients due to abnormal chloride reabsorption in the sweat duct due to the absence of CFTR. This may predispose individuals with CF to excessive salt loss in certain settings. Young children are especially susceptible to depletion, especially when exposed to warm arid climates, or when there is additional salt loss due to vomiting or diarrhea. Typically children in this circumstance present with lethargy, anorexia, and hypochloremic alkalosis and/or hyponatremia.

Overview

Considerable progress in the understanding of CF lung disease and a concerted effort to address disease manifestations in a comprehensive fashion have led to a number of new therapeutic approaches to the disease. Therapies including mucolytics, hydrators of the airway surface, inhaled antimicrobials, , systemic anti-inflammatory treatments, and nutritional support are the mainstays of current CF treatment ( Table 47-4 ). These sorts of supportive therapies, in addition to comprehensive care undertaken at organized CF clinical centers, are responsible for the steadily improving life expectancy observed over the past decade ( Fig. 47-10 ), resulting in a median life expectancy of 41 years in 2012, based on the latest analysis of the U.S. registry. Similar improvements have been observed in other developed countries. More recently, the first treatment that addresses the basic defect in CF by restoring CFTR function in patients with the G551D CFTR gating mutation has been approved, resulting in marked improvements among individuals with this mutation. Other therapies that address more common CFTR alleles or aim to replace CFTR with gene therapy are in various stages of clinical development and promise to open a new therapeutic era in the disease ( Fig. 47-11 ).

Median predicted survival in CF.

Median predicted survival for CF patients in U.S. Cystic Fibrosis centers in the CF registry from 2012. Data are depicted as medians for 5-year time spans from 1986–2012. Since the mid-1980s, predicted survival has increased from about 27 years of age to older than 36 years of age for the most recent 5-year time span.

(Data from the Cystic Fibrosis Foundation Patient Registry, Bethesda, MD; 2012.)

CF therapeutics by category.

This figure depicts the mechanisms of CF airway pathology. CF therapeutics attempt to address defective CFTR function by gene therapy or modulation of CFTR expression or function; to address the diminished airway surface liquid, abnormally viscous mucus, and disrupted mucociliary clearance; and finally, to address chronic airways infection and inflammation. When respiratory failure develops, lung transplantation is the remaining option.

Monitoring and Aggressive Approach

Aggressive monitoring and treatment of pulmonary infections and other common CF complications has been a key advance that is believed to have contributed significantly to improved clinical outcomes in the past decade. As discussed earlier, the predominant lower airway pathogen changes through the lifetime of CF patients, with S. aureus and H. influenza being the most common organisms isolated during infancy and early childhood, while P. aeruginosa typically becomes the dominant pathogen later in the disease. (See Fig. 47-9 .) New attention has focused on the efforts to delay or prevent chronic colonization with P. aeruginosa, with a number of studies evaluating various regimens to best achieve sustained clearance. Coincident with this approach is increased attention to infection control to prevent acquisition of virulent organisms, particularly in health care settings where risk of acquisition is significant. Segregation of clinics by Pseudomonas status can reduce acquisition of epidemic organisms; standard infection control procedures alone may not be sufficient, leading to even more rigorous guidelines for patient isolation and prevention against comingling. Once established, P. aeruginosa is challenging to eradicate with antimicrobial therapy alone, although it can be accomplished in selected patients. Chronic colonization with Pseudomonas is associated with a more rapid decline in respiratory status, although not as severely as more virulent organisms such as B. cepacia or B. dolosa, which can also be associated with outbreaks due to nosocomial spread. As with chronic infection, early and aggressive nutritional therapy, particularly among young children with CF, has been crucial to achieving improved outcomes. Despite advances in lung function and delays in Pseudomonas growth, delaying the progression of CF in the adolescent years remains a challenge.

Sponsored in part by the CF Foundation, U.S. centers have embarked on a rigorous quality improvement program to facilitate best practices in CF care. Outcomes are compared between centers and made publically available to encourage continuous improvement. A culture of cooperation among centers, patients, families, and the Cystic Fibrosis Foundation has led to evidence-based guidelines, protocol-driven therapies, and rapid dissemination of results to facilitate advances in health care delivery.

