Familial Pulmonary Fibrosis Research Articles

A CommonMUC5BPromoter Polymorphism and Pulmonary Fibrosis

The mutations that have been implicated in pulmonary fibrosis account for only a small proportion of the population risk. Using a genomewide linkage scan, we detected linkage between idiopathic interstitial pneumonia and a 3.4-Mb region of chromosome 11p15 in 82 families. We then evaluated genetic variation in this region in gel-forming mucin genes expressed in the lung among 83 subjects with familial interstitial pneumonia, 492 subjects with idiopathic pulmonary fibrosis, and 322 controls. MUC5B expression was assessed in lung tissue. Linkage and fine mapping were used to identify a region of interest on the p-terminus of chromosome 11 that included gel-forming mucin genes. The minor-allele of the single-nucleotide polymorphism (SNP) rs35705950, located 3 kb upstream of the MUC5B transcription start site, was present at a frequency of 34% among subjects with familial interstitial pneumonia, 38% among subjects with idiopathic pulmonary fibrosis, and 9% among controls (allelic association with familial interstitial pneumonia, P=1.2×10(-15); allelic association with idiopathic pulmonary fibrosis, P=2.5×10(-37)). The odds ratios for disease among subjects who were heterozygous and those who were homozygous for the minor allele of this SNP were 6.8 (95% confidence interval [CI], 3.9 to 12.0) and 20.8 (95% CI, 3.8 to 113.7), respectively, for familial interstitial pneumonia and 9.0 (95% CI, 6.2 to 13.1) and 21.8 (95% CI, 5.1 to 93.5), respectively, for idiopathic pulmonary fibrosis. MUC5B expression in the lung was 14.1 times as high in subjects who had idiopathic pulmonary fibrosis as in those who did not (P<0.001). The variant allele of rs35705950 was associated with up-regulation in MUC5B expression in the lung in unaffected subjects (expression was 37.4 times as high as in unaffected subjects homozygous for the wild-type allele, P<0.001). MUC5B protein was expressed in lesions of idiopathic pulmonary fibrosis. A common polymorphism in the promoter of MUC5B is associated with familial interstitial pneumonia and idiopathic pulmonary fibrosis. Our findings suggest that dysregulated MUC5B expression in the lung may be involved in the pathogenesis of pulmonary fibrosis. (Funded by the National Heart, Lung, and Blood Institute and others.).

SFTPC Mutations in Patients with Familial Pulmonary Fibrosis: Combined with Emphysema?

<i>SFTPC</i> Mutations in Patients with Familial Pulmonary Fibrosis: Combined with Emphysema?

SFTPC Mutations in Patients with Familial Pulmonary Fibrosis: Combined with Emphysema?

<i>SFTPC</i> Mutations in Patients with Familial Pulmonary Fibrosis: Combined with Emphysema?

Ancestral Mutation in Telomerase Causes Defects in Repeat Addition Processivity and Manifests As Familial Pulmonary Fibrosis

The telomerase reverse transcriptase synthesizes new telomeres onto chromosome ends by copying from a short template within its integral RNA component. During telomere synthesis, telomerase adds multiple short DNA repeats successively, a property known as repeat addition processivity. However, the consequences of defects in processivity on telomere length maintenance are not fully known. Germline mutations in telomerase cause haploinsufficiency in syndromes of telomere shortening, which most commonly manifest in the age-related disease idiopathic pulmonary fibrosis. We identified two pulmonary fibrosis families that share two non-synonymous substitutions in the catalytic domain of the telomerase reverse transcriptase gene hTERT: V791I and V867M. The two variants fell on the same hTERT allele and were associated with telomere shortening. Genealogy suggested that the pedigrees shared a single ancestor from the nineteenth century, and genetic studies confirmed the two families had a common founder. Functional studies indicated that, although the double mutant did not dramatically affect first repeat addition, hTERT V791I-V867M showed severe defects in telomere repeat addition processivity in vitro. Our data identify an ancestral mutation in telomerase with a novel loss-of-function mechanism. They indicate that telomere repeat addition processivity is a critical determinant of telomere length and telomere-mediated disease.

Will the Genes Responsible for Familial Pulmonary Fibrosis Provide Clues to the Pathogenesis of IPF?

