High Incidence of Fanconi Anemia in Patients with a Morphological Picture Consistent with Refractory Cytopenia of Childhood

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2780-2780
Author(s):  
Ayami Yoshimi ◽  
Charlotte M. Niemeyer ◽  
Irith Baumann ◽  
Stephan Schwarz-Furlan ◽  
Detlev Schindler ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Test Group ◽  
Clinical Feature ◽  
Chromosomal Breakage ◽  
Mild Phenotype ◽  
European Working ◽  
Chromosome Breaking

Abstract Abstract 2780 Introduction: Refractory cytopenia in childhood (RCC) is the most common subtype of myelodysplastic syndrome (MDS) in children. Differential diagnosis from inherited bone marrow failure (IBMF) such as Fanconi anemia (FA) remains an intriguing challenge, because most patients with RCC have a hypocellular bone marrow (BM) and dysplastic features in haematopoiesis are observed in both RCC and IBMF. Moreover the spectrum of phenotypic findings in FA is extremely wide. Some FA patients have a mild phenotype without malformation. The purpose of this study is to estimate the incidence of FA in an RCC cohort without a full clinical feature of FA, but subsequently diagnosed by chromosome breaking test. Patients and Methods: Between 01/2007 and 12/2010 reference pathologists of the European Working Group of MDS in Childhood (EWOG-MDS) provided a morphological report consistent with RCC in 137 children studied in Germany. Seventeen patients with hypercellular BM or abnormal karyotype, 2 patients, in whom dyskeratosis congenital was diagnosed after initial inclusion and one patient, in whom chromosome breaking test was not performed, were excluded. Results: Seven of remaining 117 patients had facial and/or skeletal anomalies typically noted in FA and one patient had a brother with FA. In these 8 patients, FA had been suspected by their local physicians (group FA-1). Nine patients (8.3%) without these typical anomalies were subsequently diagnosed of FA by chromosome breakage test (group FA-2). The diagnosis of RCC was finally made in the remaining 100 patients with negative chromosomal breakage test (group RCC). The clinical features of patients in each group are summarized in the Table. The mean corpuscular volume of red cells (MCV) was elevated (> +2SD) for ages in all patients with FA, but only 42 % in patient with RCC. In some children of group FA-2 additional non-haematological abnormalities were also observed. However, they were not evident and or typical to prompt the treating physicians to suspect FA. A few patients in the group RCC also had some physical anomalies, not specific for any of the known IBMF disorders. Possibly that other known or not yet described IBMF disorders remain uncovered in children with “de novo” RCC. Conclusion: Our results illustrate that the same haematological features and congenital anomalies can be noted in FA and RCC. More importantly, they indicate that the exclusion of FA by a chromosomal breakage test or other methods is mandatory in all patients prior to diagnosis RCC. Chromosomal breakage analysis may identify patients with FA in 8% of patients with a morphological description of RCC without a full clinical picture of FA. Disclosures: No relevant conflicts of interest to declare.

Unexpected Fanconi Anemia Diagnosis in Patients with Bone Marrow Failure.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1056-1056
Author(s):  
Fernando O. Pinto ◽  
Thierry Leblanc ◽  
Gwenaelle Le Roux ◽  
Helene Dastot ◽  
Moema Santos ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Clinical Features ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Somatic Mosaicism ◽  
Chromosome Breakage ◽  
Large Set ◽  
Chromosomal Breakage ◽  
Group A

