Astrocytomas and Choroid Plexus Tumors in Two Families with Identical p53 Germline Mutations
ANNE VITAL, MD, PIERRE-PAUL BRINGUIER, PHD, HUATAO HUANG, PHD, FRANÇOIS SAN GALLI, MD, JANINE RIVEL, MD, SOPHIE ANSOBORLO, MD, JEAN-MARIE CAZAURAN, MD, LUC TAILLANDIER, MD, PAUL KLEIHUES, MD, AND HIROKO OHGAKI, PHD
Abstract. Germline p53 mutations carry an increased risk of development of breast cancer, soft tissue and osteosarcomas, brain tumors, leukemia and adrenocortical carcinomas. Cerebral neoplasms are usually of astrocytic lineage and occur in 40% of affected families. This report presents clinical, neuropathological and molecular genetic data from 2 families in France with an identical p53 germline mutation in codon 248 (CGG→TGG; Arg→Trp) and a clustering of CNS tumors. The youngest patient in each family developed a malignant choroid plexus tumor while several young adults of both kindred succumbed to low-grade astrocytoma, anaplastic astrocytoma or glioblastoma. The only non-neural neoplasm was an adrenocortical carcinoma in a boy aged 4 years who developed an anaplastic choroid plexus papilloma 2 years later. Of 2 previously reported inherited choroid plexus tumors, I occurred in a family which also carried a germline mutation in codon 248. It remains to be shown whether this unusual pattern of CNS tumors is due to an organ-specific effect of this particular p53 mutation or whether it reflects the genetic background of the affected families.
Key Words: Astrocytoma; Choroid plexus carcinoma; Glioblastoma; p53 germline mutation.
INTRODUCTION
Since the discovery in 1990 of p53 germline mutations as the underlying cause of the Li-Fraumeni syndrome (1, 2), the genetics, age and tumor pattern of affected family members have been extensively studied (1-4). Our recent analysis of 599 tumors in 124 families (5) showed that breast carcinomas are most frequent (25% of all reported neoplasms), followed by sarcomas (22%), and brain tu- mors (14%). Leukemia and adrenocortical carcinoma, al- though very typical for the Li-Fraumeni syndrome, amount to less than 5% of tumors (3). Familial clustering of 4 or more cases of the same tumor type has been ob- served in 15% of kindred (5). In families with at least 1 case of brain tumor, the mean number of CNS tumors per family is 1.5 (6) and so far, only 6 families with 3 or more brain tumors have been reported (2, 7-11). We here pre- sent 2 new families with a remarkable clustering of brain tumors, including choroid plexus tumors in young children and astrocytomas in young adults. Although apparently unrelated, both families carried an identical germline mu- tation in codon 248 of the p53 tumor suppressor gene.
FAMILY AND CASE HISTORIES Family 1
Patient F (Fig. 1), a boy, and the only child of parents D and E, was well until the age of 4 when he presented
From the Departments of Neuropathology (AV, JR), Pediatrics (SA), and Neurosurgery (FSG), Victor Segalen University, 33076 Bordeaux, France, Pediatrics Department, Périgueux Hospital, 24019 Périgueux Cedex, France (JMC), Neurology Department, Saint Julien Hospital, 54000 Nancy, France, (LT), and the International Agency for Research on Cancer, 69372 Lyon, France (PPB, HH, PK, HO).
Correspondence to: Dr. Hiroko Ohgaki, Unit of Molecular Pathology, International Agency for Research on Cancer, 150 Cours Albert-Tho- mas, 69372 Lyon Cedex 08, France.
premature puberty related to a right adrenocortical car- cinoma. The tumor appeared well-circumscribed and was surgically removed in total. Regular follow-ups including abdominal echography and hormonal analyses every 3 months revealed neither recurrence nor metastasis. At the age of 6 years, the patient began complaining of cephalea that were associated with behavioral changes. Brain CT and MRI scans revealed a well-circumscribed hyperdense tumor mass without calcifications in the occipital horn of the left ventricle (Fig. 2A). There was neither hydro- cephalus nor any sign of raised intracranial pressure: The tumor was surgically removed and histologically classi- fied as atypical choroid plexus papilloma. One year after surgical resection, there are no signs of disease progres- sion.
The grandmother (B) and grandfather (A) of the patient are well but his grandmother’s sister (C) died of a brain tumor between the age of 40 and 50 years; the histolog- ical typing of the neoplasm is unobtainable. The mother (E) of patient F is well but his father (D) died of a frontal anaplastic astrocytoma when he was 23-years-old. The father’s 2 sisters are alive and well. This family is from Nancy in northeast France.
