Brain Tumors: An Overview of Current Histopathologic and Genetic Classifications

Diffuse Astrocytomas

Diffuse astrocytoma (WHO grade II) is a mildly hypercellular infiltrative neoplasm composed of well-differentiated, spindle- to stellate-shaped astrocytes with minimal nuclear atypia and separated by a loosely fibrillary (and sometimes microcystic) background. Nuclei are characteristically irregular, angulated, and hyperchromatic. Mitotic figures are either absent or rare, and there is no vascular proliferation or necrosis. Glial fibrillary acidic protein (GFAP) may be highly or weakly expressed. Two morphologic subtypes are recognized: protoplasmic and gemistocytic. Protoplasmic astrocytomas are composed of small tumor cells with scant GFAP expression and are frequently associated with a myxoid or microcystic background. Gemistocytic astrocytomas consist of a predominant population of large rounded neoplastic astrocytes with a copious amount of glassy eosinophilic cytoplasm that is strongly GFAP positive and has an eccentrically located nucleus. Despite their benign morphologic appearance, diffuse astrocytomas have an intrinsic tendency to recur, spread extensively, and undergo anaplastic progression to a higher grade. The time to recurrence and progression after initial clinical evaluation varies from case to case but ranges from months to several years.

Anaplastic astrocytoma (WHO grade III) is a hypercellular glioma composed of poorly differentiated astrocytes consisting of a mixture of pleomorphic fibrillary and gemistocytic cells with significant nuclear atypia and prominent mitotic activity. These malignant tumors may develop as a result of anaplastic progression from a preexisting, low-grade diffuse astrocytoma or may arise de novo. The mean age of patients with anaplastic astrocytoma at initial diagnosis is approximately 41 years, which falls between the age means for patients with low-grade diffuse astrocytoma and glioblastoma multiforme (GBM).

GBM (WHO grade IV) is unfortunately both the most common glioma and the most malignant primary brain tumor arising in adults. Most commonly GBMs are solitary tumors of the cerebral hemispheres, but they may develop at almost any site within the neuraxis, including the cerebellum and the spinal cord. In many cases they infiltrate across the corpus callosum or arise directly within it, with bilateral extension (butterfly tumor). Multifocal tumors are observed in about 2% of patients and are often mistaken for metastatic disease on preoperative neuroimaging studies. The necrotic tumor mass may be partially delineated on gross examination, but infiltrating glioma cells can easily be identified microscopically well beyond the apparent gross tumor boundaries. The cellular morphology of GBM cells is highly variable, with the spectrum ranging from small, tightly packed, round or elongated cells to giant bizarre and multinucleated forms, all of which are frequently encountered within a single tumor. Mitotic figures are typically readily identified, and corresponding proliferation marker indices, such as the Ki-67 antigen, show elevated levels. Positive immunostaining for GFAP is characteristic but highly variable, and can be absent in some instances. Microvascular proliferation and foci of necrosis are histologic hallmarks of GBM ( Fig. 133.1 ). The presence of necrosis is not required for a diagnosis of GBM; vascular proliferation, in conjunction with pleomorphism and increased mitotic activity, is sufficient according to WHO 2016 criteria.

The typical peripheral, ring-like zone of contrast enhancement seen on computed tomography and magnetic resonance imaging (MRI) corresponds to the highly vascularized peripheral area of the neoplasm. Vascular proliferation is defined as the presence of blood vessels with multilayered vessel walls (more than two cell layers thick). So-called glomeruloid vascular proliferation constitutes the most obvious and characteristic example of the florid microvascular proliferation seen in GBM. Large areas of necrosis are typical of GBMs, which also frequently exhibit multiple small serpiginous zones of necrosis surrounded by densely crowded tumor cells in a pseudopalisading pattern. The hypoxic perinecrotic apoptotic cells strongly express vascular endothelial growth factor, which in turn induces the characteristically prominent vascular proliferation.

