Adaptive and Innate Immunity Are Key Drivers of Age at Onset of Multiple Sclerosis.

Background and Objectives: Multiple sclerosis (MS) age at onset (AAO) is a clinical predictor of long-term disease outcomes, independent of disease duration. Little is known about the genetic and biological mechanisms underlying age of first symptoms. We conducted a genome-wide association study (GWAS) to investigate associations between individual genetic variation and the MS AAO phenotype. Methods: The study population was comprised participants with MS in 6 clinical trials: ADVANCE (N = 655; relapsing-remitting [RR] MS), ASCEND (N = 555; secondary-progressive [SP] MS), DECIDE (N = 1,017; RRMS), OPERA1 (N = 581; RRMS), OPERA2 (N = 577; RRMS), and ORATORIO (N = 529; primary-progressive [PP] MS). Altogether, 3,905 persons with MS of European ancestry were analyzed. GWAS were conducted for MS AAO in each trial using linear additive models controlling for sex and 10 principal components. Resultant summary statistics across the 6 trials were then meta-analyzed, for a total of 8.3 × 10-6 single nucleotide polymorphisms (SNPs) across all trials after quality control and filtering for heterogeneity. Gene-based tests of associations, pathway enrichment analyses, and Mendelian randomization analyses for select exposures were also performed. Results: Four lead SNPs within 2 loci were identified (p < 5 × 10-8), including a) 3 SNPs in the major histocompatibility complex and their effects were independent of HLA-DRB1*15:01 and b) a LOC105375167 variant on chromosome 7. At the gene level, the top association was HLA-C (p = 1.2 × 10-7), which plays an important role in antiviral immunity. Functional annotation revealed the enrichment of pathways related to T-cell receptor signaling, autoimmunity, and the complement cascade. Mendelian randomization analyses suggested a link between both earlier age at puberty and shorter telomere length and earlier AAO, while there was no evidence for a role for either body mass index or vitamin D levels. Discussion: Two genetic loci associated with MS AAO were identified, and functional annotation demonstrated an enrichment of genes involved in adaptive and complement immunity. There was also evidence supporting a link with age at puberty and telomere length. The findings suggest that AAO in MS is multifactorial, and the factors driving onset of symptoms overlap with those influencing MS risk.

Genome-wide study of longitudinal brain imaging measures of multiple sclerosis progression across six clinical trials.

While the genetics of MS risk susceptibility are well-described, and recent progress has been made on the genetics of disease severity, the genetics of disease progression remain elusive. We therefore investigated the genetic determinants of MS progression on longitudinal brain MRI: change in brain volume (BV) and change in T2 lesion volume (T2LV), reflecting progressive tissue loss and increasing disease burden, respectively. We performed genome-wide association studies of change in BV (N = 3401) and change in T2LV (N = 3513) across six randomized clinical trials from Biogen and Roche/Genentech: ADVANCE, ASCEND, DECIDE, OPERA I & II, and ORATORIO. Analyses were adjusted for randomized treatment arm, age, sex, and ancestry. Results were pooled in a meta-analysis, and were evaluated for enrichment of MS risk variants. Variant colocalization and cell-specific expression analyses were performed using published cohorts. The strongest peaks were in PTPRD (rs77321193-C/A, p = 3.9 × 10-7) for BV change, and NEDD4L (rs11398377-GC/G, p = 9.3 × 10-8) for T2LV change. Evidence of colocalization was observed for NEDD4L, and both genes showed increased expression in neuronal and/or glial populations. No association between MS risk variants and MRI outcomes was observed. In this unique, precompetitive industry partnership, we report putative regions of interest in the neurodevelopmental gene PTPRD, and the ubiquitin ligase gene NEDD4L. These findings are distinct from known MS risk genetics, indicating an added role for genetic progression analyses and informing drug discovery.

South Asian medical cohorts reveal strong founder effects and high rates of homozygosity.

