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Hematology 2006 63

Inherited Bone Marrow Failure Syndromes:

Molecular Features

Akiko Shimamura

The inherited marrow failure syndromes are characterized

by impaired hematopoiesis and cancer predisposition. Mostinherited marrow failure syndromes are also associated with

a range of congenital anomalies. These rare diseases offer

important insights into general mechanisms governing

human development, hematopoiesis and tumorigenesis.

First, the clinical features of four of the inherited marrow

failure syndromes: Fanconi anemia (FA), dyskeratosis

congenita (DC), Diamond-Blackfan anemia (DBA), and

Shwachman-Diamond syndrome (SDS), are briefly reviewed.

Emerging molecular themes arising from recent studies of

these syndromes are discussed. Finally, clinical applica-

tions arising from our growing understanding of molecular

pathogenesis are explored.

Clinical Features

In the past, the diagnosis of these diseases relied on therecognition of characteristic clinical features. With the ad-

vent of laboratory and genetic tests for many of these disor-

ders, our understanding of the clinical spectrum of these

disorders has broadened. Indeed, it is becoming increas-

ingly apparent that patients lacking characteristic physi-

cal stigmata may still harbor an inherited marrow failure

syndrome and develop marrow failure or malignancy. Clini-

cal presentation is no longer confined to the pediatric popu-

lation but may manifest in adults as well.

Fanconi anemia

FA is classically characterized by progressive marrow fail-

ure, congenital anomalies, and a predisposition to developleukemias and solid tumors. Marrow failure often progresses

to aplastic anemia. The leukemias are typically acute my-

elogenous leukemias (AML), though a few cases of acute

lymphocytic leukemias (ALL) have also been reported. The

solid tumors are typically squamous cell carcinomas, com-

monly involving the head and neck or female genital tract.

Associated clinical findings may include short stature, skin

pigment abnormalities (e.g., cafe au lait spots), radial ray

anomalies, genitourinary abnormalities, and microphthal-

mia. Of note, a subset of patients may lack the characteris-

Recent advances resulting from the identification ofthe genes responsible for four inherited marrow failuresyndromes, Fanconi anemia, dyskeratosis congenita,

Diamond-Blackfan anemia, and Shwachman-Diamondsyndrome, are reviewed. The interpretation of genetic

testing should be guided by an understanding of thelimitations of such testing for each disorder. The

possibility of an inherited basis for marrow failure

must be considered for adults as well as children withaplastic anemia. Shared molecular themes are

emerging from functional studies of the genes under-

lying the different inherited disorders. Genomicinstability may result from impaired DNA repair inFanconi anemia or telomere dysregulation in dyskera-

tosis congenita. Mutations affecting ribosome assem-bly or function are associated with Diamond-Blackfan

anemia, dyskeratosis congenita, and Shwachman-Diamond syndrome. These findings raise new ques-

tions about the molecular mechanisms regulating

hematopoiesis and leukemogenesis. Clinical implica-tions arising from these molecular studies are ex-

plored.

Marrow Failure Syndromes

Session Chair: Jeffrey M. Lipton, MD, PhD

Speakers: Akiko Shimamura, MD, PhD; Neal S. Young, MD; and Judith Marsh, MD

Acknowledgments: The author would like to thank David Nathan

and Monica Bessler for helpful discussions.

Correspondence: Akiko Shimamura, MD, PhD, Children’s

Hospital Boston, Karp Research Laboratories, 08214, 300

Longwood Ave., Boston MA 02115; Phone 617-919-2508; Fax

617-730-0934; Email [email protected]



64 American Society of Hematology

tic physical stigmata of FA but can be diagnosed with chro-

mosomal breakage testing (see below). In addition, patients

may first present in adulthood with marrow failure or ma-

lignancy as the primary clinical manifestation of FA.

