prognostic markers in nephroblastoma (wilms’ tumor)
TRANSCRIPT
Wcipdocpicpuiidtmetegastddpaed
FptGUS
NCN
1
REVIEW
©A
PROGNOSTIC MARKERS IN NEPHROBLASTOMA(WILMS’ TUMOR)
MAZEN A. GHANEM, GERT J. VAN STEENBRUGGE, RIEN J. M. NIJMAN, AND
THEODORUS H. VAN DER KWAST
W
sipsiAsthhlteoeAstt
W
btewctW
W
issccrm
ilms’ tumor (WT), or nephroblastoma, isone of the most common solid tumors in
hildren. The management of WT varies geograph-cally. In Europe and many other countries, therotocols of Société Internationale d’Oncologie Pé-iatrique (SIOP) are followed.1 Treatment is basedn the results of the initial imaging evaluation, andhemotherapy is then started before surgical extir-ation of the tumor. In the vast majority of centersn the United States and Canada, under the proto-ols of the National Wilms’ Tumor Study (NWTS),rimary nephrectomy is done for all but the largestnilateral tumor, and subsequent adjuvant therapy
s determined by the surgical and pathologic find-ngs.2 The assessment of the long-term risks of WTepends mainly on the histologic appearance andumor stage. However, many clinically relevant tu-or markers have been put forward that may allow
arly diagnosis and identification of high-risk pa-ients and consequently sparing of the unwantedffects of aggressive therapy in those with less ag-ressive disease.3 Limited published data are alsovailable on the possible differences in the expres-ion patterns of markers in tumors treated or notreated with chemotherapy. A tumor marker can beefined as a qualitative or quantitative alteration oreviation from normal of a molecule, substance, orrocess that can be detected by some kind of assaynd that is correlated with the prognosis.4 Differ-nt tumor markers for WT are being developed byifferent techniques (Tables I and II).
rom the Departments of Pediatric Urology and Pathology, Jose-hine Nefkens Institute, Erasmus University Medical Center Rot-erdam, Rotterdam; Department of Urology, University Hospitalroningen, Groningen, The Netherlands; and Department ofrology, Section of Paediatric Urology, Menoufiya University,hebin El-Kom, EgyptReprint requests: Gert J. van Steenbrugge, Ph.D., Josephine
efkens Institute, Room Be 430, Erasmus MC-University Medicalenter Rotterdam, P.O. Box 1738, Rotterdam 3000 DR, Theetherlands. E-mail: [email protected]: June 14, 2004, accepted (with revisions): December
T, 2004
2005 ELSEVIER INC. ULL RIGHTS RESERVED
TUMOR SUPPRESSOR GENES
T-1The WT-1 gene is localized on human chromo-
ome 11p13. It has been studied most extensivelyn WT at the mRNA level. Tumors that exhibitrimarily blastemal and epithelial differentiationhow greater levels of WT-1 expression than thosen which stromal tissue is predominantly present.lso, WT-1 mRNA has not been detected in thetroma component of WT.5 The percentage of WTshat have been shown to contain homozygous oreterozygous WT-1 mutations is relatively low,owever, and may be less than 25 and perhaps as
ittle as 5.6 The protein product of WT-1 binds tohe same DNA sequences as the protein product ofarly growth response 1 gene. Ghanem et al.7 dem-nstrated a significant increase in the expression ofarly growth response-1 in aggressive tumors.lso, the blastemal WT-1 and early growth re-ponse-1 detected immunohistochemically in pre-reated WT is related to the prognosis in additiono stage.
T-2A second locus at chromosome 11p15.5 has also
een described in WT. 11p15.5 is an importantumor-suppressor gene region showing loss of het-rozygosity (LOH) in WT. It is involved in Beck-ith-Wiedemann syndrome, which confers sus-
eptibility to WT,8 and, therefore, it may be usefulo investigate the prognostic value of 11p15.5 in
T at both the mRNA and protein level.
T-3 (16q)LOH on chromosome 16q in WT has been found
n about 17% of cases. The coincidence of micro-atellite instability in tumors exhibiting 16q LOHuggests that the loss of genetic material may beaused by errors in DNA mismatch repair and mayontribute to the pathogenesis of WT. A significantelationship between the loss of 16q and hypo-ethylation of 16q Sat2 DNA in WT was found.
