Fanconi Anemia

Etiology

Fanconi anemia is an inherited autosomal recessive disease, but about 2% is inherited in an X-linked recessive manner, with either homozygous or heterozygous mutations. An autosomal dominant subtype has been noted. DNA sequencing has identified pathogenic alleles, including point mutations, duplications, splicing defects, and deletions.[1] Mutation of Fanconi anemia genes results in impaired double-stranded DNA repair. More than 23 Fanconi anemia complementation genes (FANC) have been recognized, and all of them are involved in the DNA repair pathway.[6] Most of them are autosomal recessive except a few, such as FANCB, which is X-linked, and FANCR (RAD51), which is autosomal dominant.[7][8] 

Epidemiology

Fanconi anemia is a very rare type of anemia. Overall, an average of 1 out of 136,000 newborns has Fanconi anemia, ranging from 1 in 100,000 to 250,000 births.[9] European registries and data reveal the prevalence of Fanconi anemia is just 4 to 7 per million live births.[10] Fanconi anemia has been found in all races.[11] However, the rate is higher in South Africans, sub-Saharan Africans, and Spanish Gitanos, with rates of 1 in 40,000 births.[12] A higher carrier frequency among US Ashkenazi Jews of 1 case per 100 people has been reported. There is also a birthrate of approximately 1 per 30,000 live births.[13] Fanconi anemia has a slight predilection for men more than women.

Pathophysiology

Fanconi anemia is characterized by chromosomal damage due to the cell's inability to conduct repairs.[2][3] With biallelic mutations in the genes associated with the Fanconi anemia core complex, dysfunctionality occurs in the entire Fanconi anemia pathway. Biallelic mutations of FANCA, FANCC, and FANCG account for over 80% of cases.[14] The bone marrow failure of Fanconi anemia is thought to occur from the selective destruction of CD34+ stem cells. The primary processes affected are related to DNA repair, including homologous recombination, nucleotide excision repair, mutagenic translational synthesis, and alternate end joining. Replication processes include the stabilization of replicative forks, as well as the regulation of cytokines. 

The purpose of Fanconi anemia proteins is to maintain genomic integrity and replicative capacity. Various Fanconi anemia proteins maintain genomic stability by repairing DNA interstrand cross-links (ICLs). ICLs prevent DNA strand separation and maintain DNA integrity. Genetic defects in the Fanconi anemia pathway lead to cells that cannot properly repair DNA damage, resulting in genomic instability, subsequent pancytopenia, and an increased susceptibility to cytotoxic agents, ultraviolet radiation, spontaneous deformation, and predisposition to malignancies. Studies have shown that there is a high frequency of solid tumors (eg, head and neck squamous cell carcinomas) with FANCA variants affecting exons 27 to 30.[15]

Proteins encoded by Fanconi anemia genes include ubiquitin ligase, monoubiquitinated protein, helicase, and breast and ovarian cancer susceptibility proteins. These proteins are responsible for DNA repair and resistance against DNA damage insults. Some Fanconi anemia proteins are similar to the BRCA2 protein, such as FANCD1, which works in the Fanconi anemia-BRCA network. An interactive network of these Fanconi anemia proteins with other proteins is responsible for other rare genetic syndromes (eg, ataxia-telangiectasia, Bloom syndrome, and breast and ovarian cancer).[16][17][18] These Fanconi anemia proteins also have multiple other functions. The Fanconi anemia pathway involves FANC genes and their products to maintain genomic integrity. Fanconi cells show hypersensitivity to crosslinking agents, eg, mitomycin C, diepoxybutane, and cisplatin. Fanconi anemia proteins are also involved in other stress response pathways. The inability of the affected cells to withstand normal oxidative stress and oxygen-free radicals causes oxidative damage.[19]

Fanconi Anemia Genetics and Genomic Instability

Several Fanconi anemia genes have been identified including FANCA, FANCB, FANCC, FANCD1/BRCA2, FANCD2, FANCE, FANCI, FANCM, FANCN/PALB2, FANCO/RAD51C, FANCP/SLX4, FANCR/RAD51, FANCS/BRCA1, FANCV/REV7 and FANCW/RFWD3.[20] Proteins encoded by these genes have various roles in the DNA repair Fanconi anemia pathway. The formation of ICLs leads to Fanconi anemia core complex arrangements. Among them, the FANCD2 protein has a central role in this pathway. Core proteins FANC (A, B, C, E, F, G, L, M), more specifically, the FANCL, component acts as E3 ligase responsible for monoubiquitination, and other core proteins for phosphorylation and translocation of FANCD2 proteins in nuclear foci.[9] Monoubiquitination and phosphorylation are the activation processes after various agents damage DNA in the normal DNA repair pathway. Monoubiquitinated FANCD2 accumulates in nuclear foci with other proteins such as BRCA1, FANCD1/BRCA2, RAD51, and FANCN/PALB2, which are involved in DNA repair and resistance against DNA crosslinking and damaging agents like ionizing radiation, hydroxyurea, and UV light irradiation.

