A novel enhancer regulates MGMT expression and promotes temozolomide resistance in glioblastoma

Temozolomide (TMZ) was used for the treatment of glioblastoma (GBM) for over a decade, but its treatment benefits are limited by acquired resistance, a process that remains incompletely understood. Here we report that an enhancer, located between the promoters of marker of proliferation Ki67 (MKI67) and O6-methylguanine-DNA-methyltransferase (MGMT) genes, is activated in TMZ-resistant patient-derived xenograft (PDX) lines and recurrent tumor samples. Activation of the enhancer correlates with increased MGMT expression, a major known mechanism for TMZ resistance. We show that forced activation of the enhancer in cell lines with low MGMT expression results in elevated MGMT expression. Deletion of this enhancer in cell lines with high MGMT expression leads to a dramatic reduction of MGMT and a lesser extent of Ki67 expression, increased TMZ sensitivity, and impaired proliferation. Together, these studies uncover a mechanism that regulates MGMT expression, confers TMZ resistance, and potentially regulates tumor proliferation

Real-time methylation-specific polymerase chain reaction for MGMT promoter methylation clinical testing in glioblastoma an alternative detection method for a heterogeneous process

Objectives: To develop and evaluate a real-time methylation-specific polymerase chain reaction (RT-MSP) MGMT assay, with a particular focus on small biopsies and indeterminate testing results. Methods: We assessed formalin-fixed paraffin-embedded glioblastoma or gliosarcoma specimens (n = 641). A testvalidation group (n = 51) with previously obtained reference laboratory (RL) results was used to determine performance characteristics of the RT-MSP assay. An indeterminate (equivocal) category was established for cases that could not be clearly classified as positive or negative. Results: Overall agreement of RT-MSP and RL results was 91% (41/45 nonindeterminate cases). Discordant cases were tested by pyrosequencing, and results were most concordant with RT-MSP. Among cases with limited amounts of tissue (n = 7), six yielded valid results by RT-MSP (all negative); the single invalid result consisted of a stereotactic biopsy specimen obtained 14 years prior. A subset of indeterminate cases obtained during clinical testing (n = 18/575 [3%]) was also evaluated by pyrosequencing and showed a heterogeneous pattern of methylation across the eight interrogated CpG sites. Conclusions: The RT-MSP assay that we developed in-house is a robust clinical detection method for the heterogeneous process of MGMT promoter methylation in glioblastoma

Phase I-II trial of imatinib mesylate (Gleevec; STI571) in treatment of recurrent oligodendroglioma and mixed oligoastrocytoma. North central cancer treatment group study N0272 (ALLIANCE/NCCTG)

Purpose: To evaluate the pharmacokinetics and efficacy of imatinib in patients with recurrent oligodendroglial tumors. Methods: Patients with progressive WHO grade II-III recurrent tumors after prior RT and chemotherapy were eligible. A phase I dose-escalation study was conducted for patients on enzyme-inducing anticonvulsants (EIAC). A phase II study for non-EIAC patients utilized a fixed dose of 600 mg/D. Primary efficacy endpoint was 6-month progression-free survival (PFS6). A 2-stage design was utilized, with 90% power to detect PFS6 increase from 25 to 45%. Results: In the Phase I, maximum tolerated dose was not reached at 1200 mg/D. For phase II patients, overall PFS6 was 33% and median PFS 4.0 months (95% CI 2.1, 5.7). Median overall survival (OS) was longer in imatinib-treated patients compared with controls (16.6 vs. 8.0 months; HR = 0.64, 95% CI 0.41,1.0, p = 0.049), and longer in patients with 1p/19q-codeleted tumors (19.2 vs. 6.2 months, HR = 0.43, 95% CI 0.21,0.89, p = 0.019). Confirmed response rate was 3.9% (PR = 1; REGR = 1), with stable disease observed in 52.9%. At 600 mg/D, mean steady-state imatinib plasma concentration was 2513 ng/ml (95% CI 1831,3195). Grade 3\u20134 adverse events (hematologic, fatigue, GI, hypophosphatemia, or hemorrhage) occurred in 61%. Conclusions: Although adequate plasma levels were achieved, the observed PFS6 of 33% did not reach our pre-defined threshold for success. Although OS was longer in imatinib-treated patients than controls, this finding would require forward validation in a larger cohort. Imatinib might show greater activity in a population enriched for PDGF-dependent pathway activation in tumor tissue

