poster055.pdf

Discovery and development of
MGCD0103
an orally active, isotype selective
HDAC inhibitor
in Human clinical trials
Arkadii Vaisburg
MethylGene/Pharmion/Taiho
Istanbul, August 31, 2006
A. Vaisburg, August 31, 2006 1

Proposed Mechanism of
Action of Histone Deacetylase Inhibitors
Biochem. J. (2003), 370, 737-749
A. Vaisburg, August 31, 2006 2

Proposed Mechanism for the Deacetylation of
Acetylated lysine
HDLP active-site residues and their
proposed HDAC1 counterparts (in
parentheses) are labeled.
A. Vaisburg, August 31, 2006 3

Space-filling Representation of TSA in the
HDLP Active Site
O
NHOH
Me
N
Me Me
Me Trichostatin
(TSA)
A. Vaisburg, August 31, 2006 4
O

O
NH
S
O
HDAC inhibitors in clinical development
PXD-101
CuraGene
NH
O
SAHA
(Aton Pharma/Merck)
O
O
NHOH
O
NH
NH
NH
O
O
O
NH
O
FK228
De psipeptide
(Fujisawa)
O
NHOH
OH
N
H LAQ-82 4
(Novartis AG)
S
S
N
N
O
O
O
O
Me
Me
N
NHOH
NH
NH
O
CRA-024781
(Celera Genomics)
MS -275
(Schering AG)
O
NH Me
O
NH
The compound has been used as a
positive control in many experiments
NH
NH2
O
NHOH
LBH-589
(Novartis AG)
A. Vaisburg, August 31, 2006 5
O
NHOH

Topology of HDAC inhibitors based on the structure of
the active site of the HDAC enzyme(s)
The cap
Spacer
Pharmacophore: to chelate the Zn 2+ ion at the
bottom of the enzyme active site
Spacer: to fill-out the 11A narrow pocket
The cap: to make contacts with the rim at the
entrance to the 11A pocket
Pharmacophore
(Zn-binding
moiety)
Example:
Trichostatin
(TSA)
A. Vaisburg, August 31, 2006 6
Me
N
Me
O
Me
Me
O
NHOH

The field of small molecule HDAC Inhibitors in 1999
Me
N
Me
O
Me
Me
Trichostatin
(TSA)
O
NHOH
H
N
O
O
NHOH
SAHA
Sloan-Kettering Cancer Institute
Aton Pharma
O
S
O
N
H
Oxamflatin
Shionogi
Research
1. Establish strong IP position
2. Develop chemistry
3. Identify lead molecules, define SAR and optimize leads against target
enzyme(s), in cellular assays, in terms of physico-chemical, ADME and
safety properties
4. Study in vivo efficacy in human xenograft models in mice
5. Select a development candidate within 3-4 years
O
NHOH
A. Vaisburg, August 31, 2006 7

Me
N
Me
MethylGene Small Molecule HDAC Inhibitor
Program: Hydroxamates
O
Me
Me
Trichostatin
(TSA)
R
O
O
NHOH
H
N
O
O
NHOH
SAHA
Sloan-Kettering Cancer Institute
Aton Pharma
O
NHOH
A B
R
O
S
O
N
H
O
S
O
N
H
O
Oxamflatin
Shionogi
Research
NHOH
A. Vaisburg, August 31, 2006 8
O
NHOH

Biological Profiling Paradigm of MG HDAC Inhibitors
Inhibitor synthesis
H4-Ac
In vivo tumor
xenograft models
Cancer cell proliferation
Cell cycle
analysis
p21
induction
PK evaluation
Transcriptional profiling
Enzyme
screening
Apoptosis
A. Vaisburg, August 31, 2006 9

Hydroxamates: Hydroxamates:
Specific Examples
Compd Structure HD1
IC50
1
2
3
4
Br
MeO
N
OMe
N
O
S
O
O
S
O
N
H
N
H
O
O
O
O
NHOH
O
NHOH
O
NHOH
NHOH
A. Vaisburg, August 31, 2006 10
(µM)
0.1
0.4
0.001
0.006
MTT
HCT116
IC50 (µM)
0.8
0.2
1.0
4.0
MTT
HMEC
IC50 (µM)
16
1.0
7.0
>20
H4-Ac
EC50
(µM)

