Dati sulla sospensione della terapia

Leucemia Mieloide Cronica Dati sulla sospensione della terapia Gianantonio Rosti, Bologna

BCR-ABL Loading in CML Patients 100% 10% 1% MMR MR 4 MR 4.5

STIM study design N=100 Start Imatinib CMR Sustained CMR for 2 years STOP Q- RT-PCR from peripheral blood every month in the first year and every 2 months thereafter Five BCR ABL analyses by Q- RT-PCR during these 2 years Molecular recurrence: positivity of BCR ABL transcript in Q-RT-PCR confirmed by a second analysis point indicating the increase of one log in relation to the first analysis point, at two successive assessments, or loss of MMR at one point. Sixth datapoint checked in centralized laboratory Mahon FX et al. The Lancet Oncology, 2010;11(11): 1029-1035.

Treatment-free survival At 60 months 40% ( 95% CI: 30-49) Median follow up: 55 months (range 9-72) Mahon, ASH 2013

Follow-up of the non-molecular relapse patients (n= 39) Follow-up: mean 56, median 58 (9-72) months 38 patients are still in CMR i.e., UMRD (sensitivity > 4.5 log) 1 patient died in CMR after 9 months of imatinib cessation due to myocardial infarction

Regression model # of molecular relapse at 24 months Univariate analysis P value Multivariate analysis (final model) Gender 0.103 0.08 Sokal score 0.0076 0.0037 Interferon 0.2426 - Time to CMR 0.2880 - CMR duration 0.2345 - IM duration 0.1357 0.1 - # Acounting for competing risk factors included when p<0.2 in univariate analysis

Survival Without Molecular Relapse STIM: Low Sokal Score and Previous Imatinib 5 y Are Prognostic For Reduced Relapse Risk 1.0 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0.0 0 3 6 9 12 15 At 24 Months: Sokal Low + IM > 5 y: Others 68% (95%Cl: 45 83) 33% (95%Cl: 22 42) 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60 Months Since Discontinuation of Imatinib P =.007 Multivariate analysis (Cox model) Hazard ratio (95% CI) P value Sokal score 2.555 (1.278-5.119) 0.008 IM duration ( 5 y vs < 5 y) 0.582 (0.340-0.995) 0.047 Mahon FX, et al. Blood 2011;118:abstract 603.

Sustained CMR after stopping imatinib according to duration of CMR before cessation 78% vs. 15%, P =.0002 by Log-rank test Takahashi N et al., Haematologica, 2012; 97:903-06.

Stem cell persistence is associated with low BCR-ABL expression Apoptosis CFU Kumari A et al, Blood. 2012 Jan 12;119(2):530-9.

Hypothetical Model of CML persistence and recurrence versus extinction The eradication of the leukemic clone may depend on inherent features of the disease or on the duration of therapy, or both. Deininger, M. Nat. Rev. Clin. Oncol. advance online publication 1 February 2011; doi:10.1038/nrclinonc.2011.17

Treatment-free survival At 60 months 40% ( 95% CI: 30-49) Median follow up: 55 months (range 9-72) Mahon, ASH 2013

Summary of treatment suspension trials Study Therapy Duration of tx Inclusion criteria and duration N patients Primary Endpoint Molecular Relapse Rate STIM 1 (IFN) IM >3 years Stable CMR > 2 years STIM2 2 IM >3 years Stable CMR >2 years A-STIM 3 IM >3 years Stable CMR >2 years STOP 2G-TKI 4 2TKI: I or II line >3 years Stable CMR >2 years TWISTER 5 (IFN) IM >3 years Stable CMR >2 years EUROSKI DASFREE ENEST Freedom ENESTop IM, 2TKI I or II line DAS I or II line >3 years Stable MR4.0 >1 year > 2 years Stable MR4.5 >1 year NIL >3 years Stable MR4.5 >1 year IM NIL NIL IM NIL >3 years ENESTPath IM NIL IM 2 NIL 2 vs 3 y Stable MR4.5 >1 year Stable MR4.0 >1 vs 2 years 100 Survival without loss of CMR 124 Survival without loss of CMR 80 Survival without loss of MMR 33 Survival without loss of MMR 40 Survival without loss of CMR 500 Survival without loss of MMR 74 Survival without loss of MMR 206 Survival without loss of MMR 117 Survival without loss of MR4.0 1058 Survival without loss of MMR 61% at 5 year 38.7% at 1 year 39% at 3 years 38.9% at 1 year 52.9% at 2 years / / / / / 1.Mahon et al ASH 2013. 2.Mahon et al Blood (2013) abs #654 3.Rousselot et al J.Clin Oncol (2013) e-pub ahead of print. 4.Rea et al Blood (2012) abs #916 5.Ross et al Blood (2013) 122:515-522

