Minimal residual disease. Bruno Paiva University of Navarra, Spain

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1 Minimal residual disease Bruno Paiva University of Navarra, Spain 1st World Congress on Controversies in Multiple Myeloma. Bangkok, Thailand. May

2 Oprozomib (ONX 912) Phase I/II Autologous bone marrow transplantation High-dose therapy with autologous stem cell support Proteasome inhibitors Ixazomib (MLN 978) Phase II/III Oral melphalan and prednisone VAD (vincristine, doxorubicin, dexamethasone) Bisphosphonates Bortezomib Carfilzomib High-dose melphalan High-dose dexamethasone Thalidomide Lenalidomide Immunomodulatory drugs Pomalidomide MRD Monoclonal Antibodies Elotuzumab Phase II/III Daratumumab Phase I/II Adapted from Anderson KC, Bench to bedside translation of targeted therapies in multiple myeloma. ASCO 211.

3 What should be the treatment goal in MM patients? To search for an appropriate balance between treatment efficacy, toxicity & costs In very elderly patients (> 8-85y).to ensure QoL & avoid additional costs of expensive treatments In fit elderly patients (65-8y) & young ones with severe co-morbidities treatment goal should be to prolong survival and ensure QoL In young patients (<65y) In reference centers & large cooperative groups to investigate therapeutic schemes with a cure in the horizon

4 M-Protein, g/dl The paradigm of multiple myeloma 1 ASYMPTOMATIC SYMPTOMATIC 2 nd RELAPSE ACTIVE MYELOMA 5 2 MGUS or SMOLDERING MYELOMA PLATEAU REMISSION sustained disease control 1 st RELAPSE sustained disease control REFRACTORY RELAPSE Therapy Time Adapted from Durie BG 1 Prolongation of patients survival ultimately requires significant tumor depletion to sustaining/control minimal residual levels of disease for long periods of time 1. Durie BG. International Myeloma Foundation; 28:1-4

5 IMWG uniform response criteria Stringent CR CR VGPR PR SD CR as defined below, Normal FLC ratio and Absence of clonal PCs by immunohistochemistry Negative IFE of serum and urine Disappearance of any soft tissue plasmacytomas, and < 5% PCs in bone marrow Serum and urine M-component detectable by IFE but not on PEP 9% reduction in serum M-component plus urine <1 mg/24 h 5% reduction of serum M-protein and reduction in 24-hour urinary M-protein by 9% or to < 2 mg/24 hours If present at baseline, <5% reduction in the size of soft tissue plasmacytomas is also required Other criteria for non-measurable disease Not meeting criteria for CR, VGPR, PR, or PD PD Increase of 25% from lowest response value of serum and urine M- component Rajkumar SV, et al. Blood. 211;117(18):

6 Patients without event (%) Patients without event (%) Elderly patients (VISTA trial) RR (CR) (%): 71 (3) vs. 35 (4) TTP 1 8 CR TTP 1 8 VMP MP 6 4 PR P<,4 <PR 2 VMP: 24. months MP: 16.6 months, P< Time (months) Time (months) San Miguel JF et al. J Clin Oncol. 213;31(4): Harousseau JL, et al. Blood. 21;116(19):

7 Transplant-eligible patients (GIMEMA MMY-36 trial) Progression-free survival was significantly longer for patients receiving VTD than in those on TD (HR.63, p=.61) CR (%) VTD TD P-value After induction therapy 23% 6% <.1 After first ASCT 44% 3%.14 After second ASCT 49% 4%.131 After consolidation therapy 61% 47%.12 Cavo M, et al. Lancet. 21 Dec 18;376(9758): Cavo M, et al. Blood. 212;12(1):9-19

8 Patients responding (%) Selected induction regimens and response in MM Adapted, Stewart et al Blood 29 Courtesy of Dr. P. McCarthy. ASH Educational 213 Induction regimen 1. Lokhorst HM, et al. Haematologica. 28;93: Rajkumar SV, et al 28 J Clin Oncol 26: Harousseau JL, et al 21 J Clin Oncol 28: Rajkumar SV, et al Lancet Oncol 21; 11: Sonneveld P, et al J Clin Oncol 212; 3: Cavo M, et al Lancet 21; 376: Reeder CB, et al. Blood. 21; 115: Richardson et al. Blood 21;116: Jakubowiak AJ, et al Blood ;12: Palumbo A, et al. Blood. 212;12:[abstract 73]. 11. Kumar S, et al. Blood. 212;12:[abstract 332]. 12. Kumar S, et al. Blood :