CFTR Potentiators, Correctors, and Other Treatments for the Basic Defect

On the basis of functional consequences of various CFTR mutations, specific therapeutic strategies to restore deficient or defective protein function are being developed by altering CFTR expression or function ( Fig. 47-12 and Table 47-5 ). Because of ambitious high-throughput screening efforts, the benefits of these new CFTR modulators have begun to come to fruition for CF patients. Results in the clinic demonstrate that the rescue of the CFTR protein by the archetype CFTR modulator ivacaftor is associated with marked improvements in the clinical outcome that compare favorably with previous therapies widely used by CF patients. The CFTR potentiator ivacaftor (formerly VX-770) was the first to advance as an approved CF treatment among patients with the G551D gating mutation, an allele represented in approximately 4% of CF patients. CFTR potentiators function to activate CFTR channels located at the cell surface. They potentiate cAMP-mediated channel gating via decoupling with ATP hydrolysis and may be more broadly useful against a greater variety of CFTR missense alleles, including other class III gating mutations and other CFTR forms that exhibit residual activity at the cell surface (i.e., conductance and mild processing mutants). The highly efficacious treatment benefit observed with ivacaftor therapy has engendered considerable interest toward recapitulating its effects among other more common CFTR alleles. This includes correctors of F508del CFTR misfolding, termed CFTR correctors. CFTR correctors attempt to restore normal CFTR processing to the most common CFTR mutation. Other agents that induce readthrough (or suppression) of PTCs to induce expression of full-length CFTR are also under development and have shown promise in proof-of-concept clinical trials. Other approaches beyond these small molecule CFTR modulators are also being explored. For example, gene replacement by viral and nonviral gene therapy remains an approach under active investigation, as well as newer strategies that attempt to express CFTR through transduction of mRNA alone. In total, this class of agents that target the basic CF defect serves as both a prime example of the potential for new genetic-based approaches in CF and a seminal example for other genetic diseases. Because many patients (≈40%) are complex heterozygotes for more than one CFTR mutation, combination therapeutics addressing more than one CFTR allele or use of multidrug therapy seem likely in the future and will dictate a need for individualized therapeutics optimized for particular patients on the basis of their underlying disease and other genetic covariates.

CFTR modulators by mutation class.

Classes of defects in the CFTR gene include premature termination codons (PTCs) causing truncated protein translation (class I); misfolded CFTR, including deletion of phenylalanine at position 508 (class II, location shown with a red dot); full-length CFTR that reaches the cell surface but exhibits abnormal channel gating (class III; ATP hydrolysis is disrupted, at site identified in red) or reduced pore conductivity (class IV); and full-length CFTR with splicing errors (class V); premature termination codon suppressors (e.g., aminoglycosides, ataluren) bind to ribosomal subunits ( green star ) to allow suppression of PTCs and expression of full-length protein. Class II mutations like F508del can respond to small-molecule corrector compounds to restore folding defects and/or enhance expression of the channel at the cell membrane (e.g., lumacaftor, VX-661). Without correction, almost all class II CFTR is sent to the proteosome, leaving detectable surface protein in only rare individuals. CFTR potentiators include ivacaftor ( green chevron ) for patients with CFTR gating mutations. Future directions include exploring the use of CFTR potentiators for other mutant CFTR known to reside at the cell surface, such as noncanonical splicing mutations. Combination therapy with potentiators has also been proposed for classes I and II CFTR mutations.

(Adapted from Rowe SM, Borowitz DS, Burns JL, et al: Progress in cystic fibrosis and the CF Therapeutics Development Network. Thorax 67(10):882–890, Fig. 1, 2012.)

Table 47-5

Approved and/or Investigational Therapeutics Targeting Each CFTR Mutational Class

CFTR Modulator Class	Representative Molecules	CFTR Mutations Affected	CFTR Mutation Class
Potentiator	Ivacaftor (VX-770)	G551D, non-G551D gating mutations * , other surface localized CFTR alleles * , F508del CFTR †	Class III Possibly classes IV, II, and I
Corrector	Lumacaftor (VX-809), VX-661, N6022 (N30 Pharma)	F508del *	Class II
Premature termination codon suppressor	Ataluren (PTC124), aminoglycosides	Nonsense mutations	Class I
Splicing repair	Antisense oligonucleotides	Splicing mutations *	Class V

 

CFTR, cystic fibrosis transmembrane conductance regulator.