Abstract Early diagnosis of Fanconi Anemia (FA) in patients with bone marrow failure is critical for optimal clinical management. However, the remarkably high clinical variability and the potential emergence of revertant hematopoietic cells (somatic mosaicism) can obscure and delay the diagnosis of FA. Here we addressed FA diagnosis in a prospective series of adult and pediatric patients who presented with bone marrow failure without clear overall clinical picture of FA. Sixty-six patients were classified into three groups: (1) bone marrow failure likely to be congenital, based on dysmorphic features or a family history [n=18], (2) aplastic anemia likely to be idiopathic [n=32], (3) patients with intermediate clinical features not classified into the former groups [n=16]. Of note, FA patients with typical clinical features were not included in the present study. FA diagnosis was evaluated using chromosome breakage test and FANCD2 immunoblot in PHA-stimulated-PBL. In addition, skin primary fibroblasts were analysed in order to overcome potential hematopoietic FA reversion. For that purpose, and considering that chromosome breakage tests are barely efficient in fibroblasts, we used FANCD2 immunoblot and also developped a new flow cytometry test based on MMC-sensitivity in fibroblasts (to detect downstream FA/BRCA groups). Using these approaches, we detected FA in 4 previously undiagnosed patients: a 35-years old patient from the congenital-like group; a 10-years old patient presenting as an idiopathic aplastic anemia without any FA signs; and two patients from the intermediate group: a 10-years old patient with an isolated thrombocytopenia, and a 50-years old patient presenting with pancytopenia/MDS and complete hematopoietic reversion. Importantly, FA diagnosis was definitely excluded in all other patients. In conclusion, we could identify a few unexpected FA cases in a series of patients with bone marrow failure. Therefore, the comprehensive use of a large set of tests is useful for accurate FA diagnosis. Classical chromosomal breakage tests in PBL appeared to be sufficient to exclude FA in idiopathic aplastic anemia, whereas fibroblast analysis can be necessary to definitely diagnose or exclude FA in other patients.

Genotype-Phenotype Associations in Patients with Fanconi Anemia: National Cancer Institute Cohort

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 4-5
Author(s):  
Burak Altintas ◽  
Neelam Giri ◽  
Lisa J. McReynolds ◽  
Blanche P. Alter
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Upper Limb ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Skin Pigmentation ◽  
Autosomal Recessive Disorder ◽  
Gene Variants ◽  
Pathway Gene ◽  
Significant Difference