Summary of treatment suspension trials Study Therapy Duration of tx Inclusion criteria and duration N patients Primary Endpoint Molecular Relapse Rate STIM 1 (IFN) IM >3 years Stable CMR > 2 years STIM2 2 IM >3 years Stable CMR >2 years A-STIM 3 IM >3 years Stable CMR >2 years STOP 2G-TKI 4 2TKI: I or II line >3 years Stable CMR >2 years TWISTER 5 (IFN) IM >3 years Stable CMR >2 years EUROSKI DASFREE ENEST Freedom ENESTop IM, 2TKI I or II line DAS I or II line >3 years Stable MR4.0 >1 year > 2 years Stable MR4.5 >1 year NIL >3 years Stable MR4.5 >1 year IM NIL NIL IM NIL >3 years ENESTPath IM NIL IM 2 NIL 2 vs 3 y Stable MR4.5 >1 year Stable MR4.0 >1 vs 2 years 100 Survival without loss of CMR 124 Survival without loss of CMR 80 Survival without loss of MMR 33 Survival without loss of MMR 40 Survival without loss of CMR 500 Survival without loss of MMR 74 Survival without loss of MMR 206 Survival without loss of MMR 117 Survival without loss of MR4.0 1058 Survival without loss of MMR 61% at 5 year 38.7% at 1 year 39% at 3 years 38.9% at 1 year 52.9% at 2 years / / / / / 1.Mahon et al ASH 2013. 2.Mahon et al Blood (2013) abs #654 3.Rousselot et al J.Clin Oncol (2013) e-pub ahead of print. 4.Rea et al Blood (2012) abs #916 5.Ross et al Blood (2013) 122:515-522

BCR-ABL transcripts in patients remaining in MMR STOP 2G-TKI Patients in MMR without therapy (median follow-up 17 months: 7-37) n=23 Always undetectable 7/23 (30.4%) Occasionally detectable on 1 test 8/23 (34.8%) Occasionally detectable on 2 consecutive tests 8/23 (34.8%) Rea, ASH2013 DR

Patient characteristics A-STIM* STIM Patients, n 80 100 Sokal Score, % Gender, M/F ratio 0.52 0.6 Median age at diagnosis, y (range) 56 (27 78) 62 (29 80) Low risk 52 49 Intermediate risk 27 39 High risk 18 11 Previous therapy, n (%) Baseline characteristics in A-STIM and STIM Interferon (or IFN/Ara-C) 43 (53.1) 34 (44.9) Autologous HSCT 3 (3.7) 0 Median duration of imatinib, mo (range) 77 (30 145) 50 (36 90) Median duration of CMR, mo (range) 40.1 (20 96) 35.5 (24 85) Median follow-up after imatinib discontinuation, mo (range) 22 (1 97) 34 (9-50) Rousselot P et al. 2014 JCO Mahon FX et al., Lancet Oncol., 2010; 11:1029-35 & Blood 2011; 118 [abstract 603].

Kaplan-Meier estimate of TFS defined as loss of MMR and defined according to STIM criteria after imatinib discontinuation in 81 CML patients. 100 Percent survival 80 60 40 20 65% 37% P = 0.003 0 0 12 24 36 48 60 72 84 96 108 Months Rousselot P et al., JCO 2014 TFS= treatment free survival

Cumulative Incidence % How many patients on imatinib achieve deep responses? 100 80 60 40 20 0 Confirmed undetectable MR 4.5 52% (CI 43-60%) n=423 patients 0 1 2 3 4 5 6 7 8 Years after commencing imatinib 37% Stable 2 years Branford et al, Blood 2013;121:3818

Deep Molecular Response METHODS: - QPCR every 6 months - MR 4.0 : 0.01% BCR-ABL IS or undetectable BCR-ABL transcript with 10,000 ABL transcripts Cumulative incidence of MR 4.0 342/559 (61%) Stable MR 4.0 (> 18 mos; > 3 samples) 108/559 (19%) Fluctuating MR 4.0 in stable MMR 153/559 (27%) Episodic MR 4.0 (one sample only) 81/559 (14%) Castagnetti on begalf of GIMEMA, ASH 2013 GIMEMA CML WP

