CANCERRESEAR(@H55.4o96—4701,October15,19951

ABSTRACT

To investigate chromosomal events that underlie formation and progression of meningiomas, we have examined a set of 18 benign (WHO

grade I), 15 atypical (grade II), and 13 anaplastic/malignant (grade III)meningiomas for loss of heterozygosity (LOH) on chromosomes ip, 6p, 9q,

lOq, and 14q. Frequent loss of loci on these chromosomeswas seen Ingrade II and grade III tumors, specifically, 14q (II and III, 47 and 55%),‘p(40 and 70%), and lOq (27 and 40%). In contrast, LOH for these lociwas infrequent in benign meningiomas, specifically, 14q (0%), ip (11 %),and lOq (12%). The smallest common regions of deletion that could hedefined were 14q24—q32, 1p32.-pter, and 10q24—qter. These observationsindicate the likely presence of tumor suppressor genes in these regionsthat are involved in the development of WHO grade II and grade IIImeningiomas. Because LOH for loci on chromosomes ip and lOq wasfound in tumors of all grades and because the frequency of LOH in allthree regions increased with tumor grade, these results would support amodel for the formation of aggressive meningiomas through tumor progression.

INTRODUCTION

Meningioma is a slow-growing and generally well-circumscribedcentral nervous system tumor. Occasionally, rapid growth and infiltration of surrounding connective tissue, bone, and rarely brain can beobserved. Usually, local tumor recurrence after surgical therapy willdetermine the patient's clinical course, because systemic metastasesoccur only very rarely (1, 2).

Retrospective studies of surgically treated meningiomas have suggested that the extent of resection and the histopathological grading ofthe tumor might be important predictors of recurrent disease (3). Thecurrent WHO grading system comprises three grades: benign (grade I,M Is), atypical (grade II, M II), and anaplastic/malignant (grade III, M

III) meningioma. M II and M III are characterized by an intermediateand a high risk of recurrence, respectively, when compared to M I (4).In a recent series of 657 primary meningiomas, using the WHOcriteria, 94.3% were diagnosed as benign, 4.7% as atypical, and 1% asanaplastic (5).

The origin of M II and M III remains unclear. In some patients, afairly large tumor and a short clinical illness would point to de novoformation of M II!M III. Other patients experience local recurrence ofa tumor that exhibits progressively more anaplastic features, suggesting that tumor progression might underlie the development of their MII/M III. Because current clinicopathological parameters do not allow

for the identification of patients with benign tumors at risk for

malignant progression, the development of predictive genetic markersin such individuals might be of crucial importance to their clinicalmanagement.

The genetics of M I formation has been extensively researched, andcytogenetic and molecular genetic studies were instrumental in pinpointing a tumor suppressor locus (NF2) on chromosome 22 inmeningiomas (6—9).Mutations have been found in the NF2 gene in upto 59% of meningiomas. No specific locus has been implicated intumors without chromosome 22/NF2 mutations (10, 11).

Only limited data on molecular lesions associated with meningiomaprogression are available. A correlation of chromosome 22 loss withhistopathological criteria of aggressiveness has been reported (12).However, more recent data suggest that neither chromosome 22 lossesnor NF2 mutations play a role in the development of malignancy inmeningiomas (1 1). Other investigations have suggested that allelicdeletions on chromosomes ip and lOq may be significant eventsduring meningioma progression (13—15).

We have previously studied a collection of various tumors for LOHon chromosomes 2, 4, 14, 19, and 20. Allelic losses on chromosome14 were found in 4 of 7 M II and 3 of 3 M III, whereas with markersfrom the other chromosomes, only in one case was LOH detected ata locus on chromosome 20 (16). To further analyze the genetics of MII/M III formation, we examined a larger cohort of tumors specifically

for incidence and extent of LOH on chromosomes 14q, 10, and ip.