9 Prognostic value of depth of response in myeloma Several trials in the transplant setting 1-1, non-transplant setting, after salvage therapie 18-22, or associated to specific stages of treatment (ie. consolidation 23 ) show a correlation between depth of response and progression-free survival Lonial S & Anderson KC. Leukemia. 213 Jul 19. doi: 1.138/leu CR does not mean cure and therefore treatment is not individualized to achieve CR Some patients that do not achieve CR have excellent outcome 1. Lahuerta JJ, et al. J Clin Oncol 28; 26: Moreau P, et al. Blood 211; 117: Harousseau JL, et al. J Clin Oncol 29;27: Galli M, et al. 25; 9: Barlogie B, et al. Cancer 28; 113: Dytfeld D, et al. Leuk Lymphoma 211; 52: Gertz MA, et al. Blood 21; 115: Kapoor P, et al. J Clin Oncol 211; 29: (abstract 869). 9. Alegre A, et al. Bone Marrow Transplant 1998; 21: Morgan GJ, et al. Blood. 211 Aug 4;118(5): Oivanen TM, et al. Eur J Haematol 1999; 62: Gay F, et al. Blood 211; 117: Kyle RA, et al. Cancer 26; 16: Offidani M, et al. Blood 26; 18: Harousseau JL, et al. Blood 21; 116: Palumbo A, et al. J Clin Oncol 27; 25: Mateos MV et al. Lancet Oncol 21 Oct;11(1): Harousseau JL, et al. Haematologica 21; 95: Niesvizky R, et al. Br J Haematol 28; 143: Palumbo A, Ann Oncol 28; 19: Pineda-Roman M, Leukemia 28; 22: Quach H, et al. Leuk Lymphoma 29; 5: Mellqvist UH et al. Blood 213;121:

10 Outcome of symptomatic MM with an MGUS-like signature independent of depth of response MGUS-like signature (symptomatic MM) (N = 59) TTP 1 8 Median: Not reached Symptomatic MM (N = 639) Higher BM tumor burden 6 4 Median: Not reached 2 P =.812 Unbalance ratio in favor of clonal PCs Time from diagnosis (months) CR (n=26) < CR (n=33) Identical results by GEP in Total Therapy protocols Zhan et al. Blood. 27;19: Paiva et al. Leukemia 213 (epub ahed of print)

11 Prognostic value of depth of response in myeloma Several trials in the transplant setting 1-1, non-transplant setting, after salvage therapie 18-22, or associated to specific stages of treatment (ie. consolidation 23 ) show a correlation between depth of response and progression-free survival Lonial S & Anderson KC. Leukemia. 213 Jul 19. doi: 1.138/leu CR does not mean cure and therefore treatment is not individualized to achieve CR Some patients that do not achieve CR have excellent outcome Some patients in CR relapse early on 1. Lahuerta JJ, et al. J Clin Oncol 28; 26: Moreau P, et al. Blood 211; 117: Harousseau JL, et al. J Clin Oncol 29;27: Galli M, et al. 25; 9: Barlogie B, et al. Cancer 28; 113: Dytfeld D, et al. Leuk Lymphoma 211; 52: Gertz MA, et al. Blood 21; 115: Kapoor P, et al. J Clin Oncol 211; 29: (abstract 869). 9. Alegre A, et al. Bone Marrow Transplant 1998; 21: Morgan GJ, et al. Blood. 211 Aug 4;118(5): Oivanen TM, et al. Eur J Haematol 1999; 62: Gay F, et al. Blood 211; 117: Kyle RA, et al. Cancer 26; 16: Offidani M, et al. Blood 26; 18: Harousseau JL, et al. Blood 21; 116: Palumbo A, et al. J Clin Oncol 27; 25: Mateos MV et al. Lancet Oncol 21 Oct;11(1): Harousseau JL, et al. Haematologica 21; 95: Niesvizky R, et al. Br J Haematol 28; 143: Palumbo A, Ann Oncol 28; 19: Pineda-Roman M, Leukemia 28; 22: Quach H, et al. Leuk Lymphoma 29; 5: Mellqvist UH et al. Blood 213;121:

