1) The document discusses a symposium on trastuzumab in early breast cancer treatment. 2) Trastuzumab requires HER2 overexpression for efficacy and is associated with improved overall survival when used in the adjuvant setting for breast cancers that overexpress HER2. 3) The symposium included a question and answer session on the appropriate use and efficacy of trastuzumab in breast cancer treatment.

1. SYMPOSIUM
TRASTUZUMAB IN EARLY BREAST
CANCER
29/05/2012
DR. R. RAJKUMAR
II YR POST GRADUATE
DEPT OF MEDICAL ONCOLOGY
MADRAS MEDICAL COLLEGE
CHENNAI

2. QUESTION
• Trastuzumab:
– A. Is indicated for all adjuvant breast
cancer treatment
– B. Marginally improves overall
survival
– C. Requires Her 2 neu
overexpression for efficacy
– D. Is not yet approved for breast
cancer treatment

3. ANSWER
• Trastuzumab requires Her 2 neu
overexpression for efficacy
– Trastuzumab is indicated only for
breast cancers that overexpress Her
2 neu, is associated with a
significant improvement in overall
survival in the adjuvant setting, and
is FDA approved for adjuvant and
metastatic breast cancer
Cobleigh, MA et.al., J Clin Oncol 1999 Sep; 17(9):
2639-48
Slamon, DJ et.al., N Eng J Med 2001 Mar 15;

4. ANSWER
1. AGREE
2. DON’T AGREE
3. DON’T KNOW
4. AGREE FOR THE TIME BEING

5. ANTI HR+ & ANTI HER2+
CROSS TALK
1. AGREE
2. DON’T AGREE
3. DON’T KNOW

6. ANTI HR+ & ANTI HER2+
CROSS TALK
QUESTION
1. IT CAN BE OVERWHELMED
2. NO WAY
3. DON’T KNOW

7. HER 2 & TOP II ᾀ
CO - AMPLIFICATION
QUESTION
CHOICE OF CHEMOTHERAPY
1. ANTHARCYCLINE BASED
2. TAXANE BASED
3. COMBINATION
4. NONE

8. HER 2 & TOP II ᾀ CO
AMPLIFICATION
QUESTION
1. 50%
2. 73%
3. 35%
4. 30%

9. HER2 Gene: Background
• Localized to chromosome 17q
• Tyrosine kinase transmembrane growth factor
receptor
• Member of EGFR gene family
• In 85% of 120 publications (> 20,000 patients
total),
abnormal HER2 expression has been linked
with adverse outcome in breast cancer
Ross and Fletcher. Semin Cancer Biol. 1999;9:125.
Pegram and Slamon. Semin Oncol. 2000;27(suppl 9):13.
Data on file, Genentech BioOncology.

10. HER2 Overexpression in
Breast Cancer
HER2 is overexpressed in
~ 25% of breast cancers
Normal (1x)
~ 25,000-50,000 HER2
receptors
Overexpressed
HER2 (10-100x)
up to ~ 2,000,000
HER2 receptors
Pegram MD, et al. Cancer Treat Res. 2000;103:57-75.
Ross JS, et al. Am J Clin Pathol. 1999;112(suppl 1):
Excessive cellular division
Slamon DJ, et al. Science. 1987;235:177-182.

11. HER2 Overexpression
Shortens Survival
HER2 oncogene
amplification
HER2 oncoprotein
overexpression
Shortened survival
Median Survival From First
Diagnosis
Slamon DJ, et al. Science. HER2 overexpressing 3 yrs
1987;235:177-182. Slamon DJ, et al. HER2 normal 6-7 yrs
Science. 1989;244:707-712.

12. Methods for Testing HER2
Status
OVEREXPRESSION AMPLIFICATION
(PROTEIN) (DNA)
IHC FISH

13. Methods for Testing HER2
Status

18. ASCO/College of American Pathologists
Guidelines for HER2 Testing in Breast Cancer
Positive for HER2 is either immunohistochemistry
(IHC) HER2 3+ (defined as uniform intense
membrane staining of >30% of invasive tumor
cells) or FISH amplified (ratio of HER2 to CEP17 of
>2.2 or average HER2 gene copy number >6
signals/nucleus for those test systems without an
internal control probe)
Equivocal for HER2 is defined as either IHC 2+ or
FISH ratio of 1.8–2.2 or average HER2 gene copy
number 4–6 signals/nucleus for test systems
without an internal control probe
Negative for HER2 is defined as either IHC 0–1+ or
FISH ratio of <1.8 or average HER2 gene copy
number of <4 signals/nucleus for test systems
without an internal control probe

20. HER2 TESTING CHALLENGES
• Unclear which patients benefit most from targeted
therapy[1-3]
• Determination of HER2 status reliant on testing
infrastructure
– ie, central vs local testing, FISH vs IHC
• Potential benefit from adding trastuzumab for
treatment of patients with tumors < 3+ IHC and
FISH negative
• Disparate results with local vs central testing
– Patients in NCCTG 9831 (adjuvant chemotherapy
trastuzumab) were assessed for HER2 by local
testing
– Central testing identified subset of patients who
were protein negative and gene negative with
HR of 0.51 for DFS (P = 0.13)

21. Disease-free-Survival According
to Local Immunohistochemistry
for HER2 and Central FISH for
Patients Treated with Adjuvant
Chemotherapy with/without
Trastuzumab in the HERA Trial
_____________________
McCaskill-Stevens W, Procter M, Azambuja E, Dafni U,
Leyland-Jones B, Ruschoff J, Dowsett M, Jordan B, Dolci S,
Abramovitz M, Stoss O, Viale G, Gelber RD, Piccart-Gebhart
M,
for the HERA Study Team

22. HER2 STATUS TESTING
Local IHC 3+ → central IHC 3+
Prospective testing for
Local IHC 2+ → central FISH+
eligibility before randomization
Local FISH + → central FISH+
Central FISH results are available for:
1131 pts. prospectively (eligibility screening)
940 pts. retrospectively (assay banked
specimens)
2071 (61%) total out of the 3401 patients
central FISH+ = FISH Ratio ≥ 2.0

24. Biology of Her2 RTKs
HER family, also known as ErbB family
– HER1, HER2, HER3, HER4
– Transmembrane receptor kinases with
extracellular domain
– Receptor-specific growth factor ligands identified
for all but HER2
Activated HER molecules
– dimerize upon ligand binding
– Result in signal transduction and cell growth
Slamon D, et al. N Engl J Med. 2001;344:783-792. Valabrega G, et al. Ann Oncol.
2007;18:977-984. Pegram MD, et al. Semin Oncol. 2000;27(suppl 11):21-25. 3.

30. HER2-Targeted Agents

31. Trastuzumab Development History
1981 1985 1987 1990 1991 1992 1993 1994 1995 199 1997 1998
6
Murine Associatio Phase I Phase Phase
HER2/n n of HER2 IND for II III
eu gene with poor rhuMAb
cloned clinical HER2
outcome
Human muMA trastuzumab
HER2 b 4D5 FDA
gene approval
cloned 9/25/98

32. Trastuzumab:
Humanized Anti-HER2 Antibody
HER2 epitopes recognized by • Targets HER2 protein
hypervariable murine
antibody fragment • High affinity (Kd = 0.1 nM)
and specificity
• 95% human, 5% murine
– Decreases potential
Human for immunogenicity
IgG-1 – Increases potential for
recruiting immune effector
mechanisms
Baselga. Satellite Symposium, 23rd Annual San Antonio Breast Cancer
Symposium 2000.