Phase 1/2 trial of temsirolimus and sorafenib in the treatment of patients with recurrent glioblastoma: North Central Cancer Treatment Group Study/Alliance N0572

BACKGROUND: Mitogen-activated protein kinase (MAPK) activation and mammalian target of rapamycin (mTOR)-dependent signaling are hallmarks of glioblastoma. In the current study, the authors conducted a phase 1/2 study of sorafenib (an inhibitor of Raf kinase and vascular endothelial growth factor receptor 2 [VEGFR-2]) and the mTOR inhibitor temsirolimus in patients with recurrent glioblastoma. METHODS: Patients with recurrent glioblastoma who developed disease progression after surgery or radiotherapy plus temozolomide and with 642 prior chemotherapy regimens were eligible. The phase 1 endpoint was the maximum tolerated dose (MTD), using a cohorts-of-3 design. The 2-stage phase 2 study included separate arms for VEGF inhibitor (VEGFi)\u2013naive patients and patients who progressed after prior VEGFi. RESULTS: The MTD was sorafenib at a dose of 200 mg twice daily and temsirolimus at a dose of 20 mg weekly. In the first 41 evaluable patients who were treated at the phase 2 dose, there were 7 who were free of disease progression at 6 months (progression-free survival at 6 months [PFS6]) in the VEGFi-naive group (17.1%); this finding met the prestudy threshold of success. In the prior VEGFi group, only 4 of the first 41 evaluable patients treated at the phase 2 dose achieved PFS6 (9.8%), and this did not meet the prestudy threshold for success. The median PFS for the 2 groups was 2.6 months and 1.9 months, respectively. The median overall survival for the 2 groups was 6.3 months and 3.9 months, respectively. At least 1 adverse event of grade 653 was observed in 75.5% of the VEGFi-naive patients and in 73.9% of the prior VEGFi patients. CONCLUSIONS: The limited activity of sorafenib and temsirolimus at the dose and schedule used in the current study was observed with considerable toxicity of grade 653. Significant dose reductions that were required in this treatment combination compared with tolerated single-agent doses may have contributed to the lack of efficacy. Cancer 2018;124:1455-63. \ua9 2018 American Cancer Society

Is the blood-brain barrier really disrupted in all glioblastomas? A critical assessment of existing clinical data

The blood-brain barrier (BBB) excludes the vast majority of cancer therapeutics from normal brain. However, the importance of the BBB in limiting drug delivery and efficacy is controversial in high-grade brain tumors, such as glioblastoma (GBM). The accumulation of normally brain impenetrant radiographic contrast material in essentially all GBM has popularized a belief that the BBB is uniformly disrupted in all GBM patients so that consideration of drug distribution across the BBB is not relevant in designing therapies for GBM. However, contrary to this view, overwhelming clinical evidence demonstrates that there is also a clinically significant tumor burden with an intact BBB in all GBM, and there is little doubt that drugs with poor BBB permeability do not provide therapeutically effective drug exposures to this fraction of tumor cells. This review provides an overview of the clinical literature to support a central hypothesis: that all GBM patients have tumor regions with an intact BBB, and cure for GBM will only be possible if these regions of tumor are adequately treated

The medical necessity of advanced molecular testing in the diagnosis and treatment of brain tumor patients

Accurate pathologic diagnoses and molecularly informed treatment decisions for a wide variety of cancers depend on robust clinical molecular testing that uses genomic, epigenomic, and transcriptomic-based tools. Nowhere is this more essential than in the workup of brain tumors, as emphasized by the incorporation of molecular criteria into the 2016 World Health Organization classification of central nervous system tumors and the updated official guidelines of the National Comprehensive Cancer Network. Despite the medical necessity of molecular testing in brain tumors, access to and utilization of molecular diagnostics is still highly variable across institutions, and a lack of reimbursement for such testing remains a significant obstacle. The objectives of this review are (i) to identify barriers to adoption of molecular testing in brain tumors, (ii) to describe the current molecular tools recommended for the clinical evaluation of brain tumors, and (iii) to summarize how molecular data are interpreted to guide clinical care, so as to improve understanding and justification for their coverage in the routine workup of adult and pediatric brain tumor cases