Microsatellite instability analysis of primary human brain tumors.

Microsatellite instability, as shown by the presence of additional alleles or shifts of electrophoretic mobility at simple sequence tandem repeat loci, has been demonstrated in hereditary and sporadic colorectal tumors and many other tumor types. To study microsatellite instability in human brain tumors, we examined a total of 144 sporadic neoplasms. These included 33 astrocytic tumors, 23 oligodendrogliomas, six gangliogliomas, 41 meningiomas, 10 vestibular schwannomas and 31 pituitary adenomas. Di-, tri- and tetranucleotide repeat microsatellite markers localized on chromosome 4 and 9, X, 13 and 22, respectively, were used to assess whether instability was a significant aspect of their abnormal chromosomal pattern. Instability of microsatellite markers was detected in four oligodendrogliomas (17.4%), one pituitary adenoma (3.2%), one meningioma (2.4%), one astrocytic tumor (3.0%) and not at all in gangliogliomas and schwannomas. Therefore, our results suggest that the microsatellite instability which occurs in colorectal cancers with defective mismatch repair is infrequent in many types of human brain tumors and that the lower level of instability observed in brain tumors may be reflective of other mechanisms of genetic instability.

Dopamine D1, dopamine D2, and prolactin receptor messenger ribonucleic acid expression by the polymerase chain reaction in human meningiomas.

Previous studies have suggested the presence of high-affinity dopamine D1 receptors and prolactin receptors in human cerebral meningiomas. In this study, using the polymerase chain reaction, we report the presence of the messenger ribonucleic acid (mRNA) for the dopamine D1 and D2 receptors and the prolactin receptor in meningioma tissue specimens and cell cultures derived from meningioma tissue. Dopamine D1 receptor mRNA was present in a majority of female tissue specimens and in all male tissue specimens. D2 receptor mRNA was detected in all specimens examined. Prolactin receptor mRNA was present in a little more than half of the female and male meningioma tumor specimens. The polymerase chain reaction products were directly sequenced to confirm the identity of these receptors in meningiomas and cell cultures. Ligand binding studies confirmed the presence of the dopamine D1 receptor in meningioma tissue specimens. In contrast, receptor studies with the dopamine D2 ligand [125I]4-iodospiperone failed to detect D2 binding in meningioma membrane preparations. These results suggest the existence of active dopamine D1 receptors in cerebral meningiomas.

Steroid hormone receptors in astrocytic neoplasms.

The presence of specific steroid hormone-binding receptors has been correlated with the clinical response to hormonal therapy in a number of different neoplasias, including breast and prostate cancer. In this article, we investigated the expression of the androgen, estrogen, glucocorticoid, and progesterone receptor messenger ribonucleic acid (mRNA) and protein in a number of astrocytic neoplasms of various histological grades. Androgen and glucocorticoid receptor mRNA were detected in all astrocytic neoplasms examined, regardless of histological subtype. In contrast, progesterone receptor mRNA was observed more frequently in high-grade tumors than in low-grade tumors. Estrogen receptor mRNA was undetectable in all astrocytic tumors examined. These studies suggest a possible adjunct clinical use of hormonal therapy for the treatment of astrocytomas. Specific antagonists and agonists may allow the modulation of the growth of these tumors. Development of this body of knowledge may lead to the development of better treatment for these aggressive tumors.

Analysis of meningiomas by methylation- and transcription-based clonality assays.

