1. Novel Strategies for the Treatment of AML: Tailoring Treatment For Specific Genetic Subtypes
Martin S. Tallman, M.D.
Northwestern University Feinberg School of Medicine
Robert H. Lurie Comprehensive Cancer Center
Chicago, IL

2. Topics To Address Introduction and epidemiology
Overview of current therapy
Distinguishing genetic subtypes
Novel strategies for specific genetic subtypes
Future Directions

3. Introduction New patients/deaths in 2004: 12,000/9,000
Median age of AML: 68 years
Heterogeneity in genetics and clinical manifestations
Outcome varies by prognostic factors

4. 1Douer Blood, 1996, Br J Haematol, 2003; 2Nakase Leukemia, 2000
Epidemiology
Therapy-related (alkylating agent or topo II active agents)
Evolving from MDS or MPD
Congenital chromosomal instability syndromes
Higher frequency of APL in Latin Americans1
Higher frequency of t(8;21) in Japan2
1Douer Blood, 1996, Br J Haematol, 2003; 2Nakase Leukemia, 2000

5. Current Treatment Results In Younger Adults
STUDY N
CR %
ED %
OS % (3-5 yr)
CALGB
474 72 9 34
GAMLCG
HOVON
535
253 74 77 11 7 39 38
ALFA
345 82

6. Current Treatment Results In Older Adults
STUDY N
CR %
ED %
OS % (2-7 yr)
CALGB
388 52 25 15
ECOG
348 42 17 10
SWOG
328 43 7 19
MRC
1,314 55

7. Prognostic Factors Age
Intensity of postremission therapy (younger adults)
Cytogenetics (distinguish fav- intermed- and unfav-risk groups)

8. Overall Survival by Cytogenetic Group Years After Entering Study
Favorable %
Intermediate %
Unfavorable %
Estimate
At Risk Deaths at 5 Years
100 80 60
Cumulative Percent 40 20
Heterogeneity of 3 Groups: p<.0001 2 4 6 8
Years After Entering Study
Slovak Blood, 2000

9. Prognostic Factors Molecular Markers
Transmembrane transporter proteins which confer multidrug resistance (MDR1)
Mutations in or overexpression of specific genes-unfav. prognosis: WT1, BAX, BCL-2/BAX, BAALC, EVI1, KIT, FLT3, MLL, ERG; fav. prognosis: C/EBP, NMP1
Expressed in cells from pts with normal karyotype

10. Outcome For AML with Fav-risk Cytogenetics
CR %
DFS %
OS %
CBF
3731
3122 86 88 40 47 43 50
1613 96 52 59
APL
4634 91 85
2985 94 90 87
1Castaigne Blood, 2002; 2Appelbaum Proc ASCO, 2005; 3Marcucci J Clin Oncol, 2005; 4Sanz Pro ASCO, 2005; 5Lo Coco Blood, 2004

11. Outcome For AML in Older Adults with Unfavorable Cytogenetics
CR%
OS% (5-yr)
MRC1
145 26
2.0
ECOG2 61 23
2.5
1Grimwade Blood, 2004; 2Rowe Blood, 2004

12. Current Therapeutic Strategies
Induction with anthracycline mg/m2/day for 3 days + cytarabine 100 mg/m2/day for 7 days c.i.
Multiple cycles of high-dose ara-C consolidation
No maintenance (except APL)

13. Strategies To Improve Outcome
Dose intensification (anthracyclines)
Alternative chemotherapy
Priming with growth factors
New agents

14. Daunorubicin Dose Intensification
Study
Dauno Dose
CR%
FHCRC1
70 mg/m2 80
ALFA2
80 mg/m2 76
CALGB3
90-95 mg/m2
1Appelbaum Ann Int Med, 1984; 2Castaigne Blood, 2003; 3Kolitz Blood, 1998

15. ECOG Priming Studies Recruiting Leukemic Cells into Cell Cycle
GM-CSF
Placebo CR
38%
40%
Induction mortality
26%
17%
OS, median
5.3 mo.
8.5 mo.
DFS, median
6.9 mo.
5.1 mo.
Rowe Blood, 2004

