A brain tumor is defined as an abnormally growing cluster of cells within the brain. Glioblastoma is a tumor in the malignant group that can develop in the brain and spinal cord, consisting of glia that supports nerve cells. Glioblastoma is mostly sporadic, except in rare cases, Turcot syndrome or Li-Fraumeni syndrome. According to the 6th version classification made by the World Health Organization (WHO) in 2021, glioblastomas are Grade IV diffuse astrocytic tumors with retained isocitrate dehydrogenase (IDH) wild-type and nuclear ATRX.
According to the old classification, glioblastoma was divided into primary and secondary according to its development. Primary glioblastoma arises directly (de novo) from glial precursor cells and is seen in the older population. It usually gives clinical symptoms in the first three months.[3,4] Glioblastoma is the most common primary brain malignant tumor in adults.[5,6] It characteristically has an amplification as well as overexpression of the epidermal growth factor receptor (EGFR) and ligand. Secondary glioblastoma tumors are formed by the conversion of pre-existing lower grade astrocytomas to the more malignant grade of anaplasia. Although patients with secondary glioblastoma can be considered younger, their average age is 45 years.
NEW MOLECULAR CLASSIFICATION OF GLIOBLASTOMA
After the WHO 2021 classification of tumors of the central nervous system (CNS) and cIMPACT-NOW (the Consortium to Inform Molecular and Practical Approaches to CNS Tumor Taxonomy), the definition of glioblastoma has changed. It now integrates into the glioma classification, histological tumor typing, and grading, as well as analyzes of molecular markers.
Tissue for histopathological diagnosis is first examined for isocitrate dehydrogenase 1 (IDH1) mutation and loss of nuclear ATRX. In patients >55 years of age without a previously known low-grade glioma, with a tumor located outside the midline, and with preserved nuclear ATRX expression, the presence of immunohistochemical negativity for IDH1 R132H is sufficient for classification as IDH wild-type. Patients with the histone H3.3 G34R/V mutation with IDH wild-type but the loss of nuclear ATRX are defined as Grade IV diffuse hemispheric glioma. To identify diffuse midline gliomas, it should be evaluated for histone H3 K27M mutations and loss of nuclear K27-trimethylated histone H3 (H3K27me3). IDH wild-type diffuse astrocytic gliomas without microvascular proliferation or necrosis should be tested for +7/-10 cytogenetic signature as EGFR amplification, TERT (telomerase reverse transcriptase) promoter mutation, and molecular features of IDH wild-type glioblastomas.[2,10]
Isocitrate dehydrogenase wild-type is essential for the diagnosis of glioblastoma. Although their histological appearance is similar in IDH-mutant and wild-type gliomas, they are no longer used to refer to as IDH-mutant astrocytic gliomas because their prognosis and biological features are different. IDH-mutant astrocytomas are now divided into three WHO grades: astrocytoma, IDH-mutant, WHO Grade 2; astrocytoma, IDH-mutant, WHO Grade 3 (instead of anaplastic astrocytoma, IDH-mutant, WHO Grade 3); and astrocytoma, IDH-mutant, WHO Grade 4 (old term glioblastoma, IDH-mutant.[9,10]
Studies show that many genetic factors play a role in the development of glioblastoma. One of these genes is the IDH gene, which encodes the isocitrate dehydrogenase enzyme that plays a role in energy production and participates in the Krebs cycle. The IDH enzyme family consists of three distinct proteins found in the cytoplasm and peroxisomes (IDH1) and mitochondria (IDH2 and IDH3), involved in many cellular processes such as mitochondrial oxidative phosphorylation, glutamine metabolism, lipogenesis, glucose sensing, and regulation of cellular redox state.[12-14] IDH3 is a heterotetrameric complex enzyme that catalyzes the conversion of isocitrate to alpha-ketoglutarate (α-KG) in the tricarboxylic acid cycle in a nicotinamide adenine dinucleotide (NAD+) dependent manner. IDH1 and IDH2 are very similar homodimer enzymes that catalyze nicotinamide adenine dinucleotide phosphate (NADP+) dependent oxidative decarboxylation to α-KG. It is also the main producer in the brain of NADPH, an important cellular reducing agent that plays a role in protection against reactive oxygen deoxyribonucleic acid (DNA) damage, which is required for detoxification through the reduction of IDH1, glutathione, and thioxins and activation of catalase.