FANCI is monoubiquitylated and phosphorylated after DNA damage, but its role is not prominent. Genetic defects of any component of this pathway result in decreased resistance against damaging agents, ultimately leading to chromosome fragility. Fanconi anemia core complex proteins FANCC and FANCE localizes to nuclear foci along with FANCD2, FANCG with BRCA2, and RAD51. This nuclear foci formation and the monoubiquitination of FANCD2 proteins are the primary steps in the Fanconi anemia DNA repair pathway. The Fanconi anemia core protein complex, with the help of the FANCM protein, stimulates translocase and ATPase, which causes ATP hydrolysis and energizes translocase to move the Fanconi anemia core complex along the DNA strand. Another FAAP24 protein binds to ssDNA and the branching structures at the damaged site. In addition, FANCM contains helicase and endonuclease domains, which separate the double-stranded DNA and cleave the phosphodiester bond at the damaged site to repair the damaged part. FANCJ is also a helicase that binds and interacts with BRCA1.

Fanconi anemia core complex with other proteins (eg, BLM, RPA, and topoisomerase III-alpha) are also involved in the DNA repair pathway. Some of the Fanconi anemia proteins, for instance, FANCD1 with BRCA2 and FANCJ with BACH1 and BRIP1, work downstream on homologous recombination DNA repair. Ultimately, bone marrow failure and other complications are due to defects in the DNA repair pathway, increased oxygen-reactive species, and inflammatory cytokines.[3] Cancer predisposition has the same common reason for defective DNA repair systems.[21] 

Histopathology

Microscopic examination of a bone marrow biopsy shows hypoplasia and hypocellularity with fatty replacement characteristic of aplastic anemia. Hypocellularity is often out of proportion with cytopenias. Bone marrow biopsy at infancy may be normocellular. Some islands of hyperplastic erythroblasts and erythroid dysplasia can be seen. 

History and Physical

Clinical History

Common clinical symptoms of Fanconi anemia include shortness of breath, chest pain, dizziness, and fatigue. Additionally, a clinical history of epistaxis, petechiae, and excessive bleeding from a wound site is common due to thrombocytopenia. The risk of recurrent infections increases with the severity of leukopenia, which presents with fever and flu-like illness. A history of low birth weight is present in some cases, and a history of weight loss is essential to elicit from patients for cases complicated by cancer. Family and marriage history are crucial, primarily in populations with a high prevalence of consanguinity marriages.

Physical Examination Findings

Approximately 75% of Fanconi anemia is associated with congenital disabilities. On physical examination, the child's stature may be short, and light-brown skin pigmentation (ie, café au lait) lesions in more than 50% of cases.[22] Structural abnormalities of extremities are more common in patients with Fanconi anemia. Common upper limb abnormalities include absent, bifid, supernumerary, low set, short or hypoplastic thumb, absent or hypoplastic radii, and dysplastic ulna. Lower limb abnormalities include polydactyly, short toes, club foot, flat feet, hip dislocation, abnormal femur, and thigh osteoma. Other skeletal abnormalities consist of head and face anomalies, microcephaly and hydrocephaly, frontal bossing, flathead, micrognathia, sloped forehead, webbed and short neck, low hairline, spina bifida, scoliosis, abnormal ribs, extra vertebrae. Hypogonadism is common in both sexes and includes less genitalia development, undescended and absent testis, phimosis, hypospadias, micropenis, and vaginal atresia. Fanconi anemia complicates pregnancies. 

Other anomalies include abnormal epicanthal folds, proptosis, ptosis, cataracts, blindness, epiphora, and ear anomalies such as absent eardrums, small or large pinnae, and atresia of the ear canal. Physical signs of pancytopenia are sometimes visible, including pallor, petechiae, bruising, and coldness of the hands and feet. Gastrointestinal abnormalities such as imperforate anus, tracheoesophageal fistula, atresia of the intestine, Meckel diverticulum, megacolon, hepatocellular adenoma, and umbilical hernia are less common. Proximal renal transport impairment may occur, unlike Fanconi syndrome, which has no primary glomerular involvement. Cardiac effects include ventricular septal defects. 

