Allan H. Friedman

IDH1 and IDH2 Mutations in Gliomas

BACKGROUND A recent genomewide mutational analysis of glioblastomas (World Health Organization [WHO] grade IV glioma) revealed somatic mutations of the isocitrate dehydrogenase 1 gene (IDH1) in a fraction of such tumors, most frequently in tumors that were known to have evolved from lower-grade gliomas (secondary glioblastomas). METHODS We determined the sequence of the IDH1 gene and the related IDH2 gene in 445 central nervous system (CNS) tumors and 494 non-CNS tumors. The enzymatic activity of the proteins that were produced from normal and mutant IDH1 and IDH2 genes was determined in cultured glioma cells that were transfected with these genes. RESULTS We identified mutations that affected amino acid 132 of IDH1 in more than 70% of WHO grade II and III astrocytomas and oligodendrogliomas and in glioblastomas that developed from these lower-grade lesions. Tumors without mutations in IDH1 often had mutations affecting the analogous amino acid (R172) of the IDH2 gene. Tumors with IDH1 or IDH2 mutations had distinctive genetic and clinical characteristics, and patients with such tumors had a better outcome than those with wild-type IDH genes. Each of four tested IDH1 and IDH2 mutations reduced the enzymatic activity of the encoded protein. CONCLUSIONS Mutations of NADP(+)-dependent isocitrate dehydrogenases encoded by IDH1 and IDH2 occur in a majority of several types of malignant gliomas.

Bevacizumab Plus Irinotecan in Recurrent Glioblastoma Multiforme

PURPOSE The prognosis for patients with recurrent glioblastoma multiforme is poor, with a median survival of 3 to 6 months. We performed a phase II trial of bevacizumab, a monoclonal antibody to vascular endothelial growth factor, in combination with irinotecan. PATIENTS AND METHODS This phase II trial included two cohorts of patients. The initial cohort, comprising 23 patients, received bevacizumab at 10 mg/kg plus irinotecan every 2 weeks. The dose of irinotecan was based on the patients anticonvulsant: Patients taking enzyme-inducing antiepileptic drugs (EIAEDs) received 340 mg/m2, and patients not taking EIAEDs received 125 mg/m2. After this regimen was deemed safe and effective, the irinotecan schedule was changed to an accepted brain tumor regimen of four doses in 6 weeks, in anticipation of a phase III randomized trial of irinotecan versus irinotecan and bevacizumab. The second cohort, comprising 12 patients, received bevacizumab 15 mg/kg every 21 days and irinotecan on days 1, 8, 22, and 29. Each cycle was 6 weeks long and concluded with patient evaluations, including magnetic resonance imaging. RESULTS The 6-month progression-free survival among all 35 patients was 46% (95% CI, 32% to 66%). The 6-month overall survival was 77% (95% CI, 64% to 92%). Twenty of the 35 patients (57%; 95% CI, 39% to 74%) had at least a partial response. One patient developed a CNS hemorrhage, which occurred in his 10th cycle. Four patients developed thromboembolic complications (deep venous thrombosis and/or pulmonary emboli). CONCLUSION Bevacizumab and irinotecan is an effective treatment for recurrent glioblastoma multiforme and has moderate toxicity.

Phase II trial of bevacizumab and irinotecan in recurrent malignant glioma

Purpose: Recurrent grade III-IV gliomas have a dismal prognosis with minimal improvements in survival seen following currently available salvage therapy. This study was conducted to determine if the combination of a novel antiangiogenic therapy, bevacizumab, and a cytotoxic agent, irinotecan, is safe and effective for patients with recurrent grade III-IV glioma. Experimental Design: We conducted a phase II trial of bevacizumab and irinotecan in adults with recurrent grade III-IV glioma. Patients with evidence of intracranial hemorrhage on initial brain magnetic resonance imaging were excluded. Patients were scheduled to receive bevacizumab and irinotecan i.v. every 2 weeks of a 6-week cycle. Bevacizumab was administered at 10 mg/kg. The dose of irinotecan was determined based on antiepileptic use: patients taking enzyme-inducing antiepileptic drugs received 340 mg/m2, whereas patients not taking enzyme-inducing antiepileptic drugs received 125 mg/m2. Toxicity and response were assessed. Results: Thirty-two patients were assessed (23 with grade IV glioma and 9 with grade III glioma). Radiographic responses were noted in 63% (20 of 32) of patients (14 of 23 grade IV patients and 6 of 9 grade III patients). The median progression-free survival was 23 weeks for all patients (95% confidence interval, 15-30 weeks; 20 weeks for grade IV patients and 30 weeks for grade III patients). The 6-month progression-free survival probability was 38% and the 6-month overall survival probability was 72%. No central nervous system hemorrhages occurred, but three patients developed deep venous thromboses or pulmonary emboli, and one patient had an arterial ischemic stroke. Conclusions: The combination of bevacizumab and irinotecan is an active regimen for recurrent grade III-IV glioma with acceptable toxicity.