SAR of TSA/SAHA-Type TSA/SAHA Type Long-Chain Long Chain Hydroxamates
oxime or alkene
tolerated
hydrophobic substituents
increase activity
O
α− or β−substitutions
decrease activity
O
H
N OH
6-carbon chain length
optimal
hydroxamate
required
A. Vaisburg, August 31, 2006 11

4-Acetylamino
Acetylamino-N-(2' (2'-aminophenyl)
aminophenyl)-benzamide benzamide
(CI-994) (CI 994) – an anti-Proliferative
anti Proliferative Compound with
Ability to Differentiate Leukemic Cells (1993)
(unknown mechanism of action)
H 3C
O
H
N
O
H
N
CI-994
Parke-Davis/Pfizer
NH 2
CI-994 CI 994 was prepared by MethylGene and demonstrated to be weak
HDAC inhibitor
A. Vaisburg, August 31, 2006 12

Me
N
Me
MethylGene Small Molecule HDAC Inhibitor
Program: Aminoanilides
O
Me
Me
Trichostatin
(TSA)
R
O
O
NHOH
Me
H
N
O
O
H
N
O
H
N
O
NH2
NHOH
SAHA
Sloan-Kettering Cancer Institute
Aton Pharma
O
N
H
NH2
C D
R
CI-994
(Parke-davis)
O
S
O
N
H
O
S
O
N
H
O
Oxamflatin
Shionogi
Research
A. Vaisburg, August 31, 2006 13
N
H
NH 2
O
NHOH

Aminoanilides: Aminoanilides:
Specific examples
Compd Structure HD1
IC50
5
6
7
N
MeO
OMe
O
O
S
O
O
S
O
N
H
N
H
O
O
O
N
H
H
N
N
H
NH 2
NH 2
(µM)
5.0
0.8
MTT
HCT116
IC50 (µM)
19
1.0
MTT
HMEC
IC50 (µM)
ND
>20
H4-Ac
EC50
(µM)
A. Vaisburg, August 31, 2006 14
NH 2
1.0
15
ND
ND
10
15

R
Discovery of N-(2 (2-Aminophenyl)
Aminophenyl)-4-
((arylamino)methyl)-benzamides
((arylamino)methyl) benzamides - the Most
Advanced Class of MG HDAC Inhibitors
O
S
O
N
H
Replacement of
SO 2 NH by NH
D
O
N
H
NH 2
R
H
N
R
E
O
N
H
H
N
NH 2
From
cinnamides to
benzamides
A. Vaisburg, August 31, 2006 15
F
O
N
H
NH 2

Aminoanilides: Aminoanilides:
Specific examples (cont’d) (cont d)
Compd Structure HD1
IC50
(µM)
8
9
10
MeO
MeO
MeO
MeO
N
OMe
OMe
O
N
H
H
N NH2 N
H
N
H
N
N
O
O
N
H
N
H
NH 2
NH 2
2.0
0.3
MTT
HCT116
IC50 (µM)
2.0
2.0
MTT
HMEC
IC50 (µM)
>20
ND
H4-Ac
EC50
(µM)
A. Vaisburg, August 31, 2006 16
3.0
7.0
>20
1.0
5.0
5.0

Aminoanilides: Aminoanilides:
Specific examples (cont’d) (cont d)
Compd Structure HD1
IC50
11
12
13
14
O
N
O
MeO
MeO
O
N
H
N
N N
N
H
NH
N
N
N
N
H
N
H
N
S
NH
O
O
H
N
O
H N
H
N
NH 2
O
HN
H 2N
NH2
NH 2
(µM)
0.2
0.9
0.1
MTT
HCT116
IC50 (µM)
0.4
1.0
0.1
MTT
HMEC
IC50 (µM)
15
18
15
H4-Ac
EC50
(µM)
A. Vaisburg, August 31, 2006 17
0.2
0.6
>20
1.0
1.0
1.0
1.0

SAR of N-(2 (2-Aminophenyl)
Aminophenyl)-4-
((arylamino)methyl)-benzamides
((arylamino)methyl) benzamides
Aryl
Heteroaryl
(Optionally substituted)
H
N
X
CH2 - essential
X
Substitution
detrimental
X=CH,N;
O
N
H
Substitution
detrimental
Substitution
tolerated
NH 2 (OH)
Substitution
detrimental
A. Vaisburg, August 31, 2006 18