33. Trastuzumab: Mechanism of
Action

35. 1. Trastuzumab mediates ADCC
Once bound to the Fc domain of
trastuzumab, the NK cells
release substances…
…that perforate the tumour
cell membrane and promote
cell death
Nahta R, Esteva FJ. Breast Cancer Res 2006; 8: 215
Clynes RA, et al. Nat Med 2000; 6: 443-446
Gennari R, et al. Clin Cancer Res 2004; 10: 5650-5655
Arnould L, et al. Br J Cancer 2006; 94: 259-267

36. 2. Trastuzumab prevents
formation of p95HER2
Formation of the active
p95 fragment, through
proteolytic cleavage of
the extracellular
domain of HER2…
…is prevented by
trastuzumab
Molina MA, et al. Cancer Res 2001; 61: 4744-4749
Nahta R, Esteva FJ. Cancer Lett 2006; 232: 123-138

37. 3. Trastuzumab blocks
HER2-activated cell proliferation
HER2 signalling induces
cell proliferation
Trastuzumab interrupts
this process
Nahta R, Esteva FJ. Cancer Lett 2006; 232: 123-138
Fry MJ. Breast Cancer Res 2001; 3: 304-312
Gershtein ES, et al. Clin Chim Acta 1999; 287: 59-67
Yakes FM, et al. Cancer Res 2002; 62: 4132-4141
Longva KE, et al. Int J Cancer 2005; 116: 359-367

38. 4. Trastuzumab inhibits
HER2-regulated angiogenesis
HER2 signalling induces angiogenesis
Trastuzumab inhibits this process
Izumi Y, et al. Nature 2002; 416: 279-280
Nahta R, Esteva FJ. Cancer Lett 2006; 232: 123-138
Wen XF, et al. Oncogene 2006; 25: 6986-6996
Klos KS, et al. Cancer 2003; 98: 1377-1385

39. Trastuzumab: 1 target
4 mechanisms of action
Activation of ADCC Prevention of formation of
p95HER2
Inhibition of cell Inhibition of
proliferation HER2-regulated angiogenesis

40. ADCC is a key mechanism of Herceptin’s
antitumour activity in vivo
HER2
Tumour + Herceptin
cell
NK cell ADCC
FcgRIII
• Once bound to HER2, the Herceptin Fc domain recruits
immune cells to target and destroy the tumour

43. Lapatinib Blocks Signaling Through Multiple
Receptor Combinations
 Blocks signaling through
1+1 2+2 1+2
ErbB1 and ErbB2 homodimers
and heterodimers
 Might also prevent signaling
through heterodimers between
these receptors and other
ErbB family members
 Potentially blocks multiple
ErbB signaling pathways
Downstream signaling
cascade

44. ®
Herceptin in the adjuvant setting:
rationale
• HER2 overexpression is an early event in breast
cancer development and is associated with
aggressive disease
®
Herceptin offers
• A new mechanism of antitumour activity
• Proven clinical benefits in the metastatic setting,
including increased survival when used in
combination with chemotherapy
• Greater benefit when used earlier in metastatic
disease
• A favourable safety profile and good tolerability

45. HER2 and Adjuvant
• Benefit estimates for use of
trastuzumab available in Adjuvant!
Version 9.0
– HER2 included as a variable
• HER2 expression prognostic for breast
cancer
– Modest independent relative risk of 1.5
• Trastuzumab now included as
adjuvant therapy option
– Projections of benefit for trastuzumab
only for 3 years because of short follow-
up on current trials

46. Adjuvant! Limitations
• Many estimates are based on as yet
incomplete evidence and as yet
strongly debated assumptions
• For example
– No impact of HER2 status on estimates of
hormonal therapy efficacy
– No impact of HER2 status on estimates of
efficacy of adjuvant anthracyclines and/or
taxanes

47. Adjuvant! Version 9.0

48. Adjuvant! Projection for
Trastuzumab

50. Four major ongoing Herceptin ®
adjuvant trials
• The extensive Herceptin adjuvant
®
trial programme will
– investigate complementary strategies
– establish the efficacy and role of
Herceptin in the adjuvant setting
®
– establish the safety profile of Herceptin ®
– determine the optimal duration of
adjuvant Herceptin therapy
®

51. â
Herceptin in the adjuvant setting:
major trials
Four main trials are currently
investigating Herceptin in the
®
adjuvant setting
• HERA (Herceptin Adjuvant) Trial
®
• NSABP (National Surgical Adjuvant
Breast Project) trial B31
• Intergroup trial N9831
• BCIRG (Breast Cancer International
Research Group) trial 006

52. CURRENT CLINICAL DATA
ON ADJUVANT THERAPY
FOR HER2-POSITIVE
BREAST CANCER

56. Comparison of the four large
Herceptin adjuvant trials
®
Accrual Follow-up
Target Patient phase phase Primary
Trial accrual selection (years) (years) endpoint
NSABP B31 2,700 Node+, 4.75 15 OS
IHC 3+ or
FISH+
Intergroup 3,000 Node+, 4.5 15 DFS
N9831 IHC 3+ or
FISH+
BCIRG 006 3,000 Node+ and – NA NA DFS
FISH+
HERA Trial 3,192 Node+ and – 4 10 DFS
IHC 3+ or
FISH+

59. North American Trastuzumab
Adjuvant Trials in Breast Cancer
NSABP B-31 4 cycles
52 wks
T HD every 3 wks
4 cycles
Trastuzumab
AC 4 cycles
T HD every 3 wks
NCCTG 9831 12 wks
52 wks
T LD/wk
64 wks
4 cycles Trastuzumab
AC T LD/wk Trastuzumab
T LD/wk
Romond EH, et al. N Engl J Med. 2005;353:1673-1684.

60. NSABP TRIAL B31: TREATMENT PLAN
 Doxorubicin 60mg/m
2
 Cyclophosphamide 600mg/m
2
 Paclitaxel 175mg/m q3w
2
 Herceptin
®
– loading dose 4mg/kg on week 1
– maintenance dose 2mg/kg x 51 weeks

61. NSABP TRIAL B31:PRIMARY
OBJECTIVES
 Stage I: (n=1,000)
– evaluation of cardiac safety
 Stage II: (n=1,700; total=2,700)
– evaluation of efficacy
• survival: primary endpoint
• disease-free survival (DFS): secondary
endpoint

62. NSABP TRIAL B31: SECONDARY
OBJECTIVES
 Prognostic and predictive value of
phosphorylated HER2 receptor
 Prognostic and predictive value of shed
extracellular domain (ECD)
 Concordance between different HER2 assays, i.e.
IHC versus FISH
 Change in HER2-phosphorylated receptor, ECD
level or HER2 overexpression upon relapse

63. NSABP TRIAL B31: KEY INCLUSION
CRITERIA
 Histologically/cytologically proven invasive
adenocarcinoma of the breast
 At least one positive axillary node
 Axillary dissection AND either total mastectomy OR
lumpectomy
 HER2 overexpression (IHC 3+ or FISH positive)
 Known hormone receptor status (ER/PgR)
 No more than 84 days since prior surgery for breast
cancer
 No prior chemotherapy, radiotherapy or hormonal
therapy for breast cancer
 Normal cardiac, renal and hepatic function

64. Disease-Free Survival
B-31 N9831
100 100
90 87% 87%
85% 90
86%
Patients (%)
80 AC  T 80 78%
AC  TH 74%
70 66% 70 68%
Patients Events Treatment
60 60 AC  T (n = 807)
872 171 AC  T
864 83 AC  TH AC  TH (n = 808)
50 HR: 0.45; 2P = 1 x 10-9 50 HR: 0.55; 2P = .0005
0 1 2 3 4 5 0 1 2 3 4 5
Years From Randomization Years From Randomization
Romond EH, et al. N Engl J Med. 2005;353:1673-1684.

