1. Oncology Journal Club Addressing Imatinib-resistant CML
Frank Giles, MD, FRCPI, FRCPath
Chief, Division of Hematology and Medical Oncology
Director, Institute for Drug Development
Deputy Director, CTRC at University of Texas
Health Science Center
San Antonio, TX

2. Historical CML Survival Curve

4. Targets in CML Signaling Pathways
Ribosome
biogenesis
PI3K
Raptor
mTOR
PDK1
IRS
Perifosine
Triciribine
eIF4E
eIF3
eIF4G
PABP
TSC2
TSC1
RHEB
4E-BP1
S6K1
AKT
Cap-dependent
translation
Translation
Ribosomal
proteins P
VEGF PDGF Flt3
Cell membrane
Hif-1a
ERK
RAD001
CCI-779
AP23573
Bcr-Abl
Jak2
MK-0457
Dasatinib
Nilotinib
SKI606
Sunitinib
Surafinib
PTK787
CEP701, MLN518
PKC412

5. Human tyrosine kinases

6. Human Kinase Dendrogram
Manning. Science; 298: 1912, 2002

7. ASH 2007, Abstract 25
IRIS 6-Year Follow-Up: Sustained Survival and Declining Annual Rate of Transformation in Patients with Newly Diagnosed Chronic Myeloid Leukemia in Chronic Phase (CML-CP) Treated with Imatinib
Hochhaus A. et al

8. IRIS 6-Year Update: Overall Survival (ITT Principle)
38 (7%) patients lost to follow-up by 5 years
6 year OS is 88% (95% considering only CML-related deaths)
(incl. 3% after BMT, 4% non-CML related)
Hochhaus A. et al, Blood. 110, 11. Abstract 25. ASH 2007

9. IRIS 6-Year Update: Annual Event Rates8
7.5 7
Event Loss of CHR, Loss of MCyR,
AP/BC, Death during treatment 6
4.8 5
% Annual Rates 4
AP/BC
3.3
2.8 3 2
1.6
1.5
1.5
0.9
0.8 1
0.5
0.4
Year
1st
2nd
3rd
4th
5th
6th
Hochhaus A. et al, Blood. 110, 11. Abstract 25. ASH 2007

10. Incidence of CML by Age
SEER 2001

11. Adherence to Imatinib May Decline Over Time
685
Patients
Months
0–1
3500
3000
2500
2000
1500
1000
500
1–2
2–3
3–4
4–5
5–6
6–7
7–8
8–9
9–10
10–11
11–12
12–13
13+
2,921
Patients taking recommended dose of imatinib
In this US study, persistency* was near 100% at month 4
Persistency declined from 94% at month 5, to 23% at month 14
Imatinib plasma level testing may help identify patients who become less adherent
*Time on therapy without significant gaps in refills.
Tsang J-P, Rudychev I, Pescatore SL. Poster presented at ASCO 2006.

12. Genes Associated with CML Progression
Radich. PNAS 103: 2794, 2006

13. Dasatinib in Blast Phase CML Overall Survival
1.0
0.8
0.6
0.4
0.2 N
No. of deaths
Median (mo)
CML-MB
109 49
11.8
CML-LB 48 27
5.3
Proportion alive
Months
Martinelli. ASH Abs 745

14. Clinical Resistance to Imatinib Mechanisms
Primary resistance
Insufficient inhibition of BCR-ABL
Low plasma levels of imatinib
Activity of drug pumps
Secondary resistance
Imatinib-resistant BCR-ABL kinase-domain mutations
Overproduction of BCR-ABL
BCR-ABL-independent mechanisms
? Activation of other kinases
? Other molecular events
Giles. Hematol Am Soc Hematol Educ Program:2005: ;
von Bubnoff. Leukemia. 203;17:829.