Family 2
Clinical investigations of the family history were initi- ated after patient H (Fig. 1), a girl, at the age of 8 months, presented signs of increased intracranial pressure, behav- ioral changes, axial hypotonia, and generalized epileptic seizures. The CT scan showed a marked hydrocephalus, caused by a large parieto-occipital tumor occupying the left ventricle. On MRI scans, the lesion showed partly cys- tic, with inhomogeneous contrast-enhancement (Fig. 2B). Several additional contrast-enhancing lesions were noticed
VITAL ET AL
Family 1
A
B
C 40-50 yr Brain tumor
0 23 yr Anaplastic astrocytoma mut/-in tumor
E
F
mut/wt in blood 4 yr Adrenocorticol carcinoma mut/-In tumor 6 yr Chorold plexus papilloma mut/-in tumor
Family 2
+
A
36 yr Low-grade astrocytoma 37 yr Glioblastoma
8 47 yr Low-grade astrocytoma mut/-in tumor
C 29 yr Anaplastic astrocytoma mut/-in tumor
D
£
mut/wt In blood 29 yr
F wt/wt in blood
G
H
mut/wt in blood 8 mo
Choroid plexus carcinoma mut/-In tumor
wt/wt in blood 3mo
1
in the meninges of the posterior fossa. The tumor was surgically resected, and a ventriculoperitoneal shunt was put in place. At gross examination, the tumor appeared partly fleshy and partly cystic, with necrotic and hemor- rhagic areas. Histologically, the lesion was diagnosed as choroid plexus carcinoma (WHO Grade III). The patient underwent postoperative fractionated chemotherapy, but 2 months after surgery, symptoms of increased intracranial pressure recurred. The CT scans evidenced severe hydro- cephalus, and persistence of a heterogeneous lesion facing the superior part of the occipital horn of the left ventricle, MRI also revealed an enlargement of the secondary men-
ingeal lesions facing the posterior fossa. Malignant tumor cells were found in the ventricular CSF Clinical symptoms rapidly deteriorated, and the patient died at the age of 13 months.
The patient’s grandmother (D) is well but her husband (C) died of an anaplastic astrocytoma (WHO Grade III) at the age of 29. Two of his brothers (A and B) also died from brain tumors. In patient (A), the brain tumor was diagnosed as low-grade astrocytoma (WHO Grade II) when he was 36-years-old, but recurred as glioblastoma (WHO Grade IV) 1 year later. The other brother (B) de- veloped a low-grade astrocytoma at the age of 47 years.
DI4 38 4
A
B
The parents (E, F) and siblings (G, I) of patient H are healthy. This family is from southwestern France, and has lived in a small rural area for several generations.
MATERIALS AND METHODS Immunohistochemistry
The sections were deparaffinized in xylene and rehydrated in graded ethanol. For glial fibrillary acidic protein (GFAP) im- munohistochemistry, endogenous peroxidase was blocked by 3% H2O2 in absolute methanol for 20 min. The sections were preincubated with normal swine serum (diluted 1:5 in PBS, DAKO, Trappes, France), followed by incubation with the pri- mary antibody for 60 min (Cambridge Research Biochemicals, Cambridge, UK). After rinsing in PBS, sections were incubated with swine anti-rabbit, biotinylated for 30 min (DAKO, Trap- pes, France). To label proliferating cells, a monoclonal antibody MIB-1 against recombinant parts of the Ki-67 antigen (Diano- va, Hamburg, Germany) was used. Sections were boiled 3 times for 5 min in a microwave oven at 750 W. The slides were cooled for 30 min, rinsed twice in distilled water, and incubated in PBS for 5 min. The sections were reacted for 1 hour at room temperature with MIB-1 antibody (diluted 1:50 in PBS). Pre- albumin immunohistochemistry was carried out using rabbit anti-human prealbumin antibody (DAKO, diluted 1:1,000 in PBS). For all immunohistochemistry, the reaction was visual- ized using the Vectastain ABC kit and diaminobenzidine (Vec- tor Laboratories, Burlingame, Calif.). Sections were then coun- terstained with hematoxylin.