65. DISEASE-FREE SURVIVAL
B-31 N9831
100
100
AC->T+H AC->T+H
90
90
AC->T
AC->T
80
80
70
70
N Events N Events
60
60 AC->T 807 90
AC->T 872 171
AC->T+H 864 83 AC->T+H 808 51
HR=0.55, 2P=0.0005
50
50
HR=0.45, 2P=1x10-9
0 1 2 3 4 5 0 1 2 3 4 5
Years Years

68. NSABP B-31: CARDIOTOXICITY DATA
Years Cum Inc Cum Inc No. at
Arm 2: AC  T + H After Arm 1, % Arm 2, % Risk
6 Day 1
n = 850, 31 CHFs, Cycle 5
no cardiac deaths
0.5 0.3 2.6 1472
4.1%
Percentage
4 1.0 0.5 3.6 1202
HR: 5.9 1.5 0.5 3.9 983
Arm 1 evaluable cohort
Arm 2 evaluable cohort 2.0 0.5 4.1 775
2 Arm 1: AC  T 2.5 0.8 4.1 595
n = 814, 4 CHFs, 0.8% 3.0 0.8 4.1 405
1 cardiac death
0
0.5 1.0 1.5 2.0 2.5 3.0
Years After Day 1 Cycle 5
Tan-Chiu E, et al. J Clin Oncol. 2005;23:7811-7819. Reprinted with permission
from the American Society of Clinical Oncology.

69. Intergroup trial N9831: treatment
plan
 Herceptin®
– 4mg/kg loading dose (90 minutes i.v. infusion)
followed by 4mg/kg weekly (90 minutes i.v. infusion
or 30 minutes i.v. infusion based on toxicity)
 Doxorubicin 60mg/m2 every 3 weeks
 Cyclophosphamide 600mg/m2 every 3 weeks
 Paclitaxel 80mg/m2 weekly

70. Intergroup trial N9831: objectives
 Primary
– disease-free survival
– cardiotoxicity
 Secondary
– overall survival
– evaluation of whether sHER1 or sHER2 levels at
baseline are prognostic for disease-free and
overall survival
®
– concordance of IHC (HercepTest ) with FISH
TM
(Vysis ); disease-free survival; and overall
survival

71. Intergroup trial N9831:
inclusion criteria
• Operable, histologically confirmed
adenocarcinoma of the breast
• Node-positive disease
• Hormonal status known (ER/PgR)
• HER2 overexpression (IHC 3+ or FISH
positive)
• No prior chemotherapy
– hormonal therapy allowed for up to 4 weeks
but discontinued prior to enrolment
• No more than 84 days from mastectomy
or axillary node dissection
• LVEF normal

72. Intergroup trial N9831:
exclusion criteria
• Locally advanced tumours
• Prior history of breast cancer
• Prior chemotherapy or radiotherapy
for breast cancer
• Cardiac disease including:
– myocardial infarction
– history of congestive heart failure
– medication for arrythmia or angina
pectoris
• Prior anthracycline or taxane therapy
for any malignancy

73. HERA TRIAL: STUDY DESIGN
Primary management
(surgery, [neo]adjuvant chemotherapy
± adjuvant radiotherapy)
Stratification
Randomisation
Herceptin ®
Herceptin ®
q3w x 1 year q3w x 2 years Observation*
®
*Observation group to receive the same follow-up as the Herceptin treatment groups

74. HERA: Trastuzumab in HER2-
Positive
Early-Stage Breast Cancer
Observation*
Women with HER2- (n = 1698)
positive invasive
early-stage breast
Interim follow-up:
cancer, who
median 2 years
received surgery Trastuzumab
and adjuvant or 8 mg/kg loading dose,
neoadjuvant 6 mg/kg every 3 weeks
chemotherapy for 1 year
radiotherapy (n = 1703)
(N = 3401)
*All patients given the option to switch to trastuzumab May 2005 after positive interim data review.
Piccart-Gebhart MJ, et al. N Engl J Med. 2005 ;353:1659-1672.

75. HERA TRIAL: PRIMARY
OBJECTIVES
• Compare disease-free survival (DFS) in
patients with HER2-overexpressing breast
®
cancer who received Herceptin versus®
those who did not receive Herceptin
– in patients treated for 1 year
– and those treated for 2 years

76. HERA TRIAL: SECONDARY
OBJECTIVES
 Overall survival, relapse-free survival and
distant DFS
– 1 year of Herceptin versus observation
®
– 2 years of Herceptin versus observation
®
 Safety and tolerability – Herceptin versus ®
observation
 Incidence of cardiac dysfunction –
Herceptin versus observation
®
 Treatment duration (efficacy and safety) – 1
year versus 2 years of Herceptin ®

77. HERA TRIAL: STUDY SIZE
AND DURATION
 Sample size: 3,192 (1,064 per arm)
 Target population: women with HER2-
positive primary breast cancer (IHC 3+
or FISH positive)
 Study duration
– recruitment 48 months
– follow-up until 10 years after last patient enrolled
 Number of centres: ~600

78. HERA TRIAL: KEY INCLUSION
CRITERIA
 Invasive, non-metastatic, operable primary breast cancer –
histologically confirmed and adequately excised
 Axillary node positive, or node negative with tumour size
>1cm
 Known hormone receptor status (ER/PgR or ER alone)
 Completed 4 cycles of approved (neo)adjuvant
chemotherapy
 Baseline LVEF >55% (echocardiography or MUGA scan)
 Completed radiotherapy if indicated
 Centrally confirmed HER2 overexpression (IHC 3+ or
FISH positive) in invasive component of primary

79. HERA TRIAL: KEY EXCLUSION
CRITERIA
 Clinical T4 tumour, including inflammatory breast cancer
2
 Cumulative dose of doxorubicin >360mg/m or epirubicin
2
>720mg/m
 (Neo)adjuvant chemotherapy with peripheral blood/bone
marrow stem cell support
 Supraclavicular lymph node involvement
 Any prior malignant neoplasms (including primary invasive
breast cancer), except
– curatively treated basal/squamous cell carcinoma of skin
– curatively treated in-situ cervical carcinoma

83. HERA: Trastuzumab in HER2-
Positive
Early-Stage Breast Cancer (cont’d)
DFS (Censored) OS (Censored)
100 100
Patients Alive (%)
80 80
Patients (%)
60 60
1-year trastuzumab 1-year trastuzumab
40 Observation 40 Observation
20 3-year DFS: 80.6% vs 74.0% 20 3-year OS: 92.4% vs 89.2%
HR: 0.63 (95% CI: 0.53-0.75; P < .0001) HR: 0.63 (95% CI: 0.45-0.87; P < .0051)
0 0
0 6 12 18 24 30 36 0 6 12 18 24 30 36
Months From Randomization Months From Randomization
1703 1127 140 1703 1190 146
1698 930 114 1698 1042 126
Smith IE, on behalf of HERA. ASCO 2006. Clinical Science Symposium.

84. HERA: DFS Benefit in Subgroups
n HR
All 3387 0.54
Nodal status
Any, neoadjuvant chemotherapy 358 0.53
0 pos, no neoadjuvant chemotherapy 1100 0.52
1-3 pos, no neoadjuvant chemotherapy 972 0.51
4 pos, no neoadjuvant chemotherapy 953 0.53
Adjuvant chemotherapy regimen
No anthracycline or taxane 203 0.64
Anthracycline, no taxane 2307 0.43
Anthracycline + taxane 872 0.77
Receptor status/endocrine therapy
Negative 1674 0.51
Pos + no endocrine therapy 467 0.49
Pos + endocrine therapy 1234 0.68
0 1 2
Favors Favors
Trastuzumab Observation
HR: 1-Year Trastuzumab vs Observation
Smith IE, on behalf of HERA. ASCO 2006. Clinical Science Symposium.