16. Priming With Growth Factor (GF) in Younger Patients with Intermediate Cytogenetics
Study N
CR %
DFS %
OS %
GF/No GF
HOVON1
464
87/86
45/33
(p=.006)
45/35
(p=.02)
ALFA2
259
91/87
50/35 (p=.05)
56/47
(p=.07)
1Lowenberg NEJM, 2003; 2Thomas Blood, 2005

17. Specific Genetic Subtypes of AML To Which Therapy Can Be Tailored
APL
CBF AML
CD33 pos AML
AML with FLT3 mutatations
AML with c-kit mutations
AML with MLL PTD

18. Curative Strategies in APL
Induction: ATRA + anthracycline-based chemotherapy
Consolidation: Anthracycline-based chemotherapy for cycles to molecular negativity
Intermediate-dose ara-C for high-risk
Maintenance: ATRA +/- low-dose chemo for years; ? Role in PCR neg after consol
Mol. Monitoring: RT-PCR from PB every 3-6 months for years, prob for high-risk only
Relapse: Arsenic followed by ASCT (allo if PCR pos) (consider prophylactic IT therapy)

19. GIMEMA AIDA 2000 Overall Survival N=338 3 years: 87% Survival Years
1.0
.75
N=338
3 years: 87%
Survival
.50
.25 .5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
Years
Courtesy of F. Lo Coco

20. Advances in Therapy in APL Paradigm for tailoring therapy to specific genetic subtype
Study Group
Contribution
No. Am. Intergroup
Maintenance
PETHEMA
Elimination of ara-C
PETH/GIMEMA
ATRA in consolidation
ATO in consolidation
GIMEMA
Gemtuzumab
Shanghai
ATRA + ATO
Iran
ATO single agent
Will chemotherapy be eliminated?

21. Arsenic Trioxide for Rel/Ref APL Pilot and US Multicenter Trial
Overall and Relapse-Free Survival
100 80 60
Probability (%) 40
OS - 1st Relapse
OS - > 1st Relapse 20
RFS - 1st Relapse
RFS - > 1st Relapse 1 2 3 4
Years
Soignet J Clin Oncol, 2001; Douer, Tallman J Clin Oncol, 2005

22. Randomized Trial of Arsenic Trioxide and ATRA in Untreated APL
< .02*
11.1
26.3
Relapse (%) (median follow-up 18 months)
< .01
119 32
6.7
 PML/RARα (fold)
< .05
25  5
35  3
40  10
Time to CR (days) NS 95 90
(%) CR — P 21
ATRA + ATO 20
ATO
ATRA N
Induction/ maintenance therapy
Reference
Shen ZX, Shi ZZ, Fang J, et al. All-trans retinoic acid/As2O3 combination yields a high quality remission and survival in newly diagnosed acute promyelocytic leukemia. Proc Natl Acad Sci U S A. 2004;101:5328–5335.
*All patients also treated with consolidation chemotherapy and 6-MP + MTX as maintenance
Shen Proc Natl Acad Sci,U S A, 2004

23. Disease-Free Survival by Treatment Group
1.0
0.8
0.6
Probability
0.4
ATRA + arsenic trioxide (n = 20)
Reference
Shen ZX, Shi ZZ, Fang J, et al. All-trans retinoic acid/As2O3 combination yields a high quality remission and survival in newly diagnosed acute promyelocytic leukemia. Proc Natl Acad Sci U S A. 2004;101:5328–5335.
0.2
arsenic trioxide (n = 18)
ATRA (n = 19) 4 8 12 16 20 24 28 32
Time of Follow-up (months)
Shen PNAS, 2004