PCR-Single Strand Conformation Polymorphism (SSCP) and DNA Sequence Analyses for p53 Mutations
DNA was extracted from typical tumor areas as previously described (12). PCR for exons 5 to 8 of the p53 gene were carried out as previously described (12). For SSCP, 4 ul of PCR products were mixed with 10 ul loading buffer (95% form-
amide, 20 mM EDTA, 0.05% xylene cyanol and bromophenol blue), denatured at 95℃ for 5 min, and quenched on ice until loading. Samples were run at 40 W with cooling on a 6% ac- rylamide (49:1) containing 6% glycerol. Samples which showed abnormal mobility shifts were further analyzed by direct DNA sequencing using the Sequenase PCR Product Sequencing Kit (USB) following the manufacturer instructions (labelling with 3 dNTPs) except that the termination reaction was carried out at 42°C.
Detection of SV40 Sequences
The presence of SV40 was assessed using a PCR-Southern hybridization method that reliably distinguishes between SV40, JC and BK large T antigen sequences (13).
RESULTS Surgical Pathology
Brain tumor typing was carried out according to the WHO Classification (14) in conjunction with the St. Anne/Mayo grading criteria for diffuse astrocytomas (15).
Family 1
Patient D: The biopsy showed a histologically hetero- geneous neoplasm. In some tumor areas, the features were those of a low-grade diffuse fibrillary astrocytoma with moderate nuclear atypia, extensive microcystic de- generation but absence of mitoses. In other areas, the cel- lularity was greatly increased, with occasional binuclear cells, gemistocytic neoplastic astrocytes, and mitotic ac- tivity. There was no necrosis or microvascular prolifera- tion. Based on the more malignant areas of the neoplasm, it was designated anaplastic astrocytoma (WHO Grade III).
Patient F: The biopsy revealed a neoplasm largely re- sembling the histological features of the normal choroid plexus epithelium. In most parts, there was a single layer of epithelial cells but in other parts the architecture was less regular and multiple layers of tumor cells were oc- casionally observed (Fig. 3a). Mitoses were infrequent but MIB-1 immunohistochemistry showed focally a con- siderable accumulation of proliferating tumor cells (Fig. 3b). This lesion appeared to occupy an intermediate po- sition between the choroid plexus papilloma (WHO grade I) and carcinoma (WHO grade III) and was termed “atyp- ical choroid plexus papilloma.” The resected adrenal gland showed the typical features of a highly polymor- phic carcinoma of the adrenal cortex. Tumor cells were often multinucleated, hyperchromatic and extremely an- aplastic, but mitotic activity was moderate. The capsule of adrenal gland was largely intact and there was no ev- idence of invasion of tumor vessels.
Family 2
Patient A: The first biopsy, taken at age 36, showed an astrocytic neoplasm of low cellularity with occasional
0
a
b
C
1
e
nuclear atypia (Fig. 3c). The tumor cells appeared on the background of a loose fibrilliary matrix, which was strongly immunoreactive to GFAP. In some areas, neo- plastic gemistocytic astrocytes (gemistocytes) were ob- served. There was no evidence of necrosis or vascular
proliferation. The tumor was diagnosed as diffuse low- grade astrocytoma (WHO Grade II).
In the second biopsy 1 year later, the histological fea- tures were distinctly different. The biopsy showed a high- ly cellular tumor with a marked degree of anaplasia and
| Family | Family member* | Age at diagnosis | Tumor | p53 Mutation ** | SV40 | ||
|---|---|---|---|---|---|---|---|
| Tumor | White blood cell | Tumor | Blood | ||||
| 1 | C | 40-50 years | Brain tumor, unclassified | n.d. | n.d. | n.d. | n.d. |
| D | 23 years | Anaplastic astrocytoma | mut/- | n.d. | + | n.d. | |
| F | 4 years | Adrenocortical carcinoma | mut/- | n.d. | n.d. | n.d. | |
| 6 years | Atypical chloroid plexus papilloma | mut/- | mut/wt | + | - | ||
| 2 | A | 36 years | Low-grade astrocytoma | n.d. | n.d. | n.d. | n.d. |
| 37 years | Glioblastoma | n.d. | n.d. | n.d. | n.d. | ||
| B | 47 years | Low-grade astrocytoma | mut/- | n.d. | - | n.d. | |
| C | 29 years | Anaplastic astrocytoma | mut/- | n.d. | + | n.d. | |
| E | 29 years | None | mut/wt | + | |||
| F | None | wt/wt | 1 | ||||
| H | 8 months | Choroid plexus carcinoma | mut/- | mut/wt | + | + | |
| I | 3 months | None | wt/wt | - | |||
* Letter refers to family trees in Figure 1.