12 Importance of achieving durable complete response (Results from TT2*) 1% Survival by 3 year CR 8% 6% 4% 2% % Median Deaths/N in years SUS-CR 28/256 NR NON-CR 63/ (4,6) LOS-CR 23/ (1,2) P value: a v b<.1, b v c <.1, a v c < Years from 3 years from enrollment SUS-CR:achieved and sustained CR status NON-CR:never achieved CR status LOS-CR:attained and lost CR status *Total therapy 2 regimen: Induction: VAD, DCEP, CAD, DCEP (TT2) Double transplantation: MEL 2 x 2 Consolidation: DCEP vs DCEP/CAD Maintenance: Interferon Randomization: thalidomide throughout vs no thalidomide Barlogie et al. Cancer 28;113:

13 Prognostic value of depth of response in myeloma Several trials in the transplant setting 1-1, non-transplant setting, after salvage therapie 18-22, or associated to specific stages of treatment (ie. consolidation 23 ) show a correlation between depth of response and progression-free survival Lonial S & Anderson KC. Leukemia. 213 Jul 19. doi: 1.138/leu CR does not mean cure and therefore treatment is not individualized to achieve CR Some patients that do not achieve CR have excellent outcome Some patients in CR relapse early on Similar CR rates are associated with different outcome Different CR rates do not translate into different outcome 1. Lahuerta JJ, et al. J Clin Oncol 28; 26: Moreau P, et al. Blood 211; 117: Harousseau JL, et al. J Clin Oncol 29;27: Galli M, et al. 25; 9: Barlogie B, et al. Cancer 28; 113: Dytfeld D, et al. Leuk Lymphoma 211; 52: Gertz MA, et al. Blood 21; 115: Kapoor P, et al. J Clin Oncol 211; 29: (abstract 869). 9. Alegre A, et al. Bone Marrow Transplant 1998; 21: Morgan GJ, et al. Blood. 211 Aug 4;118(5): Oivanen TM, et al. Eur J Haematol 1999; 62: Gay F, et al. Blood 211; 117: Kyle RA, et al. Cancer 26; 16: Offidani M, et al. Blood 26; 18: Harousseau JL, et al. Blood 21; 116: Palumbo A, et al. J Clin Oncol 27; 25: Mateos MV et al. Lancet Oncol 21 Oct;11(1): Harousseau JL, et al. Haematologica 21; 95: Niesvizky R, et al. Br J Haematol 28; 143: Palumbo A, Ann Oncol 28; 19: Pineda-Roman M, Leukemia 28; 22: Quach H, et al. Leuk Lymphoma 29; 5: Mellqvist UH et al. Blood 213;121:

14 CR: 25% P =.49 CR: 2% Palumbo et al. EHA 211 (oral presentation; plenary session) Similar results presented at the IMW213 comparing CyRD vs. HDT/ASCT Gay, F et al. IMW 213 Abstract 7 (oral presentation; plenary session)

