CLONAL ANALYSIS OF GLIOMAS

In malignant gliomas, the characteristically heterogeneous features an d frequent diffuse spread within the brain have raised the question of whether malignant gliomas arise monoclonally from a single precursor cell or polyclonally from multiple transformed cells forming confluent clones. Although monoclonality has been shown in surgically resected tissues, these may not include the full spectrum of patterns seen on a utopsy material. Little is known about the clonality of low-grade glio mas from which malignant gliomas may sometimes arise. We sought to inv estigate the clonality of low-grade and malignant gliomas by using and comparing surgical and autopsy material with a Polymerase chain react ion (PCR)-based assay for nonrandom X chromosome inactivation. For tha t, purpose, archival surgical and autopsy material from 15 female pati ents (group A) (age 4 to 73 years; median, 45) with malignant gliomas (12 glioblastomas, one gliosarcoma, one anaplastic oliogastrocytoma, o ne gliomatosis cerebri), surgical material only from 21 female patient s (group S) (age 6 to 78 years; median, 60) with low-grade and maligna nt gliomas (four low-grade astrocytomas, three oligoastrocytomas, two anaplastic astrocytomas, one gemistocytic astrocytoma, four oligodendr ogliomas, seven glioblastomas) were analyzed. In group A, representati ve areas (mean = 5/patient; median = 7) were microdissected from tissu e sections and assayed by PCR amplification of a highly polymorphic mi crosatellite marker locus of the human androgen receptor gene (HUMARA) in the presence of alpha(32)P With and without predigestion with a me thylation-sensitive restriction enzyme (HhaI). Products were resolved by denaturing gel electrophoresis and autoradiographed. In group S, se lected tumor areas were used for the assay. Each patient's normal brai n tissue was used for control. The band intensity of alleles were meas ured by densitometric scanning. In group A, 13 of 15 cases were inform ative (heterozygous). The same pattern of nonrandom X chromosome inact ivation was present in all areas of solid dense and moderate tumor inf iltration in eight including all components of the gliosarcoma. Two of eight also showed focal loss of heterozygosity (LOH). One of 13 prese nted global LOH. Two of 13 showed microsatellite instability, one of w hich in a patient with Turcot syndrome, the other in gliomatosis cereb ri. Opposite skewing patterns were seen in distant areas of gliomatosi s cerebri consistent with oligoclonal derivation. Clonality remained i ndeterminate in one glioblastoma and in the anaplastic oligoastrocytom a because of skewed lyonization in the normal control. In group S, 19 of 21 cases were informative. Fifteen of 19 were monoclonal (four low- grade astrocytomas, one anaplastic astrocytoma, one gemistocytic astro cytoma, two oligodendrogliomas, one oligoastrocytoma, six glioblastoma s). Four of 19 were indeterminate. We conclude that (1) Low-grade and malignant gliomas are usually monoclonal tumors, and extensively infil trating-tumors must result from migration of tumor cells (2) Gliomatos is cerebri may initiate as an oligoclonal process or result from colli sion gliomas (3) Biphasic gliomas likely arise from a single precursor cell. (4) LOH at the HUMARA locus is probably related to partial or c omplete deletion of an X-chromosome, which occurs in malignant gliomas during clonal evolution. Copyright (C) 1997 by W.B. Saunders Company.