Abstracts from the 2011 BNOS Conference, June 29 – July 1, 2011, Homerton College, Cambridge

doi:10.1093/neuonc/nor144

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

2. GAS6/AXL-FAMILY RECEPTORS IN SCHWANNOMA PATHOLOGICAL PROLIFERATION, ADHESION AND SURVIVAL

S Ammoun ¹, L Zhou ¹, M Barczyk ¹, D Hilton ¹, S Hafizi ¹, CO Hanemann ¹

Abstract

INTRODUCTION: Tumour suppressor merlin deficiency leads to the development of schwannomas, meningiomas and ependymomas. Using our in vitro model of human schwannoma we have demonstrated that merlin-deficiency leads to increased cell proliferation, cell-matrix adhesion and survival involving ErbB2/3, platelet-derived-growth-factor-receptor-β (PDGFR-β), and Insulin-like growth factor receptor-I (IGF-IR) acting via extracellular-signal-regulated-kinase 1/2 (ERK1/2), AKT and JNK. We have inhibited increased proliferation of schwannoma by AZD6244, sorafenib 2, nilotinib, lapatinib 1 and BEZ-235. Since, schwannoma overexpress multiple receptors/signalling pathways the inhibition of a single target is not sufficient. Therefore, all relevant receptors/signalling pathways must be revealed. AXL subfamily RTKs (AXL, SKY, MER and Ron) are over-expressed in cancers correlating with poor prognosis. Expression profiling and phosphoprotein arrays showed that Axl-family receptors are overexpressed/activated in human schwannoma. METHODS: Western blotting, immunohistochemistry, primary human cell culture, proliferation/adhesion assays. RESULTS: AXL, SKY and their ligand Gas-6 are over-expressed as well as activated in human schwannoma cells and tissues leading to increased proliferation and adhesion. CONCLUSIONS: Axl, SKY and Gas-6 constitute new potential therapeutic targets in merlin- deficient tumours.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

3. CD133 GLYCOSYLATION IS AFFECTED BY HYPOXIA IN CULTURED GLIOMA STEM CELLS

Kristina S Lehnus ¹, Laura K Donovan ¹, Geoffrey J Pilkington ¹, Qian An ¹

Abstract

INTRODUCTION: The widely employed cancer stem cell (CSC) marker CD133 has come under scrutiny in gliomas as the expression and poorly characterised glycosylation patterns of CD133 could affect immunological detection of CD133+ CSCs. The present study aims to investigate the effect of oxygen deprivation on CD133 expression and glycosylation in a glioblastoma multiforme (GBM) cell line, IN699. METHODS: Cells were cultured under normoxic (21% oxygen) and hypoxic (3% oxygen) conditions. Two CD133 antibodies, CD133/1 (glycosylated) and ab19898 (unglycosylated), were used for CD133 immunodetection by flow cytometry and immunocytochemistry (ICC). RESULTS: Using flow cytometry, ab19898 detected 94.1% and 96.2% CD133+ cells under normoxia and hypoxia respectively. Hypoxia significantly increased the percentage of CD133+ cells from 69% to 92% using CD133/1 antibody (p < 0.005). Although similar percentages of CD133+ cells were detected in hypoxia irrespective of antibodies used, a significantly higher geomean fluorescence intensity (GMI) was demonstrated by ab19898 (p < 0.005) in positive cells. ICC confirmed that ab19898 binds CD133 intracellularly while CD133/1 binds the extracellular domain. CONCLUSIONS: This study demonstrates the shortcomings of CD133/1 as the most used antibody to detect CD133+ CSCs. Our data indicate that 1) previously reported CD133– cells may have been misidentified using the glycosylation-specific CD133/1 as constitutive expression of CD133 was detected by the intracellular antibody ab19898; 2) hypoxia promotes glycosylation status of CD133. Therefore, the role of CD133 should be further explored in order to elucidate its biological significance. We are currently investigating CD133-associated signal transduction pathways and the impact of CD133-KD (knockdown) on cell biology.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

4. COMPLICATIONS FOLLOWING GLIADEL WAFER INSERTION DURING SURGERY FOR GLIOBLASTOMA MULTIFORME: A SINGLE CENTRE-EXPERIENCE

Ian A Anderson ¹, Simon Thomson ¹

Abstract

INTRODUCTION & OBJECTIVE: Surgically implanted carmustine-impregnated wafers (Gliadel^®) confer a significant survival benefit in the surgical treatment of high grade glioma although complications have been reported including seizures, poor wound healing and infection. Complication rates are infrequently quantified in published reviews. We report the complication rates for all patients treated with Gliadel at a Yorkshire neurosurgical centre in a two-year period. METHODS: Retrospective review of case notes, electronic records and GP records. Outcome data collected on survival, wound breakdown, wound infection, post-operative length of stay, readmission rates, need for further surgery and seizure record. RESULTS: 11 patients identified: 8 male, 3 female; mean age 45.6 years; 10 glioblastoma multiforme, 1 anaplastic pleomorphic xanthoastrocytoma; 7 recurrent and 4 primary surgeries; 7 frontal, 2 parietal and 2 temporal tumours. 4 patients have subsequently died (mean post-operative survival 13 months, range 1–33) and 7 remain under follow-up (mean time since resection 8 months, range 2–17). 1 episode of suspected infected wound breakdown documented (9%); treated with oral antibiotics as an outpatient. No patients were re-admitted or required further surgery. Mean post-operative length of stay was 4.3 days (range 1 to 15). CONCLUSIONS: Quoted rates may over-estimate the frequency of complications following insertion of Gliadel wafers after resection of high grade glioma. Just as clinicians must scrutinise the evidence in favour of a new treatment before implementing it, we must maintain objectivity when considering the significance of any adverse effects of a product before discounting it.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

6. SURVIVAL OF PATIENTS WITH GLIOBLASTOMA – HOW DO WE COMPARE TO STUPP?

M Bailey ¹, E Lekka ¹, J Law ¹, C Davis ¹

Abstract

INTRODUCTION: The publication of the results of the EORTC-NCIC trial in 2005 and 2009 changed the standard of care for patients with histologically proven glioblastoma. Their treatment protocol of 60Gy of radiotherapy plus concomitant daily Temozolomide followed by up to six cycles of adjuvant Temozolomide has become known as the “Stupp regime”. The present study compares the survival of patients with Glioblastoma treated at our institution with the reported survival of the EORTC-NCIC trial. METHODS: We retrospectively collected and analysed data of treatment and survival of all patients with histologically proven glioblastoma for the five year period from 1/1/06 to 31/12/10. The Kaplan-Meier method was used to produce survival data and these were compared with the EORTC-NCIC trial results. RESULTS: In the 5 years from Jan 2006 to Dec 2010, 375 patients had a histological diagnosis of glioblastoma and 98 were treated with the “Stupp regime”. Of these, 68.4% underwent debulking surgery and 31.6% underwent biopsy. Median age was 56, median survival 13.0 months and the 2-year survival rate was 17.3% (compared to 14.6 months median survival and 2-year survival rate of 27.2% in the EORTC-NCIC trial). For the latter 3 years from Jan 2008 to Dec 2010 the estimated 2-year survival rate was 22%. In 2006 56% of patients underwent debulking surgery compared to 73% in 2009 and 82% in 2010. CONCLUSIONS: At our institution we have yet to achieve the survival results achieved in the EORTC-NCIC trial. However, our estimated 2-year survival rate for glioblastoma patients has increased with time, and this may be due to an increased proportion of patients receiving surgical debulking rather than biopsy.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

7. STEREOTACTIC RADIOSURGERY AT LEEDS GAMMA KNIFE CENTRE: INITIAL EXPERIENCE OF TREATING BRAIN METASTASES

KE Banfill ¹, C Loughrey ¹, P Hatfield ¹

Abstract

INTRODUCTION: Stereotactic radiosurgery for brain metastases has been carried out at the Leeds Gamma Knife Centre since March 2009. METHOD: A retrospective review of patients treated with radiosurgery between March 2009 and July 2010 was carried out (minimum 6 months follow up). Data on survival, primary tumour, Karnofsky performance status (KPS), previous whole brain radiotherapy (WBRT) and time from diagnosis to identification of brain metastases were recorded at initial presentation. Patients were then followed up with regular magnetic resonance imaging of the brain and data on toxicity and oral steroid dose were recorded. RESULTS: 58 patients (19 male/39 female) had a median survival of 50.4 weeks (95% CI, 32.6 to 68.2 weeks). Lung (36%) and breast (27%) were the most common primary tumours. 12% of patients had WBRT prior to radiosurgery and these patients had improved survival (possibly due to case selection). There was a clear but non-significant trend that patients with a disease-free interval longer than 1 year had a better outcome. The size of the largest treated metastasis seemed a better predictor of outcome than number of metastases treated.16% of patients had WBRT following radiosurgery. Data on steroid dose at first follow up were available for 45 patients. Steroid dose dropped significantly after radiosurgery (P < 0.01) and was the same or less in 91% of patients. There were only 3 cases of grade 3 toxicity (fatigue, weakness and seizure). CONCLUSION: Our study reports survival comparable with other series on radiosurgery and demonstrates a significant decrease in steroid dose following treatment.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

8. HIGH THROUGHPUT DRUG SCREENING OF PAEDIATRIC GLIOMA CELL LINES

Dorine Bax ¹, Richard Elliott ¹, Ryan Bishop ¹, Katy Taylor ¹, Lynley Marshall ¹, Nathalie Gaspar ¹, Marta Viana-Pereira ¹, Rui Reis ¹, Jane Renshaw ¹, Alan Ashworth ¹, Chris Lord ¹, Chris Jones ¹

Abstract

INTRODUCTION: The clinical outcome for children with high grade glioma is dismal, with little change in the past four decades. Novel therapies targeting the genetic alterations that drive the disease are therefore sorely needed. Recent molecular profiling data have demonstrated that paediatric high grade gliomas differ biologically from their adult counterparts, and therefore may require different targeted agents. In order to identify candidates for targeted inhibition, we have performed an in vitro high-throughput drug screen on a series of well-characterised paediatric glioma cell lines. METHODS: The drug screen consisted of commercially available small molecule inhibitors and chemotherapeutic agents, at concentrations ranging from 1nM–1 µM. The compounds were screened in triplicate on the cell lines SF188, KNS42, UW479, Res259 and Res186. The primary endpoint was cell viability as measured using the CellTiter-Glo assay. Selected hits were investigated in more detail using the MTS assay, immunoblotting, expression microarrays and cell cycle analysis. RESULTS: The drug screen data were compared to similar data generated on a panel of breast and ovarian cancer cell lines, identifying both generalisable and cell line specific hits. All glioma cell lines were found to be highly sensitive to inhibitors of aurora kinase, PI3-kinase/mTOR and the molecular chaperone HSP90. Individual cell lines were found to be sensitive to inhibition of distinct receptor tyrosine kinases related to the molecular profiles of the cells, including PDGFR, EGFR, MET and FGFR families. CONCLUSION: These data demonstrate the suitability of high-throughput drug screening to identify novel classes of targeted therapies, which may be applied to paediatric high grade glioma models. In-depth validation of target inhibition related to the underlying biology of the tumour cells will provide rationale for selecting patients most likely to benefit from specific therapeutic regimens.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

9. THE SELECTIVE ACTION OF TRICHOSTATIN A IN COMBINATION WITH THE DNA DAMAGING AGENTS, TEMOZOLOMIDE AND CISPLATIN

CA Bellamy ¹, L Shaw ¹, JE Alder ¹, AJ Shorrocks ¹, RW Lea ¹

Abstract

INTRODUCTION: Changes to the epigenome of a cell precede genetic alterations during transformation to a malignant state; hence enzymes involved in epigenetic modulation are a potential therapeutic target. One such target is histone deacetylase (HDAC), an enzyme involved in regulation of gene expression by deacetylation of histones. HDAC inhibitors (HDACi) promote DNA damage in normal and cancerous cells however, due to aberrant genetics of neoplasms, only normal cells can repair this damage. The aim of this study was to assess whether HDACi combined with the DNA damaging agents cisplatin and temozolomide, could work selectively and synergistically to increase cytotoxicity in a glioma cell line compared to a normal astrocytic culture. METHODS: 0.1 nM Trichostatin A (TSA) was used in combination with 1 µM temozolomide or 1 µM cisplatin to treat U87-MG glioblastoma and SVG-P12 normal cells. Single treatments of each drug were included for comparison. An MTS cell proliferation assay was performed to compare the cytotoxic effects of the drug combinations on the cell lines. RESULTS: A significant decrease in cell proliferation for both combinations, compared to single drug treatment, was seen in the U87-MG cell line. There was no significant difference between combination treatments and single drug treatments in the SVG-P12 cell line. CONCLUSIONS: TSA showed a significant synergistic and selective effect on the U87 glioma cell line in combination with DNA damaging agents. This highlights the potential of novel combination therapies to increase efficacy when treating glioma, which may enable dose-reduction of chemotherapeutic agents and hence reduced side-effects.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

10. MODULATING IMMUNOSUPPRESSIVE BEHAVIOUR IN GLIOBLASTOMA

Suzanne Birks ¹, Michael Burnet ¹, Geoffrey Pilkington ¹

Abstract

INTRODUCTION: Glioblastoma are remarkably adept at evading immune responses mounted against them and have been shown to manipulate immune components into promoting tumourigenesis. They achieve this through a number of synergistic methods, including the release of inhibitory cytokines, that converge to inhibit T-cell proliferation, activation and migration as well as inducing T-cell apoptosis. Administering compounds that up-regulate the expression of tumour-suppressive cytokines whilst decreasing the immunosuppressive cytokines may show some therapeutic benefits. METHODS: Using biopsy-derived cell cultures at P1 and P2 we have analysed the macrophage and myeloid components of the population as well as assessing IL-10, IL-12, IL-23 and IFN-γ expression and secretion by Western blot, flow cytometry and ICC. After treating the cells with 9 compounds (Synovo GmbH), which regulate macrophage phenotype through NFκB activation and translocation promotion, at 10, 3, 1 and 0.3 µM for 72 hours and subsequently performed the same analyses as described above. RESULTS: Using MAC387 antibody, we have shown the macrophage content of our early passage biopsy-derived cells to be around 30% and similarly quantified the expression of IL-10, IL-12, IL-23 and IFN-γ. Preliminary studies show that several of the compounds enhance the expression of the tumour-suppressor cytokines IL-12 and IFN-γ while decreasing amounts of IL-10 and IL-23. CONCLUSIONS: Initial data suggest that modulating the expression of certain cytokines may promote the anti-tumour immune response. Further investigation is required on how this manipulation affects tumour cell behaviour and survival; studies will also determine the effect on other immune components such as myeloid cells.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

12. A SINGLE CENTRE CASE CONTROL STUDY OF THE EFFICACY AND SAFETY OF 5-AMINOLEVULINIC ACID GUIDED RESECTIONS OF GRADE IV (WHO) GLIOBLASTOMAS