Dyskeratosis congenita

DC is classically characterized by the clinical triad of dys-

trophic nails, a reticular rash, and mucosal leukoplakia.Bone marrow failure is the usual cause of mortality. DC is

associated with an increased risk of leukemias and solid

tumors, particularly squamous cell carcinomas. Pulmonary

fibrosis has also been described in these patients. Addi-

tional clinical features may be variably seen, and include

short stature, dental abnormalities, esophageal stricture, pre-

mature hair loss, early graying, osteoporosis, liver cirrhosis,

urinary tract anomalies, and hyperhidrosis. Of note, the char-

acteristic clinical triad is typically absent early in life but

manifests with age. The diagnosis may be obscured in pa-

tients presenting with aplastic anemia at a young age prior to

the onset of skin, nail and mucosal findings. The clinical

spectrum is highly variable, even within a given family.

Diamond-Blackfan anemia

DBA is classically characterized by reticulocytopenic ane-

mia presenting in early infancy. The bone marrow typi-

cally shows red cell aplasia with a paucity of erythroid

precursors. Other causes of pure red cell aplasia must be

ruled out. The red cells are typically macrocytic. Red cell

adenosine deaminase (ADA) activity is elevated in most,

but not all, patients. Associated congenital anomalies, such

as craniofacial anomalies, radial ray abnormalities, renal

and cardiac defects, may also be seen. Hemoglobin levels

may improve upon treatment with adrenocortical steroids.

Spontaneous remission may occur in a subset of patients.

An increased risk of AML and osteosarcomas and likely

additional malignancies has been reported.1 Aplastic ane-

mia has also been described in a few patients with DBA.

Shwachman-Diamond syndrome

SDS is characterized by exocrine pancreatic insufficiency

and bone marrow failure. Neutropenia is the most common

manifestation of marrow failure, but anemia or thrombocy-

topenia may also occur. The pancreatic acini are largely

replaced by fatty tissue with relative sparing of the pancre-

atic ducts and islets. Pancreatic insufficiency typically

manifests in early infancy with steatorrhea and failure tothrive. Serum levels of trypsinogen or pancreatic

isoamylase are generally low. With increasing age, pancre-

atic function improves in over half of patients, which may

render the diagnosis elusive. Cytopenias may be intermit-

tent, further obscuring the diagnosis. Patients with SDS are

at increased risk of developing aplastic anemia and AML.

To date, solid tumors have not been reported in patients

with SDS.

Molecular Pathogenesis

Genomic instability and Fanconi anemia

The hallmark of FA is the inability to repair DNA damage

induced by DNA crosslinking agents such as mitomycin C

or diepoxybutane. When exposed to these agents, cells from

FA patients manifest chromosomal breaks and fusions with

characteristic radial forms (Figure 1) consistent with anunderlying genomic instability. Complex cytogenetic ab-

normalities are typically associated with the leukemias aris-

ing in FA patients. Ongoing molecular studies support a

role for the FA pathway in DNA repair.

FA is a recessive disorder with both autosomal and X-

linked patterns of inheritance. Twelve different FA comple-

mentation groups have been identified to date, and the

responsible genes for 11 of these groups have been identi-

fied (Table 1) (reviewed in 2,3). Around 84% of patients fall

within the subtypes A, C, or G, with the majority of patients

comprising subtype A. Many of the FA genes encode novel

proteins of unknown function. The identification of the

FANCD1 gene as the BRCA2 tumor suppressor gene in-volved in homologous recombination repair provided a

direct link between the FA pathway and DNA repair. The

FANCJ/BACH1/BRIP1 gene encodes a helicase, which is

Figure 1. Fanconi anemia chromosomal breakage test.Diepoxybutane-induced chromosomal breakage in a metaphaselymphocyte from a patient with Fanconi anemia. Chromosomeswith breaks and fusions are indicated with arrows. Courtesy ofLisa Moreau, Dana Farber Cancer Institute, Boston, MA.



Hematology 2006 65

an enzyme that unwinds DNA. The FANCM ( Hef ) gene

shares regions of homology with both helicases and endo-

nucleases, though these functions have yet to be directly

demonstrated. FANCM may function in DNA transloca-

tion. The FANCL/PHF9/POG gene shares homology with

other E3 ubiquitin ligases.