MATERIALS AND METHODS

Tumor and Blood Samples. Tumor tissue samples from 44 patientstreated at University of Cincinnati Medical Center, University Hospital Bonn,

University Hospital Lausanne, and University of Michigan Medical Centerwere obtained at surgery, snap frozen, and stored at —80°Cor in liquid

nitrogen. Histopathological diagnoses were made according to the current

WHO classification and reviewed on hematoxylin/eosin-stained sections by asingle neuropathologist (A. v. D.). In this system, brain invasion alone does notallow for the designation of grade III (Fig. 1; Ref. 4). In three cases (cases 56,101, and 102) slides were not available for review. All three tumors weredescribed as M III. No tissue could be obtained for additional studies. In

another case, only the diagnosis of recurrent meningioma could be made based

on the available material; the specimen excised previously was diagnosed as MII. The tumor was excluded from the analysis of deletion frequencies in tumorsof different grade. In one patient, tissues from a M I (case 700) and twoconsecutive M III recurrences (cases 788 and 834) could be obtained. Although

the patient presented with multiple meningiomas, at the time of the threesurgeries, only one site was operated on, ensuring that locally recurrent tumor

was analyzed. Available information regarding tumor location, age, and genderof the patient were obtained from the pathology reports and can be found in

Table 1. DNA was prepared from tumor and peripheral venous blood samplesaccording to standard procedures (17).

Microsatellite Loci and PCR Primers. Oligonucleotide primer pairs wereobtained from Research Genetics, Huntsville, AL, for the amplification ofmicrosatellites/variable number of tandem repeats from the following loci:

D1S162 (lp32);D1S188 (lp32);FGR (1p36.2—p36.l);D6S291 (6p2l.3—

p21.2);09S301 (9q13—q21);D9S302 (9q31—q33);D10S179 (lOp);D10S89(l0pter—pl 1.2); DIOSJJI (lopter—p11.2); DIOSJO9 (lOqi 1.2—qter); 10S169

(lOqi 1.2—qter);D10S187 (10q24—q25);D10S209 (10q24—q25);D14S80(14q12); D14S54 (14q12—q13);D14S70 (14q12—q13);D14S43 (14q24.3);

Received1/I1/95:accepted8/16/95.The costs of publication of this article were defrayed in part by the payment of page

charges. This article must therefore be hereby marked advertisement in accordance with18 U.S.C. Section 1734 solely to indicate this fact.

@ This work was funded by NIH grant NS31 145—02(to A. G. M.) and Grant DE445/2-1 from the Deutsche Forschungsgemeinschaft (to A. v. D.). M. S. is a postdoctoralfellow supported by the Deutsche Forschungsgemeinschaft.

2 To whom requests for reprints should be addressed, at Department of Molecular

Genetics, Biochemistry and Microbiology, University of Cincinnati Medical Cenler,Cincinnati, OH 45267-0524. Phone: (513) 558—5534; Fax: (513) 558—8474.

3 The abbreviations used are: M I (II, Ill), meningioma; WHO grade I (II, III); LOH,

loss of heterozygosity; NF2, neurofibromatosis type 2; MEN2, multiple endocrineneoplasia type 2.

I

4696

Allelic Losses on Chromosomes 14, 10, and 1 in Atypical and Malignant

Meningiomas: A Genetic Model of Meningioma Progression'

Matthias Simon, Andreas von Deimling, Jeffrey J. Larson, Ruth Wellenreuther, Peter Kaskel, Andreas Waha,Ronald E. Warnick, John M. Tew, Jr., and Anil G. Menon2

Departments of Molecular Genetics fM. S., J. J. L. A. G. M.J and Neurosurgery Ii. J. L. R. E. W., J. M. TI. University of Cincinnati College of Medicine, Cincinnati. Ohio45267,,ind InstitutfürNeuropathologie,UniversitätsklinikenBonn,Sigmund-Freud-Strag3e25. D-53105Bonn,GermanyIA. v. D., R. W.,P. K., A. W.J

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ALLELIC LOSSES IN MENINGIOMA PROGRESSION

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Fig. 1. Histological appearance of primary benign and recurrent anaplastic meningioma. A. tumor 700 (meningiothelial meningioma WHO grade I): indistinct cell borders, roundto oval nuclei, small and inconspicuous nucleoli, and delicate chromatin; B, tumor 834 (anaplastic meningioma WHO grade III): nuclear atypia, high nuclear cyloplasmic ratio,prominent nucleoli, and several mitotic figures. C, tumor 834: brain invasion. Islands of meningioma cells within the substance of the brain.

