Presentation on theme: "The Bristol Experience of Molecular Genetic Analysis of"— Presentation transcript:
1The Bristol Experience of Molecular Genetic Analysis of Gliomas (LOH and MGMT)for Optimisation of TreatmentCMGS Spring Meeting 27th March 2009The care of all patients with brain and other central nervoussystem (CNS) tumours should be coordinated through a specificmodel of multidisciplinary assessment and careHilary SawyerBristol Genetics Laboratory
2Outline Background to Gliomas Clinical Trials suggesting prognostic indicatorsClinical Utility of 1p19q LOH and MGMT testingBristol team dataCase studiesHow results are used clinically in BristolThe future and laboratory issues
3Brain tumours 1.6% of cancers in England and Wales High morbidity and mortalityCommon symptoms: headache with cognitive or behavioural symptoms, epilepsy, progressive focal neurological deficits, raised intracranial pressure• Treatment - surgery, radiotherapy and more recently chemotherapyTumours within the brain, such as gliomas, can rarely be completely removed because of their relation to critical structures and the infiltrating nature of the tumourMany do not demonstrate metastasis but invasion may preclude surgical resection generating ongoing management issuesSlow growing tumours may transform into more aggressive tumoursClassified by cell type, grade and location2007 WHO classification:Grade I: low grade, well circumscribed, slowly progressing, can often be cured by resectionGrade II: low grade, typically infiltrative, low proliferation but higher likelihood of recurrence Grade III and IV: high grade, malignant, grow rapidly, aggressiveThere is considerable inter-observer variation in diagnosis and classification however.Molecular tests help understand pathogenesis and improve classification.Evidence for two predictive molecular markers emerging for GLIOMAS:1p/19q LOH and MGMT
4Gliomas Tumours arising from glial cells Non-neuronal cells provide physical support and nutritionmaintain homeostasisform myelinparticipate in signal transmissionGlial cell typesAstrocytes – anchor neurons to blood supply, regulation and signalling ASTROCYTOMASOligodendrocytes - produce myelin sheath OLIGODENDROGLIOMASMixed Gliomas – OLIGOASTROCYTOMASEpendymocytes - lining and secret CSF EPENDYMOMASGlioblastomas are the most common form of astrocytic tumourAverage survival times:Grade II astrocytoma: 7 yearsAnaplastic astrocytoma: 3.5 yearsGlioblastoma: 9-11 monthsHigh-grade glioma (WHO III and IV) includes:glioblastoma, anaplastic astrocytomas, anaplastic oligodendrogliomas andanaplastic ependymomaswell documented to progress to higher grade malignancy
6OligodendrogliomaTypical ‘fried egg’ appearance (fixation artifact):Oligodendroglia - specialised CNS myelin-forming cells.Rare primary brain tumours, frequency ~0.53 cases/100,000Around 25% primary brain tumours in adults~20% are anaplasticAnaplastic oligodendroglioma (WHO grade III)Median age of onset between 40 & 50 yearsTreatment by surgical resection, followed by radiotherapy and PCV chemotherapy on recurrence.Main aims of treatment and follow-up are to increase survival while maximising a patient’s functional capability and quality of lifeRelatively good prognosis, survival of 3-10 years from diagnosis.Recurrence is common, often leading to disease progression and death.
7Molecular Markers in Oligodendrogliomas (ODs) Various trials support the clinical utility of analysis for 1p19 LOH:There had been general disillusionment with chemotherapyVarious trials have showed improved outcomesin patients with 1p19q LOH:Initial trials: Cairncross G et al (1994 and 1998)1p and 19q deletions were observed in ~66% ODsReifenberger et al Neuropath Exp Neurol 2003LOH 1p/19qNo LOHThe above were small trials or retrospectiveseries suggesting predictive value.Further trials (Phase III) were needed:From: McDonald et al (2005) Cancer. 1;104(7):1) RTOG Trial 9402: Pure and Mixed Anaplastic Oligodendroglioma: PCV + RT vs RT aloneProgression-free survival time favoured PCV + RT but 65% of patients experienced toxicityTumours with LOH 1p19q have longer median survival times (>7 v 2.8 yrs) and longer PFSBetter prognosis, time to relapse but no difference whether early v late chemotherapy2) EORTC: In newly diagnosed AODs and OAs:Adjuvant chemotherapy improves PFS but does not affect OS, therefore timing is less relevant.Oligodendrogliomas with 1p loss alone have an intermediate prognosis. Ino Y et al. Clin Cancer Res (2001).I