Fanconi anemia (FA) is a predominantly autosomal recessive disorder resulting from mutations in one of >22 genes involved in the FA/BRCA DNA repair pathway. FA is characterized by multiple congenital abnormalities, progressive bone marrow failure (BMF) and cancer predisposition. Genetic heterogeneity and diverse clinical presentations challenge early diagnosis and optimal management. We previously reviewed the genotype-phenotype associations in FA from literature cases (Fiesco-Roa MO et al. Blood Rev. 2019). We now report the results from the NCI cohort. We studied 147 patients with FA in the NCI inherited bone marrow failure syndromes Cohort Study (ClinicalTrials.gov, NCT00027274) to explore genotype phenotype associations by genes, location in the FA/BRCA pathway (upstream, ID complex, downstream), and compare information on the clinic cohort (CC) and field cohort (FC) patients. 57 patients (CC) were evaluated at the NIH Clinical Center between 2002 and 2020. Details on 90 patients in the FC were obtained from the review of medical records. The sex ratio (M:F) was similar (0.6:1 and 0.8:1). Patients in the FC were younger than in the CC (p=0.004) with median ages 27 (3-68) years for the CC and 19 (0-57) for the FC. The main genotypes in the CC were 59% FANCA, 17% FANCC, 6% FANCI and in the FC were 60% FANCA, 13% FANCC and 8% FANCG. At least one FA type physical abnormality was present in all CC patients and 73/79 (92%) FC patients (phenotype data not reported on 11 FC patients). >3/8 VACTERL-H features (Vertebral, Anal, Cardiac, Tracheo-esophageal fistula (TEF), Esophageal or duodenal atresia, Renal, upper Limb (radial ray) and Hydrocephalus) were present in 32% of CC patients and 16% of FC (p=0.04). At least 4/6 PHENOS features (skin Pigmentation, small Head, small Eyes, other central Nervous system (CNS) anomalies, Otology and Short stature) were present in 54% of CC patients and 34% FC (p=0.02). The types and frequencies of phenotypic abnormalities are shown in figure 1. 17 patients in the CC (30%) and 10 in the FC (13%) had both VACTERL-H and PHENOS (p=0.01). We excluded patients with unknown genotype or phenotype from further analysis. In the CC, cardiac abnormalities were more common in patients with FANCI or ID complex gene variants than in all others (p=0.02 and 0.001, respectively) as were VACTERL-H and structural CNS abnormalities in patients with ID complex variants (p=0.03 and 0.006, respectively). In the FC, VACTERL-H, imperforate anus and hydrocephalus were more common in patients with FANCD1 genotype (p=0.03, 0.009 and 0.004, respectively) and downstream pathway gene variants (p=0.004, <0.001 and 0.03, respectively). PHENOS, renal and neurodevelopmental abnormalities were less common in patients with upstream genes variants (p=0.001, 0.009 and <0.001, respectively). Upper limb abnormalities were less common in patients with FANCC genotype (p=0.007). BMF was present in 121/147 (88%) patients; 33% had been transfusion-dependent and 26% received androgen therapy. Clonal cytogenetic abnormalities were seen in 30%; 17% developed myelodysplastic syndrome at a median age of 17 (1.4-44) years and 6 patients developed acute myeloid leukemia at a median age of 19 (12-29) years. 72 (49%) patients underwent bone marrow transplant at a median age of 9.5 (1.5-44) years for BMF, MDS or leukemia. There was no significant difference between the FC and CC. The median survival age of our cohort is 38 (95% CI 34-43) years and at least 80% of our patients are >18 years of age. Kaplan-Meier survival estimates are presented in figure 2. Solid tumors developed in 30/135 (22%) patients with available data; median age at first cancer was 30 (2-44) years. The most common tumor was head and neck squamous cell carcinoma (n=15 patients), followed by skin (n=8) and anogenital cancers (n=6); many patients developed multiple cancers. Detailed hematologic, cancer, endocrine outcomes and survival analyses are ongoing. Overall, renal and upper limb abnormalities were reported in most of the patients in both CC and FC, as shown previously (Alter BP et al. Mol Syndromol. 2013). Data from the CC were more complete than from the review of charts from the FC highlighting that the clinical in person evaluation of patients provides detailed characterization of FA phenotypes and more accurate assessment of genotype-phenotype associations. This will facilitate timely diagnosis, surveillance and clinical management of patients with FA. Disclosures No relevant conflicts of interest to declare.

Links Between DNA Damage and Metabolism, Pathways Causing Bone Marrow Failure in Fanconi Anemia, and Therapeutic Implications

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. SCI-3-SCI-3
Author(s):  
Ketan J. Patel
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Genetic Defect ◽  
Scientific Session ◽  
Ethanol Exposure ◽  
Southeast Asians ◽  
Therapeutic Implications ◽  
Blood Stem Cells

Abstract Abstract SCI-3 Recent work from my lab has discovered that metabolism generates reactive aldehydes. These reactive molecules are potent damagers of DNA. The consequences of this are revealed by the inactivation of enzymes that detoxify these aldehydes and the Fanconi anemia DNA repair pathway in mice and vertebrate cell lines. The scientific session presentation will discuss this work and recent unpublished research on how natural aldehydes damage blood stem cells. This work has consequences for understanding how metabolism and ethanol exposure can be genotoxic, particularly in the vast population of Southeast Asians carrying a genetic defect in aldehyde catabolism (“pink flushers”). It is also relevant to the emergence of bone marrow failure and leukemia in Fanconi anemia. Disclosures: No relevant conflicts of interest to declare.