Will the Genes Responsible for Familial Pulmonary Fibrosis Provide Clues to the Pathogenesis of IPF?

TINF2 Mutations In Patients with Aplastic Anemia Result In Low TIN2 Expression In Hematopoietic Cells and Very Short Telomeres.

AbstractAbstract 1165Telomeres, the ends of chromosomes, protect against chromosomal instability. They are composed of hundreds to thousands TTAGGG double-stranded DNA repeats coated by at least six telomere-binding proteins collectively called “shelterin” (TRF1, TRF2, TIN2, TPP1, and POT1). As telomeres shorten at each cell division, highly proliferative cells express telomerase, a reverse transcriptase that elongates telomere ends. Loss-of-function mutations in genes encoding proteins and RNA of the telomerase complex are etiologic in dyskeratosis congenita (DC) and also are genetic risk factors for acquired aplastic anemia (AA), cancer, familial idiopathic pulmonary fibrosis, and liver diseases (cirrhosis and nodular regenerative hyperplasia). More recently, mutations in the shelterin component TIN2 were reported in patients with DC, further supporting that short and dysfunctional telomeres are the molecular etiology of marrow failure in dyskeratosis congenita. In order to investigate whether TINF2 mutations occurred in individuals with apparently acquired AA, we screened 190 consecutive patients diagnosed with the disease based on the International Agranulocytosis and Aplastic Anemia Study, all of whom were treated at a single institution (Hematology Branch, National Institutes of Health). All six TINF2 exons were bi-directionally sequenced using the Big Dye® sequencing method. We have observed eleven synonymous polymorphisms in TINF2 exons. Additionally four nonsynonymous TINF2 gene variants were identified in seven patients (carrier frequency, 3.7%). Three patients carried both codon Gly237Asp and codon Pro241Ser polymorphisms (one homozygous and two heterozygous); two patients carried codon Gly237Asp alone; one patient was heterozygous for a novel codon Gln120Arg TINF2 mutation; and one patient was heterozygous for a novel Thr275fs*codon316X TINF2 mutation. Patients carrying TINF2 nonsynonymous variants were significantly younger (median age, 13 years; range, 4–23 years vs. 36 years; 2–78 years; p=0.0015) but none had any signs of classic DC. Peripheral blood lymphocyte's telomere length measured by flow-fluorescence in situ hybridization was extremely short in the patient with a novel frameshift mutation (2.7 kb; expected-for-age length, 9.2 kb; below 1st percentile) and short in the patient with the novel codon Gln120Arg mutation (6.6 kb; expected-for-age, 8.0 kb; below the 10th percentile). Telomeres were short (below the 10th percentile) in three out of five patients carrying codon Gly237Asp and/or codon Pro241Ser variants. Hematopoietic cells of healthy volunteers and patients with TINF2 gene variants were probed for TIN2 expression in bone marrow samples by immunohistochemistry (polyclonal anti-TIN2 kindly provided by Dr. Titia de Lange, Rockefeller University). In normal bone marrows, TIN2 stained positive in megakaryocytes and in myeloid progenitors in a nuclear pattern. Mature granulocytes and lymphocytes were negative for TIN2 expression. Erythroid progenitors stained negative in the nucleus but some displayed a blushed positivity in the cytoplasm, which was demonstrated to be non-specific in immunoblot analyses of nuclear and cytoplasmic cell lysates. In the two patients with novel mutations, hematopoietic cells stained negative for TIN2. In patients carrying the two variants Gly237Asp and Pro241Ser, TIN2 stained weak or negative in hematopoietic cells, although it appeared in endothelial cells in the bone marrow. Patients with DKC1 or TERT mutations also were analyzed as controls and stained positive in the nucleus, similar to healthy controls, in spite of having very short telomeres. Our results describe for the first time the pattern of TIN2 expression in hematopoietic cells. TIN2 is positive with a nuclear pattern in megakaryocytes and early myeloid precursors, but not in erythroid progenitors or lymphocytes. The present findings further supports the relevance of the shelterin complex for telomere stability and for disease susceptibility; TINF2 mutations which result in short telomeres are present at relatively low frequencies in patients with apparently acquired AA but have important clinical management implications. TINF2 mutations are associated with reduced TIN2 expression in hematopoietic progenitors in the marrow. Disclosures:Lansdorp:Repeat Diagnostics: Equity Ownership.