his molecular finding has been linked to a worseROLOGY 65: 1047–1054, 2005 • 0090-4295/05/$30.00doi:10.1016/j.urology.2004.12.005 1047
oeLlsopd
p
twmdsmpecvdpWm
lpoi
mpcaooaghcpistnscdicWoet
pWpavsItmtcbotbinapppt
1
utcome of patients with WT in NWTS-5.9 Sev-ral genetic loci have been implicated in WT byOH studies or by the presence of germline trans-
ocations, including 7p, 11q, 22q, and loss of 4q. Aignificant association was noted between the gainf 1q and an increased risk of tumor relapse inatients with WT and favorable histologic findingsetected by comparative genomic hybridization.10
53In 1989, Baker et al.10 showed that deletions of
he short arm of chromosome 17 were associatedith point mutations in the p53 allele on the ho-ologous chromosome. Since then, much evi-
ence has supported that p53 is a tumor suppres-or gene involved in a wide range of humanalignancies. Conflicting reports regarding the
ossible existence of p53 mutations in WT havemerged from various studies. Most studies haveonfirmed its correlation with anaplasia and ad-anced-stage disease.12 However, Waber et al.13
emonstrated that p53 is not involved in theathogenesis of WT by examining a group of 38T cases with single-strand conformational poly-
TABLE I. Possible prognostic markers forWilms’ tumor
Tumor suppresor genesWT1WT-2WT3p53
Apoptosis-associated markersBcl-2BaxBcl-X
Proliferation markersProliferating cell nuclear antigenKi-67 (MIB-1)
Growth factorsInsulin-like growth factors (I, II)Epidermal growth factor familyPlatelet-derived growth factorBasic fibroblast growth factorMidkine expressionVascular endothelial growth factorsNerve growth factors
Cell adhesion molecules and extracellular matrix proteinsIntegrins and extracellular matrix proteinsTenascinHyluronic acidCD44
Nuclear acid content (DNA)N-myc expressionTotal plasma reninDrug-resistant markersTissue and cDNA microarray
orphism analysis. According to one report, the s
048
evel of p53 immunopositivity was not altered byreoperative chemotherapy,14 in contrast to thebservation by Moll et al.15 that chemotherapy maynfluence p53 protein expression.
APOPTOSIS-ASSOCIATED MARKERS
The control of apoptosis is complex and involvesany genes, including oncogenes and tumor sup-
ressor genes. The role and the prognostic signifi-ance of Bcl-2, Bcl-X, and Bax in nephroblastoma islso unknown. However, an increased expressionf Bcl-2 was observed in the blastemal componentf pretreated tumors with advanced stages and wasn independent prognostic marker for clinical pro-ression in addition to stage. Bcl-2/Bax expressionad prognostic significance for the prediction oflinical progression.16 Tanaka et al.17 showed thatreoperative chemotherapy did not significantlynfluence the occurrence of apoptosis or expres-ion of Bcl-2 in a group of treated WTs. However,he study of Bax mRNA expression levels inephroblastoma was not indicative that its expres-ion could have a role in the prognosis of WT. Inontrast, the anti-apoptotic Bcl-XL protein was theominant isoform of the Bcl-XS/L protein in exper-
mental human nephroblastoma, suggesting its on-ogenic potential in the development of anaplastic
T.18 The reduction of expression of the anti-ap-ptotic Bcl-L protein expression may contribute tonhanced sensitivity to radiotherapy or chemo-herapy.
PROLIFERATION MARKERS
Few reports have described the prognostic im-act of proliferating cell nuclear antigen scores inT in children. Nagoshi and Tsuneyoshi19 re-
orted no significant correlation between prolifer-ting cell nuclear antigen scores and patient sur-ival. However, Delahunt et al.20 reported aignificant correlation in postchemotherapy WT.n contrast, no prognostic impact was observed forumors resected before the administration of che-otherapy. Also, Khine et al.21 demonstrated that
he epithelial component of WT showed signifi-antly greater MIB-1 scores than those found forlastema, and a lack of MIB-1 staining has beenbserved in 11% of WTs studied. Also, some inves-igators reported a significant inverse relationshipetween blastemal MIB-1 and p27kip1 expressionn WT. This was observed in pretreated, as well asontreated, groups, because the proliferation wasffected and the MIB-1 scores were lower in theretreatment group. Blastemal MIB-1 and p27kip1
rotein expression proved to be an independentrognosticator for clinical progression in additiono stage.22 These latter results support the conclu-
ion that blastemal MIB-1 and p27kip1 are relevantUROLOGY 65 (6), 2005
mt
bko
I
cbWIketrtwL
sg
rmv
E
pEvtfhtepWspt
T
WEWW1pBBBPMpIETEPMVpGTHCDRPD
K�d
U
arkers for assessing the proliferative activity andumor cell dynamics of WT.
GROWTH FACTORS
The synthesis and actions of growth factors haveeen characterized in the developing metanephricidney. Studies have identified the abnormalitiesf growth factor expression in WT.