Phase II Trial of Gefitinib in Recurrent Glioblastoma

PURPOSE To evaluate the efficacy and tolerability of gefitinib (ZD1839, Iressa; AstraZeneca, Wilmington, DE), a novel epidermal growth factor receptor tyrosine kinase inhibitor, in patients with recurrent glioblastoma. PATIENTS AND METHODS This was an open-label, single-center phase II trial. Fifty-seven patients with first recurrence of a glioblastoma who were previously treated with surgical resection, radiation, and usually chemotherapy underwent an open biopsy or resection at evaluation for confirmation of tumor recurrence. Each patient initially received 500 mg of gefitinib orally once daily; dose escalation to 750 mg then 1,000 mg, if a patient received enzyme-inducing antiepileptic drugs or dexamethasone, was allowed within each patient. RESULTS Although no objective tumor responses were seen among the 53 assessable patients, only 21% of patients (11 of 53 patients) had measurable disease at treatment initiation. Seventeen percent of patients (nine of 53 patients) underwent at least six 4-week cycles, and the 6-month event-free survival (EFS) was 13% (seven of 53 patients). The median EFS time was 8.1 weeks, and the median overall survival (OS) time from treatment initiation was 39.4 weeks. Adverse events were generally mild (grade 1 or 2) and consisted mainly of skin reactions and diarrhea. Drug-related toxicities were more frequent at higher doses. Withdrawal caused by drug-related adverse events occurred in 6% of patients (three of 53 patients). Although the presence of diarrhea positively predicted favorable OS from treatment initiation, epidermal growth factor receptor expression did not correlate with either EFS or OS. CONCLUSION Gefitinib is well tolerated and has activity in patients with recurrent glioblastoma. Further study of this agent at higher doses is warranted.

Multicenter Phase II Trial of Temozolomide in Patients With Anaplastic Astrocytoma or Anaplastic Oligoastrocytoma at First Relapse

PURPOSE To determine the antitumor efficacy and safety profile of temozolomide in patients with malignant astrocytoma at first relapse. PATIENTS AND METHODS This open-label, multicenter, phase II trial enrolled 162 patients (intent-to-treat [ITT] population). After central histologic review, 111 patients were confirmed to have had an anaplastic astrocytoma (AA) or anaplastic mixed oligoastrocytoma. Chemotherapy-naive patients were treated with temozolomide 200 mg/m(2)/d. Patients previously treated with chemotherapy received temozolomide 150 mg/m(2)/d; the dose could be increased to 200 mg/m(2)/d in the absence of grade 3/4 toxicity. Therapy was administered orally on the first 5 days of a 28-day cycle. RESULTS Progression-free survival (PFS) at 6 months, the primary protocol end point, was 46% (95% confidence interval, 38% to 54%). The median PFS was 5.4 months, and PFS at 12 months was 24%. The median overall survival was 13.6 months, and the 6- and 12-month survival rates were 75% and 56%, respectively. The objective response rate determined by independent central review of gadolinium-enhanced magnetic resonance imaging scans of the ITT population was 35% (8% complete response [CR], 27% partial response [PR]), with an additional 26% of patients with stable disease (SD). The median PFS for patients with SD was 4.4 months, with 33% progression-free at 6 months. Maintenance of progression-free status and objectively assessed response (CR/PR/SD) were both associated with health-related quality-of-life (HQL) benefits. Adverse events were mild to moderate, with hematologic side effects occurring in less than 10% of patients. CONCLUSION Temozolomide demonstrated good single-agent activity, an acceptable safety profile, and documented HQL benefits in patients with recurrent AA.

TERT promoter mutations occur frequently in gliomas and a subset of tumors derived from cells with low rates of self-renewal

