Section 1 -

Clinical, Radiological, and Pathological Consequences of Glioma Growth Patterns Pieter Wesseling Radboud University Nijmegen Medical Centre Nijmegen , The Netherlands

Glioma Growth Patterns The group of gliomas is heterogeneous and encompasses many different histological types and malignancy grades. Based on the resemblance of the tumor cells with non-neoplastic glial cells and the presence of particular tissue elements (e.g. formation of (pseudo)rosettes), most gliomas can be typed as astrocytic, oligodendroglial, mixed oligo-astrocytic, or ependymal tumors [19]. Of these, the diffuse infiltrative astrocytic tumors are by far the most common, esp. its most malignant form i.e. glioblastoma multiforme (GBM). While meningiomas generally show compact, expansive growth, and metastatic tumors primarily show a combination of compact growth and perivascular infiltration in the brain parenchyma, most gliomas in adult patients are characterized by extensive, diffuse infiltration in the neuropil [19] . Other gliomas, however, such as ependymomas and pilocytic astrocytomas, often have a more circumscribed growth pattern. One of the pioneers in the study of glioma growth patterns was Hans-Joachim Scherer [23]. Scherer systematically described different patterns of arrangements of glioma cells that he considered to be dependent on preexisting tissue elements. Examples of such structures, designated by Scherer as "secondary structures", are perineuronal growth, surface growth, perivascular growth, and peri/intrafascicular growth. Arrangement of glioma cells that does not seem to depend on preexisting tissue but can be considered as an expression of the intrinsic architectural potential of the tumor (e.g. canalicular structures, papillary formations, whorls) were called "proper structures", while "tertiary structures" were defined by Scherer as formations brought about by the interaction of glioma cells with the proliferating mesenchymal tissue of the tumor [27] . Indeed, in diffuse gliomas the cells often seem to preferentially invade along myelinated fibers in the white matter tracts (intrafascicular growth). Furthermore, subpial and perivascular growth and perineuronal accumulation of tumor cells (satellitosis) is frequently encountered in these tumors [15] . The diffuse infiltrative growth of glioma cells in the neuropil (which has been called "stealth invasion of the brain" by some authors [32] ) is unique and suggests specific interactions between these tumor cells and their microenvironment. The exact nature of these interactions is still largely unknown [12, 25] . Apart from direct penetration of the brain tissue, gliomas may also spread via the cerebrospinal fluid pathways. Such (subarachnoid or intraventricular) dissemination may well be increasing as patients with gliomas live longer [4]. In gliomas of high grade malignancy, prominent "endothelial"/"microvascular" proliferation and necrosis emerge [31] . These changes, which are in fact used as histopathological criteria to diagnose high-grade malignancy in these tumors [19], are often spatially and temporally related. Furthermore, around areas of necrosis pseudopalissading of tumor cell nuclei is frequently present. The tumor cells in those pseudopalisading zones were shown to be less proliferative and have a higher apoptosis rate than the tumor cells more distant from necrotic areas. The perinecrotic cells also show increased expression of vascular endothelial growth factor (VEGF) and hypoxia inducible factor 1a (HIF-1a ), two factors that play a crucial role in the induction of angiogenesis [5, 6, 13] . The accumulation of tumor cells in the pseudopalisading zone may very well be explained by migration of the tumor cells away from the necrotic area [6] . It has been hypothesized that in this way necrosis may select for tumor cells that are more aggressive and more resistant to different therapeutic modalities [24] . Even though gliomas may show an extreme angiogenic response, extraneural metastases of these tumors are extremely rare. Clinical Consequences Patients with diffuse low grade gliomas may survive for multiple years, but sooner or later these tumors often progress to high grade malignancies. Other gliomas present as high-grade malignant lesions from the start. Partly because of the diffuse infiltrative growth pattern, curative treatment for these gliomas is generally impossible. Eradicating glioma cells by radiotherapy without damaging the in filtrated brain parenchyma has been difficult to achieve. Furthermore, whereas surgery of most other tumors aims at complete resection of the tumor with a margin of normal tissue, the infiltrative growth of diffuse gliomas in the brain parenchyma almost always precludes complete tumor removal. There is much debate on the optimal timing and extent of surgery, as well as on the role of radiotherapy and chemotherapy for patients with low grade diffuse gliomas [22]. Several studies revealed that maximum removal of contrast-enhancing, high grade glioma tissue is an independent prognostic factor for overall survival [26] . The surgeon's intraoperative impression is , however, a notoriously inaccurate method for assessing the extent of resection [1]. A recent study showed that fluorescence guided resection (using 5-amino-levulinic acid, a non-fluorescent pro-drug that leads to intracellular accumulation of fluorescent porphorins in malignant glioma cells) allows for more radical resection of the contrast-enhancing part of malignant gliomas, leading to a significantly improved progression free survival at 6 months [28] . Patients with malignant oligodendrogliomas, esp. when the tumors shows loss of the short arm of chromosome 1 (-1p), often show response to chemotherapy [17]. A recent study showed that patients with GBM have some