The benefits of large-scale genetic studies for healthcare of the populations studied are well documented, but these genetic studies have traditionally ignored people from some parts of the world, such as South Asia. Here we describe whole genome sequence (WGS) data from 4806 individuals recruited from the healthcare delivery systems of Pakistan, India and Bangladesh, combined with WGS from 927 individuals from isolated South Asian populations. We characterize population structure in South Asia and describe a genotyping array (SARGAM) and imputation reference panel that are optimized for South Asian genomes. We find evidence for high rates of reproductive isolation, endogamy and consanguinity that vary across the subcontinent and that lead to levels of rare homozygotes that reach 100 times that seen in outbred populations. Founder effects increase the power to associate functional variants with disease processes and make South Asia a uniquely powerful place for population-scale genetic studies.

Whole genome sequencing across clinical trials identifies rare coding variants in GPR68 associated with chemotherapy-induced peripheral neuropathy.

BACKGROUND: Dose-limiting toxicities significantly impact the benefit/risk profile of many drugs. Whole genome sequencing (WGS) in patients receiving drugs with dose-limiting toxicities can identify therapeutic hypotheses to prevent these toxicities. Chemotherapy-induced peripheral neuropathy (CIPN) is a common dose-limiting neurological toxicity of chemotherapies with no effective approach for prevention. METHODS: We conducted a genetic study of time-to-first peripheral neuropathy event using 30× germline WGS data from whole blood samples from 4900 European-ancestry cancer patients in 14 randomized controlled trials. A substantial number of patients in these trials received taxane and platinum-based chemotherapies as part of their treatment regimen, either standard of care or in combination with the PD-L1 inhibitor atezolizumab. The trials spanned several cancers including renal cell carcinoma, triple negative breast cancer, non-small cell lung cancer, small cell lung cancer, bladder cancer, ovarian cancer, and melanoma. RESULTS: We identified a locus consisting of low-frequency variants in intron 13 of GRID2 associated with time-to-onset of first peripheral neuropathy (PN) indexed by rs17020773 (p = 2.03 × 10-8, all patients, p = 6.36 × 10-9, taxane treated). Gene-level burden analysis identified rare coding variants associated with increased PN risk in the C-terminus of GPR68 (p = 1.59 × 10-6, all patients, p = 3.47 × 10-8, taxane treated), a pH-sensitive G-protein coupled receptor (GPCR). The variants driving this signal were found to alter predicted arrestin binding motifs in the C-terminus of GPR68. Analysis of snRNA-seq from human dorsal root ganglia (DRG) indicated that expression of GPR68 was highest in mechano-thermo-sensitive nociceptors. CONCLUSIONS: Our genetic study provides insight into the impact of low-frequency and rare coding genetic variation on PN risk and suggests that further study of GPR68 in sensory neurons may yield a therapeutic hypothesis for prevention of CIPN.

Large-scale rare variant burden testing in Parkinson's disease.

Parkinson's disease has a large heritable component and genome-wide association studies have identified over 90 variants with disease-associated common variants, providing deeper insights into the disease biology. However, there have not been large-scale rare variant analyses for Parkinson's disease. To address this gap, we investigated the rare genetic component of Parkinson's disease at minor allele frequencies <1%, using whole genome and whole exome sequencing data from 7184 Parkinson's disease cases, 6701 proxy cases and 51 650 healthy controls from the Accelerating Medicines Partnership Parkinson's disease (AMP-PD) initiative, the National Institutes of Health, the UK Biobank and Genentech. We performed burden tests meta-analyses on small indels and single nucleotide protein-altering variants, prioritized based on their predicted functional impact. Our work identified several genes reaching exome-wide significance. Two of these genes, GBA1 and LRRK2, have variants that have been previously implicated as risk factors for Parkinson's disease, with some variants in LRRK2 resulting in monogenic forms of the disease. We identify potential novel risk associations for variants in B3GNT3, AUNIP, ADH5, TUBA1B, OR1G1, CAPN10 and TREML1 but were unable to replicate the observed associations across independent datasets. Of these, B3GNT3 and TREML1 could provide new evidence for the role of neuroinflammation in Parkinson's disease. To date, this is the largest analysis of rare genetic variants in Parkinson's disease.

