SPINT2 is hypermethylated in both IDH1 mutated and wild-type glioblastomas, and exerts tumor suppression via reduction of c-Met activation.

PURPOSE: Both IDH1-mutated and wild-type gliomas abundantly display aberrant CpG island hypermethylation. However, the potential role of hypermethylation in promoting gliomas, especially the most aggressive form, glioblastoma (GBM), remains poorly understood. METHODS: We analyzed RRBS-generated methylation profiles for 11 IDH1WT gliomas (including 7 GBMs), 24 IDH1MUT gliomas (including 6 GBMs), and 5 normal brain samples and employed TCGA GBM methylation profiles as a validation set. Upon classification of differentially methylated CpG islands by IDH1 status, we used integrated analysis of methylation and gene expression to identify SPINT2 as a top cancer related gene. To explore functional consequences of SPINT2 methylation in GBM, we validated SPINT2 methylation status using targeted bisulfite sequencing in a large cohort of GBM samples. We assessed DNA methylation-mediated SPINT2 gene regulation using 5-aza-2'-deoxycytidine treatment, DNMT1 knockdown and luciferase reporter assays. We conducted functional analyses of SPINT2 in GBM cell lines in vitro and in vivo. RESULTS: We identified SPINT2 as a candidate tumor-suppressor gene within a group of CpG islands (designated GT-CMG) that are hypermethylated in both IDH1MUT and IDH1WT gliomas but not in normal brain. We established that SPINT2 downregulation results from promoter hypermethylation, and that restoration of SPINT2 expression reduces c-Met activation and tumorigenic properties of GBM cells. CONCLUSIONS: We defined a previously under-recognized group of coordinately methylated CpG islands common to both IDH1WT and IDH1MUT gliomas (GT-CMG). Within GT-CMG, we identified SPINT2 as a top cancer-related candidate and demonstrated that SPINT2 suppressed GBM via down-regulation of c-Met activation.

Long-Term Effects of Moderate Concussive Brain Injury During Adolescence on Synaptic and Tonic GABA Currents in Dentate Granule Cells and Semilunar Granule Cells.

Progressive physiological changes in the hippocampal dentate gyrus circuits following traumatic brain injury (TBI) contribute to temporal evolution of neurological sequelae. Although early posttraumatic changes in dentate synaptic and extrasynaptic GABA currents have been reported, and whether they evolve over time and remain distinct between the two projection neuron classes, granule cells and semilunar granule cells, have not been evaluated. We examined long-term changes in tonic GABA currents and spontaneous inhibitory postsynaptic currents (sIPSCs) and in dentate projection neurons 3 months after moderate concussive fluid percussion injury (FPI) in adolescent rats. Granule cell tonic GABA current amplitude remained elevated up to 1 month after FPI, but decreased to levels comparable with age-matched controls by 3 months postinjury. Granule cell sIPSC frequency, which we previously reported to be increased 1 week after FPI, remained higher than in age-matched controls at 1 month and was significantly reduced 3 months after FPI. In semilunar granule cells, tonic GABA current amplitude and sIPSC frequency were not different from controls 3 months after FPI, which contrast with decreases observed 1 week after injury. The switch in granule cell inhibitory inputs from early increase to subsequent decrease could contribute to the delayed emergence of cognitive deficits and seizure susceptibility after brain injury.

Correlation of commercially available quantitative MGMT (O-6-methylguanine-DNA methyltransferase) promoter methylation scores and GBM patient survival.

BACKGROUND: Between 2011 and 2016, O-6-methylguanine-DNA methyltransferase (MGMT) promoter methylation testing at University of California Los Angeles (UCLA) was performed through LabCorp, using a threshold of 2 to distinguish MGMT methylated from unmethylated tumors. In this study, we sought to determine whether the magnitude of the methylation score correlated with outcome. METHODS: We identified 165 newly diagnosed glioblastoma (GBM) isocitrate dehydrogenase (IDH) wild-type and temozolomide-treated upfront patients at UCLA and Kaiser Permanente Los Angeles with LabCorp-derived quantitative MGMT scores obtained on pretreatment tissue samples. Using LabCorp's threshold, we found 102 unmethylated and 63 methylated patients. We then further substratified each group based on the magnitude of the score, and performed Kaplan-Meier and Cox regression analyses of overall survival (OS) and progression-free survival (PFS). RESULTS: We validated that the standard LabCorp threshold of 2 could separate our cohort by survival, showing longer OS and PFS for MGMT methylated patients vs unmethylated patients. Cox regression analysis confirmed that MGMT (<1) patients had worse outcome, with OS and PFS hazard ratios of 2.375 (P = .053) and 2.463 (P = .023), respectively, when compared to the MGMT (1-1.99) patients. Contrary to our expectation, when we substratified the ≥2 (methylated) group, we did not find a dose-dependent relationship between the magnitude of MGMT methylation and improved survival. CONCLUSIONS: The MGMT unmethylated group contains a partially methylated group (greater than 1) that shares survival benefits similar to the methylated group. However, we did not demonstrate an association of very high methylation scores with increased survival. These findings will require validation in additional independent clinical data sets.

Bevacizumab at first recurrence after standard radio-chemotherapy is associated with improved overall survival in glioblastoma patients with large tumor burden.

BACKGROUND: Since its approval for use in recurrent glioblastoma (GBM), the survival benefit of bevacizumab (Bev) remains to be demonstrated. To address this issue, we retrospectively examined survival from first recurrence in patients treated with Bev, lomustine (CCNU), or Bev/CCNU. METHODS: We identified 168 primary GBM patients diagnosed at UCLA and Kaiser Permanente LA who received upfront radio-chemotherapy, followed by Bev and/or CCNU at first recurrence. Three patient groups, contemporaneously diagnosed from 2009 through 2015, were identified: (1) patients treated with Bev alone (n = 49), (2) CCNU alone (CCNU 09-15) (n = 36), and (3) Bev/CCNU (n = 53). Another CCNU control group (n = 30) diagnosed from 2001 through 2004 (CCNU 01-04) was also derived. We measured tumor size at first recurrence treatment initiation, using bidimensional (2D) and volumetric (3D) techniques, and analyzed overall survival (OS) from first recurrence. RESULTS: Among the entire cohort, larger tumor size at first recurrence was associated with poorer survival. The CCNU 01-04 group had similar tumor size as the Bev arms and low Bev crossover (7%). Treatment with Bev was associated with improved survival in patients with large tumor 2D measurements: Median OS for Bev and Bev/CCNU groups were 6.71 mo (n = 27) and 6.97 mo (n = 36) vs 4.03 mo (n = 10) in CCNU 01-04. Analysis by 3D measurement yielded similar results. Interestingly, the CCNU 09-15 group showed the highest survival, likely due to smaller tumor size and crossover to Bev (69%). CONCLUSION: Survival advantage from Bev treatment was observed only among patients with large tumor burden as determined by either 2D or 3D measurement.