15 Prognostic value of depth of response in myeloma Several trials in the transplant setting 1-1, non-transplant setting, after salvage therapie 18-22, or associated to specific stages of treatment (ie. consolidation 23 ) show a correlation between depth of response and progression-free survival Lonial S & Anderson KC. Leukemia. 213 Jul 19. doi: 1.138/leu CR does not mean cure and therefore treatment is not individualized to achieve CR Some patients that do not achieve CR have excellent outcome Some patients in CR relapse early on Similar CR rates are associated with different outcome Different CR rates do not translate into different outcome 1. Lahuerta JJ, et al. J Clin Oncol 28; 26: Moreau P, et al. Blood 211; 117: Harousseau JL, et al. J Clin Oncol 29;27: Galli M, et al. 25; 9: Barlogie B, et al. Cancer 28; 113: Dytfeld D, et al. Leuk Lymphoma 211; 52: Gertz MA, et al. Blood 21; 115: Kapoor P, et al. J Clin Oncol 211; 29: (abstract 869). 9. Alegre A, et al. Bone Marrow Transplant 1998; 21: Morgan GJ, et al. Blood. 211 Aug 4;118(5): Oivanen TM, et al. Eur J Haematol 1999; 62: Gay F, et al. Blood 211; 117: Kyle RA, et al. Cancer 26; 16: Offidani M, et al. Blood 26; 18: Harousseau JL, et al. Blood 21; 116: Palumbo A, et al. J Clin Oncol 27; 25: Mateos MV et al. Lancet Oncol 21 Oct;11(1): Harousseau JL, et al. Haematologica 21; 95: Niesvizky R, et al. Br J Haematol 28; 143: Palumbo A, Ann Oncol 28; 19: Pineda-Roman M, Leukemia 28; 22: Quach H, et al. Leuk Lymphoma 29; 5: Mellqvist UH et al. Blood 213;121:

16 Superior CR rates, but no differences in survival MRC Myeloma IX trial (non-intensive) GIMEMA MMY-36 Morgan GJ, et al. Blood. 211 Aug 4;118(5): Cavo M, et al. Blood. 212;12(1):9-19

17 Can single snapshot CR (or any depth-of-response) assessment produce a miracle? Induction Consolidation Maintenance No No R HDT/ASCT R Yes R Yes R Standard R Placebo Experimental Active drug Kinetics of response rather than a single assessment are more meaningful Historical landmark analysis do not take into consideration subsequent treatment strategies The most informative response assessments take place at the latest stages of treatment Ultimately, it needs to be really sensitive REDEFINE CR CRITERIA

18 Re-defining the CR criteria in MM < 5% PCs in bone marrow Negative IFE of serum and urine Disappearance of soft tissue plasmacytomas Cellular clonality Cellular production Cellular dissemination Immunohistochemistry Flow cytometry ASO-PCR NGS sflc Hevylite Isotype specific LC-MS/MS PET/CT WB-MRI

19 PFS According to PET-SUV Post-ASCT PET/CT in Patients Achieving CR Monitoring disease with PET/CT and WB-MRI 1. PET/CT retained prognostic relevance also in patients achieving CR (positive in 23%) % PET-SUV 1% reduction MRD by WB-MRI was positive in 18% of patients in CR, whereas MRI-CR was observed in 66% of patients in PR (and in 33% of patients with Months 3% PET-SUV <1% reduction Logrank P value=.195 SD/PD). 2 In how many patients in MRD is seen outside the BM? Is the baseline imaging mandatory? Is there inter-observer reproducibility? 1,3 False + (infection and/or inflammation) and false - (high-dose steroids) results 4 1. Zamagni et al. Blood 211;118(23): Hillengass et al. Haematologica 212; 97(11): Moreau et al. Blood 211;118(23): Zamagni et al. Br J Haematol 212;159:

20 Re-defining the CR criteria in MM < 5% PCs in bone marrow Negative IFE of serum and urine Disappearance of soft tissue plasmacytomas Cellular clonality Cellular production Cellular dissemination Immunohistochemistry Flow cytometry ASO-PCR NGS sflc Hevylite Isotype specific LC-MS/MS PET/CT WB-MRI

21 HLC ratios after treatment correlate with survival in symptomatic multiple myeloma All patients Patients in PR Patients in VGPR Ludwig et al. Leukemia 213;27(1): Early monitoring (after 2 cycles) of HLC patterns is an independent predictor of rapid response Bhutani M, et al. Blood 213;122( 21): abstract 762 HLC ratio as a surrogate marker of immune recovery after myeloablative transplantation, rather than as a marker of minimal residual disease Tovar N, et al. Biol Blood Marrow Transplant. 212;18(7):176-9.