* Investigational.

† In combination with a CFTR corrector.

CFTR Potentiators.

CFTR potentiators activate mutant CFTR localized to the cell surface by potentiating channel gating stimulated by physiologic activation of cyclic AMP. The impetus for development included the anticipated need to activate F508del CFTR once localized to the cell surface, in addition to the need to activate other mutant channels that reside at the plasma membrane but are dysfunctional due to aberrant gating, impaired conductance, or a reduced number of channels due to mild processing mutations or splice variants. Early attempts included use of genistein, a natural flavonoid molecule that demonstrates strong activation of CFTR but is poorly absorbed and exhibits other undesirable physiologic properties. Subsequently ivacaftor and other investigational CFTR potentiators were discovered via high-throughput screening approaches that identified small molecule potentiators optimized for druglike qualities through traditional medicinal chemistry approaches. Ivacaftor induced about 50% CFTR activity in G551D/F508del CFTR-expressing primary epithelial cells, an emerging benchmark for preclinical development of CFTR modulators.

In a long-term Phase III placebo-controlled trial conducted in older children and adults (age 12 and older) with CF and with at least one copy of the G551D- CFTR mutation, ivacaftor caused a roughly 10% absolute improvement in FEV ₁ % at 24 weeks, an effect durable through 48 weeks of testing, providing confirmation of prior Phase II testing. In addition, all secondary clinical end points showed meaningful and statistically significant improvements, including a 55% reduction in the probability of experiencing a pulmonary exacerbation, improved weight gain, and an improvement in respiratory symptoms. Sweat chloride testing also exhibited marked improvements (mean of sweat chloride was about 55 mEq/L, a value below the traditional diagnostic threshold). Of note the improvement in spirometry was rapid; within 2 weeks, 90% of the maximal improvement was observed, suggesting that the mechanism was clearance of mucus, rather than the reversal of longstanding structural lung disease. In a smaller study that enrolled pediatric G551D CF patients 6 to 12 years old, benefit was also documented by lung clearance index measurements in CF patients with minimal detectable lung function abnormality by spirometry, likely reflecting activity in the small airways. The degree of improvement in spirometry among participants of the Phase III trial of ivacaftor compares favorably with that of commonly used therapies for chronic CF care, including inhaled recombinant human DNase, inhaled tobramycin, azithromycin, and hypertonic saline. In all clinical studies to date, ivacaftor appeared safe and well tolerated, although monitoring of liver function tests is recommended.

In contrast to the large effect in G551D patients, ivacaftor monotherapy had no meaningful effect in CF patients homozygous for F508del CFTR , establishing that CFTR potentiation alone is unlikely to be effective without concomitant administration of a corrector molecule to bring F508del CFTR to the cell surface.

These findings have formed the basis of clinical approval for ivacaftor in CF patients with G551D mutations, and this treatment has rapidly been deployed to children and adults with the appropriate mutation. Establishing the proof of concept that CFTR modulation could have pronounced clinical effects among CF patients has also engendered considerable interest in extending these findings toward other CFTR mutations in which ivacaftor might be effective and developing CFTR modulators for other common CFTR alleles. The long-term benefit of effective modulation of CFTR will also be evaluated in ongoing long-term observational studies, which will examine the effects of ivacaftor in G551D CF patients on clinical outcome, sputum microbiology, and systemic inflammation, among other parameters.

Because ivacaftor also increased CFTR activity in other gating mutations in vitro, these non-G551D gating mutations were also tested; the beneficial effects of ivacaftor were also observed in patients with other non-G551D class III gating mutations. Patients observed a large increase in FEV ₁ and similar decrease in sweat chloride compared with that seen in patients with G551D, suggesting a class effect. Other CFTR mutations, such as conductance mutations, mild processing mutations, or CFTR splice variants allow low levels of partially active CFTR channels to reach the cell surface, conferring partial CFTR function and typically a milder CF phenotype. The effect of ivacaftor on partially active missense mutations from these CFTR classes is generally proportionate to basal CFTR function, reflecting the ability of increased gating to compensate in part for reduced surface expression or conductance, even when gating is normal in these CFTR forms. With this information in mind, ivacaftor is being tested in the archetype conductance mutation R117H in addition to individuals who exhibit residual CFTR function and a relevant CFTR allele. These results may justify use of CFTR potentiators in a broader group of individuals with CF. Combined with individuals with gating mutations, this represents approximately 10% to 15% of the CF population who ultimately might benefit from monotherapy with a CFTR potentiator.