Surfactant Protein C Mutations Are the Basis of a Significant Portion of Adult Familial Pulmonary Fibrosis in a Dutch Cohort

Familial clustering of adult idiopathic interstitial pneumonias (IIP) suggests that genetic factors might play an important role in disease development. Mutations in the gene encoding surfactant protein C (SFTPC) have been found in children and families with idiopathic pneumonias, whereas cocarriage of a mutation in ATP-binding cassette subfamily A member 3 (ABCA3) was postulated to have a disease-modifying effect. To investigate the contribution of SFTPC mutations to adult familial pulmonary fibrosis (FPF) and the disease-modifying effect of mutations in ABCA3 within their families. Twenty-two unrelated patients with FPF (10%) were identified within our single-center cohort of 229 patients with IIP. SFTPC was sequenced in 20 patients with FPF and 20 patients with sporadic IIP. In patients with an SFTPC mutation, sequencing of ABCA3 was performed. Discovered variants were typed in more than 100 control subjects and 121 additional patients with sporadic IIP. In 5/20 unrelated patients with FPF (25%; confidence interval, 10-49) a mutation in SFTPC was detected: M71V, IVS4+2, and three times I73T. No mutations were detected in the sporadic or control cohort. Patients with SFTPC mutations presented with a histopathological pattern of usual interstitial pneumonia and nodular septa thickening and multiple lung cysts in combination with ground glass or diffuse lung involvement on chest high-resolution computed tomography. Two variants in ABCA3 were found in adult patients with FPF but not in affected children. Mutations in SFTPC are a frequent cause of FPF in adult patients in our cohort. Nonclassifiable radiological patterns with cystic changes and histopathological patterns of usual interstitial pneumonia are characteristics of adult SFTPC mutation carriers.

Fibrose pulmonaire idiopathique familiale associée à une polyendocrinopathie auto-immune et à une épidermodysplasie verruciforme

Familial idiopathic pulmonary fibrosis (IPF) is a very rare and progressively fatal disease. Its pathogenesis is not fully understood and involves damage to alveolar epithelial cells of possibly immunological, microbiological or chemical origin, leading to fibrosing healing. A genetic predisposition has been demonstrated. The authors report the case of a female patient whose brother died at the age of 29 from IPF. She had epidermodysplasia verruciformis since childhood, with the absence of pubertal development. At the age of 31, she presented diffuse interstitial pneumonia. A lung biopsy confirmed the diagnosis of IPF. Endocrine explorations detected hypogonadotropic hypogonadism, primary hypothyroidism and magnetic resonance imaging revealed an empty sella turcica. The association of familial IPF, autoimmune polyendocrinopathy and genetic dermatosis caused by a cellular immune deficiency supports the hypothesis of an immune dysfunction in the pathogenesis of IPF.

Genetic Basis of Children's Interstitial Lung Disease

Specific genetic causes for children's interstitial lung disease (chILD) have been identified within the past decade. These include deletions of or mutations in genes encoding proteins important in surfactant production and function (SP-B, SP-C, and ABCA3), surfactant catabolism (GM-CSF receptor), as well as transcription factors important for surfactant production (TTF1) or lung development (Fox F1), with heterozygous deletions or loss-of-function mutations of the latter resulting in alveolar capillary dysplasia (ACD) with misalignment of the pulmonary veins. Familial pulmonary fibrosis in adults may result from mutations in genes encoding components of telomerase and SP-A2. While not yet reported in children, the expression of these genes in alveolar type II epithelial cells supports a key role for the disruption of normal homeostasis in this cell type in the pathogenesis of interstitial lung disease. The identification of specific genetic causes for chILD now allows for the possibility of non-invasive diagnosis, and provides insight into basic cellular mechanisms that may allow the development of novel therapies.

