Deleterious Germline Mutations in Telomere Maintenance Genes Identified in a Subset of Patients with Myelodysplastic Syndrome and Idiopathic Pulmonary Fibrosis

The myelodysplastic syndromes (MDS) are the most common cause of bone marrow failure in adults, with an incidence of 40,000 cases per year. Next-generation sequencing of candidate genes has led to major advances in the description of the genetic landscape of MDS, identifying recurrently mutated genes and cellular pathways involved in disease pathogenesis. However, use of targeted panels indicates more comprehensive, unbiased sequencing techniques may yet identify additional recurrently mutated genes or cellular pathways important in MDS. Diseases caused by defects in telomere maintenance (telomeropathies) are variable in clinical and genetic presentation but often involve bone marrow failure. We hypothesized that acquired mutations in telomerase maintenance genes may be a recurrent event in MDS. This is significant as identification of recurrent somatic mutations in telomerase maintenance genes would provide further insight into MDS pathogenesis and identify a potential druggable pathway for MDS patients, as novel agents targeting the telomerase pathway are currently in clinical development. First, we identified three adults who presented in middle age with MDS and idiopathic pulmonary fibrosis (IPF), two of which also had a family history of IPF. All three patients had shortened telomeres (<1^st centile), but no mutations detected in the telomerase pathway genes TERC, TERT, TINF2, or DKC2. In order to identify inherited and/or acquired genetic mutations affecting telomere biology, we performed exome sequencing on paired whole bone marrow (tumor) and skin (comparator/germline) samples from each patient. In patient 1 (see Table), we identified a heterozygous, inherited frameshift variant in RTEL1, (R132fs, not previously reported), a helicase required for telomere maintenance that has not been reported in MDS. Compound heterozygous mutations in RTEL1 have been implicated in Hoyeral-Hreiderson syndrome, and recent work showed ~5% of patients with familial IPF have heterozygous germline RTEL1 mutations (Cogan J.D. et al., Am J Resp Crit Care Med, 2015 and Stuart B.D. et al., Nat Genet, 2015). In patient 2, we identified a heterozygous inherited missense variant in ACD (V484I), a component of the telomerase shelterin complex also implicated in Hoyeral-Hreiderson syndrome that is not a common population variant (absent from 1000 genomes, the Exome Sequencing Project, and dbSNP). Analysis of constitutional alterations using the exome data from patient 3, did not identify an obvious causative variant; thus, whole genome sequencing is ongoing. These data show that inherited telomerase maintenance gene variants are found in patients with MDS and IPF. Next, to determine if telomerase complex genes were recurrently mutated in an unselected, de novo MDS population, we performed targeted next-generation sequencing of telomerase maintenance genes (15 genes) combined with sequencing of 335 myeloid malignancy-associated genes in 96 paired tumor/skin samples from MDS patients. Somatic variants were identified in the tumor compared to the paired skin using the Genome Modeling System, as previously reported (Griffith M. et al., PLoS Comput Biol. 2015). As expected, somatic analysis identified mutations in recurrently mutated MDS genes such as U2AF1, ASXL1, TET2, TP53 and others. A single, likely deleterious somatic inversion involving exon 9 of RTEL1 was identified; however, somatic analysis for telomere maintenance gene mutations was otherwise unremarkable. Evaluation of these genes for germline variants identified rare (ExAC allele frequency < 0.0007), apparently heterozygous, in-frame and frame-shift deletion variants affecting CTC1 (n=2), NOP10 (n=1) and TERT (n=1) in 4 individuals. The two CTC1 frame shift variants have been previously described in multiple patients with pediatric telomeropathies resulting from compound heterozygous mutations in CTC1. Additional heterozygous missense mutations previously associated with aplastic anemia were identified in 2 other individuals but are of uncertain significance. Collectively, these results indicate that germline mutations in telomere maintenance genes, other than TERT and TERC, underlie genetic predisposition for MDS and IPF in a subset of patients, which has not been previously appreciated, but are rare events in de novo MDS.

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