15. Kinase Selectivity Profiles Of Nilotinib And Dasatinib
4 targets
Imatinib
4 targets
Dasatinib
15 targets
IC50
< 10 nM
10-50 nM nM nM
After Fabian et al. Nature Biotech. 2005;23:329

16. “Targets” of Imatinib, Nilotinib, and Dasatinib
ABL
ARG
BCR-ABL
KIT
PDGFR
DDR1
NQO2
SRC
YES
FYN
LYN
HCK
LCK
FGR
BLK
FRK
CSK
BTK
TEC
BMX
TXK
DDR2
ACK
ACTR2B
ACVR2
BRAF
EGFR/ERBB1
EPHA2
EPHA3
EPHA4
EPHA5
FAK
GAK
GCK
HH498/TNNI3K
ILK
LIMK1
LIMK2
MYT1
NLK
PTK6/Brk
QIK
QSK
RAF1
RET
RIPK2
SLK
STK36/ULK
SYK
TAO3
TESK2
TYK2
ZAK

17. Dasatinib: SRC/ABL Kinase Inhibitor
Shah. Science 305: 399, 2004

18. Dasatinib 70 mg BID in CP-CML
Efficacy at 15.2 month Median Follow-up %
Complete hematologic response
Major cytogenetic response
CHR 91 59 11 49 80 5 75 52 13 40
CCyR
PCyR
Total
Imatinib intolerant
Imatinib resistant
Baccarani. Blood. 2006;108: Abstr 164.

19. Progression-Free Survival with Dasatinib 70 mg BID in CP CML
1.0
0.8
0.6
0.4
0.2
Proportion progression-free N
No. progressed
Intolerant 99 3
Resistant
287 37
Total
386 40
Months
* 70 mg BID
† Progression defined as confirmed AP / BC, loss of CHR / MCyR, ↑ WBC count, or death
Baccarani. Blood 2006; 108: Abstract 164.

20. Dasatinib 70 mg BID in CP-CML Dose Adjustment
Resistant
(N = 288)
Intolerant
(N = 99)
Total
(N = 387)
Reduction (%) 73 75
Interruption (%) 86 89 87
Escalation (%) 21 9 18
Median daily dose (mg)
(range)
101
(18–171)
104
(11–140)
(11–171)

21. Dasatinib 70 mg BID in CP-CML Fluid Retention/Cardiac AEs
Percentage
Any
G3-4
Pleural effusion 27 6
Peripheral edema 18
Pericardial effusion 4
<1
Pulmonary edema 1
Congestive heart failure 5 3
Other cardiac dysfunction 2
Baccarani. Blood 2006; 108: abst# 164

22. ASCO 2007, Abstract 7004
Dasatinib 50 Mg Or 70 Mg BID Compared To 100 Mg Or 140 Mg QD In Patients With CML In Chronic Phase (CP) Who Are Resistant Or Intolerant To Imatinib: One-year Results Of CA180034
Shah NP. et al.

23. Dasatinib in CP-CML Study Design
International, 139-center, Randomized, Open-label, Phase III
662 treated
670 randomized
100 mg QD (N = 165)
140 mg QD (N = 163)
50 mg BID (N = 167)
70 mg BID (N = 167)
100 mg
140 mg
Imatinib-resistant or
-intolerant
CP-CML
Accrual period: July 2005–March 2006: Total 662 pts
Minimum follow-up: 6 months
Median treatment duration, months (range): 8 (<1–15)
Shah et al .JCO 25; 2007 Abstract # 7004.

24. Dasatinib Phase III in CP-CML Failing Imatinib (N = 662)
Parameter (%)
100mg QD
N = 165
50mg
BID
N = 167
140mg
N = 163
70mg P
value
MCyR 64 58 62 NS
CCyR 46 47 50
Interruption 66 69 71
0.047
Reduction 33 45 54 57
<0.001
Neutropenia 34 43
0.123
Thrombocytopenia 22 40 38
0.003
Pleural effusion 10 16 20 18
0.058+
+ 100 QD vs 70 BID
Shah, JCO. 2007;25: Abstract 7004

25. Proportion progression free
Dasatinib Phase III in CP-CML Failing Imatinib Progression-free Survival
1.0
0.8
0.6
0.4
0.2 N
No. progressed
100 mg QD
165 16
50 mg BID
167 22
140 mg QD
163 23
70 mg BID 30
Proportion progression free
Months
Kantarjiran. Blood. 2006;108: Abstr 746

