Jesus Garcia-Foncillas MD PhD

Director

Aflibercept, ¿qué hemos aprendido y qué eslo nuevo que viene?

New insights in Angiogenesis

• Relevance and cornerstones in angiogenesis

New insights in Angiogenesis

• Relevance and cornerstones in angiogenesis

• Angiogenesis initiator

Pharmacologic Approaches to Blocking Angiogenesis

Neutralizing VEGF activity(e.g., bevacizumab, aflibercept)1,2

Inhibition of receptorkinase activity (e.g., sunitinib,sorafenib, pazopanib, etc.)3-5

Reducing expression of VEGFby inhibiting tumor growth pathways(e.g., anti-EGFR therapies)6-9

THREE GENERAL MECHANISMS OF ANGIOGENESISINHIBITORS THAT BLOCK THE VEGF PATHWAY

1. Avastin PI. 2010. Genentech Inc. 2. Ada[ted from Holash. PNAS. 2002;99:11393–11398. 3. Sutent PI. 2010. 4. Nexavar PI. 2010. 5. Votrient PI. 2010.6. Adapted from Petit. Am J Pathol. 1997;151:1523–1530. 7. Tarceva PI. 2010. 8. Erbitux PI 2010. 9. Vectibix PI. 2010.

Block VEGF receptor•Anti-VEGFR-2 TKIs

(sunitinib, etc)

VEGF

Neutralize VEGF•Aflibercept•Bevacizumab

Extracellular

VEGFR-2Tumorcell

Block VEGFexpression•Erlotinib•Cetuximab

•Panitumumab

Intracellular

ANGIOGENESIS INHIBITORS

New insights in Angiogenesis

• Relevance and cornerstones in angiogenesis

• Angiogenesis initiator

• Other angiogenesis initiators

The precise role of PlGF and VEGFR-1in angiogenesis remains unclear¹

When VEGF-A levels are reduced oractivation of VEGFR-2 is reduced byan antagonist, PlGF and VEGFR-1 maycontribute towards a rescue signalenabling pathological angiogenesis¹

VEGF-APIGF

The Potential Role of PlGF and VEGFR-1 in Angiogenesis

1. Adapted from Brave. Angiogenesis. 2010;13:337–347.

VEGF-A, VEGF-B, and PlGF are Involvedin Multiple Pathways of Angiogenic Response

ENDOTHELIAL CELLSurvival, Migration, Proliferation

VEGF-ASTROMAL CELLPERICYTE, SMC

MigrationProliferation

sVEGFR-1

MACROPHAGERecruitment and activation

Release of angiogenic factors

BM PROGENITORS LEUKEMIC CELLProliferation, Migration, Survival

DENDRITIC CELLSuppression ofantigen recognition

VEGF-A

TUMOR CELLProliferationand migrationChemoprotection

PlGF

VEGF-B

1. Adapted from Fischer. Nat Cancer Rev. 2008;8:942–956.

New insights in Angiogenesis

Relevance and cornerstones in angiogenesis

Angiogenesis initiator

Other angiogenesis initiators

Dual antiangiogenic therapy

Aflibercept: Specifically Engineered forHigh Binding Affinity and to Optimize Pharmacokinetics

Fusion protein of key domains fromhuman VEGF receptors 1 and 2 withhuman IgG Fc¹

Blocks all VEGF-A isoforms, VEGF-B andplacental growth factor (PlGF)²

High affinity—binds VEGF-A and PlGFmore tightly than native receptors

Contains human amino acidsequences¹

1. Adapted from Holash. Proc Natl Acad Sci. 2002;99:11393–11398. 2. Adapted from Tew. Clin Cancer Res. 2010;16:358–366.

New insights in Angiogenesis

Relevance and cornerstones in angiogenesis

Angiogenesis initiator

Other angiogenesis initiators

Dual antiangiogenic therapy

Is dual antiangiogenic therapy better than single therapy?

Anti-tumor response to aflibercept and Bevacizumab in Patient-Derived Xenografts (PDX) tumors grown in mice

● Goal: Differentiate Aflibercept vs bevacizumab

● Identify phenotypes of response to aflibercept vs bevacizumab

● Identify predictors of response to aflibercept vs bevacizumab

● Study design: Creating 48 PDX CRC Models

48 PDX CRC

models

Surgically removed

tumors engrafted into

NMRI nude mice

Expansion Phase

Profiling

Expression of human &

mouse VEGF-A, PlGF,

VEGFR1 in untreated

tumors

Consenting patients

(N=48) with metastatic

(N=39), recurrent (N=1) or

primary (N=8) CRC

PDX models better mimic human disease based on histology, molecular heterogeneity as well as clinical response

Efficacy endpoints: • Tumor growth inhibition

∆T/∆C value of < = 0% complete tumor stasis ∆T & ∆C: Changes in tumor volume for treated (T) and control (C) groups

• Statistical comparison of tumor volumes using student’s t-tests. P < 0.05 statistically significant.