22 Does the stringent CR criteria represent a deeper level of remission as compared to conventional CR? TTP in 115 patients in CR after induction (PETHEMA GEM5 65 & GEM5<65 trials) P=.19 P=.8 P< Time from diagnosis (months) Time from diagnosis (months) Time from diagnosis (months) stringent CR normal FLC 2-color FCM (IHC) - conventional CR abnormal FLC 2-color FCM (IHC) + Martinez-Lopez et al; IMW 213 (Abstract 415); poster presentation

23 Importance of achieving stringent CR after ASCT in multiple myeloma Fully defined stringent CR using sflc+ihc discriminates different outcomes among patients in CR Kapoor P et al. J Clin Oncol 213;31:

24 Re-defining the CR criteria in MM < 5% PCs in bone marrow Negative IFE of serum and urine Disappearance of soft tissue plasmacytomas Cellular clonality Cellular production Cellular dissemination Immunohistochemistry Flow cytometry ASO-PCR NGS sflc Hevylite Isotype specific LC-MS/MS PET/CT WB-MRI

25 IHC for the detection of (minimal?) residual disease? Trephine biopsy: BM clonality is defined when the Κ/λ ratio is >4:1 or <1:2 for Κ and λ patients, respectively, after counting 1 PCs Kappa probe X2 Normal PCs Lambda probe X2 Clonal PCs The majority of Κ/λ ratios after chemotherapy are polyclonal, and clonality can only be detected after identification of phenotypically aberrant cells - BM biopsies is not a world-wide standard procedure

26 MRD monitoring by NGS - superior applicability and sensitivity - Among CR (n=62), NGS discriminated two risk groups with different TTP (131 vs 35months, P <.1) but no differences for overall survival (both medians NR, P =.1) Similar results for TTP/OS obtained by 4-color flow cytometry NGS+ MFC- patients have inferior TTP (P =.48) compared to NGS- MFC- cases Martinez-Lopez J, et al. Blood 213;122: abstract 1848

27 MRD monitoring by 4-color flow: patients >65y 44 patients in CR after induction therapy (GEM25MAS65) TTP OS Median: NR 8 Both medians NR Median: 29m P =.49 P = Time from MRD assessment (months) Time from MRD assessment (months) Flow CR (n=24) MRD positive (n=2) Paiva B et al; J Clin Oncol. 211;29(12): (f/u updated March 214)

28 MRD monitoring by 4-color flow: patients <65y 12 patients in CR after HDT/ASCT (GEM25NENOS65) TTP 1 OS 1 Both medians NR 8 6 Median: 5m Median: 27m P =.1 P = Time from MRD assessment (months) Time from MRD assessment (months) 9 Flow CR (n=68) MRD positive (n=34) Paiva et al. Blood ; abstr 191 (f/u updated April 214)

29 MRD monitoring by 6-color flow: patients <65y 214 patients in CR after HDT/ASCT (MRC myeloma IX) PFS p-value for logrank test =.18 Non-significantly inferior OS (median 61.9 months vs. not reached; P =.928) Time from MRD assessment (months) Flow CR (n=183) MRD positive (n=31) Rawstron A, et al. J Clin Oncol. 213;31(2):254-7.

30 Why is it so difficult to see differences on OS? (GEM25MENOS65) 232 patients tested for MRD (GEM25MAS65) 153 patients tested for MRD OS OS 1 Both medians: NR 1 Both medians: NR P =.3 P = Time from MRD assessment (months) Time from MRD assessment (months) 1 Flow CR (n=111) MRD positive (n=121) Flow CR (n=34) MRD positive (n=119)

31 MRD monitoring by 4-color flow: patients <65y 125 patients in CR after HDT/ASCT (GEM2) TTP 1 OS Median: 141m Median: 62m Median: 61m P <.1 Median: 36m P < Flow CR (n=71) MRD positive (n=57) Paiva B et al; Blood. 28; 15;112(1): (f/u updated July 212)

32 Re-defining the CR criteria in MM < 5% PCs in bone marrow Negative IFE of serum and urine Disappearance of soft tissue plasmacytomas Cellular clonality Cellular production Cellular dissemination Immunohistochemistry Flow cytometry ASO-PCR NGS sflc Hevylite Isotype specific LC-MS/MS PET/CT WB-MRI How to choose?