Somatic point mutations in IDH1/2 result in increased secretion and accumulation of D-2-hydroxyglutarate (D-2HG), an oncometabolite. Overproduction of D-2HG inhibits α-KG-dependent dioxygenases, including histone and DNA methyls, leading to histone and DNA hypermethylation. As a result, it enables tumor cells to maintain their biosynthetic precursor pools and suppress mitochondrial reactive oxygen species (ROS). Suppression of reactive species, even if oxygen metabolism is disrupted, blocking of DNA repair mechanism contributes to sustaining rapid proliferation rates, accelerating new vessel formation, thus contributing to oncogenesis.[11,16-18]
Isocitrate dehydrogenase wild-type glioblastoma is much more aggressive than IDH-mutant astrocytomas. In addition, IDH1 mutation has been shown to be strongly associated with survival. It has been confirmed in many studies that the survival of people with this mutation is longer.[19,21-22]
The results obtained in the studies show that the inhibition of mutant IDH1/2 enzymes reverses the differentiation by decreasing the D-2HG level in the cell. Therefore, mutant IDH1/2 enzyme inhibitors represent a good group of drugs used in the treatment of patients with IDH/2 mutations.
O6-METHYLGUANINE-DNA METHYLTRANSFERASE (MGMT)
Another gene is the MGMT gene, which encodes the “O6-methylguanine DNA methyltransferase” (MGMT) enzyme consisting of 207 amino acids. O6-Methylguanine DNA methyltransferase is the most important of the DNA repair proteins that protect DNA from the mutagenic effects of alkylating agents. Therefore, in the presence of MGMT in the cell, the toxicity of alkylating agents is reduced and they cannot manifest their anticancer properties. This protein has a mechanism that creates resistance to drugs such as alkylating nitrosourea and temozolomide (TMZ) with its activity. In this mechanism, while alkyl groups from the guanine O6 location, which is one of the DNA alkylation sites, will be induced by chemotherapeutic agents, resistance to chemotherapeutic agents occurs due to their removal by MGMT. It was determined that tumor cells with high MGMT enzyme levels were resistant to TMZ, and cells with low enzyme levels had high TMZ sensitivity. Epigenetic silencing of MGMT by MGMT methylation will result in MGMT inactivation and reduced DNA repair activity. Various studies have reported better responses to radiation therapy (RT) and chemotherapy (ChT) as a result of methylation.[19,22,27,28]
In addition, mutations in the gene of another molecule, TERT promoter, are associated with worse survival independent of other clinical and molecular factors.[29,30] The presence of TERT mutation in IDH wild-type astrocytomas alone is diagnostic for glioblastoma.
The majority of glioblastoma patients are 65 years or older at the time of diagnosis. Among the clinical factors affecting the prognosis of the disease, it has been seen in many studies that advanced age and poor performance status are associated with shorter survival. The average survival of patients 65 years and older is approximately six months.[32,33] IDH mutations are becoming increasingly rare with age. In addition, the most important prognostic factor in patients over 70 years of age is that methylated MGMT offers better survival than unmethylated MGMT.
At diagnosis, patients with glioblastoma include new-onset epilepsy, neurocognitive disorders, focal deficits, and signs and symptoms of increased intracranial pressure. Headache is among the most prominent complaints, but the pain is intermittent, more pronounced in the morning, moderate and very severe unilateral throbbing pain. This pain usually does not respond to medical treatment. Although seizure is usually the first sign in slow-growing superficial tumors, it can also be seen in high-grade tumors such as glioblastoma. Nausea and vomiting also occur following increased intracranial pressure. As a result of neuronal damage that the tumor may cause in the brain tissue, motor and sensory disorders may develop in the innervated region.