NSULIN-LIKE GROWTH FACTORS
Insulin-like growth factors (IGFs) comprise alass of peptides, two members of this family haveeen identified: IGF-I and IGF-II. In vitro culturedT cells produce and release various forms of “big
GF-II,” with molecular masses between 10 and 25Da. It is also well documented that WT tissuexpresses high levels of IGF-II, with levels similaro those of the fetal kidney.23 Antibodies to the IGFeceptor inhibit the growth of WT in vitro andumors transplanted to athymic mice, consistentith a role for IGF in oncogenesis.24 In contrast,
TABLE II. Summarization of available da
umor Marker
Clinical Samples
Pretreated Untreated
T-1 � �GR-1 � �T-2 � �T-3 � �
p � �53 � �cl-2 � �ax � �cl-X � �CNA � �IB-1 � �
27 � �GF � �GF � �GF-� � �GF-R � �DGF � �idkine � �
EGF � �75 � �DNF � �enascin � �yluronic acid � �D44 � �NA � �enin � �-gp � �NA topoisomerase II� � �
EY: � � tested; � � not tested; ND � not determined; PCNA � proliferating cell ntransforming growth factor-alpha; EGF-R � EGF receptor; PDGF � platelet-de
erived neurotrophic factor; P-gp � permeability glycoprotein.
ittle et al.25 demonstrated that enhanced expres- E
ROLOGY 65 (6), 2005
ion of IGF-II is not a necessary event in WT pro-ression.Because WT cells can produce IGF-II and carry
eceptors for this growth factor, an autocrine loopay be hypothesized. However, the prognostic
alue of IGFs in WT is uncertain.
PIDERMAL GROWTH FACTOR FAMILY
Epidermal growth factor (EGF) is a mitogenicolypeptide hormone that, by way of its receptorGF-R, stimulates the growth of cancer cells inivo. The EGF-R is a transmembrane glycoproteinhat binds EGF, as well as transforming growthactor-alpha (TGF-�).26 Data from one immuno-istochemical study of TGF-� and EGF-R in pre-reated WTs suggested that the significant blast-mal expression of TGF-� may play a role inromoting transformation and/or proliferation ofT, perhaps by an autocrine mechanism. Expres-
ion of TGF-� in tumor vessels also provides aossible mechanism of vascular invasion. Addi-ionally, the simultaneous expression of TGF-�/
f tumor markers tested in Wilms’ tumor
Culture MediaChemotherapeutic
EffectClinical
Trial
� No �� ND ND� ND �� ND �� ND �� Controversial �� No ND� No ND� Yes ND� Controversial ND� Yes ND� Yes ND� ND ND� ND ND� No ND� No ND� ND ND� ND ND� No �� ND ND� ND ND� ND ND� ND �� ND ND� ND ND� Yes �� Yes ND� Yes ND
antigen; IGF � insulin-like growth factor; EGF � epidermal growth factor; TGF-�rowth factor; VEGF � vascular endothelial growth factor; GDNF � glial cell line
ta o
uclearrived g
GF-R was indicative of clinical progression.27
1049
P
pmhtpipTvbsmscwtp
B
tacSccvsoAFf
M
ekaoobvsWIgWWsd
V
iss
itmVtrbowArWlsmaht
N
phaticspblrcrtTwTwtlW
pnsrddt
pp
1
LATELET-DERIVED GROWTH FACTOR
Platelet-derived growth factor (PDGF) is therincipal mitogen of serum acting on culturedesenchymal cells. Both cultured WT cells and
uman fetal kidney cells produce PDGF, consis-ent with its role in normal fetal growth and neo-lastic renal cell transformations.28 PDGF-AA andts receptor PDGF-� were predominantly ex-ressed in epithelial elements of patients with WT.heir epithelial expression was indicative of a fa-orable prognosis, because patients positive foroth PDGF-AA and PDGF-� receptor had a longerurvival than those who were negative for botholecules. PDGF-AA and PDGF-� receptors were
imultaneously expressed in 40% of the epithelialomponents of WT, implying that the patientsith high PDGF-AA/PDGF-� ratio do better than
hose with a low ratio (M.A. Ghanem, et al., un-ublished data).
ASIC FIBROBLAST GROWTH FACTOR
Basic fibroblast growth factors (bFGFs) are po-ent angiogenic peptides implicated in the growthnd metastasis of solid tumors. The FGF familyonsists of nine members (FGF-1 to FGF-9).koldenberg et al.29 showed FGF-2 (bFGF) nu-lear staining in both the epithelial and stromalompartment of WTs. In patients with WT, ele-ated preoperative urinary bFGF levels raise theuspicion of aggressive disease, and elevated post-perative levels may indicate disease recurrence.ccording to the available data, the influence ofGFs and their receptors on WT growth is still not
ully understood.