86. HERA TRIAL: UNIQUE FEATURES
 Investigating the role of Herceptin independently from
®
chemotherapy regimen
 Investigating 2 years of Herceptin treatment
®
 3-weekly schedule from the start
– more convenient
– gives similar exposure to Herceptin as weekly
®
administration of lower doses
 New model of partnership between academia
and pharmaceutical industry

87. HERA: Cardiac Safety
Patients, n (%)
Observation 1-Yr Trastuzumab
Cardiac death* 1 (0.1) 0 (0)
Severe CHF* 1 (0.1) 10 (0.6)
Symptomatic CHF* (including severe) 3 (0.2) 36 (2.1)
Confirmed significant LVEF decline* 9 (0.5) 51 (3.0)
Any type of cardiac endpoint* 10 (0.6) 61 (3.6)
At least 1 significant LVEF decline†‡ 35 (2.3) 118 (7.4)
*Observation, n = 1678; trastuzumab, n = 1708.
†Observation, n = 1545; trastuzumab, n = 1600.
‡Many were single observations, not confirmed at subsequent time points.
Smith IE, on behalf of HERA. ASCO 2006. Clinical Science Symposium.

88. BCIRG 006
4 x AC 4 x Docetaxel
60/600 mg/m2 100 mg/m2
HER2+
(Central FISH) AC  T
4 x AC 4 x Docetaxel
60/600 mg/m2 100 mg/m2
N+
or High-Risk N- AC  TH
1-Yr Trastuzumab
6 x Docetaxel and Carboplatin
75 mg/m2 AUC 6
N = 3222 TCH
Stratified by nodes and hormone receptor status 1-Yr Trastuzumab
Slamon D. SABCS 2005. General Session 1.

90. BCIRG TRIAL 006: OBJECTIVES
• Primary
– disease-free survival
• Secondary
– overall survival
– safety
– cardiac toxicity
– quality of life
– prognostic value of HER2 overexpression

91. BCIRG TRIAL 006: TREATMENT PLAN
• Doxorubicin 60mg/m2
• Cyclophosphamide 600mg/m2
• Docetaxel 100mg/m2
• Platinum salt
– carboplatin AUC 6
– cisplatin 75mg/m2
• Herceptin ®
– 6mg/kg every 3 weeks

92. BCIRG trial 006: key inclusion criteria
• Histologically proven breast cancer
• Definitive surgical treatment
• Node-positive/negative disease
• HER2 overexpression (FISH positive)
• Normal renal, hepatic and cardiac function
• No prior systemic therapy or radiotherapy
for breast cancer

96. BCIRG 006 DISEASE-FREE SURVIVAL:
2ND INTERIM ANALYSIS
1.0 Absolute DFS benefits
(from Year 2 to 4):
93% AC  TH vs AC  T: 6%
0.9 87% TCH vs AC  T: 5%
92%
Disease Free (%)
83%
87% 86% TCH
0.8 82%
81% AC  TH
AC  T
77%
0.7
Patients Events
1073 192 AC  T
0.6
1074 128 AC  TH HR (AC  TH vs AC  T): 0.61 (0.48-0.76; P < .0001)
1075 142 TCH HR (TCH vs AC  T): 0.67 (0.54-0.83; P = .0003)
0.5
0 1 2 3 4 5
Year From Randomization
Slamon D. SABCS 2006. Abstract 52.

97. BCIRG 006 Overall Survival: 2nd
Interim Analysis
1.0 99%
97%
98% 92%
0.9 97% 95% AC  TH
93% 91% TCH
AC  T
86%
Survival (%)
0.8
0.7
Patients Events
1073 80 AC  T
0.6
1074 49 AC  TH HR (AC  TH vs AC  T): 0.59 (0.42-0.85; P < .004)
1075 56 TCH HR (TCH vs AC  T): 0.66 (0.47-0.93; P = .017)
0.5
0 1 2 3 4 5
Year From Randomization
Slamon D. SABCS 2006. Abstract 52.

98. BCIRG 006: EFFICACY RESULTS
 Both AC  TH and TCH arms
– Statistically significantly improved DFS compared with
AC  T (HR: 0.61 with AC  TH and 0.67 with TCH)
 At this time
– No statistically significant difference between AC  TH
and TCH
– Insufficient information to evaluate overall survival
(secondary endpoint)
Slamon D. SABCS 2006. General Session 1.

99. SECOND INTERIM ANALYSIS OF
ADVERSE EVENTS FOR PHASE III
BCIRG 006
 Adverse events less common and safety better in
anthracycline-free TCH arm of BCIRG 006
– Significantly lower rates of sensory neuropathy and myalgias
– No leukemias
– More grade 3/4 thrombocytopenia and anemia
 Benefit of anthracyclines in adjuvant treatment of breast
cancer now questioned
Slamon D, et al. SABCS 2006. Abstract 52.

100. HER2 and Topo IIα in BCIRG 006
2990 of 3222 patients analyzed
17 q 12 17 q 21.1 17 q 21.2
HER2 Topo IIα
N = 2990 Core region region
Topo IIα non- 1788 pts (60%)
coamplified
145 pts (5%)
Coamplified 1057 pts (35%)
Most recent analysis
Normal Amplified Deletion
Slamon D, et al. SABCS 2006. Abstract 52.

101. HER2/neu Overexpression:
Predictive of Response
Topoisomerase IIα gene (Topo IIα)
 Located close to HER2/neu on the 17q chromosome
 Integrally involved in the antitumor action of
anthracyclines
 Topo IIα is essential for DNA replication and
recombination
 Anthracyclines target Topo IIα enzyme

102. The Topo IIα Gene
Functions
 Resolves topological problems in DNA
 Is critical in RNA transcription from DNA
 Makes transient protein-bridged DNA breaks on one or both
DNA strands during replication
 Plays critical roles in segregation, condensation, and
superhelicity

103. Implications for HER2-Negative and
HER2-Positive Breast Cancers
• Superior efficacy benefits for anthracyclines
(when present) derives from their effects on Topo
IIα amplification and/or overexpression
• To date, Topo IIα amplification occurs only in 35%
of the 25% of breast cancer patients with HER2
amplification, ie, a subset of a subclass (tested in
> 4500 patients)
• Data support their preferential use in a HER2-
negative breast cancer population that is ~ 75% of
all breast cancers
• For HER2-positive breast cancers, trastuzumab
and lapatinib appear to replace the gained
efficacy of anthracyclines in the 1/3 of patients
with coamplification of HER2 and Topo IIα without
risking their known and well-established toxicities

105. HER2 is Predictive of Paclitaxel Benefit
By Estrogen Receptor
Disease Free Survival
n = 1322
ER DFS: Her2 CB11 < 50%
Neg ER DFS: Her2 CB11 < 50%
Pos
/ ER negative / ER positive
1.0
1.0
paclitaxel
paclitaxel
0.8
0.8
HER2 NEG
No paclitaxel
0.6
0.6
Proportion
Proportion
0.4
0.4
No paclitaxel n=390 n=703
No Taxol No Taxol
0.2
0.2
Taxol Taxol
(29%) (53%)
0.0
0.0
0 2 4 6 8 10 0 2 4 6 8 10
Years
DFS: Her2 Years >= 50%
CB11 DFS: Her2 CB11 >= 50%
/ ER negative / ER positive
1.0
1.0
paclitaxel
HER2 POS
paclitaxel
0.8
0.8
0.6
0.6
Proportion
Proportion
0.4
0.4
n=144 No paclitaxel
No paclitaxel
No Taxol n=79
(11%) No Taxol
0.2
0.2
Taxol Taxol
(6%)
0.0
0.0
0 2 4 6 8 10 0 2 4 6 8 105
10
Hayes D.F., et al. N Engl Years Years 357:1496-506, 2007
J Med. Years

106. FINHER TRIAL
First randomization Second randomization
Trastuzumab
Docetaxel once wkly for 9 wks;
100 mg/m2 3 cycles, first dose 4 mg/kg
followed by 3 cycles CEF then 2 mg/kg with CEF +
(n = 502) docetaxel or vinorelbine
Patients with node- (n = 116)
positive or node-
negative disease; tumor Patients with HER2
> 20 mm and PgR- amplification
negative (n = 232)
Vinorelbine
(N = 1010) CEF +
25 mg/m2 8 cycles, then
docetaxel or vinorelbine
3 cycles CEF
(n = 115)
(n = 507)
Joensuu H. SABCS 2006. Abstract 2.

