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.

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.

Impaired Spindle Assembly Checkpoint In Vivo Promotes MDS/AML in a Novel Mouse Model of Fanconi Anemia

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1211-1211
Author(s):  
Donna Cerabona ◽  
Zahi Abdul Sater ◽  
Elizabeth Sierra Potchanant ◽  
Ying He ◽  
Zejin Sun ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mouse Model ◽  
Genomic Instability ◽  
Chromosome Segregation ◽  
Fanconi Anemia ◽  
Chromosomal Instability ◽  
Spindle Assembly Checkpoint ◽  
Bone Marrow Failure

Abstract The Fanconi anemia (FA/BRCA) signaling network prevents bone marrow failure and cancer by protecting genomic integrity. Biallelic germline mutations within this gene network result in Fanconi anemia, an inherited bone marrow failure syndrome characterized by genomic instability and a predisposition to bone marrow failure, myelodysplasia and cancer, particularly acute myelogenous leukemia (AML). Heterozygous inborn mutations in the BRCA branch of FA network increase risk of breast and ovarian cancers as well as other tumors, and somatic mutations of FA/BRCA genes occur in malignancies in non-Fanconi patients. Thus, disruption of FA/BRCA signaling promotes malignancies in the inherited genetic syndromes and in the general population. The FA/BRCA network functions as a genome gatekeeper throughout the cell cycle. In interphase, the FA/BRCA network provides a crucial line of defense against mutagenesis by coordinating DNA damage response to a variety of genotoxic insults, from endogenous aldehydes to replication errors and mutagen exposure. Less is known about the role of the FA/BRCA pathway during mitosis. However, FA signaling has recently been implicated in multiple aspects of cell division, including the spindle assembly checkpoint (SAC) that ensures high-fidelity chromosome segregation at metaphase to anaphase transition; cytokinesis; centrosome maintenance and repair of ultrafine anaphase bridges. Although chromosomal instability due to mitotic errors is a hallmark of cancer, the in vivo contribution of abnormal mitosis to malignant transformation of FA-deficient hematopoietic cells remains unknown. To determine whether error-prone chromosome segregation upon loss of FA signaling contributes to abnormal hematopoiesis and cancer, we generated a novel murine FA model by genetically weakening the SAC in the FA-deficient background. The resulting mice were viable and born at expected Mendelian ratios, but exhibited increased baseline in vivo chromosomal instability evidenced by elevated red blood cell micronucleation, increased frequency of chromosome missegregation and DNA breakage in microscopy-based cytome assays, and augmented bone marrow karyotype instability. Importantly, unlike FA or SAC control animals, the FA-SAC mice were prone to premature death due to the development of myelodysplasia and AML at young age, recapitulating disease manifestations of human Fanconi anemia. This study provides the in vivo evidence supporting the essential role of compromised chromosome segregation in the development of myelodysplasia and acute leukemia due to impaired FA signaling. Our observations provide novel insights into complex mechanisms of genomic instability and carcinogenesis due to FA deficiency. Impaired mitosis is a well-established therapeutic target, and our independent ex vivo experiments using FA patient-derived primary cells show that exposure to antimitotic chemotherapeutics is synthetic lethal with loss of the FA network. Thus, our findings may have implications for future precision strategies against FA-deficient, chromosomally unstable hematopoietic cancers. The FA-SAC mouse model offers a preclinical platform to systematically test this hypothesis in vivo. Disclosures No relevant conflicts of interest to declare.

Phenotypic correction of Fanconi anemia group C knockout mice

Blood ◽  
2000 ◽  
Vol 95 (2) ◽  
pp. 700-704 ◽  
Author(s):  
Kimberly A. Gush ◽  
Kai-Ling Fu ◽  
Markus Grompe ◽  
Christopher E. Walsh
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Mitomycin C ◽  
Fanconi Anemia ◽  
Bone Marrow Cells ◽  
Bone Marrow Failure ◽  
Genetic Disorder ◽  
Hematopoietic Stem ◽  
Marrow Cells