Cumulative Incidence % Factors associated with deep responses 100 80 60 40 20 strongest independent predictor was the 3 month BCR-ABL1 value, P <.001 0 Stable undetectable MR 4.5 for 2 years n=423 Susan Branford, GOLS 2014 0 1 2 3 4 5 6 7 8 Years after commencing imatinib MMR, 78% >0.1-1.0%, 53% >1-10%, 29% >10%, 9% Branford et al, Blood 2013;121:3818

Patients With MR 4.5, % Patients With MR 4.5, % Proportion of Patients With MR 4.5 by BCR-ABL Levels at 3 Months Nilotinib 300 mg BID Imatinib 400 mg QD BCR-ABL IS 1%: 56% of pts BCR-ABL IS 1%: 16% of pts 100 90 80 70 60 50 40 30 20 10 0 BCR-ABL Level 1% > 1% to 10% > 10% Pts 144 89 24 58% P =.0001 28% P =.0135 4% MR 4.5 by 4 Years a MR 4.5 by 5 Years a 70% 52% 8% P =.0046 P =.0001 0 1 2 3 4 5 6 Time Since Randomization, Calendar Years 100 90 80 70 60 50 40 30 20 10 0 BCR-ABL Level 1% > 1% to 10% > 10% Pts 43 133 88 MR 4.5 by 4 Years a 65% P <.0001 24% 5% MR 4.5 by 5 Years a 67% 34% P =.0001 P =.0016 P = 15%.0001 0 1 2 3 4 5 6 Time Since Randomization, Calendar Years Patients with BCR-ABL 1% at 3 months have significantly higher rates of MR 4.5 by 5 years More patients achieve BCR-ABL 1% at 3 months on nilotinib 300 mg BID vs imatinib a Cumulative response rates reported consider each year to consist of twelve 28-day cycles. 20 Saglio G, et al. Blood. 2013:[abstract 92].

Cumulative Rate of MR 4 and MR 4.5 Dasatinib 100 mg QD Imatinib 400 mg QD % With MR 4 60 50 40 30 20 10 0 P=0.0021 12% 5% MR 4 MR 4.5 47% 53% 35% 42% 28% 35% 22% 17% % With MR 4.5 60 50 40 30 20 10 0 P=0.030 23% 18% 3% 12% 9% 2% 34% 21% 37% 30% 0 12 24 36 48 60 Months 0 12 24 36 48 60 Months MR 4 = BCR-ABL (IS) 0.01%; MR 4.5 = BCR-ABL (IS) 0.0032%; IS = International Scale.

We know that: Treatment discontinuation is a real opportunity: No exposure to long term toxicities Better QOL Reduced costs of treatment CMR is the absolute prerequisite Second generation TKIs allow to a much higher rate of CMR We do not know something relevant: Why the duration of TFR is so different? Which mechanisms contribute to a sustained TFR? TFR rates in a head-to-head comparison of IMA vs 2^ Gen TKIs 2^ Gen TKIs: same (long) duration of tx as for ima to allow a long TFR? What about the impact of the risk (Sokal/Euro) on TFR duration? The added value of IFN or drugs active against leukemic SCs under.

Grazie per l attenzione Leucemia Mieloide Cronica Dati sulla sospensione della terapia Gianantonio Rosti, Bologna

Defining Molecular Response Requirements for Potential Treatment Discontinuation Response at Time of Treatment Discontinuation Patients With Molecular and/or Cytogenetic Relapse, % CCyR 2 100% MMR 3 100% MMR, CCyR, MCyR 4 100% CMR for 2 years on imatinib 5-8 ~30%-65% CCyR, complete cytogenetic response; CMR, complete molecular response; MCyR, major cytogenetic response; MMR, major molecular response. 1. Milojkovic D, et al. Blood. 2011;118(21): abstract 605. 2. Goh HG, et al. Leuk ymphoma. 2009;50(6):944-951.3. Koskenvesa P, et al. Blood. 2008;112(11):738 [abstract 2121]. 4. Kuwabara A, et al. Blood. 2010;116(6):1014-1016. 5. Mahon FX, et al. Blood. 2011;118(21): [abstract 603] 6. Rousselot P, et al. Blood. 2011;118(21): abstract 3781. 7. Goh H-G, et al. Blood. 2011;118(21): abstract 2763. 8. Matsuki E, et al. Blood. 2011;118(21): abstract 3765.