; @4

@.

D14S76 (14q24.3.-q31); D14S48 (14q24.3—q31); D14S59 (14q23—q31);

D14S81 (14q31); D14S45 (14q32.1—q32.2); D14S51 (14q32.i—q32.2), !GHJ

(14q32.33, variable number of tandem repeats). Genetic maps were obtained

from recent publications (18, 19) and the Genome Data Base (as of August1995).

PCR Conditions. PCR amplification of microsatelliteswas performedwith approximately 20 ng of template DNA, 20 pmol each of forward andreverse primer, 1.25 unit Taq Polymerase, 24 nmol deoxynucleotides, and 5%DMSO in the appropriatebufferin a final volume of 20 pJ in 96-well plateson a Programmable Thermal Cycler (Mi Research, Inc., Watertown, MA).PCR reactionswere set up on ice and immediatelydenaturedat 94°Cfor 2.5mm. This was followed by 35 cycles of 30 s at 94°C,1 mm at annealingtemperature, 40 s at 72°C, and a final extension step at 72°C for 5 mm. PCR

conditions for each primer pair were optimized with respect to buffer (60 msi

Tris-HC1, 15 mM (NH4)25O4, 1.5—3.5mM@ pH 8.5—10;20 mM Tris-HCI,50 mt@iKC1, 1.5—2.5mM Mg2@,pH 8.4) and annealing temperatures (52°C-63°C).

Silver Staining and Detection of Allele Loss. PCRproductswere mixedwith 10 gd of formamide loading dye (17), and 15 pAof the resulting mixturewere separated by electrophoresis through 8% polyacrylamide-50% urea gelsin 1 X TBE@ (0.13 M Tris, 0.045 M boric acid, 2.7 mM EDTA, pH 8.9).

Polyacrylamide gels were silver stained using a protocol described recently(20),dried,andkeptasa permanentrecord.In eachcase,allelelosswasassessed on dried gels by visually comparing the relative intensity of allelicband patterns from tumor DNA using the band pattern from blood DNA as astandard (Fig. 2; Ref. 21). PCR products from the JGHJ locus were mixed with5 p.1 glycerol loading dye (17), separated by electrophoresis through 1.2%agarose gels in 1 X ThE (17), and visualized with ethidium bromide.

Statistical Analysis. The association of different deletions was tested forsignificance using the two-tailed Fisher exact test (22). For the purposes ofstatistical analysis and the calculation of allele loss frequencies, consecutiveanaplastic local recurrences from the same patient (cases 788/834 and 2062/2064) were counted as one tumor-harboring LOH, if allele loss was seen in anyof the samples.

RESULTS

Frequency of Allelic Deletions in Meningiomas of DifferentGrades. Among M I, LOH was detected only for loci on chromosomes ip (1 1%) and lOq (12%). The most prevalent aberration

observed in M II was LOH for loci on chromosome l4q (47%),followed by chromosome Ip (40%) and lOq (27%). Among M III,allele losses were detected most frequently on chromosome 1p (70%);LOH for markers on chromosome 14q and lOq was seen less frequently (55 and 40%, respectively). Notably, chromosome 14 deletions were observed at equal or almost equal frequencies in M II andM III, whereas LOH on chromosomes ip and lOq was detected morefrequently among M III (Fig. 3). Interestingly, LOH for loci onchromosome 14 showed a significant correlation with LOH for loci onchromosome ip (P < 0.002) and lOq (P < 0.02). Also, the correlationbetween chromosome ip and lOq allele loss was statistically significant (P < 0.02). In contrast to the loci mentioned above, LOH formarkers on chromosomes 9q and 6p was observed only rarely intumors of all grades (M I, 0 and 0%; M II, 14 and 14%; M III, 20 and0%, respectively). Brain invasion was seen in 2 of 15 M II and 7 of10 M III. Both M II with brain invasion showed no allelic losses onany of the chromosomes examined. The results are summarized inTable 1.