Eight of the FA proteins (A, B, C, E, F, G, L, and M)

associate in a nuclear core complex and together with theFanconi anemia I protein are required for monoubiquitin-

ation of the Fanconi anemia D2 (FANCD2) protein (Figure

2).3 Monoubiquitinated FANCD2 carries a ubiquitin pro-

tein covalently linked via post-translational modification

to a specific FANCD2 lysine residue (K561). Abrogation of

any of the nuclear complex components disrupts FANCD2

monoubiquitination. Although the Bloom syndrome heli-

case, BLM, associates with the FA core complex,4 BLM is

not required for FANCD2 monoubiquitination.5 FANCD2

is monoubiquitinated during S phase of the cell cycle and

in response to DNA damaging agents such as mitomycin C

or radiation (ultraviolet or ionizing). The monoubiquitin-

ated FANCD2 protein localizes to chromatin, where it as-sociates with other DNA repair proteins such as FANCD1/

BRCA2, BRCA1, RAD51, and NBS1. In addition to func-

tioning to promote ubiquitination of FANCD2, the FA core

complex is required for translocation of ubiquitinated

FANCD2 to chromatin and for resistance to interstrand

crosslinking agents.6 Indeed, the FA core complex proteins

FANCA, FANCC, and FANCG also associate with chroma-

tin particularly in response to DNA damage.7 The precise

molecular function of the FA pathway in DNA repair re-

mains unclear.

The FA pathway interacts with additional DNA repair

pathways involved in tumor suppression.2,3 Monoubiquitin-

ated FANCD2 co-localizes with the tumor suppressor pro-

teins FANCD1/BRCA2, BRCA1, and NBS1 in chromatin-

bound foci. The ataxia-telangiectasia protein, ATM kinase,

phosphorylates FANCD2 to induce an S-phase cell cycle

checkpoint in response to radiation. This checkpoint is

important to arrest DNA synthesis under conditions of DNA

damage. In addition, the ATR/CHK1 pathway has been im-

plicated in the activation of the FA pathway in response to

DNA damage.

Genomic instability and dyskeratosis congenita

Dyskeratosis congenita may be inherited in an X-linked

recessive, autosomal dominant, or autosomal recessive form.

The X-linked recessive form is usually more severe with an

earlier clinical onset than the autosomal dominant form.

Cutaneous manifestations may be mild or missing in the

autosomal dominant form. The clinical phenotype may vary

widely, even within a given family. The gene for the auto-

somal recessive form has not yet been identified.

Mutations in DKC1, which encodes the protein

dyskerin, are associated with the X-linked form of DC.8

Most DKC1 mutations are missense mutations. Dyskerin isan evolutionarily conserved protein that is expressed

throughout all tissues. Dyskerin associates with the box H/

ACA class of RNAs, which includes the telomerase RNA,

and is important for their nuclear accumulation and stabil-

ity. Dyskerin also shares structural similarities to pseudo-

uridine synthases.

In support of a role for telomerase dysfunction in DC,

patients with the autosomal dominant form of DC harbor

mutations in the RNA component of telomerase (TERC )9,10

or the telomerase reverse transcriptase (TERT ).11 Data indi-

cate that the clinical phenotype results from haplo-insuffi-

Figure 2. The Fanconi anemia pathway.FA proteins (A, B, C, D1, D2, E, F, G, I, J, L, M) are depictedwith shaded circles. Additional DNA repair proteins interactingwith the FA pathway are denoted in open circles. Followingactivation of the FA complex by DNA damage or the cell cycle,the D2 protein is monoubiquitinated. Monoubiquitinated D2protein translocates to chromatin where it co-localizes withadditional DNA repair proteins.

Table 1. Fanconi anemia (FA) complementation groups.