Patients, % ENESTnd 5-Year Update BCR-ABL Categories at 3 Months* 100 80 60 BCR-ABL 10% 91 67 >1-10% Imatinib (n=264) Nilotinib 300 mg BID (n=258) BCR-ABL >10% 40 20 0 >1-10% 1% 1% n 176 234 88 24 BCR-ABL Level at 3 Months 33 9 Reasons for unevaluable samples included: Atypical transcripts: 5 patients on nilotinib, 2 patients on imatinib Missing samples: 4 patients on nilotinib, 5 patients on imatinib Discontinuation: 15 patients (including 1 progression) on nilotinib, 12 patients (including 1 progression) on imatinib *Calculated from total number of evaluable patients with PCR assessments at 3 months. Data cutoff: May 22, 2013 25 Saglio G, et al. Blood. 2013:[abstract 92].

Characteristics of patients included in the STIM Study A prospective, multicentre, non-randomized study with 19 participating institutions in France: 100 patients enrolled between July 2007 and Dec 2009 Median age (range): 63 years (29 80) Gender distribution: 48 males, 52 females Patients with previous IFN treatment: 51 De novo CML patients: 49 Median follow up: 55 months (range 9-72)

Frequent and sustained drug-free remission in the Australasian CML8 trial of Imatinib withdrawal Median follow-up of 42 months (range 15 72 months) Ross DM et al. Blood. 2013 Jul 25;122(4):515-22.

Survival Probability CML-IV: Survival by Confirmed Response at 4 Years 1.0 4-year survival, 93% 8-year survival MR 4.5 at 4 years 0.8 No MMR at 4 years 0.6 0.4 0.2 0 No. at risk < 0.0032% 0.0032%-0.01% 0.01%-0.1% 0.1%-1% > 1% 0 1 All patients with primary imatinib (n = 1,194) 8-year survival (%) BCR-ABL IS < 0.0032% 92 0.0032% -0.01% 90 0.01%-0.1% 88 0.1%-1% 83 > 1% 78 2 3 4 5 6 7 8 Time Since Diagnosis, Years 198 191 260 55 44 Achievement of confirmed MR 4.5 at 4 years predicted significantly higher survival probabilities at 8 years vs 0.1% to 1% BCR-ABL IS (92% vs 83%; P =.047) NS NS NS 163 149 206 41 32 P.047.001 69 69 106 24 16 40 41 67 17 8 9 MR 4.5 vs no MMR 92% vs 83%, P <.047 Hehlmann R, et al., JCO, 2014;32(5):415-423.

Disease burden and cytogenetic response Number of Leukemic Cells 10 12 10 11 CHR ELN recommendations (optimal response) 3 months 10 10 CCyR Overall survival 12 months 10 9 10 8 MMR Stable response No risk of progression 18 months 10 7 CMR (Complete molecular response) 10 6 Undetectable BCR-ABL mrna transcript by real-time quantitative and/or nested PCR in 2 consecutive blood Baccarani M, et al. J Clin Oncol. 2009;27(35):6041-6051 Radich JP. Blood. 2009;114:3376-3381 samples of adequate quality (sensitivity > 10 4 )*

Survival without MMR loss % STOP 2G-GKI experience 2 nd generation TKI discontinuation 100 Survival without MMR loss at 12 months 80 61.1% 95% CI: 45.6-76.6) 60 40 20 0 0 6 12 18 24 30 36 42 Months since 2G-TKI discontinuation As of November 30, 2012, 39 patients with a minimum follow-up of 6 months (median 17 months, range: 7-38) were analyzed 16/39 patients lost MMR Median time to MMR loss was 3 months (1-25) Rea et al. Blood (ASH) 2012; 120: Abstract 916

Patients Without Progression, % Patients Without Progression, % PFS (AP/BC or Death) on Study by BCR-ABL Levels at 3 Months ENESTnd 5-Year Update Nilotinib 300 mg BID Imatinib 400 mg QD EMR Failure EMR Failure 100 90 80 70 60 50 40 30 20 10 0 BCR-ABL Level 1% > 1% to 10% > 10% Censored Observations Pts Evt Cen 145 7 138 89 4 85 24 6 18 PFS by 5 Years a 94.6% 95.3% 78.3% 0 1 2 3 4 5 6 Time Since Randomization, Calendar Years P =.9814 P =.0010 100 90 80 70 60 50 40 30 20 10 0 BCR-ABL Level 1% > 1% to 10% > 10% Censored Observations Pts Evt Cen 43 2 41 133 2 131 88 16 72 0 1 2 3 4 5 6 Time Since Randomization, Calendar Years PFS by 5 Years a 98.5% P =.2338 95.3% P <.0001 80.1% Patients with EMR failure (BCR-ABL > 10% at 3 months) have significantly worse 5-year PFS Rates of EMR failure are lower on nilotinib 300 mg BID vs imatinib Cen, censored; EMR, early molecular response; Evt, events; Pts, patients. a PFS rates reported consider each year to consist of twelve 28-day cycles. Data cutoff: May 22, 2013 31 Saglio G, et al. Blood. 2013:[abstract 92].