Treatment Phase in 48 PDX models

Ziv-aflibercept

25 mg/kg sc

n=8

Bevacizumab

25 mg/kg IV

n=8

Placebo

n=8

Treated twice per week for 3-4 weeks

Subcutaneous tumor measurements were recorded twice per week

Anti-tumor response to aflibercept and Bevacizumab in Patient-Derived Xenografts (PDX) tumors grown in mice

Distribution of tumor growth inhibition (dT/dC) induced by aflibercept and bevacizumab across PDX models

-60

-50

-40

-30

-20

-10

0

10

20

30

40

50

60

70

80

90

100

Aflibercept

Bevacizumab

DT

DC

(%)

CX

F2

069

CX

F2

070

CX

F2

025

CX

F2

061

CX

F1

58

CX

F2

068

CX

F1

784

CX

F2

081

CX

F2

036

CX

F2

163

CX

F2

039

CX

F2

067

CX

F1

753

CX

F1

034

CX

F2

102

CX

F2

032

CX

F2

073

CX

F23

3

CX

F2

02

9

CX

F1

99

1

CX

F1

08

6

CX

F1

10

3

CX

F2

43

CX

F1

72

9

CX

F2

80

CX

F2

12

9

CX

F2

06

5

CX

F19

72

CX

F20

83

CX

F53

3

CX

F20

48

CX

F50

4

CX

F67

6

CX

F64

7

CX

F1

78

3

CX

F7

42

CX

F9

75

CX

F2

04

9

CX

F1

09

6

CX

F1

25

6

CX

F1

04

4

CX

F1

78

8

CX

F1

29

7

CX

F26

0

CX

F2

12

7

CX

F2

06

6

CX

F20

85

CX

F6

09

Inactive

MostActive

* * * * * * *

The relative changes of tumor volume in response to treatment were determined at end point in 48 patient-derived xenograft (PDX) models of colorectal cancer

TU

MO

R V

OLU

ME

, m

m³

DAYS AFTER RANDOMIZATION

200 4 8 12 16

1200

600

300

0

900

PDX Tumor Model: CXF2067 (N=30)

TU

MO

R V

OLU

ME

, m

m³

DAYS AFTER RANDOMIZATION

200 4 8 12 16

400

200

100

0

300

PDX Tumor Model: CXF2048 (N=9)

TU

MO

R V

OLU

ME

, m

m³

DAYS AFTER RANDOMIZATION

200 4 8 12 16

1600

800

400

0

1200

PDX Tumor Model: CXF1729 (N=8)

TU

MO

R V

OLU

ME

, m

m³

DAYS AFTER RANDOMIZATION

200 4 8 12 16

1600

800

400

0

1200

PDX Tumor Model: CXF2069 N=1

Ziv-aflibercept is active whereas bevacizumab is not

Phenotye A: Ziv-aflibercept activity is greater than bevacizumab (39 tumors)

Phenotype B : Ziv-aflibercept activity is equivalent to bevacizumab (9 tumors)*

Ziv-aflibercept and bevacizumab are both inactive

Ziv-aflibercept is more active than bevacizumab

Ziv-aflibercept and bevacizumab are both active

Aflibercept Bevacizumab Placebo

Identification of two phenotypes based on differential response to aflibercept and bevacizumab.

Mice bearing subcutaneous (SC) patient-derived tumors of colorectal cancer (CRC) were treated with aflibercept () SC injection, 2x/week, 25 mg/kg), placebo () (SC injection, 2x/week), or bevacizumab (▲) (IV injection, 2x/week, 25 mg/kg) for a total of 3 weeks. Tumor measurements were recorded twice per week

* the extent of the response to Aflibercept is greater than Bevacizumab

VEGF-A levels

New insights in Angiogenesis

Relevance and cornerstones in angiogenesis

Angiogenesis initiator

Other angiogenesis initiators

Dual antiangiogenic therapy

Is dual antiangiogenic therapy better than single therapy?

Aflibercept biomarkers

The biomarkers sample population characteristics do not differ markedly from the overall VELOUR population

characteristics

Demographics: Age, Gender, Race, Country & ECOG status characteristics are similar

Notable: patients with prior bevacizumab therapy may be more highly represented in the Biomarker group compared to the VELOUR group

OS:HR = 0.809 in Biomarkers groupHR = 0.817 in VELOUR group

PFS:HR = 0.752 in Biomarkers groupHR = 0.758 in VELOUR group

VEGF PATHWAY MEMBERS

VEGF-A

interaction p-value: 0.026

VEGF Low / F

VEGF Low / F+ZT

VEGF High / F+ZT

VEGF High / F

VEGF Low / F

VEGF Low / F+ZT

VEGF High / F+ZT

VEGF High / F

13.1 mo

12.8 mo

9.7 mo

12.5 mo

5.5 mo

6.8 mo

3.9 mo

6.6 mo

Median: 142 pg/ml (min, max: 25, 2350 pg/ml)

