Eric T. Wong

Updated Response Assessment Criteria for High-Grade Gliomas: Response Assessment in Neuro-Oncology Working Group

Currently, the most widely used criteria for assessing response to therapy in high-grade gliomas are based on two-dimensional tumor measurements on computed tomography (CT) or magnetic resonance imaging (MRI), in conjunction with clinical assessment and corticosteroid dose (the Macdonald Criteria). It is increasingly apparent that there are significant limitations to these criteria, which only address the contrast-enhancing component of the tumor. For example, chemoradiotherapy for newly diagnosed glioblastomas results in transient increase in tumor enhancement (pseudoprogression) in 20% to 30% of patients, which is difficult to differentiate from true tumor progression. Antiangiogenic agents produce high radiographic response rates, as defined by a rapid decrease in contrast enhancement on CT/MRI that occurs within days of initiation of treatment and that is partly a result of reduced vascular permeability to contrast agents rather than a true antitumor effect. In addition, a subset of patients treated with antiangiogenic agents develop tumor recurrence characterized by an increase in the nonenhancing component depicted on T2-weighted/fluid-attenuated inversion recovery sequences. The recognition that contrast enhancement is nonspecific and may not always be a true surrogate of tumor response and the need to account for the nonenhancing component of the tumor mandate that new criteria be developed and validated to permit accurate assessment of the efficacy of novel therapies. The Response Assessment in Neuro-Oncology Working Group is an international effort to develop new standardized response criteria for clinical trials in brain tumors. In this proposal, we present the recommendations for updated response criteria for high-grade gliomas.

Outcomes and Prognostic Factors in Recurrent Glioma Patients Enrolled Onto Phase II Clinical Trials

PURPOSE To determine aggregate outcomes and prognostic covariates in patients with recurrent glioma enrolled onto phase II chemotherapy trials. PATIENTS AND METHODS Patients from eight consecutive phase II trials included 225 with recurrent glioblastoma multiforme (GBM) and 150 with recurrent anaplastic astrocytoma (AA). Their median age was 45 years (range, 15 to 82 years) and their median Karnofsky performance score was 80 (range, 60 to 100). Prognostic covariates were analyzed with respect to tumor response, progression-free survival (PFS), and overall survival (OS) by multivariate logistic and Cox proportional hazards regression analyses. RESULTS Overall, 34 (9%) had complete or partial response, whereas 80 (21%) were alive and progression-free at 6 months (APF6). The median PFS was 10 weeks and median OS was 30 weeks. Histology was a robust prognostic factor across all outcomes. GBM patients had significantly poorer outcomes than AA patients. The APF6 proportion was 15% for GBM and 31% for AA, whereas the median PFS was 9 weeks for GBM and 13 weeks for AA. Results were also significantly poorer for patients with more than two prior surgeries or chemotherapy regimens. CONCLUSION Histology is a dominant factor in determining outcome in patients with recurrent glioma enrolled onto phase II trials. Future trials should be designed with separate histology strata.

The Cerebrospinal Fluid Provides a Proliferative Niche for Neural Progenitor Cells

Cortical development depends on the active integration of cell-autonomous and extrinsic cues, but the coordination of these processes is poorly understood. Here, we show that the apical complex protein Pals1 and Pten have opposing roles in localizing the Igf1R to the apical, ventricular domain of cerebral cortical progenitor cells. We found that the cerebrospinal fluid (CSF), which contacts this apical domain, has an age-dependent effect on proliferation, much of which is attributable to Igf2, but that CSF contains other signaling activities as well. CSF samples from patients with glioblastoma multiforme show elevated Igf2 and stimulate stem cell proliferation in an Igf2-dependent manner. Together, our findings demonstrate that the apical complex couples intrinsic and extrinsic signaling, enabling progenitors to sense and respond appropriately to diffusible CSF-borne signals distributed widely throughout the brain. The temporal control of CSF composition may have critical relevance to normal development and neuropathological conditions.

