A Phase I Study of ENMD-2076, a Unique Aurora A, Angiogenic Kinase

Inhibitor, Administered Orally to Patients with Advanced Cancer

B.R. Bastos1, J. Diamond2, R. Hansen3, D. Gustafson3, J. Arnott4, M. Bray4, C. Sidor4, W. Messersmith2, G. Shapiro1

1Dana-Farber Cancer Institute, Boston, MA; 2University of Colorado Health Science Center, Aurora, CO; 3Colorado State University, Fort Collins, CO;

4EntreMed, Inc., Rockville, MD

ENMD-2076 – A Kinase Inhibitor With a Unique Target Profile

• Orally active, multi-kinase inhibitor currently in Phase 1 clinical

trials

• Unique combination of target activities

– Angiogenic Kinases (VEGFRs, FGFRs)

– Cell Cycle Kinases (Aurora A)

– Growth Factor Kinases (c-Kit, Flt-3, CSF1R)

• Excellent preclinical antitumor activity as single agent or in

combination

• Acceptable/manageable toxicity profile

ENMD-2076 is an Orally Available Aurora A and Angiogenic Kinase Inhibitor

AngiogenicKinases

(Flt-3, c-Kit, CSF1R)

Aurora Kinases

ENMD-2076

Growth Factor Kinases

(Aurora A)

(VEGFR; FGFR)

ENMD-2076 Experimental Results Consistent with Selective Inhibition of Aurora A In Vitro & In Vivo

• ENMD-2076 is selective for the Aurora A isoform

– Recombinant enzyme IC50s: Aur A = 14 nM, Aur B = 290 nM

– Cellular enzyme IC50s: Aur A = 130 nM, Aur B/C = 450 nM

• ENMD-2076 induces G2/M cell cycle arrest & apoptosis rather than endoreduplication/aneuploidy characteristic of Aur B/C inhibition

• ENMD-2076 induces abnormal mitosis, dose-dependent decrease in centrosomal distance, & monospindle phenotype characteristic of Aur A inhibition

• ENMD-2076 induces dose dependent increase in phospho-histone H3, decrease in anaphase profiles & inhibition of Aur A & TACC3 co-localization in treated tumors

ENMD-2076 PharmacologyMDA-MB-231 Human Tumor Xenograft Model

Female CB17 SCID mice - 10/group

Tumor implanted mfp on Day 0

Rx initiated on Day 32

0

500

1000

1500

2000

2500

Days Following Tumor Implant

Tum

or V

olum

e (m

m3 )

30 35 40 45 50Rx

CD31 (Day 49)

Vehicle

50 mg/kg

200 mg/kg

32,8058,208AUC (h∗ng/mL)

3,8701,483Cmax (ng/mL)

70% Regression51% TGIResponse

200 ( )50 ( )

Dose (mg/kg)

Tumor Regression and Inhibition of Angiogenesis in ENMD-2076 Treated HT-29 Xenografts

ENMD-2076 200 mg/kg/d PO Day 28

HT-29 Colon Cancer XenograftVehicle Control Day 28

J. Tentler UCDHSC

ENMD-2076 Induces Regression of MV4;11 AML (Flt-3-ITD) Tumor Xenografts

Female NCr nude mice -10/group

Tumor implanted sc on Day 0

Tum

or V

olum

e (m

m3 )

• Dose escalated to 150 mg/kg in 15 & 30 mg/kg cohorts

• Rx in 75 & 150 mg/kg arms stopped

No Rx

Vehicle

15 mg/kg

30 mg/kg

75 mg/kg

150 mg/kg

Study Termination

0

200

400

600

800

1000

1200

1400

1600

20 25 30 35 40 45 50 55 60 65 70 75 80 85 90

Days Following Tumor Implant

RxStartedDay 24

Rx changed

Free Base used

75 & 150 mg/kg doses of ENMD-2076 produce regressions; large tumorsgrown under lower exposures respond rapidly to increased dose

Antiangiogenic Effects Demonstrated with DCE-MRI Analysis of HT-29 Tumor Xenografts In Mice Treated With ENMD-2076

Pre-contrast 2.5 min

VEH - BL

ENMD - BL

VEH - WK1

ENMD - WK1

VEH - WK3

ENMD - WK3

0.0

0.2

0.4

0.6

0.8

1.0

AUC

Κep

Vehicle ENMD-2076Pre-contrast 2.5 min

Baseline

Day 7 Day 7

Day 21 Day 21

Baseline

VEH - BL

ENMD - BL

VEH - WK1

ENMD - WK1

VEH - WK3

ENMD - WK3

0.0

0.2

0.40.6

1.1

1.6

Ktr

ans (m

in-1

)

IAUC

Ktrans

K ep

VEH - BLENMD - BLVEH - WK1

ENMD - WK1VEH - WK3

ENMD - WK3

0

1

2

3

IAU

C (x

104 )

VEH - BLENMD - BLVEH - WK1

ENMD - WK1VEH - WK3

ENMD - WK3

0

10

20

30

40

50

AU

C (x

104 )

• Toxicology

– 28-day continuous oral dosing (rats & dogs)• Dose-proportional increases in Cmax & AUC

• GI effects observed in both rats & dogs

• Reversible hard tissue effects in rats suggestive of kinase & antiangiogenic activities

• Reversible LFTs & hematological effects in rats

• Safety Pharmacology

– IC50 hERG channel = 300 nM

– Telemetry study in non-anesthetized dogs demonstrated no effects on cardiovascular or respiratory system