CFTR Correctors.

Significant effort has been directed toward the goal of correcting the folding of F508del CFTR, thus restoring ion channel activity to the misfolded protein. Early attempts include the evaluation of agents such as 4-phenyl butyrate to down-regulate HSC70 (or other protein processing chaperones), a pathway central to the protein folding process that has been shown to augment F508del CFTR expression in vitro and represent an early example of compounds tested in the clinic. Curcumin and 8-cyclopentyl-1,3-dipropylxanthine are examples of F508del CFTR processing correctors that did not successfully translate from in vitro studies to clinical results. More recent efforts have resulted from high-throughput library screens for chloride channel function following incubation of test compounds with F508del expressing cells. A number of these strategies have identified F508del correctors that may address cell biogenesis through chaperone pathways. Pharmacologic activity of such agents has also been reported to augment F508del CFTR half-life in the plasma membrane through altered surface recycling attributed to features of the cellular processing machinery or reduced endocytic trafficking. This class of agents may be potential drug development candidates if their safety in vivo is confirmed. Other compounds have been shown to directly interact with CFTR and may offer greater specificity than agents that alter general aspects of cell folding or cellular quality control.

Success toward effective correction of F508del CFTR was seen during initial Phase II testing of lumacaftor (formerly VX-809), a putative F508del CFTR corrector. Results demonstrated modest decreases in sweat chloride compared with placebo but were not accompanied by improvements in spirometry or other clinical measures. Although this established that rescue of F508del CFTR in human subjects is achievable by a systemically delivered small molecule, the degree of CFTR rescue was insufficient to confer clinical improvement. Because F508del CFTR also exhibits abnormal channel gating, in addition to aberrant cellular processing, an approach to address insufficient activity is to coadminister a CFTR potentiator with a CFTR corrector. This approach is substantiated by in vitro results demonstrating that combination therapy increased CFTR function in F508del CFTR homozygous HBE cells. Recently a Phase II trial demonstrated that coadministration of lumacaftor and ivacaftor resulted in significant improvements in FEV ₁ (≈6% treatment effect), although changes in sweat chloride did not correlate with additional benefit seen following addition of the potentiator. Lesser effects were observed in CF patients heterozygous for F508del CFTR , providing evidence of a gene dose effect. The potential benefit of ivacaftor and lumacaftor in combination among F508del homozygous CF patients is presently being pursued in two large international Phase III trials. If positive, results could justify use of corrector-potentiator combination therapy for the most common CFTR mutation. An alternative corrector, VX-661, which has a similar mechanism as lumacaftor but more advantageous pharmacologic properties, also demonstrated additive benefit when used in combination with ivacaftor in individuals homozygous for F508del CFTR alleles in Phase II testing, providing additional confidence that the combination of corrector and potentiator agents could be effective among individuals expressing F508del CFTR. The results also suggest that ivacaftor may be useful to augment the rescue of other mutant forms of CFTR , such as premature termination codons or other processing mutants (e.g., class II mutations) sensitive to the effects of CFTR correctors.

Other strategies to identify agents that augment F508del CFTR folding are also being developed, and interest has accelerated with recent successes in clinical studies. Using a trafficking assay based on epitope-tagged CFTR, phosphodiesterase inhibitors including sildenafil and other active analogues have been shown to improve surface localization of the mutant protein. The same agents augment short-circuit current in F508del CFTR-expressing cell lines and enhance NPD in CF mice, as did the related compound vardanafil. Because misfolded F508del CFTR exhibits two fundamentally distinct properties that alter its processing (namely nucleotide binding domain 1 stability and interdomain assembly), it has subsequently been recognized that compounds that address these mechanisms independently can exhibit additive or synergistic effects of correction of F508del CFTR misfolding. Moreover, the global cellular response to misfolded protein may also represent a target. For example, treatment of CF cells with histone deacetylase (HDAC) inhibitors can modulate ER stress, and HDACs such as suberoylanilide hydroxamic acid (SAHA), as well as siRNA-silencing, increase levels of F508del CFTR in the cell membrane. Additive or synergistic rescue of F508del CFTR using more than one such strategy may offer hope of achieving ion transport activity sufficient to confer a normal phenotype in CF respiratory epithelia.