NovoTTF-100A versus physician's choice chemotherapy in recurrent glioblastoma: A randomised phase III trial of a novel treatment modality

PURPOSE NovoTTF-100A is a portable device delivering low-intensity, intermediate frequency electric fields via non-invasive, transducer arrays. Tumour Treatment Fields (TTF), a completely new therapeutic modality in cancer treatment, physically interfere with cell division. METHODS Phase III trial of chemotherapy-free treatment of NovoTTF (20-24h/day) versus active chemotherapy in the treatment of patients with recurrent glioblastoma. Primary end-point was improvement of overall survival. RESULTS Patients (median age 54 years (range 23-80), Karnofsky performance status 80% (range 50-100) were randomised to TTF alone (n=120) or active chemotherapy control (n=117). Number of prior treatments was two (range 1-6). Median survival was 6.6 versus 6.0 months (hazard ratio 0.86 [95% CI 0.66-1.12]; p=0.27), 1-year survival rate was 20% and 20%, progression-free survival rate at 6 months was 21.4% and 15.1% (p=0.13), respectively in TTF and active control patients. Responses were more common in the TTF arm (14% versus 9.6%, p=0.19). The TTF-related adverse events were mild (14%) to moderate (2%) skin rash beneath the transducer arrays. Severe adverse events occurred in 6% and 16% (p=0.022) of patients treated with TTF and chemotherapy, respectively. Quality of life analyses favoured TTF therapy in most domains. CONCLUSIONS This is the first controlled trial evaluating an entirely novel cancer treatment modality delivering electric fields rather than chemotherapy. No improvement in overall survival was demonstrated, however efficacy and activity with this chemotherapy-free treatment device appears comparable to chemotherapy regimens that are commonly used for recurrent glioblastoma. Toxicity and quality of life clearly favoured TTF.

Differential Gene Expression Analysis Reveals Generation of an Autocrine Loop by a Mutant Epidermal Growth Factor Receptor in Glioma Cells

The epidermal growth factor receptor (EGFR) gene is commonly amplified and rearranged in glioblastoma multiforme leading to overexpression of wild-type and mutant EGFRs. Expression of wild-type EGFR ligands, such as transforming growth factor-alpha (TGF-alpha) or heparin-binding EGF (HB-EGF), is also often increased in gliomas resulting in an autocrine loop that contributes to the growth autonomy of glioma cells. Glioblastoma multiformes express a characteristic EGFR mutant (EGFRvIII, de 2-7) that does not bind ligand, signals constitutively, and is more tumorigenic than the wild-type receptor. However, the downstream signals that mediate this increased tumorigenicity are not well understood. We hypothesized that signals induced specifically by EGFRvIII and not the wild-type receptor are more likely to mediate its increased tumorigenic activity and examined the gene expression profiles resulting from inducible expression of comparable levels of either wild-type EGFR or EGFRvIII in a U251-MG glioma cell line. Expression of EGFRvIII resulted in specific up-regulation of a small group of genes. Remarkably, all these genes, which include TGFA, HB-EGF, EPHA2, IL8, MAP4K4, FOSL1, EMP1, and DUSP6, influence signaling pathways known to play a key role in oncogenesis and function in interconnected networks. Increased expression of EGFRvIII-induced genes was validated by real-time PCR. The mutant receptor does not bind ligand, and EGFRvIII-induced expression of TGF-alpha and HB-EGF suggests that EGFRvIII plays a role in generating an autocrine loop using the wild-type EGFR in glioma. It also raises the possibility that EGFRvIII may signal, at least in part, through the wild-type receptor. Indeed, we show that inhibiting the activity of HB-EGF, a potent mitogen, with neutralizing antibodies reduces cell proliferation induced by expression of EGFRvIII. This suggests that the EGFRvIII-HB-EGF-wild-type EGFR autocrine loop plays an important role in signal transduction by EGFRvIII in glioma cells. We also show by immunohistochemistry that HB-EGF expression correlates with the presence of EGFRvIII in glioblastoma multiforme. Thus, our study provides a new insight into oncogenic signaling by EGFRvIII and improves our understanding of how autocrine loops are generated in glioma.

BEAMing and Droplet Digital PCR Analysis of Mutant IDH1 mRNA in Glioma Patient Serum and Cerebrospinal Fluid Extracellular Vesicles

Development of biofluid-based molecular diagnostic tests for cancer is an important step towards tumor characterization and real-time monitoring in a minimally invasive fashion. Extracellular vesicles (EVs) are released from tumor cells into body fluids and can provide a powerful platform for tumor biomarkers because they carry tumor proteins and nucleic acids. Detecting rare point mutations in the background of wild-type sequences in biofluids such as blood and cerebrospinal fluid (CSF) remains a major challenge. Techniques such as BEAMing (beads, emulsion, amplification, magnetics) PCR and droplet digital PCR (ddPCR) are substantially more sensitive than many other assays for mutant sequence detection. Here, we describe a novel approach that combines biofluid EV RNA and BEAMing RT-PCR (EV-BEAMing), as well droplet digital PCR to interrogate mutations from glioma tumors. EVs from CSF of patients with glioma were shown to contain mutant IDH1 transcripts, and we were able to reliably detect and quantify mutant and wild-type IDH1 RNA transcripts in CSF of patients with gliomas. EV-BEAMing and EV-ddPCR represent a valuable new strategy for cancer diagnostics, which can be applied to a variety of biofluids and neoplasms.