benefit from radiotherapy combined with temozolomide treatment [29]. However, these patients are far from being cured, and there is an urgent need for other therapeutic approaches [26]. The blood-brain barrier (BBB) may form an obstacle for optimal delivery of chemotherapeutics to diffuse infiltrative gliomas, especially in areas in which the original tissue architecture is relatively preserved. Furthermore, the marked intratumoral heterogeneity of gliomas may hamper the success of chemotherapy [26]. Because of the florid, sometimes glomeruloid microvascular changes in high grade gliomas, these tumors have since long been considered as good candidates for anti-angiogenic therapy [14] . However, in diffuse infiltrative gliomas many intratumoral vessels may be incorporated rather than newly formed. Indeed, in the most extreme example of diffuse infiltrative neoplastic growth in the brain, i.e. gliomatosis cerebri, quantitative analysis of the brain microvasculature suggested a complete lack of angiogenesis [3]. Moreover, the contribution of the aberrant, e.g. glomeruloid microvessels to the perfusion of the tumor tissue is unclear [30]. In experimental glioblastoma models the blockade of vascular changes by anti-VEGF therapy resulted in increased vessel coöption by the tumor [20, 21] and the diffuse infiltrative part of the tumor was relatively unaffected [11] . Radiological Consequences Magnetic resonance imaging (MRI) is now the gold standard for defining brain tumor anatomy in a clinical setting [10]. One important feature in the MR diagnosis of gliomas is the presence of contrast-enhancement in T1-weighted MR images using Gadolineum-DTPA, this phenomenon indicates increased vascular permeability. Pilocytic astrocytomas are radiologically generally relatively discrete, contrast-enhancing tumors with or without cyst formation, in this glioma category contrast-enhancement does not imply malignant behavior [19]. In diffuse infiltrative gliomas, on the other hand, contrast enhancement generally means a more malignant biological behavior. Low grade diffuse gliomas are non-enhancing, hypointense lesions on T1-weighted MR images (and hyperintense on T2 images). The presence of vessel coöption, the lack of neovascularization, and the apparently limited changes to the preexistent, incorporated vessels in the latter category explain the absence of contrast-enhancement in these tumors. The central area in "ring-enhancing" high-grade diffuse gliomas represents necrosis, while the enhancing rim contains vital glioma tissue with vascular changes and increased vascular permeability. The periphery of the diffuse infiltrative gliomas is, however, difficult to delineate by radiological techniques. Correlation of whole brain histological sections of high grade gliomas with CT scans revealed that tumor cells were present even outside the peritumoral areas of low density [8]. In the near future new MR modalities (e.g. dynamic and diffusion MRI, MR spectroscopy) may contribute to better radiological classification and delineation of brain tumors. Furthermore, in situations where more extensive resection of the tumor tissue is not an option, such modalities may assist in assessing the best spot for a biopsy [10]. Pathological Consequences Up till now, the histopathological diagnosis is the gold standard for the classification of brain tumors. Recognition of the diffuse infiltrative versus other categories of glial tumors has important prognostic and therapeutic implications. While the exact growth pattern of gliomas can not always be assessed in biopsy specimens, some of the histopathological features mentioned above (esp. intrafascicular growth, perineuronal satelitosis, subpial accumulation of tumor cells) strongly favor a diffuse infiltrative nature of the glial neoplasm. In contrast, other features (esp. Rosenthal fibers, eosinophilic granular bodies, biphasic pattern with compacted and loose textured areas) favor a diagnosis of another variant of glioma (esp. pilocytic astrocytoma) [19]. Both diffuse infiltrative and pilocytic astrocytomas can show growth in the subarachnoid/leptomeningeal compartment, and in itself this this phenomenon does not imply malignant progression. It is also important to realize that the presence of florid endothelial/microvascular proliferation in diffuse infiltrative gliomas indicates high-grade malignancy, while in pilocytic astrocytomas this feature is fully consistent with a WHO grade I tumor [19]. Diffuse infiltrative gliomas are often heterogeneous with marked spatial differences in cellular phenotype and malignancy grade. A post mortem study investigating whole brain sections showed that multifocal glioblastomas can emerge in the background of a better differentiated astrocytic neoplasm [9] . Since molecular genetic studies demonstrated a common origin in different components of diffuse gliomas, these heterogeneous gliomas are considered as clonal lesions [2, 19] . The heterogeneity does have implications for the accuracy of the histopathological diagnosis. A diagnosis based on a needle biopsy material should be interpreted with caution, preferably in a multidisciplinary setting with all relevant clinical and radiological information available. Obviously, histologic assessement of the nature of the glioma is likely to be more accurate when large amounts of tissue are provided to the (neuro)pathologist. In a recent study of 81 consecutive patients treated in a single institution, it was shown that even with expert neuropathological review there was a discrepancy in 38% of the cases between diagnoses made from small biopsy specimens and those made from resected specimens, affecting the therapeutic regimen in 26% and the prognostication in 38% of the cases [16] . 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