Inhibiting membrane rupture with NINJ1 antibodies limits tissue injury.

Plasma membrane rupture (PMR) in dying cells undergoing pyroptosis or apoptosis requires the cell-surface protein NINJ11. PMR releases pro-inflammatory cytoplasmic molecules, collectively called damage-associated molecular patterns (DAMPs), that activate immune cells. Therefore, inhibiting NINJ1 and PMR may limit the inflammation that is associated with excessive cell death. Here we describe an anti-NINJ1 monoclonal antibody that specifically targets mouse NINJ1 and blocks oligomerization of NINJ1, preventing PMR. Electron microscopy studies showed that this antibody prevents NINJ1 from forming oligomeric filaments. In mice, inhibition of NINJ1 or Ninj1 deficiency ameliorated hepatocellular PMR induced with TNF plus D-galactosamine, concanavalin A, Jo2 anti-Fas agonist antibody or ischaemia-reperfusion injury. Accordingly, serum levels of lactate dehydrogenase, the liver enzymes alanine aminotransaminase and aspartate aminotransferase, and the DAMPs interleukin 18 and HMGB1 were reduced. Moreover, in the liver ischaemia-reperfusion injury model, there was an attendant reduction in neutrophil infiltration. These data indicate that NINJ1 mediates PMR and inflammation in diseases driven by aberrant hepatocellular death.

A whole genome sequencing study of moderate to severe asthma identifies a lung function locus associated with asthma risk.

Genome-wide association studies (GWAS) have identified many common variant loci associated with asthma susceptibility, but few studies investigate the genetics underlying moderate-to-severe asthma risk. Here, we present a whole-genome sequencing study comparing 3181 moderate-to-severe asthma patients to 3590 non-asthma controls. We demonstrate that asthma risk is genetically correlated with lung function measures and that this component of asthma risk is orthogonal to the eosinophil genetics that also contribute to disease susceptibility. We find that polygenic scores for reduced lung function are associated with younger asthma age of onset. Genome-wide, seven previously reported common asthma variant loci and one previously reported lung function locus, near THSD4, reach significance. We replicate association of the lung function locus in a recently published GWAS of moderate-to-severe asthma patients. We additionally replicate the association of a previously reported rare (minor allele frequency < 1%) coding variant in IL33 and show significant enrichment of rare variant burden in genes from common variant allergic disease loci. Our findings highlight the contribution of lung function genetics to moderate-to-severe asthma risk, and provide initial rare variant support for associations with moderate-to-severe asthma risk at several candidate genes from common variant loci.

Genome-wide association study and functional validation implicates JADE1 in tauopathy.

Primary age-related tauopathy (PART) is a neurodegenerative pathology with features distinct from but also overlapping with Alzheimer disease (AD). While both exhibit Alzheimer-type temporal lobe neurofibrillary degeneration alongside amnestic cognitive impairment, PART develops independently of amyloid-ß (Aß) plaques. The pathogenesis of PART is not known, but evidence suggests an association with genes that promote tau pathology and others that protect from Aß toxicity. Here, we performed a genetic association study in an autopsy cohort of individuals with PART (n = 647) using Braak neurofibrillary tangle stage as a quantitative trait. We found some significant associations with candidate loci associated with AD (SLC24A4, MS4A6A, HS3ST1) and progressive supranuclear palsy (MAPT and EIF2AK3). Genome-wide association analysis revealed a novel significant association with a single nucleotide polymorphism on chromosome 4 (rs56405341) in a locus containing three genes, including JADE1 which was significantly upregulated in tangle-bearing neurons by single-soma RNA-seq. Immunohistochemical studies using antisera targeting JADE1 protein revealed localization within tau aggregates in autopsy brains with four microtubule-binding domain repeats (4R) isoforms and mixed 3R/4R, but not with 3R exclusively. Co-immunoprecipitation in post-mortem human PART brain tissue revealed a specific binding of JADE1 protein to four repeat tau lacking N-terminal inserts (0N4R). Finally, knockdown of the Drosophila JADE1 homolog rhinoceros (rno) enhanced tau-induced toxicity and apoptosis in vivo in a humanized 0N4R mutant tau knock-in model, as quantified by rough eye phenotype and terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) in the fly brain. Together, these findings indicate that PART has a genetic architecture that partially overlaps with AD and other tauopathies and suggests a novel role for JADE1 as a modifier of neurofibrillary degeneration.