Exclusion of a Proton ATPase from the Apical Membrane Is Associated with Cell Polarity and Tip Growth in Nicotiana tabacum Pollen Tubes

Polarized growth in pollen tubes results from exocytosis at the tip and is associated with conspicuous polarization of Ca2+, H+, K+, and Cl− -fluxes. Here, we show that cell polarity in Nicotiana tabacum pollen is associated with the exclusion of a novel pollen-specific H+-ATPase, Nt AHA, from the growing apex. Nt AHA colocalizes with extracellular H+ effluxes, which revert to influxes where Nt AHA is absent. Fluorescence recovery after photobleaching analysis showed that Nt AHA moves toward the apex of growing pollen tubes, suggesting that the major mechanism of insertion is not through apical exocytosis. Nt AHA mRNA is also excluded from the tip, suggesting a mechanism of polarization acting at the level of translation. Localized applications of the cation ionophore gramicidin A had no effect where Nt AHA was present but acidified the cytosol and induced reorientation of the pollen tube where Nt AHA was absent. Transgenic pollen overexpressing Nt AHA-GFP developed abnormal callose plugs accompanied by abnormal H+ flux profiles. Furthermore, there is no net flux of H+ in defined patches of membrane where callose plugs are to be formed. Taken together, our results suggest that proton dynamics may underlie basic mechanisms of polarity and spatial regulation in growing pollen tubes.

The role of brevican in glioma: promoting tumor cell motility in vitro

BackgroundMalignant glioma is a common primary tumor of the central nervous system. Brevican, an abundant extracellular matrix component in the adult brain, plays a critical role in the process of glioma. The mechanisms for the highly invasive behavior of gliomas are still poorly understood. The aim of this study was to examine whether brevican is a predictor of glioma and its roles in glioma cell motility.MethodsIn this study, immunohistochemistry staining for brevican expression was performed in malignant gliomas and benign controls. We also explored the effects of brevican on cell adhesion and migration in brevican-overexpressed cells. Knockdown of brevican expression was achieved by stable transfection of U251 cells transduced with a construct encoding a short hairpin DNA directed against the brevican gene, which correspondingly, down-regulated the proliferation, invasion and spread of brevican-expressing cells. Moreover, the role of brevican in the growth and progression of glioma was demonstrated by in vivo studies.ResultsOur results provide evidence for the molecular and cellular mechanisms that may underlie the motility-promoting role of brevican in the progression of glioma. The role of brevican as a target for immunotherapy might be taken into consideration in future studies.ConclusionsThis study suggests that expression of brevican is associated with glioma cell adhesion, motility and tumor growth, and also is related to glioma cell differentiation, therefore it may be a marker for malignance degree of glioma

An Open Label Trial of Sustained-release Cytarabine (DepoCyt™) for the Intrathecal Treatment of Solid Tumor Neoplastic Meningitis

Drugs currently available for intrathecal administration are cleared rapidly from the CSF. DepoCyt is a slow-release formulation of cytarabine that maintains cytotoxic concentrations of free cytarabine in the CSF for >14 days following a single injection. DepoCyt was administered to 110 patients with a diagnosis of neoplastic meningitis based on either a positive CSF cytology (76) or neurologic and CT or MRI scan findings sufficient to document neoplastic meningitis (34). Patients were treated with DepoCyt 50 mg every 2 weeks for 1 month of induction therapy by either lumbar puncture (LP) or intraventricular (IVT) injection. Patients without neurologic progression were candidates to receive an additional 3 months of consolidation therapy. All patients received dexamethasone 4 mg BID on days 1–5 of each cycle. Median time to neurologic progression was 55 days; median overall survival was 95 days. Among the 76 patients with a positive CSF cytology at baseline, 70 were evaluable for response, and of this group19 (27%) attained the criteria for response (cytologic response in the absence of neurologic progression). The most important adverse events were headache and arachnoiditis. When drug-related, these were largely low grade, transient, and reversible. Drug-related grade 3 headache occurred on 4% of cycles; grade 3 or 4 arachnoiditis occurred on 6% of cycles. No cumulative toxicity was observed. DepoCyt injected once every 2 weeks produced a response-rate comparable to that previously reported for methotrexate given twice a week. The once in every 2-week-dosing interval offers an advantage over conventional schedules (2–3 doses/week) used for other agents available for intrathecal injection.