IDKINE EXPRESSION
A new family of heparin-binding growth/differ-ntiation factors has recently been described. Mid-ine (MK), the first to be discovered, is a product ofretinoic acid responsive gene. A significant levelf MK gene expression was observed in all samplesf WT examined. The inhibition of WT cell growthy an anti-MK antibody implies that MK is in-olved in the growth regulation of these cells. Thisuggests a role of MK in the biologic behavior of
T and its potential use as a diagnostic marker.30
n addition, high expression of retinol-relatedenes such as CRABP2, tubulin, and stathmin 1 inT may be related to the high chemosensitivity ofT and patients with a poor outcome. These data
uggest that the retinoid acid may be a potentialrug.31
ASCULAR ENDOTHELIAL GROWTH FACTORS
The close relationship between tumor vascular-ty and tumor growth is well established. Onetudy, using a murine model, reported overexpres-
ion of vascular endothelial growth factor (VEGF) i050
n a highly metastatic WT line, and anti-VEGF an-ibody (A4.6.1) successfully suppressed both pri-ary tumor growth and metastases.32 The level ofEGF in the serum of patients with WT was three
imes greater than that in controls. VEGF and Flt-1eceptor was mainly localized to the cytoplasm ofoth blastemal and epithelial cells of WT. The focif high vascularization occurred most frequentlyithin the tumor stroma, as defined by CD31.lso, Ghanem et al.33 confirmed a significant cor-
elation of VEGF and Flt-1 receptor expression inT and survival, with their expression closely re-
ated to microvessel density. The preliminary re-ults of VEGF and Flt-1 staining in untreated tu-or showed that the overall scores of the blastemal
nd epithelial cells were greater, but these patientsad a similar clinical outcome as those in the pre-reatment group.
ERVE GROWTH FACTORS
Neurotrophins and their receptors regulate cellroliferation, differentiation, and death of normaluman kidney and neuronal cells. The p75 low-ffinity nerve growth factor receptor has been iden-ified on epithelial WT cells in culture. In addition,t has been demonstrated that high-affinity Trk re-eptors are expressed in histologic structures in aeries of 10 WT specimens. The level of p75 ex-ression was high in areas mainly consisting oflastemal and relatively low in regions with epithe-ial differentiation. In contrast, the trk-A and trk-Beceptors were primarily found within stromalomponents, and the TRK C and trk C= (truncated)eceptors were present within epithelial struc-ures.34 Eggert et al.35 found that high levels ofrkB-full mRNA expression in WT is associatedith a worse outcome. Conversely, low levels ofrkB-truncated mRNA expression are associatedith a favorable outcome. Cells that express func-
ional, full-length TrkB may be susceptible to pro-iferation and/or survival signaling that leads to
T progression.31
Glial cell-line derived neurotrophic factor is aotent proliferation and survival factor of dopami-ergic neurons. Séguier-Lipszyc et al.36 demon-trated the presence of glial cell-line derived neu-otrophic factor in the epithelial cells of the mostifferentiated neuroectodermal tube and in the en-othelial cells of 20 patients with chemotherapeu-ically treated WT.In summary, the nerve growth factors appear to
lay a role in WT development and may have arognostic impact in clinical WT.
CELL ADHESION MOLECULES ANDEXTRACELLULAR MATRIX PROTEINS
Cell-matrix interactions play an important role
n normal renal development, as well as in tumorUROLOGY 65 (6), 2005
imcna
I
gelsstbtocenmooppctepmgt
T
sttccWw
H
EatdHfaitbc
C
tmrlCenCsit
ungsitptpwltfltiaoasrb
wLmngoAwspRebptN
U
nvasion and metastasis, and these interactions areediated by adhesion receptors. Several families of
ell surface receptors (eg, cadherins, selectins, theeural N-CAM, integrins, and CD44) govern thesedhesive interactions.