26. Dasatinib: Pleural Effusions in CML
138 patients: Phase I (50); Phase II (88)
Pleural effusion: 48 patients (35%; grade 3/4 in 23 [17%]). 29% in CP, 50% in AP, 33% in BP
MVA risk factors: History of cardiac disease, hypertension, twice-daily schedule
Exudative in 78% of assessable cases
Increased RV systolic pressure documented
Management included:
Drug interruption - 83%
Diuretics - 71%
Corticosteroids - 27%
Thoracentesis - 19%
Quintás-Cardama. JCO 25: 3908, 2007

27. ABL Binding Surfaces Imatinib Nilotinib Weisberg. Ca Cell 7:129, 2005
Manley. Biochem Bio Acta 1754:3, 2005 27

28. Potency and Selectivity
Nilotinib / Imatinib:
Potency and Selectivity
Nilotinib
(cell prolif. IC50)
ABL
(25 nM)
>
PDGFR
(53 nM)
KIT
(158 nM)
Imatinib
(cell prolif IC50)
(39 nM)
(98 nM)
(649 nM)
Nilotinib has no significant effect on other kinases evaluated, including Src, FLT3, VEGFR, EGFR, InsR, RET, MET , IGFR at concentrations <3000 nM.
Mestan. Blood a: Abs 1978, 2004
Weisberg. Cancer Cell 7:129, 2005

29. Nilotinib: Pre-clinical Activity
More potent, more selective, inhibitor of Bcr-Abl auto, substrate phosphorylation than imatinib
Selectively induces apoptosis, inhibits proliferation of Bcr-Abl transfected and primary leukemia cells
Increases survival in murine Bcr-Abl MPD models including imatinib-resistant models
Inhibits PDGFRα,ß and KIT ~ Imatinib
STAT, CRKL inhibitor
Verstovsek et al. Cancer. 2005;104:1230
Golemovic et al. Clin Ca Res. 2005;11:4941 Griffin et al. Blood. 2004;104:160a Abs# 551
Le Coutre et al. Blood. 2004;104:218a Abs# 76 Mahon et al. Blood. 2004;104:251b Abs# 4670
Martinelli et al. Blood. 2004;104:255b Abs# 4687
Weisberg et al. Br. J. Cancer 2006, 94, O’Hare et al. Cancer Res. 2005;65(11):4500-5
Manley et al. Biochim Biophys Acta (1-2)
Scuto et al. Blood. 2004;104:546a Abs# 1977
Weisberg et al. Cancer Cell. 2005;7(2):129-41 29

30. Nilotinib Phase I Study CML Hematologic Responses
Diagnosis N
Evaluable
OR (%)
CHR MR
RTC CP 12
11 (92) 11 – AP 46
33 (72) 21 3 9
AP clonal evolution only 5
5 (100)
Myeloid BP 24
10 (42) 2 6
Lymphoid BP
3 (33) 1
Kantarjian*, Giles* et al. N Engl J Med. 354:2542, 2006
* Indicates co-first author. 30

31. Nilotinib Phase I Study: PK
Daily
Twice daily
60,000
20,000
40,000
AUC (ng/ml/hr) 50
100
200
400
600
800
1200
Dose (mg)
Median time to peak concentrations 3 hours post dose
Mean apparent half-life = 15 hours
Steady state by day 8 in both QD and BID regimens
All trough levels > IC50 for cellular Bcr-Abl phosphorylation
PK parameters dose proportional to 400 mg QD
Exposure is greatest in the 600 mg BID group
Kantarjian*, Giles* et al. N Engl J Med. 354:2542, 2006.
*Indicates co-first author. 31

32. Clinical Resistance to Imatinib Mechanisms
Primary resistance
Insufficient inhibition of BCR-ABL
Low plasma levels of imatinib
Activity of drug pumps
Secondary resistance
Imatinib-resistant BCR-ABL kinase-domain mutations
Overproduction of BCR-ABL
BCR-ABL-independent mechanisms
? Activation of other kinases
? Other molecular events
Giles. Hematol Am Soc Hematol Educ Program:2005: ; von Bubnoff. Leukemia. 203;17:829.