Julius D Bruch ¹, Jasmine Ho ¹, Colin Watts ¹, Stephen J Price ¹

Abstract

INTRODUCTION: This is a study to assess the safety and efficacy of using 5-aminolevulinic acid (5ALA) to guide the resection of grade IV glioblastomas based on the data obtained in a single UK centre. METHODS: 74 patients with a primary histologically-confirmed grade IV glioblastoma who underwent 5ALA guided resection were included in the study. Each patient was matched by age and tumour location to a control patient with similar performance status who had a resection without 5ALA. Outcome measures include residual tumour on post-op MRI, post-op deficits and access to chemoradiotherapy. RESULTS: The proportion of gross total resections (GTRs) in 5ALA patients was 53% (n = 74) and thus significantly better compared to the control group with 30% (n = 57), (p = 0.01). The absence of residual fluorescence at the end of the resection predicted a significantly better GTR rate of 72%, compared to a GTR rate of 19% in the presence of residual fluorescence (p < 0.0001). Two cases with mild adverse effects could be attributed to 5ALA (3%). There was no significant difference in median length of hospital stay and new neurological deficits between the two groups. 5ALA treatment did significantly not compromise access to chemoradiotherapy (p < 0.05). CONCLUSIONS: 5ALA guided resection of grade IV glioblastomas results in a significantly better GTR rate compared to conventional surgery. This result reflects the findings of the previous stage III clinical trial. The procedure also appears to be safe with no significant increase in hospital stay, post-operative complications and only few side effects.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

13. REALTIME INTRAOPERATIVE THREE DIMENSIONAL ULTRASOUND IN BIOPSY/RESECTION OF INTRINSIC BRAIN LESIONS

SJ Camp ¹, V Apostolopoulos ¹, A Mehta ¹, F Roncaroli ¹, D Nandi ¹

Abstract

INTRODUCTION: Accurate tissue diagnosis and near total resection influence management/outcome in intrinsic brain lesions, hence intraoperative imaging has gained popularity. It allows readjustment of biopsy needle positioning, visualisation of vital structures and eloquent regions, progression of tumour resection, and management of immediate complications. Realtime three dimensional ultrasound (3DUS) is a cost effective alternative to intraoperative magnetic resonance imaging (MRI), which despite finer images requires expensive resources and prolongs surgery. METHODS: This ongoing study assessed the benefits of realtime intraoperative 3DUS in patients with intrinsic brain lesions. Thirty six patients were evaluated: 15 underwent multiple biopsies; 19 maximum resection. Data from 2 patients were not analysed due to poor image quality. In biopsy patients preoperative scans and intraoperative 3DUS images were correlated with the descriptive characteristics of histological samples. The images of patients in whom maximum resection was achieved, were correlated with postoperative MRI. RESULTS: Discrepancies between navigation based on preoperative imaging and intraoperative 3DUS were observed due to brain shift. 3DUS produced a high biopsy yield. In a patient with gliomatosis cerebri 3DUS detected areas of differing echogenicity, correlating with varying cellularity. Radical or complete resection of low grade lesions was also facilitated by realtime 3DUS. CONCLUSIONS: Intra-operative 3DUS can be employed as an adjunct to preoperative imaging. It improves the yield of biopsy and maximises the resection of intrinsic brain lesions, with few changes required to existing theatre costs, space and scheduling.

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14. THROMBOCYTOPENIA DURING CHEMORADIOTHERAPY FOR GLIOBLASTOMA: POTENTIAL IMPLICATIONS FOR ADJUVANT TREATMENT AND OUTCOME

B Clark ¹, M Mackinnon ¹, N MacLeod ¹, W Stewart ¹, A Chalmers ¹

Abstract

INTRODUCTION: Concurrent chemoradiotherapy with oral temozolomide, followed by up to 6 cycles of adjuvant temozolomide is standard of care following resection of glioblastoma (GBM) in fit patients under 70 years old. Myelosuppression, particularly thrombocytopenia, is common during and after concurrent treatment. We investigated the incidence of thrombocytopenia during concurrent chemoradiotherapy and its impact on subsequent delivery of adjuvant chemotherapy and overall survival. METHODS: We retrospectively examined the medical records of 130 consecutive patients treated with concurrent chemoradiotherapy for GBM at a single institution between 2005 and 2010. Survival was calculated from date of surgery and analysed by the Kaplan-Meier method using SPSS version 16. RESULTS: Complete haematological results were available for 117 patients. Median duration of follow-up from date of surgery was 14.4 months. The incidence of any grade thrombocytopenia during the concurrent phase was 29.9%, grade 3 or 4 incidence was 6.0%. Of patients who developed grade 1 or higher thrombocytopenia during concurrent treatment, 14.3% completed 6 adjuvant cycles, median number of cycles was 1. Median OS was 13.5 months, 2-year OS 15.2%. Of those who maintained normal platelet counts, 42.7% completed 6 cycles, median number of cycles was 5. Median OS was 19.3 months, 2-year OS 31.0% (log rank p = 0.02). CONCLUSIONS: Thrombocytopenia is common during concurrent chemoradiotherapy for GBM and is associated with a poorer outcome. This may reflect difficulty in administering adjuvant temozolomide and subsequent lines of palliative chemotherapy. It also lends support to the importance of the adjuvant phase of treatment.

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15. ADHERENCE TO NICE GUIDANCE IMPROVES SURVIVAL FOR GBM PATIENTS IN NORTHERN IRELAND

AJ Cole ¹, GG Hanna ¹, K Bailie ¹, DS Conkey ¹, JA Harney ¹

Abstract

INTRODUCTION: Outcomes following radiotherapy in the treatment of Glioblastoma Multiforme (GBM) are poor. A recent phase III study comparing concurrent chemoradiotherapy with temozolomide versus radiotherapy alone has demonstrated increased median survival with the use of chemoradiotherapy. NICE guidance advises radical radiotherapy with the use of concurrent temozolomide in the treatment of histologically proven WHO Grade IV primary brain tumours and in those patients with ECOG performance status (PS) of <2. We report our two year institutional experience. METHODS: In a retrospective audit of neurooncology registrations in Northern Ireland from June 2007 to March 2009, patients with grade IV Glioblastoma were identified. Baseline patient characteristics, prior surgical management (biopsy/debulking/radiological) and treatment received were recorded. The completion rate of chemoradiotherapy was documented. Kaplan meier estimates of overall survival (OS) were calculated and outcomes in either group compared using the log rank test. RESULTS: We identified 86 NUMBERS? patients with GBM, 19 received chemoradiotherapy, 9 had radical radiotherapy. There was 100% completion of concomitant temozolomide. 12/19 (63.1%) completed the adjuvant component of treatment. In those treated with chemoradiotherapy, good PS and prior debulking surgery were associated with improved OS (p < 0.0001). Median OS for those receiving chemoradiotherapy was 22.9 months versus 7.5 months for radical radiotherapy alone (p < 0.0001). CONCLUSION: In our series, baseline PS, prior surgical debulking and chemoradiotherapy are associated with improved overall survival. Concurrent chemoradiotherapy should be offered to appropriate patients with grade IV malignant brain tumours.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

16. THE EFFECT OF CORTICOSTEROIDS ON THE T2 APPEARANCE OF GLIOBLASTOMA MULTIFORME

CA Darlow ¹, S Chapman ¹, LA Mohsen ¹, SJ Price ¹

Abstract

BACKGROUND: Glioblastoma multiforme (GBM) are known for their invasion, which is a key reason for their poor prognosis. On MRI they are characterised by T1 enhancing lesion, surrounded by a peri-tumoural T2 enhancing region, traditionally thought to be oedema or non-enhancing tumour. Corticosteroids are used in the treatment of brain tumours for their oedema relieving effects. We hypothesise that this T2 enhancing region is representative of non-T1 enhancing invasive tumour. METHODS: T2 weighted MRI scans from patients with glioblastoma (n = 29) and brain metastases (n = 14) before and after corticosteroid therapy were compared to see the effect of the treatment on the size of the T2 enhancing region. Measurements of the T2 weighted area were measured at the axial level of the largest T1 enhancing lesion cross-section area. RESULTS: The change in size of the T2 weighted lesion as a ratio of the size of the T1 enhancing lesion after corticosteroid therapy was insignificant in the GBM cohort (p = 0.19) but significant in the metastases cohort (p = 0.037). CONCLUSION: If the T2 enhancing region is purely oedema, it should be reduced by corticosteroid therapy. In the non-invasive brain metastases, this is the case. However, in the invasive GBM cohort, the T2 enhancing region doesn't significantly reduce with corticosteroid therapy. This is evidence supporting our hypothesis that the T2 enhancing lesion may be invasive tumour. This is supported by histological studies that tumour cells are present at the periphery of the T2 enhancing region. Further work needs to be done assessing how the T2 abnormalities affect outcome and recurrence patterns.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

17. INDUCING GLIOBLASTOMA CANCER STEM CELL DIFFERENTIATION WITH DIBUTYRYL CYCLIC-AMP AND THEOPHYLLINE, PITUITARY ADENYLATE CYCLASE ACTIVATING POLYPEPTIDE AND ALL-TRANS RETINOIC ACID

Laura Donovan ¹, Suzanne Birks ¹, Geoffrey Pilkington ¹

Abstract

INTRODUCTION: Glioblastomas represent a considerable therapeutic challenge due to their remarkable heterogeneity and adaptability and potent ability to evade or resist current treatment strategies. Recent studies point to a small subset of cells – the cancer stem cell population – that may mediate many of the clinical and biological phenomenon associated with glioblastoma – for example, angiogenesis, immunosuppression, tumour self renewal etc. Inducing differentiation in the cancer stem cell population is an attractive approach which may not only eradicate much of the tumorigenic behaviour associated with these cells but render them more resistant to therapy. METHODS: Using an early passage biopsy-derived human paediatric cell line grown in neurobasal medium under normoxic conditions, we have treated the cells for 72 hours with either db-cAMP and theophylline (1mM), PACAP (1nM) or ATRA (10nM) before analysing the expression of GFAP, O4, Tu-20, CD133, Nestin, Mushashi 1, Olig 2 and Sox2 by flow cytometry and immunocytochemistry. Viability, proliferation, live cell imaging and cell cycle analyses were also carried out. RESULTS: Of the three agents, db-cAMP and theophylline had the most striking effect, with cells differentiating along an astrocytic lineage within 48 hours as shown by the increased expression of GFAP and down-regulation of the stem cell markers. Treatment with ATRA and PACAP showed similar but less pronounced effects (where p = 0.05). CONCLUSIONS: Differentiation agents, in particular db-cAMP and theophylline, appear to reduce the stem cell like behaviour of glioblastoma cells treated in vitro. Future studies will include elucidating signal transduction pathways as well as determining post-differentiation therapeutic sensitivity to nuclear and mitochondrial agents.

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18. RETROSPECTIVE BASELINE AUDIT OF PATIENTS RECEIVING LOW DOSE PALLIATIVE RADIOTHERAPY FOR GLIOBLASTOMAMULTIFORME (GBM) IN NHS TAYSIDE

Helen Dyer ¹, Hannah Lord ¹

Abstract

INTRODUCTION: GBM is an aggressive CNS malignancy with especially poor prognosis in older and less fit patients. The optimal treatment of this patient group remains controversial. Low dose radiotherapy using 30Gy in 6 fractions is a recognised treatment choice. We review outcomes from this treatment option in the NHS Tayside population. METHOD: Patients receiving low dose radiotherapy (30Gy in 6 fractions; 2 weeks) were identified from radiotherapy booking forms from 2007–2010 Patient age, gender, date of radiotherapy, performance status and date of death were recorded. Post- radiotherapy quality of life and radiation toxicity noted. RESULTS: Eleven patients received this radiation schedule. 9 males: 2 females. Median age 69.5 years, range 55–77 years. Overall survival from diagnosis was 105 days (range 37–289 days). Patients less than 65 years (n = 2) had an overall survival of 134.5 days, and survival from date of radiotherapy was 95 days; patients older than 75 years (n = 5) overall survival was 98 days and from date of radiotherapy 78 days. CONCLUSION: Our survival rates are in keeping with published data. Older age at diagnosis has a negative impact on survival. Younger patients live longer, both from diagnosis and after radiotherapy. Small sample size precludes statistical analysis of other demographic factors. Qualities of life after radiotherapy and radiation toxicity were poorly recorded and will now be introduced. This is important as palliative radiotherapy can be toxic for older patients with less cerebral reserve and chemotherapy or best supportive care may be more appropriate.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

19. AN INTERPRETATIVE PHENOMENOLOGICAL ANALYSIS OF THE PATIENT EXPERIENCE OF AWAKE CRANIOTOMY - FROM DIAGNOSIS TO DISCHARGE

Kimberley Fletcher ¹, Roshan das Nair ¹, Jamie MacNiven ¹, Surajit Basu ¹, Paul Byrne ¹

Abstract

INTRODUCTION: Previous research exploring the patient experience of awake craniotomy has yielded inconsistent results. In the current study the authors aimed to explore the lived experience of having undergone an awake craniotomy in the United Kingdom. METHODS: Seven participants took part in the study, which used a qualitative research methodology. Participants were recruited from a Department of Neurosurgery in the United Kingdom. Each participant was interviewed using a semi-structured interview schedule. Interviews were audio recorded and transcribed verbatim. Transcripts were analysed using interpretative phenomenological analysis. RESULTS: Analysis of transcripts yielded three super-ordinate themes: self-preservation, operation environment and information. The super-ordinate themes were interpreted as interconnected with each other, as well as embedded in a core theme: relationship with the neurosurgeon. CONCLUSIONS: The relationship with the neurosurgeon appears crucial to the patient experience of awake craniotomy. The relationship could impact on the participants' decision to have the awake craniotomy, their experience of the awake operation and their acceptance of the information given. This knowledge requires consideration in clinical settings and could lead to improved delivery of care in the future.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

20. SURGICAL TREATMENT OF CEREBRAL METASTASES

LJ Glancz ¹, G Critchley ¹

Abstract

INTRODUCTION: The aim of our study was to elucidate the proportion of cancer operations performed for metastatic disease and the survival of these patients. METHODS: Data were collected on the number of operations performed for treatment of cerebral malignancy between January 2007 and December 2010 at our institution. All patients undergoing surgery for excision of cerebral metastases were included. This information along with basic demographics, location of lesion, presentation and histology was ascertained from histology forms. Survival was calculated from date of surgery to date of death or last follow-up, updated to March 2011. RESULTS: 410 operations were undertaken for treatment of cerebral malignant tumours over this 4 year period. Of these 71 (17.4%) were for metastatic disease. The most common histology of metastases was bronchial carcinoma (42%) followed by breast carcinoma (17%), malignant melanoma (13%), gastro-intestinal (11%) and renal cell carcinoma (9%). Greatest survival was observed in the breast cancer group (median survival of 9.5 months) followed by the renal cell carcinoma (8.5 months), melanoma (6 months), gastro-intestinal (5 months) and lung carcinoma groups (4 months). CONCLUSIONS: A significant proportion of operations are performed at our unit for treatment of intracranial metastases. This reflects an increasing number of patients with cerebral metastases being offered surgery. Although survival is poor, our data corroborate an increasing literature that surgery in combination with other treatment is indispensable in significantly extending lifespan for these patients when selected for appropriately.

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21. SURVIVAL ANALYSIS FOR APPARENT DIFFUSION COEFFICIENT MEASURES IN CHILDREN WITH EMBRYONAL BRAIN TUMOURS

M Grech-Sollars ¹, D Saunders ¹, K Phipps ¹, JD Clayden ¹, CA Clark ¹

Abstract

INTRODUCTION: Apparent Diffusion Coefficient (ADC) measures have been used in various tumour analyses. We investigated a previously described ADC measure, the Apparent Transient Coefficient in Tumour (ATCT), to study whether it correlates with survival outcome in children with embryonal brain tumours. The ATCT is a measure of the gradient change of ADC from the peri-tumoural oedema into the tumour core. METHOD: 55 patients with histologically proven embryonal brain tumours and who had diffusion-weighted imaging (DWI) as part of their clinical imaging were enrolled in a retrospective study looking at relating ADC measures with survival. A multivariate survival analysis was carried out considering as covariates: ATCT, mean ADC, minimum ADC, age at diagnosis, tumour location and patients less than 3 years old at diagnosis. RESULTS: Results from the multivariate analysis indicated that the significant covariates were: ATCT, tumour location and children less than 3 years old at diagnosis (p = 0.00022, 0.032, 0.049 respectively). Survival analysis using Kaplan-Meier curves, dividing the patients into four groups of increasing magnitudes of ATCT, showed statistically significant results of poorer prognosis for more negative values of ATCT (p = 0.00000068). CONCLUSIONS: A statistically significant difference was observed for survival data in children with embryonal brain tumours with respect to the change in ADC from oedema into the tumour volume. The results indicate that more negative values of ATCT are related to lower survival, whilst changes closer to zero indicate a higher survival. ATCT is a sensitive biomarker that correlates with survival in childhood embryonal brain tumours.