NTEGRINS AND EXTRACELLULAR MATRIX
Peringa et al.37 examined the expression of inte-rin subunits �1 to �6, �1, and �4, and of thextracellular matrix (ECM) proteins fibronectin,aminin, and collagen I and IV in 20 frozen WTamples and in 5 fetal kidney and 2 adult kidneypecimens. They found that their distribution inumor blastema differed strongly from the fetallastema. Undifferentiated blastema was charac-erized by expression of �3 and �6 and the absencef ECM proteins. Tumor blastema with epithelialommitment was demonstrated, having increasedxpression of �3 and �6, and production of lami-in. Tumor blastema with mesenchymal commit-ent showed loss of �3 and �6 and the expression
f �1, �4, and �5. It is speculated that the inabilityf the undifferentiated blastema to produce ECMroteins is related to its relatively high metastaticotential compared with epithelium and mesen-hyme. Integrin �-8 has been found to be the novelarget gene of WT-1 by an experiment with WT-1-xpressing fibroblasts. Most WTs showed high ex-ression of integrin �-8, and it was significantlyore highly expressed in high-stage tumors, sug-
esting that integrin �-8 may be involved in theumorigenesis and progression of WT.31
ENASCIN
Tenascin is an ECM protein prominently in thetroma of several epithelial malignancies. Its role inumor spread is unclear, but it may facilitate epi-helial cell migration during embryogenesis andarcinogenesis, by reducing rather than increasingell matrix interactions. The human sarcomatoid
T cell line induced a strong stromal responseith increased deposition of tenascin.38
YALURONIC ACID
Hyaluronic acid is a prominent component of theCM of WT. It is markedly elevated in the serumnd urine of some patients with WT. Stained WTissues with hyaluronic acid-binding protein probeemonstrated a prominent blastemal staining.39
yaluronic acid may serve as a biochemical markeror diagnosing and monitoring tumor recurrence;n immunodiagnostic test (eg, radioimmunoassay)s currently under development. The major advan-age of this test is that it will circumvent the cum-ersome and time-consuming steps inherent to a
ell culture bioassay. pROLOGY 65 (6), 2005
D44CD44 is a group of transmembrane glycopro-
eins formed by alternative splicing of a singleRNA. Membranous immunostaining of normal
enal tubular structure and blastemal and epithe-ial components of WT was demonstrated forD44s, CD44v5, and CD44v10. CD44v6, how-ver, was neither expressed in normal renal tissueor in WT. Increased blastemal expression ofD44v5 in pretreated WT correlated with progno-
is. Therefore, its expression may be of value indentifying patients with a high propensity for dis-ant metastasis.40
NUCLEAR ACID CONTENT
Flow cytometric analysis of DNA ploidy has beensed successfully to predict patient survival in aumber of malignancies. Previous studies investi-ated the relationship between DNA ploidy andurvival in WT.41,42 Douglass et al.41 found a strik-ng correlation between hyperdiploid tumor cells,he histologic subtype of anaplastic WT and poorrognosis. However, Layfield et al.42 found no sta-istically significant relationship between DNAloidy and patient survival, and anaplastic tumorsere more likely aneuploid or tetraploid than dip-
oid in their series. In contrast, Falkmer43 studiedhe method and potential problems of applyingow cytometry and static cytometry techniques forhe measurement of nuclear DNA content (ploidy)n WT. According to his study, many discrepanciesmong published series were documented as beingf methodologic origin. Because the advantagesnd disadvantages of both flow cytometry andtatic cytometry outweighed each other, Falkmer43
ecommended that the two measurements shoulde combined if applied for prognostic purposes.
N-myc EXPRESSION
The myc family of cellular oncogenes has threeell-characterized members (C-myc, N-myc, and-myc), as well as several additional members. N-yc is characteristic of several tumors that origi-ate from primitive cell lineage. WT exhibitedreatly enhanced levels of N-myc expression thatccurred in the absence of gene amplification.lso, a high level of N-myc expression is associatedith markedly diminished C-myc expression. In
itu hybridization, using [35S]-labeled RNArobes, confirmed that the high levels of N-mycNA were predominantly present in the blastemallements in WT. Attempts to detect N-myc proteiny immunologic methods have been unsuccessful,robably because of the extremely short half-life ofhe polypeptide compared with the mRNA. Thus,-myc mRNA detection may prove to be more
ractical than protein detection for diagnostic pur-1051
paWaab
hwiTantfiolm
MP
ccdtswaWDsoc
D
tcWsh
drnsnptt
igad
cnnAalmmctertanssp
himccwshsasmptutnancn
sC
Ws
o
1
oses. Using the Atlas of Human Cancer 1.2cDNArrays, the expression of C-myc was upregulated in
T-1 mutant tumors and was confirmed using re-l-time reverse transcriptase-polymerase chain re-ction. These data strongly suggest that C-myc is aiologically relevant target of WT1 in WTs.44
TOTAL PLASMA RENIN
Carachi et al.45 has demonstrated abnormallyigh levels of total plasma renin in most WTs,hich may have been due to increased levels of
nactive renin (prorenin), rather than active renin.hese high levels decreased to normal after surgerynd chemotherapy. The cells that contain immu-oreactive renin showed a typical perivascular dis-ribution in WT tissues. Also, Lindop et al.46 con-rmed the presence of immunoreactive renin in 10f 19 WTs and renin-specific mRNA (1.6 kb inength) by Northern blot analysis, similar to the
RNA detected in normal kidney tissue.