Metastatic Breast Cancer in a Patient with Fanconi Anemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5164-5164
Author(s):  
Jeffrey Graham ◽  
Debjani Grenier ◽  
Arjuna Ponnampalam
Keyword(s):  
Breast Cancer ◽  
Bone Marrow ◽  
Fanconi Anemia ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Genetic Defect ◽  
Susceptibility Gene ◽  
Metastatic Breast ◽  
Morphological Evidence ◽  
Breast Cancer Susceptibility Gene

Abstract Fanconi anemia (FA) is a rare inherited disorder characterized by progressive bone marrow failure, congenital malformations and a propensity for developing malignancies at an early age. The underlying genetic defect in FA creates a state of cellular hypersensitivity to many traditional chemotherapy agents, making the treatment of malignancies in this population particularly challenging. We describe a 42-year-old female who presented with a solitary mass in her left breast. Core biopsy revealed an invasive ductal carcinoma that did not express estrogen (ER) or progesterone receptors (PR), but did express human epidermal growth factor receptor 2 (HER2). Staging work-up revealed diffuse skeletal metastatic disease. At her initial consultation with medical oncology, she was discovered to be pancytopenic. Further history revealed a sibling with aplastic anemia and that she had undergone chromosomal breakage testing for FA in the past, which was subsequently confirmed to be positive. She underwent a bone marrow aspirate and biopsy that showed metastatic marrow infiltration by non-hematopoietic cells. In addition there was morphological evidence of dyserythropoiesis and cytogenetic abnormalities on karyotyping, features suggestive of FA. She was initially started on trastuzumab monotherapy. Low dose radiation therapy was added due to local tumor progression. Combined HER2 directed therapy was to be implemented, but was held due to a functional decline in the patient. To date, she has not received definitive genetic testing to determine which FA subgroup she belongs to. This case highlights two important aspects of FA. The first is the inherent increase in susceptibility to neoplasms in this group, including solid tumors such as breast cancer. The genes associated with FA are involved in deoxyribonucleic acid (DNA) repair pathways, including mutations in the breast cancer susceptibility gene, BRCA2. The second is the heightened sensitivity to the toxic effects of many standard chemotherapy and radiation treatments. This creates unique challenges in the treatment of malignancies in this population and stresses the importance of targeted therapies. Disclosures No relevant conflicts of interest to declare.

scholarly journals Case Series: Bone Marrow Failure in Teen Siblings with Unique RPS19 Variant

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4311-4311
Author(s):  
Alexandra Prosser ◽  
Erin Marie Hall ◽  
Lauren E Amos
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Bone Marrow Biopsy ◽  
Untranslated Region ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Case Series ◽  
Laboratory Evaluation ◽  
Unrelated Donor ◽  
Chromosomal Breakage

Abstract Aplastic anemia occurs when there is a lack of hematopoiesis in the bone marrow leading to peripheral pancytopenia. It is a rare entity and is often idiopathic. However, upon presentation, inherited bone marrow failure syndromes and acquired etiologies must be considered. Investigating causality is particularly important when multiple family members are affected, especially in a short amount of time. It is also essential to identify novel causative genetic variants of bone marrow failure in order to direct treatment in these patients. In our case series, we describe two siblings who presented two weeks apart with severe pancytopenia. The first patient is a 13 year-old non-binary female (prefers pronouns they/them/theirs) who presented from clinic after getting routine labs with white blood cell (WBC) 1.83 x10(3)/mcL, absolute neutrophil count (ANC) 0.16 x10(3)/mcL, hemoglobin (Hgb) 4.8 gm/dL, platelet 13 x10(3)/mcL. Bone marrow biopsy revealed marked hypocellularity (0-10%) with hypoplasia. The second patient is their 16 year-old brother who presented two weeks later with new-onset petechial rash and was found to have WBC 4 x10(3)/mcL, ANC 2.19 x10(3)/mcL, Hgb 6.3 gm/dL, platelet 16 x10(3)/mcL. His bone marrow biopsy demonstrated variable cellularity (10-70%), but after months of transfusion-dependence he met criteria for severe aplastic anemia. Laboratory evaluation for acquired etiologies such as infection was negative. Both patients had a shared medical history of depression briefly treated with fluoxetine, but otherwise no potential medical triggers or environmental exposures were identified. Telomere length analysis was normal and chromosomal breakage analysis was negative. Upon genetic evaluation, both patients were found to have a heterozygous variant of unknown significance of RPS19 (c.-163>T), which substitutes a moderately conserved nucleotide in the noncoding exon 1 in 5' untranslated region of RPS19. Although this variant has not been classified as pathogenic, three other variants in the 5' untranslated region of RPS19 have been reported in patients with Diamond-Blackfan Anemia (DBA). In contrast to classic DBA, these patients did not present in infancy or early childhood. Likewise, they lacked congenital anomalies or other classic phenotypic characteristics of disease. Lab studies showed normal erythrocyte adenosine deaminase levels, though it should be noted that these levels could not be obtained prior to transfusion in both patients. Given lack of other identified etiologies, however, an inherited bone marrow failure syndrome, possibly due to this variant, was presumed. With suspected but potentially unidentifiable genetic predisposition, matched sibling donor transplant was deferred and both patients underwent a matched unrelated donor bone marrow transplant with reduced-intensity conditioning. With these cases, we aim to share a unique presentation of aplastic anemia that reveals a potentially novel pathogenic variant as well as to provide our approach to medical management in pediatric aplastic anemia in the setting of uncertainty. Identification of other patients with bone marrow failure and this genetic variant will be important to determine its pathogenicity. Disclosures No relevant conflicts of interest to declare.