33. BCR-ABL Mutations Associated With Imatinib Resistance
F486S
E255K/V
M244V
M351T/L
M343T
Y253F/H
E279K
F317L
E355G/D
F359V/C/D/I
H396R/P
Q252H/R
S417Y
E459K/Q
E450G/Q/K
M388L
G250E/A/F
D276G
T277A/N
L387F/M
V379I
A397P
P-loop
Activation loop
T315I**
F311L/I/V
V289A/I
L248V
F382L
E281A
V299L
L364I
G383D
L298V
E292V
E453G/K/A/V
Q447R
S438C
G236E
D241G
M237I
L324Q
K357R
K285N
E275K
S348L
A344V
A350V
M472I
I418V

34. IC50 Cell Proliferation (nM)
Nilotinib Inhibits 32 of 33 Imatinib-resistant BCR-ABL Mutant Phenotypes With IC50 < 1 Um
P-loop
Nilotinib resistant:
>10,000 nM
F317C
G250V
M388L
E255D
S348L
F317V
E275K
M237I
E355A
M351T
L387F
E355G
E281K
E255R
K285N
G250A
Q252H
M244V
F486S
D276G
E292K
F317L
L248V
G250E
F311V
F359V
A380S
F359C
E255K
Y253H
E255V
T315I
500
1,000
1,500
10,000
IC50 Cell Proliferation (nM)
Nilotinib sensitive:
Range 19–791 nM
H396R 34

35. Nilotinib: Phase I Study Grade 3/4 Possibly Related Laboratory AEs
Overall (N=119)
400 BID (N=32)
600 BID (N=18) 0%
11% 5% 3% 9%
Amylase
ALT/AST
Lipase
Bilirubin
Kantarjian*, Giles* et al. N Engl J Med. 354:2542, 2006
* Indicates co-first author

36. ASH 2007, Abstract 471 and 735
Nilotinib Is Highly Active and Safe in Chronic Phase Chronic Myelogenous Leukemia (CML-CP) Patients with Imatinib-Resistance or Intolerance
Le Coutre P. et al. (471)
Kantarjian HM. et al. (735)

37. Nilotinib: Phase II Baseline Demographics and Disease Characteristics
CML-CP
(N = 321)
CML-AP
(N = 127)
Median age, years 58
(21-85)
(22-82)
Additional chromosomal abnormalities, %
24.4 31
Median duration of CML, months
(5-75) 71
(2-298)
Median duration of prior imatinib use, months 33 29
Imatinib-resistant/intolerant, %
71/29
80/20
Kantarjian HM et al. ASH abstract 735, Blood 2007: 110 (11).
Le Coutre P et al. ASH abstract 471, Blood 2007: 110 (11). 37

38. Nilotinib Phase II Studies: Dose Intensity (mg/day)
800
797
787
Planned
Delivered
(CP)
N = 316
(AP)
N = 64
le Coutre et al. ASH 2006; Abst #165
Kantarjian et al. ASH 2006; Abst #2169 38

39. Nilotinib Phase II CML-CP Study Response Rates
Hematologic
Response
Cytogenetic
ASH'06
Patients with ≥ 6 months of follow-up (N = 279)
ASCO'07
Patients with ≥ 6 months of treatment (N = 320)
ASH'07
Patients with ≥ 11 months of treatment (N = 321)
CHR at baseline / *No CHR at baseline, N
115 / 206
Time to first CHR*
1 month
Time to first MCyR (N=178)
2.8 months
Median duration of MCyR has not been reached at the time of data cutoff

40. Nilotinib Phase II CML-AP Study: Response Rates
Hematologic
Response
Cytogenetic
ASH'06
Patients with ≥ 8 months of treatment (N = 64)
ASCO'07
Patients with ≥ 6 months of treatment (N = 119)
ASH'07
Patients with ≥ 6 months of treatment (N = 129)