The anamnesis taken during the physical examination of the patient is very important in establishing the diagnosis. Considerable attention should be paid to the neurological symptoms here. Hereditary diseases such as neurofibromatosis, Li-Fraumeni, Von Hippel-Lindau, Turcot syndrome should be questioned. Additionally, the high number of deep vein thrombosis and pulmonary thromboembolism cases of malignant gliomas should not be ignored, and the findings related to these conditions should also be paid attention.
The contrast-enhanced magnetic resonance imaging (MRI) technique is used in the diagnosis of glioblastoma. In contrast-enhanced MRI, the gadolinium used as a contrast agent carries minimal allergy risk. Gadolinium allows visualizing tumor formations by enhancing the contrast in areas where the blood-brain barrier is disrupted. A pre-contrast, T1 (longitudinal relaxation) weighted image is taken to observe the anatomy of the brain. In addition, fluid-attenuation inversion recovery (FLAIR) or T2 (transverse relaxation) weighted images are required to detect edema, parenchyma, or corpus callosum invasion.
Peripheral contrast enhancement with a central necrotic cavity in the middle, in the form of a ring with irregular borders, and the appearance of edematous lesions on T2 images around it are typical for glioblastoma.[35,36] 95% of the cases can be diagnosed with imaging techniques.
Perfusion MRI and amino acid positron emission tomography (PET) can be helpful in tumor prediagnosis by identifying metabolic hotspots. It can be used in cases where tumor resection is difficult. Histopathological diagnosis is necessary to determine the prognosis and treatment of the disease. The tissue required for histopathological diagnosis is taken by surgical resection or stereotactic biopsy.
Glioblastoma, which has intense mitosis, vascular proliferation, and necrosis features in its histopathology, has infiltrated the surrounding tissues extensively and is usually located supratentorial. Metastasis outside of the CNS is not a common condition. Although it is not a common situation, if metastasis is detected, this situation has occurred by hematogenous route or by direct spread of the tumor.
In glioblastoma treatment, following the widest possible surgical resection, simultaneous chemo-RT followed by adjuvant ChT is the main treatment. In surgical treatment, safe maximal resection is aimed without damaging the normal tissues as much as possible. The maximum size of the tumor that can be resected may vary depending on its localization. It has been reported that as the percentage of the resected part of the tumor increases, survival is positively affected.
Due to the invasive nature of glioblastoma, the tumor cannot be completely removed in most surgeries. Even the microscopic size of the tumor that cannot be removed after surgery can cause recurrence. For this reason, RT and ChT constitute the basic treatment after surgery.[41,42]
In cases where the localization of the tumor is risky for the surgery, a stereotactic biopsy can be beneficial. Could be used when the lesion is not suitable for resection, the tumor tissue cannot be removed in a meaningful way, or the general clinical condition of the patient is not suitable for surgery. In cases such as pons glioma, where biopsy is also life-threatening, radiological diagnosis is sufficient for treatment decision.
Under normal conditions and in glioblastoma that is not critically localized, it is sufficient to use a microscope during surgery. However, it can be difficult to distinguish tumors from a normal brain during surgery with conventional light microscopy. In cases where glioblastoma is localized in the speech center, in the control pathways of various limbs, or in the center of understanding; the use of neuronavigation, ultrasound, cortical stimulation, or fluorescein becomes mandatory. Tissue fluorescence after oral administration of 5-ALA (5-aminolevulinic acid) has high sensitivity, specificity, and positive predictive values for identifying malignant glioma tumor tissue. Thus, it helps to reduce postoperative residual tumor volumes while protecting the risk of new tumors.
Unlike patients with meningioma, for whom awake surgery is recommended on the grounds that this high technology is not sufficient in some cases, this opportunity is not available for patients with glioblastoma. The reason for this is that the consciousness will not be clear during surgery because glioblastoma has made a lot of edema in the brain tissue.
