Telomere length is associated with disease severity and declines with age in dyskeratosis congenita

Abstract

Background: Dyskeratosis congenita is a cancer-prone bone marrow failure syndrome caused by aberrations in telomere biology.

Design and methods: We studied 65 patients with dyskeratosis congenita and 127 unaffected relatives. Telomere length was measured by automated multicolor flow fluorescence in situ hybridization in peripheral blood leukocyte subsets. We age-adjusted telomere length using Z-scores (standard deviations from the mean for age).

Results: We confirmed that telomere lengths below the first percentile for age are very sensitive and specific for the diagnosis of dyskeratosis congenita. We provide evidence that lymphocytes alone and not granulocytes may suffice for clinical screening, while lymphocyte subsets may be required for challenging cases, including identification of silent carriers. We show for the first time using flow fluorescence in situ hybridization that the shortest telomeres are associated with severe variants (Hoyeraal-Hreidarsson and Revesz syndromes), mutations in DKC1, TINF2, or unknown genes, and moderate or severe aplastic anemia. In the first longitudinal follow up of dyskeratosis congenita patients, we demonstrate that telomere lengths decline with age, in contrast to the apparent stable telomere length observed in cross-sectional data.

Conclusions: Telomere length by flow fluorescence in situ hybridization is an important diagnostic test for dyskeratosis congenita; age-adjusted values provide a quantitative measure of disease severity (clinical subset, mutated gene, and degree of bone marrow failure). Patients with dyskeratosis congenita have accelerated telomere shortening. This study is registered at www.clinicaltrials.gov (identifier: NCT00027274).

Figures

Figure 1

Telomere length according to age in patients with DC and their relatives. The vertical axis represents telomere length in kb. The curved lines in the figures indicate the 1st, 10th, 50th, 90th, and 99th percentiles of results from 400 normal controls. Colored symbols represent patients with DC and their relatives. Red circles: classical DC patients; green triangles: Hoyeraal-Hreidarsson; black diamonds: revesz syndrome; blue squares: silent carriers; open black squares: DC relatives in families with unknown genes; open black triangles: DC relatives without mutations in the probands’ genes. Top panels show granulocytes, lymphocytes, and CD45RA-positive/CD20-negative naïve T cells. Bottom panels show CD45RA-negative memory T cells, CD20-positive B cells, and total NK/NKT cells.

Figure 2

Telomere length Z-scores in DC patients according to various categories, and relatives. (A) Typical DC patients, silent carriers with mutations in DC genes, Hoyeraal-Hreidarsson (HH), and Revesz Syndrome (RS). Telomeres for those with HH and RS are much shorter than in other DC patients and silent carriers after age-adjustment by Z-scores; P=0.002 by rank sum. All DC patients have lower Z-scores when compared with relatives, P<0.0001 by Student’s t-test. Outliers among relatives with low Z-scores may include silent carriers of DC in families in which genes have not been identified. Telomere Z-scores in relatives in families without known genes are lower than in those in families with known mutations, P=0.02 by Student’s t-test. (B) Telomere Z-scores are lower in those with mutations in unknown genes, DKC1, and TINF2 than in those with mutations in TERC, TERT, and WRAP53; P=0.01 by rank sum. (C) The Z-scores are lower in those with increased severity of bone marrow failure (MAA: moderate aplastic anemia; SAA: severe aplastic anemia); P for trend=0.001. Z-scores in patients without aplastic anemia are similar to those with MAA, P=0.1; SAA are lower than MAA, P=0.05; and SAA are much lower than those with no AA, P<0.001. (D) Telomere Z-scores are not significantly different in those with two or three features of the clinical triad than in those with none or only one such feature; P=0.08.

Figure 3

Regression lines for telomere length according to age. The short lines indicate the change in telomere lengths in 9 DC patients over 5-year intervals. Red: NCI-6-1 and NCI-6-2 are mother and son with a mutation in TERC; Blue: NCI-26-1 and NCI-26-2 are identical twins with a mutation in TERT; Teal blue: NCI-74-3 and NCI-74-5 are brothers with a mutation in TINF2; Green: NCI-106-1 and NCI-106-3 are brothers with a mutation in DKC1; Black: NCI-114-1 with a mutation in TERC. The slope of the cross-sectional data for telomere length vs. age for the DC patients (_ _ _) is not significantly different from 0 in any lineage. In contrast, the slope for the cross-sectional results in DC relatives (_ _ _) is significantly negative (P<0.001) in all lineages.