The clonal derivation of tumors can be determined by X chromosome inactivation analysis based on differential expression of genes or differential methylation of cytosine residues in CpG islands near polymorphic loci. In this report, we compared a transcription-based RNA analysis with a methylation-based DNA assay to determine clonality of meningiomas. Both clonality assays use PCR-based analysis at the hunan androgen-receptor gene (HUMARA) on the X chromosome. Among 23 meningiomas from female patients, 19 were informative heterozygotes at this locus (83%). The patterns of X chromosome inactivation in four patients were extremely skewed towards one allele in blood (unequal Lyonization), which precluded clonality determination in the tumor samples. Concordant clonality results with methylation- and transcription-based clonality assays were demonstrated in 9 of 13 informative tumors expressing the androgen receptor. Seven meningiomas were monoclonal, but surprisingly, two pathologically documented cases of meningiomas were polyclonal. There was disparity in 4 of 13 tumor specimens that were polyclonal by the methylation-based assay but monoclonal by the transcription assay. Clonality examination of these tumors by the methylation-based phosphoglycerate kinase assay provided identical results to the methylation-based analysis at the HUMARA locus. In addition, loss of heterozygosity (LOH) studies of chromosome 22, which is frequently deleted in meningiomas, showed that four of four informative samples of the six polyclonal tumors had partial LOH in tumor tissues. However, complete LOH was observed in primary cultured cells, which were also monoclonal by the methylation assay. Taken together, these data suggest that the disparity of the two assays in these four cases may be due to differences in the level of expression of the androgen receptor gene in tumors. Therefore, we conclude that: (a) clonal derivation of meningiomas determined by both transcription- and methylation-based clonality assays are in full agreement in many (9 of 13) but not all cases (4 of 13); and (b) most meningiomas (9 of 15) are monoclonal in origin, whereas some meningioma samples (6 of 15) are polyclonal or may contain heterogeneous components.

Progesterone and glucocorticoid receptor activation in meningiomas.

The possibility that the female sex steroid progesterone plays a role in meningioma proliferation has been suggested by a number of investigators, and it has been shown that many meningiomas have high-affinity progesterone binding sites. There has been a long-standing debate in the literature as to whether the progesterone receptors that are present in meningiomas are functional. We recently showed, by the use of immunohistochemistry, that the progesterone receptor in meningiomas is localized to the nucleus, suggesting that the receptor is in a location to be activated. In this study, eight meningioma cell cultures were transiently transfected with a construct that contains two palindromic progesterone/glucocorticoid response elements in front of the thymidine kinase promoter and the chloramphenicol acetyl sequence of the tyrosine aminotransferase gene. In all meningioma cell cultures, an increase in the transcription of the progesterone response element construct was observed in the presence of dexamethasone, suggesting that the glucocorticoid receptor in meningiomas is functional. An increase in transcription was observed with the addition of promegestone (R5020), a progesterone agonist, only in meningioma cell cultures that were expressing the progesterone receptor. These data show that both the progesterone and the glucocorticoid receptor in meningiomas are functional and support the concept that progestins and glucocorticoids may play an important role in meningioma growth.

Expression of platelet derived growth factor and platelet derived growth factor receptor mRNA in a glioblastoma from a patient with Li-Fraumeni syndrome.

Expression of platelet derived growth factor (PDGF) and PDGF-receptor mRNA was examined from a glioblastoma taken from a patient with Li-Fraumeni syndrome. Northern blot analysis and in situ hybridisation showed very high concentrations of both PDGF-A and PDGF alpha-receptor mRNA in the tumour. The overall pattern of PDGF expression was similar to those found in sporadic glioblastomas. Mutations in p53 has been implicated as an early pathogenic event leading to sporadic low grade astrocytomas, and is the third most common tumour type in patients with Li-Fraumeni syndrome, where they are predisposed due to a germline mutation in the p53 tumour suppressor gene. This study suggests that progression towards a glioblastoma in both the general population and in patients with Li-Fraumeni syndrome may involve potential autocrine and paracrine stimulation by growth factors such as PDGF.

Androgen receptor expression in meningiomas.

The predominance of meningiomas in females, the accelerated growth of these tumors during the luteal phase of the menstrual cycle and during pregnancy, and the association between meningiomas and breast cancer have led to a number of studies examining the potential role of steroids in the growth of meningiomas. The possibility that androgens play a role in meningioma proliferation has been suggested by a small number of investigators. The aim of this study was to examine the expression of androgen receptor messenger ribonucleic acid (mRNA) and correlate it using immunochemistry with the nuclear localization of androgen receptor in a large number of meningiomas. Thirty-nine meningiomas were examined by Northern blot analysis for the presence of measurable amounts of androgen receptor mRNA and eight of these were analyzed by immunohistochemistry for receptor protein. Sixty-seven percent of the meningiomas expressed androgen receptor mRNA. There was a marked predominance of women among the patients whose tumors expressed androgen receptor; 69% were women and 31% were men. The immunohistochemical data correlated with Northern blot analysis of mRNA. The staining was predominantly nuclear, suggesting that the androgen receptor resides in a location that can activate gene expression.

