CML Treatment Failure: More Threatening Than It Appears. Mutation Testing

Transcription

1 CML Treatment Failure: More Threatening Than It Appears Mutation Testing

2 Content Mutations and treatment failure Single mutations Compound mutations Mutation testing Guideline recommendations Summary 2

3 Mutations and Treatment Failure

4 BCR-ABL1 mutation can confer TKI resistance and is a mechanism of treatment failure in CML Resistance BCR-ABL1 Mutation (~40%) 1,2 Others (~60%) Activation of SRC family kinases 3 Drug-drug interactions 4 BCR-ABL1 dependent Drug binding, inadequate drug concentration, or epigenetic modifications 3 Increased drug efflux or influx 3,5 Inability of TKIs to eradicate mutant stem cells or cross the blood-brain barrier 6,7 BCR-ABL1 independent BCR-ABL1 gene amplification 3 1. Soverini S, et al. Clin Cancer Res. 2006;12: Cortes J, et al. Blood. 2007;109: Bixby D, et al. Leukemia. 2011;25: Bowlin SJ, et al. ASCPT Quintás-Cardama A, et al. Cancer Control. 2009;16: Valent P. Biologics. 2007; 1: Corbin AS, et al. J Clin Invest. 2011;12:

5 How do BCR-ABL1 mutations emerge? Mutations can occur in cancer cells where genetic instability is high and accumulation of further abnormalities is likely 1,2 TKI therapy can select for BCR-ABL1 mutations in unstable cancer cells Small cell populations containing mutations may have a survival advantage during TKI therapy and emerge later as the dominant clone 1,2 In an in vitro simulation, sequential use of TKIs increases the probability that mutations will arise, and the sequence of TKIs used may influence the types of mutations that emerge 3 Sensitive to TKI Resistant to TKI Treat with TKI Sensitive to TKI Resistant to TKI 1. Soverini S, et al. Blood. 2011;118: Sierra JR, et al. Molec Cancer. 2010;9: Bauer RC, et al. Clin Cancer Res. 2013;19:

6 Mutation-related TKI resistance is common in patients with CML Up to one in five newly-diagnosed CP-CML patients who start first-line TKI therapy will develop a BCR-ABL1 mutation Ai J, et al. Ther Adv Hematol. 2014;4:

7 Physicians do not always test for mutations according to guideline recommendations According to a 2010 survey of 507 physicians treating patients with CML 1 * Nearly half would not test for BCR-ABL1 KD mutations in patients not achieving MMR two years after the initiation of TKI therapy 1 9% indicated that they were unfamiliar with, never ordered, or did not have access to the test for BCR-ABL1 KD mutations 1 Many clinicians do not appreciate the role of mutation analysis in the overall management of CML 2 *Prospective United States-based, noninterventional, cross-sectional study conducted through an online survey in December Kantarjian HM, et al. Clin Lymphoma Myeloma Leuk. 2013;13: Ai J, Tiu RV. Ther Adv Hematol. 2014;5:

8 Single Mutations

9 Over 90 BCR-ABL1 point mutations have been identified that affect sensitivity to certain TKIs 1242T M244V L248V G250E/R Q252R/H Y253F/H E255K/V D276G T277A E279K V280A V289A/I F311L/I T315I F317L/V/I/C Y320C L324Q F359V/I/C/L D363Y L364I A365V A366G L370P V371A E373K S417F/Y I418S/V A433T S438C E450K/G/A/V E453G/K/V/Q E459K/V/G/Q P-loop SH3 contact SH2 contact A-loop M237V E258D W261L L273M E275K/Q E292V/Q I293V L298V V299L Y342H M343T A344V A350V M351T E355D/G/A Data from patients resistant to first-generation TKI therapy, collated from 27 studies published between 2001 and V379I A380T F382L L384M L387M/F/V M388L Y393C H396P/R/A A397P M472I P480L F486S E507G Star indicates amino acid position reported to be directly involved in first-generation TKI binding via hydrogen bonds or van der Waals interactions. Soverini S, et al. Blood. 2011;118:

10 Spectrum and frequency of BCR-ABL1 KD mutations recovered after TKI therapy Lighter color corresponds to the first amino acid change; darker color corresponds to the second amino acid change, if applicable. Cortes J, et al. Blood. 2007;110:

