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Reference reagents for monitoring minimal residual disease

Reference reagents for monitoring minimal residual disease. Jean Gabert, MD, PhD ERT MEIDIA & APHM Marseille, France Gabert.j@jean-roche.univ-mrs.fr http://meidia.nord.univ-mrs.fr SoGAT XVII, Paris, May 27th. Acute leukemia and CML Concept of minimal residual disease (MRD). Cell number.

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Reference reagents for monitoring minimal residual disease

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  1. Reference reagents for monitoring minimal residual disease Jean Gabert, MD, PhD ERT MEIDIA & APHM Marseille, France Gabert.j@jean-roche.univ-mrs.fr http://meidia.nord.univ-mrs.fr SoGAT XVII, Paris, May 27th

  2. Acute leukemia and CMLConcept of minimal residual disease (MRD) Cell number Morphological detection limit 1012 1010 PCR detection limit 106 I Time

  3. New molecular-based therapies • Therapeutic breakthrough O Brien et al NEJM, 2003; 348; 994-1004 Gleevec a tyrosine kinase inhibitor and CML (BCR ABL) • Therapeutic follow up on cytogenetic: t(9;21) or Ph Chr. • - CCR status (18 months): • Interferon + ARA C: 14.4% • Gleevec 74% p<0,0001 • Biological survey: • Hughes TP et al NEJM 2003; 349: 1399-401 • 3 laboratories • reference value: pool of 30 CML at diagnosis • Responder: reduction of at least 3 logs by 12 months. • Sudden need of reference biological material and Quality Controls

  4. • • • • • • • • • • • • • • • • • ‘Europe Against Cancer’ Network • Fusion Transcripts (n=9) & RQ PCR • For leukemias • Standardization • Quality Controls • • • • • • • • • • • • • • • • • • • • • • • Partners: • SANCO European Commission • Applied Biosystems • ABI 7 700 • IPSOGEN • plasmid calibrators • • • • • • • • • • • •

  5. E. Beillard et al & Gabert J et al Leukemia, dec 2003

  6. developpment of a biological reference material • Rational • Allows data comparison between and within plat-forms / laboratories • Needed for research, • Strategic for network programs, • Indispensable for medical use (ex. therapeutic decision made on gene profiling signatures) • Collaboration ERT MEIDIA & NIBSC (UK) • NAT Standards in virology (Dr. J. Saldanha) • Freeze dried cells? • can be sent at room temperature worldwide • French patent n°03/03199

  7. Phase 3 International study Seattle Los Angeles Sendai Houston Uppsala Turku Adelaide Stockholm Arhus Dublin Rotterdam London Frankfurt, Mannheim Paris Vienna Torino, Monza Bordeaux Bologna Marseille Salamanca Naples Athens

  8. Freeze-dried cell lines International standards Ratio BCR ABL/ABL dosage by RQ PCR Neat 1/10 1/100 => reproducible

  9. Accelerated ageing experiments NCN = normalized copy number => stable for 3 to 5 years

  10. Control population 97 clones IPSOGEN Gene profilingreproducibility(2 K562 freeze dried cell vials)on nylon membrane, cDNA (10 000 spots) 54 clones

  11. MRD by gene expression analysis (RQ PCR)and Hematological malignancies • Towards the definition of new biological makers • Potentially available wapon within the medical arsenal against hematological malignancies • at diagnosis (Lossos et al NEJM, 2004) • For therapeutic efficiency monitoring (MRD)

  12. reference materials • Required forinnovative technologies development : • gene expression profiling, CGH array, SNP … • Proteomics, epigenetic event analysis… • Pool of freeze-dried cell lines • Stable material (RT) • Reproducible • Standardization tool +++ • Controls each step of the process from the extraction => WHO international standards for K562 freeze dried cells => the SAQUITO project & national cancéropôle network

  13. MEIDIA laboratory E. Beillard C. Picard M Silvy N. Chaigneau U387 INSERM C. Genre IPSOGEN V. Fert French networks E. Macintyre (GBMHM) JM Cayuela (Costly Innovation) EAC network JJM van Dongen International study J. Saldanha (NIBSC) Acknowledgments

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