Common Region of Deletions. An investigation of 11 loci onchromosome 14 identified 10 tumors with probable monosomy forthis chromosome. In two cases, however, terminal 14q allelic ddetions could be demonstrated (cases 3351 and 2275). One tumor (case2062) had probably undergone LOH for most of chromosome 14 but

retained heterozygosity at 14q32. A previous study identified aninterstitial chromosome 14 deletion in case 56 (between D14S43 andD14S23) using RFLP or PCR markers for D14S43, D14S13, D14S16,and D14S23 (16). Combining these cases, the cytogenetic region14q24.3—q32.33 (between D14S48 and D14S23) could be defined asthe smallest common region of deletion (Table 2).

In tumor 2275, LOH was observed for loci centromeric fromD14S54 and telomeric from D14S70 (Fig. 2; Table 2), indicating tworegions of chromosomal losses, one of which overlaps with thedeletions seen in cases 56, 2062, and 3351. Alternatively, a singlehomozygous deletion encompassing D14S54 and D14S70 would appear as retention of heterozygosity at these loci, because in theabsence of tumor sequences, normal alleles from ubiquitous stromal

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ALLELIC LO5SES IN MENINGIOMA PROGRESSION

Table 1 Clinicalfeatures and allelic losses of 47 meningiomas

ChromosomeSex,age

Tumor― (yr)― Grade and classificationc Location― ip 6p 9q lop lOq l4qe

2380 F, 56 I, t, rec Cavernous sinus 12 12 12 12 12 123328 F, 75 I, m Sphenoid wing 12 12 12 12 12 123044 F, 51 I, f Posterior fossa 12 11 12 12 12 123106 M, 39 I, m, mull Anterior fossa 12 11 12 ii 123150 F, 63 I, t, rec Middle fossa 12 12 11 11 12 123010 F,40 I,f Convexity 12 11 12 12 122534 M, 37 I, t Petroclival 12 12 12 12 12 123302 F, 42 1, m Foramen magnum 12 11 11 12 12 12

2423 F, 57 1, seer Anterior clinoidal 12 11 12 12 12 122615 F,55 I,m Sphenoidwing 12 11 12 12 12 122561 F, 57 I, t Tuberculum sellae 12 11 12 12 12 123354 F I, t, mult N/A 12 12 12 12

3089 F, 57 I, f, mult Parasagittal 12 12 12 123321 F,70 I,f Falx 12 12 12 12 12 122155 F,37 1,chord Convexity LOH 12 12 12 12 122282 M, 69 I, f Petroclival LOH 11 12 12 12 123075 F, 48 I, t Sphenoid wing 12 12 12 LOH 127m* F, 69 I, m, mult Faix 12 1 12 1 LOH 12

2/18 0/6 0/12 0/16 2/17 0/18

2066 F, 64 II, f Cerebellopontine angle 12 12 12 12 12 124058 F, 73 II, m Convexity 12 12 12 12 12610 F,54 II,m Convexity 12 11 12 12 12 122060 F, 37 II, f Tentorium 12 12 ii 12 12 12812 F, 70 II, m Parasagittal LOH 11 12 12 12 12582 M, 21 II, m, NF2 Convexity 12 12 LOH 12 12 122443 M, 71 II, m, rec Infra-orbital LOH 11 12 12 12 LOH3351 F, 29 II, m Cavernous sinus 12 LOH 12 12 LOH

2275 F, 45 II, m, rec Tentorium LOH 12 12 12 LOH LOH2068 F, 56 II, f, rec Tentorium LOH 11 LOH 12 LOH LOH3138 M, 63 II, m Convexity LOH 11 12 LOH LOH LOH2576 M, 64 II, m, rec Tentorium LOH 11 12 LOH LOH LOH2220 M, 48 Il,m, rec,VR Convexity 12 ii 12 12 12 LOH2313 F, 56 II, m, mv Convexity 12 11 12 12 12 123030 M, 23 II, m, mv Convexity 12 12 12 12 12 12

6/15 1/7 2/14 2/13 4/15 7/15

56 N/A III N/A LOH596 F, 48 III, f Cerebellopontine angle 12 12 12 12 12 122070 M, 50 III, m Convexity 12 12 12 12 12 122064@° F, 48 III, m/f, 2nd rec Convexity 12 12 12 12 122062°° F, 49 III, ru/f, 3rd rec Convexity 12 12 12 LOH LOH LOH878 F, 49 III, t, mv Convexity LOH 11 LOH 12 12 12