Evaluation

Fanconi anemia leads to pancytopenia, the consequence of bone marrow failure. Pancytopenia characteristic of Fanconi anemia is usually evident in serum laboratory studies demonstrating a decrease in all 3 blood cell lines, including red blood cells (RBCs), platelets, and leukocytes. Diagnosis may be delayed until the bone marrow failure develops. Different studies have summed up the average age of diagnosis as 7 years, although earlier diagnosis has been increased due to disease awareness and prenatal screening diagnostic enhancement. Early diagnosis may prevent severe complications [23]. 

Fanconi anemia should be evaluated in patients presenting with signs and symptoms of pancytopenia with or without characteristic malformations and in patients with a family history of bone marrow failure or unbalanced translocations identified during a diagnostic leukemia evaluation. Furthermore, Fanconi anemia should be considered in patients with early-onset tumors or excessive toxicity after standard-dose chemotherapy. A Fanconi anemia diagnosis is typically confirmed with a chromosomal fragility test, which remains the gold standard.[5]

Laboratory Studies

A complete blood count reveals the level of RBCs, white blood cells (WBCs), and platelets. Raised mean corpuscular volume (MCV) indicates macrocytosis, and there can be high fetal hemoglobin levels due to increased stress. Serum erythropoietin is increased due to the low level of blood cells and the low response of hematopoietic stem cells. Bone marrow aspiration and biopsy reveal hypocellularity, aplasia with fatty marrow, and absence of myeloid, erythroid, and megakaryocyte stem cell lines. Myelodysplastic cases show hypo- or hyper granularity, hypopigmentation of myeloid precursors, and hypo-lobulated or hyper-lobulated megakaryocytes. Clonic malformations are seen in leukemic transformations.

The chromosomal breakage/stress cytogenetics test is a diagnostic test indicated in those with severe pancytopenia defined as an absolute neutrophil count of <100 mL, hemoglobin <10 g/dL, reticulocyte <40,000 mL, bone marrow cellularity <25%, and a platelet count of <50,000 mL. Testing with DNA cross-linkers such as diepoxybutane or mitomycin C stimulates the breakage of DNA in the absence of a DNA repair system. These agents increase the chromatids' breaks, gaps, reduplications, or rearrangements. Cultured fibroblasts also show chromosome fragility and are preferred in patients with negative T-lymphocyte chromosome fragility testing or those who have already undergone hematopoietic cell transplantation.[24]

Cell cycle analysis/flow cytometry is an alternative to the chromosomal breakage test. In this test, those cells with impaired DNA repair undergo G2 arrest following DNA cross-linking agent exposure. Immunoblotting of GANCD2 mutation is less commonly used. Fanconi anemia gene sequencing is recommended in patients with positive chromosomal breakage testing. The identification of genetic defects is confirmatory and excludes other chromosome breakage syndromes. Prenatally, abnormal chromosome breakage can be analyzed with amniotic fluid cells or chorionic villous biopsy. An elevated level of serum alpha-protein is a rapid screening diagnostic test, but it may not be identified in many cases.[25]

Additional imaging surveys may be done to evaluate for other structural and congenital disability abnormalities. A skeletal x-ray can detect the size and type of bone defects. The x-ray of the head shows a crewcut appearance. An abdominal ultrasound can assess liver and kidney abnormalities. Additionally, magnetic resonance imaging (MRI) is crucial to identify CNS abnormalities (eg, absence of the corpus callosum, cerebellar and pituitary hypoplasia). 

Treatment / Management

Supportive Therapy

Blood transfusions are the best supportive therapy for Fanconi anemia. Packed RBCs and platelet transfusions have an immediate effect. RBC transfusion from family members should be avoided due to alloimmunization and graft-versus-host disease. Leukopenia responds well to granulocyte-colony-stimulating factor, but this is reserved for those patients with an ANC<200/mL. Extensive transfusion has poor outcomes in patients with hematopoietic cell transplantation.

