Schuuring SKML deelnemersdag feb 2013 EQA QCMD HPV
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Human Papillomavirus EQA Pilot Program (QCMD HPVDNA09/10/11/12) Ed Schuuring Pathology, UMCG, Groningen, NL Scientific expert and advisor for QCMD HPV Program Lid SKML-beleid, sector Pathologie Presentatie SKMLdeelnemersvergadering 5 februari 2013 HPV in scrapings of (pre)malignant cervical lesions ~4% high-risk HPV-positive ~85% high-risk HPV-positive Normal cervix CIN III 100% high-risk HPV-positive Nieuwe praktijkrichtlijn BVO cervixcytologie: indicatie HPV-onderzoek bij Pap2/3A1 Pap 1 97.1% Retour BVO 5 jaar HPV- Pap 2 Pap 3A1 2.1% Herhalen 6 maanden + HPV Pap 3A2 > 0,6% Bulkmans, the Lancet, 2007 Bais, Int J Cancer, 2005 Rebolj, Int J Cancer, 2007 Pap 1 70% Pap 2 Pap 3A1 25% Pap 3A2> 5% HPV+ HPV49% Herhalen 12 maanden HPV+ 51% Gynaecoloog J Eijsink The early detection of cervical cancer in scraping population-based screening programs worldwide 1) morphocytology only •The classical approach 2) primary morphocytology and HPV reflex testing •Presently used commonly (eg Dutch guidelines) 3) morphocytology/HPV co-testing •New guideline in USA (Saslow 2012) 4) primary HPV testing and reflex morphocytology •New guideline in NL (under review with minister) in 2013 ? Commercial HPV tests • • • • • • • • • • • • • • • • • • • • • • • • • • • • • • Hybrid Capture 2 Qiagen Digene HPV genotyping RH kit Qiagen Digene HPV genotyping LX kit, Qiagen Roche Amplicor HPV Test Roche Linear array HPV Genotyping test Innogenetics INNO-LiPA HPV Genotyping test NucliSens EasyQ HPV Biomerieux Aptima Gen-Probe Human Papilloma Virus reagents Third Wave BIOPAP QTS HPV Kit Loxo Reveal HPV Real-Time HPV Detection Kit GenoID AID STD assay GenID AID HPV screening kit GenID AID HPV typing kit GenID Linear ArrayExtra HPV Genotyping Kit Innogenetics PCR Human Papillomavirus Detection Set Takara Mirus Bio HPV DNA Chip Biomedlab Array Papillomavirus Genomica ProDect Chip HPV typing Bcs Biotech S.P.A PapType Genera Biosystems LCD Array HPV 3.5 Chipron Seeplex HPV Genotyping Seegene Viroactiv Virofem HPV OncoTest Invirion Diagnostics Genpoint Tm HPV test Dako-Oxoid Abbott RealTime High Risk HPV Abbott Luminex HPV Genotyping, Multimetrix/Progen Greiner PapilloCheck HPV Screening PreTect HPV Proofer Norchip ……… Analytical and clinical sensitivity of HPV-detection assays Clinical Clinical sensitivity sensitivity of of HC2 HC2 is is 5000 5000 HPV HPV copies copies Analytical Analytical sensitivity sensitivity of of PCRPCRbased based methods methods detecting detecting <10 <10 HPV HPV copies copies Adapted from Snijders et al. Journal of Pathology 2003; 201:1-6 Clinical validated HPV-tests (in the Netherlands) Digene HC2 HPV test (Qiagen) GP5+/GP6+ PCR EIA (Vumc/Qiagen) Cobas 4800 HPV test (Roche) Cervista hrHPV test (Hologic) GenProbe-Aptima hrHPV assay (Hologic) Abbott realtime HR HPV assay (Abbott) Kwantitatieve multiplex RT HPV test (PON) FDA-approved clinical HPV tests Digene HC2 HPV test (Qiagen) Cervista HPV HR test (Hologic) > internal control Cervista HPV 16/18 test (Hologic) > internal control GenProbe-Aptima hrHPV assay (Hologic) > based on RNA Cobas 4800 HPV test (Roche) > HPV16/18 separate and internal control available HPV EQA platforms 1) QCMD HPVDNA: • using established cell lines in LBC (4 HPV types) 2) WHO HPV panel: • Plasmid DNA spiked into cell line DNA (>30-45 types) 3) NEQAS UK: • Patient samples (~4 samples) Fagan, JClinVirol2010 QCMD 2009-2012 Human Papillomavirus DNA QCMD-HPV EQA Pilot Program Internationale rondzending: * Analytische sensitiviteit * Klinische sensitiviteit UMCG: Ed Schuuring (Scientific Advisory Board) UMCG: Lorian Slagter-Menkema (preparation/validation) Reference-labs: UMCG, UMCU, VUMC, UMCRadboud QCMD: Paul Wallace, Catherina di Lorenze QCMD = Quality Control for Molecular Diagnostics (Scotland) QCMD 2009/10/11/12 Human Papillomavirus DNA EQA Pilot Program doel Primaire doelstelling: •Aanbieden van een panel om de klinische-relevante sensitiviteit van de in lab gebruikte test te valideren Secondaire doelstellingen: •Aanbieden van een panel om de analytische sensitiviteit van de in lab gebruikte test te valideren •Aanbieden