Journal of Neuro-Oncology 10: 129-132, 1991. © 1991 Kluwer Academic Publishers. Printed in the Netherlands.

Special Paper

A summary of the Third Preuss Foundation Conference on brain tumor research February 21-24, 1990

Charles Wilson Professor & Chairman of Neurological Surgery at the University of California, San francisco, 505 Parnassus Ave, San Francisco, CA 94143, USA

Summarizing this meeting has been a daunting but gratifying assignment. Although I performed this role in the past meetings, I had a new sense of excitement as I unfolded the presentations of the last two and one-half days. Whereas the formal presentations were outstanding, some of the dis- cussions were even better.

Throughout the meeting, the astrocyte - fetal and adult, normal and neoplastic - has been the focus of attention. This is understandable, because by their sheer numbers and the opportunity that they provide for studying tumor progression and diversity, astrocytomas completely dominate pri- mary tumors of the CNS.

In this summary, omission of a presentation does not reflect its lack of importance but rather my inability to weave it into the overall fabric of the meeting's major themes.

Appropriately, the meeting began with a search for astrocyte-specific genes and gene products. Al- ready familiar is the enormous value of GFAP. For two decades, GFAP has been used to answer the question, 'what is an astrocyte?' Lipocortin 2 - LC 2 - is another such genetic marker. It is expressed in early fetal development but, with maturation, the gene is no longer expressed in adult astrocytes. Neither is it expressed in differentiated astrocyto- mas, but expression has been found in a small number of anaplastic astrocytomas and in every glioblastoma examined to date. The gene is located on the long arm of chromosome 10 and may con- tribute to the enhanced aggressiveness of glioblas- tomas, which are known to have alterations of this chromosome.

Now there is solid evidence that the consistent finding of genetic markers associated with specific grades of astrocytoma is compatible with the con- cept that tumor progression occurs in association with a series of sequential and cumulative genetic alterations. More specifically, the evolution of an astrocytoma from a benign to a malignant pheno- type is associated with aberration on chromosome 17, followed by amplification and alteration of the EGF receptor, and finally, in glioblastoma multi- forme, the loss of chromosome 10 alleles.

The study of an hereditary neoplastic syndrome, neurofibromatosis, exemplifies a human model for the investigation of multistep mechanisms of pro- gression, whether this is activation of an oncogene or inactivation (elimination) of a tumor suppressor gene. As one example, mutation of the NF-1 gene on chromosome 17q is associated with the clinical expression of neurofibromatosis, type I. A second mutation involving the loss of a gene on chromo- some 17 p leads to the evolution of a neurofibrosar- coma, an uncommon malignant progression of or- dinarily benign neurofibromas. The significance of this observation is the identification of a molecular change reflected in the conversion of a benign tu- mor to a malignant one.

The most completely characterized tumor sup- pressor gene is the retinoblastoma (RB) gene. In a study of molecular interactions of the RB gene product with genes important for the regulation of gene expression during cell proliferation, a 90 base pair region of DNA in the upstream regulatory region of the fos gene confers responsiveness to RB gene expression - mediated by the suppression of

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transcription. This is yet another step in tracing the cascade of molecular interactions by which muta- tions lead to the changes that we see as malignancy.

We were treated to a technical innovation, namely needle micro-inj ection of the ras oncogene product into cultured cells. This technique pro- vides another means of studying genes encoding proteins important in the earliest events following the initiation of cell division.

In the PDGF (platelet derived growth factor) system, 3 dimeric forms containing either A and/or B chains interact with A and B receptors to pro- duce 16 distinct phenotypes, 8 of which are never seen in gliomas. The different combinations of growth stimulating ligand and receptor provide a system that can be finely tuned and which may be different in different glial cells and may contribute to the histologic cellular and regional heterogenei- ty that characterizes malignant gliomas.

Several studies contributed to our appreciation of the complexity of the extracellular matrix (ECM) and its pivotal role in the modulation of cellular processes and ultimately cellular behavior. Reconstituted basement membrane (RBM) on the one hand promotes cell and organ differentiation, e.g., promotes differentiation of endothelial cells to form capillaries, yet on the other hand, RBM promotes tumor cell growth in vivo. One of the best studied components of RBM is laminin, which pro- motes a variety of cellular changes such as the flattening of oat cell carcinoma cells; at the same time, it confers resistance to chemotherapy, pos- sibly explaining the clinical phenomenon of the exquisite sensitivity of oat cell carcinoma to a range of chemotherapeutic agents and yet its resistance to cure. Decorin, a newly identified component of ECM, is a proteoglycan that binds TGF beta (transforming growth factor-beta), although TGF beta stimulates its synthesis. It appears that growth factors and components of ECM may form a feed- back loop for the continuous modification of growth proliferating signals. Two models of tumor cell invasion - reconstituted basement membrane and organotypic cultures - may provide clues to future anti-invasive therapies, e.g., the use of met- alloprotease inhibitors. Clearly, tumor invasion of

normal tissue requires modulation of the normal ECM; metalloproteases are examples of candidate molecules involved in that process.

Is the astrocytoma a disorder of precursor cells? As a concept, the notion of disordered behavior of progenitor cells is inviting, particularly the experi- mental immortalization of 0-2A cells. The condi- tionally transgenic mouse - immortomouse - is a cleverly conceived model system that, while limit- ed by its interferon dependence and temperature requirements, provides a unique new tool for the study of differentiation and the parallel underlying molecular events.

The use of transgenic mice has provided a new understanding of tissue organization by the use of lineage-specific promoters, such as the neurofila- ment gene promoter.

