Cytogenetic differentiation of Fanconi anemia, “idiopathic” aplastic anemia, and Fanconi anemia heterozygotes

Cervenka, J; Ba, Hirsch

doi:10.1002/ajmg.1320150205

Cited by 72 publications

(54 citation statements)

References 19 publications

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“…Chromosomal study using Mitomycin-C (MMC) was done to diagnose patients of Fanconi anemia. [5] Two milliliters of venous blood was collected from cases and healthy controls. Five to six drops of heparinized blood (0.1 mL of heparin/2 mL of blood) was mixed with growth media and incubated in tissue culture fl ask at 37°C for 96 h. Mitomycin-C was added in concentration of 40 ng/mL of media for the duration of culture.…”

Section: Methodsmentioning

confidence: 99%

A study of bone marrow failure syndrome in children

et al. 2008

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Section: Methodsmentioning

confidence: 99%

A study of bone marrow failure syndrome in children

et al. 2008

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“…28 A diagnosis of FA was confirmed in the 29 patient-derived fibroblasts cell lines by cytogenetic analysis of sensitivity to MMC (Sigma-Aldrich, St Louis, MO) and DEB (SigmaAldrich) as previously described (Table 1). 4,57,58 Briefly, approximately 50 Wright-stained metaphase spreads from each of the following cultures were assayed for chromosomal breakage and radial formation: (1) patient-derived DEB (multiple concentrations) and MMC-(multiple concentrations) treated fibroblasts and (2) untreated patient-derived fibroblasts. A diagnosis of FA was confirmed if Ն20% of fibroblasts demonstrated radials.…”

Section: Cytogenetic Analysismentioning

confidence: 99%

Validation of Fanconi anemia complementation Group A assignment using molecular analysis

Moghrabi¹,

Johnson²,

Yoshitomi³

et al. 2009

Genetics in Medicine

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Purpose: Fanconi anemia is a genetically heterogeneous chromosomal breakage disorder exhibiting a high degree of clinical variability. Clinical diagnoses are confirmed by testing patient cells for increased sensitivity to crosslinking agents. Fanconi anemia complementation group assignment, essential for efficient molecular diagnosis of the disease, had not been validated for clinical application before this study. The purpose of this study was (1) confirmation of the accuracy of Fanconi anemia complementation group assignment to Group A (FANCA) and (2) development of a rapid mutation detection strategy that ensures the efficient capture of all FANCA mutations. Methods: Using fibroblasts from 29 patients, diagnosis of Fanconi anemia and assignment to complementation Group A was made through breakage analysis studies. FANCA coding and flanking sequences were analyzed using denaturing high pressure liquid chromatography, sequencing, and multiplex ligation-dependent probe amplification. Patients in which two mutations were not identified were analyzed by cDNA sequencing. Patients with no mutations were sequenced for mutations in FANCC, G, E, and F. Results: Of the 56 putative mutant alleles studied, 89% had an identifiable FANCA pathogenic mutation. Eight unique novel mutations were identified. Conclusion: Complementation assignment to Group A was validated in a clinical laboratory setting using our FANCA rapid molecular testing strategy. Genet anconi anemia (FA; MIM no. 227650), the most common inherited bone marrow disorder, has an overall prevalence of 1-5 per million and an estimated carrier frequency of 1 in 200 to 1 in 300 in most populations. 1,2 Demonstrating either an autosomal or X-linked recessive mode of inheritance, FA is characterized by childhood progressive bone marrow failure and predisposition to acute myelogenous leukemia; older patients are at increased risk for squamous cell carcinomas of the head, neck, and genitourinary tract. [3][4][5][6][7] Congenital abnormalities are present in approximately 70% of FA patients and may include radial ray defects; café au lait spots or hypopigmentation; short stature; microphthalmia; malformations of kidneys, gastrointestinal tract, and heart; mental retardation; and hearing defects. 8 Because of the high degree of phenotypic variability exhibited by FA patients, diagnosis may be difficult on the basis of clinical manifestations alone. Because FA patient-derived lymphocytes and fibroblasts exhibit hypersensitivity to DNA crosslinking agents such as diepoxybutane (DEB) 9 and mitomycin C (MMC), 10 resulting in a high rate of chromosomal breakage and radial formation, analysis on the basis of this hypersensitivity has been routinely used to confirm clinical diagnosis.Molecular diagnosis of FA has been challenging because of the genetic heterogeneity associated with the disease; FA is multigenic, with 13 complementation groups and associated genes having been characterized (A, B, C, D1, D2, E, F, G, I, J, L, M, N). [11][12][13][14][15][16][17][18][19][20][21][...