** CGG → TGG with amino acid substitution of Arg → Trp in codon 248.
n.d., not determined.
heterogeneity. Tumor cell nuclei were often spindle- shaped and arranged in a storiform pattern. There was brisk mitotic activity in virtually all tumor areas. Addi- tional features were marked microvascular proliferation and large areas of ischaemic necrosis. Occasional multi- nucleated giant cells were observed. GFAP expression varied considerably. Occasionally, there was a sharp de- lineation between GFAP expressing glioma cells and ar- eas with very high mitotic activity that lacked GFAP ex- pression (Fig. 3d), suggestive of the emergence of a new tumor clone through the acquisition of an additional al- teration. The neoplasm fulfilled all criteria for the diag- nosis of a glioblastoma multiforme (WHO Grade IV).
Patient B: The biopsy showed the histological features of a low-grade diffuse astrocytoma (WHO Grade II) with low cellularity and moderate nuclear atypia. Tumor cells were embedded in a loose fibrilliary matrix with marked GFAP expression. There was extensive microcystic de- generation, but mitotic activity, necrosis, and microvas- cular proliferation were absent.
Patient C: The biopsy revealed an astrocytic tumor of moderate to high cellularity. Tumor cell nuclei showed a considerable degree of atypia with occasional multinu- cleated giant cells. Whilst most tumor cells resembled the histological features of a fibrillary astrocytoma, some ar- eas showed a marked admixture of gemistocytes. There was strong GFAP expression throughout the biopsy. Mi- totic activity was present but there was no necrosis and no vascular proliferation, allowing the diagnosis of an anaplastic astrocytoma (WHO Grade III).
Patient H: The biopsy revealed a highly pleomorphic, anaplastic epithelial tumor. Its papillary structure resem- bling choroid plexus was still discernible but in some
areas, the tumor had become compact and irregularly structured. There were multiple layers of epithelia with brisk mitotic activity and invasion of neighboring brain structures (Fig. 3e). Immunoreactivity to prealbumin was observed in isolated tumor cells and focally in tumor cell clusters (Fig. 3f). Because of its high degree of anaplasia, mitotic activity and invasive nature, the tumor was di- agnosed as choroid plexus carcinoma (WHO Grade III).
p53 Mutations
In Family 1 (Fig. 1), SSCP was carried out in the an- aplastic astrocytoma of patient D, and the adrenocortical carcinoma, anaplastic choroid plexus papilloma, and pe- ripheral white blood cells (WBC) of patient F. Autora- diographs showed mobility shifts in exon 7 in all these samples, suggesting the presence of a p53 mutation. DNA sequencing revealed that all tumors and the WBC of pa- tient F contained a CGG→TGG mutation in codon 248, resulting in an amino acid substitution of Arg→Trp (Ta- ble 1; Fig. 4).
In Family 2 (Fig. 1), SSCP was carried out on DNA from the low-grade astrocytoma of patient B, the ana- plastic astrocytoma of patient C, and the choroid plexus carcinoma and WBC of patient H. In all tumors analyzed, SSCP autoradiographs showed only abnormally shifted bands, suggesting a loss of the wild-type allele (Table 1; Fig. 5). DNA sequence autoradiographs showed a mu- tation in codon 248 identical to that in Family 1 (CGG- >TGG, Arg→Trp). This mutation was detected in all tu- mor samples and in WBC from patient H. DNA from WBC of the healthy proband E showed bands represent- ing normal and mutated alleles, whereas WBC DNA
ACGTACGT
/C/T G
\G
Tumor
Blood
from nonaffected family members I and F only contained the 2 wild-type DNA strands (Table 1; Fig. 5).
Detection of SV40 Sequences in Brain Tumors
Sequences identical to SV40 large T antigen were identified in 4 out of 5 brain tumors analyzed and in leukocyte DNA from patient H and her father E in Family 2 (Table 1). JC and BK virus sequences were not detect- ed.
Family
2
1
CBEIFIHF
4wt
mut
4 wt
Tu Tu WBC WBC WBC N WBC Tu Tu
DISCUSSION
The 2 families presented in this study show a remark- able clustering of brain tumors. Within 3 generations, 3 CNS neoplasms developed in Family 1, and 4 brain tu- mors in Family 2. In both kindreds, gliomas of astrocytic origin prevailed; in addition, the youngest affected mem- ber of each family developed a malignant choroid plexus tumor.