33 Published/reported data EXCLUSIVELY among patients in CR PFS OS stringent CR 1 Flow cytometry 2-9 ASO-PCR 8,1,11 NGS 9 PET/CT Kapoor P et al. J Clin Oncol 213;31: Rawstron AC, et al. Blood 22; 1(9): Paiva B et al; Blood. 28; 15;112(1): Paiva B et al. Blood 21;116: abstract Paiva B et al; J Clin Oncol. 211;29(12): Paiva B, et al. Blood. 212;119: Rawstron A, et al. J Clin Oncol. 213;31(2): Puig N et al. Leukemia doi: 1.138/leu Martinez-Lopez J, et al. Blood 213;122: abstract Martinelli G, et al. J Clin Oncol 2;18: Corradini P, et al. Blood. 23;12: Zamagni et al. Blood 211;118(23):

34 Unmet need to consistently go beyond 1-5 GEM2/GEM25 GEM21 GEM events 2.. events > 4.. events MRD+ ve :.1% (1-5 )

35 % of tumor cells Higher sensitivity uncovers the unmeet need for higher resolution Polyclonal Clonal Background Protein monoclonal vs. oligoclonal spep atypical abnormal sflc ratios Imaging false + (infection; inflammation) false - (high-dose steroids) Cellular new normal phenotypes disproportional amplification of different rearrangements

36 Standardization of multidimensional flow cytometry to implement Flow-CR as a clinical end-point Software guided PC identification and computerized classification of normal vs. aberrant PCs Myeloma PC reference Normal PC reference

37 Do I have available baseline sample/studies? Can imaging- or NGS- based MRD monitoring be performed w/o baseline evaluation? CLL/MBL-like clone in ~5% of MM patients Ig sequencing Paiva B, et al. unpublished observations

38 In which patients and where should MRD be investigated?

39 GEM2+GEM25<65y: Impact on survival of achieving immunophenotypic CR according to cytogenetics PFS 1 8 Standard-risk / MRD- (n=88) High-risk / MRD- (n=82) Standard-risk / MRD+ (n=119) High-risk / MRD+ (n=25) 6 4 Standard-risk / MRD- vs Standard-risk / MRD+; P <.1 vs High-risk / MRD-; P =.462 High-risk /MRD- vs Standard-risk / MRD+; P=.135 vs High-risk / MRD+; P = Identical results in the MRC myeloma IX study Rawstron A, et al. J Clin Oncol. 213;31(2):254-7.

40 Where could Flow-CR be clinically relevant? Pretreatment prognostic factors MRD HDT/ASCT TD Induction TD TD Consolidation

41 Not all patients in CR after induction look the same at the MRD level 22 patients in CR after HDT/ASCT: monitoring stem cell harvests PFS: GEM2 & GEM25<65y (patients in CR after induction) TTP Flow CR (n=6) MRD (-) MRD (-) 1 1 MRD (+) MRD (-) MRD positive (n=16) MRD (+) MRD (+) 8 6 Median: 85m TTP 8 6 P =.84 4 Median: 1m P = Time from MRD assessment (months) Time from diagnosis (months) Similar results by PCR: Takamutsu H, et al. Exp Hematol. 213;41(1): Paiva B, et al. (unpublished)

42 Where could Flow-CR be clinically relevant? Pretreatment prognostic factors MRD HDT/ASCT TD F/up TD Induction TD TD Maintenance Consolidation

43 Proportion event free MRC myeloma IX trial: What about maintenance therapy after HDT/ASCT? Progression-free survival by 1-day outcome assessed in all patients by MRD/maintenance; T: thalidomide; NM: no maintenance p-value for logrank test =.36 T; MRD+ NM; MRD Rawstron A, et al. J Clin Oncol. 213;31(2):254-7.