scholarly journals Genomic Instability in Fanconi Anemia Results from a Combination of Chromosome Mis-Segregation in Mitosis and Unresolved Interphase DNA Damage

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 357-357 ◽  
Author(s):  
Donna Cerabona ◽  
Zahi Abdul Sater ◽  
Rikki Enzor ◽  
Grzegorz Nalepa
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Genomic Instability ◽  
Fanconi Anemia ◽  
Chromosomal Instability ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Genetic Disorder ◽  
Biological Significance

Abstract Fanconi anemia (FA) is a complex genetic disorder characterized by bone marrow failure, multiple congenital anomalies, and genomic instability resulting in predisposition to cancer. Disruption of the FA signaling network impairs multiple genome-housekeeping processes, including DNA damage recognition and repair in interphase, DNA replication as well as high-fidelity chromosome segregation during mitosis. Recent data published by several groups, including our work (J Clin Invest 2013; 123: 3839-3847), implicated FA signaling in the control of several cell division events essential for chromosomal stability, including the spindle assembly checkpoint (SAC), centrosome maintenance, resolution of ultrafine anaphase bridges and cytokinesis. Understanding the mechanistic origins of chromosomal instability leading to carcinogenesis and bone marrow failure has important scientific and clinical implications. However, the relative contribution of the interphase and mitotic events leading to genomic instability in Fanconi anemia has not been systematically evaluated. In this work, we dissected the origins and mechanistic significance of chromosomal instability in Fanconi anemia ex vivo and in vivo. We employed the cytochalasin micronucleus assay to quantify the patterns of spontaneous and chemotherapy-induced genomic lesions in FA-A patient-derived primary fibroblasts and Fancc-/- mouse embryonic fibroblasts (MEFs). In this assay, dividing cells are treated with cytochalasin to inhibit cytokinesis and generate binucleated daughter cells. The presence of micronuclei in the resulting cells is indicative of genomic instability caused by either interphase DNA damage or chromosome mis-segregation. Centromere-negative micronuclei (CNMs) represent chromosomal fragments due to unresolved ds-DNA damage. Centromere-positive micronuclei (CPMs) result from whole-chromosome mis-segregation during mitosis. The frequency of both CPMs and CNMs was significantly increased in FA-deficient human and murine cells compared to gene-corrected isogenic control cells. These results indicate that genomic instability in FA is caused by a combination of interphase DNA damage and disordered mitosis. We confirmed the biological significance of these findings by showing that FA patient cells are hypersensitive to low concentrations of taxol (a spindle checkpoint-activating chemotherapeutic) similarly to mitomycin C (a cross-linking agent). Finally, we found increased frequency of micronuclei in Fancc-/- murine red blood cells compared to age-matched wild-type mice, which indicates that spontaneous chromosome mis-segregation occurs in FA-deficient bone marrow in vivo. Our study supports the emerging model of the FA family of proteins as holistic guardians of the genome during interphase and mitosis (see figure based on F1000Prime Rep. 2014; 6: 23, modified). This model furthers our understanding of genomic instability in Fanconi anemia and FA-deficient cancers, and opens new inroads towards targeted therapeutic interventions in these diseases. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

