compounds related to 3-alkylamine-1h-indolyl acrylate and

XIII Encuentro de Cooperación Farma-Biotech

Barcelona, october 20th 2015

Compounds related to 3-alkylamine-1H-indolyl acrylate and their use for the treatment of neurodegenerative diseases

Rafael León Ph.D.

Content

1.   The Institution

2.   The Product a)   Target Indications b)   Innovative mechanisms of action c)   Differential features facing the market d)   Current status of development e)   IPR protection f)   Pitfalls & Risks to be considered

3.   Partnering Opportunities


INSTITUTION: IIS HOSPITAL UNIVERSITARIO LA PRINCESA

ITH 8: Rafael León Ph.D (Medicinal Chemistry and biological screening)

Dr. Leon MedChem group: Current members

IIS H. La Princesa(Fundación I. B.)

Area 1: Francisco Sánchez-Madrid Area 2: Antonio García García Area 3: Isidoro González Álvaro

•  Giammarco Tenti: Postdoctoral Student

•  Zulay Pardo: Postdoctoral Student (Juan de la Cierva fellowship)

•  Patrycja Michalska: Ph.D. Student (FPU fellowship)

•  Isabel Gameiro: Ph.D. Student (FPI fellowship)

•  Sheila Abril: Ph.D. Student (FPU fellowship)

•  Michelle Satriani: Erasmus Student

•  Guillermo Furones: Pharmacology Master Student

•  Alberto Ruiz Priego: Pharmacology Master Student

Neuropharmacology and neuroprotection

Hippocampal Neurotransmision

Clinical pharmacology and pharmacogenomics Affective disorders Epilepsy

neurosurgery Cardiovascular

diseases

Dr. Leon MedChem group: Financial support

Content





Neurodegenerative diseases: Cognitive degeneration and chronic neuroinflammation

Neurodegenerative and inflammatory diseases

•  Common physiopathological hallmarks: Directed drug design

q  Aberrant protein aggregates q  Mitochondrial dysfunction

q  Extensive oxidative stress

q  Neuroinflammation q  Autophagy failure

q  Detoxification pathways failure

q  Synaptic deregulation

Target indications

Neurodegenerative and inflammatory diseases

•  Neurodegenerative diseases (600 pathologies described as NDDs)

q  Alzheimer disease (claim 11) q  Parkinson’s disease (claim 12)

q  Huntington’s disease (claim 13)

q  Multiple sclerosis (claim 14) q  Cerebral ictus (claim 15)

q  Peripheral neuropathic diseases (claim 16)

q  Amyotrophic lateral sclerosis (claim 17)

Potential target indications •  Wide range of application not included in the first patent due to its mechanism of action

q  Cancer q  Chronic obstructive pulmonary disease (seeking financial support, ITH-Evgen pharma, UK)

q  Retinal neurodegenerative diseases (In vitro and in vivo studies in progress, UA-ITH-Bayer Healthcare)

q  Chronic kidney disease (In vitro and in vivo studies in progress, FJD-ITH) q  Crohn disease

Content





Nrf2-ARE pathway… combined with:

Nrf2-ARE pathway as innovative mechanism of action

Oxidative stress regulation and neuroinflammation

Nrf2

HS

HS

HS

SH

SH

SH

Ub

Proteasome Degradation

EpRE

Maf

Gene transcription

A) Normal conditions

Phase II Antioxidant genes- Direct antioxidant enzymes: HO-1- Glutation homeostasis: GCLc, GCLm- Detoxifiying enzymes: GST, NQO1- Thiol homeostasis: Txn, TrxR1, Srx1- Inhibition of inflammation: HO-1

Keap1dimer

Proteasome 26S

Nrf2Ub

Cul3

BTB BTB

Nrf2

HS

HS

HS

SH

SH

SH

Ub

CuI3

BTB BTB

B) Oxidative conditions

Nrf2

S

HS

HS

SH

SH

BTB BTB

Nrf2

HS

HS

HS

SH

SH

SH

BTB BTB

C) Receptor modulation

STRESORSElectrophilesROSHeavy metals

Direct Nrf2-Keap1 inhibitors

R

Hinge and latch mechanism

D) Chronic inflammation

AKT

P38

GSK3β

HDAC

Fyn Nrf2P

CuI3

nAChR-α7 MT 1/2 recptors

PI3K

Jak2

GSK3β

Nrf2

Nrf2

Nrf2

ERK

Prolonged inflammation

León et al, Pharm. & Therap. 2015, in press.