DRUG RESISTANCE MARKERS
ULTIDRUG RESISTANT-ASSOCIATED
ERMEABILITY-GLYCOPROTEIN
Multidrug resistant-associated permeability-gly-oprotein (P-gp) is a 170-kDa transmembrane gly-oprotein involved in the cellular redistribution ofrugs in the cytoplasm. Untreated patients and pa-ients with classic WT showed no P-gp overexpres-ion, except in areas of tubular differentiation, asas also seen in normal kidney.47 In contrast, Sola
nd associates48 detected P-gp in 12 nontreatedTs. Patients with WT treated with actinomycinand vincristine showed increased P-gp expres-
ion.47 However, the variation in method and usef different antibodies prohibited obtaining con-lusive results.45,46
NA TOPOISOMERASE II�Granzen et al.49 reported lower levels of DNA
opoisomerase II� in both blastemal and stromalomponents of treated compared with untreated
T. Takahashi et al.31 reported that high expres-ion of topoisomerase II� may be related to theigh chemosensitivity of WT.
TISSUE AND cDNA MICROARRAY
A recent technological advance was made by theevelopment of tissue microarrays, allowing theapid immunohistochemical analysis of a largeumber of potential prognostic markers on a largeet of tumor samples. The application of this tech-ique might have led to the investigation of therognostic impact of a much larger set of markershan those reported in this review.49 A disadvan-
age of the tissue microarray technique is the lim- 2052
ted amount of tumor tissue that can be investi-ated. Particularly for heterogeneous tumors suchs WT, this could lead to a lack of sensitivity toetect potentially clinically relevant markers.Another promising approach is the technology of
DNA microarrays studying the expression of largeumbers of genes at the mRNA level, exploring theew world of the genome with DNA microarrays.fter extraction of mRNA from a number of WTsnd its hybridization on a cDNA array containing aarge number of genes, the level of expression of a
ultitude of genes in WT can be investigated. Itay be anticipated that this technique will dis-
over prognostic markers for WT in addition tohose described in this report. However, mRNAxpression levels do not necessarily correlate di-ectly with protein levels, and a number of poten-ially important markers may be missed by thispproach. cDNA microarray analysis of WT is alsoow yielding important information on the expres-ion of molecules with a role in chemotherapy re-istance, such as the recently reported value of to-oisomerase II� and stathmin.31
CONCLUSIONS
For almost all the reviewed markers, except foristopathologic features and tumor stage, conflict-
ng results have been published. However, thearkers identified as yet have failed to provide
onsistent predictive information regarding thelinical outcome of WT. A large number of studiesere impaired by methodologic shortcomings,
uch as the small number of samples studied, aeterogeneous patient population with regard totage distribution, inappropriate statistics, or thebsence of multivariate analysis. Another, more ba-ic, problem in the identification of prognostic tu-or markers is the difference in chemotherapeutic
rotocols and their effects on the expression ofissues markers. In general, data of a preoperativelyntreated group of patients with WT demonstratedhat the expression patterns of several markers didot significantly differ and showed the same trends those in a treated group of patients (M.A. Gha-em, unpublished results) Additional studies ofhemotherapeutically untreated patients are defi-itely needed, however.
REFERENCES1. De Kraker J, Weitzman S, and Voute PA: Preoperative
trategies in the management of Wilms’ tumor. Hematol Oncollin North Am 9: 1275–1285, 1995.
2. D’Angio GJ, Breslow N, Beckwith JB, et al: Treatment ofilms’ tumor: results of the Third National Wilms’ tumor
tudy. Cancer 64: 349–360, 1989.3. Wen JG, van Steenbrugge GJ, Egeler RM, et al: Progress
f fundamental research in Wilms’ tumor. Urol Res 25: 223–
30, 1997.UROLOGY 65 (6), 2005
gt
iw
lC
at2
ai
hJ
dS
tc2
iaa
p7
cu
mab
atC
b1
tt2
arc1
o1
apaL
MiC
lt
tW1
sW
te
ati
tm1
AJ
icc
pt
ct3
Egt
to
ta2
G2
et
mt8
cect
en
a
U
4. Hayes DF, Bast RC, Desch CE, et al: Tumor markerrading system: a frame work to evaluate clinical utility ofumor markers. J Natl Cancer Inst 88: 1456–1468, 1996.
5. Kikuchi H, Akasaka Y, Nagai T, et al: Genomic changesn the WT-gene (WT1) in Wilms’ tumors and their correlationith histology. Am J Pathol 140: 781–786, 1992.
6. Slater RM, and Mannens MMA: Cytogenetics and mo-ecular genetics of Wilms’ tumor of childhood. Cancer Genetytogene 61: 111–121, 1992.
7. Ghanem MA, Van der Kwast TH, Den Hollander JC, etl: Expression and prognostic value of WT-1 and EGR-1 pro-eins in nephroblastoma. Clin Cancer Res 6: 4265–4271,000.
8. Karnik P, Chen P, Paris M, et al: Loss of heterozygosityt chromosome 11p15 in Wilms’ tumors: identification of twondependent regions. Oncogene 17: 237–240, 1998.