Hematopoeitic Stem Cell Transplantation

Cure of aplastic anemia and prevention of myelodysplastic syndrome can be achieved through bone marrow, peripheral blood cells, and cord blood transplantation. Bone marrow transplantation from an HLA-matched sibling is the preferred method. Recent clinical trials utilized a low-intensity, fludarabine-based preparative regimen.[26] The 5-year overall survival was higher with Fanconi patients having aplastic anemia as compared with those having underlying myelodysplasia or acute promyelocytic leukemia. Chromosome breakage testing of siblings or other related donors should be performed to exclude Fanconi anemia from donors. However, hematopoeitic stem cell transplantation (HCT) is not accessible to everyone and is typically reserved for patients with severe myelodysplastic syndrome, leukemia, or treatment failure. Approximately 50% to 75% of patients respond, and it is a more permanent treatment.[27] Alternate approaches using autologous transplants are being investigated to prevent the adverse effects of allogeneic hematopoietic stem cell transplant (eg, graft versus host disease).[28][29] One methodology utilizes "digital" genomic editing, or correction, of the FANCA mutation (Fanconi Anemia Complementation Group A) through cytosine and adenosine base editors. 

Androgen Therapy

Androgen therapy may be considered in patients who are not good candidates for hematopoeitic stem cell transplantation. Oxymetholone is the most commonly used androgen. Other androgens less widely used are danazol and oxandrolone. Androgens stimulate hematopoietic stem cell proliferation, but it is not curative. Oxymetholone is initiated with a starting dose of 2 to 5mg/kg/day and then tapered to avoid toxicity. Patients with severe bone marrow hypocellularity frequently have a poor response to androgen therapy. Red blood cells respond well, but platelets and leukocytes are less likely to respond. Some sources advocate using androgens only in children as a bridge therapy to transplantation or in patients ineligible for transplant.[14]

Surgical Therapy

Surgery is performed only for the management of structural deformities. Splitting of anomalies of the hand should be repaired early in life to avoid functional delays. Other surgeries include congenital heart defect surgery, repair of trachea-esophagus fistulas, and imperforate anus. Surgery may be required for cancers as well.

Gene Therapy

Gene therapy is a developing treatment that replaces an abnormal gene with a normal gene. The correction of CD34+ in affected cells is now feasible.[30] However, gene therapy reportedly does not fully correct the problem.[14](B3)

Differential Diagnosis

Fanconi anemia resembles a large variety of diseases. Other hematological problems that manifest clinical features of Fanconi anemia and congenital structural defects associated with Fanconi anemia should be excluded.[23][31]

Acquired aplastic anemia: This condition is due to acquired hematopoietic stem cells' destruction of bone marrow following various toxicogenic agents. Pancytopenia is the cardinal feature of Fanconi anemia due to bone marrow failure. Hypocellularity of the bone marrow is sufficient for diagnosing acquired aplastic anemia, which does not show chromosome fragility by chromosome breakage test. Fanconi anemia shows chromosome fragility test positive, and gene sequencing is required to confirm the diagnosis.[32][33]

Other inherited bone marrow failure syndromes: Because bone marrow failure is distinct in Fanconi anemia, differentiation of other causes of bone marrow failure is essential. Diseases with a shortened telomerase, such as dyskeratosis congenital, are associated with bone marrow aplasia. Bone marrow aplasia caused by reticular dysgenesis is a combined immunodeficiency disease due to AK2 gene mutation. However, sensorineural hearing loss and adaptive immune deficiency in infancy are prominent features of reticular dysgenesis. Diamond-Blackfan anemia is pure red cell aplasia that typically presents with bone marrow abnormalities.[34] Platelets and white blood cells are generally normal, but reticulocytes are decreased. Schwachman-Diamond syndrome is bone marrow aplasia primarily associated with neutropenia in 88% to 100% of cases.[35] There is also associated pancreatic insufficiency. Congenital amegakaryocytic thrombocytopenia principally affects platelets.[36] All these diseases may show trilineage aplasia in the bone marrow after an extended period.

Denovo myelodysplastic syndrome: This condition is a clonal myelodysplastic disease characterized by ineffective hematopoiesis and peripheral cytopenias. Denovo myelodysplastic syndrome (MDS) is also associated with Fanconi anemia, but de novo MDS is not susceptible to chromosome breakage tests and does not have FA mutation or congenital anomalies. Suspected MDS patients with associated FA should be tested with skin fibroblast chromosome breakage tests rather than peripheral lymphocytes. MDS typically presents with bilineage cytopenias.

Drug-induced or infection-associated pancytopenia: Exposure to many cytotoxic drugs, mainly cancer chemotherapy, viral and bacterial infections, and chemicals cause transient bone marrow hypoplasia and pancytopenia in children, but unlike FA, this pancytopenia lacks congenital anomalies and also pancytopenia is reversible with removal of the agents. The chromosomal breakage test is negative.