van een panel om de accuraatheid van de genotypering van de in lab gebruikte test te valideren •Door simulatie van klinische samples mbv “established” BMKH-cellijnen in dunne-laag-cytologie QCMD 2009/10/11/12 Human Papillomavirus DNA EQA Pilot Program participation HPVDNA09 HPVDNA10 HPVDNA11 HPVDNA12 (not finished) Participants 108 155 167 171 136 149 153 Responders 98 Countries 26 26 27 31 Deelnemers in Nederland 14 21 27 ? Datasets: - Analytical - Clinical - Genotyping 113 113 66 44 77 84 88 133 114 91 144 115 QCMD 2009 Human Papillomavirus DNA EQA Pilot Study panel composition of first EQA Clinical testing • 10 samples in original PreservCyt tubes • 7 dilutions of cervical cancer cell lines with approx 400 HPV16 copies/cells: HC2 was used to determine the clinical-relevant threshold • 2 positive controls (HPV18 and HPV67) and single HPV-negative control • all samples tested in a second reference-lab (VuMC, Amsterdam, Peter Snijders) QCMD 2011 Human Papillomavirus DNA EQA Pilot Study panel composition of first EQA Clinical testing • 9 samples in original PreservCyt tubes • 5 dilutions of cervical cancer cell lines with approx 400 HPV16 copies/cells HC2 was used to determine the clinical-relevant threshold • 2 positive controls (HPV18 and HPV45) and two HPV-negative controls • all samples pre-tested in reference-labs by PCR (1x), HC2 (2x), cobas (1x) HPVDNA11 – Clinical results 1: All participants HPVDNA11 – Clinical results 1: All participants 2: The Netherlands (8x HC2 en 15 PCR-based HPV tests) Rapportage van lab dat clinical testing uitvoert performance-score Rapportage van lab dat clinical testing uitvoert (Cumulative %) performance of clinical interpretation Clinical performance of all versus Dutch participants QCMD-HPVDNA2011 Performance of all participants Performance of Dutch participants QCMD 2011 Human Papillomavirus DNA EQA Pilot Study overall clinical performance Overall performance poor Expected to be below the HC2=1.0 threshold (representing analytical-pos/clin-neg sampeles Performance clicinal test poor (should be negative in all 100%) Performance analytical test is poor (should be negative in clinical setting (100%) QCMD 2011 Human Papillomavirus DNA EQA Pilot Study panel composition of first EQA Analytical testing • 9 samples in original PreservCyt tubes • 5 dilutions of cervical cancer cell lines with approx 400 HPV16 copies/cells HC2 was used to determine the clinical-relevant threshold • 2 positive controls (HPV18 and HPV45) and two HPV-negative controls • all samples pre-tested in reference-labs by PCR (1x), HC2 (2x), cobas (1x) 1: All participants 2: The Netherlands Analytical performance of all versus Dutch participants QCMD-HPVDNA2011 Performance of all participants Performance of Dutch participants QCMD 2009/10/11 Human Papillomavirus DNA EQA Pilot Study overall performance 2009 2010 2011 Analytical 100% Analytical 1x incorrect Analytical FP nd nd 5.1% 40.6% 77.0% 14.1% 59.1% 81.8% 4.5% Clinical 100% Clinical 1x incorrect Clinical FP nd nd 5.1% 45.5% 67.6% 13.0% 5.3% 27.9% 4.5% 1) High FP rate > 2/10 samples are true negative controls 2) Very low overall clinical performance > only high positive (Caski) with 50-500 HPV16copies/cell) and very low copy numbers (SiHa with ~2-3 HPV16-copies/cell Pre-test threshold van klinisch-relevante HPV assays Clinical HPV-tests: HC2: 1 pg/ml = ~5000 kopieën Abbott: = ~5000 kopieën Cobas = ~300 kopieën (afhankelijk van HPV-type) Dus eigenlijk kunnen we geen panel definiëren met een klinische threshold omdat HC2 niet meer de standaard is QCMD HPV testen vanaf 2013 (planning) Vanaf 2013 alleen toetsing performance van analytische interpretatie HPV-positieve, klinische relevante samples alleen als educatief samples in panel (eigen interpretatie op basis van performance van andere met vergelijkbare testen) Vanaf 2013 verwoording mbt klinische interpretatie verbeteren Kleinere panels en meer rondzendingen/jaar Andere matrices (SurePath, DNA, freeze-dry) Andere HPV-types, dubbel-/triple-infecties Ontwikkelen van referentie/calibratie-sets Advisory board is aangevuld met internationale assessors Dank voor uw aandacht Vragen ? 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