24. Induction With Single-Agent Arsenic Trioxide: Untreated APL
Study N
CR (%)
PCR neg (%)
Postremission therapy
Zhang – China1
124 88 NR
Chemotherapy
Ghavamzadeh – Iran2,3,4
111 86 92
ATO  1
Chandy – India5,6,7
References
Zhang P. The use of arsenic trioxide (As2O3) in the treatment of acute promyelocytic leukemia. J Biol Regul Homeost Agents. 1999;13:195–200.
Ghavamzadeh A, Alimoghaddam K, Aghdami N, et al. Treatment of new cases of acute promyelocytic leukaemia by arsenic trioxide. Presented at: 39th Annual Meeting of the American Society of Clinical Oncology; May 31–June 3, 2003; Chicago, Ill. Abstract 2285.
Ghavamzadeh G, Ghavamzadeh A, Alimoghaddam K, et al. Treatment of new cases of acute promyelocytic leukaemia by arsenic trioxide. Presented at: 9th Annual Congress of the European Hematology Association; June 10–13, 2004; Geneva, Switzerland. Abstract 093.
George B, Mathews L, Balasubramanian P, Shaji RV, Srivastava A, Chandy M. Molecular remission with arsenic trioxide in patients with newly diagnosed acute promyelocytic leukemia. Haematologica. 2004;89:1266–1267.
George B, Mathews L, Balasubramanian P, Shaji RV, Srivastava A, Chandy M. Treatment of newly diagnosed patients with acute promyelocytic leukemia using intravenous arsenic trioxide. Presented at: 9th Annual Congress of the European Hematology Association; June 10–13, 2004; Geneva, Switzerland. Abstract 085. 51 80 95
ATO  6
4Ghavamzadeh Ann Oncol, 2005
5George Haematologica, 2004
6George European Hematol Assoc, 2004
7Mathews Blood, 2005 (abstr)
1Zhang J Biol Regul Homeost Agents, 1999
2Ghavamzadeh ASCO, 2003[abst]
3Ghavamzadeh EHA, 2004[abst]

25. North American Intergroup Trial
Induction
Consol #1
Consol #2
Maint
ATRA 6-MP
MTX
ATRA Dauno Ara-C
ATO
ATRA Dauno
Untreated APL CR
No ATO
ATRA 7 days, QOW
Tests arsenic as early consolidation and 2 maint regimens

26. ATRA Daunorubicin Ara-C Gemtuzumab Ozogamicin
Proposed North American Intergroup Study-Phase III Low- and Intermediate-Risk
Induction
Consol #1
Consol #2
Maint
ATRA 6-MP
MTX
neg
ATRA Daunorubicin Ara-C
ATRA Daunorubicin
PCR
ATO
OBS
pos
Gemtuzumab Ozogamicin
HSCT
Tests benefit of maint in PCR neg patients

27. Proposed North American Intergroup Study-Phase II High-Risk
Induction
Consol #1
Consol #2
Consol #3
Maint
ATRA ATO Gemtuzumab Ozogamicin
Gemtuzumab Ozogamicin
ATRA 6-MP
MTX
ATRA Daunorubicin
ATO
Tests benefits of 3 nonchemotherapy agents in induction

28. Incidence of CBF AML by Age
Total
inv(16)
t(8;21)
P-value
16-30
27%
43%
57%
0.056
31-40
26%
55%
45%
41-55
59%
41%
56-65 9%
50%
66-83
10%
62%
38%
Appelbaum FR, Kopecky KJ, Tallman MS, et al (submitted)

29. Outcome of t(8;21) AML Study N CR OS Postrem Palmeri 17 82% 79%
HiDACx3
Nishii 85
95%
52%
No I-or HDAC
Baer 29
90%
45%
>/=1 HiDAC
Marcucci
139
89%
46%
S-, I-, HiDAC
Appelbaum
Schlenk
174
191
85%
87%
65%
Variable
HiDAC
Palmeri Leuk Res, 2002; Nishii Leukemia, 2003; Baer Blood, 1997; Marcucci J Clin Oncol, 2005; Appelbaum (submitted); Schlenk J Clin Oncol, 2004

30. Outcome of inv16/t(16;16) AML
Study N CR OS
Postrem
Marcucci
164
87%
54%
S-, I-HiDAC
Delaunay
110
93%
58%
variable
Appelbaum
196
89%
50%
Schlenk
201
60%
HiDAC
Marcucci J Clin Oncol, 2005; Delaunay Blood 2003; Appelbaum (submitted); Schlenk J Clin Oncol 2004

