ALBERT

Description: Coccidioidomycosis (CM) is a fungal infection endemic in the southwestern United States (US). In California, CM incidence increased more than 213% (from 6.0/100,000 (2014) to 18.8/100,000 (2017)) and continues to increase as rates in the first half of 2018 are double that of 2017 during the same period. This cost-of-illness study provides essential information to be used in health planning and funding as CM infections continue to surge. We used a “bottom-up” approach to determine lifetime costs of 2017 reported incident CM cases in California. We defined CM natural history and used a societal approach to determine direct and discounted indirect costs using literature, national datasets, and expert interviews. The total lifetime cost burden of CM cases reported in 2017 in California is just under $700 million US dollars, with $429 million in direct costs and $271 million in indirect costs. Per person direct costs were highest for disseminated disease ($1,023,730), while per person direct costs were lowest for uncomplicated CM pneumonia ($22,039). Cost burden varied by county. This is the first study to estimate total costs of CM, demonstrating its huge cost burden for California.

Description: Abstract 415 Loss-of-function mutations in the telomerase complex genes, the enzyme responsible for maintaining the length of telomeres, are etiologic in patients with dyskeratosis congenita, an inherited bone marrow failure syndrome associated with mucocutaneous abnormalities, pulmonary fibrosis, esophageal stricture, liver cirrhosis, and an increased susceptibility to cancer, especially acute myeloid leukemia and squamous cell carcinoma. Heterozygous mutations abrogate telomerase's enzymatic ability to elongate telomeres by a mechanism of haploinsufficiency. Patients show accelerated telomere shortening in leukocytes, which eventually cause cellular senescence and apoptosis of hematopoietic precursors. Critically short telomeres lose their ability to sustain chromosomal stability, promoting end-to-end fusions and breakage-fusion-bridge cycles. Constitutional telomerase mutations also are found in patients with apparently acquired aplastic anemia, pulmonary fibrosis, and acute myeloid leukemia (AML). In AML, telomerase mutations correlate with cytogenetic abnormalities, especially trisomy 8 and inversion 16, and it appears that dysfunctional telomeres limit normal stem cell proliferation and predispose for leukemia cells by selection of stem cells with defective DNA damage responses that are prone to chromosomal instability. In the present study, we investigated whether mutations in genes for components of telomerase also occur in patients with myelodysplastic syndromes (MDS), and correlated mutation status with chromosomal changes. We screened DNA extracted from blood or bone marrow cells from 359 patients diagnosed with MDS and 528 healthy individuals for sequence variations in the telomerase reverse transcriptase (TERT) and telomerase RNA component (TERC) genes. Fourteen patients (4%) were heterozygous for telomerase gene variants in comparison to 7 healthy controls (1.3%; P=0.02, Fisher's exact test). Three mutant patients had refractory anemia; three had secondary AML; three had refractory anemia with ringed sideroblasts; two had chronic myelomonocytic leukemia; one had refractory anemia with excess blasts-2; one had refractory cytopenia with multilineage dysplasia; and one had MDS-unclassified. Vectors containing the TERT gene were mutagenized with novel gene variants and introduced into VA13 cells along with the TERC gene (VA13 cells are deficient for telomerase expression and maintain their telomeres via asymmetric telomeric recombination). Cell lysates from cells transfected with MDS-associated TERT showed reduced telomerase enzymatic activity in comparison to wild-type TERT. Using conventional karyotyping, six mutant patients had abnormal cytogenetics, and trisomy 8 was the most common cytogenetic abnormality observed in telomerase-mutant patients (three cases). Single nucleotide polymorphism array (SNP-array) analysis was performed in twelve telomerase-mutant patients (6.0 array in nine, and 250K array in three patients). Eleven of the twelve patients analyzed showed large lesions or copy number variations that have not been previously reported in normal individuals. More common were large losses in the long arm of the acrocentric chromosome 13, chromosome 7, and gains in the long arm of chromosome 3 and trisomy 8. One patient also showed somatic uniparental dissomy. Our results indicate that hypomorphic telomerase mutations may predispose to the development myelodysplastic syndromes. Myelodysplastic cells of patients carrying a telomerase mutation showed significant number of chromosomal abnormalities either by conventional cytogenetics or SNP-array in a proportion higher than that usually observed for MDS, suggesting increased chromosomal instability. In conclusion, short and dysfunctional telomeres in hematopoietic progenitors reduce telomere protection, compromising chromosomal stability and integrity, and our results indicate that these genomic changes may predispose for the selection of abnormal clones and the development of myelodysplastic syndromes. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 1165 Telomeres, 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.