788 F, 70 III, m, rec, mult, mv FaIx LOH 1 12 1 LOH 12

834 F, 70 III, m, 2nd rec, mult, mv Faix LOH 1 12 1 LOH 12102 F III,mv N/A LOH 12 12 11 LOH 12864 M, 65 III, f, 4th rec, mv Convexity LOH 12 12 12 LOH3464 M, 73 III, mv Tentorium LOH 11 12 12 LOH101 F III, rec, mv N/A LOH 12 LOH 12 12 LOH2226 F, 70 III, m, mv Middle fossa LOH 11 12 LOH LOH LOH

7/10g 0/5k 2/10e 21r1g 4/iOu 6/llg

3408 M, 40@ rec Cavernous sinus LOH 12 LOH LOH LOH

a *@ consecutivelocalrecurrencesfromthesamepatient.b F, female; M, male; N/A, not available.

C I (II, III), WHO grade; m, meningiothelial; f, fibroblastic; I, transitional; seer, secretory; chord, chordoid meningioma; (2nd, 3rd, 4th) rec, (2nd, 3rd, 4th) recurrence; mult, patient

presented with multiple meningiomas; VR, infiltration of Virchow-Robin spaces (not considered brain invasion); mv, brain invasion present by histopathological observation.d N/A, not available.

e LOH, loss of heterozygosity; 1 1, homozygous for all markers tested; 12, heterozygosity retained for at least one marker and no LOH; 1, mono-allelic band pattern, no constitutional

control available. Tumors 700, 788, and 834 showed a mono-allelic band pattem with two markers from chromosome lOp and three markers from lOq. The tumors were thereforeclassified as harboring LOH for loci on chromosome iOq.

‘@Onlythe diagnosis of recurrence was possible based on the available material. The previously excised specimen was diagnosed as atypical meningioma.S Locally recurrent malignant tumors 788/834 and 2064/2062 were counted as one tumor each. Tumors were classified as harboring LOH, if allele loss could be demonstrated in

any of the subsequent tumors.

cells would be amplified during PCR. Such a deletion would not share centromeric locus (D1S188) defining 1p32—ptertelomeric to D1S188a region of overlap with the deletions observed in cases 335 1 and 56, as target region.unless the presence of large homozygous losses is masked as centro- Allelotype after Progression. In one patient with three consecumeric regions with retained heterozygosity in these tumors. tive tumors, allele loss at the FGR locus on chromosome lp could be

In 4 of 12 tumors with chromosome 10 allele loss, deletions were detected in both anaplastic recurrences (cases 788 and 834) using therestricted to loci on chromosome lOq. Case 102 showed a terminal benign primary tumor (case 700) as a control. Chromosome 10 lossallelic deletion of lOq telomeric to D10S187. Therefore, a common was present in all three consecutive tumors, because a hemi/homozytarget would have to be located in the region 10q24—qtertelomeric to gous band pattern was seen after amplification of five chromosome 10D10S187 (Table 3). Among 17 tumors with LOll for loci on chro- loci. The very high frequency of heterozygosity at these loci makes itmosome ip, one tumor (case 2068) retained heterozygosity for the highly unlikely that this patient was a compound homozygote. Lack of

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ALLELIC LOSSES IN MENINGIOMA PROGRESSION

A NTNT

B NTNT

on chromosome 14 in M II/M III. We provide data on four cases ofpartial deletions on chromosome 14, resulting in the delineation of asmallest common region harboring a putative tumor suppressor in MII/M III. The region defined in this study (14q24.3—q32.33, between

D14S48 and D14S23) would allow for the exclusion of the putativetarget of chromosome 14 deletions in colon cancer as the geneinvolved in M II/M III formation (14q32.33—qter, telomeric toD14S23; Ref. 24). On the other hand, the region described in thisstudy overlaps with the smallest common region of deletion reportedin neuroblastoma (14q32—qter, telomeric to D14S13; Ref. 25), ovarian cancer (14q12—14q32.2, between D14S49 and D14S51; Ref. 28),and possibly renal cell carcinoma (14q24.3—q31, between D14S76and D14S73; Ref. 30).