108. FINHER TRIAL: EFFICACY
 At 36 months of median follow-up, the following was
observed in the trastuzumab arm
– 58% improvement in DFS
– A trend for improvement in OS
– No major increase in cardiotoxicity
 Established short duration trastuzumab as an option for
patients unable to complete a 1-year course
Joensuu H, et al. N Engl J Med. 2006;353:809-820..

111. ECOG TRIAL E2198:
INCLUSION CRITERIA
• Histologically confirmed stage II or IIIa
adenocarcinoma of the breast
• HER2 overexpression (IHC 2+/3+)
• Axillary node dissection AND mastectomy or
lumpectomy within 12 weeks prior to
enrolment
• No prior chemotherapy, hormonal therapy (at
least one year since tamoxifen therapy) or
radiotherapy
• No history of cardiac disease

112. ECOG TRIAL E2198:
OBJECTIVES
• Evaluate the incidence of cardiotoxicity
associated with paclitaxel plus
®
Herceptin in women with HER2-
positive breast cancer
• Assess the long-term safety of
®
Herceptin in this patient population

113. ECOG TRIAL E2198:
CARDIOTOXICITY
LVEF LVEF LVEF
>10% < normal grade 3/4
Post paclitaxel 9.5 (18/189) 2.1 (4/189) –
+ Herceptin®
Post AC 12.5 (16/128) 5.5 (7/128) 8

114. BIG 2.06/N063D Adjuvant HER2+
(paclitaxel) trastuzumab (trast for 1 yr)
HER2+ BC R
Tumors 1 A
cm after N (paclitaxel) lapatinib (lap for 1 yr)
completion D
of O (paclitaxel) trastuzumab+ lapatinib
anthracycline M (trast + lap for 1 yr)
based I
therapy with Z (paclitaxel) trastuzumab (12 weeks),
LVEF 50% E 6-week wash out , lapatinib (34 weeks)
Treatment Schema 1: No taxane: all neoadjuvant/adjuvant
chemo before targeted therapy.
N = 8,000
Treatment Schema 2: Taxane included: targeted therapy after
neoadjuvant/adjuvant anthracycline-based chemo, and
concurrent with weekly paclitaxel.

115. Adjuvant Regimens Prescribed for
HER2+ Disease
AC-TH Vinorelbine/trastuzumab*
Endocrine Rx +/- trastuzumab*
Vin/trastuz. then FEC
AC/EC then trastuzumab
FAC/FEC then trastuzumab
Trastuzumab alone*
TCH
Docetaxel/cyclophos + trastuzumab*
Chemo then “short course” trastuzumab*
*not based on phase III data

116. PRECLINICAL RATIONALE FOR
HERCEPTIN TREATMENT BEYOND
PROGRESSION IN HER2-POSITIVE
BREAST CANCER

117. HERCEPTIN TREATMENT BEYOND
PROGRESSION ENHANCES
EFFICACY OF COMBINATION
CHEMOTHERAPY
• HER2 remains overexpressed and
active in progressive disease
• HER2 may contribute to an even more
aggressive tumour growth if
Herceptin treatment is discontinued
• Inhibition of HER2 signalling may
sensitise tumours to chemotherapy in
tumours progressing on Herceptin
alone

118. HERCEPTIN TREATMENT BEYOND
PROGRESSION ENHANCES EFFICACY OF
COMBINATION THERAPY WITH TARGETED
AGENTS
• Herceptin synergistically enhances the
antitumour effect of Avastin in tumours
progressing on Herceptin
• Herceptin synergistically enhances the
antitumour effect of pertuzumab in
tumours progressing on Herceptin
• Lapatinib enhances the antitumour
effect of Herceptin
Scheuer et al 2006; Friess et al 2006; Scaltriti et al 2008

119. TREATMENT OPTIONS
AFTER PROGRESSION ON
TRASTUZUMAB

120. Treatment Options After
Trastuzumab
 Trastuzumab use after disease recurrence has not been
evaluated in clinical studies
 In a retrospective evaluation[1]
– Response rate was 26% when trastuzumab was used in
the second-line setting vs 43% in the first-line setting
– In another review, TTP was extended from 7.1 months to
10.2 months in patients who continued trastuzumab
 A phase III study of lapatinib plus capecitabine compared
with capecitabine alone provides evidence for lapatinib
therapy following progression on trastuzumab[2]
1. Extra JM, et al. SABCS 2006. Abstract 2064.
2. Geyer C, et al. N Engl J Med. 2006;355:2733-2743.

121. PERTUZUMAB
 Monoclonal antibody to HER2
– Recognizes different epitope than trastuzumab
– Inhibits homo- and heterodimerization of HER2
– Potentially useful for patients who have progressed
on trastuzumab
 Interim phase II study results combining
trastuzumab and pertuzumab indicate combination
is well tolerated
– Overall response rate is 18.2% in this pretreated
population
– Results suggest new HER2 monoclonal antibodies
are promising in HER2-positive breast cancer

122. PERTUZUMAB AND TRASTUZUMAB BIND
TO DISTINCT EXTRACELLULAR HER2
EPITOPES
Pertuzumab-HER2 Complex Trastuzumab-HER2 Complex
Pertuzumab
I I Dimerization domain
I I
II I II I
I I Trastuzumab
I I
V V
 Inhibits HER2 dimerization with other HER  Activates ADCC
family receptors (particularly HER3)
 Inhibits HER-mediated signaling pathways
 Activates ADCC
 Prevents HER2 domain cleavage
 Inhibits multiple HER-mediated signaling
pathways
Hubbard SR. Cancer Cell. 2005;7:287-288.

123. ESTABLISHED CHEMOTHERAPY
RESISTANCE MECHANISMS
 Impaired drug uptake
 Active drug efflux, eg by ABC transporters
(P-glycoprotein, MDR2, BCRP, MRP1-6 etc)
 Enhanced drug metabolism, eg by P450
enzymes
 Alterations of intracellular target, eg tubulin
 Upregulation of DNA repair in tumour cells
 Upregulation of signalling pathways,
eg anti-apoptotic genes (bcl-2, XIAP etc)

124. Hypothetical mechanisms of
resistance to Herceptin (1)
• Selection of HER2-negative cells in a
heterogeneous tumour
– Outgrowth of HER2-negative tumours from an
originally mixed tumour cell population
• Defective interaction of Herceptin with HER2
– Masking of Herceptin-binding epitope of HER2
– Alterations in Herceptin-binding epitope of HER2
– Loss of HER2 ECD by shedding or alternative
initiation of translation on HER2 gene
Kunitomo et al 2004; Nagy et al 2005;
HER2, human epidermal growth factor Tanner et al 2004; Stephens et al 2004;
receptor 2; ECD, extracellular domain Stephens et al 2005; Anido et al 2006