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22. ROLE OF BMI1 OVEREXPRESSION IN DNA DAMAGE IN NEURAL STEM CELLS AND IN GLIOBLASTOMA INITIATING CELLS

Azzura Greco ¹, Serena Acquati ¹, Silvia Marino ¹

Abstract

INTRODUCTION: The Polycomb group (PcG) protein Bmi1 is an epigenetic gene silencer which plays a key role in self-renewal of embryonic and adult neural stem cells (NSC). It is overexpressed in several types of brain tumours such as for example in high grade gliomas. Here, we investigated how Bmi1 overexpression influences cellular properties of normal and glial cancer stem cells. METHODS: A Cre/LoxP-based conditional transgenic approach was used to induce gain of function of Bmi1 in embryonic and postnatal NSC and in differentiating neural progenitor cells (NPC) originating thereof. Immunostaining, qRT-PCR, immunoprecipitation, and chromatin immunoprecipitation (ChIP) were performed on these cells. RNA interference for Bmi1 was also carried out in high grade glioma initiating cells (HGG-IC) isolated from a murine model where p53 and PTEN were inactivated. RESULTS: Conditional activation of Bmi1 induces ectopic proliferation and oxidative stress induced DNA damage followed by apoptotic death in NPC but not NSC. Furthermore, we observed a cell type specific downregulation of Survivin, a member of the inhibitor of apoptosis protein family, both in vivo and in cultured NPC. This is due to enhanced direct binding of Bmi1 to the survivin promoter, as shown by ChIP. Interestingly, no downregulation of survivin is observed in NSC and in HGG-IC overexpressing Bmi1. CONCLUSIONS: We show a differentiation stage specific role of Bmi1 overexpression in neural stem/progenitor cells, which highlights a potential parallel between the role of Bmi1 in normal and glial cancer stem cells.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

24. THERMAL MEASUREMENT OF INTRACRANIAL GLIOMAS USING MRI SPECTROSCOPY

Salah Hammouche ¹, Simone P Wilkins ¹, Trevor Smith ¹, Andrew Brodbelt ¹

Abstract

INTRODUCTION: MRI spectroscopy (MRS) can be used successfully to non-invasively measure brain temperature in a number of neurological pathologies. Lower temperatures in these conditions correlated with increased blood flow, decreased metabolism and better prognosis. Similar studies have not been conducted in the case of intracranial gliomas. This study aims to use MRS to determine whether there is a difference in brain temperature between the centres of glioma masses compared with normal neural tissues. METHODS: Twenty six eligible patients who had MRS imaging at 3 Tesla using a Philips Achieva MR Scanner were identified. All patients had histologically confirmed glioma (16 WHO grade II, 2 WHO grade III, and 8 WHO grade IV); 1:1 male/female ratio. Images were processed using Philips embedded post processing software. Using self-referencing water proton resonance frequency shift technique, the temperature of central voxels of the tumour masses and mirrored voxels on the normal-appearing hemispheres were calculated. Paired t-test was utilised to determine the statistical significance in temperature differences. RESULTS: In grade II gliomas, the temperature of the centers of tumour masses was significantly lower by one degree Celsius when compared with the contralateral normal-appearing hemispheres (p = 0.0128). Although similar differences were not identified in other grades, the small number of processed voxels might have limited such correlation. In fact, the used processing software was able to calculate the temperature in only 50% of the examined voxels. CONCLUSIONS: WHO grade II gliomas are cooler than the contralateral brain. The significance of this is not clear. Temperature measurement is easy to perform and applicable in a busy clinical setting, but is limited by current processing software. This technique may be useful in the future to provide additional prognostic information on patients presenting with gliomas. Further research is needed.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

25. LONG-TERM SURVIVAL ANALYSIS OF ATYPICAL MENINGIOMAS: SURVIVAL RATES, PROGNOSTIC FACTORS, OPERATIVE AND RADIOTHERAPY TREATMENT

Salah Hammouche ¹, Simon Clark ¹, Alex Hie Lin Wong ¹, Paul Eldridge ¹, Jibril Osman Farah ¹

Abstract

INTRODUCTION: Due to the rarity and inconsistent criteria for defining atypical meningioma prior to the WHO 2000 classification, its management and prognostic factors are poorly understood. Only few articles addressed survival rates of WHO-classified atypical meningiomas; with small numbers or disproportionate representation of irradiated patients. Thus, it was difficult to assess the impact of radiotherapy treatment. This study evaluated whether extent of surgical resection and receiving postoperative radiotherapy along with other patient characteristics had influenced recurrence and survival rates of atypical meningiomas. METHODS: Clinical and surgical notes of the 79 patients with grade II atypical meningioma treated at our centre between 1996 and 2009 were retrospectively evaluated. The histology grading was consistent with WHO 2000 classification. Simpson grading system was used to assess the extent of surgical resection. Kaplan Meier analysis, Cox multivariate regression analysis and Log-rank test were conducted. RESULTS: The average age at the time of initial operation was 58 years, 54 % were males. The mean follow up period was 50 months. In Cox multi-variate analysis, only Simpson grading was predictive of recurrence (hazard ratio = 2.22/1 increase in Simpson grade. p = 0.003). Simpson grade I patients had a relapse-free survival rate of 97%, 74% at 1 and 5 years respectively compared with 88%, 32 % in the subtotal resection group (Simpson grades II to IV). There was no statistically significant correlation between recurrence and subjecting patients to postoperative radiotherapy. Apart from Simpson grade I patients, there was a general trend for worse outcomes in irradiated patients. CONCLUSIONS: The most important prognostic factor in determining recurrence was Simpson grading. There was no statistically significant impact of adjuvant radiotherapy on the recurrence of atypical meningiomas. Meta-analysis for the existing literature is needed.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

26. THE USE OF INTRA-OPERATIVE CARMUSTINE CHEMOTHERAPY WAFERS IN THE MANAGEMENT OF HIGH GRADE GLIOMAS: AN ASSESSMENT OF COMPLICATIONS AND ACCESS TO FURTHER THERAPY

J Ho ¹, J Bruch ¹, C Watts ¹, SJ Price ¹

Abstract

INTRODUCTION: NICE have approved the use of Carmustine wafers and Temozolomide in the treatment of high-grade gliomas, but have not commented on the combined use. The concern is that Carmustine wafers will lead to increased complications that will prevent patients receiving Temozolomide. This study aims to see if this is happening. METHODS: Retrospective review of high-grade glioma patients in whom the intention was to insert Carmustine wafers. Outcome data assessed complication rates and subsequent access to oncological treatment. Those patients where Carmustine was not inserted were used as controls. RESULTS: 83 patients were included, 35 received Carmustine wafers and 48 did not. Out of the 48, 10 had diagnoses that were not high-grade gliomas (HGG). Comparing HGG patients only, 9/35 (25.7%) in the Carmustine group and 7/38 (18.4%,) in the control group not receiving Carmustine wafers experienced complications, showing no significant difference between the two groups (p = 0.574). Subsequently, 30/35 (85.7%) of those receiving Carmustine wafers went on to have chemoradiotherapy compared to 24/38 (63.2%) of those not receiving Carmustine wafers. CONCLUSION: In our series, we found that there is no difference in complication rates between patients given Carmustine wafers and those who were not. The use of Carmustine wafers also did not compromise the subsequent access to radical oncological follow-up treatment.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

27. IMPLEMENTATION OF VMAT FOR INTRA-CRANIAL TUMOURS: A PLANNING COMPARISON

G Lamb ¹, S Smith ¹, A James ¹, M Glegg ¹

Abstract

INTRODUCTION: This planning study compared conventional RT with multiple VMAT (Varian Solution) treatment plans for intra-cranial sites with planning target volumes (PTVs) overlapping organs at risk (OARs). METHOD: 6 patients were planned using conformal RT, & using 3 VMAT techniques. The 1st VMAT planning technique utilised a single complete arc, the 2nd used 2 concentric coplanar arcs & the 3rd technique utilised 4 non coplanar semi arcs, positioned to avoid entry through eyes. The PTV was separated into high & lower dose regions receiving 60Gy & 54Gy in 30 fractions respectively. The lower dose region encompassed overlapping OARs. The objectives were to cover 95% of the PTVs to 95% (CI95) of the dose & 99% of the PTVs to 90% (CI90) of the dose, without violating OAR constraints. Dose volume histograms for PTV & OARs were compared. RESULTS: Semi arc treatment plans achieved all CI90 & CI95 PTV & OAR dose constraints for all patients. Double arc technique achieved all the constraints for only 1 out of 6 patients while the single arc plans did not achieve all constraints for any patient. Double & single arc plans were able to achieve the mean brain dose constraint for only 1 patient, other OAR constraints were met. Conformal RT plans never achieved PTV or brain dose constraints. Normal tissue conformity index (NI CI 95%) was comparable for all VMAT treatment plans (0.97–0.99) & significantly reduced for conformal RT (0.65–0.75). CONCLUSIONS: VMAT achieved better PTV coverage when compared with conventional RT. Similar OAR doses were achieved, except for normal brain tissue, where the dose was greater with conventional RT, single & double arc VMAT plans. Semi arc non-coplanar treatment plans produced better PTV coverage than single or double arc VMAT plans while achieving OAR dose constraints. Semi arc non-coplanar treatment plans will now be used as a class solution when planning primary brain tumours that have PTVs overlapping OARs.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

28. IMMUNOHISTOCHEMICAL ANALTSIS OF EMMPRIN AND CYCLOPHILIN A EXPRESSION IN ADULT GLIOMA

T Jeffcote ¹, S Boulos ¹, P Robbins ¹, N Knuckey ¹

Abstract

INTRODUCTION: Extracellular matrix metalloproteinase inducer (EMMPRIN), a member of the immunoglobulin superfamily, is present on the surface of tumor cells. It is closely associated with Cyclophilin A (CypA) a cytoplasmic and extracellularly expressed molecule upregulated in cerebral hypoxia. We have analyzed the clinicopathological characteristics of EMMPRIN and CypA expression in normal brain tissue, low grade and high grade glioma tissue. METHODS: 50 paraffin embedded samples were sectioned and processed following the standard automated immunohistochemistry protocols at Sir Charles Gairdner Hospital (Pathwest). 20 high grade, 20 low grade and 10 normal samples were treated with commercially available antibodies and scored by two trained investigators. RESULTS: Both EMMPRIN and CypA were expressed in normal brain and glioma tissues with different levels of malignancy. The intensively positive expression rates of EMMPRIN in anaplastic astrocytoma and glioblastoma tissues were significantly higher (92% of samples/Mean Allred score – 4.5) than those in normal brain and low-grade astrocytoma tissues (0%/0 and 7.5%/0.9, respectively). No consistent relationship between glioma grade and CypA expression could be established. The positive expression of EMMPRIN was associated with higher grade gliomas. DISCUSSION: Based on these results, we conclude that EMMPRIN is avidly expressed in high grade glioma alone. A correlation between CypA expression and glioma grade could not be established by this study. The detection of EMMPRIN expression is feasible using standard high throughput immunohistochemical techniques and may facilitate diagnosis of high grade glioma where histological differentiation is inconclusive.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

29. INTERSTITIAL RADIOSURGERY FOR CEREBRAL METASTASES

Adonye Banigo ¹, Andrew R Brodbelt ¹, Michael D Jenkinson ¹

Abstract

INTRODUCTION: Brain metastases are a significant cause of morbidity and mortality. Treatment modalities include surgery, stereotactic radiosurgery and whole brain radiotherapy. We investigated the outcome of interstitial radiosurgery with radio-iodine seeds (brachytherapy) for metastatic brain tumours. METHODS: Retrospective analysis of patients treated with interstitial radiosurgery between 2001–2004. Patient demographics, ECOG performance status, Recursive Partitioning Analysis (RPA) class, primary tumour type, size/number of brain metastases, seed dose, duration of implant and overall survival were determined. RESULTS: 19 patients, median age 60.6 years (range: 41–77), were treated. 18 had a single brain metastasis, 1 had two metastases. Median tumour diameter was 15mm (range: 10–30). Four patients had extracranial metastases. 2 patients had an ECOG of 1, the remainder were ECOG 2. Primary tumour types were: lung (n = 10), melanoma (n = 3), breast (n = 2), oesophagus (n = 2), renal (n = 1), colorectal (n = 1). 4 patients were RPA class I, 15 were RPA class II. Radio-iodine125 seeds (median radioactivity 19.5 mCi) were implanted for 14 to 34 days to deliver a reference dose of 60Gy. Complications included seizures (n = 2) and transient arm monoparesis (n = 1). 4 patients had adjuvant WBRT. Median overall survival from date of implant to death was 4.3 months (range: 0.5–47.1 months). There was no statistically significant difference in overall survival according to RPA class or ECOG. CONCLUSION: Interstitial radiosurgery is an alternative local therapy to resection or stereotactic radiosurgery. In our experience, it has a low complication rate, but does not afford a survival advantage compared to other treatment options.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

30. MICRORNA EXPRESSION IN PAEDIATRIC BRAIN TUMOURS

Jennie N Jeyapalan ¹, Muhammad A Mumin ¹, Tim Forshew ¹, Andrew RJ Lawson ¹, Ruth G Tatevossian ¹, Thomas S Jacques ¹, Denise Sheer ¹

Abstract

INTRODUCTION: MicroRNAs are key players in the regulation of gene expression. Aberrant microRNA expression has been identified in a number of cancers but few studies have looked in paediatric brain tumours. The aim of this study is to improve our understanding of the role of miRNAs in paediatric brain tumours. METHODS: MiRNA and mRNA expression were analysed on paediatric brain tumours (n = 60) and normal brain controls (n = 16) using Illumina MicroRNA expression array (v2) and Human HT-12 Beadchips. The Illumina Genome Studio software, Sylarray and Sylamer algorithms (http://www.ebi.ac.uk/enright/sylamer) used for data analysis. RESULTS: MicroRNA profiles were produced for each tumour type. Pilocytic astrocytomas and ependymomas showed the greatest differential expression compared to control brain. MiR-124 was identified as a key player in all tumours. MiR-124 and other miRNAs involved in neuronal differentiation, miR-128, miR-129 and miR-133b were also down-regulated. MiR-216 cluster was up-regulated specifically in medulloblastoma, which has recently been shown to target PTEN. MiR-450a was up-regulated in astrocytomas and miR-31 up-regulated in ependymomas. MiR-146, linked to inflammation and senescence, was up-regulated in pilocytic astrocytomas. Well-characterised microRNAs in adult glioblastomas, miR-92b and miR-10a, plus novel microRNAs were identified in the paediatric glioblastomas. CONCLUSIONS: This study identified aberrantly expressed miRNAs in a range of paediatric brain tumours. These novel findings will be investigated further to elucidate the targets and pathways altered in the tumours.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

31. CHARACTERISATION OF GENOMIC IMBALANCES IN PAEDIATRIC EPENDYMOMA IDENTIFIES A PROGNOSTIC ROLE FOR PRUNE.

JP Kilday ¹, K Wright ¹, S Leavy ¹, J Lowe ¹, EC Schwalbe ¹, SC Clifford ¹, R Gilbertson ¹, B Coyle ¹, RG Grundy ¹