N-(2 (2-Amino Amino-phenyl) phenyl)-4-[(4 [(4-pyridin pyridin-3-yl yl-pyrimidin pyrimidin-2-ylamino) ylamino)-
methyl]-benzamide methyl] benzamide MGCD0103
(di di-hydrobromide
hydrobromide salt)
- MethylGene HDAC Inhibitor in Clinical Development
N
N
N
N
H
x2HBr
MGCD0103
O
H
N
Chemical Formula: C 23H 20N 6O
Molecular Weight: 396.44
Log P: 2.6
PSA: 104.7 A 2
M.p. 191.5-194.5 o C
A. Vaisburg, August 31, 2006 19
NH 2

H 2 N
HCl
N
I
N
Synthesis of MGCD0103
(free base)
O
N
OMe
N
H
NH2
NH2
BOP
NEt 3/DMF
66%
N
N
HCl
O
Me
N
HN NH2
DMF/DIPEA
77%
H2N
NH
N
H
OMe
N
Me
III
Iso-Propyl
II
O
alcohol/MS
reflux
52%
O
OMe
N
LiOH/THF
MeOH/H 2O
40 o C
N
N
95%
IV V
N
H
MGCD0103
free base
A. Vaisburg, August 31, 2006 20
O
N
H N
NH2
N
N
N
H
O
OH

In vitro profile of MGCD0103
N
Potency against HDAC
enzymes IC50 (µM)
N
N
N
H
O
H
N
NH 2
Inhibition of cell proliferation
(MTT assay, IC50 µM)
HD1 HD2 HD3 HD11 HCT116 A549 DU145 HMEC
0.1 0.2 ~ 2.0 ~ 2.0 0.2 1.0 0.8 >20
MGCD0103 unlike hydroxamates
does not inhibit HD4, HD5, HD6, HD7, HD8, HD9 and HD10
A. Vaisburg, August 31, 2006 21

SAR of MGCD0103
MG No Structure
MGCD0103
free base
15
16
17
18
N
N
N
N
N N H
N N H
N
N
N
N
H
O
N N H
N
N
N
N
N
N
H
N
H
O
O
O
H
N
N
H
H
N
NH 2
O NH 2
N
H
NH 2
NH 2
HD1
IC50 (µM)
MTT
HCT116
IC50 (µM)
0.1 0.2
>25 >50
>20 25
>20 43
2 2
A. Vaisburg, August 31, 2006 22

Induction of Histone Acetylation in DU145 Tumors in
Mice Treated with MGCD0103 in vivo by Oral
Administration
vehicle
Me
40mg/kg BID
O
H
N
CI-994 MGCD0103 (free base)
80mg/kg QD
O
H
N
NH 2
40mg/kg BID
80mg/kg QD
H3Ac
H4Ac
A. Vaisburg, August 31, 2006 23

Acetylated Histone H3 Levels in HCT116 Tumors
Densitometric Area (A.U.)
1.4
1.2
1.0
0.8
0.6
0.4
0.2
0
Cont 2h
MS-275 = 30 mg/kg
MGCD0103L = 90 mg/kg
MGCD0103H = 120 mg/kg
Cont 8h
Cont 24h
Histone H3 acetylation
MS275 2h
MS275 8h
MS275 24h
A. Vaisburg, August 31, 2006 24
MGCD0103 H 2h
MGCD0103 H 8h
MGCD0103 H 24h
MGCD0103 L 2h
MGCD0103 L 8h
MGCD0103 L 24h

Densitometric Area (A.U.)
Induction of p21 in HCT116 Tumors
1.6
1.4
1.2
1
0.8
0.6
0.4
0.2
0
Vehicle
Vehicle
MS275
MS-275
30 mg/kg
*
MG5837 L
MGCD0103
90 mg/kg
*
MG5837 H
MGCD0103
120 mg/kg
A. Vaisburg, August 31, 2006 MG 25

PK summary of MGCD0103
Species T ½ (hours) F (%)
Mouse 0.6 12
Rat 0.7 42
Dog 1.3 4 (10 mg/kg)
92 (30 mg/kg)
A. Vaisburg, August 31, 2006 26

Tumor Volume (mm 3 )
Inhibition of of A549 Human Non Small NonCell Small Human Cell Lung Lung Cancer Cancer by by
MGCD0103, Administration MS-275, of SAHA MGCD0103, and LAQ-824 MS-275, SAHA in Mouse and LAQ-284 Xenografts
1400
1200
1000
800
600
400
200
0
0 2 4 6 8 10 12 14 16
A. Vaisburg, August 31, 2006 Days
27
Saline
MS-275 20mg/kg P.O.
MGCD0103 60mg/kg P.O.
SAHA 30mg/kg I.P.
LAQ-284 30mg/kg I.P.