Within 24-48 hours after the surgery, the extent of the surgery should be evaluated with MRI (or computed tomography [CT] if MRI is not possible), with and without contrast; it should also include diffusion-weighted arrays for the assessment of perioperative ischemia.
After surgery, in patients with tumors that cannot be completely resected, the risk of progression within one year is reduced by 37%, and the risk of death by 45%. It was determined that the average survival rate was 6.6 months in patients who underwent biopsy only, 10.4 months in partial resection, and 11.3 months incomplete resection.
The purpose of adjuvant RT is to improve local control without inducing neurotoxicity and after surgery is to minimize the postoperative recurrence due to macroscopic or microscopic residues. However, in cases where surgical resection cannot be performed, post-biopsy radiotherapy is the only treatment.
In glioblastoma, 50-60 Gy RT at a fraction dose of 1.8-2 Gy/day is applied for six weeks. During the treatment, fractions are given on weekdays, and the patient is rested at the weekend to help restore normal cells. No randomized data support that doses >60 Gy improve survival. Hypofractionated radiotherapy with a higher dose per fraction and the lower total dose is appropriate in elderly patients and patients with poor performance status.
Surgical bed area plus residual tumor area defined on T1-weighted, T2-weighted, and FLAIR MRI sequences is defined as gross tumor volume (GTV). For microscopic invasion, the clinical target volume (CTV) is established, which is usually modified to include edema on T2-weighted MRI and subtracted from anatomically normal tissues. GTV is given a margin of 1-2 cm. Finally, a margin planning target volume (PTV) of 0.3-0.5 cm is established for motion or uncertainties during treatment.
O6-Methylguanine DNA methyltransferase methylation in glioblastoma leads to DNA repair protein gene silence and loss of expression; it has been observed that this increases the benefit of ChT and has a positive effect on survival. In glioblastoma patients under 70 years of age with MGMT methylation; adjuvant TMZ is recommended in the continuation of a treatment consisting of a combination of RT and TMZ. In the recommended treatment, adjuvant TMZ has been shown to improve survival.[40,52] In addition, there is another alternative treatment created by adding lomustine to the RT and TMZ combination in young and resistant patients, but since it has been observed that it has higher toxicity in the observations, standard RT and TMZ combination therapy is considered more appropriate. In glioblastoma patients without MGMT methylation, there was no statistically significant change in survival when the recommended treatment was applied.
Temozolomide, which is used as a pill, modifies many regions of DNA and is an alkylating agent, is applied both simultaneously with RT and as monotherapy after RT. Temozolomide is an oral alkylating agent dosed according to body surface area (BSA). During radiation, TMZ is given daily (seven days a week) at a dose of 75 mg/m2.  Temozolomide is taken on an empty stomach at least two hours after the last meal. Temozolomide should be taken two hours before RT to maximize synergy with radiation.
Bevacizumab, on the other hand, does not directly target tumor cells but targets the vessels that feed the tumor cells and are their oxygen source. Bevacizumab receives the vascular endothelial growth factor (VEGF) signal, which provides new vessel formation and growth, and blocks its formation. So, bevacizumab, a monoclonal antibody that binds to VEGF, is not recommended for routine use in patients with diagnosed glioblastoma. Although bevacizumab has potent antiedema effects that can heal and reduce glucocorticoid requirements in selected patients with unresectable tumors to control refractory edema and mass effect that may occur during or shortly after RT, it has been reported that it does not improve overall survival and increases the risk of toxicity when used as part of initial therapy. Bevacizumab also called a type of rescue therapy, is a chemotherapeutic that is usually administered when the expected success of treatment is not achieved or in relapses.
It is one of the cases reported that the application of RT, which is applied after surgery during the treatment, together with ChT, prolongs the survival by an average of two months.[55,56] According to another report, the survival rate up to two years is 10.4% with the application of RT alone, while this rate is 26.5% when ChT and RT are applied together.