31. Overall Survival by Abnormality Disease-Free Survival by Treatement
Outcome for CBF Leukemias
100
Overall Survival 80
5Year
N Deaths Estimate
All Patients % 60
Percent 40 20 5 10 15 20 25
Years
Overall Survival by Abnormality
Disease-Free Survival by Treatement
5Year
N Events Estimate
FA %
HCT %
HDAC %
Other %
Years
100
Percent 40 60 80 20
5Year
N Events Estimate
inv %
t(8;21) % 10 25 5 15
100 80 60
Percent 40 20 5 10 15 20 25
Years After Start of Post-CR Regimen
Appelbaum et al.(submitted)

32. CALGB Protocol 19808: Induction +/- MDR Modulation Followed by Cytogenetic Risk-Adapted Intensification in Younger Adults
HiDAC
Consolidation
Therapy x 3
ADE
Favorable Cytogenetics
Obs.
HiDAC
G-CSF VP-16
If able to
Receive PSCT
BU/VP-16
PSCT
Stem Cell
Mobilize
Unfavorable Cytogenetics
IL-2
If unable to
Receive PSCT
HiDAC
Consolidation
Therapy x 2
ADEP
VP-16
HiDAC
G-CSF
Tests HiDAC for fav. Cyto and IL-2 post-ASCT

33. Specific Genetic Subtypes Are Heterogeneous: t(8;21)
CD56 expression may confer poor prognosis
Associated trisomy 4 is a distinct subtype with poor prognosis
Receptor tyrosine kinase pathway mutations in 49% and confer poor prognosis
Baer Blood, 1997; Nishii Leukemia 2003; Nanri Leukemia 2005

34. Valproic acid, SAHA, depsipeptide Deoxyadenosine analogs
New Agents
Class
Agent
Target
Antibodies
Gemtuzumab
CD33
MDR inhibitors
PSC833, Zosuquidar
P-gp
FT inhibitors
Tipifamib
Lamin A, HJJ-2
FLT3 inhibitors
PKC-412, CEP-701, MLN518, SU11248
FLT3 ITD
HDAC inhibitors
Valproic acid, SAHA, depsipeptide
HDAC
Antiangio agents
Bevacizumab
VEGF
Apoptosis inhibitors
Genasense
BCL-2
Deoxyadenosine analogs
Nucleoside analogs
Clofarabine
Tiazofurin
DNA
IMPDH

35. Gemtuzumab Ozogamicin
Structure
hP humanized anti-CD33 antibody
Blue- linker
Periwinkle - calicheamicin
hP67.6 HN S O
NHN Me
Approved for adults > 60 in first relapse
Induces CR + CRp in ~ 30%1,2
Rare VOD/SOS if allo < 3.5 mo.3 S H HO O
OCH 3 NH N Et OH CH HN I
1Sievers J Clin Oncol, 2001; 2Larson Leukemia, 2002; 3Wadleigh Blood, 2003

36. Gemtuzumab OzogamicinCR
MD Anderson1
Single Agent 8%
With IL-11
36%
Northwestern2
27%
EORTC3
23%
Followed by Chemo
35%
1Estey Blood, 2002; 2Nabhan Leukemia Res, 2004; 3Amadori Blood, 2004 [abstr]

37. Gemtuzumab Ozogamicin in Induction
Study
Chemotherapy
GO Dose CR
DeAngelo
Dauno/Ara-C
6 mg/m2 d4
83%
Kell
DAT or FLAG-ida
3 mg/m2 d1
85%
Prompts SWOG Phase III trial of chemotherapy +/- GO d4
DeAngelo Blood, 2003 (abstr); Kell Blood, 2003

38. Daunorubicin 45 mg/m2/day Daunorubicin 90 mg/m2/day
ECOG Protocol E1900: Dose Intensification in Induction and Gemtuzumab Ozogamicin (GO) pre-ASCT in Younger Adults with Untreated AML
Daunorubicin 45 mg/m2/day +
Cytarabine
High Risk
Allogeneic HSCT CR
Autologous
ASCT
HiDAC x 2
PBSH after 2nd course
Daunorubicin 90 mg/m2/day +
Cytarabine GO
6 mg/m2 IV day 1
Tests anthracycline dose intens and GO as in vitro purge pre- ASCT