Required for PercentageFA FANCD2 of FASubtype Gene Ubiquitination Patients*

A FANCA yes 66%

B FANCB yes rare

C FANCC yes 10%

D1 FANCD1/BRCA2 no 3%

D2 FANCD2 yes 3%

E FANCE yes 3%

F FANCF yes 2%

G FANCG/XRCC9 yes 9%

I ? yes 2%

J FANCJ/BACH1/BRIP1 no 2%

L FANCL/PHF9/POG yes rare

M FANCM/Hef yes rare

*From Levitus M, Rooimans MA, Steltenpoo, J, et al. Blood.2004;103:2498-2503




ciency of TERC or TERT rather than from dominant-nega-

tive effects. Patients with DC generally exhibit markedly

shortened telomeres.12 Family members with autosomal

dominant DC manifest “disease anticipation” with progres-

sive disease severity and earlier clinical onset over succes-

sive generations in the context of inheriting both progres-

sively shorter telomeres in association with a mutated copy

of TERC or TERT from generation to generation.13,14

Telomeres stabilize the chromosome ends to prevent

their shortening during replication, to distinguish chromo-

some ends from DNA damage-induced breaks, and to in-

hibit end-to-end fusions. Telomeres are comprised of 6-

basepair repeated sequences (TTAGGG) bound by specific

telomere-associated proteins. The telomeres are maintained

by the telomerase enzyme, which is composed of an RNA

component (TERC ) complexed with a reverse transcriptase

(TERT ) as well as additional proteins including dyskerin,

GAR1, NHP2, and NOP10 (Figure 3). The TERC RNA aligns

with the 3' end of the chromosomal telomeric repeats and

serves as a template for the TERT polymerase to replicate

the 3' chromosomal terminus.Telomerase is highly expressed in tissues with a high

replicative state, such as hematopoietic cells, germ cells,

and tissue stem cells. The clinical phenotype of DC mirrors

those tissues where there is a high rate of cell turnover and

telomerase is highly expressed. Telomerase is also present

in many cancer cells. When telomerase levels are low or

absent, as is the case in most somatic tissues, telomeres

progressively shorten with each cell division. When a criti-

cally short telomere length is reached, a checkpoint is trig-

gered causing the cells to stop dividing and senesce to

prevent chromosomal rearrangements. Thus, progressive

telomere shortening limits the replicative capacity of most

somatic tissues.

Telomerase RNA-null mice (mTR– / –)15,16 are viable and

manifest successive telomere shortening over subsequent

generations. Telomere shortening is accompanied by hair

greying, alopecia, and genetic instability.16 Impaired he-

matopoietic recovery following 5-FU treatment is ob-

served.16 Later generations of mTR– / – mice develop early

tumors that manifest a high incidence of cytogenetic ab-

normalities.15,16

Mutations abrogating Dkc1 expression resulted in

embryonic lethality in mouse models. Mice carrying a tar-

geting vector integrated downstream and in opposite ori-

entation to the Dkc1 gene demonstrated diminisheddyskerin expression levels, presumably from transcription

interference.17 These Dkc1m mice showed early phenotypic

abnormalities, including severe anemia, lymphopenia and

decreased marrow cellularity. Bone marrow colony-form-

ing assays showed diminished CFU-E and CFU-preB colo-

nies, but no difference in proliferation rate measured by

[3H]-thymidine incorporation. A high incidence of tumors

involving the lung, mammary gland, and kidney was ob-

served. Interestingly, although the clinical manifestations

were apparent in early generations of Dkc1m mice, signifi-

cant telomere shortening was not seen until later genera-

tions, suggesting that additional functions of dyskerin, such

as ribosome modification (discussed below), may also playan important role in the clinical spectrum of DC.

Ribosomal dysfunction in dyskeratosis congenita

Since dyskerin associates with other box H/ACA RNA spe-

cies in addition to the TERC RNA, additional effects of

dyskerin mutations may also contribute to the pathobiology

of DC. Box H/ACA RNAs18 complex with proteins to form

ribonucleoparticles (RNPs). Box H/ACA RNAs share a com-

mon secondary structure comprised of two hairpin stem-

loop structures separated by a hinge region and followed

by a tail containing the sequence ACA located three resi-

dues from the 3' terminus (Figure 4). The nomenclature

reflects the hinge region (box H) and the conserved ACA

triplet sequence. Several classes of box H/ACA RNAs have

been identified. The small nucleolar RNA (snoRNA) class

of box H/ACA RNAs function in ribosomal RNA (rRNA)

pseudouridylation and rRNA maturation. The telomerase

RNA (TERC or hTR) associates in the telomerase complex

to maintain telomeres. The U17/E1 RNAs function in rRNA

processing. The small Cajal body RNAs (scaRNA) func-

tion in pre-mRNA splicing. There are additional orphan

Figure 3. Telomerase and H/ACA small nucleolarribonucleoprotein complexes.Reproduced with publisher permission from Vulliamy TJ, et al.Blood. 2006;107(7):2680-2685.Abbreviations: sno, small nucleolar; RNP, ribonucleoprotein.