Fanconi anemia (FA) is a genetic disorder characterized by bone marrow failure, congenital anomalies, and a predisposition to malignancy. FA cells demonstrate hypersensitivity to DNA cross-linking agents, such as mitomycin C (MMC). Mice with a targeted disruption of the FANCC gene (fancc −/− nullizygous mice) exhibit many of the characteristic features of FA and provide a valuable tool for testing novel therapeutic strategies. We have exploited the inherent hypersensitivity offancc −/− hematopoietic cells to assay for phenotypic correction following transfer of the FANCC complementary DNA (cDNA) into bone marrow cells. Murine fancc −/− bone marrow cells were transduced with the use of retrovirus carrying the humanfancc cDNA and injected into lethally irradiated recipients. Mitomycin C (MMC) dosing, known to induce pancytopenia, was used to challenge the transplanted animals. Phenotypic correction was determined by assessment of peripheral blood counts. Mice that received cells transduced with virus carrying the wild-type gene maintained normal blood counts following MMC administration. All nullizygous control animals receiving MMC exhibited pancytopenia shortly before death. Clonogenic assay and polymerase chain reaction analysis confirmed gene transfer of progenitor cells. These results indicate that selective pressure promotes in vivo enrichment offancc-transduced hematopoietic stem/progenitor cells. In addition, MMC resistance coupled with detection of the transgene in secondary recipients suggests transduction and phenotypic correction of long-term repopulating stem cells.

Gender-related differences in the oxidant state of cells in Fanconi anemia heterozygotes

2011 ◽  
Vol 392 (7) ◽  
Author(s):  
Sandra Petrovic ◽  
Andreja Leskovac ◽  
Jelena Kotur-Stevuljevic ◽  
Jelena Joksic ◽  
Marija Guc-Scekic ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Genetic Disorder ◽  
Significant Difference ◽  
Heterozygous Carriers ◽  
Male Carriers ◽  
Female Carriers

Abstract Fanconi anemia (FA) is a rare cancer-prone genetic disorder characterized by progressive bone marrow failure, chromosomal instability and redox abnormalities. There is much biochemical and genetic data, which strongly suggest that FA cells experience increased oxidative stress. The present study was designed to elucidate if differences in oxidant state exist between control, idiopathic bone marrow failure (idBMF) and FA cells, and to analyze oxidant state of cells in FA heterozygous carriers as well. The results of the present study confirm an in vivo prooxidant state of FA cells and clearly indicate that FA patients can be distinguished from idBMF patients based on the oxidant state of cells. Female carriers of FA mutation also exhibited hallmarks of an in vivo prooxidant state behaving in a similar manner as FA patients. On the other hand, the oxidant state of cells in FA male carriers and idBMF families failed to show any significant difference vs. controls. We demonstrate that the altered oxidant state influences susceptibility of cells to apoptosis in both FA patients and female carriers. The results highlight the need for further research of the possible role of mitochondrial inheritance in the pathogenesis of FA.

Tumor Necrosis Factor-Mediated Inflammation Exacerbates Genomic Instability and Promotes Leukemia Development.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4265-4265
Author(s):  
June Li ◽  
Daniel P. Sejas ◽  
Xiaoling Zhang ◽  
Reena Rani ◽  
Qishen Pang
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Tumor Necrosis Factor ◽  
Genomic Instability ◽  
Tumor Necrosis ◽  
Bone Marrow Failure ◽  
Genetic Disorder ◽  
Leukemic Transformation ◽  
Leukemia Development ◽  
Necrosis Factor

Abstract A correlation has been shown between elevated circulating pro-inflammatory cytokines and anemia in patients with leukemia-related diseases but direct evidence for the mechanistic link between inflammation and leukemia is lacking. We have investigated the role of the pro-inflammatory cytokine tumor necrosis factor a (TNF-a) in leukemic development using the disease model of Fanconi anemia, a genetic disorder clinically characterized by congenital anomalies, progressive bone marrow failure, and a high risk of developing leukemia and other cancers. We demonstrate that long-term TNF-a exposure of Fanconi bone marrow progenitors enhances inflammatory response, promotes clonal proliferation, and ultimately leads to leukemia development. NF-kB activation is required for TNF-a-promoted progenitor growth and early stage of leukemia development but is dispensable for the maintenance of leukemic transformation. Pharmacological elimination of TNF-a-induced reactive oxygen species reduces inflammation and delays leukemia development, suggesting that oxidative damage may serve as a link between inflammation and leukemia. In addition, TNF-a-promoted leukemic cells show persistent DNA damage response and increased genomic instability. Furthermore, correction of Fanconi genetic deficiency prevents clonal progenitor proliferation and leukemic transformation by eliminating oxidative DNA damage. Thus, inflammation can exacerbate genomic instability and contribute to leukemia development. This may explain, at least in part, why cells with genomic instability have high predisposition to cancer. Our study underscores therapeutic value of anti-oxidants and anti-inflammation towards tumorigenesis.

