Fanconi Anemia via the FANCD2 Gene

Fanconi Anemia (FA) is an inherited anemia associated with bone marrow failure (aplastic anemia), however the clinical features of FA can expand well beyond hematologic anomalies. FA is characterized by a range of physical abnormalities, bone marrow failure (aplastic anemia), pancytopenia, and predisposition to cancers - particularly acute myelogenous leukemia (AML), gynecologic and GI tract cancers, and cancers of the head and neck (Auerbach 2009). FA patients are up to 800 fold more susceptible to AML than the general population with a median age of onset of 13 years (Rosenberg et al 2003). Physical abnormalities include radial ray defects (absent thumb or radius), skin pigmentation defects, short stature, microphthalmia, renal and urinary tract defects, genital defects (males in particular), gastrointestinal malformations (atresia), congenital heart disease, hearing and central nervous system defects, and general developmental delay (Tischkowitz and Hodgson 2003; Dokal 2000). About one-third of FA patients have no obvious physical abnormalities and are diagnosed only after a family member is diagnosed, or after developing hematologic anomalies such as thromobocytopenia, leukopenia, and anemia (Giampietro et al. 1997). A hallmark of FA is hypersensitivity of chromosomes to inter cross-strand linkage (ICL) agents such as diepoxybutane (DEB) or mitomycin C (MMC) (Sasaki and Tonomura 1973). Exposure of primary cell cultures from FA patients to DEB or MMC results in chromosomal aberrations (breaks, radials, rearrangements) due to damaged DNA repair mechanisms that require functional products of the Fanconi anemia genes. For example, the FANCA, -B, -C, -E, -F, -G, -L, and -M proteins are part of a nuclear core complex that regulates monoubiquitination of the FANCD2 and FANCI proteins (ID complex) during S-phase and after exposure to DNA crosslinking agents (Moldovan and D'Andrea 2009). In unaffected individuals, ubiquitination helps localize the ID complex to sites of DNA damage and facilitate repair (Grompe and van de Vrugt 2007; Smogorzewska et al. 2007), but in FA patients, this mechanism is impaired.

FA is a genetically heterogeneous disorder. To date, 21 FA or FA-like genes have been discovered. Inheritance is primarily autosomal recessive or X-linked, however a case of heterozygous FA-like syndrome was associated with a dominant-negative variant in the RAD51 (FANCR) gene (Ameziane et al. 2015). Approximately 86% of all cases are attributed to variants in three genes: FANCA (~60%), FANCC (~16%) and FANCG (~10%) (Auerbach 2009). Since variants in FANCA are the most common cause of FA, it is important to note that large deletions make up over one-third of all reported pathogenic variants in FANCA. In the United States, the carrier frequency for Fanconi anemia is estimated at 1 in 181 and the incidence rate is estimated at 1 in 131,000 (http://www.fanconi.org/; Rosenberg et al. 2011). Nearly 95% of all FA cases are attributed to variants in eight genes, FANCA, -C, -G, -D1 (aka BRCA2), -D2, -E, -F, and –L that are either part of the core complex required for ID complex ubiquitination and facilitation of DNA repair or function directly in ICL recognition and repair (Grompe and van de Vrugt 2007). FA is phenotypically diverse even among related patients that harbor a common variant; null alleles however are reported to result in more severe phenotypes (Faivre et al. 2000). FA affects males and females roughly equally and affects all ethnic groups.

Nearly 95% of all Fnconi Anemia cases are attributed to variants in eight genes, FANCA, -C, -G, -D1 (aka BRCA2), -D2, -E, -F, and –L that are either part of the core complex required for ID complex ubiquitination and facilitation of DNA repair or function directly in ICL recognition and repair (Grompe and van de Vrugt 2007). ~ 3-6% of FA patients are assigned to the FANCD2 complementation group (Kalb 2007; www.ncbi.nlm.nih.gov/). Gross deletions or duplications account for ~ 10% of the reported variants in the FANCD2 gene (www2.rockefeller.edu/fanconi/genes/).

Exons 12-28 of the FANCD2 gene share high sequence similarity with several other genomic regions outside of the FANCD2 gene (paralogs). In order to ensure we are sequencing exons 12-28 of the FANCD2 gene and not the paralogous regions, exons 12-28 are sequenced using Sanger sequencing and employing primers unique to the functional FANCD2 gene.

This test provides full coverage of all coding exons of the FANCD2 gene plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define full coverage as >20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).

NOTE: Due to the presence of paralogous regions in the genome, exons 12-28 of the FANCD2 gene are difficult regions to cover using aCGH. Consequently, our aCGH test for the FANCD2 excludes exons 12-28.