NK CELL COUNTS AT THE TIME OF IMATINIB DISCONTINUATION NK cells NK cells CD56 dim NK cells CD56 bright p=0.0135 p=0.011 p=0.366 CD3 - /CD56 + /mm 3 800 600 400 200 0 No relapse Relapse CD3 - /CD56 dim /mm 3 800 600 400 200 0 No relapse Relapse CD3 - /CD56 bright /mm 3 40 30 20 10 0 No relapse Relapse All No relapse Relapse Median CD3 - CD56 + /mm 3 (range) 174 (67-727) 233 (70-727) 145 (67-450) Median CD3 - CD56 dim /mm 3 (range) 162 (55-723) 216 (55-723) 139 (58-438) Median CD3 - CD56 bright /mm 3 (range) 7 (2-31) 7 (2-30) 8 (2-31) Delphine Rea et al for STIM Investigators, ASH 2013 (abs 856)

Patients With MMR, % Cumulative Incidence of MMR ENESTnd 5-Year Update Nilotinib 300 mg BID (n = 282) Nilotinib 400 mg BID (n = 281) Imatinib 400 mg QD (n = 283) 100 90 By 1 Year a By 4 Years a By 5 Years a 80 70 60 50 55%, P <.0001 51%, P <.0001 76%, P <.0001 73%, P <.0001 Δ 17% to 20% 56% 77%, P <.0001 77%, P <.0001 Δ 17% 60% 40 Δ 24% to 28% 30 20 27% 10 0 0 1 2 3 4 5 6 Time Since Randomization, Calendar Years MMR, major molecular response (BCR-ABL IS 0.1%). a Cumulative response rates reported consider each year to consist of twelve 28-day cycles. Data cutoff: May 22, 2013 33 Saglio G, et al. Blood. 2013:[abstract 92].

Patients With MR 4.5, % ENESTnd 5-Year Update Cumulative Incidence of MR 4.5 100 90 Nilotinib 300 mg BID (n = 282) Nilotinib 400 mg BID (n = 281) Imatinib 400 mg QD (n = 283) 80 70 By 4 Years a By 5 Years a 60 50 By 1 Year a 40%, P <.0001 54%, P <.0001 52%, P <.0001 40 37%, P =.0002 Δ 21% to 23% 30 20 10 0 11%, P <.0001 Δ 14% to 17% 31% Δ 6% to 10% 7%, P <.0001 1% 23% 0 1 2 3 4 5 6 Time Since Randomization, Calendar Years MR 4.5, molecular response 4.5-logs (BCR-ABL IS 0.0032%). a Cumulative response rates reported consider each year to consist of twelve 28-day cycles. Data cutoff: May 22, 2013 34 Saglio G, et al. Blood. 2013:[abstract 92].

Patients, n ENESTnd 5-Year Update Progression to AP/BC on Core Treatment a 25 P =.0185 P =.0085 20 P =.0059 17 P =.0009 15 12 10 5 0 5 2 3 3 4.2% 0.7% 1.1% 6.0% 1.1% 1.8% Including Clonal Evolution Imatinib 400 mg QD (n = 283) Nilotinib 300 mg BID (n = 282) Nilotinib 400 mg BID (n = 281) No new progressions on core treatment (excluding or including clonal evolution) in year 5 a Includes progression to AP/BC or deaths in patients with advanced CML occurring on core treatment. Data cutoff: May 22, 2013 35 Saglio G, et al. Blood. 2013:[abstract 92].