9. Mason BE, Goodfellow PJ, Grundy PE, et al: 16q loss ofeterozygosity and microsatellite instability in Wilms’ tumor.Pediatr Surg 35: 891–897, 2000.10. Baker SJ, Fearon ER, Nigro JM, et al: Chromosome 17
eletions and p53 gene mutations in colorectal carcinomas.cience 244: 217–221, 1989.
11. Klamt B, Schulze M, Thate C, et al: Allele loss in Wilms’umors of chromosome arms 11q, 16q, and 22q correlates withlinicopathological parameters. Genes Chromosome Cancer2: 287–294, 1998.12. Cheah PL, Looi LM, and Chan LL: Immunohistochem-
cal expression of p53 proteins in Wilms’ tumors: a possiblessociation with the histological prognostic parameter ofnaplasia. Histopathology 28: 49–54, 1996.
13. Waber PG, Chen J, and Nisen PD: Infrequency of ras,53, WT1, of Rb gene alterations in Wilms’ tumors. Cancer2: 373–378, 1993.14. Lahoti C, Thorner P, and Malkin D: Immunohisto-
hemical detection of p53 in Wilms’ tumors correlates withnfavorable outcome. Am J Pathol 148: 1577–1589, 1996.15. Moll UM, Ostermeyer AG, Ahomadegbe J-C, et al: p53ediated tumor cell response to chemotherapeutic DNA dam-
ge: a preliminary study in matched pairs of breast canceriopsies. Hum Pathol 26: 1293–1301, 1995.16. Ghanem MA, Van der Kwast TH, Den Hollander JC, et
l: The prognostic significance of apoptosis-associated pro-eins Bcl-2, Bax and Bcl-X in clinical nephroblastoma. Br Jancer 85: 1557–1563, 2001.17. Tanaka K, Granata C, Wang Y, et al: Apoptosis and
cl-2 oncogene expression in Wilms’ tumor. Pediatr Surg Int5: 243–247, 1999.18. Re GG, Hazen-Martin DJ, El-Bahtimi R, et al: Prognos-
ic significance of Bcl-2 in Wilms’ tumors and oncogenic po-ential of Bcl-Xl in rare tumours cases. Int J Cancer 84: 192–00, 1999.19. Nagoshi M, and Tsuneyoshi M: Expression of prolifer-
ting cell nuclear antigen in Wilms’ tumors and other pediat-ics renal tumors: the correlation between histologic classifi-ation and proliferative activity. J Surg Oncol 55: 114–121,994.20. Delahunt B, Farrant GJ, Bethwaite PB, et al: Assessment
f proliferative activity in Wilms’ tumor. Anal Cell Pathol 7:27–138, 1994.21. Khine NM, Aung W, Sibbons PD, et al: Analysis of rel-
tive proliferation rates of Wilms’ tumor components usingroliferating cell nuclear antigen and MIB-1 (Ki-67 equivalentntigen) immunostaining and assessment of mitotic index.ab Invest 70: 125–129, 1994.22. Ghanem MA, van der Kwast TH, Sudaryo MK, et al:IB-1(Ki-67) proliferation index and cyclin dependent kinase
nhibitor p27kip1 protein expression in nephroblastoma. Clin
ancer Res 10: 591–597, 2004. aROLOGY 65 (6), 2005
23. Schmitt S, Ren-Qiu Q, Torresani T, et al: High molecu-ar weight forms of IGF-II (“big-IGF-II”) released by Wilms’umor cells. Eur J Endocrin 137: 396–401, 1997.
24. Gansler T, Furlanetto R, Gramling TS, et al: Antibody toype I insulin-like growth factor receptor inhibits growth of
ilms’ tumor in culture and in athymic mice. Am J Pathol35: 961–966, 1989.25. Little MH, Ablett G, and Smith PJ: Enhanced expres-
ion of insulin-like growth factor II is not a necessary event inilms’ tumor progression. Carcinogenesis 8: 865–868, 1987.26. Rogers SA, Ryan G, and Hammerman MR: Metanephric
ransforming growth factor-� is required for renal organogen-sis in vitro. Am J Physiol 262: F533–F539, 1992.
27. Ghanem MA, Van Der Kwast TH, Den Hollander JC, etl: Expression and prognostic value of epidermal growth fac-or receptor, transforming growth factor-alpha and C-erbB-2n nephroblastoma. Cancer 92: 3120–3129, 2001.