Clinical research was a small but essential part of the program. Clearly identified were two glaring deficiencies in the translation of the new biology into the treatment of patients with brain tumors. First, and of greater urgency, is the critical impor- tance of establishing tissue banks that provide im- mediate access to large numbers of characterized tumors and the retrospective availability of proper specimens for the correlation of genetic markers with observed behavior and therapeutic respon- siveness of human gliomas. The second major defi- ciency is in the present classification of glial tu- mors, the astrocytomas in particular. Classical his- tology is inadequate to distinguish the different pathological states. The classification of gliomas should be at least as complex as that of leukemias and lymphomas, two neoplastic processes that have been enlightened by the specificity of current- ly available markers. Even now, for therapeutic purposes glial tumors can be characterized as MER positive or MER negative with a high probability of predicting the tumor's sensitivity to ethyl nitrosu- reas. Although not yet of known therapeutic pre- dictive value, the markers defined by Webster Ca- venee, lipocortin 2, and the presence or absence of the p53 mutations can be determined on surgically obtained specimens. The p53 mutation is of consid- erable interest because of its presence in younger patients. These genetic differences between ne-

oplastic and non-neoplastic astrocytes must be cor- related in a way that will translate into definable biologic behavior and therapeutic responsiveness.

Currently ongoing and planned clinical trials are variations on past themes with few new modifica- tions. It is unlikely these newer trials will give as much as 25% improvement over current 'best known therapy'. We know predictors of outcome, but until we can design more specific and effective forms of therapy, we will continue to conduct trials looking for small incremental gains in survival. Even so, a continued aggressive approach to treat- ment is mandatory. We must caution nihilistic col- leagues against tearing apart the very structure that, with new therapeutic tools, would allow us to evaluate and implement new forms of treatment that someday will be curative. Treatment of the disease should be inextricably interlocked with the basic research that ultimately will lead to its suc- cessful treatment.

A novel therapeutic approach described in this meeting was accelerated fractionated irradiation in conjunction with the use of a radiopotentiator. In another trial, the introduction of a new drug into a phase III trial without prior demonstrated effec- tiveness is a new approach to evaluating promising chemotherapy. Both innovations represent inno- vative clinical research, and I applaud the investi- gators.

Finally, what are the truly unconventional ap- proaches to therapy of the future? First on the list of potential therapeutic approaches based on lab- oratory investigation is the inhibition of tumor an- giogenesis, without which a tumor's growth into a mass is precluded. Several inhibitors of angiogene- sis have been characterized. The requirement of continuous administration should be no more con- founding than the diabetic's dependence on a con- tinuous supply of exogenous insulin.

The transfection of an immunoglobulin (IgM) gene into a glial cell has demonstrated the amazing efficiency of glioma cells in the production of IgM. The production of antibody by a non-lymphoid cell opens up the possibility of functional inactivation of extracellular gene products as well as selected intracellar proteins. For example, such antibodies

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could be used to block autocrine secretion of growth factors, an intriguing possibility, but one that must wait for further development in animal models.

Genetically modified cells, transfected by sever- al currently available techniques, present another therapeutic possibility. Engrafting of functionally active tissue has recent precedence in the treatment of Parkinsonism, and the potential of grafting ge- netically modified cells into, or adjacent to, brain tumors is not in the realm of science fiction.

Can systems be devised for the delivery of new genes to glioma cells in vivo? We are shown a strategy involving an HSV-1 defective mutant virus for packaging a gene to be expressed specifically in glioma cells. This kind of gene therapy may be possible, but problems of efficiency and specificity seem formidable. Finally, immunotherapy belongs in the category of innovative treatment. One can- not help but be impressed by the effectiveness of intrathecally administered IL-2 in the treatment of tumors that have metastasized to the meningeal space. This and similar approaches should be pur- sued with continued thoughtfulness and scientific scrutiny.

The power of monoclonal antibodies is no longer in doubt - - the question is not if they will be effec- tive, but how can their use be optimized and when better antigens will be identified and exploited. More than likely, the final therapeutic product will be a non-antigenic cocktail with radioactive war- heads attached to glioma-associated antibodies, such as an antibody to a mutated EGF receptor. The prospect of constructing libraries of human antibodies from which ones of therapeutic value might be found is particularly promising in this regard. I for one have become a convert to the potential of treatment using monoclonal antibod- ies.

I came to this meeting with the sober perspective of having attended brain tumor conferences for the past 20 years. The quality of the science in this meeting has been stunning, and I say this quite literally. The number of investigators now working with glial tumor far exceeds the expectation of just a few years ago. This meeting differs from all previ-

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ous meeting because of the quantity of molecular science concerning brain tumors. Phenomenology has been reduced to specific molecules and specific cellular processes. For someone like me, this rapid acquisition of new information is difficult to be- lieve.

I am exhilarated by the reality of truly new ther- apy based on specific molecular mechanisms. Without doubt new forms of molecular therapy will be entering clinical trials within a decade. We must be prepared to change our thinking from curing, i.e., destroying tumors, to modulating and render- ing genetically impotent the tumors that are the object of our investigations. Peaceful coexistence is preferable to a war that cannot be won.

I want to applaud three persons. Lorraine Marin had a key role in selecting the speakers and their topics, Shannon Leeper-Tatum took care of us in her characteristically efficient and charming man- ner and Peter Preuss made the meeting possible. Peter conceived the idea of bringing together a diverse group of scientists because of his single minded dedication to finding a cure for brain tu- mors and the insight to know that this will come through fundamental research and the interaction of serious investigators from a number of unrelated disciplines. This meeting is testimony to his vision, determination, energy and generosity. It is the best meeting that I have ever attended and, for all of us, I thank Peter Preuss.