Malignant cells, like all actively growing cells, must maintain their telomeres, but genetic mechanisms responsible for telomere maintenance in tumors have only recently been discovered. In particular, mutations of the telomere binding proteins alpha thalassemia/mental retardation syndrome X-linked (ATRX) or death-domain associated protein (DAXX) have been shown to underlie a telomere maintenance mechanism not involving telomerase (alternative lengthening of telomeres), and point mutations in the promoter of the telomerase reverse transcriptase (TERT) gene increase telomerase expression and have been shown to occur in melanomas and a small number of other tumors. To further define the tumor types in which this latter mechanism plays a role, we surveyed 1,230 tumors of 60 different types. We found that tumors could be divided into types with low (<15%) and high (≥15%) frequencies of TERT promoter mutations. The nine TERT-high tumor types almost always originated in tissues with relatively low rates of self renewal, including melanomas, liposarcomas, hepatocellular carcinomas, urothelial carcinomas, squamous cell carcinomas of the tongue, medulloblastomas, and subtypes of gliomas (including 83% of primary glioblastoma, the most common brain tumor type). TERT and ATRX mutations were mutually exclusive, suggesting that these two genetic mechanisms confer equivalent selective growth advantages. In addition to their implications for understanding the relationship between telomeres and tumorigenesis, TERT mutations provide a biomarker that may be useful for the early detection of urinary tract and liver tumors and aid in the classification and prognostication of brain tumors.

Immunologic Escape After Prolonged Progression-Free Survival With Epidermal Growth Factor Receptor Variant III Peptide Vaccination in Patients With Newly Diagnosed Glioblastoma

PURPOSE Immunologic targeting of tumor-specific gene mutations may allow precise eradication of neoplastic cells without toxicity. Epidermal growth factor receptor variant III (EGFRvIII) is a constitutively activated and immunogenic mutation not expressed in normal tissues but widely expressed in glioblastoma multiforme (GBM) and other neoplasms. PATIENTS AND METHODS A phase II, multicenter trial was undertaken to assess the immunogenicity of an EGFRvIII-targeted peptide vaccine and to estimate the progression-free survival (PFS) and overall survival (OS) of vaccinated patients with newly diagnosed EGFRvIII-expressing GBM with minimal residual disease. Intradermal vaccinations were given until toxicity or tumor progression was observed. Sample size was calculated to differentiate between PFS rates of 20% and 40% 6 months after vaccination. RESULTS There were no symptomatic autoimmune reactions. The 6-month PFS rate after vaccination was 67% (95% CI, 40% to 83%) and after diagnosis was 94% (95% CI, 67% to 99%; n = 18). The median OS was 26.0 months (95% CI, 21.0 to 47.7 months). After adjustment for age and Karnofsky performance status, the OS of vaccinated patients was greater than that observed in a control group matched for eligibility criteria, prognostic factors, and temozolomide treatment (hazard ratio, 5.3; P = .0013; n = 17). The development of specific antibody (P = .025) or delayed-type hypersensitivity (P = .03) responses to EGFRvIII had a significant effect on OS. At recurrence, 82% (95% CI, 48% to 97%) of patients had lost EGFRvIII expression (P < .001). CONCLUSION EGFRvIII-targeted vaccination in patients with GBM warrants investigation in a phase III, randomized trial.

Medical complications of aneurysmal subarachnoid hemorrhage: A report of the multicenter, cooperative aneurysm study

OBJECTIVES This report examines the frequency, type, and prognostic factors of medical (nonneurologic) complications after subarachnoid hemorrhage in a large, prospective study. The influences of contemporary neurosurgical, neurological, and critical care practice on mortality and morbidity rates after aneurysmal subarachnoid hemorrhage are evaluated. DESIGN A study of medical complications observed in the placebo limb of a large, randomized, controlled trial of the calcium antagonist, nicardipine, after subarachnoid hemorrhage. SETTING Patients were recruited from 50 hospitals in 41 neurosurgical centers in the United States and Canada. PATIENTS A total of 457 patients with subarachnoid hemorrhage, > or = 18 yrs of age, were randomly assigned to the placebo group. All patients arrived at the participating center within 7 days (mean 1.0 +/- 1.8 [SD] days) of rupture of an angiographically documented saccular aneurysm. MEASUREMENTS AND MAIN RESULTS The frequency rates of symptomatic vasospasm, rebleeding, and total mortality rate after subarachnoid hemorrhage at 3-month follow-up were 46%, 7%, and 19%, respectively. The frequency of having at least one severe (life-threatening) medical complication was 40%. The proportion of deaths from medical complications was 23%. This value was comparable with the proportion of deaths attributed to the direct effects of the initial hemorrhage (19%), rebleeding (22%), and vasospasm (23%) after aneurysmal rupture. The frequency of life-threatening cardiac arrhythmias was 5%; less ominous rhythm disturbances occurred in 30% of the patients. There was an increased frequency of cardiac arrhythmias on the day of, or day after, aneurysm surgery. Pulmonary edema occurred in 23% of the patients, with a 6% occurrence rate incidence of severe pulmonary edema. There was a wide variation from center to center, with the greatest frequency on days 3 through 7. There was a nonsignificant association of pulmonary edema with the use of hypertensive hypervolemic therapy (p = .10), and a significant association with the timing of surgery (p < .05). Some degree of hepatic dysfunction was noted in 24% of patients, the majority with only mild abnormalities of hepatic enzymes with no clinical accompaniment (4% frequency of severe hepatic dysfunction). Thrombocytopenia occurred in 4% of patients, usually in the setting of sepsis. Renal dysfunction was reported in 7% of the patients, with 15% of that figure deemed to be of life-threatening severity. There was an association (p = .001) with antibiotic therapy. CONCLUSIONS Potentially preventable medical complications after ruptured cerebral aneurysm add to the total mortality rate of patients, and may increase length of hospital stay in the critical care setting. The proportion of deaths after subarachnoid hemorrhage from medical complications equals those deaths from either direct effects, rebleeding, or vasospasm individually. Pulmonary complications are the most common nonneurologic cause of death. Cardiac arrhythmia, although frequent, was not associated with significant mortality. The frequency of cardiac arrhythmia and pulmonary edema increased on the day of, or day after, aneurysm surgery. Renal and hepatic dysfunction, and blood dyscrasias, were also observed, underscoring the need for meticulous monitoring for metabolic and hematologic derangements.