Inhibition of SW48 Human Colon Cancer by
administration of MGCD0103 in Mouse Xenografts
Tumor volume (mm 3 )
700
600
500
400
300
200
100
0
MGCD0103
0 2 4 6 8 10 12 14 16
Day
Control
MGCD0103 60mg/kg p.o.
MGCD0103 90mg/kg p.o.
MGCD0103 120mg/kg p.o.
A. Vaisburg, August 31, 2006 28

Chemical
Class
MGCD0103*
(MethylGene)
SAHA
(Merck)
FK-228
Depsipeptide
(Gloucester)
Aminonilide Hydroxamate Macrocyclic
Compound
MS-275
(Schering AG)
Stage/Phase Phase I / II PH IIb (NDA) Phase II (Pivotal) Phase II
Indication AML, MDS,
Solid tumors
CTCL, CRC,
Solid tumors
RCC, prostate
thyroid, CTCL,
PTCL
PXD101
(CuraGen/
TopoTarget)
Aminoanilide Hydroxamate
Solid tumors/
lymphoma
Route Oral Oral IV Oral
Dose 45-70 mg/m 2
3x / wk
400mg/day
300mg/bid
14 mg/m 2 days
1, 8, 15
8 mg/m 2 /wk
Half Life 9 hr 2.5 hrs 4 hrs 39-80 hrs
HDAC
inhibition
Reverses by
48-72 hr
Side effects Fatigue
G.I.
HDAC Clinical Compounds
Reverses by
12 hrs
Fatigue
G.I.
Thrombocyte
Pul. Embol
Cardiac.
Reverses by
24 hrs
Fatigue
G.I.
Thombocyte
Neutropenia
Cardiac
Hypocalcemia
Reverses by
24 hrs
Fatigue
G.I.
Thrombocyte
Neutropenia
Hypophos.
Phase II
MM, TCL, CRC,
Ovarian/
Meso/AML
IV / Oral
1000mg/m 2
30min by IV for
5 days/3 weeks
1-2 hrs
Reverses by
6-24 hrs
Fatigue
G.I.
Phlebitis
Cardiac
*Reported as of Nov 05 at EORTC, ASH in December 05 and ASCO in June 06
G.I. includes nausea, vomiting, diarrhea, anorexia
A. Vaisburg, August 31, 2006 29

• Signs of single-agent efficacy
Efficacy
– Refractory AML: 3 of 9 patients treated at 60-80 mg/m 2 3x/week have
demonstrated normalization of bone marrow blasts.
– Thymic carcinoma: tumor shrinkage and prolonged stable disease (7 cycles)
– Non-Small Cell Lung Cancer: shrinkage of central lung lesion
– Colon Cancer: prolonged stable disease in patient (4 cycles)
– Renal Cell Carcinoma: 3 patients with prolonged stable disease (>4 cycles)
• Support for combinations
– Most effective cancer treatments involve combinations of agents that target
different cancer processes
– MGCD0103 mechanism of action and preclinical studies suggest it will be
promising in combination with other agents
A. Vaisburg, August 31, 2006 30

Florescence units
30.0
25.0
20.0
15.0
10.0
5.0
0.0
MGCD0103 in Leukemia
Whole-Cell HDAC Activity
Baseline Week 1 Cycle 2
RFU
Histone Acetylation
25000
20000
15000
10000
5000
Bone Marrow Blast Counts
% BM Blasts
Uninhibitable
Pool
25
20
15
10
5
0
Baseline Cycle 2
0
Baseline Week 1
A. Vaisburg, August 31, 2006 31

Summary of Clinical Development of MGCD0103
• Safe and manageable dosing schedule
• Dose-dependent pharmacokinetics with exposures
comparable to xenograft activity.
• Exposure-dependent pharmacodynamics indicate
MGCD0103 is inhibiting its target long after serum
levels have dissipated
• Evidence of anticancer activity in a variety of solid
and hematological tumors, especially AML.
• Future studies in solid tumors will combine with
other anticancer agents
• Future monotherapy Phase II trials in hematologic
cancers
A. Vaisburg, August 31, 2006 32