125. Hypothetical mechanisms of
resistance to Herceptin (2)
• Changes in downstream signalling proteins
which eventually disconnect growth
regulation from HER2
– PIK3CA mutations resulting in constitutively
active
PI3-kinase
– Loss of PTEN function leading to persistent
signalling activity via the PI3K/Akt survival
pathway
– Changes in cyclin-dependent kinase inhibitor
p27kip1
Berns et al 2007; Nagata et al 2004; Crowder et al 2004;
Pandolfi 2004; Kute et al 2004; Nahta et al 2004

131. IN VITRO STUDIES ARE NOT
PREDICTIVE OF IN VIVO
RESISTANCE
• In vitro resistance was observed in cell
lines exposed to Herceptin
• In vitro resistance models tend to focus
on just one biological feature
• In vitro resistance represents intrinsic
insensitivity or artificial manipulation of
cells
• Conclusions from in vitro resistance
models cannot be translated to clinical
settings
– ADCC is a key mechanism of Herceptin
efficacy in vivo
ADCC, antibody-dependent Gennari et al 2004; Arnould et al 2006;
cellular cytotoxicity Musolino et al 2008; Gianni 2008

136. Trastuzumab-DM1: Novel Antibody-
Drug Conjugate
Target expression: HER2
Monoclonal antibody: Trastuzumab Trastuzumab
Cytotoxic agent: DM1
Highly potent cytotoxic agent DM1 MCC
Linker: SMCC
T-DM1
Average drug:antibody
Systemically stable ratio ≅ 3.5:1

138. Targeted Agents for HER2+
Breast Cancer
Trastuzumab
Bevacizumab
phase III VEGF T-DM1
phase III
Sunitinib EGFR Pertuzumab
phase II
VEGFR HER2 phase III
P P P P
P P PI3-K P P
Akt/PKB
Lapatinib
PTEN phase III
Everolimus
phase III mTOR Neratinib
phase III
4E-BP1
S6K1 Gefitinib
elF-4E
phase II
Protein synthesis
Cell growth, proliferation, survival, metastasis, angiogenesis

139. Clinical Significance of
Polysomy 17 in the HER2+
NCCTG N9831 Intergroup
Adjuvant Trastuzumab Trial
Reinholz MM, Jenkins RB, Hillman D, Lingle
WL, Davidson N, Martino P, Kaufman P, Kutteh
L, and Perez EA.
NCCTG, ECOG, SWOG, CALGB
Reinholz et al: SABCS 2007 (abstract #36)
CP1270832-139

140. Adjuvant Trastuzumab May Benefit Pts
with Normal HER2 Breast Tumors (n=103)
IHC 0,1,2+ HER2 FISH ratio < 2.0
100 100
AC→T+H
90 90 AC→T+H
80 p = 0.26
Percent
80 p = 0.12
Percent
70 AC→T
N Events DFS 70 N Events DFS AC→T
60 3 yr 5 yr 3 yr 5 yr
60
50 AC→T 142 20 88.2 67.6 AC→T 74 19 82.0 63.7
AC→T+H 191 19 89.1 82.3 50 AC→T+H 82 11 91.0 80.8
40
40
0 1 2 3 4 5
0 1 2 3 4 5
Time (years)
IHC 0,1,2+ and HER2 FISH ratio <2.0 Time (years)
100
90
AC→T+H
80
Percent
p = 0.14
70
N Events DFS AC→T
60 3 yr 5 yr
50 AC→T 44 14 82.6 60.9
AC→T+H 59 9 90.2 81.2
40
0 1 2 3 4 5
Time (years) CP1270832-140

141. •p-value for interaction = 0.38 (HER2 copy ≥ 4 only)

142. RR of ACTH/ACT for DFS (NSABP B-31)
FISH+ (1588)
Categories (N)
FISH- (207)
Interaction p=0.60 for FISH
IHC 3+ (1488) Interaction p=0.26 for IHC
IHC <3 (299)
FISH- & IHC <3 (174)
0.00 0.25 0.50 0.75 1.00 1.25 1.50
RR
Note: RR adjusted for ER and nodal status

143. HER2 Amplification and
Polysomy
• Retrospective tissue analysis of CALGB 9840
patient subset
• Polysomy 17 may be associated with increased
response to trastuzumab
• More study warranted to evaluate this
response marker
• Counting centromeres may not correlate with
degree of HER2 amplification
Response Rate, % P Value
Paclitaxel Paclitaxel +
Trastuzumab
Polysomy 17 and FISH ratio < 2 (n = 38) 26 63 .043
Kaufman PA, et al. ASCO 2007. Abstract 1009.
CEP 17 < 2.2 and FISH ratio < 2 (n = 103) 36 36 NS

144. This Situation is Quite Common
• Common clinical scenarios:
– FISH neg and IHC 1+/2+ = 40% of cases
– FISH ratio 1-2 = 25%-40% of cases
– Polysomy 17 = 8%-27% of cases
• Does give one pause…
– Retest negatives?
– Consider trastuzumab if the FISH
– ratio = 1-2, or if polysomy 17?

145. ANTI HR + & ANTI HER2 +
CROSS TALK
• TAMOXIFEN – Oldest targeted
agent (1896/1960)
• TRASTUZUMAB- Newest targeted
agent (1998)

146. CROSS TALK
• Endocrine resistance presents
major problem
• 70% Percent ER positive
• develop endocrine resistance
eventually

147. The ER Pathway
Estrogen
Cell Surface
E
R Cytoplasm
Co
Nucleus
E E A
R R
DNA
.
Roop R., Ma C., Future Oncology, In press Transcription of genes

148. Steroid receptor coactivators and
ER-dependent gene transcription
Histone
P/CAF Acetylase
CBP Activity
SRC
Family AIB1
Transcription
Estradiol-bound ER

149. SERM sensitive
estrogen
tamoxifen Corepressors
Coactivators
N-CoR/SMRT
NH2 A/B C D E/F COOH
AF-1 DBD AF-2/HBD
transcription

150. SERM resistant
estrogen
Coactivators tamoxifen
(AIB1,etc.)
Corepressors
NH2 A/B C D E/F COOH
AF-1 DBD AF-2/HBD
transcription

151. HER2/neu
JNK PI3K-Akt
src MAPK estrogen
Coactivators N-CoR
(AIB1,etc.) tamoxifen
SMRT
PSer118
NH2 A/B C D E/F COOH
AF-1 DBD AF-2/HBD
transcription

152. Non-classic Effects of ER
RTK: FGFR, IGF-
Estrogen 1R, EGFR, HER2
Cell Surface
E
R E
Cytoplasm R Ad
P
E E
R E
p R
R P MAP
AKT
C K
Nucleus
E
CoA R E
E E T T R
R R F F
DNA
Roop R, Ma C. Future
Oncology, In press.
Transcription of genes

154. Cross-talk between signal transduction
and endocrine pathways
Growth factor
Estrogen IGFR
HER2 Trastuzumab
Plasma
P P
membrane P P
P
AI P SOS
PI3-K RAS
RAF
Cell
P
survival Akt MEK
P
ER
p90RSK P
MAPK
P
Cytoplasm Cell
P P P Basal growth
P transcription
ER p160 CBP machinery
ER
Nucleus ERE ER target gene transcription
Adapted from Johnston

155. Ligand
E
ErbB ErbB
P P
P E
p85
p110 Ras ER
Akt MAPK
P E
P ER
Transcription
ER-Responsive Element

156. Crosstalk with TK pathways
• Endocrine resistance
– Cross talk with growth factor (GF) pathways
• EGFR, HER2, AKT, MAPK, PI3K
• Ligand independent pathway
– GF pathways also cause ER independent
endocrine resistance
– Novel targeted agents to inhibit these
pathways
• Goal of restoring endocrine sensitivity

157. Her2 and Endocrine Resistance
• ER+ and Her2+ breast cancer = 10%
– Less than you expect by chance
• Interaction between Her2 and ER expression
• ERE exist on promoter region of HER2 gene
• Her2 pathway facilitates endocrine
resistance
– Increases ER phosphorylation
– Disrupt interaction of ER and co-repressors
– AKT and MAPK pathways (both activated by
ER and HER2)

159. Adjuvant Endocrine Therapy Study TransATAC:
Time to Recurrence by HER2 Status
Tamoxifen Patients Anastrozole Patients
35 35
HER2+ n=839 HER2+ n=877
30 HER2 HR=2.30 30 HER2 HR=3.23
P=0.001 P<0.0001
Patients (%)
25 25
20 20
15 15
10 10
5 5
0 0
0 1 2 3 4 5 6 0 1 2 3 4 5 6
Years Years
• HER2+ status was significantly associated with reduced time to recurrence for
both tamoxifen and anastrozole
HR = hazard ratio.
Update of Dowsett and Allred. Breast Cancer Res Treat. 2006;100(suppl 1):S21. Abstract 48.