Familial Interstitial Pulmonary Fibrosis: A Large Family with Atypical Clinical Features

A large kindred of familial pulmonary fibrosis is reported. Six members from the first two generations of this particular kindred were described more than 40 years previously; six more individuals from the third and fourth generations have also been evaluated. The proband, now 23 years of age, has mild disease; the other 11 documented affected family members all died from their disease at an average age of 37 years (range 25 to 50 years). The pathology was that of usual interstitial pneumonia, as is typical in idiopathic pulmonary fibrosis. However, the initial radiographic pattern in many of these individuals was upper lobe and nodular and, along with the young age, was atypical for idiopathic pulmonary fibrosis. Several genetic abnormalities have been associated with familial pulmonary fibrosis. The present study examined the genes coding for surfactant protein-C, ATPbinding cassette protein A3 and telomerase, and found no abnormalities.

Erosion of Telomeric Single-Stranded Overhang in Patients with Aplastic Anemia Carrying Telomerase Mutations.

Abstract 1079Poster Board I-101Loss-of-function mutations in telomerase complex genes reduce telomerase activity and shorten overall telomere length in leukocytes, and they can clinically manifest as bone marrow failure (aplastic anemia and dyskeratosis congenita), familial pulmonary fibrosis, and hepatic cirrhosis. The double-stranded tandem telomeric TTAGGG repeats are followed by a 3' G-rich single-stranded overhang, a crucial structural component responsible for protective t-loop formation. We investigated the length of telomeric overhangs in 25 healthy individuals from 0 to 76 years, 16 patients with aplastic anemia, and 13 of their immediate relatives using a non-denaturing in-gel method and the telomere-oligonucleotide ligation assay (T-OLA). Among healthy controls, overall telomeric length of leukocytes shortened as a function of age. It was longest in umbilical cord blood samples and eroded as a function of a third-order polynomial until 76 years (R2=0.9517; Spearman=0.90; P<0.0001). However, the lengths of single-stranded overhangs were constant with age, as determined by both non-denaturing and the T-OLA methods. In contrast, most patients with marrow failure carrying a telomerase gene mutation showed marked erosion of telomeric overhangs associated with critically short telomeres (P<0.05). Seven telomerase complex-mutant aplastic patients had overhang lengths below the 5% percentile established for healthy volunteers. In aplastic patients with normal genotypes, normal overall telomere lengths, and who effectively responded to immunosuppressive therapy, telomeric overhangs were maintained and were all within the range established for control individuals. In conclusion, telomeric overhang erosion does not participate in physiological aging but eroded telomeric overhangs and abnormal telomere structure appear in pathologic shortening of telomeres, especially caused by loss-of-function telomerase mutations. Disrupted telomere structure caused by short telomeric overhangs may contribute to the mechanisms of abnormal hematopoietic compartment senescence and chromosomal instability in human bone marrow failure. Disclosures:No relevant conflicts of interest to declare.

Erosion of telomeric single‐stranded overhang in patients with aplastic anaemia carrying telomerase complex mutations

Loss-of-function mutations in telomerase complex genes reduce telomerase activity and shorten overall telomere length in leucocytes, and they can clinically manifest as bone marrow failure (aplastic anaemia and dyskeratosis congenita) and familial pulmonary fibrosis. Telomeres are constituted of double-stranded tandem TTAGGG repeats followed by a 3' G-rich single-stranded overhang, a crucial telomeric structural component responsible for the t-loop formation. We investigated the length of telomeric overhangs in 25 healthy individuals from 0 to 76 years of age, 16 patients with aplastic anaemia, and 13 immediate relatives using a non-denaturing in-gel method and the telomere-oligonucleotide ligation assay. Telomeric overhang lengths were constant from birth to eighth decade of life in healthy subjects, in contrast to overall telomere length, which shortened with ageing. Most patients with marrow failure and a telomerase gene mutation showed marked erosion of telomeric overhang associated with critically short telomeres; in other aplastic patients with normal genotypes, normal overall telomere lengths and who responded to immunosuppressive therapy, telomeric overhangs were maintained. Telomeric overhang erosion does not participate in physiological ageing but support a role for eroded telomeric overhangs and abnormal telomere structure in pathological shortening of telomeres, especially caused by loss-of-function telomerase mutations. Disrupted telomere structure caused by short telomeric overhangs may contribute to the mechanisms of abnormal haematopoietic compartment senescence and chromosomal instability in human bone marrow failure.

