1. MRD in myeloma UKMF Spring Day Assessment of disease response, CR and beyond. Roger Owen St James’s Institute of Oncology Leeds, UK

2. Myeloma trials: the challenges Complex multicomponent therapy Increasing survivalM protein issues

3. What do we need? Applicability Consensus on methodology Predicts PFS and OS including CR pts Cytogenetic risk groups Transplant eligible and ineligible Upfront and relapse Independent of treatment

4. Flow cytometry in MM. Minimum four colour method Minimum four colour method Gating using CD38, CD138 and CD45 Gating using CD38, CD138 and CD45 MRD+ defined by a minimum of 100 events (10 6 total events acquired for a sensitivity of 0.01%) MRD+ defined by a minimum of 100 events (10 6 total events acquired for a sensitivity of 0.01%) Clonality assessment suboptimal for MRD due to the presence of normal cells in post treatment samples Clonality assessment suboptimal for MRD due to the presence of normal cells in post treatment samples Aberrant phenotype defined by CD19 and CD56 Aberrant phenotype defined by CD19 and CD56 Leeds - CD138/CD38/CD45/CD19/CD56/CD27 Applicability ~97%

5. Value of flow cytometry in the routine setting Confirmation of a diagnosis of myeloma - good practice c.f. acute leukaemia - immunohistochemistry on trephine sections only needed in limited situations - saves time and money Differential diagnosis of MGUS and MM Outcome prediction – MGUS, smouldering MM and plasmacytoma Amyloidosis Rare / difficult cases Response assessment

6. Flow cytometry in AL amyloidosis. 97% of patients have aberrant phenotype PCs Paiva B et al. Blood 2011;117:3613-3616

7. Perez-Persona, E. et al. Blood 2007;110:2586-2592 Updated BJ Haem epub October 2009 Progression in MGUS (A) and SMM (B) – Salamanca data Adverse risk defined by >95% aberrant phenotype plasma cells.

8. 2002!

10. MRC Myeloma IX— Trial Design Intensive Clodronate CVAD Clodronate CVAD Zoledronic acid CVAD Zoledronic acid CVAD Clodronate CTD Clodronate CTD Zoledronic acid CTD Zoledronic acid CTD MEL-200 ASCT MEL-200 ASCT –Thal +Thal Non-intensive Clodronate MP Clodronate MP Zoledronic acid MP Zoledronic acid MP Clodronate CTDa Clodronate CTDa Zoledronic acid CTDa Zoledronic acid CTDa Maximal Response Maximal Response –Thal +Thal N = 1,960 RANDOMISATION

13. Effect of MRD according to cytogenetic risk profile

14. Paiva B et al. Blood 2012;119:687-691

16. What about salvage Rx? Ashcroft et al, ASH 2013 Paiva et al, Haematologica. 2015;100(2):e53-5.

17. Impact of therapy received 12345678 20 40 60 80 100 TIME (YEARS) % PFS MRD- CVAD n = 113 MRD+ CVAD n = 95 MRD- CTD n = 134 MRD+ CTD n = 55 2 3 = 24.30 P <.00001 de Tute et al, submitted

18. Outcome determined by level of disease not treatment received

20. CR patients only?

21. MRD predicts outcome in CR patients. Paiva et al. Blood 2008;112:4017-4023

22. MRD and M protein response Rawstron et al, 2015

23. Bruno Paiva et al. Blood 2008;112:4017-4023 MRD and M protein response

24. MRD and M protein response.

25. Multivariate analysis. Rawstron et al, 2015

28. MRD: Comparison of induction regimens. CVADCTD Post induction (n=252) 13%25%P=0.004 Day 100 (n=397) 54%71%P<0.0001

29. Ongoing role of ASCT? Rawstron et al, J Clin Oncol. 2013 ;31(20):2540-7 Paiva B et al. Blood 2008;112:4017-4023

30. Maintenance

31. No change in conventional response with thalidomide maintenance but clear differences in neoplastic plasma cell levels “Using electrophoresis and immunofixation as a monitoring technique, there was no difference between the thalidomide maintenance and no maintenance arms in the percentage of patients that upgraded response status over time (P.19).” (1) 27.6 96 3.4 68.8 0 20 40 60 80 100 Become MRD negativeRemain MRD negative Thalidomide maintenance No maintenance (2) 1.Morgan et al, Blood 2012, 119(1): 7-15 2.Rawstron, JCO 2013; 31(20):2540-7

33. 123456789100 0.2 0.4 0.6 0.8 1.0 0 Years from Diagnosis Proportion free from disease progression P=0.003 Normal phenotype plasma cells Neoplastic phenotype plasma cells Outcome prediction in SPB Hill et al, Blood 2014 IMWG – “solitary plasmacytoma with minimal marrow involvement” IDRIS study of Len-Dex in high-risk patients

34. Conclusions. MRD assessment is highly predictive of outcome CR patients Standard and adverse risk cytogenetics Presentation and relapse ASCT and non-ASCT Assessment of individual components of multicomponent treatment schedules and maintenance strategies Thalidomide / Bortezomib eradicates MRD in a proportion of patients

35. University of Birmingham MT Drayson K Walker A Adkins N Newnham Wessex Regional Genetics Laboratory, Salisbury F Ross L Chieccio LTHT, Leeds G Cook S Feyler D Bowen HMDS, Leeds RG Owen AC Rawstron R de Tute M Dewar S Denman ICR, London FE Davies M Jenner B Walker D Johnson D Gonzalez N Dickens K Boyd P Leone L Brito A Avridromou MRC Leukaemia Trial Steering Committee MRC Leukaemia Data Monitoring and Ethics Committee NCRI Haematological Oncology Clinical Studies Group NIHR, through the National Cancer Research Network UK Myeloma Forum Clinical Trials Committee Myeloma UK Funding Medical Research Council Pharmion Novartis Chugai Pharma Bayer Schering Pharma OrthoBiotech Celgene Kay Kendall Leukaemia Fund Chief Investigators JA Child GJ Morgan GH Jackson CTRU, Leeds K Cocks W Gregory A Szubert S Bell N Navarro Coy F Heatley P Best J Carder M Matouk D Emsell A Davies D Phillips A Gillman L Flanagan C Tyas and others Acknowledgements