11 Higher number of mutations were associated with poor survival and 4 year EFS According to a single-center study of 207 patients with chronic phase, accelerated phase, or blast phase CML who failed first-line TKI, those with >1 BCR-ABL1 mutation exhibited worse response rates and long-term outcomes with TKI therapy compared with those with 1 BCR-ABL1 mutation. Among patients with 0, 1, or >1 BCR-ABL1 mutation 1 : 4-year EFS were 56%, 49%, and 0%, respectively (P=0.02) Overall survival rates were 91%, 69%, and 75%, respectively (P=0.13) 1. Quintas-Cardama A, et al. Haematologica. 2011;96(6):

12 NCCN Clinical Practice Guidelines In Oncology (NCCN Guidelines ) treatment recommendations based on BCR-ABL1 mutations Mutation present 2L and subsequent therapy options Y253H E255K/V F359V/C/I F317L/V/I/C V299L T315A T315I DAS, BOS DAS, BOS DAS, BOS NIL, BOS NIL NIL, BOS PON, OMA, HCT, or clinical trial Adapted with permission from the NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines ) for Chronic Myeloid Leukemia V National Comprehensive Cancer Network, Inc. All rights reserved. The NCCN Guidelines and illustrations herein may not be reproduced in any form for any purpose without the express written permission of NCCN. To view the most recent and complete version of the NCCN Guidelines, go online to NCCN.org. The NCCN Guidelines are a work in progress that may be refined as often as new significant data becomes available. NCCN makes no warranties of any kind whatsoever regarding their content, use or application and disclaims any responsibility for their application or use in any way. 12

13 ELN guidelines: Treatment recommendations based on BCR-ABL1 mutations In vitro sensitivity based on select BCR-ABL1 mutations* Mutation present Second-generation TKI IC50, range (nm) NIL DAS BOS PON Y253H NA 6.2 E255K E255V F359V F317L F317V 350 NA NA 10 V299L NA T315A NA 760 NA 1.6 T315I 697 to >10, to > *Per the ELN guidelines, one factor to take into consideration when considering switching TKIs is the presence and type of mutation. Baccarani M, et al. Blood. 2013;122:

14 The probability of TKI-resistant mutations may increase as the line of therapy increases and CML disease progresses Although patients may develop resistance to TKI therapy at any time, the risk of developing additional mutations may increase as patients move to later lines of TKI therapy and as the disease progresses 1 Up to 80% of patients with BP-CML have mutations 2 83% of first-generation TKI-resistant patients who relapsed while on a secondor third-line TKI experienced an emergence of newly acquired BCR-ABL1 mutations 3 1. Bauer RC, et al. Clin Cancer Res. 2013;19: Soverini S, et al. Blood. 2011;118: Soverini S, et al. Blood. 2009;114:

15 Compound Mutations

16 Compound mutations CML cells BCR-ABL1 compound mutations 1 Treatment with multiple TKIs may select for compound mutations ( 2 mutations in the same BCR-ABL1 molecule) that confer resistance to multiple TKIs 1,2 In vitro data suggest that compound mutations can be highly resistant to certain TKIs 3 1. Khorashad JS, et al. Blood. 2013;121: Shah NP, et al. J Clin Invest. 2007;117: O Hare T, et al. Nat Rev Cancer. 2012;12:

17 Sequential therapy may increase the probability of compound mutations 1 1. Shah et al. J Clin Invest. 2007;117:

18 Mutation Testing

19 Sensitivity A variety of mutational analysis techniques exist Method Sensitivity Pros Cons Direct (Sanger) 15-25% Mutation characterization Semiquantitative Bidirectional conformation of mutations Least sensitive (but sensitive enough for general use) High Resolution Melt (HRM) analysis 5-10% Pyrosequencing 5% Denaturing High Pressure Liquid Chromatography (DHPLC) High throughput Cost effective DNA Spiking not required No additional steps required after the PCR step Ideal for large scale screening High sensitivity and specificity Quantitative Internal quality and negative controls Allele-specific, High sensitivity Abbreviations: oligonucleotide AP, accelerated phase; % BP, blast phase; Quantitative CP, chronic phase. (ASO)-PCR Easy to perform 1% Alikian M, et al. Am J Hematol. 2012; 87: High throughput Cost effective Good for large scale studies Unable to characterize the mutations Optimal function requires small amplicon size which implies several (4-5) PCR reactions Short read length Prior mutations knowledge is required Labor intensive Unable to characterize the mutations Wild Type DNA Spiking required Prone for contamination Requires several PCR amplifications to obtain amplicons of ideal size Occurrence of non-specific peaks making data interpretation difficult Sensitivity could be compromised by closely located mutations Requires prior knowledge of mutations False positives issue Labor intensive if screening for multiple mutations 19