Two subsets of GBM, primary and secondary, have been described based on clinical and genetic data. Primary GBM arises de novo (i.e., without evidence of a preexisting lower grade astrocytoma), typically in patients in the sixth decade of life and beyond. In contrast, secondary GBM tends to develop in younger adults (<45 years old) by malignant progression from a diffuse astrocytoma of WHO grade II or III ( Table 133.1 ).

Although the histologic grading criteria to distinguish among diffuse astrocytoma WHO grade II, anaplastic astrocytoma WHO grade III, and glioblastoma WHO grade IV are still utilized in the WHO 2016 classification system, the increased understanding of the molecular alterations that drive astrocytomas challenges the prognostic value of grading on histologic grounds alone. Mutations in the isocitrate dehydrogenase 1 and 2 genes ( IDH1 and IDH2 ) have been recognized as key genetic alterations in diffuse and anaplastic astrocytomas. The presence or absence of IDH1/DIH2 mutations in an astrocytoma is a more powerful predictor of tumor behavior than grading based on histologic features alone. IDH1 or IDH2 mutations in astrocytomas frequently coexist with mutations in TP53 and ATRX . Therefore, the molecular signature of low-grade astrocytomas, which have better prognosis, is IDH1 or IDH2 mutant, TP53 mutant, and ATRX mutant ( Fig. 133.2 ). In contrast, IDH-wildtype astrocytomas have worse prognosis and frequently have mutations in the promoter of the TERT gene, EGFR amplification, and PTEN mutations. IDH-mutant astrocytomas occur in younger patients and typically present as grade II or III astrocytomas, with later progression to GBM. In contrast, IDH-wildtype astrocytomas present later in life, and frequently as grade IV tumors. Therefore, in molecular terms, IDH-wildtype GBM is the equivalent of primary GBM and IDH-mutant GBM is the equivalent of secondary GBM. Although rarely, IDH-mutant GBM will present de novo. Under the WHO 2016 classification system, diffuse astrocytoma, anaplastic astrocytoma, and GBM must be further classified as IDH-mutant or IDH-wildtype. The most common mutation in IDH-mutant astrocytomas is IDH1 p.R132H, and it is present in approximately 95% of IDH-mutant tumors. There is an antibody that recognizes the IDH1 p.R132H mutant protein (see Fig. 133.2A ) and is routinely used in clinical practice. In addition, an antibody that recognizes the ATRX protein is frequently used to aid in the diagnosis of lower grade astrocytomas. The majority of astrocytomas with an ATRX mutation will demonstrate loss of protein expression that can easily be demonstrated with immunohistochemistry (see Fig. 133.2B ). Large-scale genomic studies have shown that ATRX mutations and 1p/19q co-deletion are mutually exclusive events. Therefore the presence of an ATRX mutation or loss of ATRX expression, demonstrated by immunohistochemistry in a diffuse glioma, essentially excludes the diagnosis of oligodendroglioma.

Molecular alterations of GBM include TERT promoter mutations, combined whole chromosome 7 gain and whole chromosome 10 loss (+7/−10), and EGFR amplification, among others. Given the evidence that molecular alterations correlate better with patient prognosis than histologic grading, it is possible to make a diagnosis of “diffuse astrocytoma with molecular alterations of glioblastoma” in a patient with an infiltrating astrocytoma without the presence of vascular proliferation or necrosis, if the critical genetic alterations ( TERT promoter mutation, EGFR amplification, or +7/−10) can be demonstrated.