Our recent analyses of 124 families carrying a p53 germline mutation showed that of a total of 85 brain tu- mors, 50 had been classified histologically (5). Of these, 34 (68%) were of astrocytic origin (low-grade astrocy- toma, anaplastic astrocytoma and glioblastoma), followed by medulloblastomas and related primitive neuroectoder- mal tumors (PNET) of childhood (14%). This corre-
| No. of gener- ations ana- lyzed* | Choroid plexus tumor | Location | Age/sex of patient | p53 germline mutation | Other tumors in the family | Reference |
|---|---|---|---|---|---|---|
| 3 | Choroid plexus car- cinoma | not specified | 15 mo/M | splice site in in- tron 5 | Breast cancer, lung cancer, ovarian carcinoma, colon carcinoma | (17) |
| 2 | Choroid plexus tu- mor | not specified | 1 yr | codon 248 CGG - CAG, Arg -> Gin | Breast cancer, lym- phoma, lung can- cer | (18) |
| 3 | Atypical choroid plexus papilloma | Occipital horn of the left ventricle | 6 yr/M | codon 248 CGG -> TGG, Arg -> Trp | Brain tumor (un- specified), ana- plastic astrocyto- ma, adrenocortical carcinoma | This study (Family 1) |
| 3 | Choroid plexus car- cinoma | Parieto-occipital left ventricle | 8 mo/F | codon 248 CGG-> TGG, Arg -> Trp | Low-grade astrocy- toma, anaplastic astrocytoma, glio- blastoma | This study (Family 2) |
* All the choroid plexus tumors were identified in the last generation.
CLUSTERING OF BRAIN TUMORS IN FAMILIES WITH p53 GERMLINE MUTATIONS
| No. of genera- tions ana- lyzed | p53 Germline mutation | Brain tumors (Age/sex of patient) | Other tumors (Age/sex of patient) | Reference | |
|---|---|---|---|---|---|
| Codon | Mutation | ||||
| 3 | 245 | GGC ++ GAC Gly -> Asp | Unclassified brain tumors (3 cases), schwannoma | Breast cancers (3 cases), os- teosarcoma, polycythae- mia, soft tissue sarcoma, leukemia, colon cancer | (2) |
| 5 | 242 | TGC -> TAC Cys -> Tyr | Unclassified brain tumors (2/M, 21/F), ependymoma (8/M) | Gynecologie tumor (37/F), lung cancer (15/M), sarco- ma (83/M), cancer (58/F), nasopharyngeal tumor (26/M), ovarian cancer (23/F), osteosarcoma (18/ M), cervical cancer (53/F) | (9) |
| 3 | 252 | CTC -> CCC Leu -> Pro . | Unclassified brain tumors (9, 26/M, 38/M) | Breast cancers (32/F, 27/F), soft tissue sarcoma (2/F), osteosarcomas (15/F, 18, 22/M, 22/M), leukemia (17M) | (8) |
| 3 | 236 | 3bp deletion | Anaplastic astrocytoma (27/F), glioblastoma (5/M, 10/F), unclassified brain tumor (9/M) | Leukemia (47/M), adreno- cortical carcinoma (27/F) | (7) |
| 4 | 133 | ATG -> AAG Met -> Lys | Unclassified brain tumors (9/M, 10/F, 25/M) | Breast cancers (26/F, 40/F), gastric cancer (F), malig- nant lymphoma (37/M) | (10) |
| 3 | 213 | CGA + CCA Arg -> Pro | Unclassified brain tumors (5/M, 13/M, 7/F) | Gastric cancer, endometrial carcinoma (48/F), soft tis- sue sarcoma (43/F), breast cancer (48), acute myeloid leukemia (11/F) | (11) |
| 3 | 248 | CGG -> TGG Arg -> Trp | Anaplastic choroid plexus papilloma (6/M), brain tu- mor (40-50/F), anaplastic astrocytoma (23/M), | Adrenocortical carcinoma (4/M) | This study (Family 1) |
| 3 | 248 | CGG - TGG Arg -> Trp | Glioblastoma (37/M), low- grade astrocytoma (36/M, 47/M), anaplastic astrocy- toma (29/M), choroid plexus carcinoma (<1/F) | None | This study (Family 2) |
sponds remarkably well to sporadic brain tumors, in which p53 mutations prevail in astrocytic tumors, and, less frequently, in medulloblastomas/PNETs (16). This is also true for the 2 families reported in this study. Again, most brain tumors were diffuse astrocytomas (Table 1; Fig. 1), ranging from low-grade fibrillary astrocytoma (2 cases), to anaplastic astrocytoma (2 cases) and glioblas- toma (1 case).