VEGF-A may be a predictive biomarkerOS: Patients with high plasma VEGF who were treated with ZALTRAP do better than control group (blue vs green lines): HR= 0.644, 95% CI: 0.49 – 0.85; p value: 0.0013

PFS: comparable results with HR= 0.599, 95% CI: 0.453 - 0.792, p value: 0.0002

sVEGFR2

VEGFR2 may be a predictive biomarker

OS: Patients with high plasma VEGFR2 who were treated with ZALTRAP may do better than control group (blue vs green lines): HR= 0.686, 95% CI: 0.49 – 0.85; p value: 0.008,

PFS: comparable results with HR= 0.679, 95% CI: 0.516 - 0.893; p value: 0.005

VEGF-R3 Low / F

VEGF-R3 Low / F+ZT

VEGF-R3 High /F+ZT

VEGF-R3 High / F

11.0 mo

12.1 mo

11.3 mo

14.5 mo

VEGF-R2 Low / F

VEGF-R2 Low / F+ZT

VEGF-R2 High /F+ZT

VEGF-R2 High / F

11.0 mo

12.1 mo

11.3 mo

14.5 mo

sVEGFR3

VEGFR3 may be a predictive biomarker

– OS: Patients with high plasma VEGFR3 who were treated with ZALTRAP may do better than control group (blue vs green lines): HR= 0.686; p value: 0.006

– PFS: comparable results with HR= 0.711, 95% CI: 0.545 – 0.927, p value: 0.01

VEGF-R3 Low / F

VEGF-R3 Low / F+ZT

VEGF-R3 High /F+ZT

VEGF-R3 High / F

11.9 mo

12.7 mo

10.7 mo

12.8 mo

SPD AND MIF MAY BE PREDICTIVE IN VELOUR

MIF

MIF Low / F

MIF Low / F+ZT

MIF High /F+ZT

MIF High / F

13.8 mo

12.9 mo

9.5 mo

12.7 mo

MIF may be a predictive biomarker

OS: Patients with high plasma MIF who were treated with ZALTRAP may do better than control group (blue vs green lines): HR= 0.67, 95% CI: 0.512 –0.875; p value: 0.003

PFS: similar results with HR= 0.607, 95% CI: 0.463 – 0.797; p value: <0.001

Response rate: comparable results -response increased from 10.5% (control) to 20.9% (ZT)

Surfactant Protein D

SPD Low / F

SPD Low / F+ZT

SPD High /F+ZT

SPD High / F

9.7 mo

13.9 mo

13.0 mo

12.3 mo

SPD (low) may be a predictivebiomarker

OS: Patients with low plasma SPD who were treated with ZALTRAP may do better than control group (red vs black lines): HR= 0.598, 95% CI: 0.453 –0.791; p value: <0.001

PFS: comparable results with HR= 0.581, 95% CI: 0.437 - 0.773, p value: <0.001

Response rate: comparable results with low SPD group ORR increased from 8.1% (control) to 25.6% (ZT)

INTERLEUKINE 8 (IL-8) MAY BE PREDICTIVE AND PROGNOSTIC IN VELOUR

Interleukine-8 (IL-8)

Prognostic

IL-8 Low / F

IL-8 Low / F+ZT

IL-8 High / F+ZT

IL-8 High / F

19.8 mo

18.8 mo

8.0 mo

9.4 mo

IL-8 Low / F

IL-8 Low / F+ZT

IL-8 High / F+ZT

IL-8 High / F

6.7 mo

8.3 mo

3.9 mo

4.9 mo

IL-8Low / F

IL-8Low /F+ZT

IL-8High /F+ZT

IL-8 High / F

Median: 20 pg/ml (min, max: 2, 4505 pg/ml)

IL-8 may be a prognostic biomarker– OS: Patients with low plasma IL-8 levels may do better in VELOUR (red vs blue & black vs green lines)

– Prognostic effect (Risk) High IL-8: HR=2.319 in ZT; 4.48 in control, p<0.001

IL-8 may be a predictive biomarker– PFS: comparable results with HR= 0.694, 95% CI: 0.534 – 0.902; p value: 0.005

– Response: comparable results with high IL-8 group response increased from 6.9% (F) to 11.9% (F+ZT)

New insights in Angiogenesis

Relevance and cornerstones in angiogenesis

Angiogenesis initiator

Other angiogenesis initiators

Dual antiangiogenic therapy

Is dual antiangiogenic therapy better than single therapy?

Aflibercept biomarkers

New biological features

Aflibercept increased the sensitivity of HCT-116/5-FU cells to 5-FU under hypoxia.

Improved sensitivity to 5-FU

Angiogenesis is a cornerstone in the carcinogenesis where multiple typesof cells and signaling pathways are involved.

VEGF is relevant but is not enough an efficient target where otherdifferent molecules may initiate angiogenesis in an independent way

Dual antiangiogenic therapy provides better possibilities of controllingneoangiogenesis and tumor proliferation

Aflibercept offers more biological advantages than other anti-angiogenicapproaches currently available