28. Fraizer GE, Bowen-Pope DF, and Vogel AM: Produc-ion of platelet-derived growth factor by cultured Wilms’ tu-or cells and fetal kidney cells. J Cell Physiol 133: 169–174,
987.29. Skoldenberg EG, Christiansson J, Sandstedt B, et al:
ngiogenesis and angiogenic growth factors in Wilms’ tumor.Urol 165: 2274–2279, 2001.30. Tsutsui J, Kadomatsu K, Matsubora S, et al: A new fam-
ly of heparin-binding growth/differentiation factors: in-reased midkine expression in Wilms’ tumor and other humanarcinomas. Cancer Res 53: 1281–1285, 1993.
31. Takahashi M, Yang X, Lavery T, et al: Gene expressionrofiling of favorable histology Wilms tumors and its correla-ion with clinical features. Cancer Res 62: 6598–6605, 2002.
32. Kayton ML, Rowe DH, O’Toole KM, et al: Metastasisorrelates with production of vascular endothelial growth fac-or in a murine model of human Wilms’ tumor. J Pediatr Surg4: 743–748, 1999.33. Ghanem MA, van Steenbrugge GJ, Sudaryo MK, et al:
xpression and prognostic relevance of vascular endothelialrowth factor (VEGF) and its receptor (Flt-1) in nephroblas-oma. J Clin Pathol 56: 107–113, 2003.
34. Donovan MJ, Hempstead B, Julie-Huber L, et al: Iden-ification of the neurotrophin receptors p75 and trk in a seriesf Wilms’ tumors. Am J Pathol 145: 792–801, 1994.35. Eggert BA, Grotzer MA, Ikegaki N, et al: Expression of
he neurotrophin receptor TrKB is associated with unfavor-ble outcome in Wilms’ tumor. J Clin Oncol 19: 689–696,001.36. Séguier-Lipszyc E, El-Ghoneimi A, Brinon C, et al:
DNF expression in Wilms’ tumor. J Urol 165: 2269–2273,001.37. Peringa J, Molenaar WM, and Timens W: Integrins and
xtracellular matrix-proteins in the different components ofhe Wilms’ tumor. Virchows Arch 425: 113–119, 1994.
38. Talts JF, Aufderheid E, Sorokin L, et al: Induction ofouse tenascin expression by a human sarcomatoid Wilms’
umor cell line growing in nude mice. Int J Cancer 54: 868–74, 1993.39. Roth J, Zuber C, Wagner P, et al: Presence of the long
hain form of polysialic acid of the neural cell adhesion mol-cule in Wilms’ tumor: identification of a cell adhesion mole-ule as an oncodevelopmental antigen and implications forumor histogenesis. Am J Pathol 133: 227–240, 1988.
40. Ghanem MA, Van Steenbrugge GJ, Van der Kwast TH,t al: Expression and prognostic value of CD44 isoforms inephroblastoma (Wilms’ tumor). J Urol 168: 681–686, 2002.41. Douglass EC, Look AT, Webber B, et al: Hyperdiploidy
nd chromosomal rearrangements define the anaplastic vari-
nt of Wilms’ tumor. J Clin Oncol 4: 975–981, 1986.1053
sn
at1
isi3
a2
g1
ad1
to
eac
1
42. Layfield LJ, Ritchie AWS, and Ehrlich R: The relation-hip of deoxyribonucleic acid content to conventional prog-ostic factors in Wilms’ tumor. J Urol 142: 1040–1043, 1989.43. Falkmer UG: Methodologic sources of errors in image
nd flow cytometric DNA assessments of the malignancy po-ential of prostatic carcinoma. Hum Pathol 23: 360–367,992.44. Udtha M, Lee S-J, Alam R, et al: Upregulation of c-Myc
n WT1-mutant tumors: assessment of WT1 putative tran-criptional targets using cDNA microarray expression profil-ng of genetically defined Wilms’ tumors. Oncogene 23: 3821–826, 2003.45. Carachi R, Lindop GBM, and Leckie BJ: Inactive renin:tumor marker in nephroblastoma. J Pediatr Surg 22: 278–
80, 1987. 2054
46. Lindop GBM, Duncan K, Millan DWM, et al: Reninene expression in nephroblastoma. J Pathol 161: 93–97,990.47. Re GG, Willingham MC, El-Bahtimi G, et al: Anaplasia
nd drug selection-independent overexpression of the multi-rug resistant gene, MDR1, in Wilms’ tumor. Mod Pathol 10:29–136, 1997.48. Sola JE, Wojno KJ, Dooley W, et al: P-glycoprotein sta-
us of favorable histology Wilms’ tumor predicts treatmentutcome. J Pediatr Surg 29: 1080–1083, 1994.49. Granzen B, Efferth T, Keller UMA, et al: Differential
xpression of the drug resistance markers DNA Topoisomer-se II� and Glutathione S-transferase-� in the histologicalompartments of Wilms’ tumors. Anticancer Res 21: 771–776,
001.UROLOGY 65 (6), 2005