160. TransATAC: Time to Recurrence
by HER2 Status
Patients Events HR
HER2–
1526 149 0.66
HER2+ 190 45 0.92
Combined 1786 200 0.72
0.5 1.0 2.0
HR (ANA:TAM) and 95% CI
ANA TAM
better better
• HER2+ statuswas associated with substantially reduced benefit in time
to recurrence with adjuvant anastrozole compared with tamoxifen
Update of Dowsett and Allred. Breast Cancer Res Treat. 2006;100(suppl 1):S21. Abstract 48.

161. TANDEM
• Combination ER/HER2 blockade
(TANDEM)
– 207 patients postmenopausal ER+/HER2+
– Anastrazole +/- trastuzumab for metastatic
disease
– PFS prolonged for combination
• 4.8 vs. 2.4 months
– Clinical Benefit Rate 40.7 vs. 20.3
– Increased SAE’s (28% vs. 16%)
• Mostly GI toxicities, arthralgias

162. Kaufman, B., et. al., J Clin Oncol, 27:5529-5537 (2009).
TANDEM

163. ER+/HER2+ br ca is less responsive to endocrine therapy
N=1,925
De Laurentiis et al. Clin. Cancer Res. 11:4741, 2005

164. ER+/PR tumors are resistant to
tamoxifen (ATAC)
From Cui et al. JCO 23:7721, 2005

165. Negative PR is a marker of high HER1/HER2
levels and tamoxifen resistance
ER+/PR+
ER+/PR+
Arpino et al. JNCI 97:1254, 2005

166. Negative PR is a marker of high HER1/HER2
levels and tamoxifen resistance
ER+/PR+ ER+/PR
ER+/PR+ ER+/PR
Arpino et al. JNCI 97:1254, 2005

167. Randomized Phase II Trial of Tamoxifen ±
Gefitinib in MBC (ZD1839IL/0225)
Primary Endpoint
274 Pts R • TTP
a Tamoxifen + Gefitinib
20 mg/day 250 mg/day
n Secondary
Strata 1 d
•No prior Tam o • ORR & CBR
•Prior Tam > m
1yr i
Tamoxifen + Placebo Exploratory
Strata 2 z
20 mg/day • IHC study of
•Prior AI e
downstream effectors
of erbB family and ER
& co-activators (AIB1)
Study Chair: Kent Osborne, Baylor College Medicine.
PI CK Osborne, Baylor College Medicine

168. Tamoxifen-resistant breast tumors acquire
ErbB receptor overexpression
Tam-S Tam-R
EGFR
HER2
Knowlden et al. Endocrinology 144:1032, 2003
10% ‘conversion rate’ to HER2 overexpression in breast cancers
that recur (early) on adjuvant tamoxifen (Gutierrez et al. J. Clin.
Oncol. 23:2469, 2005)

169. Neoadjuvant aromatase inhibitors (AI) are better
than tamoxifen against ER+/HER2+ breast cancer
Osborne and Schiff J. Clin. Oncol. 23:1616, 2005

170. Gefitinib blocks HER2 signaling and restores
letrozole-
mediated growth inhibition of breast cancer
cells
HER2
IP:HER2 P-Tyr
Akt 40
35
Ser473 P-Akt
Colonies (10-1)
30
25
MAPK 20
15
P-MAPK 10
5
ER 0
AD - + + + +
Ser167 P-ER Letrozole - - + - +
Gefitinib - - - + +
Ser118 P-ER
LY294002 - + - -
U0126 - -+ -
Gefitinib - - - +
Shin et al. Submitted

171. Is EGFR/HER2 signaling upregulated after
escape from estrogen depletion?
 Estrogen depletion upregulates EGFR transcription
(EGFR gene contains a 96-bp intron fragment that is
repressed by estradiol)
– Wilson and Chrysogelos, J. Cell Biochem. 85:601, 2002
 ER+ breast cancer cells selected for resistance to
fulvestrant show EGFR and P-MAPK levels
– McClelland et al., Endocrinology 142:2776, 2001
 Resistance to fulvestrant does not occur if selection is
done in the presence of gefitinib or MAPK inhibitors
 MCF-7/aromatase cells that become resistant to
letrozole overexpress HER2 and P-MAPK; resistance is
reversed by gefitinib or MEK inhibitors
– Jelovac et al. Cancer Res. 65:5380, 2005; Sabnis et al. ibid
65:3903, 2005

172. Serum HER2 converts to positive at disease
progression
in patients with breast cancer on hormonal therapy
Letrozole 29/111 (26%)
Tamoxifen 32/129 (25%)
Lipton et al. Cancer 104:257, 2005

173. EGFR
 Interaction of with ER less studied than HER2
 Like HER2 can activate downstream pathways
– MAPK
 Can form heterodimers with HER2 receptors
– Gefitinib prevented heterodimer
formation/phosphorylation
– Reversed tamoxifen resistance MCF-7 cell line

174. Clinical data
 56 postmenopausal patients ER+ and EGFR+
– Gefitinib + Anastrazole or placebo
– 4-6 weeks prior to surgery
– Primary endpoint cell cycle inhibition (Ki67)
– Combination arm showed higher Ki67 reduction
– 5.6% relative difference P=0.0054
– Tumor response rates were similar
– Showed that combination is tolerable
Polychronis, A., et. al., Lancet oncology 6:383-91 (2005).

175. IGF-1 Pathway
 TK receptor
 TK activity on transmembrane subunit
 Has homology with the insulin receptor
– IGF-1R and IR can form hybrid receptors
– IGF-1R and IR can bind each other’s ligands
 IGF-1R expressed in ~45% of breast cancers (by IHC)

176. Belinsky, M.G., et. al., Cell Cycle 7:19, 2949-2955 (2008).

177. IGF-1 system and Endocrine
Resistance
 Crosstalk with ER pathway similar to other TK
 IGFR inhibitors + endocrine therapy
– Negative trials (two monoclonal Ab)
– At least two ongoing (one small molecule TKI, one
monoclonal Ab)
 Data too sparse to make any judgments

178. FGF Pathway
 Fibroblast growth factor pathway TKR
 FGF1-FGF4 (4 different genes)
– Alternate splicing results in many isoforms
– At least 18 ligands that can bind FGF receptors
– Activates downstream pathways similar to other TKI

179. FGF pathway
18 different ligands
FGFR(1-4)
Cell Surface
Cytoplasm
PI3
Ras
K
MAP
AKT
K
Gene
Growth TF
p Expression Invasion
Roop R, Ma C., Future Oncology, In press.