DNA mismatch repair and O6-alkylguanine-DNA alkyltransferase analysis and response to Temodal in newly diagnosed malignant glioma

PURPOSE We evaluated the response to Temodal (Schering-Plough Research Institute, Kenilworth, NJ) of patients with newly diagnosed malignant glioma, as well as the predictive value of quantifying tumor DNA mismatch repair activity and O6-alkylguanine-DNA alkyltransferase (AGT). PATIENTS AND METHODS Thirty-three patients with newly diagnosed glioblastoma multiforme (GBM) and five patients with newly diagnosed anaplastic astrocytoma (AA) were treated with Temodal at a starting dose of 200 mg/m2 daily for 5 consecutive days with repeat dosing every 28 days after the first daily dose. Immunochemistry for the detection of the human DNA mismatch repair proteins MSH2 and MLH1 and the DNA repair protein AGT was performed with monoclonal antibodies and characterized with respect to percent positive staining. RESULTS Of the 33 patients with GBM, complete responses (CRs) occurred in three patients, partial responses (PRs) occurred in 14 patients, stable disease (SD) was seen in four patients, and 12 patients developed progressive disease (PD). Toxicity included infrequent grades 3 and 4 myelosuppression, constipation, nausea, and headache. Thirty tumors showed greater than 60% cells that stained for MSH2 and MLH1, with three CRs, 12 PRs, three SDs, and 12 PDs. Eight tumors showed 60% or less cells that stained with antibodies to MSH2 and/or MLH1, with 3 PRs, 3 SDs, and 2 PDs. Eleven tumors showed 20% or greater cells that stained with an antibody to AGT, with 1 PR, 2 SDs, and 8 PDs. Twenty-five tumors showed less than 20% cells that stained for AGT, with 3 CRs, 12 PRs, 4 SDs, and 6 PDs. CONCLUSION These results suggest that Temodal has activity against newly diagnosed GBM and AA and warrants continued evaluation of this agent. Furthermore, pretherapy analysis of tumor DNA mismatch repair and, particularly, AGT protein expression may identify patients in whom tumors are resistant to Temodal.

Increased Regulatory T-Cell Fraction Amidst a Diminished CD4 Compartment Explains Cellular Immune Defects in Patients with Malignant Glioma

Immunosuppression is frequently associated with malignancy and is particularly severe in patients with malignant glioma. Anergy and counterproductive shifts toward T(H)2 cytokine production are long-recognized T-cell defects in these patients whose etiology has remained elusive for >30 years. We show here that absolute counts of both CD4(+) T cells and CD4(+)CD25(+)FOXP3(+)CD45RO(+) T cells (T(regs)) are greatly diminished in patients with malignant glioma, but T(regs) frequently represent an increased fraction of the remaining CD4 compartment. This increased T(reg) fraction, despite reduced counts, correlates with and is sufficient to elicit the characteristic manifestations of impaired patient T-cell responsiveness in vitro. Furthermore, T(reg) removal eradicates T-cell proliferative defects and reverses T(H)2 cytokine shifts, allowing T cells from patients with malignant glioma to function in vitro at levels equivalent to those of normal, healthy controls. Such restored immune function may give license to physiologic antiglioma activity, as in vivo, T(reg) depletion proves permissive for spontaneous tumor rejection in a murine model of established intracranial glioma. These findings dramatically alter our understanding of depressed cellular immune function in patients with malignant glioma and advance a role for T(regs) in facilitating tumor immune evasion in the central nervous system.