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“…The FA genes clearly protect cells from genotoxic stress. Mutant cells exhibit spontaneous chromosomal breakage 38 and increased sensitivity to the development of cytogenetic anomalies on in vitro treatment with cross-linking agents 39 such as MMC 40 and DEB. 41 Additional functions for FA proteins have also been described, including modulation of cytokine-mediated signaling, [42][43][44][45][46][47][48][49][50] responsiveness to oxidative stress, [51][52][53][54][55][56][57] and chromatin remodeling.…”

Section: Genetic Analysis Of a Lymphoblastoid Cell Line From The Samementioning

confidence: 99%

Acquired FANCA dysfunction and cytogenetic instability in adult acute myelogenous leukemia

Lensch¹

2003

Blood

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Myelodysplastic and leukemic stem cell clones that evolve in children and adults with Fanconi anemia universally bear complex cytogenetic abnormalities. The abnormalities are generally recurring deletions or chromosomal loss and involve precisely the same chromosomes with the same frequency as has been described in marrow cells from patients with secondary acute leukemia induced by alkylating agents. Reasoning that acquired Fanconi anemia protein dysfunction might contribute to cytogenetic instability in secondary acute myelogenous leukemia (AML) cells, we analyzed leukemic cells bearing characteristic complex cytogenetic defects obtained from a 68-year-old man whose lymphoblasts showed no evidence of Fanconi anemia. Unlike the lymphoblasts, this myeloid leukemia cell line (UoC-M1) was hypersensitive to mitomycin-C (MMC) and diepoxybutane (DEB) and exhibited a marked decrease in nuclear FANCA, FANCG, and FANCD2-L. Retroviral transduction of FANCA significantly reduced MMC sensitivity but FANCF, FANCG, and FANCC did not. Overexpression of FANCA restored levels of both FANCA and FANCG, whereas overexpression of FANCG or IntroductionA variety of molecular defects have been discovered in patients with myeloproliferative syndromes and acute myelogenous leukemia, many of which result in the activation of autonomous signal transduction pathways for growth and survival. [1][2][3][4][5] Although oncogenic mutations in such pathways are sufficient to induce myeloproliferative disorders in transgenic mice, 2,3 the development of acute leukemia requires additional mutations that ultimately interfere with myeloid differentiation. 6 The risk of this second type of mutation would be necessarily enhanced by cytogenetic instability, a manifestation of which in the leukemic clone would be multiple cytogenetic defects. That such complex cytogenetic defects universally occur in the myeloid leukemic clones of children with Fanconi anemia, a cytogenetic instability syndrome, supports this notion. In fact, because the clonal abnormalities found in this clinical context exactly recapitulate those found in patients with secondary myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML), 7,8 and high-risk MDS, 9 we hypothesized that acquired Fanconi anemia (FA) protein dysfunction might serve as a progression factor for the evolution of cytogenetically unstable AML clones, even in patients without hereditary evidence of Fanconi anemia. To test this notion, we examined a variety of AML cells and cell lines, including a cell line from a 68-year-old patient with cytogenetic defects characteristic of secondary AML. The patient's lymphoblasts were normal, but the leukemic cell population exhibited hypersensitivity to mitomycin C, a feature of Fanconi anemia. Nuclear FANCC, FANCG, and FANCA levels were markedly reduced, and retroviral complementation indicated that the leukemic cells exhibited a secondary defect of FANCA function. Because lymphoblasts from this same patient showed normal levels of nuclear Fanconi proteins, we propo...

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Cytogenetic differentiation of Fanconi anemia, “idiopathic” aplastic anemia, and Fanconi anemia heterozygotes

Cited by 72 publications

References 19 publications

A study of bone marrow failure syndrome in children

A study of bone marrow failure syndrome in children

Validation of Fanconi anemia complementation Group A assignment using molecular analysis

Acquired FANCA dysfunction and cytogenetic instability in adult acute myelogenous leukemia

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