Abstract

INTRODUCTION: Paediatric ependymomas remain a clinical challenge with a relatively poor prognosis. An improved understanding of ependymoma biology may identify new correlates of outcome and potential therapeutic targets. METHODS: Affymetrix 500K SNP arrays were used to identify genomic imbalances in 42 primary and 21 recurrent paediatric ependymomas. Selected gene copy number alterations were validated by quantitative real-time polymerase chain reaction (qPCR). Candidates from the SNP analysis were incorporated into a panel of putative prognostic markers assessed by FISH and immunohistochemistry across a tissue microarray cohort of 107 primary ependymomas treated according to two age defined paediatric clinical trials (CCLG 1992 04 and SIOP 1999 04). RESULTS: Chromosome 1q gain was the most common anomaly in primary and recurrent intracranial ependymomas, while tumours from different locations harboured characteristic aberrations. Unsupervised hierarchical clustering of cytoband anomalies identified a sample group characterised by chromosome 1q21.3 gain which demonstrated a worse five year event-free survival (0 v 56.1%; univariate analysis p = 0.003, multivariate cox regression analysis p = 0.02). Within this region, gain of PRUNE was associated with a reduced five year overall survival when assessed across the primary cohort (65.6 v 88.1%; p = 0.025, multivariate p = 0.045). Immunohistochemistry identified PRUNE overexpression as an independent marker of reduced event-free and overall survival in infant intracranial ependymomas (multivariate p = 0.007). It was also associated with reduced overall survival in older children (univariate p = 0.02). CONCLUSION: This study identifies PRUNE as the first prognostic correlate in uniformly treated paediatric intracranial ependymomas. Confirmation in an independent European infant trial could result in the incorporation of PRUNE into future trial decisions regarding therapeutic stratification of children with this tumour.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

32. MIR-23B TARGETS THE X-LINKED INHIBITOR OF APOPTOSIS GENE IN GLIOBLASTOMA

Paula Kinsella ¹, Martin Clynes ¹, Verena Amberger-Murphy ¹, Niall Barron ¹

Abstract

INTRODUCTION: micro (mi) RNAs are highly conserved noncoding RNAs that control gene expression post-transcriptionally, by degradation of target mRNAs or the inhibition of protein translation. As they control important processes such as differentiation, cell growth and cell death, miRNAs hold great promise for targeted cancer therapy. METHODS: using the high through-put screening method of Taqman Low Density Array analysis (TLDA), we examined the expression of 365 miRNAs in primary glioblastoma cultures. We transiently over-expressed miR-23b in a glioblastoma cell line, SNB-19; following this we examined the effect on proliferation, and the effect on protein expression of the X-linked inhibitor of apoptosis (IAP) by quantitative real time PCR (qRT-PCR) and western blot analysis. RESULTS: we found lower (5.3 fold) expression of miR-23b in glioblastoma cultures with a high proliferation rate, in comparison to cultures with low proliferation rates. By increasing the expression of miR-23b in the established glioma cell line SNB-19, we achieved a 30% decrease in proliferation, with qRT-PCR we found a 65.3% knockdown of XIAP; and using western blot analysis we confirmed XIAP was strongly inhibited by miR-23b at the protein level. CONCLUSION: the ability of cancer cells to resist apoptosis is crucial for development, progression, and treatment resistance. It is possible that a decrease of miR-23b expression in glioblastomas, and subsequent unregulated expression of XIAP, has contributed to the ability of glioblastomas to evade apoptosis. Therefore, miR-23b might be a promising candidate for the development of new targeted treatment options in glioma.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

33. GENE EXPRESSION PROFILING OF PILOCYTIC ASTROCYTOMA

SR Lambert ¹, DTW Jones ¹, D Pearson ¹, I Ichimura ¹, VP Collins ¹

Abstract

INTRODUCTION: Pilocytic astrocytomas (PA) are the most frequent central nervous system tumour in paediatric patients. They are most commonly characterised by RAF fusion genes (with occasional gene mutations) that result in activation of the mitogen activated protein kinase (MAPK) pathway. METHODS: We have used Illumina's HT12 v3 expression microarrays to investigate the genomewide expression profile of 30 PA. For comparison, 3 normal adult cerebella were analysed. Following QC analysis, differentially expressed probes were identified using Bayesian statistics in R. Quantitative real-time PCR was used on a subset of these genes to confirm the microarray findings. RESULTS: In total, 2871 probes were differentially expressed between normal cerebellum and PA using an adjusted p value of <0.001. Gene set enrichment revealed a significant number of genes related to the MAPK pathway, phosphatidylinositol signalling and the ErbB signalling pathway, suggesting these pathways are altered in PA pathogenesis. Furthermore, gene ontology showed enrichment for terms associated with cell death, adhesion and metabolic processes, suggesting these are disrupted in PA. Half of the genes (6/12) that were present on the microarray within the region of common tandem duplication at 7q34 were overexpressed in PA, including HIPK2. Comparison of PAs by location (infratentorial or supratentorial) confirmed previous reports of overexpression of LHX2 and SIX6 in supratentorial PAs. CONCLUSION: We have identified a large number of differentially expressed genes in PA in comparison to normal cerebella. Further characterisation of the pathways that are disrupted in PA may lead to new opportunities for therapeutic intervention.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

34. GLUCOSE-RESPONSIVE MICRORNA NETWORKS ARE ESSENTIAL FOR AGGRESSIVE TUMOUR GROWTH IN GLIOBLASTOMA

Abstract

INTRODUCTION: In order for aggressive tumours such as glioblastoma multiforme to grow rapidly, cells must be able to survive periods of stress such as hypoxia, acidity and nutrient deprivation. We previously identified a pathway in which miR-451 acts as a glucose responsive switch regulating glioma proliferation, migration and survival. METHODS: Microarray analysis of microRNA expression was performed using custom arrays, and validated by qRT-PCR. Target identification was done using bioinformatics followed by qRT-PCR, Western blotting, and analysis of 3'UTR reporters. Cell-based assays were performed using synthetic microRNA mimics, and in vivo studies were carried out using X12 glioma cells stably expressing miR-451/144. RESULTS: We have identified a miR-451 regulated network that mediates responses to glucose alterations in glioblastoma. We also report that miR-451 is co-expressed with miR-144 in patient samples (R2 = 0.718), and that miR-144 may support the miR-451 network. Overexpression of miR-451/144 in X12 patient-derived glioblastoma cells causes unusual tumour growth patterns and completely blocks invasion into brain tissue in vivo, suggesting that fluctuating miR-451/144 levels in glioblastoma in response to glucose are necessary for rapid growth. We have also identified several additional glucose-regulated microRNAs using microarrays. The significance of these alterations and emergence of a glucose-responsive microRNA network in glioblastoma will be discussed. CONCLUSIONS: The dynamic regulation of microRNAs in response to fluctuating glucose levels in the glioblastoma microenvironment is essential for rapid tumour growth and dissemination, and is revealing novel targets for therapeutic intervention.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

35. IS STEREOTACTIC RADIOSURGERY UNDER-UTILISED IN THE TREATMENT OF SURGICALLY EXCISABLE CEREBRAL METASTASES?

Maggie K Lee ¹, Mohsen Javadpour ¹, Michael D Jenkinson ¹

Abstract

INTRODUCTION: The limited availability of stereotactic radiosurgery (SRS) in the UK means surgical resection is the only local treatment option for most patients. Recent capitol investment in new SRS services has increased availability. We determined the proportion of surgically resected metastases that would be suitable targets for SRS. METHODS: Patients undergoing surgery for cerebral metastases between April 2007 and March 2009 were identified. Primary tumour type, adjuvant therapy, complications and length of stay were recorded. Estimated tariffs for elective SRS and craniotomy were obtained. Pre-operative CT/MRI were reviewed to determine tumour size (mm), composition (solid -vs- cystic) and location, presence of hydrocephalus or midline shift and number of metastases. Suitable SRS targets were defined as: <30mm diameter, no hydrocephalus, no symptomatic mass effect, solid tumour. RESULTS: 116 patients (median age 63 years) underwent surgical resection; 86 received adjuvant whole brain radiotherapy (WBRT); 30 did not receive WBRT due to poor performance status. 102 cases (88%) were solitary metastasis; 14 (12%) had multiple metastases. Median tumour diameter was 34mm (range: 12–70mm). 41 cases (35%) were suitable targets for SRS. Size (n = 65), cystic tumour (n = 18) and hydrocephalus (n = 7) alone and in combination were the commonest reasons for lack of suitability. Seven patients (6%) had surgical complications. Median post-operative length of stay was 7 days (range: 2–51). Median overall survival (OS) was 7.7 months for those suitable for SRS and 5.4 months for those not (Log Rank: P = 0.52). The current tariffs for elective SRS and craniotomy are £7777 and £6129 respectively. CONCLUSIONS: One third of surgically excisable brain metastases are suitable targets for SRS. Day case SRS would minimise length of stay and complication rates, but there would be no crude financial benefit. Full health economic analysis warrants further investigation.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

36. CYTOKINE MEASUREMENT IN PRE AND IMMEDIATEVPOSTOPERATIVE SERUM SAMPLES OF HIGH-GRADE GLIOMA PATIENTS USING LUMINEX TECHNOLOGY

E Lekka ¹, P Abel ¹, T Dawson ¹, B Lea ¹, C Davis ¹

Abstract

INTRODUCTION: There is a growing interest in the potential use of cytokine measurement as a biomarker in serum samples of patients with Grade 4 Gliomas. This study aimed to investigate the effect of surgery on cytokine levels perioperatively. MATERIALS AND METHODS: We prospectively collected serum samples immediately preoperatively and within 24 hours after surgery, from 36 patients who underwent biopsy or debulking of tumour. Control serum samples were collected pre and immediately postoperatively from 36 patients with no known history of malignancy, who underwent elective surgery for benign pathology. The control samples were pooled according to age. We evaluated the levels of 8 cytokines using the Bioplex analyser supplied by BioRad. RESULTS: There were significant differences between pre and postoperative levels for three of the eight cytokines IL-6, IL-8, IL-10 (p < 0.05) measured when compared to the control samples. There was no correlation between the cytokine levels and the extent of surgery, sex or survival. CONCLUSIONS: These results suggest that cytokine levels fluctuate throughout the disease process and in the immediate postoperative period. Further studies should examine the cytokine levels in samples taken during the course of the disease at regular intervals.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

37. EFFECTIVENESS AND OUTCOMES OF SURGERY FOR CEREBRAL METASTASES

Christina Sae-Kyung Lim ¹, Paul L Grundy ¹

Abstract

INTRODUCTION: Recently, local commissioners have questioned the effectiveness of surgery for cerebral metastases, attempting to discontinue this service. Two RCTs had reported a survival benefit of surgery but a further trial demonstrated no benefit. This study aims to assess the effectiveness of surgery for cerebral metastases in current practice. METHODS: A retrospective review was performed for a single surgeon series of consecutive patients undergoing primary surgery for cerebral metastases between June 2005 and April 2010. The main outcome measure was the survival time after surgery. RESULTS: 123 patients (61 males, 62 females) were identified with a mean age of 58.4 years. 83 patients (67%) were under 65 years. The overall 30-day mortality rate was 2.4%. The overall median survival was 10 months. There were 26 (21%) breast cancers with median survival of 405 days, 32 (26%) NSCLC with 249 days, 24 (19%) melanomas with 201 days, 13 (11%) colorectal cancers with 193 days, and 11 (9%) renal cell cancers with 409 days. The differences were not significant (p > 0.05). However, when the breast cancer group was compared to the NSCLC group, the difference was significant (p = 0.005). The median survival differences were not significant (p > 0.05) with regards to the RPA class, the site (supratentorial or infratentorial) and the number of metastases (single or two). CONCLUSIONS: Median survival in this cohort was identical to those in the 2 RCTs that showed survival benefits from surgery. This was significantly longer than that (5.6 months) in the single series demonstrating no benefit. Our results support the previous evidence of improved outcomes with surgery and form a basis for informed discussion with local commissioners.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

38. EPENDYMOMA: PREVALENCE, TREATMENT AND MANAGEMENT IN NHS TAYSIDE

M Pendleton ¹, HK Lord ¹

Abstract

INTRODUCTION: There is a paucity of published data on treatment and patient outcomes for ependymoma. We present a retrospective audit of prevalence, treatment and outcomes of patients diagnosed with ependymoma in NHS Tayside in 13 year period. METHODS: Cases identified from Scottish Cancer Registry. Age, gender, site of disease, grade of pathology, degree of surgery, post operative radiotherapy, date of progression, second line treatment and survival recorded. RESULTS: Nine patients diagnosed with ependymoma between 1997 and 2010, 7 adults and 2 paediatric cases. The latter were excluded. Age range 19–64, median 38. Three WHO Grade 1, 4 Grade 2 and none Grade 3. Six localised, one metastatic. Two occurred in cervical - thoracic spine; 2 in thoraco-lumbar spine, 1 in frontal lobe distant from lateral ventricle; 1 involved left lateral ventricle and one 4th ventricle with deposits in cervico- thoracic and lumbo-sacral spine. All patients treated with surgery. All received post-operative radiotherapy due to residual disease. None received post operative chemotherapy. Radiotherapy delivered to local tumour site in 6 patients, to cranial spinal axis in 1. 3 patients relapsed, all at site of original disease and in one spread to the third ventricle. All treated with further de-bulking surgery. One received intra-operative radiation. None received chemotherapy. Time to relapse 7 months, 11 months, and 5 years, calculated from date of treatment finishing to date of follow up MRI demonstrating relapse. Survival 100%. CONCLUSION: Low incidence and good survival with surgical resection and post operative radiotherapy, but significant recurrence in Tayside population. Notes gave limited information on patient quality of life (QoL). Larger prospective studies essential. Treatment at relapse to be better defined. Will introduce patient QoL scoring.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

39. RADICAL CHEMORADIATION IN ELDERLY PATIENTS WITH GLIOBLASTOMA: QUALITY OF LIFE (QOL) VERSUS SURVIVAL BENEFIT

Mairi Mackinnon ¹, Aoife Williamson ¹, Allan James ¹, Willie Stewart ¹, Brian Clark ¹, Anthony Chalmers ¹

Abstract

BACKGROUND: Subgroup analysis in the original Stupp study indicated that patients aged >60 derived least benefit from intensive multimodal therapy. The aim of this study was to investigate the impact of this treatment on QoL. METHODS: Medical records were examined retrospectively. WHO PS was extracted at 3 timepoints: (1) 1st appointment with oncologist, (2) at start of adjuvant chemotherapy and (3) cycle 6 of adjuvant treatment. Survival data were calculated from date of surgery by Kaplan-Meier method using SPSS version 16. RESULTS: Of the 130 patients, 40 were > 60 years. 10 patients did not receive adjuvant treatment, 14 completed 1–3 cycles, 9 completed 4–5 cycles and 7 completed adjuvant treatment. Median survival was 12.3 months. One year overall survival (OS) was 55.1%. WHO PS data at the timepoints were available for 40, 40 and 7 patients respectively: (1) 20 pts PS 0, 20 pts PS 1; (2) 10 pts PS 0, 29 pts PS 1, 1 pt PS 2; (3) 2 pts PS 0, 3 pts PS 1, 2 pts PS 2. CONCLUSIONS: The majority of patients' PS dropped by 1 point during treatment, although we cannot exclude more significant drops in PS causing cessation of therapy. These preliminary data indicate that, with appropriate patient selection and careful on treatment review, radical chemoradiation can be delivered to patients aged 60–70 with acceptable deterioration in PS. Future studies will prospectively assess QoL and duration of stable/responding disease after completion of adjuvant treatment.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

