Clinical Applicability of FANCD2 Mono-Ubiquitination Test for Fanconi Anemia Diagnosis and a Suggestion for

Objectives: Fanconi anemia is caused by mutations in related FANC genes that are involved in DNA repair pathways. Diepoxybutane induced chromosome break test in lymphocytes if positive is affirmative for the diagnosis. However, in 20-25% of the cases the test results have intermediate values which are mostly attributed to somatic mosaicism and cause an ambiguity for diagnosis. Here, we utilize the relatively new and rarely used FANCD2 mono-ubiquitination test to facilitate and resolve the diagnostic problem in a clinical scheme. Methods: Fifteen patients with different diepoxybutane test results (positive, intermediate, negative) and four control subjects were included to the study. Western blot analyses in cultured peripheral blood lymphocyte-isolates were done with a specific antibody to discriminate between ubiquitinated and non-ubiquitinated forms of FANCD2. Results: The test confirmed the diagnosis in one of the diepoxybutane positive patients. In four cases out of seven with intermediate test values we were able to make the right diagnoses. For the other three in this group, FANCD2 test neither confirmed nor rejected the diagnoses which may be due to high somatic mosaicism or due to a defect in the downstream part of the Fanconi pathway. The ubiquitinated isoform of FANCD2 were detected in all six diepoxybutane negative cases. Conclusion: It is expected that adopting FANCD2 mono-ubiquitination Western blot analysis as a diagnostic tool with the proposed algorithm could facilitate the management of Fanconi Anemia without compromise.