Bevacizumab Reverses Cerebral Radiation Necrosis

A 43-year-old Chinese woman, with a 6-month history of hearing loss, was diagnosed with biopsy-proven T3, N1, M0, stage III undifferentiated nasopharyngeal carcinoma. Her initial head magnetic resonance imaging (MRI) scan showed a mass in the nasopharynx (Fig 1A, asterisk) extending into the left sphenoid sinus (Fig 1B, arrow), and there was fluid accumulation in the left mastoid process. She received cisplatin and involved-field intensity-modulated radiotherapy (IMRT) to 70 Gy, followed by an additional three cycles of cisplatin and fluorouracil. Her inferior temporal lobes received up to 68.5 Gy of radiation (Fig 1C, red isodose line). Two years later, she experienced verbal memory loss, together with fatigue and mild imbalance. Her neurological examination was notable for psychomotor slowing and deficit in immediate recall. She also had a mini–mental status examination score of 26 (from a total of 30), suggesting moderate cognitive dysfunction. A gadolinium-enhanced head MRI, taken for evaluation of her neurocognitive deficits, showed enhancement in the inferior left temporal lobe with increased fluid-attenuated inversion recovery signals in the surrounding brain parenchyma (Figs 2A and 2B). She underwent [F]fluorodeoxyglucose (FDG)–positron emission tomography and thallium–single photon emission computed tomography, and revealed no uptake of either radionuclide in the corresponding region of gadolinium enhancement. The inferior left temporal lobe did not have increased blood flow, as measured by arterial spin labeling. Because the findings suggest radiation necrosis of the brain, she was treated with 5 mg/kg of bevacizumab every other week. After four doses, the enhancement on MRI was nearly gone (Fig 3A & 3B) and her mini–mental status examination score increased to 30. Her imbalance and fatigue disappeared. These neurocognitive improvements persisted at the time of follow-up 6 months later. Temporal lobe radiation necrosis was common before the adoption of IMRT for head and neck cancer. The spectrum of this disorder ranged from edema noted as hyperintense fluid-attenuated inversion recovery, T2 signals in the temporal lobes, and hemorrhagic transformation, to necrotic cysts causing mass effect. The development of cerebral radiation necrosis from fractionated radiotherapy is a function of total dose, dose per fraction, and time from completion of radiation. The higher the total dose or dose per fraction, the sooner cerebral radiation necrosis would appear in patients. The mechanism appears to be a result of radiation damage to vascular endothelial cells, causing endothelial cell proliferation, telangiectatic vessels, and fibrinoid necrosis with accompanying perivascular exudation and edema. In experimental settings, microhemorrhages was noted in the brains of rabbits when they were treated with proton stereotaxic radiosurgery. Similarly, in the gastrointestinal tract of mice treated with 15 Gy of whole-body irradiation, radiation-induced endothelial cell apoptosis was the primary mechanism leading to stem-cell dysfunction, crypts damage, and death. The secondary stem-cell dysfunction was probably a result of damage to the vascular niche where the stem cells reside. Although the kinetics of this damage is unknown in experimental animals, patients who received radiation at 62.5 Gy or more have greater than 25% probability of radiation necrosis within 5 years. Our patient received a dose of up to 68.5 Gy in the inferior temporal lobe, making her high risk for developing radiation necrosis despite the tight conformality of IMRT-limiting scattered radiation to the temporal lobes. The gadolinium-enhancing abnormality was not FDG or thallium avid, suggesting that it was unlikely to be tumor or infection. A diagnosis of cerebral radiation necrosis was also supported by more than 2 years of delayed development of her temporal abnormality. Conventional treatment consisted of corticosteroid to limit the extent of cerebral edema and, if necessary, drainage of compressive cysts. Unfortunately, due to the unrelenting process of necrosis, patients in the past typically succumbed to complications of