Genetic ablation of Gpnmb does not alter synuclein-related pathology.

The gene GPNMB is known to play roles in phagocytosis and tissue repair, and is upregulated in microglia in many mouse models of neurodegenerative disease as well as in human patients. Nearby genomic variants are associated with both elevated Parkinson's disease (PD) risk and higher expression of this gene, suggesting that inhibiting GPNMB activity might be protective in Parkinson's disease. We tested this hypothesis in three different mouse models of neurological diseases: a remyelination model and two models of alpha-synuclein pathology. We found that Gpnmb deletion had no effect on histological, cellular, behavioral, neurochemical or gene expression phenotypes in any of these models. These data suggest that Gpnmb does not play a major role in the development of pathology or functional defects in these models and that further work is necessary to study its role in the development or progression of Parkinson's disease.

Genetic variation associated with thyroid autoimmunity shapes the systemic immune response to PD-1 checkpoint blockade.

Activation of systemic immune responses using PD-1 checkpoint inhibitors is an essential approach to cancer therapy. Yet, the extent of benefit relative to risk of immune related adverse events (irAE) varies widely among patients. Here, we study endocrine irAE from 7 clinical trials across 6 cancers where atezolizumab (anti-PD-L1) was combined with chemotherapies and compared to standard of care. We show that atezolizumab-induced thyroid dysfunction is associated with longer survival. We construct a polygenic risk score (PRS) for lifetime risk of hypothyroidism using a GWAS from the UK Biobank and apply this PRS to genetic data collected from 2,616 patients of European ancestry from these trials. Patients with high PRS are at increased risk of atezolizumab-induced thyroid dysfunction and lower risk of death in triple negative breast cancer. Our results indicate that genetic variation associated with thyroid autoimmunity interacts with biological pathways driving the systemic immune response to PD-1 blockade.

Polygenic risk for skin autoimmunity impacts immune checkpoint blockade in bladder cancer.

PD-1 and PD-L1 act to restrict T cell responses in cancer and contribute to self-tolerance. Consistent with this role, PD-1 checkpoint inhibitors have been associated with immune-related adverse events (irAEs), immune toxicities thought to be autoimmune in origin. Analyses of dermatological irAEs have identified an association with improved overall survival (OS) following anti-PD-(L)1 therapy, but the factors that contribute to this relationship are poorly understood. We collected germline whole-genome sequencing data from IMvigor211, a recent phase 3 randomized controlled trial comparing atezolizumab (anti-PD-L1) monotherapy to chemotherapy in bladder cancer. We found that high vitiligo, high psoriasis, and low atopic dermatitis polygenic risk scores (PRSs) were associated with longer OS under anti-PD-L1 monotherapy as compared to chemotherapy, reflecting the Th17 polarization of these diseases. PRSs were not correlated with tumor mutation burden, PD-L1 immunohistochemistry, nor T-effector gene signatures. Shared genetic factors impact risk for dermatological autoimmunity and anti-PD-L1 monotherapy in bladder cancer.

Integration of eQTL and a Single-Cell Atlas in the Human Eye Identifies Causal Genes for Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) is a leading cause of vision loss. To better understand disease pathogenesis and identify causal genes in GWAS loci for AMD risk, we present a comprehensive database of human retina and retinal pigment epithelium (RPE). Our database comprises macular and non-macular RNA sequencing (RNA-seq) profiles from 129 donors, a genome-wide expression quantitative trait loci (eQTL) dataset that includes macula-specific retina and RPE/choroid, and single-nucleus RNA-seq (NucSeq) from human retina and RPE with subtype resolution from more than 100,000 cells. Using NucSeq, we find enriched expression of AMD candidate genes in RPE cells. We identify 15 putative causal genes for AMD on the basis of co-localization of genetic association signals for AMD risk and eye eQTL, including the genes TSPAN10 and TRPM1. These results demonstrate the value of our human eye database for elucidating genetic pathways and potential therapeutic targets for ocular diseases.