•  High CNS vulnerability •  Crosslinks between

oxidative stress and protein aggregates

•  Mitochondrial dysfunction

Neuronal loss

Sultana, R.; Butterfield, D. A. J Alzheimers Dis 2010, 19, 341-53.

Glass et al, Cell 2010, 140, 918-34.

•  Oxidative stress • Neuroinflammation

•  Glial overactivation (ROS and protein agregates)

•  Pro-inflammatory cytokines and infiltration

Nrf2-ARE pathway target of BG12 (Tecfidera®)



Glass et al, Cell 2010, 140, 918-34.

2nd generation of Nrf2 inducers are generating great interest



Glass et al, Cell 2010, 140, 918-34.

q  XenoPort: Biopharmaceutical company is developing a “ME TOO” derivative of Tecfidera

Content





3-alkyl-1H-indolyl acrylate derivatives Vs BG12 (Tecfidera®)

Advantages and differences of NCE facing the market

•  Bardoxolone RTA-402

•  Clinical Trial phase III BEACON NCT00664027

•  Result: TERMINATED (Safety concerns October 2012)

q  Nrf2 induction potency control: 2nd Strong Nrf2 induction is TOXIC¡¡¡

q  Nrf2 induction is not enough: 1st Combination of mechanisms (Nrf2 induction – free radical scavenger – anti-inflammatory)

Advantages and differences of NCE facing the market

q  3rd Improved activity without increasing toxicity might led to: q  Lower incidence/ less sever side effects

q  Improved compliance, fewer treatment failures

q  Onset and/or magnitude of immunomodulation q  Earlier onset of immunomodulation


Glass et al, Cell 2010, 140, 918-34.




q  4th Improvement in efficacy due to combination of action mechanism: q  Dosing frequency

q  Once-a-day rather than BID (TECFIDERA)

q  5th Novel chemical entity with many substitution possibilities: q  Pharmacokinetic and pharmacodynamics properties modulation

Content





Development status 3-alkylamine-1H-indolyl acrylate derivatives

In vitro characterization: Nrf2 induction

R = crotonic

Basal 0,3 3 10 300

10

20

30

40

1 (µM)

***

**

***

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.) R = p-Cl

Basal 0,3 3 10 300

5

10

15

20

10 (µM)

***

***

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.) R = m-OMe, p-OH

Basal 0,3 3 10 300

1

2

3

4

7 (µM)

***

***

*

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.)

Basal 0.3 3 10 300.0

0.5

1.0

1.5

Melatonin (µM)

***

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.)

Basal 0.3 3 10 300.0

0.5

1.0

1.5

Ethyl cinnamate (µM)

*****

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.)

Basal 0.3 3 10 300.0

0.5

1.0

1.5

2.0

Et. Cin. + Mel (µM)

***

Activ

idad

Luc

ifera

sa(ta

nto

por 1

)

R = crotonic

Basal 0,3 3 10 300

10

20

30

40

1 (µM)

***

**

***

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.) R = p-Cl

Basal 0,3 3 10 300

5

10

15

20

10 (µM)

***

***

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.) R = m-OMe, p-OH

Basal 0,3 3 10 300

1

2

3

4

7 (µM)

***

***

*

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.)

Basal 0.3 3 10 300.0

0.5

1.0

1.5

Melatonin (µM)

***

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.)

Basal 0.3 3 10 300.0

0.5

1.0

1.5

Ethyl cinnamate (µM)

*****

Luci

fera

se A

ctiv

ity(F

old

incr

ease

a.u

.)