39. Inhibition of Multidrug Resistance
P-gp-mediated MDR plays major role in clinical resistance to chemotherapy
P-gp correlates inversely with CR
71% of patients > age 60 express moderate to high P-gp1
Major limitation is alteration in pK of concomitant chemotherapy
1Leith Blood, 1994

40. MDR Modulation Studies in AML De novo > 60 years
Group
Modulator
Regimen
Outcome
CALGB1
PSC-833
ADE+PSC-833
Closed due to toxicity
HOVON2
DA+PSC-833
DFS, OS not improved
1Baer Blood, 1999;
2 Van der Holt Blood, 2004 (abstr)

41. Inhibition of Multidrug Resistance
LY (Zosuquidar)
Selective P-gp inhibitor with high affinity1
In vitro conc of nM circumvent P-gp-mediated resistance 2,3
Does not alter PKs of co-administered drugs2
Phase II trial in poor-risk AML CR or mCR 42%4
ECOG phase III trial
1Sato Cancer Res, 1991; 2Dantzig Cancer Res, 1996;
3Green Biochem Pharmacol, 2001; 4Cripe Blood, 2001 (abstr)

42. ECOG Protocol E3999: Dauno + Cytarabine +/- Zosuquidar in Older Adults
Induction
Consolidation I
Daunorubicin
Cytarabine
Zosuquidar E V A L U T E
CR or MR
Cytarabine
Consolidation II
Daunorubicin
Cytarabine
Placebo
Daunorubicin
Cytarabine
Zosuquidar
Daunorubicin
Cytarabine
Placebo
Tests novel MDR modulator

43. Ras Ras mutations Activating mutations in 10-30% of AML1,2
Frequent in t(3;5) and inv(16)3
Active inhibition of farnesyl transferase (FT), inhibits ras protein
Inhibitors of FT active in AML4,5
Gene expression profiling may predict response to Tipifarnib6
1Radich Blood, 1990; 2Neubauer Blood, 1999; 3Bowen Blood, 2005;
4Karp Blood, 2001; 5Lancet Blood, 2002; 6Raponi Blood, 2005

44. Tipifarnib-Phase II Trial in Untreated High-risk AML/MDS
Med age 74 yrs (46-85)
CR in 21%
Med CR dur 5-8 mo. ( )
Gr. 4 neutropenia 13%
Inhibition of FT in 74% of samples
Encouraged US Intergr Phase II trial of 2 different doses/schedules in older adults
Lancet Blood, 2003 (abstr)

45. S0432: US Intergroup Phase II Study Zarnestra for Previously Untreated AML in Patients > Age 70
Randomization
Arm 1
Arm 2
Arm 3
Arm 4
Zarnestra
600 mg
bid x 21 days
q 28 days
Zarnestra 600 mg bid x 7 days every other week q 28 days
Zarnestra 300 mg
bid x 7 days every other week q 28 days

46. AML With Mutant RAS and Cytarabine Intensification
wtRAS
mutRAS
LoDAC
HDAC
Yrs to relapse
0.8
1.1
0.6 NR RR
82%
71%
100%
45%
Neubauer ASCO, 2005

47. Prognostic Significance of FLT3 ITD
100 80 60
Survival (%)
ITD- n=627
44% 40
ITD+ n=227
32% 20
P<0.001 1 2
Years 3 4 5
Kottaridis Blood, 2001

48. Tyrosine Kinase Inhibitors
PKC412
Staurosporine-derived, targets PKC, KDR, VEGF-R2, PDGFR, c-KIT, FLT3
Phase II study1 – 28 pts with FLT3 mutation, HI in 50%, but no CR or PR
CEP-701
Indolcarbazole alkyloid-targets TrkA, VEGFR, FLT3
Phase I/II study2-14 pts with FLT3 mutation, biologic activity in 35%, but no CR or PR
1Estey Blood, 2003; Smith Blood, 2004