Expression of PDGF and PDGF receptors in human astrocytoma operation specimens supports the existence of an autocrine loop.

Established cell lines derived from human malignant astrocytomas typically express a combination of platelet-derived growth factor (PDGF) and PDGF receptor which could form an autocrine loop. In this study, we screened for the essential components of a PDGF autocrine loop in fresh surgical isolates of human astrocytomas, using in situ hybridization and immunohistochemical techniques. Eight malignant astrocytomas (6 glioblastomas and 2 anaplastic astrocytomas), 5 low-grade astrocytomas and 4 non-neoplastic glial specimens (mesial temporal sclerosis) were evaluated. Malignant astrocytomas, and to a lesser extent low-grade astrocytomas, expressed more PDGF-A and PDGF-B than non-neoplastic glia. PDGF-alpha-receptor expression was elevated both in malignant and in low-grade astrocytomas. These data support the argument that PDGF autocrine loops contribute to the unregulated growth of human astrocytomas. Expression of PDGF and PDGF receptor in low-grade astrocytomas suggests that activation of PDGF autocrine loops may be an early event in the pathogenesis of malignant astrocytomas.

Platelet-derived growth factor expression and stimulation in human meningiomas.

The platelet-derived growth factor (PDGF) family consists of subunits A and B and receptors alpha and beta. This paper evaluates the potential role of the homodimer PDGF-BB as a growth factor in meningiomas. It analyzes the expression of messenger RNA in members of the PDGF family in these tumors, measures the growth response of meningiomas to exogenous PDGF-BB in culture, and examines the induction of the c-fos proto-oncogene by PDGF-BB. Northern blot analysis was carried out on tissue from 20 meningiomas to measure the expression of PDGF-A, PDGF-B, PDGF-alpha receptor (PDGF-alpha-R) and PDGF-beta receptor (PDGF-beta-R). All tumors expressed PDGF-A and PDGF-B subunits. Nineteen of the 20 tumors expressed PDGF-beta-R and none expressed PDGF-alpha-R as measured by this technique. Because the beta receptor is selectively sensitive to stimulation by the PDGF-B subunit, these data suggest that meningiomas might be susceptible to stimulation by PDGF-BB. To test this hypothesis, the effect of exogenous PDGF-BB on meningioma growth was evaluated by incubating cells from 10 human meningiomas. Tritiated thymidine incorporation was used to evaluate stimulation of growth over a 48-hour period using PDGF-BB concentrations of 1, 3, or 6 ng/ml. Linear regression analysis and multiple-factor analysis of variance were used to measure PDGF-BB effects. Three of the 10 tumor specimens responded significantly to PDGF-BB, with a three- to sixfold increase in thymidine incorporation over 72 hours of exposure, and there was a significant overall growth-stimulating effect of PDGF-BB in the 10 tumor specimens tested. In the last set of experiments, the functionality of the PDGF-beta-R was determined by examining the induction of the proto-oncogene c-fos by PDGF-BB in meningioma cell cultures. A significant increase in c-fos protein was observed 3 hours after PDGF-BB addition. These findings demonstrate that PDGF-A, PDGF-B, and PDGF-beta-R are expressed in meningiomas and suggest that the beta receptor is functional: when it is activated, c-fos levels are increased, and an increase in meningioma cell division is observed after the addition of PDGF-BB. These studies support the hypothesis that PDGF acts as a growth factor in meningiomas.

Messenger ribonucleic acid expression of platelet-derived growth factor subunits and receptors in pituitary adenomas.