40. BMI-1 GENE SILENCING INFLUENCES CELL ADHESION AND INVASIVE PROPERTIES OF MEDULLOBLASTOMA CELLS VIA INHIBITION OF BMP PATHWAY

A Merve ¹, X Zhang ¹, S Marino ¹

Abstract

INTRODUCTION: Medulloblastoma (MB) is the commonest malignant childhood CNS tumour with a high metastatic potential. We have recently demonstrated that during cerebellar development the polycomb group gene Bmi-1 regulates cell adhesion properties through repression of BMP pathway. BMPs are known to inhibit MB growth in mouse models. Here we assess whether overexpression of Bmi-1 in MB may contribute to tumour growth by modulation of cell adhesion. METHODS: 2 MB cell lines DAOY and D458 known to overexpress Bmi-1 were used and Bmi-1 was downregulated using Short interfering RNA (SiRNA). Immunolabelling for phosphorylated SMAD proteins (pSMADs) was carried. Traswell migration assays (TW) and time lapse experiments (TL) were performed to assess the invasive properties. The results were compared with the scrambled SiRNA treated cells (controls). Statistical analysis was performed using student t-test. Immunohistochemical staining (IHC) for cell adhesion molecules were performed on cell blocks. RESULTS: We observed an increase labelling index for pSMAD among the Bmi-1 downregulated cells compared to controls – 72.85% vs 57.51% in DAOY and 83.93% vs 75.69% in D458. Interestingly we also found an increased number of multicellular aggregates (average/field) - 2.36 vs 0.55 in DAOY and 4.78 vs 1.61 in D458. Upon concomitant BMP pathway inhibition with Noggin, there was phenotypic rescue noted. The TW assay showed a lower number of cell migration (average/field) – 222 vs 147 in DAOY. The TL experiment demonstrated a decreased motility (average distance in μm) of Bmi-1 downregulated cells – 2.69 vs 2.07 in DAOY and 2.01 vs 1.65 in D458. Finally IHC revealed a differential expression of CD44 and Thrombospondin in Bmi-1 silenced D458 compared to controls. CONCLUSIONS: Our data support the notion that Bmi-1 overexpression contributes to aggressiveness of medulloblastoma probably by repression of BMP pathway.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

41. GENOME WIDE MOLECULAR CHARACTERISATION OF CENTRAL NERVOUS SYSTEM PRIMITIVE NEUROECTODERMAL TUMOUR AND PINEOBLASTOMA

Suzanne Miller ¹, Hazel A Rogers ¹, Paul Lyon ¹, Vikki Rand ¹, Martyna Adamowicz-Brice ¹, Steven C Clifford ¹, James T Hayden ¹, Sara Dyer ¹, Stefan Pfister ¹, Andrey Korshunov ¹, Marie-Anne Brundler ¹, James Lowe ¹, Beth Coyle ¹, Richard G Grundy ¹

Abstract

INTRODUCTION: CNS PNET and pineoblastoma are highly malignant embryonal brain tumours of poor prognosis. Current therapies are based on the treatment of paediatric medulloblastoma, even though these tumours are distinct at both the anatomical and molecular level. CNS PNET and pineoblastoma have a worse clinical outcome than medulloblastoma, thus improved therapies based on an understanding of the underlying biology of CNS PNET and pineoblastoma are needed. METHODS: Affymetrix SNP arrays were used to characterise the genomic alterations of 36 paediatric CNS PNETs and 8 pineoblastomas. Gene copy number alterations were validated using qPCR. RESULTS: Overall the majority of CNS PNETs contained a greater degree of genomic imbalance than pineoblastomas. Novel gene copy number alterations were identified; gain of PCDHGA3, 5q31.3 in 62.1% primary CNS PNETs and all primary pineoblastomas and FAM129A, 1q25 in 55.2% primary CNS PNETs and 50% primary pineoblastomas. Utilizing the collection of 5 primary and recurrent CNS PNET pairs, we found gain of 2p21 was maintained at relapse in 80% of cases. Novel gene copy number losses included OR4C12, 11p11.12 in 48.2% of primary CNS PNETs and 50% primary pineoblastomas. Loss of CDKN2A/B (9p21.3) was identified in 14% primary CNS PNETs and significantly associated with older children (p = 0.05). CADPS, 3p14.2 was lost in 27.6% primary CNS PNETs and was associated with poor prognosis (p = 0.043). CONCLUSION: This genome-wide analysis has revealed the marked molecular heterogeneity of CNS PNETs and enabled the identification of novel genes and clinical associations potentially involved in the pathogenesis of these tumours.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

42. QUALITY OF LIFE AFTER TREATMENT FOR BRAIN METASTASES: INTERIM DATA FROM THE MRC QUARTZ CLINICAL TRIAL.

Matthew Nankivell ¹, Paula Mulvenna ¹, Rachael Barton ¹, Paula Wilson ¹, Corinne Faivre-Finn ¹, Cheryl Pugh ¹, Ruth Langley ¹

Abstract

INTRODUCTION: QUARTZ addresses a longstanding clinical question: what is the role of whole brain radiotherapy (WBRT) in patients with non-small cell lung cancer (NSCLC) and inoperable brain metastases? All patients receive steroids and optimal supportive care (OSC), and are randomised to WBRT or not. The primary endpoint is quality adjusted life years (QALYS), reflecting the importance of quality of life as well as survival in this very poor prognosis patient group. It is a non-inferiority trial, with a non-inferiority margin of 1 week in QALYS seen as clinically important. QUARTZ requires 534 patients and began recruiting in March 2007. Despite widespread support from 77 UK and Australian centres, recruitment was lower than anticipated and the trial was under threat of closure. An issue contributing to the poor recruitment was thought to be the lack of preliminary data regarding the omission of WBRT. METHODS: Permission to release data on the first 151 QUARTZ patients was given by the Trial Steering Committee, who were unaware of the results at the time. Data were analysed and presented to participating centres. RESULTS: 60% of patients were male; the median age was 67 (range 38–85); 50% had good performance status (KPS >70) and 39% had a solitary brain metastasis on CT or MRI scan. The average QALY was 30 days in the OSC alone group, and 31 days in the OSC + WBRT group. Median survival was 7.3 weeks for patients receiving OSC alone, compared to 7.0 weeks for those receiving OSC + WBRT. Treatment with WBRT did not appear to have an obvious effect on QoL. CONCLUSIONS: These data provide preliminary evidence that omitting WBRT is not detrimental to this group of patients and can be used as a base for trial decisions and discussion. They are not definitive results but provide a strong rationale for the trial continuing, to answer this longstanding and important clinical question.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

43. INTRA-OPERATIVE DETERMINATION OF NADH REDOX POTENTIAL IN PATIENTS WITH INTRACRANIAL TUMOURS

D Ngoga ¹, D Tennant ¹, A Williams ¹, P Moss ¹, G Cruickshank ¹

Abstract

INTRODUCTION: There has been a resurgence in interest in the “Warburg effect”; cellular metabolic adaptations that result in a reliance on accelerated glycolysis in the presence of abundant oxygen, first described in 1924. Recent studies on isocitrate dehydrogenase 1 (IDH1) gene mutations, have highlighted the importance of a balanced redox potential (the balance between oxidized and reduced cellular metabolites) to the development and behaviour of cancers. Nicotinamide adenine dinucleotide (NAD+) and NAD phosphate (NADP+) are involved in numerous cellular redox reactions, the failure of which is associated with the growth, chemoresistance and invasion of malignant cells In this study, we examine NAD(P)H redox potentials in a number of CNS tumours correlating these with histological diagnosis and grade with comparison made with normal tissue. METHODS AND RESULTS: Patients with different intracranial tumours requiring surgical resection were recruited. Intra-operative tumour samples were analysed to establish their NAD(P)H redox status using a novel combined gated pulse fluorescent excitation and nanosecond time-resolved fiberoptic fluorescence detector. Samples of normal (muscle) tissue were also obtained for comparison. Samples were then sent for standard histological analysis. Preliminary data from this ongoing study has confirmed proof of principle with SD values at reasonable levels and correlation of NAD(P)H redox status and histological diagnosis and grade. DISCUSSION: There are suggestions that cytosolic NADH imbalance might be a sensitive controller of nuclear transcription. The role of IDH1 mutation on NAD(P)H redox status and the contribution of this to improved prognosis for low and high grade glioma is still not fully understood. The ability to asses NADH in tissues is a crucial step in advancing our understanding of tumour metabolism, offering potential biomarker, intra-operative prognostic information potentially new redox targeted therapies.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

44. ESTIMATES OF REGIONAL CNS TUMOUR INCIDENCE: A COMPARISON BETWEEN HOSPITAL-BASED DATA AND THE NATIONAL CANCER REGISTRY ESTIMATES FOR 2009

K Owusu-Agyemang ¹, S Bell ¹, W Stewart ¹, J StGeorge ¹

Abstract

INTRODUCTION: The National Cancer Intelligence Network (NCIN) is developing data collection methods to support a national brain tumour registry. The accuracy of such datasets in recording tumour incidence, and hence the optimum method of data capture, remains unclear. The neuropathology and MDT databases, two sources of data for a potential hospital-based cancer registry (HBCR), were interrogated and compared to national, centrally recorded data. METHODS: New tumours of the brain, meninges and other CNS sites encountered by the neuro-oncology service during 2009 were identified in the respective databases of the department of neuropathology and the neuro-oncology MDT; these were combined to form a HBCR. The estimated tumour incidence for the equivalent period was calculated from the National Cancer Registry (NCR) data for the preceding three years (2006–2008). The data derived from these sources were then assessed and compared. RESULTS: 364 cases of primary (79%) and secondary tumours were captured by the HBCR. The 3-year average data published by the NCR estimated 374 new cases of primary tumours versus the 289 in the HBCR. Similar proportions of primary tumours were recorded as high grade (WHO grade III or IV) in the HBCR and NCR. Notification patterns varied according to age at diagnosis, diagnostic group and WHO grade/ICD-10 code. An inverse male-to-female ratio was observed between the HBCR and NCR datasets (1.2 v 0.8; respectively). CONCLUSION: Previous studies suggest regional cancer registries may underestimate the incidence of CNS tumours. However, our results suggest a considerable number of new cases are not captured by the data being gathered through existing hospital datasets.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

45. FLUORESCENCE GUIDED RESECTION IDENTIFIES DISTINCT COMPARTMENTS OF TUMOUR-INITIATING CELLS IN HUMAN GLIOBLASTOMA

Sara GM Piccirillo ¹, Colin Watts ¹

Abstract

INTRODUCTION: Recent evidences in human glioblastoma (GBM) suggest that distinct areas of the same tumour contain “cancer stem cells” (CSCs) endowed with different tumorigenicity. Cells in the tumour margin are supposed to be responsible for the recurrences but the investigation of this area has been carried out based on standard surgical resection. This approach is not objective and is hampered by the infiltrative nature of tumour cells. As a result, there is no consensus on how it is possible to interrogate the tumour edges in GBM. METHODS: Primary culture, culture propagation, cell line establishment, multiple immunofluorescence and in vivo experiments were performed as described by Piccirillo et al., 2009 and according to the Cambridge protocol. RESULTS: Here we show that different regions of the same GBM can be objectively identified by the use of 5-aminolevulinic acid. This technique objectively discriminates distinct compartments (mass, necrotic and margin) and does not alter the 90% efficiency of derivation of CSCs from GBM. Of note, only the cells derived from the mass give rise to multipotent long-term expanding CSCs. Margin cells are tumorigenic in vivo but in vitro do not grow under stem cell conditions nor possess the stem cell molecular signature. CONCLUSIONS: Our findings suggest that margin cells are tumorigenic but do not fulfil the criteria of CSCs. Further experiments are ongoing to better characterize the phenotype and genotype of margin cells versus cells derived from the mass and to evaluate response to conventional therapies. Histological analysis will be used to characterize disease in non fluorescent margin.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

46. THE ROLE OF SERIAL STEREOTACTIC BIOPSY IN ADULTS WITH BRAINSTEM LESIONS

SRM Qadri ¹, E Pirola ¹, MD Jenkinson ¹, A Brodbelt ¹

Abstract

INTRODUCTION: Brain stem tumours are rare in adults and are associated with a poor prognosis. Histological diagnosis is required before administration of adjuvant therapy. We investigated the role of stereotactic biopsy in the management of brainstem lesions. METHODS: Retrospective analysis of adult patients with brainstem lesions between 2001–2010. Frame-based serial stereotactic biopsy was used in all cases. Histological diagnosis, adjuvant therapy and clinical outcome were investigated. RESULTS: Twenty-eight patients (13 female; 15 male; age range: 20–76 years) were biopsied. ECOG performance status at presentation was 0–1 (n = 17) and 2 (n = 11). Histopathology diagnoses included: grade II astrocytoma (n = 11), high grade glioma (n = 5), lymphoma (n = 5), pilocytic astrocytoma (n = 2), metastasis (n = 2), non-diagnostic (n = 3) . Median survival in grade II astrocytoma was 91% at one year, 68% at 2 years and 55% at 5 years. Adjuvant therapy included radio-iodine seed implant (n = 3), fractionated radiotherapy (n = 6) and temozolomide (n = 1). Seven patients are still alive. Patients with anaplastic astrocytoma (n = 4) received radiotherapy; there was a 76% median survival at 6 months and 50% at 1 year. In lymphoma cases (n = 5) the median survival was 40% at 1 year following radiotherapy +/− methotrexate. Median survival for brainstem metastases was 3.5 months. Three patients had non-diagnostic biopsies – one patient subsequently developed metastatic breast cancer; two patients showed lesion resolution suggestive of infarct. One patient (3.5%) had worsening deficit following biopsy. CONCLUSIONS: Brainstem lesions in adults are a heterogeneous group. Stereotactic biopsy has a low complication rate and is accurate. Histological diagnosis is important for guiding adjuvant therapy and determining prognosis. Low grade brainstem gliomas have an indolent course and a good prognosis following adjuvant radiotherapy.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

47. EVALUATING NOVEL POLYMERIC MICROPARTICLE-BASED INJECTABLE MATRICES FOR LOCAL CHEMOTHERAPEUTIC DELIVERY

R Rahman ¹, C Rahman ¹, S Smith ¹, D MacArthur ¹, F Rose ¹, K Shakesheff ¹, RG Grundy ¹

Abstract

INTRODUCTION: We have investigated a novel formulation based on poly(lactic-co-glycolic acid) (PLGA; FDA-approved) and poly(ethylene glycol) (PEG) blended microparticles for local chemotherapeutic drug release. The biodegradable microparticles inject at room temperature and solidify into a matrix upon delivery into the brain, potentially allowing persistent tissue contact in the resection cavity lining and sustained multiple drug release. METHODS: Viability/growth rate of paediatric brain tumour cells cultured on matrices made from PLGA/PEG microparticles was assessed (Live/dead staining; SEM; alamar blue proliferation assay). UV absorbance was used to determine single/multiple release of etoposide, methotrexate and the histone deacetylase inhibitor trichostatin A (TSA) from matrices. Cytotoxic response from cells cultured on drug-loaded matrices was evaluated using alamar blue assay. RESULTS: Brain tumour cells were biocompatible with PLGA/PEG matrices, with both the apical and basal surface of cells amenable to drug penetration. Sustained TSA and etoposide release over 3 weeks from PLGA/PEG matrices was achieved by diffusion kinetics and followed near zero-order kinetics after an initial burst. Release profiles were similar for matrices loaded with high and low concentrations of drug. CONCLUSIONS: No adverse toxicity from the PLGA/PEG co-polymer alone was observed and matrices permitted sustained drug release in vitro. Combinatorial release of TSA, etoposide and methotrexate from a single formulation will be discussed. In addition, ongoing studies measuring the cytotoxic response of brain tumour cells cultured on drug-loaded matrices will be described. The PLGA/PEG, injectable, self-assembling drug delivery system will be widely applicable for brain tumours for which complete surgical resection is not achievable.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