GBM is highly resistant to therapy, largely because of its invasive properties, which preclude surgical cure; the presence of a population of neural stem cell–like tumor cells that may harbor resistance mechanisms distinct from those of most nonneural stem cell–like tumor cells; and retention of abundant DNA repair mechanisms that abrogate the effectiveness of chemotherapy and radiotherapy. In 2005, a randomized phase 3 trial of temozolomide and radiation therapy in patients with newly diagnosed GBM determined that patients whose tumors exhibited methylation of the MGMT gene promoter survived longer after treatment than did those without evidence of promoter methylation. MGMT is a ubiquitous cellular enzyme largely responsible for resistance to the cytotoxic action of DNA alkylating agents such as temozolomide, carmustine, lomustine, and procarbazine. Inactivation of MGMT impairs the ability of the cell to repair DNA damage. To date, MGMT promoter methylation status is the single most important prognostic marker for predicting benefit from chemotherapy with alkylating agents.

Gliomatosis cerebri has historically been considered to be a unique glial tumor entity, but mounting evidence from molecular studies supports that gliomatosis cerebri is a pattern of exceptionally diffuse involvement of the central nervous system (CNS) that can be seen with any of the diffuse glioma subtypes, including oligodendroglioma.

The “diffuse midline glioma, H3 K27M-mutant” (DMG-K27M) is a newly recognized entity defined in the WHO 2016 classification system as a highly malignant infiltrative midline glioma with a K27M mutation in either H3F3A or HIST1H3B/C genes. The H3 K27M mutation is found in the majority of diffuse intrinsic pontine gliomas and nearly half of diffuse gliomas arising at any other midline location of the CNS (mostly the thalamus, brainstem, and spinal cord). DMG-K27M tumors are most common in children, but they can occur at any age. These tumors may present a variety of imaging findings as well as histologic appearances at biopsy, including bland hypocellular tumors with features of low-grade gliomas. Although K27M mutations have also been described in the context of other histologies such as pilocytic astrocytoma, ganglioglioma, and ependymoma and in tumors away from the midline, their highly aggressive behavior is closely associated with DMGs. Therefore only the midline infiltrative gliomas bearing the K27M mutation qualify for the DMG-K27M diagnosis. The mutation can be readily demonstrated by immunohistochemistry with antibodies targeting the mutant protein ( Fig. 133.3 ). Another mutation in the H3 histone family, the H3F3A G34R/V mutation, has been recently described in hemispheric high-grade gliomas in children and young adults.

Circumscribed Astrocytomas

Pilocytic astrocytoma (WHO grade I) typically arises in children and young adults. Pilocytic astrocytomas are macroscopically circumscribed and grow very slowly. In contrast to diffuse astrocytomas, pilocytic astrocytomas exhibit very little tendency for anaplastic progression. Pilocytic astrocytomas preferentially affect the cerebellum, brainstem, optic nerves, and third ventricular region. The tumor often manifests as a mural nodule within a cyst. Invasion of the overlying leptomeninges is common, but this feature does not constitute a negative prognostic factor. Microscopically, many pilocytic astrocytomas exhibit a biphasic architectural pattern consisting of compacted areas of elongated, piloid (hair-like) cells alternating with loosely textured and microcystic areas populated by scattered stellate cells. The neoplastic pilocytes of pilocytic astrocytoma are GFAP positive. Rosenthal fibers, which are brightly eosinophilic compacted masses of glial intermediate filaments with entrapped cytosolic proteins and eosinophilic granular bodies (EGBs), are histologic hallmarks of pilocytic astrocytoma, although they are neither a constant nor a pathognomonic feature. Hyalinized and glomeruloid vessels are frequently observed. Vascular proliferation is responsible for the contrast enhancement seen on preoperative MRI studies and can occasionally cause histologic diagnostic confusion with a high-grade glioma (e.g., GBM), particularly in small biopsy samples. In pilocytic astrocytomas, mitotic activity, vascular proliferation, and necrotic foci do not have the same ominous prognostic significance as when present in diffuse astrocytomas. In favorable anatomic locations, such as the cerebellum, surgical resection of pilocytic astrocytoma has the potential to be curative.

