Rapid assessment of fanconi pathway function: A new diagnostic approach to fanconi anemia

W. Nicholas Haining, Lisa Moreau, Toshiyasu Taniguchi, Rocio Montes De Oca, Irene Garcia-Higuera, Eva C. Guinan, Markus Grompe, Alan D. D'Andréa

Research output: Contribution to journalArticlepeer-review


Fanconi Anemia (FA) is an autosomal recessive cancer susceptibility syndrome characterized by cellular sensitivity to DNA crosslinking agents, such as DEB (diepoxybutane). The DEB (chromosome breakage) test is a highly sensitive, specific, and reproducible method for diagnosing new patients with FA. However, it has several disadvantages: 1) it requires a high degree of technical expertise; 2) it cannot detect heterozygous carriers of FA; and 3) it can generate false negative results in cases of FA mosaicism. In an attempt to develop an improved diagnostic test for FA, we have exploited advances in the molecular understanding of the FA pathway. We have recently demonstrated that the Fanconi Anemia protein complex, consisting of the FANCA, FANCC, and FANCG proteins, is required for the downstream monoubiquitination of the FANCD protein. Biallelic mutation of any of the other upstream FA genes (FANCA, FANCB, FANCC, FANCE, FANCF, and FANCG) blocks this post-translational modification of FANCD. A western blot of the FANCD protein readily distinguishes the unubiquitinated (FANCDS) and ubiquitinated (FANCD-L) forms of the FANCD protein. This provides a convenient test of the integrity of the FA pathway. In the current study we have devised a rapid diagnostic test for FA, using peripheral blood samples and an anti-FANCD immunoblot. Individuals with upstream defects in the FA pathway are unable to ubiquitinate FANCD which is evident by an absence of FANCD-L on a western blot. The assay has comparable sensitivity and specificity to the DEB test, but is more rapid and robust, and does not involve hazardous chemicals. We have also employed a combination of retroviral gene transfer and FANCD immunoblotting to provide a rapid subtyping analysis of newly diagnosed FA patients. This screening test provides a simple but powerful tool to assess FA pathway function, and make the diagnosis of FA. Furthermore, it may be able to distinguish subtle differences in pathway function that may exist between carriers of FA gene defects and unaffected individuals. This functional probe of the FA pathway may also lead to an advanced understanding of the role of as yet uncharacterized FA pathway defects in myelodysplastic syndrome, aplastic anemia or cancer susceptibility syndromes.

Original languageEnglish (US)
Pages (from-to)228a-229a
Issue number11 PART I
StatePublished - Dec 1 2000
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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