Little is known about the expression of growth factors and their receptors in pituitary tumors or the relationship of growth factors to pituitary neoplasia. Platelet-derived growth factor (PDGF) is a potent mitogen that has been postulated to stimulate tumor growth through autocrine and/or paracrine loops in a number of human tumors. In the present study we demonstrate messenger ribonucleic acid expression of the PDGF subunits and receptors in a variety of human pituitary adenomas and in a normal human anterior pituitary gland. Northern blot analysis performed on 34 pituitary adenomas showed that all tumors expressed the PDGF-A and PDGF-B subunits, the majority (94%) expressed the PDGF-beta receptor, and a subset (44%) expressed the PDGF-alpha receptor. The normal anterior pituitary studied expressed all of the PDGF subunits and receptor subunits. Clinically, there was no correlation between expression of the PDGF subunits or receptors and tumor size or with invasion into adjacent structures (cavernous sinus, sphenoid sinus, or clivus). A higher proportion of the endocrinologically active adenomas expressed the PDGF-alpha receptor (5 of 8) compared to the endocrine inactive tumors (10 of 26). The function of PDGF in the pituitary is currently not known. The finding that PDGF subunit and receptor messenger ribonucleic acids are coexpressed in pituitary adenomas and normal anterior pituitary suggests that autocrine and/or paracrine loops involving PDGF may exist in pituitary adenomas and normal anterior pituitary.

Progesterone receptor expression in meningiomas.

The possibility that the female sex steroid progesterone plays a role in meningioma proliferation has been suggested by a number of investigators; and it had been shown that many meningiomas have high affinity progesterone binding sites. The aim of this study was to examine the expression of progesterone receptor mRNA and correlate it with the nuclear localization of progesterone receptor by immunocytochemistry in a large number of meningiomas. Thirty-three meningiomas were examined for the presence of measurable amounts of progesterone receptor mRNA by Northern blot analysis and 11 of these were analyzed for receptor protein by immunohistochemistry. These were compared with normal arachnoid and an arachnoid cell line. Sixty-four % of the meningiomas expressed progesterone receptor mRNA. This occurred in a similar pattern to what has previously been shown for T-47 D cells, a breast carcinoma cell line with 11.4-, 6.1-, 5.2-, 4.5-, 3.2-, and 2.5-kilobase mRNA species. This suggests that progesterone receptor expression is not tumor specific. There was a marked predominance of women among those patients whose tumors expressed progesterone receptor; 81% were female and 19% were male. The immunohistochemistry data correlate well with the Northern blot analysis. The staining was clearly nuclear, suggesting that the receptor is in a location to be activated. These data suggest that progesterone receptor mRNA and protein is expressed in meningiomas and support the concept that progesterone may play an important role in meningioma growth.

MRI guided interstitial laser therapy in a rat malignant glioma model.

We have used MR monitoring to guide and evaluate the effects of the Nd:YAG interstitial laser on a well-characterized rat brain tumor model (C6). MRI was used to determine the tumor size, verify the location of the interstitial probe, and evaluate the size and location of the laser-induced lesion during and after treatment. During laser irradiation, an irreversible loss of signal intensity at the fiber tip and a reversible decrease in signal intensity around it were observed with MRI. None of the treatment protocols affected mean rat survival significantly. Although MRI-guided interstitial laser therapy appears to be safe and easy, it does not provide a curative treatment for spatially disseminated gliomas where a "target volume" cannot be adequately defined. Better results can be expected, especially in well-defined tumors, with improvements of both the imaging techniques and the laser treatment protocol.

Distribution of hematoporphyrin derivative in canine glioma following intraneoplastic and intraperitoneal injection.

Hematoporphyrin derivative (HPD) is a photosensitizing agent that has been used to locate and kill tumors. The distribution of tritiated (3H)-HPD was studied in a transplantable canine glioma model following intraperitoneal or direct intraneoplastic injection. Compared to intraperitoneal administration of 3H-HPD, direct injection resulted in levels that were more than 2.5 times higher in tumor tissue and approximately 10 times lower in skin. Dose-corrected analysis of the data indicated that outside the central nervous system (CNS) the distribution of 3H-HPD is dose-related, regardless of route of injection. Within the CNS, direct injection leads to more efficient uptake of 3H-HPD, especially at the tumor periphery. Fluorescence microscopy confirmed the selective biodistribution of HPD fluorescence within the cytoplasm of tumor cells.