Figure 4. Schematic diagram of box H/ACA RNA structure.H/ACA RNAs are characterized by two hairpin stem-loopstructures separated by a hinge region and followed by a tailcontaining the sequence ACA located three residues from the 3 '

terminus.



Hematology 2006 67

box H/ACA RNAs whose functions are not yet understood.

The different classes of box H/ACA RNAs associate with

the same four highly conserved core proteins: dyskerin

(NAP57, CBF5p), GAR1, NHP2, and NOP10.18 Studies in

yeast indicate that all four core proteins are essential.

Dyskerin, the protein mutated in dyskeratosis

congenita, shares homology with pseudouridine synthases.

Mutations of the yeast homolog CBF5P reduce rRNApseudouridylation and impair growth.19 In patients with X-

linked DC, mutations in DKC1 are typically missense mu-

tations. Although the mutations are located in scattered

regions of the DKC1 gene, analysis of the crystal structure

of the yeast homolog CBF5 revealed that the mutations

clustered around a common spacial region of the folded

protein structure (the PUA domain) that is predicted to play

a role in binding H/ACA RNAs.20

The bulge in the hairpin of snoRNAs basepairs with

complementary sequences flanking a uridine in the RNA

species targeted for pseudouridylation. Pseudouridylation

is a post-transcriptional modification resulting in a differ-

ent isomer of uridine at specific residues of RNA molecules.Pseudouridylation has been postulated to affect rRNA sec-

ondary structure and binding. Pseudouridylation of rRNA

is seen throughout evolution and tends to occur in func-

tionally important regions of the rRNA. Pseudouridylation

of specific regions of rRNA is essential for ribosome func-

tion and growth in yeast.21 A defect in translation initiated

by internal ribosome entry site sequences (IRES) has been

observed in Dkc1m mice and in cells from X-linked DC pa-

tients wherein rRNA pseudouridylation was diminished.22

Patients with autosomal dominant forms of DC, who har-

bor mutations affecting telomerase function but not ribo-

some function, typically manifest milder clinical pheno-

types than do patients with DKC1 mutations. It has been

proposed that ribosomal dysfunction might affect clinical

severity in the context of telomerase dysfunction. In sup-

port of this model, some DC patient–derived point muta-

tions in DKC1 have been demonstrated to result in differ-

ent effects on telomerase activity versus pseudo-

uridylation.10,23

Ribosomal dysfunction in Diamond-Blackfan anemia

Around 20-25% of patients with DBA harbor heterozygous

mutations in the gene for the ribosomal protein RPS19.24

The disease likely results from haploinsufficiency of RPS19

expression rather than from a dominant negative effect.

25

An autosomal dominant pattern of inheritance may be seen

in families, but many cases appear to arise spontaneously.

RPS19 is a component of the small 40S eukaryotic riboso-

mal subunit and localizes to the nucleolus, the major cellu-

lar site of rRNA transcription and ribosome biogenesis. The

eukaryotic ribosome is composed of many more proteins

than the prokaryotic ribosome, and there is no prokaryotic

ortholog for RPS19. The roles of many of the ribosomal

proteins are not well understood in higher eukaryotes. Most

such studies have been performed in yeast.

Yeast have two RPS19 orthologs. Yeast carrying dele-

tions of both copies of the RPS19 gene are nonviable. De-

letion of either RPS19 gene results in decreased growth

with diminished production of the 40S ribosomal subunit.