The majority of pilocytic astrocytomas, in particular those occurring in the cerebellum, show a characteristic tandem duplication in chromosome 7 (7q34) that results in a fusion between the BRAF gene and the poorly characterized gene KIAA1549. ^, The fusion results in a constitutively active BRAF kinase domain. Other genetic alterations in pilocytic astrocytomas include point mutations in BRAF (e.g., BRAF p.V600E), NRAS, and KRAS. This has led to the understanding that pilocytic astrocytomas occur as a result of dysregulation of the mitogen-activated protein kinase (MAPK) pathway.

Pilomyxoid astrocytoma is a histologic variant of pilocytic astrocytoma that has been found to be clinically more aggressive than ordinary pilocytic astrocytoma. It occurs most commonly in the hypothalamic or chiasmatic region. Distinctive histologic features include a monomorphic population of neoplastic pilocytes in a prominent myxoid background stroma. Unlike the situation with most pilocytic astrocytomas, in pilomyxoid astrocytoma, Rosenthal fibers and EGBs are either very inconspicuous or completely absent, although tumors with overlapping features or maturation into classic pilocytic astrocytoma over time have been reported. Because of the more aggressive clinical course, pilomyxoid astrocytomas were previously designated as WHO grade II. However, it is now believed that the more aggressive clinical course results from difficulties in obtaining complete surgical resections in the hypothalamic/chiasmatic region, and the grading of these tumors is now uncertain, although they remain in the low-grade category (I versus II).

Pleomorphic xanthoastrocytoma (PXA; WHO grade II) is another circumscribed variant of astrocytoma that typically arises in a superficial cerebral location in children and young adults with a history of seizures. The temporal lobe is a favored anatomic site. Histologically, PXA mimics GBM in its strikingly pleomorphic tumor cells, but the paucity of mitotic figures and absence of vascular proliferation and necrosis are clues to the correct diagnosis, and the characteristic presence of EGBs, which are also seen in other circumscribed tumors (such as pilocytic astrocytoma and ganglioglioma), is an even stronger signature of PXA. The pleomorphic, giant, and often multinucleated cells may display a variable xanthomatous change in their cytoplasm because of intracellular accumulation of lipids. Most of these pleomorphic cells are GFAP positive, but neuronal marker proteins can also be detected in some tumors, indicative of dual differentiation. In addition to prominent EGBs, dense intercellular and pericellular reticulin and perivascular lymphocytic infiltrates are also typical microscopic features of PXA. As with pilocytic astrocytoma, invasion of the overlying meninges is characteristic and does not constitute a negative prognostic factor. PXAs generally exhibit indolent behavior; however, approximately 15% will recur and undergo anaplastic progression to high-grade diffuse astrocytoma. The majority of PXAs have a characteristic mutation in the BRAF gene (BRAF p.V600E). Recently, the most frequent molecular alterations in anaplastic PXAs have been described as BRAF p.V600E, CDKN2A deletion, and TERT promoter mutations. BRAF p.V600E is not specific to PXAs; it can be present in pilocytic astrocytomas, gangliogliomas, and GBMs. Specifically, the BRAF p.V600E mutation can be seen in approximately 50% of GBMs with epithelioid morphology (i.e., epithelioid GBM subtype).

Subependymal giant cell astrocytoma (WHO grade I) is an intraventricular tumor composed of large cells that resemble gemistocytes or ganglion cells, or both, morphologically and immunohistochemically. This tumor is almost invariably associated with tuberous sclerosis, although this condition is often not known at initial evaluation. Surgical resection is curative.

Desmoplastic infantile astrocytoma (WHO grade I) is a rare tumor of infancy with morphologic and clinical features that overlap those of another rare tumor termed desmoplastic infantile ganglioglioma. Both lesions share a superficial cerebral cortical location, large size, circumscribed growth pattern, and development during infancy. A marked desmoplastic meningeal reaction is characteristic. Surgical resection is the treatment of choice, and the prognosis is generally more favorable than would otherwise be suggested by the typically very large size of most of these tumors at diagnosis.