41. Months Since Start of Treatment
Nilotinib Phase II Study: CML-CP Overall Survival Post Imatinib Failure
95%
91%
Months Since Start of Treatment
Alive, %
Total = 321
Failed = 25
lll = Censored observations
Kantarjian et al. ASH 2007 abstract 735

42. Phase II CML-AP Study Overall Survival Post Imatinib Failure
Patients = 129
Number deaths = 27
92%
81%
Alive, %
Months Since Start of Treatment
Le Coutre et al. ASH 2007 abstract 471

43. Phase II CML-CP Study Grades 3/4 Biochemical Laboratory Abnormalities
AST 2
ALT 4
Bilirubin (total) 8
Bilirubin (direct) 5
Creatinine 1
Hypocalcemia
Hypomagnesemia
<1
Hypophosphatemia 14
Lipase elevation 15
Hyperglycemia 13
Kantarjian et al. ASH 2007 abstract 735

44. Phase II CML-CP Study Possibly Related Non-hematologic AEs (frequency >10%)
All Grades (%)
Grades 3/4 (%)
Rash 30 2
Pruritus 25
<1
Nausea 24
Fatigue 20 1
Headache 18
Vomiting 12
Constipation
Diarrhea
Kantarjian et al, ASH 2007 abstract 735

45. Cross-intolerance Between Imatinib and Nilotinib
CML-CP (N=94) / CML-AP (N=23)
Patients enrolled in nilotinib study 2101 with grade 3/4 imatinib intolerance
Patients with cross-intolerance (grade 3/4) after switching to nilotinib
Rash / skin toxicity
Fluid retention
GI intolerance
Liver toxicity
Myalgias / Arthralgias
Cortes et al. ASH abstract 29, Blood 2007:110 (11) 45

46. Nilotinib Cross Intolerance in Patients with Imatinib Intolerance Nonhematologic AEs
Reason for Imatinib Intolerance
Imatinib Intolerant*
Grade 3/4 AE or persistent Grade 2 AE on nilotinib**
Gr. 3/4 AE on Nilotinib***
AE that led to dose reduction of nilotinib
D/C nilotinib due to AE N
CML-CP
N = 95
Non-hematologic 57 4 1
Rash/Skin 26
Fluid Retention 17 GI
- diarrhea 3
Liver Toxicity
- ALT
- AST 12
Myalgia/arthralgia 9

47. Nilotinib Phase II Study CML-CP Myelosuppresion
All Grades
Grade 3/4
ASH'06
Patients with ≥ 6 months of follow-up (N = 316)
ASCO'07
Patients with ≥ 6 months of treatment (N = 320)
ASH'07
Patients with ≥ 11 months of treatment (N = 321)
Most Frequent Newly Occurring or Worsening Hematologic Lab Abnormalities
Regardless of Causality
Grade 3/4
Median Duration (days) 9 15 23
Median Onset (days) 61 55 42

48. Nilotinib Phase II Study CML-AP Myelosuppresion
All Grades
Grade 3/4
ASH'06
Patients with ≥ 8 months of treatment (N = 64)
ASCO'07
Patients with ≥ 6 months of treatment (N = 127)
ASH'07
Patients with ≥ 11 months of treatment (N = 136)
Most Frequent Newly Occurring or Worsening Hematologic Lab Abnormalities
Regardless of Causality
Grade 3/4
Median Duration (days) 8 15 27
Median Onset (days) 14 21

49. CML CP After Dasatinib and Imatinib
Nilotinib:
CML CP After Dasatinib and Imatinib
Giles F et al. Presented at: 43rd ASCO Annual Meeting; June 1-5, 2007; Chicago, Ill. Abstract 7038.
Giles F et al. Presented at: 12th Congress of the EHA; June 7-10, 2007; Vienna, Austria. Abstract 554. 49

50. Cautions With Dasatinib / Nilotinib
QTc prolongation: Both – Baseline EKG, Close K and Mg monitoring
Past pancreatitis: Nilotinib C/I
Hypertension, COPD, CCF, Chest wall injury, Asthma, Pneumonia, GI bleeding, Auto-immune disorders, aspirin: Dasatinib C/I