Yeast RPS19 is required for maturation of the 3' end of the

18S rRNA. Deletion of RPS19 leads to accumulation of

aberrant pre-40S rRNA species that accumulate in the

nucleus. Introduction of patient-derived RPS19 point mu-tations into yeast leads to similar defects in 18S rRNA matu-

ration and 40S ribosomal subunit assembly.26 Additional

potential nonribosomal functions for RPS19 are currently

under investigation.27

Ribosomal dysfunction in

Shwachman-Diamond syndrome

Around 90% of patients who meet the clinical criteria for

Shwachman-Diamond syndrome harbor mutations in the

SBDS gene.28 SBDS is highly conserved throughout evolu-

tion and appears to be ubiquitously expressed throughout

all tissues. The majority of SBDS mutations appear to arise

from a gene conversion event with its adjacent pseudogene.To date, no patients homozygous for two null SBDS alleles

have been reported, suggesting that complete abrogation

of SBDS function is likely to be lethal. In support of this

hypothesis, mice lacking both sbds alleles exhibit early

embryonic lethality.29 The SBDS protein is localized

throughout the cell, but shuttles into the nucleolus, the

major cellular site of ribosome biogenesis, in a cell cycle–

dependent manner.30 Based on inferences from orthologs,

SBDS protein has been proposed to function in RNA me-

tabolism.28 Studies in yeast further suggest that SBDS may

function in rRNA maturation.31-34 The role of ribosome dys-

function in SDS remains to be ascertained.

Bone Marrow Failure

Genomic instability and marrow failure

It is currently surmised that defects in telomerase likely

result in premature senescence and apoptosis of the rapidly

cycling hematopoietic stem cells to result in marrow fail-

ure for patients with DC. The molecular mechanisms un-

derlying marrow failure in FA patients remains unclear.

While it is possible that the hematopoietic compartment of

FA patients might be particularly sensitive to endogenous

and exogenous genotoxic stressors, it is striking that mar-

row failure is not a typical feature of most other chromo-somal instability syndromes, such as ataxia-telangiectasia.

FA proteins, FANCC in particular, possess additional func-

tions that might additionally contribute to the pathogen-

esis of marrow failure.35 Cells from FA patients are hyper-

sensitive to apoptotic stimuli, and this may result in pre-

mature loss of hematopoietic stem cells. It is not clear

whether this heightened apoptosis represents a primary

defect of inappropriate apoptosis causing marrow failure

or a secondary checkpoint response to delete damaged cells

that cannot achieve proper DNA repair. The distinction




between these two possibilities carries profound clinical

implications. Loss or inhibition of apoptotic checkpoints

might promote the survival of cells that have acquired on-

cogenic mutations.

Ribosomal dysfunction and marrow failure

It remains unclear how haploinsufficiency for a ribosomal

protein manifests primarily as a defect in erythropoiesis inDBA. It has also been postulated that, in theory, differen-

tial expression levels of specific ribosomal proteins might

render some ribosomal proteins limiting for ribosome as-

sembly in certain tissues.36 As with all marrow failure syn-

dromes, increased levels of apoptosis have been observed

in marrows of DBA patients, though whether this repre-

sents a direct effect of ribosomal insufficiency or a second-

ary response to eliminate damaged or stressed cells is yet

unclear. It has been proposed that diminished translation

of mRNAs for anti-apoptotic factors might contribute to

marrow failure in DC.22

Another possibility that has yet to be explored is a

model wherein insufficiency of one ribosomal protein mightlead to a relative excess of other ribosomal proteins. The

equimolar generation of ribosomal proteins appears to be a

highly regulated process in the cell systems examined to

date. Given the orderly sequential assembly of ribosomal

proteins onto the maturing ribosomal RNAs, it is possible

that a block in one step of ribosome synthesis might lead to

inappropriate accumulation of unassembled ribosomal pro-

teins or precursor complexes that could either be directly

toxic to the cell or might trigger a checkpoint response.

Such a situation would be reminiscent of that seen in the

thalassemias, where erythroid demise results in part from

excessive accumulation of free alpha or beta globin pro-

tein in the setting of the relative paucity of the partner

globin protein. Since red cells contain high concentrations

of ribosomes to permit high levels of production of hemo-

globin, it is possible in this model that red cells are particu-

larly sensitive to limiting amounts of specific ribosomal

proteins. Such a model might also apply to the situation in

DBA, where potential defects in rRNA maturation might

result in inappropriate accumulation of rRNA precursors,

unassembled ribosomal proteins or RNP intermediates to

trigger a checkpoint response leading to the observed in-

crease in apoptosis of erythroid precursors. Additional po-

tential extra-ribosomal functions for RPS19 are under in-

vestigation. Further studies are needed to elucidate themechanisms promoting marrow failure.