A High Throughput Small Molecule Screening Method for Fanconi Anemia.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3191-3191
Author(s):  
Anur Praveen ◽  
Jeffrey W Tyner ◽  
Scott Vanderwerff ◽  
Winifred Keeble ◽  
Grover C. Bagby
Keyword(s):  
Bone Marrow ◽  
Small Molecules ◽  
High Throughput ◽  
Fanconi Anemia ◽  
High Throughput Screening ◽  
Conflicts Of Interest ◽  
Lentiviral Vector ◽  
Bone Marrow Failure ◽  
Screening Method ◽  
Hematopoietic Cells

Abstract Abstract 3191 Poster Board III-128 The Fanconi Anemia (FA) proteins play an important role in regulating genome stability, but there is little evidence that the loss of the genoprotection per se, in FA cells accounts for the molecular pathogenesis of the bone marrow failure characteristic of this disease. Indeed, there is evidence that at least some of these proteins are multifunctional and participate in canonical signaling pathways in hematopoietic cells. FANCC deficient cells, for example, are hypersensitive to the apoptotic effects of TNFαa. In addition, FA-C cells over-produce TNFαa at least in part because FANCC ordinarily suppress the activation potential of toll-like receptor 8 (TLR8) (abstract submitted to this meeting). There is clear evidence that over-production of TNFαa and hypersensitivity to TNFαa in hematopoietic cells of Fancc-/- mice results in bone marrow hypoplasia (Sejas et al, 2007 and Zhang et al 2007) and that long-term ex-vivo exposure of murine Fancc -/- hematopoietic cells to both growth factors and TNFαa results in the evolution of cytogenetically marked preleukemic clones (Li et al 2007). Because the hematopoietic phenotype of FA may evolve from the overproduction of precisely the cytokine to which FA stem cells are hypersensitive, we reasoned that suppression of TNFαa production by FA cells might enhance hematopoiesis. So we sought to develop a strategy to permit high throughput screening of small molecules designed to suppress TNFαa production specifically in FANCC deficient cells. Methods THP1 Blue cells (THP1B) have a stably integrated NF-kappaB reporter gene, secreted embryonic alkaline phosphatase (SEAP) and express SEAP and TNFαa in response to TLR ligands including the TLR8 ligand (R848). Each of five samples of THP1B cells were transduced with one of five lentiviral vectors expressing FANCC targeted shRNA. One of these vectors suppressed FANCC expression (by immunoblotting and RT-RT-PCR), suppressed FANCD2 levels in MMC exposed THP1B cells, induced chromosomal instability in the MMC assay and markedly enhanced R848-induced TNFαa production when compared to THP1B cells transduced with a non-targeted shRNA lentiviral vector. In multiwell plates, THP1B-shFANCC cells were exposed to multiple doses one of 81 small molecules including steroid hormones and inhibitors of tyrosine or serine threonine kinases. TNFαa (ELISA) and SEAP (QUANTI-blue colorimetry) were quantified in the supernatant media 24 hours after exposure to R848. Results 15 agents suppressed SEAP production without cytotoxicity and all of these suppressed TNFαa production as well. The same agents suppressed TNFαa production in two patient-derived FANCC-deficient cell lines (HSC536 and PD149) both of which over-express TNFαa in the ground state. Four p38 inhibitors (100nM-10μM) were analyzed and at 500 nM all suppressed SEAP and TNFαa by 90% or more. The Src family kinase inhibitor, Dasatinib (500nM) was also effective. Using Fancc-deficient mice exposed to TLR activating agents, in vivo preclinical studies designed to test the effectiveness of Dasatinib and one p38 inhibitor are underway, as are mechanistically focused multiplex assays in which known target molecules of these agents are suppressed using RNAi. Conclusions We have developed a reliable screening tool based upon the TNFαa-overproduction phenotype of FANCC deficient cells. Using it, we have identified inhibitors of p38 MAPK and Src family kinases that suppress TNFαa-overproduction in patient derived FANCC-deficient cells. The identification of these agents provides not only an opportunity to discover novel biochemical roles played by FANCC in innate immunity but also a strong rationale for evaluating such agents in preclinical models for marrow failure in FA. Disclosures No relevant conflicts of interest to declare.