Translational Readthrough.

Readthrough of premature termination codons (PTC) represents a potential approach to address CF caused by this mechanism, which could also be applicable to many other genetic disease caused by nonsense mutations. The approach was identified when certain aminoglycoside antibiotics were found to interact with the eukaryotic rRNA within the ribosomal subunits. Through this interaction, the fidelity of eukaryotic translation can be altered by interrupting the normal proofreading function of the ribosome. Insertion of a near cognate amino acid (AA) at a PTC allows protein translation to continue normally. Specificity is conferred by greater termination codon fidelity at the authentic (3′) end of mRNA and has been established in vitro by demonstrating that there is no detectable elongation beyond native termination codons. This has been bolstered by a good safety profile in both preclinical and clinical studies and has subsequently been adopted in a number of relatively common genetic diseases aside from CF in which premature termination codons are relatively prevalent, including Duchenne muscular dystrophy, Hurler syndrome, ceroid lipofuscinosis, nephropathic cystinosis, and expression of mutated p53.

Proof of concept experiments with aminoglycosides established that PTCs within CFTR in human subjects can be suppressed, resulting in the synthesis of full-length, functional CFTR protein. The approach has also demonstrated success in mouse models of CF. Following two small pilot trials indicating restoration of chloride secretion in CF subjects harboring PTCs, a double-blind, placebo-controlled trial conducted in Israel showed correction of nasal ion transport specifically in subjects with nonsense mutations following topical administration of gentamicin, and as expected, not in CF controls homozygous for F508del. A trial examining systemic gentamicin in seven French subjects with Y122X CFTR , a mutation highly susceptible to readthrough, also indicated rescue of CFTR activity in the airway and sweat duct. Not all aminoglycoside trials in CF have demonstrated success, suggesting low levels of protein correction. Regardless, due to the known toxicity and poor bioavailability of aminoglycosides, more efficacious agents that avoid undesirable properties of aminoglycosides will be needed for the long-term genetic treatment of CF. One promising approach uses medicinal chemistry optimization to identify the antimicrobial, toxic, and readthrough effects of aminoglycoside scaffolds, a strategy demonstrating initial success using in vitro reporters of efficacy and toxicity, and cell- and animal-based models of CFTR rescue.

Others have attempted to identify novel compounds that address the disadvantages of aminoglycosides. One such molecule is ataluren (formerly PTC124), an investigational agent resulting from high-throughput screening efforts to identify molecules that induce translational readthrough. Ataluren is an orally bioavailable agent that has demonstrated efficacy in vivo in a transgenic mouse model of nonsense-mediated CF. The drug is well tolerated in normal and CF subjects, leading to a series of clinical trials examining ataluren in CF individuals harboring nonsense alleles. Results thus far have been mixed. Two studies, conducted in Israel among adults and France/Belgium in pediatric subjects, detected rescue of CFTR activity (as detected by the NPD) in open-label, two-dose crossover Phase II trials in CF subjects possessing at least one premature termination codon. The former study included follow-up testing that examined the effect of ataluren for 3 months in 19 subjects previously studied for 2 weeks; significant and time-dependent improvement in CFTR activity following the treatment period were observed. In contrast, a nearly identical trial conducted in the United States did not demonstrate improvement in CFTR function, raising questions about efficacy. Subsequent Phase III testing conducted over 1 year was negative, although a trend toward improved FEV ₁ and exacerbation frequency was observed, which was statistically significant in a subgroup of individuals not exposed to inhaled tobramycin, a concomitant medication later shown to attenuate the beneficial effect of ataluren in vitro. Future studies will evaluate the effect of ataluren in CF subjects with nonsense mutations that are not exposed to tobramycin. The effects of ataluren are likely also modulated by genetic founder effects including the degree of CFTR mRNA expression at baseline as modulated by nonsense mediated decay. Poor selection and optimization of ataluren efficacy has also been suggested to have affected successful development of this agent, due in part to its stabilization of firefly luciferase, which induces a paradoxical and off-target increase in the reporter assay used to select the agent. More recent studies have highlighted its relatively poor activity. Given new knowledge regarding the best models to identify and optimize CFTR drug candidates that has emerged since the identification of ataluren, it is possible that identification of new chemicals using alternate readthrough assays might yield more efficacious compounds.