Malignancy

Genomic instability and malignancy

Multiple lines of evidence support a role for genomic in-

stability in tumorigenesis.37 FA patients manifest cytoge-

netic abnormalities consistent with an underlying defect

in DNA repair. The protein products of many DNA repair

genes interact with the FA pathway (Figure 2). Indeed, the

FANCD1 gene is identical to the DNA repair gene BRCA2,

which is a tumor suppressor. Cancers arising in FA patients

typically harbor complex cytogenetic abnormalities, con-

sistent with an underlying genomic unstability.38 Genes

causing chromosomal instability syndromes, such as ataxia-

telangiectasia and xeroderma pigmentosa, also confer can-

cer predisposition. One model whereby genomic instabil-

ity has been postulated to promote tumorigenesis is throughthe facilitated acquisition of oncogenic mutations as well

as mutations promoting inappropriate survival of prema-

lignant cells.

Telomere loss results in the generation of unprotected

DNA ends that are unstable. These exposed DNA ends are

prone to exonucleolytic degradation or may undergo end-

to-end fusion with other chromosomes. The resulting di-

centric chromosomes do not experience proper chromo-

somal segregation and are prone to additional chromosomal

breakage during mitosis. Aneuploidy and end-to-end chro-

mosomal fusions were observed in the telomerase knock-out

mouse models. These events may facilitate the acquisition of

genetic mutations predisposing to cancer formation.

Ribosomal dysfunction and malignancy

Evidence supports a correlation between oncogenesis and

upregulation of ribosome biogenesis and translation,

though a direct oncogenic role for ribosome dysfunction

has yet to be ascertained.39 Dysregulation of specific sub-

sets of mRNAs, such as increased translation of growth-

promoting mRNAs, has been observed in response to on-

cogenic stimuli. The tumor suppressor protein, Arf, inhibits

the production of rRNA,40 while the oncogenic protein,

nucleophosmin, promotes rRNA biosynthesis.41 An intrigu-

ing mutagenesis screen for tumor suppressors in zebrafish

revealed a high incidence of heterogyzous mutations in ribo-

somal protein genes among the zebrafish lines exhibiting

tumor predisposition.42 Whether these ribosomal protein

mutations are directly oncogenic remains to be determined.

Extra-ribosomal functions for specific ribosomal pro-

teins are becoming increasingly apparent. For example, ri-

bosomal protein L11 binds and inhibits HDM2, resulting

in stabilization and activation of p53.43 Ribosomal protein

RPL26 binds the 5' untranslated region of p53 mRNA after

DNA damage to upregulate p53 expression.44 The large

ribosomal component L13a is phosphorylated in response

to gamma interferon and binds to the 3' untranslated region

of the ceruloplasmin mRNA to inhibit its translation.

45

Thus,possible oncogenic roles for specific ribosomal proteins

may include functions beyond that of the ribosome.

Molecular Studies: Diagnostic ImplicationsWith the identification of some of the genes underlying

these disorders, genetic testing for the purposes of diagno-

sis has become possible (Table 2). The role of genetic test-

ing in clinical management is an evolving field, and re-

sults must be considered critically. Interpretation of newly

identified sequence alterations in the absence of functional



Hematology 2006 69

data must be tempered with caution since they may not

necessarily represent pathogenic mutations, but rather may

constitute rare polymorphisms. The absence of apparent

mutations should be evaluated critically and within the

context of the limitations of the experimental assays uti-

lized. For example, an exon-directed sequencing approach

may miss potential deletions, inversions, or promotor mu-

tations. For the SBDS gene, extended gene conversion

events may be missed if the affected region is not spannedby the standard exon-directed primers. Gene identification

is still ongoing, so the absence of mutations in currently

known genes does not necessarily preclude a given diag-

nosis if the patient fits clinical criteria.