180. FGF and endocrine resistance
 Amplification of FGFR1 in 10% breast CA
– FGFR1 amplified cell line resistant to tamoxifen
 FGFR1 amplified ER+ tumors usually PR-
– Tend to have higher Ki67
– ? Role in luminal B breast cancers
 FGFR3 activation in cell lines decreased sensitivity to
endocrine therapy
 FGFR4 transcription predicts Tamoxifen sensitivity
clinically

181. PI3K
 Activated by TK or G-protein
 Divided into 3 subclasses (I-III)
– Subclass I divided Ia and Ib
 Heterodimers p110 and p85
– Three isoforms p110 exist (α, β, δ)
 PI3K activation
– Activates AKT and interacts with mTOR

182. Roop R., Ma C., Future
Oncology, In press.
PI3K/AKT/mTOR Pathway
RTK: FGFR, IGF-
1R, EGFR, HER2
PIP
p8
2 PTE
5
RAS p110 N
PIP
3
PDK1
MAPK mTOR-
AKT mTOR-
C2
C1
Downstream Target
growth, Invasion
Proteins

183. Increased PI3K Pathway Activity
 Promotes endocrine resistance (LTED cells)
 Ways PI3K can be overactivated
– PIK3CA (encodes p110α) mutated 30-40% time
– Loss of PTEN
– Amplification PIK3CB (encodes p110β)
– Mutations in AKT

184. PI3K Inhibition
 Cell line experiments
– ER+ and PIK3CA or PIK3CB silenced (RNAi)
– Apoptosis and growth inhibition
– PIK3CA ≥ PIK3CB
– Combined PIK3CA and PIK3CB greatest
– Apoptosis dependent on estrogen depletion
– ER negative cell line had no effect

185. Clinical trials PI3K
 A few clinical trials underway
– Endocrine therapy + BKM120 or BEZ235
– Letrozole + XL147 or XL765
 Data is very early for this class of drugs

186. mTOR
 Downstream of PI3K pathway
 Rational target for treating endocrine resistance
 Phase III letrozole +/- temsirolimus (metastatic)
– negative
 Phase II trial neo-adjuvant letrozole +/- everolimus
– positive
 TAMRAD – phase II tamoxifen +/- everolimus
– positive

187. ONCOGENES AND SIGNALING
MOLECULES THAT BEEN ASSOCIATED
WITH ANTIESTROGEN RESISTANCE
Ha-Ras Cox-2 IGF-II
Src Heregulin FGF-1/4
Erk (MAPK) VEGF p38Mapk
IGF-I receptor
Cyclin D1 AIB-1 (SRC-1)
and IRS-1
Cyr61 (ligand for
Activated Akt p130Cas
avb3)

188. Summary/Conclusions
 High ER and PR+ predict good response to tamoxifen
 Negative PR and high EGFR/HER2 predict early
escape
 ER+/HER2+ tumors are initially more responsive to
AIs than tamoxifen
 EGFR/HER2 overexpression occurs at the time of
escape from hormonal therapy
 Blockade of EGFR/HER2 is one of many approaches to
enhance hormonal therapy action
 We need new clinical paradigms to elucidate the
preferential mechanisms of escape from endocrine
therapy as well as to prioritize combinatorial
molecular strategies

189. In summary
Her2+ Her2-
Agressive Non agressive Agressive Non
agressive
Trastuzumab Trastuzumab +
RH+ + CT Hormono PolyCT Hormono
Trastuzumab Sequentiel
Trastuzumab PolyCT
RH- + CT
+/- CT
MonoCT,
PolyCT?

190. LESSONS LEARNED
SO FAR
190

191. Prognosis for Patients With
HER2+ Breast Cancer
• HER2 positivity is an independent predictor of poor prognosis
• HER2 positivity predicts response and survival
• HER2 positivity also correlates with other clinical pathologic variables
– Short disease-free interval
– Larger tumour size
– Positive nodal status
– Ductal rather than lobular histology
– Ploidy
– High S-phase fraction
– High nuclear grade
– Mutated p53
– Decreased ER and PgR expression
HER = human epidermal growth factor receptor; ER = oestrogen receptor; PgR = progesterone receptor.
Berger et al. Cancer Res. 1988;48:1238; Chazin et al. Oncogene. 1992;7:1859; Hynes and Stern. Biochim Biophys Acta.
1994;1198:165; O’Reilly et al. Br J Cancer. 1991;63:444; Paik et al. J Clin Oncol. 1990;8:103; Press et al. J Clin Oncol.
1997;15:2894; Slamon et al. Science. 1987;235:177; van de Vijver et al. N Engl J Med. 1988;319:1239.

192. All HER2/neu Might Not Be
Created Equal . . .

193. Multiple Approaches to Targeting the
HER Pathways
Extracellular Trastuzumab
domain binding site
HER2 receptors
Lapatinib
Intracellular binding site
domain
Activation mediates multiple processes

194. Multiple Approaches to Targeting the
HER Pathways (cont’d)
Truncation of HER2
Truncated HER2 continues
to mediate multiple
processes

195. Adjuvant Node+, HER2+
Unresolved Clinical Questions
• Concurrent vs sequential trastuzumab
• Which chemotherapy regimen?
• Anthracycline or not?
• Duration of trastuzumab?
• Endocrine therapy plus trastuzumab only?
• Trastuzumab alone?

196. Traditional
approach Node
Node positive
negative
New Approach
ER-, PR- ER+
HER-2+ ER++
HER-2- Luminal A Luminal B
Virulent Basal-like
Virulent Indolent Virulent

197. Indolent ER and PR-Positive Breast
Cancer
E2
ER
Breast Cancer
•Tamoxifen and
P Aromatase
Inhibitor-
responsive
PR

198. More Virulent ER+ Breast Cancer
TAM
IGFR
Her- Her-2
1
ER
+
AIB1
An estrogen response
coactivator
PR
Schiff R, J Natl Cancer Inst 2003;95:353 - 361

199. Cancer stem cells: are we missing the target?
Jones et al. JNCI 96:583, 2004
Courtesy of Jenny Chang, MD

200. Cancer stem cells: are we missing the target?
Jones et al. JNCI 96:583, 2004

201. Cancer stem cells: are we missing the target?
Jones et al. JNCI 96:583, 2004

202. Breast Stem Cell Survival
Hedgehog
Notch family TGFβ
family family
Wnt Growth
family Hormone
/Insulin-like GF
EGF
Estrogen
family
Progesterone
Self-renewal
FGF
family Prolactin
Stem Cell
Modified from Clarke et al 2005

203. Breast Cancer Lab Report of the Future
Jones, Mary A. DOR: 01/31/25
MR#:555690 Dx: Breast cancer
Receptor status: ER-, Her2-, PR-, AR-
Activated pathway: Insulin-like growth factor receptor, AKT,
mTOR Basal-like Breast Cancer
Percentage of patients that have had this pathway
activated in breast cancer: 16%

204. Breast Cancer Lab Report of the Future
Jones, Mary A. DOR: 01/31/25
MR#:555690 Dx: Breast cancer
Receptor status: ER-, Her2-, PR-, AR-
Activated pathway: Insulin-like growth factor
receptor, AKT, mTOR Basal-like Breast Cancer
Percentage of patients that have had this pathway
activated in breast cancer: 16%
Potential therapies:
RAD0001
Imatinib
Anti-IGFR antibody
Metformin
Exercise, low fat diet

205. Breast Cancer Mortality
Will the Progress Continue?
Queen-
Clinical size panty
Trials in Hose –
Breast one
Cancer size does not fit
Subtypes all !

206. The Future Is Possible
I don’t know if heavier than air flight
is possible, but I’m committed to living
my life dedicated to its possibility.
- Wilbur Wright

207. QUESTIONS ???