In contrast, choroid plexus tumors are rare in families with a p53 germline mutation (3, 6), with only 2 cases reported to date (17, 18). Jolly et al (17) reported a cho- roid plexus carcinoma in a 15-month-old boy from a fam- ily with a splice-site p53 mutation in intron 5, which lead to deletion of exon 6 and a premature stop codon in gene
transcripts (Table 2). Other affected family members de- veloped synchronous bilateral breast cancer, lung cancer, ovarian carcinoma and colon cancer (17). Frebourg et al (18) reported a choroid plexus tumor in a 1-year-old child with a p53 germline mutation in codon 248 (CGG- >CAG) with amino acid substitution Arg→Gln. Breast cancer, lymphoma and lung cancer also developed in this family. It is of interest to note that 3 out of the 4 choroid plexus tumors associated with a p53 germline mutation occurred in the same codon 248 (Table 2). This could reflect a target cell specificity of this genetic alteration for choroid plexus epithelia since the association of co- don 248 mutations with choroid plexus tumors is signif- icant when compared with the frequency of codon 248
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germline mutations in families with astrocytic brain tu- mors (3 out of 27, p < 0.01, Fisher’s exact test). On the other hand, codon 248 mutations are rather frequent, amounting to 14% of all p53 germline mutations reported to date (3), and a similar clustering of codon 248 muta- tions was observed in families with medulloblastomas/ PNETs (3 out of 5).
Familial clustering due to gene-environment interac- tions, e.g. exposure of families to similar carcinogens or life-style factors has been discussed for stomach and breast cancer (3, 5), but is less likely to play a role in the evolution of brain tumors. Details of the neoplasms which developed in 8 families with 3 or more brain tu- mors, including the families presented in this study, are summarized in Table 3. Of these, only 4 families showed a pattern of target organs typical for the Li-Fraumeni syn- drome, i.e. a frequent development of breast cancer, os- teosarcomas and soft tissue sarcomas (19, 20). However, brain tumors, leukemia and adrenocortical carcinomas, although less frequent, also fit well into this familial can- cer syndrome. It is noteworthy that in 6 out of 8 families with clustering of 3 or more brain tumors, p53 germline mutations were located in a relatively small region of exon 7 between codons 236 and 252, which encodes a loop structure in the p53 protein (loop 3) that contains all the residues in direct contact with the minor groove of target DNA (21). It remains to be elucidated to what extent the various combinations of tumors observed in affected families reflect the location of mutations within the p53 gene or the genetic background of the respective kindred, which may contain additional genetic alterations or polymorphism in transformation-associated genes oth- er than p53.
SV40 sequences have recently been detected in brain tumors, particularly in choroid plexus tumors and epen- dymomas (13, 22-24). SV40 is neuro-oncogenic in ham- sters (25). Transgenic mice expressing the SV40 large T antigen under transcriptional control of the viral enhancer develop choroid plexus papillomas (26). We identified SV40 sequences in both choroid plexus tumors that de- veloped in the 2 children carrying a p53 germline mu- tation (Table 1). Furthermore, the blood samples from a child with choroid plexus carcinoma (Family 2, H) and her father (Family 2, E) also contained SV40 sequences. The children and their parents live(d) in different regions of France so that a cross-infection is highly unlikely. However, SV40 contaminated poliovaccine was used from 1955 until 1963 in USA, Canada, and Europe, and this is the generally accepted source of infection (27). It is currently unclear whether SV40 contributes signifi- cantly to the process of malignant transformation in the human CNS or whether its presence constitutes a by- stander infection due to an intra-tumoral microenviron- ment that favors viral replication in individuals with a latent SV40 infection.
ACKNOWLEDGMENTS
This work was supported by a grant from the Foundation for Pro- motion of Cancer Research, Japan. We thank Dr. Jean Floquet. Labor- atoire d’Anatomie et de Cytologie Pathologiques, Centre Hospitalier Universitaire de Nancy and Dr. Jean-Michel Vignaud, Laboratoire d’Anatomie Pathologique, Hôpital Central, Nancy. for providing his- tological specimens.
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Received April 14, 1998 Revision received July 30, 1998 Accepted July 31. 1998