PTCD1 Is Required for Mitochondrial Oxidative-Phosphorylation: Possible Genetic Association with Alzheimer's Disease.

In addition to amyloid-ß plaques and tau tangles, mitochondrial dysfunction is implicated in the pathology of Alzheimer's disease (AD). Neurons heavily rely on mitochondrial function, and deficits in brain energy metabolism are detected early in AD; however, direct human genetic evidence for mitochondrial involvement in AD pathogenesis is limited. We analyzed whole-exome sequencing data of 4549 AD cases and 3332 age-matched controls and discovered that rare protein altering variants in the gene pentatricopeptide repeat-containing protein 1 (PTCD1) show a trend for enrichment in cases compared with controls. We show here that PTCD1 is required for normal mitochondrial rRNA levels, proper assembly of the mitochondrial ribosome and hence for mitochondrial translation and assembly of the electron transport chain. Loss of PTCD1 function impairs oxidative phosphorylation and forces cells to rely on glycolysis for energy production. Cells expressing the AD-linked variant of PTCD1 fail to sustain energy production under increased metabolic stress. In neurons, reduced PTCD1 expression leads to lower ATP levels and impacts spontaneous synaptic activity. Thus, our study uncovers a possible link between a protein required for mitochondrial function and energy metabolism and AD risk.SIGNIFICANCE STATEMENT Mitochondria are the main source of cellular energy and mitochondrial dysfunction is implicated in the pathology of Alzheimer's disease (AD) and other neurodegenerative disorders. Here, we identify a variant in the gene PTCD1 that is enriched in AD patients and demonstrate that PTCD1 is required for ATP generation through oxidative phosphorylation. PTCD1 regulates the level of 16S rRNA, the backbone of the mitoribosome, and is essential for mitochondrial translation and assembly of the electron transport chain. Cells expressing the AD-associated variant fail to maintain adequate ATP production during metabolic stress, and reduced PTCD1 activity disrupts neuronal energy homeostasis and dampens spontaneous transmission. Our work provides a mechanistic link between a protein required for mitochondrial function and genetic AD risk.

Previously reported placebo-response-associated variants do not predict patient outcomes in inflammatory disease Phase III trial placebo arms.

In clinical trials, a placebo response refers to improvement in disease symptoms arising from the psychological effect of receiving a treatment rather than the actual treatment under investigation. Previous research has reported genomic variation associated with the likelihood of observing a placebo response, but these studies have been limited in scope and have not been validated. Here, we analyzed whole-genome sequencing data from 784 patients undergoing placebo treatment in Phase III Asthma or Rheumatoid Arthritis trials to assess the impact of previously reported variation on patient outcomes in the placebo arms and to identify novel variants associated with the placebo response. Contrary to expectations based on previous reports, we did not observe any statistically significant associations between genomic variants and placebo treatment outcome. Our findings suggest that the biological origin of the placebo response is complex and likely to be variable between disease areas.

Enabling genome-wide association testing with multiple diseases and no healthy controls.

MOTIVATION: While large-scale whole genome sequencing is feasible the high costs compel investigators to focus on disease subjects. As a result large sequencing datasets of samples with different diseases are often readily available, but not healthy controls to contrast them with. While it is possible to perform an association study using only diseases, the associations could be driven by a disease acting as a control and not the focal disease. METHODS: We developed a genotype-on-phenotype reverse regression with a Bayesian spike and slab prior to enable association testing in datasets with multiple diseases. This method, referred to as revreg, flagged associations (both common and rare) that were driven by diseases that were not of primary interest. RESULTS: Based on simulations, revreg had 80% power to detect an odds ratio of 1.74 for common variants (3500 samples total) and 3.73 for rare variants (14,000 samples total), with minimal type I error. For common variants, we tested this method on 3657 whole genome sequenced samples aimed at discovering variants associated with disease risk of Chronic Obstructive Pulmonary Disease using three other diseases as controls. We demonstrated detection of six highly significant associations likely due to Age-Related Macular Degeneration. In an exome dataset of 8836 samples aimed at characterizing rare variants associated with disease risk of Asthma, using five other diseases as controls, we detected and removed genic regions due to AMD (C3, CFH, CFHR5, CFI, and DNMT3A) and RA (KRTAP13-4).