Disease burden and cytogenetic response Number of Leukemic Cells 10 12 10 11 CHR ELN recommendations (optimal response) 3 months 10 10 CCyR Overall survival 12 months 10 9 10 8 MMR Stable response No risk of progression 18 months 10 7 CMR (Complete molecular response) 10 6 Undetectable BCR-ABL mrna Possibility transcript to by real-time quantitative and/or nested PCR discontinue in 2 consecutive blood samples of adequate quality (sensitivity therapy > 10 4 )* Baccarani M, et al. J Clin Oncol. 2009;27(35):6041-6051 Radich JP. Blood. 2009;114:3376-3381

Cumulative Incidence of MR 5.0 Achievement According to 6 Month Response Cumulative incidence of MR 5 1.0 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 6 mo < 0.1% IS at 4.5 to 7.5 months, n = 255 0.1%-1% IS at 4.5 to 7.5 months, n = 254 1%-10% IS at 4.5 to 7.5 months, n = 211 > 10% IS at 4.5 to 7.5 months, n=107 MMR at 6 mo No MMR at 6 mo 0 0 1 2 3 4 5 6 7 8 Years after diagnosis Muller MC, et al. Blood. 2013: 122(21): [abstract 4008].

Study Schema Patients treated with imatinib for 2 years who achieved CCyR but have detectable BCR-ABL a (N = 207) R A N D O M I Z E Nilotinib 400 mg BID (n = 104) Crossover from imatinib to nilotinib for: Detectable BCR-ABL a at year 2 Treatment failure, confirmed loss of response Year 2 Imatinib continue same dose (n = 103) 4-year study a By RQ-PCR with sensitivity of 4.5 logs. BID, twice daily; CCyR, complete cytogenetic response; MMR, major molecular response; RQ-PCR, real-time quantitative polymerase chain reaction.

The kinetics of molecular relapse 10 BCR-ABL / ABL(%) BCR-ABL / ABL(%) 1 1 Logarithmic 10 1 2 Cyclic No imatinib 0,1 0,1 0,01 0,01 0,001 0,001 3 10 1 0,1 BCR-ABL/ABL (%) Fluctuating No imatinib re-treated 0,01 0,001 Mahon ASH 2009

Patients With MR 4.5, % Cumulative Incidence of MR 4.5 ENESTcmr 3-year follow-up in Patients Without MR 4.5 at Baseline (ITT analysis) Nilotinib Imatinib Crossover to nilotinib P =.0006 P =.0453 P =.0020 Δ 14% Δ 19% Δ 22% 33% n 98 96 98 96 98 96 By 12 Months By 24 Months By 36 Months In a subgroup analysis when only responses up to crossover were counted, 47% versus 24% of patients in the nilotinib and imatinib arms, respectively, achieved MR 4.5 (P =.0003) Leber B, et al. Blood. 2013;122(21):[abstract 94]. ITT, intention-to-treat.

Follow-up of the molecular-relapse patients (n=61) Among the 57 patients alive another attempt of TKI discontinuation was proposed for 16 patients in sustained CMR Patients Treatment in progress Molecular response Sustained treatment 40 27 Imatinib 8 Nilotinib 5 Dasatinib 31 CMR 9 MMR 2 nd Stop without molecular relapse 9 NA 7 CMR 2 MMR 2 nd Stop with molecular relapse 7 5 Imatinib 1 Nilotinib 1 Dasatinib 3 CMR 1MMR No response* 3 rd Stop 1 NA 1 CMR No treatment 1 NA 1 CMR Deaths 4 stroke, mesothelioma, gastric carcinoma, elderly woman general deterioration

Strategies to Try to Silence / Eradicate Ph+ Stem Cells - Ph+ stem cell eradiction may occur spontaneously after TKI therapy in some cases (how many?) - It may be stimulated by immunomodulatory agents like IFN - We may try to force it: - Through vaccination processes - Adoptive immunotherapy with antibodies directed against Ph+ Stem cells antigens (IL1R*) - Drugs affecting stem cell activated pathways *Fioretos et al. Blood (ASH) 2010.

CD34-PE CML Stem Cells are Resistant to Imatinib: Experimental Approaches Clonal exhaustion: Telomerase inhibitors Recruitment out of niche/into cycle: + IFN-alpha + Plerixafor +/- G-CSF + PLGF-Inhibitor... - imatinib + imatinib Imatinib/GF insensitive CML stem cells Reversible G1 arrest anti-proliferative effect accumulation CFSE Graham. Blood 2002;99(1):319-25 (modified) 10mM 10x IC50 +/- GFs Targeting of quiescent LSC: + JAK2 Inhibitors. + IFN-alpha + Vaccination + Autophagy inhibitors + earlier acting TKIs

JAK2-Mediated Extrinsic Survival of CML Stem Cells Stromal cells protect CML stem/progenitors cells by producing cytokines which activate JAK2 pathway: rational for combining Bcr-Abl and JAK2 inhibitor Traer E, Mackenzie R, Snead J et al. Blood 2010; 116: abstract 199