Beyond MGCD0103: Towards the
Second Generation
MG HDAC Inhibitors
A. Vaisburg, August 31, 2006 33

Structural analogues of MGCD0103
Replacement of pyridine ring
by substituted benzene, or 5- and
6-member heteroaromatics
N
N
N
N
H
MGCD0103
Replacement of pyrimidine ring
by benzene or 6-member
heteroaromatics
A. Vaisburg, August 31, 2006 34
O
H
N
NH 2

Structural Analogues of MGCD0103: Specific examples
Compd Structure HD1
IC50 (µM)
19
20
21
22
23
O
N
S
O
N
HOH 2C
N
N
N
O
N
N
N
N
N
H
N
N N
H
N
N
H
N
H
N
N
N
H
O
O
O
H
N
O
H
N
H
N
H
N
NH 2
NH 2
NH 2
NH 2
O
H
N
NH 2
MTT
HCT116
IC50 (µM)
MTT
HMEC
IC50 (µM)
H4-Ac
EC50 (µM)
P21
EC50 (µM)
0.1 0.1 14 50 1 1
0.2 0.4 50 1 0.6
0.3
0.1
0.4 11

Structurally different HDAC Inhibitors:
Specific Examples
Compd Structure HD1
IC50
24
25
26
27
MeO
MeO
F3CO
MeO
O
O
O
N
H
N
HN
N
N
N
S
O
O
N
HN
O
O
HN
H 2N
O
H N
HN
O
H 2N
H 2N
NH 2
(µM)
0.7
2.0
0.7
MTT
HCT116
IC50 (µM)
1.0
1.0
1.0
MTT
HMEC
IC50 (µM)
>20
ND
>20
H4-Ac
EC50 (µM)
A. Vaisburg, August 31, 2006 36
0.1
8.0
>20
2.0
>5
2.0
ND

Towards Greater HDAC Potency:
Anilides versus Aryl(heteroaryl)
Aryl(heteroaryl)
Anilides
Compd Structure HD1
IC50
11
28
29
30
MeO
MeO
MeO
MeO
NH
NH
HN
HN
H 2N
O
H 2N
O
HN
H 2N
MeO NH
O
S
MeO
MeO
MeO
NH
HN
H 2N
O
O
(µM)
0.2
0.06
0.05
0.05
HD2
IC50
(µM)
0.4
0.04
0.05
0.03
MTT
HCT116
IC50 (µM)
0.4
0.3
0.3
0.3
MTT
HMEC
IC50 (µM)
Aryl(heteroaryl)-substituted anilides are more potent against HD1 and HD2 than
the parent anilide!
A. Vaisburg, August 31, 2006 37
15
12
>50
>50

Features of the second generation compounds
1. Considerable structural differences
2. Different HDAC enzyme selectivity
3. Different transcriptional profile
4. Acceptable physico-chemical physico chemical properties (solubility,
stability, ionization state, etc.)
5. Favorable ADME properties
6. Safety
7. Therapeutic window
8. In vivo efficacy in multiple human xenograft models in
mice
9. The development candidate to complement MGCD0103 –
the second generation MG HDAC inhibitor is expected to
be selected at the end of 2006
A. Vaisburg, August 31, 2006 38

Medicinal Chemistry:
Nancy Zhou,
Sylvie Frechette,
Isabelle Paquin,
Frederic Gaudette,
Naomy Bernstein,
Giliane Bouchain,
Silvana Leit,
Stephane Raeppel,
Oscar Moradei
Elie Abou-Khalil,
Rico Lavoie,
Carl Thibault,
Rejean Ruel,
Soon Hyung Woo,
Daniel Delorme
Acknowledgements
(pre-clinical studies)
Molecular Biology:
Marielle Fournel,
Claire Bonfils,
Ann Kalita,
Pu T. Yan,
M-C. Trachy-Bourget,
Jianhong Liu
Yu Hou
Zuomei Li
PK and in vivo studies:
Zhiyun Jin,
Roussen Pascal
Jeff Gillespie,
James Wang
Enzymology:
Carole Beaulieu,
Aihua Lu,
Jubrail Rahil
Cell biology & pharmacology:
Cindy Lalancette,
Julie-Pare
Marie-France Robert,
Sylvain Lefevbre,
A. Robert MacLeod
Jeffrey M. Besterman
A. Vaisburg, August 31, 2006 39