Interestingly, LOH centromeric and telomeric from D14S49 wasfound in a case of ovarian carcinoma, while heterozygosity at D14S49was retained (28). A similar result was obtained in case 2275 from thisstudy. In this tumor, two alleles were detected at D14S54 and D14S70,whereas LOH could be shown for telomeric and centromeric loci (Fig.2; Table 2), implying the existence of more than one target for

deletions in meningiomas on chromosome 14. It is also conceivablethat interstitial retention of heterozygosity reflects amplification ofalleles derived from contaminating normal tissue in the presence of ahomozygous deletion of the tumor alleles. Such a homozygous deletion likely would not overlap with the smallest common region ofchromosomal losses described for the other meningiomas. Thus, thisinterpretation suggests the involvement of a different target gene incase 2275.

Frequent LOH for loci on chromosome lp32—p36 has been described in M II/M III (13, 14). Our results confirm these findings andindicate a common region of deletion harboring a potential tumorsuppressor gene. Other reported neoplasms with non-random chromosome ip losses include oligodendroglioma (32), MEN2 associatedtumors (33), colon (34), liver (35), and breast cancer (36), male germcell tumors (37), melanoma (38), malignant mesothelioma (39), andneuroblastoma (25, 40). Interestingly, some evidence suggests theexistence of a locus for familial melanoma on chromosome lp36 (41).

Deletions on chromosome 10 generally including the distal part oflOq have been reported to be frequent in glioblastomas (42—44),renalcell carcinomas (45), prostate cancer (46), uterine cancer (47), andmelanomas (48). LOH for loci on chromosome 10 has also been

DISCUSSION

This study demonstrates that meningiomas with morphologicalsigns of malignancy are characterized by frequent deletions on chromosomes 14q, ip, and lOq. In contrast, these deletions are rare amongbenign tumors. The high rate of deletions detected in M II and M IIIdoes not appear to reflect random genomic instability, because LOHfor markers on chromosomes 6p and 9q in this series was observedonly rarely (6p: 1 of 7 M II and 0 of 5 M III; 9q: 2 of 14 M II and 2of 10 M III). Because frequent nonrandom allelic deletions are generally believed to reflect the loss of tumor suppressor sequences (23),our findings suggest the presence of tumor suppressor genes onchromosomes 14q, lOq, and ip involved in the formation of M II andM III.

Frequent chromosome 14 deletions have been detected in advancedcolon cancer (24), neuroblastoma (25, 26), endometrial and ovariancancer (27, 28), renal cell carcinoma (29, 30), and non-Hodgkin'slymphoma (31). In this study, we report on frequent LOH for markers

4 i. J. Larson, N. de Tribolet, i. M. Tew, Jr., H. R. van Loveren, R. E. Wamick, M. G.

Balko, and A. G. Menon. Clonal origin of primary and recurrent meningiomas, submittedfor publication.

70%

60%

50%

40%

30%

20%

10%

0%

Fig. 3. Histogram showing the frequency of allelic losses on chromosomes ip, iOq, and14q in meningiomas of different grades. The proportion of tumors with LOH is shown asa column for each chromosome and tumor grade. LOH for loci on chromosomes ip andlOq becomes more frequent with increasing grade, while the proportion of tumors withchromosome 14 allele loss does not significantly differ between M II and M III. M I (II.III), meningioma WHO grade I (II, III); LOH, loss of heterozygosity.

4699

D14S80

@i@ @_1D14S70Fig. 2. Examples of allele loss detection and deletion mapping with PCR-amplified

microsatellites. Arrows, allelic losses. The faint signals in the tumor lanes might be dueto contaminating normal tissue or tumor heterogeneity. A, D14S80: cases 2275 (Lanes Iand 2) and 3089 (Lanes 3 and 4). B, DI4S7O:cases 2275 (Lanes I and 2) and 3138 (Lanes3 and 4). Note that in tumor 2275, LOH can be detected at DI4SSO, while heterozygosityis retained more telomeric at D14S70. N, normal; T, tumor; LOH, loss of heterozygosity.

a normal constitutional control did not allow for deletion mapping inthese tumors.