Basal 0.3 3 10 300.0

0.5

1.0

1.5

2.0

Et. Cin. + Mel (µM)

***

Activ

idad

Luc

ifera

sa(ta

nto

por 1

)

Nrf2-induction properties are easily tuneable


In vitro characterization: Antioxidant effect

Mel

Et. Cina

Mix 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 160

2

4

6

8

10

Antio

xida

nt a

ctiv

ity(T

rolo

x eq

.)

In vitro characterization: Anti-inflammatory effect (LPS response blockade)

Basal LPS 30 60 10075

100

125

150

175

Mn 3/24 ( µM) Pm 15 (µM) Pm 19 (µM)

###

****** ***

1 (µM)

R = Crotonico

LPS (1 µg/mL)

% N

itrite

pro

duct

ion

(% V

s Ba

sal)

Basal LPS 30 60 10075

100

125

150

175

Mn 3/24 (µM)

###

******

12 (µM)

R = m,p-di-Cl

LPS (1 µg/mL)

***

% N

itrite

pro

duct

ion

(% V

s Ba

sal)

Basal LPS 30 60 10075

100

125

150

175

Mn 3/24 ( µM) Pm 15 (µM) Pm 19 (µM)

###

*** ******

7 (µM)

R = m-OMe; p-OH

LPS (1 µg/mL)%

Nitr

ite p

rodu

ctio

n(%

Vs

Basa

l)

Basal 3 10 30

100

150

200

Mel (µM)

****

###

LPS (1 µg/ml)

% N

itrite

pro

duct

ion

(% V

s Ba

sal)

Basal 3 10 30

100

150

200

LPS (1 µg/ml)

Etil Cin (µM)

*###

% N

itrite

pro

duct

ion

(% V

s Ba

sal)

Basal 10 30 100

100

150

200

MIX (µM)

###*

***

**

LPS (1 µg/ml)

% N

itrite

pro

duct

ion

(% V

s Ba

sal)

Basal LPS 3 10 30

100

120

140

160

Mn 3/24 ( µM) Pm 15 ( µM) Pm 19 ( µM)

LPS (1 ng/mL)

###

7 (µM)

***

***

***

R = m-OMe; p-OH

% N

itrite

pro

duct

ion

(% V

s Ba

sal)




•  Result: TERMINATED (Safety concerns october 2012)


Glass et al, Cell 2010, 140, 918-34.




In vitro characterization: Neuroprotective effect against oxidative stress

Control 0.3 1 3 350

75

100

OGD (15 Min) + 2 h reox

###

***

*** ****

(µM)

Compound 7Melatonin

% C

ell V

iabi

lity

(MTT

Red

uctio

n)

q  Cerebral ictus (hippocampal slices) q  Tau hyperphosphorylation

Neurodegenerative diseases specific models

Control 0.3 1 3 3

50

75

100

Okadaic acid (30 nM)

*** ******

###

(µM)

Compound 7Melatonin

% C

ell V

iabi

lity

(MTT

Red

uctio

n)

BASAL Mel 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 160

20

40

60

80

100

R/O (30/10 µM) + Compound (1 µM)Pre-incubation (1 µM)

R/O 30/10 µM

###

******

*** *** *** *** *** *** *** *** ******

******

******

***

% C

ell V

iabi

lity

(MTT

Red

uctio

n)

BASAL Mel 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 160

20

40

60

80

100

Menadione (8 µM) + Compound (1 µM)Pre-incubation (1 µM)

Menadione 8 µM

###

***

***

*** *** *** ***

***

***

***

***

***

***

***

*** ****** ***

% C

ell V

iabi

lity

(MTT

Red

uctio

n)



Glass et al, Cell 2010, 140, 918-34.

q  Amyloid β and Okadaic acid combination

Basal 1 10 300

20

40

60

80

100

Mn 3/24 (µM) Pm 15 (µM) Pm 19 (µM)

Aβ (0.5 µM) /Okadaic acid (10 nM)