KIT Blockade Is Sufficient to Sustain Donor Hematopoietic Stem Cell Engraftment in Fanconi Anemia Mice

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1206-1206
Author(s):  
Shanmuganathan Chandrakasan ◽  
Rajeswari Jayavaradhan ◽  
Ernst John ◽  
Archana Shrestha ◽  
Phillip Dexheimer ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Fanconi Anemia ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Conditioning Regimen ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Increased Risk

Abstract Background: Fanconi anemia (FA) is the most common cause of inherited bone marrow failure (BMF). Currently, the only curative option for the BMF in FA is an allogenic hematopoietic stem cell transplant (HSCT). However, due to the underlying DNA repair defect, FA patients poorly tolerate alkylating chemotherapy or irradiation based conditioning, which is necessary for donor engraftment. However, this results in significant short and long term morbidity/mortality and augments the inherent increased risk of malignancies in FA patients. To overcome the adverse effects associated with alkylating conditioning agents, alternate experimental approaches exploiting the inherent hematopoietic stem cell (HSC) defect in FA are of utmost clinical necessity. Objective: To develop a safe KIT blocking antibody (KIT-Ab) based HSCT conditioning regimen for FA that does not involve chemotherapy or irradiation. Method: High purity KIT-Ab was made from the ACK2 hybridoma and its specificity to KIT binding was validated using mast cell assay. Baseline peripheral blood cells and the bone marrow hematopoietic stem and progenitor cell (HSPC) compartment (Lin-Kit+Sca+ and Lin-Kit+Sca+CD150+CD48- cells) of FANCA-/- and FANCD2-/- murine models were analyzed. Mechanistic studies using sorted FA bone marrow HSPC were performed ex vivo. This was followed by definitive primary and secondary transplants experiments following injection of KIT-Ab. Results: Several features of FA hematopoietic stem/progenitor cells (HSPC) suggested their susceptibility to KIT-Ab blockade-mediated killing: (a) Expression of KIT was significantly lower in FANCA-/- HSPC, while expression of its ligand was higher in bone marrow stroma; (b) Moreover, genes associated with apoptosis/senescence, stress and inflammatory signaling that were upregulated in WT-HSPC following KIT-Ab blockade, were upregulated in FANCA-/- HSPC at baseline; (c) Furthermore, FANCA-/- HSPC demonstrated increased susceptibility to KIT-Ab mediated apoptosis and had a reduced proliferative capacity. In-vivo studies following ACK2 injection showed a marked reduction of colony-forming units (CFU-C) from both FANCA-/- and FANCD2-/- mice one week following injection, when compared to WT mice (48% and 76% decrease in CFU-C, respectively). Based on these findings, we evaluated the role of ACK2 as a sole HSCT conditioning regimen in FANCA-/- and FANCD2-/- mice. Indeed, definitive HSCT in both FANCA-/- and FANCD2-/- mice using KIT-Ab based conditioning resulted in donor HSC engraftment with multi-lineage chimerism, which progressively increased to 22-24% by 4-months, and was sustained in secondary transplants. Overall, we show that KIT-blockade alone is an adequate non-genotoxic HSPC-targeted conditioning in FA mice, and its clinical translation could circumvent the extensive transplant-related morbidity/mortality in this disease. Disclosures No relevant conflicts of interest to declare.