We have shown previously by X chromosome inactivation studiesthat local tumor recurrence and progression in meningiomas generallyreflect regrowth of the same neoplastic clone.4 That such clonalprogression took place in tumors 700, 788, and 834 is strongly

suggested by the results of X chromosome inactivation studies of thephosphoglycerate kinase locus, which showed an identical inactivation pattern in all three tumors4 and the identical mono-allelic bandpattern obtained with all five chromosome 10 markers. Some of theseband patterns likely represent LOH affecting the same allele in allthree tumors, because the probability is very low that the patient washomozygous at all five loci.

In one M II (case 2068), which recurred after subtotal excision ofa M I 3.5 years earlier (no tissue available), chromosome ip, lOq, and14q allele loss was shown. The observations made in cases 700, 788,834, and2068 mayindicatethatLOH for loci on chromosomesip,lOq, and 14q is reflected in the morphological progression seen inthese tumors.

1pLOH 1OqLOH 14qLOH

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Table 2 Extent of chromosome 14 allele loss in 14meningiomasTumor/Grade―2068/

2220/ 2443/ 2576/ 3 138/ 2275/ 3351/ 2062/ 56/ 864/ 101/ 2226/3464/Locus―l..ocation II II II II II II II III III III Ill III Ill3408'@D14S80

14q12 LOH LOH LOH LOH LOH LOH 12 11 11 LOH11D14S5414q12-ql3 11 LOH 12 11 11LOHD14S70l4q12-ql3 LOH LOH LOH LOH LOH 12 LOH LOH 11LOHDI4S43l4q24.3 LOH LOH LOH 12 LOHLOHD14S76l4q24.3-q3 I LOH LOH 11 11 LOH LOH I 1 LOHLOHD14S48l4q24.3-q3l 11 11 LOH LOH LOH LOH 12 LOH LOH 11 LOH LOHIiD14S5914q23-q31 LOH LOH LOH LOH 11 LOH LOH LOH LOH LOH 11LOHDJ4S8I14q3l 11 LOH LOU 11 LOH LOH 11 LOH LOHLOHD14S45l4q32.1-q32.2LOHDJ4S5I14q32.1-q32.2 11 11 LOH LOH 11 LOH11D14S13l4q32.1-q32.2LOHD14SI6l4q32.32-q32.33LOHDI4S23l4q32.3312IGHJI4q32.33 I I LOH LOH LOH 11 11 LOH 12 LOH LOH LOH LOHLOH(1

Genetic map of the loci: /, relativeordernot

known.hI (II, III), WHO grade; LOH, loss of heterozygosity; 1 1, not informative; 12, informative and noLOH.t

No grade could be assigned based on the available sample. Previous specimen was diagnosed as atypicalmeningioma.described

in 5 of 15 atypical and malignant meningiomas. In two does not provide further evidence linking a subgroup of M I to MhIMcases,LOH was observed on chromosome lOq only (15). In thisIII.study,we describe an additional 12 cases of meningioma with chro- The results of this investigation may suggest a genetic modelofmosome

10 allele loss, including 4 cases in which LOH was confined malignant progression in meningiomas. We found correlations beto chromosome lOq, and define a minimal region on distal chromo- tween allelic losses on chromosomes 14q, lOq, and ip and notedthatsome

lOq (10q24—qter) deleted in all cases with LOH for marker on the frequency of deletions in all three chromosomal regions increasedchromosome 10. Thus, our results map a potential tumor suppressor with grade. The deletion frequencies observed among tumors of

locus to a region commonly deleted in a variety of tumors. different grades indicate a relatively “early―and stage-specific role forRecently, Rempel et a!. (15) have reported frequent loss of markers loci on chromosome 14. In contrast, LOH for loci on chromosomes ip

on chromosome 10 in morphologically malignant meningiomas. In and lOq appears to be predominantly (although not exclusively, comtheir study, such deletions were not seen in five cases with brain pare cases 700, 788, and 834) selected for during the later stages of theinvasion but without other features of malignancy. Based on these development of malignancy. Interestingly, chromosome ip and lOqresults, it was suggested that different genetic loci might be respon- allele loss also appear to be associated with late stages of tumorsible for the development of brain invasion and morphological malignancy in meningiomas (15). Our data neither support nor detract progression in melanomas and astrocytomas (38, 42—44).