20 Clinical conditions and the presence of BCR-ABL1 mutations Mutation analysis in 399 patients with CP-CML receiving first-line, first-generation TKI Clinical condition Patients tested for mutations, n Presence of 1 BCR-ABL1 mutations (%) Failure (27) No CHR at 3 mo 16 3 (19) No CyR at 6 mo 9 1 (11) No PCyR at 12 mo 24 4 (17) No CCyR at 18 mo 36 6 (17) Loss CCyR (31) Loss CHR (50) Suboptimal (5) No CyR at 3 mo 15 1 (7) No PCyR at 6 mo 20 1 (5) No CCyR at 12 mo 51 5 (8) No MMR at 18 mo 52 0 (0) Loss MMR (but not CCyR) 95 4 (4) Soverini S, et al. Blood. 2011;118:abstract

21 Sanger sequencing is the most extensively used technique Direct (Sanger) sequencing is currently the most extensively used technique to detect BCR-ABL1 mutations 1,2 Least sensitive method, but sufficient for general use Several contract labs offer this service Sanger can be useful for predicting the best course of treatment for TKI-resistant patients and for monitoring resistant mutations in subsequent treatment settings 2,3 1. Baccarani M, et al. Blood. 2013;122: Alikian M, et al. Am J Hematol. 2012; 87: Soverini S, et al. Blood. 2011;118:

22 Sanger sequencing may not detect all mutations present T315I F359V F317L Y253H E255V E255K F359C Mutations detectable by Sanger sequencing (n=169) Low-level mutations detectable by massspectrometry assay only (n=132) Mutation testing performed after failure with a firstgeneration TKI prior to treatment with secondgeneration TKIs (n=220) Mutations that would influence therapeutic decisions after failure with a first-generation TKI were found in more patients with mass spectrometry than direct sequencing (32% vs 23%; P=0.03) Frequency of mutations (%) 10 Parker WT, et al. J Clin Oncol. 2011;29:

23 Low-level mutations can influence failure-free survival in CP-CML patients treated with second-generation TKIs after failure with a first-generation TKI Probability of failure-free survival (%) 100 No mutations (n=38) 80 Mutations by sequencing (n=9) Low-level mutations by mass spectrometry (n=6) Months since start of second-line therapy Parker WT, et al. J Clin Oncol. 2011;29:

24 NGS provides a higher level of sensitivity to detect clinically relevant BCR-ABL1 mutations that are not detected by Sanger sequencing 1 NGS performs ultra-deep sequencing (UDS) of the BCR-ABL1 kinase domain 2 UDS provides increased sensitivity and dissects qualitatively and quantitatively the clonal texture of the mutated BCR-ABL1 positive subpopulations 2 NGS is comprehensively selective for mutations that likely contribute to disease pathogenesis 3 For example, NGS identifies potential driver mutations (genetic alterations that provide the cell with a survival advantage), while filtering out passengers (mutations that can be expected to have no effect on cell survival) 3 1. Molecular MD. Accessed December Baccarani M et al. ASCO. 2014: Shuen A, Foulkes WD. Curr Onc. 2010;17(5):

25 Proportion of patients, % NGS detects more mutations than Sanger sequencing Sanger NGS No mutations Mutations 2 mutations Baseline mutation status. 267 patients with CP-CML resistant/intolerant to at least 2 prior TKIs or with the T315I mutation. Deininger MW, et al. Blood. 2016;127:

26 Deep sequencing (DS) may detect mutations earlier than conventional sequencing 51 patients treated with second-generation TKIs for a median of 9 months following failure of a first-generation TKI acquired BCR-ABL1 mutations detected by conventional sequencing Previously collected samples were analyzed for mutations using DS In 23 patients (45%), DS identified mutations that may confer resistance following failure of a first-generation TKI Median interval between detection by DS and conventional sequencing: 3 months (range, 1-9 months) Response status (as per ELN 2013 guideline) at the time of mutation detection by DS was Optimal (n=1), Warning (n=13), Failure (n=4); 5 patients had mutations at baseline (i.e., after failure of a firstgeneration TKI) Results suggest that mutation testing using DS may be useful for patients with Warning during second-line therapy Soverini S, et al. Blood. 2014;124: Abstract