49. FLT3 Inhibitors and Chemotherapy in AML
CEP-7011
N=34 in first relapse
MEC or HiDAC +/- CEP-701
10/17 CR with CEP-701
PKC4122
N=19 de novo
Dauno + Ara-C + PKC412
CR 71-75%
100% FLT3 mut
62% FLT3 wt
1Lewis, Blood, 2005 (abstr);
2 Stone, Blood, 2005 (abstr)

50. 1Stone Blood, 2005; Levis, Blood 2005
FLT3 Inhibitors
In vitro data strong
Modest clinical activity-few, if any, CRs
How best to develop?
Focus on combinations with chemotherapy1, 2; Phase III US Intergroup trial planned (dauno + ara-C +/- PKC412)
Will combining agents with in vitro activity, but modest clinical activity, be effective?
1Stone Blood, 2005; Levis, Blood 2005

51. Genasense (Oblimersen Sodium)
Bcl-2 associated with poor outcome in AML
Phosphorothioate 18-mer antisense oligonucleotide directed at first 6 codons of Bcl-2
Genasense + dauno/cytarabine in high-risk de novo pts
26 pts, med. age 67
CR in 45.4%; no unexpected toxicity
CALGB phase III trial underway
Marcucci Blood, 2003

52. CALGB 10201: Daunorubicin + Cytarabine +/- G3139 (Genasense) in Older Adults
High-Dose
Cytarabine CR CR
Cytarabine +
Daunorubicin
High-Dose Cytarabine
Remission
Induction
Consol
I & II
Tests bcl-2 antisense strategy

53. Clofarabine
Intentionally designed to incorporate the favorable properties of fludarabine/cladribine
Multiple mechanisms of action
Inhibits DNA replication and repair
Disrupts mitochondrial function leading to apoptosis
Active in both dividing and non-dividing cells

54. Clofarabine Trials in AML
Regimen N
Population
CR/OR %
Clo 31
Rel/Ref 01 28
Untreated
59/752
Clo +IDAC 25
22/383
Clo + IDAC 60
52/604
Clo + LoDAC 32
59/625
1Kantarjian Blood, 2003;2Faderl Blood, 2005; 3Burnett Blood, 2005; 4Faderl Blood, 2005; 5Faderl Blood, 2005

55. Proposed ECOG Trial
HLA-iden sibling
Mini-MUD alloSCT
CClofarabine
Low-dose cytarabine
CClofarabine
Low-dose cytarabine
Age
60-69 R
CCytarabine 1.5 mg/m2 IV Q12 d1-6
DDaunorubicin
Cytarabine C R
RESPONSE
CCytarabine 1.5 g/m2 IV Q12 d1-3
Age
70-79 R
CClofarabine
Clofarabine
Can we eliminate conventional chemotherapy in older adults?
No CR
Off Study

56. Specific Genetic Subtypes Of AML To Target In The Very Near Future
C-KIT AML: Imatinib, Dasatinib, FLT3 inhibitors
MLL PTD AML: HDAC and DMT inhibitors
Bcr-abl pos AML: IMP dehydrogenous inhibitor (Tiazofurin )
Whitman Blood, 2005; Malek Leuk Res, 2004

57. Other Active Agents Awaiting Assignement To Specific Genetic Subtypes
Choretazine (VNP40101M): sulfonylhydrazine alkylating agent1
Tandutinib: small molecule inhibitor or type II receptor tyrosine kinases2
XL99: inhibits multiple receptor kinases(VEGFR-2, PDGFR-alpha, PDGFR-beta, c-KIT, SRC, FGFR1, FLT4, FLT3)
Low-dose decitabine3
Arsenic plus low-dose ara-C4
1Giles Blood 2005 (abstr); 2Deangelo Blood, 2004 (abstr);
3Lubbat Blood, 2005 (abtsr); 4Robosz Blood, 2005(abstr)

58. Future Directions
Gene expression profiling to determine signatures characteristic of specific genetic subtypes, identify cooperating mutations and perturbed pathways and predict treatment response
Further characterization of normal karyotype AML and adverse karyotype AML
Increased collaboration among cooperative groups to study myriad of new agents to tailor to specific genetic subtypes