48. PREVALENCE, PREDICTORS AND ASSOCIATIONS OF APATHY IN ADULT SURVIVORS OF AN EARLY CHILDHOOD POSTERIOR FOSSA BRAIN TUMOUR

Cliodhna Carroll ¹, Peter Watson ¹, Mike Hawkins ¹, Helen Spoudeas ¹, David Walker ¹, Tony Holland ¹, Howard Ring ¹

Abstract

INTRODUCTION: Apathy is a disorder of diminished motivation defined as a deficiency in behavioural, emotional and cognitive components of goal-directed behaviour. It occurs in several neurological pathologies and is associated with pervasive and disadvantageous effects on daily life. In this study we examined prevalence, predictors and associations of apathy in adult survivors of a childhood posterior fossa tumour. METHODS: 118 adult survivors of early childhood (diagnosed before 5 years of age) posterior fossa tumours, and 62 of their siblings, were assessed an average of 32 years after initial tumour treatment, using the Weschler Abbreviated Scale of Intelligence, the Marin apathy evaluation scale and the Composite International Diagnostic Interview. RESULTS: Apathy scores reached or exceeded clinical cut-off in 35% of survivors compared to 18% of the comparison group. In the survivors this was associated with decreased employment, decreased income and lower self-ratings of physical and mental health. Apathy scores at or above cut-off were associated with lower verbal IQ scores and with a current or previous psychiatric diagnosis, but not with age at tumour treatment (all were < 5 years), duration of follow-up, history of radiotherapy or tumour type (astrocytoma or medulloblastoma). Clinically significant apathy in the survivors was not associated with a diagnosis of depression or with performance IQ measures. CONCLUSIONS: Clinically significant apathy occurs relatively often in adult survivors of childhood brain tumours and is associated with impaired social functioning and increased psychopathology. Further research should determine whether any tumour or treatment-related variables increase risk of apathy.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

49. THE FREQUENCY, CLINICAL ASSOCIATIONS AND LONGITUDINAL COURSE OF MAJOR DEPRESSIVE DISORDER IN ADULTS WITH CEREBRAL GLIOMA

AG Rooney ¹, S McNamara ¹, M Mackinnon ¹, M Fraser ¹, R Rampling ¹, A Carson ¹, R Grant ¹

Abstract

INTRODUCTION: Currently there is little high-quality evidence as to (1) the frequency, (2) independent clinical associations, and (3) the longitudinal course of DSM-IV Major Depressive Disorder (MDD) in adults with primary cerebral glioma. METHODS: This was a twin-centre, prospective, observational cohort study with six-month follow-up. Adults with newly diagnosed cerebral glioma received the Structured Clinical Interview for DSM-IV (SCID) to diagnose MDD. Interviews occurred shortly after the start of radiotherapy (T1), three months later (T2) and six months later (T3). Independent associations between MDD and clinical variables were analysed using logistic regression. RESULTS: At T1 n = 155, at T2 n = 108, and at T3 n = 88. The frequency of MDD at T1 was 13.5% ± 5.4%; at T2 was 14.8% ± 6.7%; and at T3 was 6.8% ± 5.3%. Overall, 32 individuals were diagnosed with MDD during the study period (20.6% ± 6.4%). Inter-rater diagnostic agreement for MDD was good (κ = 0.81, 95% CI 0.60–1.00). Clinical variables which independently predicted the development of MDD were functional impairment (OR = 3.6, 95% CI 1.4–9.4) and a previous history of depression (OR = 3.8, 95% CI 1.5–9.8). In patients who could be followed up, MDD usually persisted for at least three months. CONCLUSIONS: To date, this is the largest study of MDD in a consecutively presenting cohort of glioma patients. Although most did not become depressed, MDD was a clinically significant complication of glioma, affecting up to one in five patients during primary treatment. Patients with severe functional impairment or previous depression are at higher risk and may benefit from closer follow-up. Depression in glioma can last for months, and clinicians should actively seek and treat it.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

50. OLDER PATIENTS WITH GLIOMA HAVE A HIGH FREQUENCY OF A RARE CDKN2A P14/P16 ALLELE

JA Royds ¹, S Al Nadaf ¹, A Ahn ¹, Y-J Chen ¹, A Wiles ¹, D Jellinek ¹, AW Braithwaite ¹, BC Baguley ¹, MR MacFarlane ¹, NA Hung ¹, TL Slatter ¹

Abstract

INTRODUCTION: Immortal cells are thought to require a telomere maintenance mechanism (TMM) to prevent critical shortening during repeated cell cycles. However, we have shown that a significant fraction of high-grade gliomas (HGG) do not have either elevated telomerase or evidence of the Alternate Lengthening of Telomeres (ALT). The TMM in the latter is referred to as “none”. We now wish to determine the underlying genotype for the cyclin-dependent kinase inhibitor 2A (CDKN2A) rs11515 (C500G) polymorphism in HGG, its relation to TMM and how it affects patient outcome. METHODS: CDKN2A 3' UTR SNP, C > G 500, was analysed in blood leucocyte DNA from 107 patients with HGG along with 150 normal controls. TMM and CDKN2A gene dosage were determined for the HGGs. The results were compared with patient outcomes. RESULTS: Of the 107 GBMs, 16% were ALT, 47% telomerase and 37% “none”. 19 of the 107 were heterozygous and 2 homozygous for G500. The G500 allele had an increased frequency in non telomerase-ALT tumors (0.29) compared to the general population (0.13, p = 0.001) or to patients with telomerase (0.12) and ALT (0.09) positive tumors. In the non telomerase-ALT tumor group, carriers of the G500 allele were associated with poorer survival (median survival 3 versus 9 months, p = 0.004), CDKN2A loss (p = 0.02) and old age (p = 0.04) compared to patients who were homozygous for C500. CONCLUSIONS: The G500 CDKN2A allele, which increased the likelihood of p16INK4a/p14ARF loss and poorer survival, is a risk factor associated with tumourigenesis in older patients with non telomerase-ALT HGG. Screening for the G500 allele may aid prognosis.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

51. PREDICTING OUTCOMES OF VOCATIONAL REHABILITATION IN PATIENTS WITH BRAIN TUMOURS: A PILOT STUDY

SL Rusbridge ¹, NC Walmsley ¹, SB Griffiths ¹, PA Wilford ¹, JH Rees ¹

Abstract

INTRODUCTION: The multi-disciplinary Vocational Rehabilitation (VR) Service for patients with brain tumours was funded for one year by Macmillan Cancer Support to provide assistance to patients of working age to return to work, remain in work, or decide to stop work. The impact of VR on work rates and quality of life (QoL) in patients with brain tumours has not been systematically examined to date. The purpose of the current study was to measure the outcome of VR and to create a predictive model of vocational outcomes. METHODS: To date, 39 patients have been seen in the service. Of these, 11 have been discharged with pre- and post-intervention data collected. 8 patients were diagnosed with low grade glioma, 1 with high grade glioma, and 2 with brain metastases. Outcomes examined included work status at entry, hours of work and occupational level. In addition, measures of QoL, fatigue, and mood were assessed. The predictive validity of demographic variables, tumour and treatment details, functional ability, and VR interventions were examined for vocational outcome. RESULTS: Preliminary findings are presented. All 11 patients were in work pre-morbidly and 6 patients were in work at referral (46%). Following intervention, 8 patients were in work (67%) and 3 patients were not working (33%). Paired t-tests revealed no significant differences in QoL, fatigue and mood following intervention. CONCLUSIONS: VR may be useful in assisting patients with brain tumours to return to or remain in work. Due to limited sample size, additional data are being collected to establish further VR and QoL outcomes and to create a predictive model of VR outcome, to be presented at BNOS 2011.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

52. HUMAN INDUCED PLURIPOTENT STEM (IPS) CELL DERIVED NEURAL STEM CELLS IN MALIGNANT GLIOMA. AN AUTOLOGOUS TROJAN HORSE IN THERAPEUTICS DELIVERY

D Ryan ¹, C Watts ¹, P Liu ¹

Abstract

INTRODUCTION: A theoretical panacea in neuro-oncology would be the ability to selectively target the tumour population and leave the normal brain uninjured. Neural stem cells (NSCs) have innate tumour tropism and not only migrate to the tumor mass, but selectively ‘track down’ microsatellite deposits often quite distant from the primary focus. Using unique induced pluripotent stem cell technology we sought to investigate whether iPS cell derived neural stem cells possessed this property, representing a potentially autologous therapeutics vector in our armamentarium against malignant glioma. METHODS: Following cellular reprogramming, generated Sanger Human induced Pluripotent Stem cells (SH-iPS cells) were successfully differentiated to neural stem cells using mouse monolayer differentiation. Tumour tropism was assessed using a transmigration assay towards conditioned media from glioma. Using PKH26 fluorescent labeling we sought to investigate whether iPS derived NSCs would migrate towards the individual tumour cell. RESULTS: Quantitative assessment of iPS cell derived NSCs showed superior tumor tropism capabilities relative to the negative control (fibroblasts) at 300% migration efficiency as a percentage of the control (100%). PKH26 fluorescent labeling demonstrated that iPS NSCs migrated to the level of the individual glioma cell and relative to the current gold standard, fetal neural stem cells, derived NSCs demonstrated a higher efficacy of migration. CONCLUSIONS: Our preliminary proof of concept data has demonstrated that iPS cell derived NSCs have inherent tumour tropism and represent a potentially powerful autologous therapeutics vector in malignant glioma.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

53. THE METABOLIC AUTOPHAGY PATHWAY REGULATES MIGRATION AND INVASION IN GLIOMA

Sara Galavotti ¹, Maya Shaked-Rabi ¹, Eugene Tulchinsky ¹, Sebastian Brandner ¹, Chris Jones ¹, Paolo Salomoni ¹

Abstract

INTRODUCTION: High-grade gliomas (HGG) remain among the poorest prognosis primary tumours of adults. One of the factors underlying such dismal prognosis is the ability of glioma cells to invade the brain parenchyma, which makes surgical debulking almost invariably insufficient. Glioma is highly heterogeneous and can be classified into molecular subclasses. Among them, the mesenchymal subtype is characterised by mesenchymal/angiogenic markers and is associated with poor survival. The metabolic autophagy pathway has been proposed to contribute to disease progression in aggressive cancers. However, its precise role in glioma has not been fully elucidated. METHODS: We analysed autophagy status in glioma by using publicly available gene expression datasets from over 400 patient samples. Next, we studied autophagy in glioma-initiating cells and determined the impact of its suppression on growth, migration/invasion and bioenergetics. Finally, we investigated the involvement of RAS/MAPK as an upstream regulator of autophagy. RESULTS: We found that a number of autophagy genes, such as DRAM1 and p62 are highly expressed in glioma tumours belonging to the mesenchymal subclass. Patients expressing high levels of the autophagy inducer DRAM1 have poor prognosis. Autophagy appears to be under the control of the RAS/MAPK pathway in glioma. In glioma-initiating cells, suppression of autophagy impairs migration/invasion without overt effects on survival. Autophagy is induced in cells undergoing epithelial-to-mesenchymal transition (EMT) and regulates EMT-triggered migration. Finally, autophagy suppression impinges on mitochondrial respiration. CONCLUSIONS: Taken together, these findings shed new light on the role of autophagy in cancer and propose a new role of this intracellular degradation pathway in the control of migration/invasion.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

54. ISOGENIC GLIOMA STEM CELL LINES WITH DIFFERENT EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR) AMPLIFICATION STATUS FROM THE SAME TUMOUR

Alexander Schulte ¹, Hauke S Günther ¹, Svenja Zapf ¹, Sabine Riethdorf ¹, Manfred Westphal ¹, Katrin Lamszus ¹

Abstract

INTRODUCTION: EGFR amplification is present in almost 50% of all glioblastomas (GBM) and is frequently associated with expression of a truncated, constitutively active variant, EGFRvIII. Experimental limitations to study these alterations exist as both are rapidly lost when GBMs are taken into culture. We developed conditions which facilitate the growth of cell lines that either maintain high-level EGFR amplification/EGFRvIII expression or not, allowing direct comparison between cells with a heterogeneous EGFR status derived from the same original tumour. METHODS: Fresh tumour material was cultured using glioma stem cell conditions with modifications. The resulting matched pairs of cell lines and original tumours were analyzed by RT-PCR, Western Blot, FACS and FISH for different levels of EGFR gene amplification, EGFRvIII expression, EGFR/EGFRvIII protein, and in vivo tumorigenicity. RESULTS: From 2 freshly resected GBM pairs of cell lines were generated that differed in (i) EGFR gene amplification levels, (ii) expression of EGFRvIII, and (iii) levels of EGFR/EGFRvIII protein. The first pair consisted of one cell line with high-level EGFR amplification and high EGFR protein expression, whereas the isogenic sister-line lacked EGFR amplification. The second pair comprised one line showing EGFRvIII expression, EGFR amplification and high EGFRvIII/EGFR protein levels, whereas these alterations were absent in the sister-line. Nude mice injected with EGFR-amplified cell lines died significantly earlier than those xenografted with non-amplified lines, and the EGFR/EGFRvIII status was stably maintained in vivo. Xenograft tumors resembled their parental primary tumours on protein as well as on genomic levels. CONCLUSIONS: Our cell lines provide a model to study the function of EGFR amplification/EGFRvIII expression in glioma cells and assess the impact of intratumoral EGFR status heterogeneity on the response to EGFR-targeting and other agents.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

55. OUTCOME AND PROGNOSTIC FEATURES IN ADULT PINEOBLASTOMAS: ANALYSIS OF CASES FROM THE SEER DATABASE

Senthil K Selvanathan ¹, Salah Hammouche ¹, Heidi J Salminen ¹, Michael D Jenkinson ¹

Abstract

INTRODUCTION: Adult pineoblastomas are rare central nervous system tumours. Patient and treatment factors associated with outcome are poorly defined and limited to small retrospective case series and single case reports. Using the Surveillance, Epidemiology, and End Results (SEER) cancer registry, we investigated clinicopathological factors associated with outcome in adult pineoblastomas. METHODS: Adult patients (≥16 years old) with pineoblastomas diagnosed between 1990–2007 were identified from the SEER database. Kaplan-Meier survival analysis and Cox models were used to examine the effect of variables on overall survival. The variables analysed included patient's age at diagnosis, gender, race, tumour location, uni-focal or multi-focal tumour, tumour size, surgical resection and the use of adjuvant radiotherapy. RESULTS: Ninety-five patients were identified, with a median age at diagnosis of 39.2 years. Sixty-one patients (64%) underwent surgery and forty-four patients (44%) received adjuvant radiotherapy. Forty-two patients (44%) had both surgery and radiotherapy. The median overall survival was 176 months. Univariate analysis identified younger age at diagnosis, uni-focal and localised disease as important predictors of overall survival. On multivariate analysis, only age at diagnosis and localised disease emerged as important prognostic factors. CONCLUSIONS: This study represents the largest analysis of adult pineoblastomas to date. Clinically relevant prognostic factors were younger age of diagnosis and localised disease. Surgery and adjuvant radiotherapy did not influence overall survival.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