Germline genetic polymorphisms influence tumor gene expression and immune cell infiltration.

Cancer immunotherapy has emerged as an effective therapy in a variety of cancers. However, a key challenge in the field is that only a subset of patients who receive immunotherapy exhibit durable response. It has been hypothesized that host genetics influences the inherent immune profiles of patients and may underlie their differential response to immunotherapy. Herein, we systematically determined the association of common germline genetic variants with gene expression and immune cell infiltration of the tumor. We identified 64,094 expression quantitative trait loci (eQTLs) that associated with 18,210 genes (eGenes) across 24 human cancers. Overall, eGenes were enriched for their being involved in immune processes, suggesting that expression of immune genes can be shaped by hereditary genetic variants. We identified the endoplasmic reticulum aminopeptidase 2 (ERAP2) gene as a pan-cancer type eGene whose expression levels stratified overall survival in a subset of patients with bladder cancer receiving anti-PD-L1 (atezolizumab) therapy. Finally, we identified 103 gene signature QTLs (gsQTLs) that were associated with predicted immune cell abundance within the tumor microenvironment. Our findings highlight the impact of germline SNPs on cancer-immune phenotypes and response to therapy; and these analyses provide a resource for integration of germline genetics as a component of personalized cancer immunotherapy.

Paired Immunoglobulin-like Type 2 Receptor Alpha G78R variant alters ligand binding and confers protection to Alzheimer's disease.

Paired Immunoglobulin-like Type 2 Receptor Alpha (PILRA) is a cell surface inhibitory receptor that recognizes specific O-glycosylated proteins and is expressed on various innate immune cell types including microglia. We show here that a common missense variant (G78R, rs1859788) of PILRA is the likely causal allele for the confirmed Alzheimer's disease risk locus at 7q21 (rs1476679). The G78R variant alters the interaction of residues essential for sialic acid engagement, resulting in >50% reduced binding for several PILRA ligands including a novel ligand, complement component 4A, and herpes simplex virus 1 (HSV-1) glycoprotein B. PILRA is an entry receptor for HSV-1 via glycoprotein B, and macrophages derived from R78 homozygous donors showed significantly decreased levels of HSV-1 infection at several multiplicities of infection compared to homozygous G78 macrophages. We propose that PILRA G78R protects individuals from Alzheimer's disease risk via reduced inhibitory signaling in microglia and reduced microglial infection during HSV-1 recurrence.

Genetic analysis of isoform usage in the human anti-viral response reveals influenza-specific regulation of ERAP2 transcripts under balancing selection.

While genetic variants are known to be associated with overall gene abundance in stimulated immune cells, less is known about their effects on alternative isoform usage. By analyzing RNA-seq profiles of monocyte-derived dendritic cells from 243 individuals, we uncovered thousands of unannotated isoforms synthesized in response to influenza infection and type 1 interferon stimulation. We identified more than a thousand quantitative trait loci (QTLs) associated with alternate isoform usage (isoQTLs), many of which are independent of expression QTLs (eQTLs) for the same gene. Compared with eQTLs, isoQTLs are enriched for splice sites and untranslated regions, but depleted of sequences upstream of annotated transcription start sites. Both eQTLs and isoQTLs explain a significant proportion of the disease heritability attributed to common genetic variants. At the ERAP2 locus, we shed light on the function of the gene and how two frequent, highly differentiated haplotypes with intermediate frequencies could be maintained by balancing selection. At baseline and following type 1 interferon stimulation, the major haplotype is associated with low ERAP2 expression caused by nonsense-mediated decay, while the minor haplotype, known to increase Crohn's disease risk, is associated with high ERAP2 expression. In response to influenza infection, we found two uncharacterized isoforms expressed from the major haplotype, likely the result of multiple perfectly linked variants affecting the transcription and splicing at the locus. Thus, genetic variants at a single locus could modulate independent gene regulatory processes in innate immune responses and, in the case of ERAP2, may confer a historical fitness advantage in response to virus.