27 Guideline Recommendations

28 Mutational analysis may provide additional information for patients with inadequate response The presence of mutations remains a consideration when making treatment decisions 1 Routine monitoring of BCR-ABL1 transcripts, in conjunction with cytogenetic evaluation, provides important information about long-term disease control in patients with CML 2 National Comprehensive Cancer Network (NCCN ) and ELN guidelines do not recommend a specific technique 1,2 Recommendations on When to Perform Mutational Analysis ELN 1,3 : In case of treatment failure or progression to AP or BP NCCN 2 : CP-CML - Failure to reach response milestones - Any sign of loss of response (defined as hematologic or cytogenetic relapse) - 1-log increase in BCR-ABL1 transcript levels and loss of MMR Disease progression to AP or BP IS, International Scale. 1. Baccarani M, et al. Blood. 2013;122: Referenced with permission from the NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines ) for Chronic Myeloid Leukemia V National Comprehensive Cancer Network, Inc All rights reserved. Accessed January 20, To view the most recent and complete version of the guideline, go online to NCCN.org. 3. Soverini S, et al. Blood. 2011;118:

29 Analysis of mutations in CP-CML Mutations studies can help make treatment decisions in the event of cytogenetic or hematologic relapse 1,2 Certain mutations confer resistance to certain TKIs 1,2 There is currently no role for mutation analysis at diagnosis or in patients with adequate response to therapy 1,2 1. Referenced with permission from the NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines ) for Chronic Myeloid Leukemia V National Comprehensive Cancer Network, Inc All rights reserved. Accessed January 20, To view the most recent and complete version of the guideline, go online to NCCN.org. 2. Soverini S, et al. Blood. 2011;118:

30 Treatment options for TKI-resistant CML The results of mutational analysis is one of many factors (eg, efficacy, safety, patient comorbidities, cost) in making treatment decisions 1 For patients with TKI-resistant CML, potential treatment options include 2-4 : Alternate TKI Protein synthesis inhibitors Allogeneic hematopoietic stem cell transplantation (HCT) Clinical trial 1. Ai et al. Ther Adv Hematol. 2014;5: Baccarani M, et al. Blood. 2013;122: Referenced with permission from the NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines ) for Chronic Myeloid Leukemia V National Comprehensive Cancer Network, Inc All rights reserved. Accessed January 20, To view the most recent and complete version of the guideline, go online to NCCN.org. 4. Baccarani M, et al. Ann Oncol. 2012;23(Suppl7):vii

31 Summary

32 Summary Mutations in the BCR-ABL1 kinase domain may emerge at any time during TKI therapy and confer treatment resistance 1-7 The type of mutation can help determine the most appropriate subsequent therapy 8 Despite its recommendation in current treatment guidelines, mutational analysis is not always performed in patients with suspected TKI resistance 8,9 Mutational analysis, as recommended in current treatment guidelines, should be considered a standard part of monitoring CML patients treated with TKIs 8, Soverini S, et al. Clin Cancer Res. 2006;12: Cortes J, et al. Blood. 2007;109: Bixby D, et al. Leukemia. 2011;25: Bowlin SJ, et al. ASCPT Quintás-Cardama A, et al. Cancer Control. 2009;16: Valent P. Biologics. 2007; 1: Corbin AS, et al. J Clin Invest. 2011;12: Referenced with permission from the NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines ) for Chronic Myeloid Leukemia V National Comprehensive Cancer Network, Inc All rights reserved. Accessed January 20, To view the most recent and complete version of the guideline, go online to NCCN.org. NCCN makes no warranties of any kind whatsoever regarding their content, use or application and disclaims any responsibility for their application or use in any way. 9. Kantarjian HM, et al. Clin Lymphoma Myeloma Leuk. 2013;13: Ai J, Tiu RV. Ther Adv Hematol. 2014;5: Baccarani M, et al. Blood. 2013;122: Soverini S, et al. Blood. 2011;118:

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