Paroxysmal nocturnal hemoglobinuria: Paroxysmal nocturnal hemoglobinuria results from mutations of the PIGA gene encoding anchoring protein glycosylphosphatidylinositol (GPI), mainly in hematopoietic progenitor cells, which leads to nocturnal hemolysis due to activation of the complement system in the acidic environment during shallow breathing at night. Patients are anemic and have an increased risk of thrombosis, hemoglobin in the urine, and jaundice. Bone marrow failure is mainly due to acquired autoimmunity rather than being inherited. Chromosomal breakage testing results are negative.

Other rare chromosomal breakage syndromes: Many chromosomal instability diseases in the presence of irradiation, DNA cross-linkers, and chemicals are evident but rare, which mimic FA and necessitate their exclusion for an accurate diagnosis and treatment plan. These disorders include Bloom syndrome,[37] LIG4 syndrome, Ataxia-telangiectasia, Nijmegen breakage syndrome, Seckel syndrome, NHEJ1 deficiency, Warsaw breakage syndrome, and cohesinopathies (eg, Robert syndrome).[38][39][38][40][41][42] Similar to Fanconi anemia, these diseases show an abnormal chromosomal breakage test and congenital anomalies (eg, short stature, microcephaly, and malignancy). They can be quite difficult to differentiate. However, bone marrow failure is not typical of rare chromosomal breakage syndromes, and there are also spectral differences in the congenital anomalies and malignancies associated with Fanconi anemia and other chromosomal breakage syndromes. Genomic sequencing of mutation may be utilized to differentiate these conditions. 

Pertinent Studies and Ongoing Trials

Several clinical trials are ongoing to investigate the therapy and prevention of Fanconi anemia. Clinicians should be aware of these trials and refer their patients accordingly.[14][43][44]

Prognosis

The prognosis of Fanconi's anemia is poor.[2] Severe aplastic anemia is the main cause of mortality that leads to death before 10 years of age in the absence of a diagnosis. Many children with Fanconi Anemia die of bacterial and fungal infections.[3] Survival has been improved in developed countries due to reduced mortality by bleeding or infection complications. However, living well into adulthood has increased the chances of cancer development. Most individuals eventually develop acute myelogenous leukemia or myelodysplasia.[45] Allogeneic BMT is the only curative option for these patients, but regular monitoring is needed for the possible relapse in those who have undergone a transplant. In a small study, a relapse rate of 16% was seen, and a graft failure rate of 16% was also evident.[44] Most of the patients have other associated birth defects, including developmental delays, kidney problems, and microcephaly.   

Complications

The major complications of Fanconi anemia are aplastic anemia, myelodysplastic syndrome (MDS), acute myeloid leukemia, and specific solid tumors.[2][46] 

Fanconi anemia is associated with various cancers, as the defective FA gene is associated with various cancers.[47] Defects of genes of the Fanconi anemia pathway responsible for DNA repair and cell cycle checkpoints are involved in the hypersensitivity of cancer cells to chemotherapeutic drugs, viruses, and radiation. Impairment of the DNA repair system and lack of stability in cell cycle checkpoints leads to the uncontrolled proliferation of cells. Common variants of cancer include squamous cell carcinomas of the head, neck, and upper esophagus and carcinomas of the vulva, anus, and cervix, which have a 50-fold higher risk as compared to the absence of Fanconi anemia association. Myelodysplastic syndrome is the most common finding, which has a 6000-fold higher chance as compared to the general population. Acute myelogenous leukemia is the second most common cancer, which has a 700-fold higher chance as compared to the general population. Clonal mosaicism in Fanconi anemia has been demonstrated in aging and cancers.[48] The presence of +1q or 7Q- are harbingers of AML transformation.[14]

Additionally, Fanconi anemia registries reveal around 90% of patients suffer from bone marrow failure by the age of 40 years.[49] Bone marrow failure ultimately leads to pancytopenia.[50] A wide range of endocrine disorders are either intrinsic to molecular defects or associated HCT and androgen therapies. Structural disruption of the hypothalamus-pituitary axis may lead to short stature due to growth hormone deficiency, hypothyroidism is found in around 60% of patients, adrenal dysfunction that responds to exogenous ACTH, and dysfunction of pancreatic islets associated with glucose intolerance, dyslipidemia, and infertility due to hypogonadism. Androgen therapy causes benign and malignant liver tumors and peliosis hepatis.[51][52]