• In vitro mutagenicity studies negative

ENMD-2076 – Toxicology Results

A Phase I Study of ENMD-2076, a Unique Aurora A, Angiogenic KinaseInhibitor, Administered Orally to Patients with Advanced Cancer

• Objectives– To assess safety and tolerability and to determine the maximum

tolerated dose (MTD) of ENMD-2076 administered orally in patients with advanced cancer over a range of doses

– To determine plasma pharmacokinetics of ENMD-2076 administered orally

– To investigate clinical efficacy in patients with advanced cancer

• Primary Eligibility Requirements– Advanced cancer with progression on current therapy or for which no

curative therapy exists– ECOG PS 0-1– Adequate organ function

• No uncontrolled severe heart failure, Afib, ↑QTc• No uncontrolled severe hypertension• No > 2+ proteinuria or nephrotic syndrome

– Modified RECIST criteria or clinically evaluable disease– Age ≥18 years– Signed informed consent

Demographics and Dose Escalation

25Enrolled

12:13Male: Female

Colorectal = 8Ovarian = 7

Renal cell = 3Pancreatic = 2

Thyroid = 1 Neuroendocrine = 1

Urachal = 1Melanoma = 1Cervical = 1

Tumor Type

61Median Age 60 mg/m2

80 mg/m2

120 mg/m2

200 mg/m2

160 mg/m2

3 (or 4) + 3 (or 2) dose escalation design

ENMD-2076 given orally once daily in 28-day cycles

Dose-Limiting Toxicities (DLT)

PendingPending4160 mg/m2

Grade 3 HTNGrade 3

Neutropenia277200 mg/m2

033120 mg/m2

04480 mg/m2

Grade 3 CholecystitisGrade 4 HTN

16760 mg/m2

DLT DLT (#)EvaluableEnrolledENMD-2076 Dose

Treatment-Related Toxicities (n=22)

1 (5%)1 (5%)Pain

2 (9%)2 (9%)Nausea2 (9%)2 (9%)Neutropenia

5 (22%)5 (22%)Diarrhea

1 (5%)1 (5%)Elevated SGOT

1 (5%)1 (5%)Cholecystitis1 (5%)1 (5%)Elevated SGPT

2 (9%)2 (9%)Proteinuria2 (9%)2 (9%)Mucositis2 (9%)2 (9%)Pruritus

3 (14%)3 (14%)Headache3 (14%)3 (14%)Fatigue

8 (36%)1 (5%)2 (9%)5 (22%)HypertensionTotal*Grade 4Grade 3Grade 1/2Toxicity

*Grade 1 or 2 treatment-related AEs in ≥ 2 patients and all Grade 3 or 4 treatment–related AEs

Pharmacokinetic Results (Day 28)

Dose (mg/m2/day)

0

100

200

300

400

500

600

700

800

900

C max(ng/mL)

0

10000

20000

30000

40000

50000

60000

70000

20060 80 120 20060 80 120

AUC 0

→inf(ngx hr/m

L)

t1/2

CL/F

Vd/F

= 49.19 hr

= 12.62 L/hr

= 844.27 L

Steady state concentrations of ENMD‐2076 approached or exceeded 1μM (375 ng/mL) with a half‐life of 40 to 60 hours

Dose (mg/m2/day)

Response of Plasma Soluble KDR to Treatment with ENMD-2076 (Cohorts 1 - 4)

Mean plasma soluble KDR went from 9977 pg/mL (95% CI 8901,10992) at Baseline to 7109 pg/mL (95% CI 6429, 7789) at C1D28

9977

7109*

*p<0.00010

2000

4000

6000

8000

10000

12000

14000

Baseline D28

Plasma sKDR(pg/mL)

Waterfall Plot of RECIST Measurements (n=19)

-30%

-20%

-10%

0%

10%

20%

30%

40%

50%

60%

color

ecta

l (1)

panc

reat

ic (1

)

rect

al (4

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colon

(2)

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(3)

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(4)

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Bes

t Res

pons

e fro

m B

asel

ine

(1) 60 mg/m2/d cohort(2) 80 mg/m2/d cohort(3) 120 mg/m2/d cohort(4) 200 mg/m2/d cohort

CEA+95.5%

CEA+57.7%

CEA+99.2%

CEA-0.82%

CA-125-15.4%

CEA-8.8%

CEA+56.3% CEA

+28.7%CA-125-16.8%

CEA-79.2%

CEA-59.0%

CA-125-13.7%

CA-125-40.9%

Study 2076-CL-001 Results/Conclusions

• Dose limiting toxicities of hypertension and neutropenia are observed at the 200 mg/m2/d dose level

• Steady state concentrations of ENMD-2076 approach 1μM with a half life of 40 to 60 hours; an active metabolite, ENMD-2060, is present at concentrations of 0.3μM with a half life of 100 hours

• Plasma soluble KDR reductions are seen in all patients when compared to their baseline

• Clinical benefit has been demonstrated with reductions in tumor volume, reductions in tumor markers, and improvement in cancer-related symptoms in melanoma, renal cell, ovarian, and colorectal cancer patients

• Expanded Phase 1 cohorts at the MTD and trials in myeloma and leukemia are underway that incorporate additional PD assessments

• ENMD-2076 represents an active and unique oral kinase inhibitor that targets a combination of Aurora A and angiogenesis kinases