scholarly journals Bone Marrow Failure in the Fanconi Anemia Group C Mouse Model After DNA Damage

Blood ◽  
1998 ◽  
Vol 91 (8) ◽  
pp. 2737-2744 ◽  
Author(s):  
Madeleine Carreau ◽  
Olga I. Gan ◽  
Lili Liu ◽  
Monica Doedens ◽  
Colin McKerlie ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Dna Damage ◽  
Mouse Model ◽  
Fanconi Anemia ◽  
Bone Marrow Failure ◽  
Blood Parameters ◽  
Cross Linking ◽  
Inherited Disease

Fanconi anemia (FA) is a pleiotropic inherited disease that causes bone marrow failure in children. However, the specific involvement of FA genes in hematopoiesis and their relation to bone marrow (BM) failure is still unclear. The increased sensitivity of FA cells to DNA cross-linking agents such as mitomycin C (MMC) and diepoxybutane (DEB), including the induction of chromosomal aberrations and delay in the G2 phase of the cell cycle, have suggested a role for the FA genes in DNA repair, cell cycle regulation, and apoptosis. We previously reported the cloning of the FA group C gene (FAC) and the generation of a Fac mouse model. Surprisingly, the Fac −/− mice did not show any of the hematologic defects found in FA patients. To better understand the relationship of FA gene functions to BM failure, we have analyzed the in vivo effect of an FA-specific DNA damaging agent in Fac −/− mice. The mice were found to be highly sensitive to DNA cross-linking agents; acute exposure to MMC produced a marked BM hypoplasia and degeneration of proliferative tissues and caused death within a few days of treatment. However, sequential, nonlethal doses of MMC caused a progressive decrease in all peripheral blood parameters of Fac −/− mice. This treatment targeted specifically the BM compartment, with no effect on other proliferative tissues. The progressive pancytopenia resulted from a reduction in the number of early and committed hematopoietic progenitors. These results indicate that the FA genes are involved in the physiologic response of hematopoietic progenitor cells to DNA damage.

HSC Exit From Dormancy Provokes De Novo DNA Damage, Leading To Bone Marrow Failure If Unresolved By The Fanconi Anemia Pathway

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 799-799
Author(s):  
Dagmar Walter ◽  
Amelie Lier ◽  
Anja Geiselhart ◽  
Sina Huntscha ◽  
David Brocks ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Aplastic Anemia ◽  
Dna Damage Response ◽  
Fanconi Anemia ◽  
De Novo ◽  
Bone Marrow Failure ◽  
Damage Response ◽  
Fold Induction