Impaired Bone Marrow Microenvironment in Fanconi Anemia Murine Model Is Responsible for the Defective Hematopoietic Stem/ Progenitor Cell Mobilization.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3629-3629
Author(s):  
Yan Li ◽  
Shi Chen ◽  
Yongzheng He ◽  
Xiaohong Li ◽  
Fengchun Yang
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Fanconi Anemia ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Genetic Disorder ◽  
Bone Marrow Microenvironment ◽  
Hematopoietic Stem

Abstract Abstract 3629 Poster Board III-565 Fanconi anemia (FA) is a heterogeneous genetic disorder characterized by progressive bone marrow failure (BMF) and acquisition of malignancies. The only cure for BMF is a human leukocyte antigen (HLA)-matched BM transplantation from a family member or autologous stem cells before BMF develops. Therefore, mobilization of hematopoietic stem/progenitor cells (HSPCs) from BM into peripheral blood (PB) for collection has been a prerequisite for the therapy. However, patients with FA show a markedly decreased HSPC mobilization in response to the traditional mobilizing drug G-CSF and the mechanism(s) underlying the defect remains unknown. Mesenchymal stem/progenitor cells (MSPCs) have been known to be the common progenitor of a variety of cellular components in the bone marrow microenvironment. MSPCs express/secrete cytokines, extracellular matrix proteins and cell adhesion molecules, which regulate the homing, migration, proliferation and survival of HSPCs in vitro and in vivo. Recently, we reported that Fancg-/- MSPCs have a defect in hematopoietic supportive activity both in vitro and in vivo (Li et al. Blood, 2009). In the current studies, we show that Fancg-/- MSPCs have significant reduction in HSPC recruitment as compared to WT MSPCs in a transwell assay. Furthermore, Fancg-/- MSPCs have an alteration in the production of multiple cytokines/chemokines. Application of a neutralizing antibody to the cytokine blocked WT MSPC mediated HSPC migration in vitro. Furthermore, administration of the specific cytokine significantly increased HSPC mobilization in the Fancg-/- mice in vivo. These results demonstrated that an impaired BM microenvironment, specifically MSPCs in Fancg-/- mice, is contributory to defective HSPC mobilization. This study provides evidence of alternative clinical therapeutics for the mobilization of HSPCs in FA patients. Disclosures: No relevant conflicts of interest to declare.

A novel diagnostic screen for defects in the Fanconi anemia pathway

Blood ◽  
2002 ◽  
Vol 100 (13) ◽  
pp. 4649-4654 ◽  
Author(s):  
Akiko Shimamura ◽  
Rocio Montes de Oca ◽  
John L. Svenson ◽  
Nicholas Haining ◽  
Lisa A. Moreau ◽  
...  
Keyword(s):  
At Risk ◽  
Bone Marrow ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Chromosomal Breakage ◽  
Retroviral Gene Transfer ◽  
Bone Marrow Failure Syndromes ◽  
Populations At Risk ◽  
Healthy Control ◽  
Or Gene

Fanconi anemia (FA) is an autosomal recessive chromosomal instability syndrome characterized by congenital abnormalities, progressive bone marrow failure, and cancer predisposition. Although patients with FA are candidates for bone marrow transplantation or gene therapy, their phenotypic heterogeneity can delay or obscure diagnosis. The current diagnostic test for FA consists of cytogenetic quantitation of chromosomal breakage in response to diepoxybutane (DEB) or mitomycin C (MMC). Recent studies have elucidated a biochemical pathway for Fanconi anemia that culminates in the monoubiquitination of the FANCD2 protein. In the current study, we develop a new rapid diagnostic and subtyping FA assay amenable for screening broad populations at risk of FA. Primary lymphocytes were assayed for FANCD2 monoubiquitination by immunoblot. The absence of the monoubiquitinated FANCD2 isoform correlated with the diagnosis of FA by DEB testing in 11 known patients with FA, 37 patients referred for possible FA, and 29 healthy control subjects. Monoubiquitination of FANCD2 was normal in other bone marrow failure syndromes and chromosomal breakage syndromes. A combination of retroviral gene transfer and FANCD2 immunoblotting provides a rapid subtyping assay for patients newly diagnosed with FA. These new FA screening assays would allow efficient testing of broad populations at risk.

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