51. Addressing Imatinib-resistant CML
Future Directions

52. Young. Ca Res 66;1007, 2006

53. MK-0457 Phase I: Patient with T315I CML BP
Cycles of therapy
WBC (x109/L) 2 7 6 5 4 3 8 9 10
12 mg/m2/hr
20 mg/m2/hr
16 mg/m2/hr
Giles. Blood. 109,

54. Number of leukemia cells (log10)
Ph Chromosome And BCR-ABL Transcript Numbers As Measures Of ‘Residual’ Leukemia During Treatment
Decreasing residual
leukemia
Number of leukemia cells (log10) 1 2 3 4 5 6 7 8 9 10 11 12 13
6.0
5.0
4.0
3.0
1.0
Log reduction from baseline
Leukocytosis
Ph-chromosome pos
RQ-PCR <3 log
Cure ?
2.0
CCyR
MMR
RQ-PCR negative
(undetectable)
RQ-PCR >3 log

55. Nilotinib Newly Diagnosed CML-CP
Nil 400mg BID 10 20 30 40 50 60 70 80 90
100 49 48 46
202
190
181 14 13 11 N=
9 mos
3 mos
6 mos
% CCyR 93 61 37 85 54 92 67
IM 400mg/day
IM 800mg/day
Jabbour E. ASCO Abstract 2172

56. Dasatinib in ECP CML Non-Hematologic Adverse Events
Percentage G1 G2 G3 34 20 26 29 40 9 3 17 37 14 23 6 11
Number of patients

57. ASCO 2007, Abstract 7023
Efficacy Of Dasatinib In Chronic Phase Chronic Myelogenous Leukemia Patients After Imatinib Failure According To Baseline BCR-ABL Mutations
Hochhaus A. et al.

58. Dasatinib Response: Cellular IC50 Of Post-imatinib Mutation (CP-CML)
Cellular IC Dasatinib Imatinib N
nM nM
H396P/R
M351T
M244V
G250E
Y253F/H >
L387M
F359V
Q252H
E255K/V
F317L
T315I > >
Complete CyR
Partial CyR Complete HR
No response
Hochhaus et al. JCO 25; 2007 Abstract # 7023

59. ASCO 2007, Abstract 7024
Response Dynamics To Nilotinib Depend On The Type Of Bcr-abl Mutations In Patients With Chronic Myelogenous Leukemia (CML) After Imatinib Failure
Mueller MC. et al.

60. CML-CP: Best Response Within 6 Months by Cellular IC50 to Nilotinib
Baseline Cellular Mutation IC50 (nM) T315I >10,000
Y253H 700
E255K 548
F359C 161
F317L 91
D276G 77
M244V 67
E355G 47
H396R 41
M351T 38
IC50 >10,000 nM:
4 no response
IC50 >100 nM:
2 PCyR 2 CHR 5 no response
IC50 <100 nM:
8 CCyR 2 PCyR 7 CHR 1 no response
Mueller et al.JCO 25; 2007 Abstract # 7024

61. CML: Potential Antigen Targets
Junction Peptides
b3a2-p210Bcr/Abl
b2a2-p210Bcr/Abl
e1a2-p190Bcr/Abl
Non-specific
Hsp70
Tissue-specific Antigens
Proteinase 3
Tryptase
Cathepsin G
Leukotriene-B4 omega hydroxylase
C-pim, C-fes
MRP14
GM-CSF receptor  chain

62. CML in 2008
Imatinib (IM) optimal frontline therapy: No role for frontline AlloSCT in adults
Dasatinib / nilotinib are effective in IM-failure in equivalent % patients with approved regimens
Toxicities should dictate order of use of dasatinib and Nilotinib. Each will generate new patterns of resistance
MK-0457 is active in T315I phenotype patients
Need to avoid manufactured discontent
Need to focus on cure

63. Questions on Patients and/or Studies with/of Hematologic Malignancies or Refractory Solid Tumors?