Abstract Long-term quiescence has been proposed to preserve the genomic stability of hematopoietic stem cells (HSCs) during aging. The current models of HSC aging are limited in their ability to observe both DNA damage in vivo and the consequences of this damage upon hematopoiesis. Fanconi Anemia (FA) is a hereditary multisystem disorder, characterized by defective DNA damage response and progressive bone marrow failure in most patients. However, the existing genetic models of FA do not develop aplastic anemia, suggesting that cell-extrinsic factors may play a causal role. We sought to identify whether physiologic mediators of HSC activation could be used as agonists to provoke DNA damage and HSC attrition in vivo. Mice were treated with a range of agonists that promote the in vivo exit of HSC from a dormant state into active cycling (polyI:polyC; Interferon-α; G-CSF; TPO; and serial bleeding). Highly purified HSC demonstrated a rapid 3-5-fold induction of DNA damage after treatment with all agonists (p<0.01), as assessed by both enumerating γ-H2AX foci and by alkaline comet assay. Mechanistically, stress-induced exit from quiescence correlated with increased mitochondrial metabolism in HSC, as evaluated by elevated mitochondrial membrane potential (2-fold increased, p<0.01) and superoxide levels (1.5-fold increased, p<0.05). Critically, we could directly implicate these reactive oxygen species in DNA damage as we observed a 1.4-fold increase in 8-Oxo-dG lesions in HSC that had been activated into cycle in vivo(p<0.05). At 48 h post-treatment, γ-H2AX levels began to decrease and this repair was concomitant with an induction of the FA signaling pathway in HSC, as demonstrated by both increased levels of FA gene expression and elevated FANCD2 foci (4-fold induction, p<0.01). Treatment of Fanca-/- mice with polyI:polyC led to a HSC proliferative response comparable to wild type (WT) mice but resulted in a 2-fold higher level of activation-induced DNA damage (p<0.05), demonstrating that this repair pathway is involved in resolving activation-induced DNA damage. Four rounds of serial in vivo activation led to a permanent depletion of the most primitive label-retaining Fanca-/- HSC and this correlated with a 4-fold depletion of functional HSC (p<0.01) as defined by competitive repopulation assays. Subsequent rounds of HSC activation with polyI:polyC resulted in the onset of a severe aplastic anemia (SAA) in 33% of treated Fanca-/- mice but not in any of the WT controls. SSA was characterized by a dramatic reduction in bone marrow (BM) cellularity, profound thrombocytopenia (21-246x106 platelets/ml), leukocytopenia (0.4-0.5x106 WBC/ml), neutropenia (0.03-0.1x106/ml) and anemia (1.5-2.3 g/dL Hb). Examination of BM HSC/progenitors demonstrated nearly complete loss of HSC, MPP, CMP and CLP (depletion of ≥33x, 8x, 4x and 12x respectively compared to PBS-treated Fanca-/-controls). Taken together, these data demonstrates that enforced exit from dormancy in vivo leads to de novo DNA damage in HSC, which is repaired by activation of a FA-dependent DNA damage response. Furthermore, the highly penetrant bone marrow failure observed in Fanconi anemia patients can be recapitulated by the serial application of a physiologic HSC activating signal to Fanca-/- mice. This suggests that the BM failure in FA may be caused by an aberrant response to HSC activation, most likely during exposure to infection or other physiologic stressors. These data provides a novel link between pro-inflammatory cytokines, DNA damage and HSC dysfunction and may have important clinical implications relevant to both prevention of BM failure in FA and in the study of age-related hematopoietic defects in non-FA patients. Moreover, these data provide the first evidence that FA knockout mouse models accurately recapitulate and provide novel insights into the etiology of BM failure in patients with FA. Disclosures: No relevant conflicts of interest to declare.

scholarly journals New Bone Marrow Failure Genes: DNAJC21 and ERCC6L2

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. SCI-22-SCI-22
Author(s):  
Inderjeet Dokal
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Nuclear Export ◽  
Conflicts Of Interest ◽  
Excision Repair ◽  
Bone Marrow Failure ◽  
Retinal Dystrophy ◽  
Ribosomal Subunit ◽  
Genotoxic Stress ◽  
Biallelic Mutations