83) NOA-4 study Phase III trial Multicentre randomised trial in Germany of sequential radiochemotherapyof oligoastrocytic WHO grade III tumours with PCV or temozolomideWolfgang WW et al (2008) J Clin Oncol 26Patients with any grade III tumour: anaplastic oligodendroglioma, oligoastrocytoma or astrocytoma with orwithout deletions Either 6 week course RT or 4x6 week cycles of either PCV or temozolomideLOH 1p/19q and hypermethylation of MGMT large risk reduction for time totreatment failure (TTF) regardless of histologyOligodendroglial histology better than astrocytic (NB 1p/19q LOH is present in higher% of cases of AOD than AOA, also low in mixed OAs with predominant OD)No difference in TTF between patients on RT v chemotherapy or which is used firstMGMT methylation status may have been more importantThese trials confirmed that 1p/19q loss improved outcome following RT with or without chemotherapyi.e. appears to be a prognostic factor for survival regardless of treatmenttype or timingODs with 1p19q LOH also respond more favourably to temozolomide(an alkylating agent)
9Loss of Heterozygosity Analysis LOH (Bristol) D1S468D1S214D1S2736D1S19919q1pD19S408D19S412D19S926D19S418Ino Y et al. (2001) Clin Cancer Res 7,Smith J et al (2000). J Clin Oncol:Cairncross et al (1998) J Natl Cancer Inst 90,Marker profile of tumour DNA compared to blood DNA to determine LOH. Panel of 8 markers in 3 multiplexes and one simplexFresh/frozen tumours used in Bristol v PPFEAs oligodendrogliomas may be diffuse tumours, problems with mixed tissue can occur (require 60-90% tumour tissue)DNA from peripheral blood:DNA from tumour tissue:Allelic loss
10Bristol LOH 1p19q Data 2005 to early March 2009 300 patients Bristol LOH 1p19q Data 2005 to early March patients. Approx 50 per year but increasingThe majority of pure AODscarry 1p/19q deletions: predictive,prognosticTumourTypeNo LOH%LOH1p/19qLOH 1pLOH 19qPartialPartial LOH1p+19qUn-ReportableTotalNumberOligodendroglioma7.582.5540Oligoastrocytoma34.6467.73.826Astrocytoma22.214.171.124.851Malignant astrocyticglioma502Infiltrating astrocytictumour85.77.114Glioblastoma57.61.21610.15.10.6158Gliosarcoma2514NeuroectodermalLiponeurocytoma100300Some OAs have loss andrespond wellGrey = low no casesThere is a good response ofglioblastomas with 19q deletionPatient with rare histologywith 1p/19q LOH and goodresponse to PCVLiterature: Where co-deletion of 1p and 19q is present this is usually found throughout the tumour
11Clinical Case 1 Aug 02 Male aged 68 at presentation Confusion and left sided weaknessRight parieto-occipital massPartial resectionHistology : ganglioglioma with foci of mitotically active primitive neuroectodermal tumourOct/Nov 02 Radical radiotherapyDec 02 Massive recurrenceFurther surgery but prognosis poor
12Clinical Case 1 (con)Dec 02 1p19q deletion detected and offered palliative PCVFeb/Oct 03 PCV with complete radiological responseJun 06 relapse treated with stereotactic radiosurgeryDec 06 Rapid declineFeb 07 Died1p19q analysis correctly identified a chemosensitive tumour, even when the morphology was confusing and the clinical course appeared aggressive.Patient had a 3.5 year remission with a good quality of life due to this intervention
13Clinical Case 2 Dec 06 40 yr old female Presented with headaches and drowsinessExtensive tumour in right hemisphere and thalamusJan 07 Partial resectionHistology: Central liponeurocytomaRare tumour- c. 25 reported casesno clear guidance on treatment1p19q deletions detectedFeb/Apr 07 Radical radiotherapyOct 07 Recurrence –further surgeryHistology unchangedNov/Jul 08 PCV chemotherapyJul 08 MRI clear1p19q encouraged the use of PCV where no data was availableIt has already proved of more durable adjuvant benefit than radiotherapy
14Glioblastomas Commonest primary brain tumour: ~5/100,000 annum Either develop from lower malignancy grade tumour or de novo (different genes/same cell pathway)50% respond to alkylating agent temozolamideGlioblastoma (WHO grade IV)Responsive tumours show promoter methylation (inactivation) of the MGMT (O6-methylguanine-DNA methyltransferase) gene (10q26)MGMT is a DNA repair protein (suicide enzyme) that removes alkyl groups from guanine, reversing the effect of temozolamideMGMT methylation may predict responsiveness to temozolamide treatment.