56. GOLD NANOPARTICLE MEDIATED RADIOSENSITIZATION WITH CONCOMITANT TEMOZOLOMIDE FOR GLIOBLASTOMA THERAPY

Sonali Setua ¹, Colin Watts ¹, Mark E Welland ¹

Abstract

INTRODUCTION: Glioblastoma is the most common primary malignant brain tumour in adults. Despite recent therapeutic advances, the treatment of glioblastoma remains inadequate. Moreover, present evidences suggest that; glioblastoma contains tumour-initiating cancer stem cells. They are resistant to the conventional radiation and chemotherapy which results in tumour recurrence. Gold nanoparticle mediated enhanced X radiation therapy can be a viable solution in this regard. Because of its high atomic number, gold preferentially absorbs kV X- rays compared to the soft tissue, resulting in the emission of ionizing Auger electrons. The enhanced free radicals generated by the gold nanoparticle under this condition and temozolomide induced DNA damage can lead to the effective demolition of the malignant tissue. METHODS AND RESULTS: A novel nanoconjugate made of human serum albumin and polyethylenimine capped gold nanoparticle is developed. DLS, electron microscopic and UV-Visible spectrophotometric analysis suggest that the nanoconjugates are extremely stable at physiological pH. Cell viability assay showed that the nanoconjugates are nontoxic. However, the uptake of this nanosystem by glioblastoma tumour-initiating cells yet to be analyzed by EDX spectroscopy and confocal microscopy. CONCLUSION: Preliminary results indicate that the nanoconjugates are suitable for biomedical applications. In future we will analyze the potential of this nanosystem for gold nanoparticle mediated radiosensitization in combination with temozolomide induced DNA damage for glioblastoma therapy. Moreover, by functionalizing this nanosystem with appropriate targeting moieties more efficient tumour-initiating cell targeted therapy can be developed.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

57. RECOMBINANT HUMAN HEAT SHOCK PROTEIN HSP70 AS POSSIBLE ADJUVANT IN TREATMENT OF MALIGNANT BRAIN TUMOURS IN CHILDREN

MA Shevtsov ¹, WA Khachatryan ¹, AV Kim ¹, KA Samochernych ¹, AV Pozdnyakov ¹, IV Guzhova ¹, IV Romanova ¹, BA Margulis ¹

Abstract

INTRODUCTION: Despite progress in the treatment of malignant brain tumours, patient prognosis remains very poor with a median survival of less than 15 months. One of the promising therapeutic strategies can be based on immunomodulatory activity of molecular chaperons, particularly Hsp70. METHODS: Using the C6 intracranial glioma model rats were intratumourally treated with Hsp70 on the 14 day after implantation of C6 cells. Cytotoxic activity of lymphocytes (CTL–test), CD3 + , CD4 + , CD8+ cell infiltration of animal brains in control and treated groups were assessed. Tumour volume was assessed using magnetic resonance imaging (MRI). All treated rats were followed for survival. In addition, 10 patients with a diagnosis of malignant brain tumour were treated by 5 intratumoural injections of Hsp70. Immunological assays were made both before and after Hsp70 treatment. RESULTS: Intratumoural injection of purified Hsp70 increased the survival rate of animals from 20 ± 2 to 28 ± 1 days (P<0.01). This correlated with increased CTL response and massive infiltration with CD3 + , CD4+ and CD8+ cells, both in the zone of injection and in tumour itself. Delay of tumour volume growth was detected by MRI in the Hsp70 treated group. Injection of Hsp70 was well tolerated by patients. There were no serious adverse effects. In peripheral blood an increase of markers of T-cell mediated immunity was observed. CONCLUSIONS: Our results suggest that Hsp70 has a high antitumour effect due to activation of cell-mediated immunity and thus can become a useful therapeutic strategy in treatment of malignant brain tumours.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

58. PAEDIATRIC BRAIN TUMOURS DEMONSTRATE VASCULOGENIC MIMICRY IN A THREE DIMENSIONAL CULTURE SYSTEM

SJ Smith ¹, R Rahman ¹, C Rahman ¹, JH Barrow ¹, DC Macarthur ¹, F Rose ¹, RG Grundy ¹

Abstract

INTRODUCTION: Two-dimensional (2D) monolayer cell cultures represent highly reductionist tumour models due to the loss of physiological extracellular matrix (ECM), making complex three dimensional (3D) processes such as angiogenesis difficult to model effectively. We therefore present a novel 3D culture system to create macroscopic tumour aggregates. We describe the formation of a substantial ECM, upregulation of angiogenic pathways, and derivation of vessel like structures from tumour cells (vasculogenic mimicry). METHODS: The Rotary Cell Culture System (RCCS) was used to cultivate ∼8mm diameter aggregates of paediatric high grade glioma and medulloblastoma cell lines. Expression of 84 angiogenesis and 84 ECM related genes was assessed by RT-PCR. Immunohistochemistry was performed against vessel related antigens, ECM and angiogenic components. Genome wide gene expression analysis was performed. RESULTS: The tumour aggregates possess a complex structure with distinct proliferating, senescent and necrotic regions, replicating primary tumour, including a similar cell population density. Tubular structures within the aggregates, with expression of antigens such as CD105, represent vasculogenic mimicry by tumour cells. Many key angiogenic and ECM genes are greatly upregulated in 3D compared to 2D culture, including EGF, IGF1 and TGF beta, (p-values less than 0.05). CONCLUSIONS: The RCCS creates 3D tumour aggregates recapitulating many key features of paediatric brain tumours, including histology and gene expression. The phenomenon of vasculogenic mimicry will be discussed with respect to our study. Our findings contribute to understanding angiogenesis and possible mechanisms of resistance to current anti-angiogenic therapies.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

59. A WALK IN THE PARP? THE POTENTIAL TO TARGET DNA REPAIR IN PAEDIATRIC HIGH GRADE GLIOMA

SJ Smith ¹, A Long ¹, JH Barrow ¹, DC Macarthur ¹, B Coyle ¹, RG Grundy ¹

Abstract

INTRODUCTION: Paediatric high grade gliomas (pHGG) remain tumours with a poor prognosis for which novel therapeutic strategies are needed. Poly (ADP-ribose) polymerase (PARP) is known to have multiple functions in tumours including single strand DNA repair and induction of caspase independent apoptosis. It has been suggested as a therapeutic target in adult malignancies and this study examines whether it could be a potential target in pHGG. METHODS: Tissue microarrays containing 150 formalin fixed pHGG were examined by immunohistochemistry for levels of PARP and apoptosis inducing factor (AIF) expression. Full retrospective clinical data were also available for this cohort and statistical analysis was performed to assess for the effect of PARP status on prognosis. Copy number of 1q was assessed by array comparative genomic hybridisation and fluorescent in situ hybridisation. RESULTS: Level of PARP immunopositivity had a statistically significant positive correlation with survival in supratentorial paediatric high grade glioma. AIF staining was notable for its absence in the majority of tumours but with higher levels of expression in non-neoplastic brain. Extent of surgical resection and administration of chemotherapy or radiotherapy were the other significant factors on multi-variate analysis. No correlation was found between gain of 1q and PARP immunopositivity levels. CONCLUSIONS: PARP is expressed at significant levels in many pHGG and has a significant correlation with prognosis, though this increased expression is not DNA copy number driven. In these tumours the ability of PARP to activate AIF appears to have been lost. PARP may therefore represent a promising therapeutic target for these lesions.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

60. MEASURING ELASTICITY OF HIGH-GRADE HUMAN BRAIN TUMOUR CELLS

Zaynah Maherally ¹, James R Smith ¹, Luke Dickson ¹, Geoffrey J Pilkington ¹

Abstract

INTRODUCTION: CD44 is over-expressed on cancer cells and plays a key role in cell adhesion, migration and invasion. The increased action of CD44 facilitates the structural scaffold of cancer cells. METHODS: We have investigated the mechanical properties of high-grade human glioma cells (SNB-19) using atomic force microscopy. Normal human astrocytes (CC-2565) were used as a control. We also examined siRNA CD44 knock-down SNB-19 cells. An AFM probe was pressed into cell surfaces (on and away from nuclei; n = 2 × 50) and Young's Modulii (E) determined. Fluorescence images, using F-actin and GFAP staining, were also acquired. RESULTS: Nuclei regions of all cells types had low E-vales (SNB-19 and CC-2565 cells, 0.67 ± (sem)0.05 kPa and 0.64 ± 0.06 kPa, respectively; siRNA CD44 knock-down, 0.28 ± 0.03 kPa), attributed to ease of nucleus displacement. Away from nuclei, SNB-19 cells were more rigid than CC-2565 cells (2.34 ± 0.58 kPa and 1.09 ± 0.13 kPa, respectively), whilst siRNA CD44 cells were less compliant (0.61 ± 0.08 kPa) than SNB-19 and CC-2565 cells. DISCUSSION: Greater rigidity of SNB-19 cells may be explained by the probe sensing the underlying hard glass surface, since malignant cells with active CD44 are thinner due to extension of invadopodia. Results are discussed with reference to cell cytoskeleton and AFM literature. AFM nanoindentation could serve as a potential tool for testing anti-cancer drug efficacy and understanding tumour invasion.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

61. TOWARDS ESTABLISHING THE EFFECTS AND MECHANISM OF ACTION OF A SERIES OF INDOLES IN AN IN VITRO CHEMOSENSITIVITY SYSTEM FOR GLIOMA TREATMENT

Saurabh Prabhu ¹, Frederick Harris ¹, Robert Lea ¹, Timothy J Snape ¹

Abstract

INTRODUCTION: Substituted indoles and related structures have been shown to exhibit potent anticancer activity against breast cancer cell lines. Here, the effects of structurally similar substituted indoles against the human glial cancer cell lines, 1321N1 and U87MG, have been investigated by comparing the effects of these compounds to conventional anti cancer drugs. METHODS: Cell viability in the presence of the test compounds was measured using an MTS assay and corroborated by an ATP cell proliferation assay as well as a Trypan blue exclusion test. The significance of reactive oxygen species (ROS) in the process was determined using an Image-iT^® LIVE ROS kit from Invitrogen. RESULTS: Both cell lines were treated with four commercial anticancer drugs and IC50 values were only reached at concentrations of 20 µM for cisplatin and 50 µM for gemcitabine over 48hrs on the 1321N1 cell line. However, the more malignant U87MG cell line was resistant to all the drugs, except for cisplatin where the IC50 value was reached at 300 µM after treatment for 48hrs. Similar studies were carried out with various substituted indoles and the cytotoxicity results on both cell lines showed that the IC50 value was reached within 90 minutes for the most potent compound at a concentration of 600 µM (1321N1) and 800 µM (U87MG). The idea that the mechanism of action of these compounds may work through the generation of ROS was investigated and this was confirmed over a similar time course using a suitable fluorogenic marker. Moreover, it was shown that the addition of an antioxidant (ascorbic acid) abolished the potency of the most active compound. CONCLUSION: Here, it has been demonstrated that certain substituted indoles are able to have a rapid, deleterious effect on the viability of two glioma cell lines and indicated that ROS generation may induce cell death.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

65. AN AUDIT OF REFERRAL PATHWAYS FOR CNS IMAGING OF COMMON NEUROLOGICAL CONDITIONS IN CHILDREN (<18 YEARS) AGAINST ROYAL COLLEGE OF PAEDIATRICS AND CHILD HEALTH (RCPCH) BRAIN TUMOUR REFERRAL GUIDELINES

Maya Sussman ¹, Sophie Wilne ¹, William Whitehouse ¹, Gabby Chow ¹, Jo-Fen Liu ¹, David Walker ¹

Abstract

INTRODUCTION: Brain tumours constitute a differential diagnosis of many childhood neurological presentations; CNS imaging is essential for diagnosis or exclusion of tumour as a cause. Previous UK studies identify prolonged symptom intervals compared to other European health systems. This project aimed to evaluate actual referral pathways to CNS imaging against the RCPCH Guidelines. METHODS: Data was collected from 41 paediatric neurology clinic patients and 11 newly diagnosed patients with brain tumour using a structured interview tool combined with case-note review. Compliance with referral and CNS imaging guidelines conditional on headache, vomiting, seizure and motor symptoms was measured. Hospital appointment and CNS imaging waiting times were calculated. Handoff maps were used to visualise pathways to CNS imaging. RESULTS: Compared with RCPCH referral guidelines, there was poor compliance with referral guidelines of 64.1% and CNS imaging guidelines of 39.0% (p = 0.003). Non compliance with CNS imaging guidelines was associated with young age < 5yrs (26.1% p = 0.03) and presenting symptom (p = 0.007); those with seizures (29.0%) or motor disturbances (32.4%) being least compliant. Handoff maps showed many patients were involved in tortuous routes to CNS imaging, causing avoidable cumulative delay. CONCLUSIONS: The results support the need for a national awareness campaign linked to the RCPCH guidelines. The “HeadSmart - be brain tumour aware” campaign will be launched in June 2011 in the UK in response. The structured interview tool and handoff maps will be used as part of the HeadSmart campaign's evaluation and educational outreach programme (http://www.rcpch.ac.uk/bpp).

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

66. EXPLOITING CONFORMATIONALLY RESTRICTED UREAS AS BIOLOGICALLY ACTIVE C=C DOUBLE BOND ANALOGUES AGAINST GBM CELLS IN VITRO

TJ Snape ¹, A Karakoula ¹, F Rowther ¹, TJ Warr ¹

Abstract

INTRODUCTION: The combretastatin family of biologically active stilbenes possess promising antimitotic activity through interaction with the colchicine binding site on tubulin. We have synthesised 5 novel combretastatin analogues which exploit conformationally restricted bond rotations and thus mimic the cis-geometry of combretastatin. We compared the in vitro activity of these analogues to the combretastatin phosphate salt, CA4P. METHODS: Established synthetic and analytical techniques were employed to prepare the compounds and analyse their structures and conformations. A tubulin polymerisation assay was used to assess in vitro microtubule interactions. The cytotoxic and anti-proliferative effects of all compounds were evaluated in 3 GBM short-term cell cultures and the established glioma cell line U251. RESULTS: CA4P had higher activity than the 5 analogues in all in vitro assays. It was the most efficient inhibitor of tubulin polymerisation with a Vmax of 0.43 at 10 µM compared to the analogues (range 0.64–0.76). CA4P also induced the highest levels of cytotoxicity, apoptosis and mitotic arrest. Of the analogues, the two most similar in shape and functionality to CA4P were the best tubulin inhibitors, and better than colchicine. Conversely, the analogue least like CA4P was relatively inactive in all cell cultures and a poor inhibitor of tubulin. Gratifyingly, the analogue most like inactive trans-combretastatin was the worst tubulin inhibitor. CONCLUSIONS: Although the biological activity of the analogues was lower than CA4P, the structure-activity relationship of these novel compounds enhances our understanding for future drug development. We have demonstrated an excellent correlation between conformation and functionality and validated this new class of combretastatin analogue.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

67. TO INVESTIGATE WHETHER THERE IS EVIDENCE OF A CORRELATION BETWEEN THE PTV AND THROMBOCYTOPENIA DURING CONCOMITANT TREATMENT IN PATIENTS WITH NEWLY DIAGNOSED GBM