Clinical Year in Review I: Interstitial Lung Disease, Pulmonary Vascular Disease, Pulmonary Infections, and Cardiopulmonary Exercise Testing and Pulmonary Rehabilitation

Telomeres, regions of repetitive DNA at the end of chromosomes, shorten with cell division. Short telomeres limit tissue renewal capacity and are thought to play a role in diseases of aging. Telomerases are reverse transcriptase enzymes that add telomere repeats to chromosomes. Germ-line mutations in the components of telomerase (hTERT, hTR) have been associated with familial pulmonary fibrosis in some families (1). However, the role of telomerase mutations in idiopathic pulmonary fibrosis (IPF) is not known. Alder and colleagues (2) examined telomere lengths in leukocytes and alveolar epithelial cells in 64 patients with sporadic idiopathic interstitial pneumonia (IIP), also screening for mutations in hTERT and hTR, and reviewing cases for features of a telomere syndrome such as aplastic anemia or cryptogenic hepatic fibrosis. Telomere length was shorter in IIP cases (both IPF and nonIPF) than in age-matched control subjects, and this was mirrored by shortening of telomeres in leukocytes and alveolar epithelial cells that was of similar amplitude to that observed in familial IPF. A telomerase mutation was observed in only one case, although several individuals had a cluster of idiopathic pulmonary fibrosis and cryptogenic hepatic fibrosis, suggesting a telomere syndrome. These findings suggest that short telomeres are a signature of idiopathic interstitial pneumonia. The linkage between short telomeres and diseases related to aging supports the concept that IPF (and possibly other IIPs) can be viewed as a form of premature aging of the lung. Because known telomerase mutations did not account for short telomere length in this study, further studies are needed to identify the determinants of telomere shortening. The association of short telomere length with both IPF and other IIPs alike suggests the possibility of shared pathogenetic mechanisms between the IIPs. Diagnosis: Nonspecific Interstitial Pneumonia

Familial Pulmonary Fibrosis in 2 Mexican Sisters With Hermansky-Pudlak Syndrome

Hermansky-Pudlak syndrome is an autosomal recessive disorder commonly found in individuals of Puerto Rican ancestry. We present 2 cases of familial pulmonary fibrosis in 2 Mexican sisters with Hermansky-Pudlak syndrome. Pulmonary fibrosis was biopsy-proven in 1 of the patients. This report shows that Hermansky-Pudlak syndrome may occur in individuals of Mexican ancestry.

Prevalence of Telomere Shortening in Familial and Sporadic Pulmonary Fibrosis Is Increased in Men

Prevalence of Telomere Shortening in Familial and Sporadic Pulmonary Fibrosis Is Increased in Men

Fibrosis pulmonar familiar en 2 hermanas mexicanas con síndrome de Hermansky-Pudlak

Hermansky-Pudlak syndrome is an autosomal recessive disorder commonly found in individuals of Puerto Rican ancestry. We present 2 cases of familial pulmonary fibrosis in 2 Mexican sisters with Hermansky-Pudlak syndrome. Pulmonary fibrosis was biopsy-proven in 1 of the patients. This report shows that Hermansky-Pudlak syndrome may occur in individuals of Mexican ancestry.

Familial idiopathic pulmonary fibrosis in association with bone marrow hypoplasia and hepatic nodular regenerative hyperplasia: a new “trimorphic” syndrome

This is the first report of familial idiopathic pulmonary fibrosis associated with hepatic nodular regenerative hyperplasia and bone marrow hypoplasia. Four members of one family presented with this triad of...

Gene expression profiling: can we identify the right target genes?

Gene expression profiling allows the simultaneous monitoring of the transcriptional behaviour of thousands of genes, which may potentially be involved in disease development. Several studies have been performed in idiopathic pulmonary fibrosis (IPF), which aim to define genetic links to the disease in an attempt to improve the current understanding of the underlying pathogenesis of the disease and target pathways for intervention.Expression profiling has shown a clear difference in gene expression between IPF and normal lung tissue, and has identified a wide range of candidate genes, including those known to encode for proteins involved in extracellular matrix formation and degradation, growth factors and chemokines.Recently, familial pulmonary fibrosis cohorts have been examined in an attempt to detect specific genetic mutations associated with IPF. To date, these studies have identified families in which IPF is associated with mutations in the gene encoding surfactant protein C, or with mutations in genes encoding components of telomerase.Although rare and clearly not responsible for the disease in all individuals, the nature of these mutations highlight the importance of the alveolar epithelium in disease pathogenesis and demonstrate the potential for gene expression profiling in helping to advance the current understanding of idiopathic pulmonary fibrosis.