Alternative electric fields, also a method used in glioblastoma treatment; is a portable medical device that is applied to the scalp and produces Tumor Treating Fields (TTFields).[57,58] Monthly use of the device with TMZ in newly diagnosed glioblastoma patients has been shown to improve both progression-free and overall survival.[59,60] Although it is not a treatment that can be applied to every patient because it creates a potential load, it is applied to the shaved scalp in suitable patients with a four transducer array connected to a portable battery or a device working with a power source during the treatment. The device should be applied continuously or for at least 18 hours a day, on the scalp that is kept shaved. It has been reported experimentally that the treatment is a source of antimitotic action, in which alternating electric fields are generated that exert forces on the charged tubulin subunits, thereby interfering with the formation of the mitotic spindle.[61,62] Clinicians who will prescribe this treatment must be trained and certified.
Scores from 0 to 100 on a criterion called the “Karnofsky Performance Scale” determine the patient’s activity. According to this scale; patients with a score of 80-100 can continue their normal activities and do not need care, patients with a score of 50-70 can not work in their social life while they can fulfill most of their personal needs, patients with a score of 0-40 are completely in need of care and their diseases progress much faster. The prognostic factors are primarily affected by Karnofsky’s performance status, age, good mental status, and complete resection of the tumor at a rate of 98%.
The treatment process in older adults may vary depending on factors such as comorbid disease, polypharmacy, increased susceptibility to side effects, and socioeconomic vulnerability. In addition, maximal surgical resection compatible with the preservation of neurological function is recommended instead of biopsy in elderly patients. In the study; A two- or three-month survival advantage was seen after subtotal resection adjusted for gross total resection, tumor size, location, and RT administration. Simultaneous and adjuvant TMZ with RT is also recommended in older adults with good performance status and uneventful comorbidity.
In general, there are no formal clinical studies that define the optimal frequency of followup in patients after treatment. The National Comprehensive Cancer Network (NCCN) guidelines recommend repeat MRI in glioblastoma patients approximately four weeks after completion of RT, then every four months for two to three years, and then less frequently.
Despite the developing technology, the genetic structure and prognosis of glioblastoma, which is the most rapidly progressing and deadly known, still has not been fully determined. In addition, although a complete treatment is not possible yet, the patient's quality of life and survival could not be increased significantly. Even if recurrence is very common, there is currently no application that prevents this.
According to the studies so far, the best treatment is RT and ChT to be applied together after a large-scale resected surgery. In addition to MGMT methylation, young age, a good Karnofsky score, ChT, and RT performed together after surgery is important in terms of prolonging the survival and defining a better clinical prognosis of the patient diagnosed with glioblastoma.
According to the findings in our article, survival time is longer in patients with high Karnofsky scores than in patients with low scores. At the same time, it has been clearly seen that the maximum percentage of tumors resected by surgical operation and the survival time are directly proportional.
In general, no situation was encountered that affected the survival time depending on sex and tumor localization. It is among the findings that patients with MGMT methylation have a better prognosis than patients without MGMT methylation, regardless of treatment. And also, it is among the findings that patients without TERT gene mutation have a better prognosis than patients with TERT gene mutation.
In conclusion, glioblastoma is cytogenetically and genetically heterogeneous. Primary glioblastoma, which develops as de nova and is frequently observed in elderly individuals, has a worse prognosis than those with secondary structure seen in younger patients. Prominence and exacerbation of glioblastoma symptoms depend on the amount of local edema and intracranial pressure. They have a more favorable prognosis with short-term and mild symptoms. Despite treatment options, glioblastoma is still quite mortal. The best prognosis belongs to the tumor masses resected with the most extensive surgery. The fact that the studies up to the present period are not yet sufficient for a definitive treatment clearly reveals that many more studies are needed. Discovering the genetic structure of glioblastoma in detail will affect the treatment process in a very positive way.