Aoife Williamson ¹, Mairi Mackinnon ¹

Abstract

INTRODUCTION: Subgroup analysis in the original Stupp study indicated that patients aged >60 derived least benefit from intensive multimodal therapy. The impact of radical concomitant treatment on quality of life (QoL) of these patients has not been studied. The aim of this study was to investigate the QOL at 3 separate time points (using WHO performance status (PS) as a surrogate of QOL). METHODS: Medical records were examined retrospectively. WHO PS was extracted at 3 timepoints: (1) 1st appointment with oncologist, (2) at start of adjuvant chemotherapy and (3) cycle 6 of adjuvant treatment. Survival data were calculated from date of surgery by Kaplan-Meier method using SPSS version 16. RESULTS: Of the 130 patients, 40 were > 60 years. 10 patients did not receive adjuvant treatment, 14 completed 1 -3 cycles, 9 completed 4–5 cycles and 7 completed adjuvant treatment. Median survival was 12.3 months. One year overall survival (OS) was 55.1%. WHO PS data at the timepoints were available for 40, 40 and 7 patients respectively: (1) 20 pts PS 0, 20 pts PS 1; (2) 10 pts PS 0, 29 pts PS 1, 1 pt PS 2; (3) 2 pts PS 0, 3 pts PS 1, 2 pts PS 2. CONCLUSIONS: The majority of patients' PS dropped by 1 point during treatment, although we cannot exclude more significant drops in PS causing cessation of therapy. These preliminary data indicate that, with appropriate patient selection and careful on treatment review, radical chemoradiation can be delivered to patients aged 60–70 with acceptable deterioration in PS. Future studies will prospectively assess QoL and duration of stable/responding disease after completion of adjuvant treatment.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

68. MALIGNANT CNS PRIMARY LYMPHOMAS. A CHALLENGE OF MANAGING. EXPERIENCE OF A 20 YEARS FOLLOW UP

AK Zisakis ¹, V Varsos ¹, A Panteli ¹, O Karypidou ¹, AM Zampethanis ¹

Abstract

INTRODUCTION: Primary central nervous system lymphoma (PCNSL) is a rare form of the non-Hodgkin's lymphoma arising within and confined to the CNS. PCNSL is of a particular interest for several reasons. First this tumour has increased in incidence over the past several decades. Therefore, although it remains relatively rare, the differential diagnosis remains very important from other intracranial or spinal mass lesions. Second unlike many primary brain tumours, PCNSL is well responsive to treatment and aggressive management may lead to prolonged remission or cure. Finally, the long term consequence of aggressive therapy was result in significant neurologic dysfunction. METHODS AND RESULTS: In our study within the period 1990–2010, 25 patients have been diagnosed with PCNSL. 17 out of them were cerebral PCNSL and 8 were spinal PCNSL. From the 17 cerebral cases, 16 were high grade diffuse large B-cell lymphomas and 1 was characterised as low grade lymphoma. All the patients underwent surgical operation for a biopsy or gross total excision and treated with corticosteroids and radiotherapy (50Gy). Chemotherapy (methotrexate) applied in 20 of the patients. Patients were monitored with monthly MRI scans after initiation of methotrexate chemotherapy. CONCLUSIONS: The optimal treatment for PCNSL has not been established. Radiation therapy alone produces a median survival duration of only 18 months. Combination chemotherapy and radiation therapy more than doubled survival time (up to 50 months) but such success was achieved at the price of a greater than 50% incidence of dementia in those who survived more than 18 months of these regimens.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

69. Y-BOX BINDING PROTEIN-1 (YB-1) INHIBITION TRIGGERS DIFFERENTIATION OF NORMAL AND CANCER STEM CELLS FROM THE BRAIN

Abbas Fotovati ¹, Samah Abu-Ali ¹, Pei-Shan Wang ¹, Loic Deleyrolle ², Cathy Lee ¹, Joanna Triscott ¹, James Y Chen ¹, Sonia Franciosi ¹, Yasuhiro Nakamura ³, Yasuo Sugita ⁴, Takeshi Uchiumi ³, Michihiko Kuwano ⁵, Blair R Leavitt ¹, Sheila K Singh ⁶, Alexa Jury ⁷, Chris Jones ⁷, Hiroaki Wakimoto ⁸, Brent A Reynolds ², Catherine J Pallen ¹, Sandra E Dunn ¹

Abstract

INTRODUCTION: The Y-box-binding protein-1 (YB-1) is up-regulated in many human malignancies including glioblastoma (GBM). It is also essential for normal brain development, suggesting that YB-1 is part of a neural stem cell (NSC) network. MATERIALS AND METHODS: The techniques used to investigate this project included the use of transgenic mice, isolation of primary neural stem cells from mice, differentiation using small interfering RNAs against YB-1, 10% FBS, BMP-4 or BNDF, immunofluorescence, flow cytometry, immunoblotting, neurosphere growth assays and the analysis of stem cell markers by immunostaining tumour tissue microarrays. RESULTS: Here we show that YB-1 was highly expressed in the subventricular zone (SVZ) of mouse fetal brain tissues but not in terminally differentiated primary astrocytes. Conversely, YB-1 knock-out mice had reduced Sox-2, nestin and musashi-1 expression in the SVZ. While primary murine neurospheres were rich in YB-1, its expression was lost during glial differentiation. Likewise, YB-1, Sox-2, mushashi-1, Bmi-1 and nestin are coordinately expressed in SF188 cells and 9/9 GBM patient-derived primary CSCs. Silencing YB-1 with small interfering RNA attenuated the expression of these NSC markers, reduced neurosphere growth and triggered differentiation via coordinate loss of GSK3-b. Further, differentiation of CSCs with 1% serum or bone morphogenetic protein-4 (BMP-4), suppressed YB-1 protein expression. Likewise, YB-1 expression was lost during differentiation of normal human NSCs. Consistent with these observations, YB-1 was preferentially expressed in highly undifferentiated primary gliomas based on the analysis of WHO grade I-IV tumors (n = 49 cases). YB-1 was also co-expressed with Bmi-1 (Spearmans 0.80, p > 0.001) and Sox-2 (Spearmans 0.66, p > 0.001) based on the analysis of 282 cases of high-grade gliomas. These CSC markers were also highly expressed in 10/15 (67%) of GBM patients that subsequently relapsed. CONCLUSIONS: YB-1 is a key feature of stem cells where it functions to suppress differentiation.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

70. TAILORING DECISION NAVIGATION (DN) TO HIGH GRADE GLIOMA PATIENT'S NEEDS

SC Shepherd ¹, SE Scott ¹, D Bowyer ¹, LM Wallace ¹, B Hacking ¹

Abstract

INTRODUCTION: In 2008–10 Decision Navigation (question-listing, audio-recording, note taking1) was piloted with Prostate cancer patients. Qualitative evaluation shows DN helped patients participate in treatment decisions; quantitative measures report navigated patient's confidence in decision making was significantly enhanced. The National Institute of Health and Clinical Excellence2 (NICE) recommends brain tumour patients should be involved in making decisions about healthcare. This study aims to develop and tailor DN to the needs of patients with high grade glioma (HGG). METHODS: Two consecutive focus groups were conducted with Brain tumour patients and relatives (n = 7). Seven HGG patients took part in in-depth interviews post-surgery prior to their initial oncology appointment. Data was analysed using framework analysis3. RESULTS: Focus groups reported: an “excruciating” wait for a diagnosis they were not prepared for. Retaining information was difficult, while thinking of and asking questions was frightening. Patients described feeling like a statistic and being part of the medical process without being consulted. Pre-diagnostic interviewees reported: honest and open information from surgeon's pre-surgery allowed patients to feel more in control, although waiting to discuss pathology was an uncertain and anxious period, not helped by a lack of clear procedural information from post surgery to diagnosis. CONCLUSION: The period of waiting for diagnosis is fraught with uncertainty. The navigation intervention should be implemented at this early stage as a form of support and dialogue. Facilitation to think ahead about relevant questions, encouragement to voice questions, and the provision of personalised information from the consultation would be greatly beneficial for both patients and carers.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

71. NORMALIZED CEREBRAL BLOOD VOLUME IN THE PERI-TUMOURAL REGION HELPS IDENTIFY INFILTRATION

LA Mohsen ¹, R Jena ¹, JH Gillard ¹, SJ Price ¹

Abstract

INTRODUCTION: The undetected invasion of glioblastomas is a leading cause for failure of current treatments and poor prognosis. Post-mortem studies showed that glioma cells may extend for less than 1 cm from the tumour edge in 25% of patients and for more than 3 cm in further 20%. As GBMs invade, they need to develop blood supply. In a previous study1, we showed that it is possible to detect the invasive margin of GBM by comparing the isotropic (p-map) and anisotropic diffusion (q-map) abnormalities. AIM: In this study we aim to see if angiogenesis can be detected in invasive regions by comparing the rCBV in this infiltrative region and other peri-tumoural regions. MATERIALS AND METHODS: 20 GBM patients (mean age 61), were imaged pre-operatively. All patients had standard anatomical MRI sequences and standard DTI and DSCI perfusion MRI. DTI series were analyzed by FSL (Oxford, UK), while DSCI series were analyzed by Nordic Ice (NordicNeuroLab, Norway). The p and q maps were calculated as previously described2. The p and q abnormalities were outlined on their respective maps. The ROIs were overlapped on the rCBV maps. More ROIs were applied to the regions where the p abnormality exceeds the q abnormality by >1 cm. The maximum rCBV (normalized to the contra-lateral centrum semi-ovale) was recorded. The same was done for the peri-tumoral regions immediately outside the p ROIs but away from the proposed infiltrative region. We excluded regions in the vicinity of cortical vessels or choroid plexus (falsely elevated rCBV). The results from all patients were compared using a paired t-test. RESULTS: Normalized rCBV values recorded from the proposed infiltrative region ranged had a range of 2.1–8.2, while the values recorded from peri-tumoural region outside the isotropic abnormality ranged from 1.2–3.8. Comparing these values revealed significant difference between the presumed infiltrative region and other peri-tumoural regions (p = 0.00015). CONCLUSION: The current results add strong evidence to the ability of the DTI to outline the infiltrative regions around the GBM, proving that there is significant difference of rCBV between what is presumed to be the infiltrative margin of the GBM and other peri-tumoural regions. This still needs more validation by biopsy.

Neuro Oncol. 2011 Oct;13(Suppl 2):ii1–ii14.

72. TARGETING POLO-LIKE KINASE (PLK) FOR THE TREATMENT OF BRAIN TUMOURS AND ITS UNIQUE CAPACITY TO ELIMINATE CANCER STEM CELLS THROUGH SOX-2 INHIBITION

C Lee ¹, A Fotovati ¹, M Verraeult ¹, H Wakimoto ¹, B Reynolds ¹, C Dunham ¹, M Bally ¹, J Hukin ¹, S Singhal ¹, S Singh ¹, SE Dunn ¹

Abstract

INTRODUCTION: In a genome-wide siRNA library screen of 691 kinases, we previously reported that polo-like kinase 1 (PLK1) inhibition suppressed the growth of paediatric glioblastoma cells in vitro. PLK1 was a particularly attractive molecular target for paediatric oncology because BI2636, a PLK small molecule inhibitor, has been evaluated in phase I and II clinical trials for adult cancers. In this study, we examined PLK inhibitors for the treatment of gliomas and specifically addressed the impact on brain tumour-initiating cells (BTICs) given that they are often resistant to current therapies and as such are believed to facilitate relapse. METHODS: PLK was inhibited with small interferring RNAs and BI2536 to assess their impact on tumour cell growth and viability. Primary BTICs were isolated from patients and the explanted cells were challenged with PLK inhibitors in a dose-dependent manner. Neurosphere and monolayer growth was assessed over time. The consequence of PLK inhibiton on Sox-2, musashi and BMI-1 was assessed by immunoblotting and qRT-PCR. Further, the ability of PLK inhibitors to suppress tumour growth was assessed orthotopically in mice. RESULTS: PLK inhibition with siRNA or BI2536 significantly blocked the growth, caused G2/M arrest and induced apoptosis of GBM and medulloblastoma cell lines (IC50 = 2–5 nM). However, PLK1 inhibition exerted very modest effects on the growth of immortalized human astrocytes. Of note, PLK1 inhibitors were cytotoxic to the GBM cell line SF188 that is TMZ-resistant. PLK1 was readily detectable in primary patient-derived BTICs (n = 8–10 samples) and moreover inhibiting it suppressed the stem cell marker Sox-2, neurosphere growth and induced cell death. Further, BI2536 crosses the blood brain barrier and prolonged survival of mice with GBM. CONCLUSIONS: We have demonstrated for the first time that primary BTICs are exquisitely sensitive to PLK inhibition and that BI2536 has in vivo activity using an orthotopic model of GBM. Similarly, primary medulloblastoma BTIC cells are dependent upon PLK for survival and as such further pre-clinical development is justified. To conclude, inhibition of PLK1 may help overcome drug resistance and prevent disease relapse, which are two major challenges for long-term survival of patients.

PERMALINK

Abstracts from the 2011 BNOS Conference, June 29 – July 1, 2011, Homerton College, Cambridge

2. GAS6/AXL-FAMILY RECEPTORS IN SCHWANNOMA PATHOLOGICAL PROLIFERATION, ADHESION AND SURVIVAL

S Ammoun

L Zhou

M Barczyk

D Hilton

S Hafizi

CO Hanemann

Abstract

3. CD133 GLYCOSYLATION IS AFFECTED BY HYPOXIA IN CULTURED GLIOMA STEM CELLS

Kristina S Lehnus

Laura K Donovan

Geoffrey J Pilkington

Qian An

Abstract

4. COMPLICATIONS FOLLOWING GLIADEL WAFER INSERTION DURING SURGERY FOR GLIOBLASTOMA MULTIFORME: A SINGLE CENTRE-EXPERIENCE

Ian A Anderson

Simon Thomson

Abstract

6. SURVIVAL OF PATIENTS WITH GLIOBLASTOMA – HOW DO WE COMPARE TO STUPP?

M Bailey

E Lekka

J Law

C Davis

Abstract

7. STEREOTACTIC RADIOSURGERY AT LEEDS GAMMA KNIFE CENTRE: INITIAL EXPERIENCE OF TREATING BRAIN METASTASES

KE Banfill

C Loughrey

P Hatfield

Abstract

8. HIGH THROUGHPUT DRUG SCREENING OF PAEDIATRIC GLIOMA CELL LINES

Dorine Bax

Richard Elliott

Ryan Bishop

Katy Taylor

Lynley Marshall

Nathalie Gaspar

Marta Viana-Pereira

Rui Reis

Jane Renshaw

Alan Ashworth

Chris Lord

Chris Jones

Abstract

9. THE SELECTIVE ACTION OF TRICHOSTATIN A IN COMBINATION WITH THE DNA DAMAGING AGENTS, TEMOZOLOMIDE AND CISPLATIN

CA Bellamy

L Shaw

JE Alder

AJ Shorrocks

RW Lea

Abstract

10. MODULATING IMMUNOSUPPRESSIVE BEHAVIOUR IN GLIOBLASTOMA

Suzanne Birks

Michael Burnet

Geoffrey Pilkington

Abstract

12. A SINGLE CENTRE CASE CONTROL STUDY OF THE EFFICACY AND SAFETY OF 5-AMINOLEVULINIC ACID GUIDED RESECTIONS OF GRADE IV (WHO) GLIOBLASTOMAS

Julius D Bruch

Jasmine Ho

Colin Watts

Stephen J Price

Abstract

13. REALTIME INTRAOPERATIVE THREE DIMENSIONAL ULTRASOUND IN BIOPSY/RESECTION OF INTRINSIC BRAIN LESIONS

SJ Camp

V Apostolopoulos

A Mehta

F Roncaroli

D Nandi

Abstract

14. THROMBOCYTOPENIA DURING CHEMORADIOTHERAPY FOR GLIOBLASTOMA: POTENTIAL IMPLICATIONS FOR ADJUVANT TREATMENT AND OUTCOME

B Clark

M Mackinnon

N MacLeod

W Stewart

A Chalmers

Abstract

15. ADHERENCE TO NICE GUIDANCE IMPROVES SURVIVAL FOR GBM PATIENTS IN NORTHERN IRELAND

AJ Cole

GG Hanna