44 Maintenance alone did not overcome unsustained CR TTP after day+1 HDT/ASCT OS after day+1 HDT/ASCT 1y 2.5y 5y % 8% 89% 76% 1 8 Median: NR % Median: 83m 4 Median: 47m 2 % 2 Median: 21m Median: 6m Median: 28m P <.1 P < Months Months 8 1 * t(4;14), t(14;16) and/or del(17p) Standard-risk FISH + MRD negative (n=58) High-risk* FISH OR MRD positive (n=45) High-risk* FISH + MRD positive (n=7) Paiva B, et al. Blood. 212;119:

45 Where could Flow-CR be clinically relevant? Pretreatment prognostic factors MRD HDT/ASCT TD F/up TD Induction TD TD Maintenance Consolidation TD F/up

46 Impact of MRD after consolidation with VTD Patients: (n=39) with VGPR after consolidation Ladetto M et al. J Clin Oncol. 21;28:277. Ladetto M et al. Blood. 211;118: Abstract 827.

47 Where could Flow-CR be clinically relevant? Pretreatment prognostic factors MRD HDT/ASCT TD F/up TD TD Induction TD TD Maintenance TD Preventive treatment Consolidation TD F/up TD

48 Can we turn myeloma into CML? Treatment prolonged until MRD clearance or stabilization (plateau) R Continuous treatment consolidation/maintenance (+/- 4 or 6 m) and stop

49 Should these patients be re-treated (e.g.: MoAb±IMiD?) 1. Rocci A et al. Blood 213;122: abstract Ferrero S et al. Blood 213;122: abstract Kosuri S et al. Blood 213;122: abstract Ladetto M et al. Blood 211;118: abstract 827 Can we turn myeloma into CML? 31 patients in CR + Flow CR after up-front treatment MRD monitoring during f/up MRD positive: 85% clinical relapse Flow CR: 39% clinical relapse P =.1 Paiva B et al. manuscript ijn preparation Immunophenotypic relapse anticipate clinical relapse 1 There is a 9 months lag between molecular CR loss and need for salvage treatment 2 Flow cytometry permits the detection of MRD preceding frank relapse 3 A dynamic increase in molecular tumor burden detectable by RQ-PCR, predicts late disease relapses several months before clinical recurrence 4

50 Where could Flow-CR be clinically relevant? Pretreatment prognostic factors MRD Relapse HDT/ASCT TD F/up TD TD Induction TD TD Maintenance TD Preventive treatment Salvage treatment TD Consolidation TD F/up TD

51 MRD monitoring by 4-color flow: R/R patients 43 patients in CR after salvage therapy (81% treated w/o novel agents; 63% received SCT) TTP 1 OS 1 Myeloma-related deaths: 13m vs. 41m (P =.5) Median: 25m 6 Median: NR 4 4 Median: 38m 2 Median: 13m 2 P =.35 P = Time from MRD assessment (months) Flow CR (n=2) Time from MRD assessment (months) MRD positive (n=23) BSBMT/Ukmf Myeloma X (patients in CR after salvage therapy) TTP: 25 vs. 1m (P <.1) Ashcroft J et al; Blood 213;122 (21): abstract Paiva B. et al. EHA 213; abstract P219; median f/up: 16 months

52 Where else can MRD be of help in MM? OS Standard-risk (8%): 6-8 years median OS 1 8 High-risk (2%): 2-3 years median OS - t(4;14) - t(14;16) - del(17p) - GEP signature 6 4 Many novel agents being developed - monoclonal antibodies - deacetylase inhibitors - many more 2 Wait 8-years to demonstrate improvement in OS of standard-risk patients? P < Time from diagnosis (months) 125 Wait for early dead to demonstrate lack of improvement OS in high-risk patients?

53 PETHEMA/GEM & MRC: same PFS independent of previous induction regimen PFS GEM2: MRD - (n=71) MRD + (n=54) GEM25<65y: MRD - (n=68) MRD + (n=34) Medians: 62m & 59m P =.21 PFS 4 2 Medians: 36m & 39m P = Time from diagnosis (months) Rawstron A, et al. IMW 213 Abstract 16 (oral presentation; plenary session) Paiva et al. Blood ; abstr 191. (updated July 212)

54 State of the art therapy should go along with state of the art response criteria HDT/ASCT Thalidomide Bortezomib Lenalidomide Carfilzomib Pomalidomide MoAbs?? Year Criteria EBMT (Blade) 1 IMWG 2 IMWGv2 3? Depth of response CR Stringent CR Flow CR Molecular CR Flow CR Molecular CR NGS CR PET/CT Flow CR NGS CR Imaging CR