15The Stupp Trial N=573 MS (months) 2 year survival (%) Recruited patients with GBM post surgery randomised toRT alone or RT with concomitant temozolomideN=573MS (months)2 year survival (%)3 year survival (%)RT alone12.110.43RT and Temo14.626.517Addition of chemotherapy to radiotherapy significantly prolongs survival among patients with newly diagnosed glioblastomaIncrease in survival rate at 2 years
16MGMT gene silencing and benefit from temozolomide in glioblastoma MGMT gene silencing and benefit from temozolomide in glioblastoma. Hegi ME, et al New Eng J Med 2005;352:Randomized trial comparing RT alone with RT combined with concomitant and adjuvant temozolamideFrom Hegi:206 tumours44.7% methylated55.3% unmethylatedMGMT methylated tumours: Median Survival 18.2 monthsNo methylation: Median Survival 12.2 monthsIrrespective of treatment, MGMT promoter methylation was an independentfavorable prognostic factor
17Bristol MGMT methylation analysis Main assay is a diplex of unmeth and meth productMuchofliteratureas separatesimplexesUnmethresultMethresultBristol MGMT analysis data to Late Jan 09To late Jan 09388 casesHegi et al N=206 Methylated 45%RTOG study Methylated 50%
182005 MGMT Survival data n=21 Methylated (n=12) Median Survival 15.5 mthsRange 0-31 mths 1 patient aliveUnmethylated (n=9)Median survival 10 mthsRange 1-36 months 1 patient aliveThe above date from 2005, before concomitant temozolomide was routinely available on the NHSThe above did not receive concomitant temozolomideNow NICE approval obtained for concomitant temozolamideIt is expected that survival gap will widen.
19How are the clinicians using these MGMT data now? Subgroups in Hegi study were too small to exclude a possible un-seen benefit for unmethylated patients so NICE recommendation is that all patients with GBM receive concomitant treatment.However – the outlook is clearly poor for patients in the unmethylated group, therefore the challenge is either to enhance the effectiveness of temozolomide or to find a better option.RTOG ‘dose dense’ temozolomide study – can MGMT be saturated with more prolonged treatment schedule?Bristol highest recruiters to trial in Western Europe
20Clinical Management in Bristol All patients with grade II-IV tumours have 1p19q analysis and MGMT analysedManagement is not affected by morphology but determined by grade and genetic resultsGenetic results may be particularly useful when the histological diagnosis is unclearGrade II: Surgery and follow up1p19q deletion informed of better prognosisAt progression offer BR13 trial (randomised for radiotherapy or temozolamide as initial treatment to look at OS,PFS and QOL )Trial declined? PCV if 1p deletions, RT if no deletionsGrade III: Surgery and immediate oncology treatmentIf have 1p19q deletions recommend PCV as first line treatment as patients with good prognosis may suffer late effects if cranial radiotherapyNo deletions recommend initial radiotherapyPatient factors and choice importantGrade IV: Surgery and immediate radiotherapy with concomitant temozolamide Standard regimen whilst awaiting results of dose dense study.Patients uncertain about chemotherapy are informed of methylation statusand Hegi data to inform choiceWhere MGMT promoter is unmethylated, patients advised that this tumour is particularly ‘aggressive’May consider using chemotherapy in patients with rare tumours and 1p19q LOH
21The Future Require robust funding streams and staffing – previous from LOH and MGMT testing now indicated as valuable predictive markers inRCPath Dataset for tumours of the CNS (2nd edition) April 2008Require robust funding streams and staffing – previous fromcharitable funds/training (NICE suggest 2500 annual tests and up to 30technical staff across England and Wales)Local clinicians are proud of the accurate information patients are given.Bristol is one of the biggest testing centres in UKIt is hoped to have better therapies for all patientsBristol patients in other trials –therapy for low grade gliomas (BR13 etc)1p and 1p19q LOH appears to be a marker for improved PFS in grade II gliomasRoutinely collecting fresh tissue for these assays has allowed Bristol to be prominent in national and international trials to inform future treatment (BR13 and CATNON trials etc). The local team will be able to implement any new guidelines speedily.Need to underpin this work through discussion of laboratory testing standards and guidelines to improve analysis, including tissue type (fresh v PPFE), choice of assay method(s), EQA and through availability of control reference materials
22Brain cancer network MDT We are part of the wider Bristol MDT network as outlined by NICELaboratory-histopathology-geneticsNeurosurgeonNeuropathologistNeuro-radiologistSpecialistoncologistPalliative careClinicalnurse specialistEpilepsy nurseAHPs for speech therapy, OT, physioBrain cancer networkMDTNeuro-psychologist
23Thanks to the Bristol Team BGL team Mark GreensladeElena MavrakiSarah Burton-JonesSuzanne O’SheaLaura YarramThais SimmonsPaula WaitsKayleigh McDonaghMaggie Williams(Emma Ryan/Karen Meaney/Meera Parmar)Neuropathology Seth Love, Neuropathology, FrenchayNeurooncology Kirsten Hopkins, Bristol Oncology CentreHugh Newman, UH Bristol