Respiratory Therapies

Other Supportive Care

Treatment of Lung Complications

Treatments directed to control airway infection, limit inflammation, and optimize airway clearance are the cornerstone of therapy in both early and advanced CF lung disease. Hypoxemic respiratory insufficiency should be recognized and treated with supplemental oxygen. Low-flow oxygen is effective at relieving nocturnal, exertional, and resting hypoxemia and does not usually cause significant hypercapnia. The development of nocturnal or resting hypoxemia is strongly associated with the onset of pulmonary hypertension and is associated with increased mortality. Diuretics, inotropic agents, and theophylline provide little to no benefit in CF and are rarely used. Cor pulmonale is an end-stage finding in advanced disease with few viable treatment options beyond those to stabilize lung disease and symptomatically treat pulmonary hypertension.

Acute respiratory failure due to reversible insults can be stabilized by either non invasive or invasive ventilatory support. Although the associated mortality with mechanical ventilation is high, it is not absolute. It has been reported that one third of CF patients with advanced lung disease survive events requiring mechanical ventilation. Mechanical ventilation can also be used as a bridge to lung transplant at appropriate centers. Extracorporeal membrane oxygenation has also been used for this purpose.

Atelectasis is managed by escalating the intensity and frequency of airway clearance and usual therapy directed at pulmonary exacerbations. Systemic or inhaled corticosteroids may be helpful in the presence of asthma, ABPA, or significant airway inflammation refractory to antimicrobial therapy. There is little evidence that bronchoscopy and lavage are effective in treating atelectasis. However, bronchoscopic evaluation of atelectasis and treatment of mucoid impaction in association with endobronchial aspergillosis and ABPA have been described. ABPA typically responds to standard doses of systemic corticosteroids. Inhaled corticosteroids, suppressive oral antifungal therapy, and omalizumab may be useful in limiting the systemic steroid burden.

When small and minimally symptomatic, a pneumothorax can be observed with expectation of spontaneous resolution. Pneumothoraces of greater volume (e.g. >20%) or that compromise ventilation or physiologic stability require chest tube decompression. Recurrent pneumothoraces are common in advanced lung disease and are associated with higher mortality. Patients with chronic pneumothorax who are clinically unstable or experience significant morbidity may benefit from chemical and/or mechanical pleurodesis. Prior pleurodesis is not considered a contraindication to lung transplantation.

Major hemoptysis is treated with antibiotics and chest rest, although reinstitution of therapy has become more aggressive over time. Supplemental vitamin K should be given if the prothrombin time is prolonged due to malabsorption. Clot stabilization with tranexamic acid and systemic blood pressure reduction with beta-blockade may have some clinical utility. Massive hemoptysis may resolve with conservative therapy, but bronchial artery embolization provides more definitive clinical control and can also be considered in recurrent cases of submassive bleeding.

Lung Transplantation

Sequential double-lung transplantation is an accepted therapy for respiratory failure secondary to CF. Living lobar transplantation is an effective alternative to conventional cadaveric lung transplants but is only performed in selected centers. More than 2300 lung transplants have been performed for CF around the world, and the rate is increasing. The 5-year survival is just above 50%, which compares favorably to the survival of lung transplant performed in other lung diseases. Given the limited survival and complex treatment required, consideration of lung transplantation requires a careful psychological and social evaluation. Should lung transplantation be chosen, patients should be thoroughly evaluated and referred when they have a survival benefit or predicted mortality in the range of 2 to 3 years; prediction formulas can assist with this process. Following successful transplantation, the patient can experience a dramatically improved respiratory status and QOL, but significant medical treatment burden can remain.

Key Points

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