Analysis of protein-altering variants in telomerase genes and their association with MUC5B common variant status in patients with idiopathic pulmonary fibrosis: a candidate gene sequencing study.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) risk has a strong genetic component. Studies have implicated variations at several loci, including TERT, surfactant genes, and a single nucleotide polymorphism at chr11p15 (rs35705950) in the intergenic region between TOLLIP and MUC5B. Patients with IPF who have risk alleles at rs35705950 have longer survival from the time of IPF diagnosis than do patients homozygous for the non-risk allele, whereas patients with shorter telomeres have shorter survival times. We aimed to assess whether rare protein-altering variants in genes regulating telomere length are enriched in patients with IPF homozygous for the non-risk alleles at rs35705950. METHODS: Between Nov 1, 2014, and Nov 1, 2016, we assessed blood samples from patients aged 40 years or older and of European ancestry with sporadic IPF from three international phase 3 clinical trials (INSPIRE, CAPACITY, ASCEND), one phase 2 study (RIFF), and US-based observational studies (Vanderbilt Clinical Interstitial Lung Disease Registry and the UCSF Interstitial Lung Disease Clinic registry cohorts) at the Broad Institute (Cambridge, MA, USA) and Human Longevity (San Diego, CA, USA). We also assessed blood samples from non-IPF controls in several clinical trials. We did whole-genome sequencing to assess telomere length and identify rare protein-altering variants, stratified by rs35705950 genotype. We also assessed rare functional variation in TERT exons and compared telomere length and disease progression across genotypes. FINDINGS: We assessed samples from 1510 patients with IPF and 1874 non-IPF controls. 30 (3%) of 1046 patients with an rs35705950 risk allele had a rare protein-altering variant in TERT compared with 34 (7%) of 464 non-risk allele carriers (odds ratio 0·40 [95% CI 0·24-0·66], p=0·00039). Subsequent analyses identified enrichment of rare protein-altering variants in PARN and RTEL1, and rare variation in TERC in patients with IPF compared with controls. We expanded our study population to provide a more accurate estimation of rare variant frequency in these four loci, and to calculate telomere length. The proportion of patients with at least one rare variant in TERT, PARN, TERC, or RTEL1 was higher in patients with IPF than in controls (149 [9%] of 1739 patients vs 205 [2%] of 8645 controls, p=2·44 × 10-8). Patients with IPF who had a variant in any of the four identified telomerase component genes had telomeres that were 3·69-16·10% shorter than patients without a variant in any of the four genes and had an earlier mean age of disease onset than patients without one or more variants (65·1 years [SD 7·8] vs 67·1 years [7·9], p=0·004). In the placebo arms of clinical trials, shorter telomeres were significantly associated with faster disease progression (1·7% predicted forced vital capacity per kb per year, p=0·002). Pirfenidone had treatment benefit regardless of telomere length (p=4·24 × 10-8 for telomere length lower than the median, p=0·0044 for telomere length greater than the median). INTERPRETATION: Rare protein-altering variants in TERT, PARN, TERC, and RTEL1 are enriched in patients with IPF compared with controls, and, in the case of TERT, particularly in individuals without a risk allele at the rs35705950 locus. This suggests that multiple genetic factors contribute to sporadic IPF, which might implicate distinct mechanisms of pathogenesis and disease progression. FUNDING: Genentech, National Institutes of Health, Francis Family Foundation, Pulmonary Fibrosis Foundation, Nina Ireland Program for Lung Health, US Department of Veterans Affairs.