Telomere shortening in familial and sporadic pulmonary fibrosis.

Heterozygous mutations in the coding regions of the telomerase genes, TERT and TERC, have been found in familial and sporadic cases of idiopathic interstitial pneumonia. All affected patients with mutations have short telomeres. To test whether telomere shortening is a frequent mechanism underlying pulmonary fibrosis, we have characterized telomere lengths in subjects with familial or sporadic disease who do not have coding mutations in TERT or TERC. Using a modified Southern blot assay, the telomerase restriction fragment length method, and a quantitative polymerase chain reaction assay we have measured telomere lengths of genomic DNA isolated from circulating leukocytes from normal control subjects and subjects with pulmonary fibrosis. All affected patients with telomerase mutations, including case subjects heterozygous for newly reported mutations in TERT, have short telomere lengths. A significantly higher proportion of probands with familial pulmonary fibrosis (24%) and sporadic case subjects (23%) in which no coding mutation in TERT or TERC was found had telomere lengths less than the 10th percentile when compared with control subjects (P = 2.6 x 10(-8)). Pulmonary fibrosis affectation status was significantly associated with telomerase restriction fragment lengths, even after controlling for age, sex, and ethnicity (P = 6.1 x 10(-11)). Overall, 25% of sporadic cases and 37% of familial cases of pulmonary fibrosis had telomere lengths less than the 10th percentile. A significant fraction of individuals with pulmonary fibrosis have short telomere lengths that cannot be explained by coding mutations in telomerase. Telomere shortening of circulating leukocytes may be a marker for an increased predisposition toward the development of this age-associated disease.

MMP1 and MMP7 as Potential Peripheral Blood Biomarkers in Idiopathic Pulmonary Fibrosis

BackgroundIdiopathic pulmonary fibrosis (IPF) is a chronic progressive fibrotic lung disease associated with substantial morbidity and mortality. The objective of this study was to determine whether there is a peripheral blood protein signature in IPF and whether components of this signature may serve as biomarkers for disease presence and progression.Methods and FindingsWe analyzed the concentrations of 49 proteins in the plasma of 74 patients with IPF and in the plasma of 53 control individuals. We identified a combinatorial signature of five proteins—MMP7, MMP1, MMP8, IGFBP1, and TNFRSF1A—that was sufficient to distinguish patients from controls with a sensitivity of 98.6% (95% confidence interval [CI] 92.7%–100%) and specificity of 98.1% (95% CI 89.9%–100%). Increases in MMP1 and MMP7 were also observed in lung tissue and bronchoalveolar lavage fluid obtained from IPF patients. MMP7 and MMP1 plasma concentrations were not increased in patients with chronic obstructive pulmonary disease or sarcoidosis and distinguished IPF compared to subacute/chronic hypersensitivity pneumonitis, a disease that may mimic IPF, with a sensitivity of 96.3% (95% CI 81.0%–100%) and specificity of 87.2% (95% CI 72.6%–95.7%). We verified our results in an independent validation cohort composed of patients with IPF, familial pulmonary fibrosis, subclinical interstitial lung disease (ILD), as well as with control individuals. MMP7 and MMP1 concentrations were significantly higher in IPF patients compared to controls in this cohort. Furthermore, MMP7 concentrations were elevated in patients with subclinical ILD and negatively correlated with percent predicted forced vital capacity (FVC%) and percent predicted carbon monoxide diffusing capacity (DLCO%).ConclusionsOur experiments provide the first evidence for a peripheral blood protein signature in IPF to our knowledge. The two main components of this signature, MMP7 and MMP1, are overexpressed in the lung microenvironment and distinguish IPF from other chronic lung diseases. Additionally, increased MMP7 concentration may be indicative of asymptomatic ILD and reflect disease progression.