The availability of a functional test for FA has greatly

facilitated diagnosis for this disease. Increased chromosomal

breakage in response to the interstrand crosslinking agents

mitomycin C or diepoxybutane is the diagnostic hallmark

of FA. Somatic mosaicism, a state in which a somatic cell

clone has undergone genotypic reversion to a non-Fanconi

phenotype, may be seen in hematopoietic cells, thus ob-

scuring the diagnosis. In such cases, the diagnosis may be

ascertained by assessing a different tissue, typically skin

fibroblasts. It is currently unknown whether somatic mosa-

icism might confound diagnosis in other inherited marrow

failure syndromes. Patients with Nijmegen’s breakage syn-

drome may exhibit increased mitomycin C–induced chro-

mosomal breakage, leading to confusion with FA. Loss of

FANCD2 monoubiquitination is a useful adjunct diagnos-

tic screen but is currently only available on a research ba-

sis. A limitation to this approach is that a rare subset of

patients with subtypes D1 and J, where the affected FA

proteins function distal to FANCD2 monoubiquitination,

would be missed. Flow cytometry to assess cell cycle arrest

at G2 /M has been reported as a diagnostic test for FA. Ad-

vantages include the ability to rapidly screen large num-

bers of cells and relative ease of the assay. The sensitivity

and specificity of this assay for the different FA subtypes

remains to be ascertained.

Since patients with DC generally have shorter telom-

eres compared with age-matched controls, telomere lengthanalysis is currently under investigation as a diagnostic

screen for DC. The sensitivity and specificity of telomere

length analysis for diagnostic purposes remains to be as-

certained. The optimal cell population to be assayed for

diagnostic purposes also remains to be determined. This

test is currently only available on a research basis.

A careful patient history, including family history of

any physical anomalies, hematologic abnormalities, and

cancer predisposition, as well as a thorough physical exam

to assess for clinical stigmata of the inherited marrow fail-

ure syndromes together with supportive laboratory tests,

are critically important to the diagnostic workup of these

syndromes and may aid in the choice of genetic tests pur-

sued. The results of genetic testing should be interpreted

carefully within the context of the entire clinical picture of

the patient and the patient’s family.

Molecular Studies: Therapeutic ImplicationsRecently, attention has focused on the potential role of

acquired FA gene mutations or epigenetic silencing in the

somatic cells within the non-FA population.46 Studies indi-

cate that acquired somatic defects in FA gene expression,

Table 2. Tests for inherited marrow failure syndromes.

Helpful AdditionalDisease Gene Inheritance Pattern Laboratory Tests Research Tests

Fanconi Anemia FANCA Autosomal Recessive MMC or DEB chromosomal breakage FANCD2 mono-ubiquitinationFANCC Autosomal RecessiveFANCD1 Autosomal RecessiveFANCD2 Autosomal Recessive

FANCE Autosomal RecessiveFANCF Autosomal RecessiveFANCG Autosomal RecessiveFANCI Autosomal RecessiveFANCJ Autosomal RecessiveFANCL Autosomal RecessiveFANCM Autosomal RecessiveFANCB X-linked Recessive?

Dyskeratosis DKC1 X-linked Recessive None Telomere lengthCongenita TERC Autosomal Dominant

TERT Autosomal Dominant? Autosomal Recessive

Diamond-Blackfan RPS19 Autosomal Dominant Elevated red cell adenosine deaminaseAnemia ? (ADA)*

Shwachman- SBDS Autosomal Recessive Decreased serum trypsinogen and/or SBDS protein expressionDiamond Syndrome ? pancreatic isoamylase

*ADA levels are variably elevated in patients with Diamond-Blackfan anemia.




either through gene mutation or epigenetic silencing, might

contribute to tumorigenesis in a subset of non-FA patients.2,3

Since cells deficient in DNA repair are generally hypersen-

sitive to chemotherapeutic agents and to radiation, agents

that block DNA repair pathways such as the FA pathway in

tumors might be useful therapeutic agents to influence drug

or radiation sensitivity of tumors.46

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