A significant number of cases with bone marrow failure present with one or more extra-hematopoietic abnormality. This suggests a constitutional or genetic basis, and yet many of them remain uncharacterized. Through exome sequencing, we have recently identified two sub groups of these cases, one defined by germline biallelic mutations in DNAJC21 (DNAJ homolog subfamily C member 21) and the other in ERCC6L2 (excision repair cross complementing 6 like-2). Patients with DNAJC21 mutations are characterized by global bone marrow failure in early childhood. They can also have a variable number of extra-hematopoietic abnormalities such as short stature and retinal dystrophy. The encoded protein associates with ribosomal RNA (rRNA) and plays a highly conserved role in the maturation of the 60S ribosomal subunit. Lymphoblastoid patient cells exhibit increased sensitivity to the transcriptional inhibitor actinomycin D and reduced levels of rRNA. Characterisation of mutations has revealed impairment in interactions with cofactors (PA2G4, HSPA8 and ZNF622) involved in 60S maturation. DNAJC21 deficiency results in cytoplasmic accumulation of the 60S nuclear export factor PA2G4, aberrant ribosome profiles and increased cell death. Collectively these findings demonstrate that biallelic mutations in DNAJC21 cause disease due to defects in early nuclear rRNA biogenesis and late cytoplasmic maturation of the 60S subunit. Patients harbouring biallelic ERCC6L 2 mutations are characterized by bone marrow failure (in childhood or early adulthood) and one or more extra-hematopoietic abnormality such as microcephaly. Knockdown of ERCC6L2 in human cells significantly reduces their viability upon exposure to the DNA damaging agent irofulven but not etoposide and camptothecin suggesting a role in nucleotide excision repair. ERCC6L2 knockdown cells and patient cells harbouring biallelic ERCC6L2 mutations also display H2AX phosphorylation that significantly increases upon genotoxic stress, suggesting an early DNA damage response. ERCC6L2 is seen to translocate to mitochondria as well as the nucleus in response to DNA damage and its knockdown induces intracellular reactive oxygen species (ROS). Treatment with the ROS scavenger, N-acetyl-cysteine, attenuates the irofulven-induced cytotoxicity in ERCC6L2 knockdown cells and abolishes its traffic to mitochondria and nucleus in response to this DNA damaging agent. Collectively, these observations suggest that ERCC6L2has a pivotal rolein DNA repair and mitochondrial function. In conclusion, ERCC6L2 and DNAJC21 have an important role in maintaining genomic stability and ribosome biogenesis, respectively. They bring into focus new biological connections/pathways whose constitutional disruption is associated with defective hematopoiesis since patients harbouring germline biallelic mutations in these genes uniformly have bone marrow failure. Disclosures No relevant conflicts of interest to declare.

Xeroderma Pigmentosum as a Model for Treatment of Bone Marrow Failure in Fanconi Anemia and Aplastic Anemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4235-4235
Author(s):  
W. Clark Lambert ◽  
Santiago A. Centurion
Keyword(s):  
Cell Cycle ◽  
Bone Marrow ◽  
Fanconi Anemia ◽  
Xeroderma Pigmentosum ◽  
Bone Marrow Failure ◽  
S Phase ◽  
White Female ◽  
Cross Linking

Abstract We have previously shown that the primary cell cycle defect in the inherited, cancer-prone, bone marrow failure associated disease, Fanconi anemia (FA), is not in the G2 phase of the cell cycle, as had been thought for many years, but rather in the S phase. FA cells challenged with the DNA cross-linking agent, psoralen coupled with long wavelength, ultraviolet (UVA) radiation (PUVA), fail to slow their progression through the S phase of the subsequent cell cycle, as do normal cells. FA cells are extremely sensitive to the cytotoxic and clastogenic effects of DNA cross-linkers, such as PUVA, so much so that the diagnosis of FA is based on an assay, the “DEB test”, in which cells are examined for clastogenic and cytotoxic effects of diepoxybutane (DEB), a DNA cross-linking agent. More recently, we have shown that artificially slowing the cell cycle of FA cells exposed to PUVA by subsequent treatment with agents which slow their progression through S phase leads to markedly increased viability and reduced chromosome breakage in vitro. We now show that similar results can be obtained in vivo in patients with another DNA repair deficiency disease, xeroderma pigmentosum (XP), a recessively inherited disorder associated with defective repair of sunlight induced adducts in the DNA of sun-exposed tissues followed by development of numerous mutations causing large numbers of cancers in these same tissues. We treated two patients with XP, a light complected black male and a white female, both 14 years of age, in sun-exposed areas with 5-fluorouracil, an inhibitor of DNA synthesis, daily for three months. In contrast to normal patients, who only show clinical results if an inflammatory response is invoked, marked improvement in the clinical appearance of the skin was seen with no inflammation observed. This effect was confirmed histologically by examining epidermis adjacent to excised lesions in sun-exposed areas and further verified by computerized image analysis. Treatment with agents that slow progression through S phase, such as hydroxyurea, may similarly improve clinical outcomes in patients with FA or others who are developing bone marrow failure.

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