The detection of chromosome 10 allele loss in a benign tumor thatfrom this hypothesis, because the present series includes only twononanaplastic tumors with brain invasion (cases 2313 and 3030). subsequently underwent malignant transformation (cases 700, 788,However, in agreement with the findings of Rempel et a!., we ob- and 834) might indicate that in M I, the low frequency of LOH at lociserved no deletions on chromosome lOq in these cases. Both tumors associated with malignant progression does not necessarily reflect

random losses. These mutations might well predispose to or reflectalso retained heterozygosity for loci on chromosomes ip and 14q.LOH for loci on chromosome 22 and mutations in the NF2 locus ongoing malignantprogression.probably

do not play a role in the development of malignancy in The observations made in the tumors of two patients with documeningiomas. Based on the high and roughly equal frequency of allele mented dedifferentiation from a benign meningioma (cases 700,788,loss

for markers from chromosome 22 and NF2 mutations, a common 834, and 2068) indicate that LOH for loci on chromosomes 14q,ip,originfor M III, M II, and the fibroblastic and transitional subtypes of and lOq can be reflected in morphological progression. ThissuggestsM

I was recently proposed (1 1). In our series, the low incidence of a common genetic pathway for the formation of M hIM III. ItisLOHamong M I for markers on chromosomes ip, lOq, and 14q did conceivable that either a de novo M II/M III or a primary M Iandnot

allow for a correlation with histological subtypes. Thus, this study subsequent tumor progression would be observed, depending ontheTable

3 Extent of chromosome 10 allele loss in ninemeningiomas°Tumor/Grade'3075/

2068/ 2275/ 2576/ 3138/ 102/ 2062/2226/I.,ocus@'I..ocation I II II II II III III III3408'@D10S179

lOp 12 12 11 11 LOH 11 LOH LOHLOHD10S89lOpter—pll.212DIOSII1I0pter—pll.2 12 12 12 LOH 11 11 LOHLOHD1OSI69lOqll.2—qter LOH11D10S18710q24-q25 LOH LOH LOH LOH 12 LOH LOHLOHD10S209l0q24-q25 LOH LOH LOH LOH 11 LOH LOH LOHIia

Tabledoesnotincludetumors700,788,and834.LOHonchromosome10couldbedemonstratedinthesetumors,becauseahemi/homozygousbandpatternwasseenatfiveloci.Thehigh frequency of heterozygosity at these loci makes it unlikely that this patient was a compound homozygote. Lack of normal control did not allow for deletion mapping inthesetumors.h

Genetic map of the Id: pter-D1OSI 79-D10S89-D1OSI II-cen-D10S169-DIOSIS7-DIOS2O9-qter.(

I (II, III), WHO grade; LOH, loss of heterozygosity; ii, not informative; 12, informative and no LOH.

d No grade could be assigned based on the available sample. Previous specimen was diagnosed as atypicalmeningioma.4700

ALLELIC LOSSES IN MENINGIOMA PROGRESSION

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ALLELIC LOSSES IN MENINGIOMA PROGRESSION

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4701

time elapsed between mutations leading to tumor formation andmutations necessary for the development of malignancy.

In conclusion, the results of this investigation outline a model ofmeningioma progression in which specific chromosomal deletionshave been implicated. Further characterization of these mutationsmight facilitate the early identification of those tumors with a highintrinsic risk ofprogression. In these cases, timely and aggressive surgicalintervention, or even adjuvant treatment of the tumor, might preventmalignant progression and its associated adverse clinical course.

ACKNOWLEDGMENTS

We thankProf.N. de TriboletandDr.D. A. Ross fortumorsamplesandDr.M. 0. Balko for helpful suggestions. We are grateful to Xiaomin Wu forskillful technical assistance.

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1995;55:4696-4701. Cancer Res   Matthias Simon, Andreas von Deimling, Jeffrey J. Larson, et al.   ProgressionMalignant Meningiomas: A Genetic Model of Meningioma Allelic Losses on Chromosomes 14, 10, and 1 in Atypical and

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