###

Melatonin

% C

ell V

iabi

lity

(MTT

Red

uctio

n)

RO

S Ib

a-1

Basal 1 10 300

20

40

60

80

100

***

###

**

***

Mn 3/24 (µM)

Aβ (0.5 µM) /Okadaic acid (10 nM)Compound 1 (µM)

% C

ell V

iabi

lity

(MTT

Red

uctio

n)

Neuroinflammation inhibition (Primary rat glial cultures)

q  Nitrite release reduction

Basal 3 10 30

100

120

140

160

Mn 3/24 (µM) Pm 15 (µM) Pm 19 (µM)

LPS (1 µg/mL)

###

Compound 7

***

***

***

% N

itrite

pro

duct

ion

(% V

s Ba

sal)

q  TNFα release inhibition

q  Lipopolysaccharide model of inflammation

Basal LPS 3 10 300

100

200

300

400

Mn 3/24 (µM) Pm 15 (µM)

LPS (1 µg/mL)

###

7 (µM)

***

% T

NFα

pro

duct

ion

(pic

ogra

ms/

ml)

***



Glass et al, Cell 2010, 140, 918-34.

Compound 7 Vs BG-12

Wilms, H.; J. Neuroinflammation 2010, 7, 30.

q  Nrf2 induction

LPS 3 10 300

20

40

60

80

100

Mn 3/24 (µM)

LPS (1 µg/mL)

###

Compound 7 (µM)

***

***

***

**

% T

NFα

pro

duct

ion

(pic

ogra

ms/

ml)

Treatment with 10 ngr/mL LPS and 10 µM BG-12

q  TNFα release inhibition q  TNFα release inhibition

q  Nitrite release reduction q  Nitrite release reduction

Compound 7 BG-12

LPS 3 10 300

20

40

60

80

100

Mn 3/24 (µM)

LPS (1 µg/mL)

###

Compound 7 (µM)

***

***

***

% N

itrite

pro

duct

ion

(Nor

mal

ized

to L

PS)



Glass et al, Cell 2010, 140, 918-34.

Differentiating aspects facing commercialization:

100

120

140Nrf2 +/+Nrf2 -/-

LPS (1 µg/ml)7 (10 µM)

ns

ns

###

###

******

Basal% N

itrite

pro

duct

ion

(% V

s B

asal

)

q  Nitrite release reduction

Anti-inflammatory effect in Nrf2 KO mouse (Primary mouse glial cultures)

q  iNOS expression inhibition

0

1000

2000

3000

4000Nrf2 +/+Nrf2 -/-


ns

ns

###

###

******

Basal

iNO

S/β

-act

in

q  HO-1 expression

0

500

1000

1500 Nrf2 +/+Nrf2 -/-


###

***

Basal

HO

-1/β

-act

in

THE ANTI-INFLAMATORY EFFECT IS NOT Nrf2-DEPENDENT

Content





IPR status

3-alkylamine-1H-indolyl acrylate derivatives

•  Patent status: Derivatives of 3-alkylamine-1H-indolyl acrylate and their use for the treatment of neurodegenerative diseases

q  19 claims

q  Chemical core structure protected

q  Chemical substructure protected

q  Biological activity protected

q  Chemical substituents included

q  Anti-inflammatory effect protected

q  Use as drugs for Neurodegenerative diseases included as claim

q  Common formulations, salts and excipients included for core structure

q  Use in COPD, kidney disease or macular degeneration suitable for new patent applications

•  Priority date: 15/10/2014 •  Application number: P201400810 •  Priority country: Spain •  PCT application: 15/10/2015 •  Priority country: Europe

•  Holder: IS Hospital La Princesa / •  Universidad Autonoma de Madrid •  DNS Neuroscience

•  Authors: Rafael León, I. Buendia, E. Navarro, P. Michalska, I. Gameiro, A. López, J. Egea, A. García, M. García.

Content





Pitfalls & risk to be considered



•  Result: TERMINATED (Safety concerns october 2012)


Glass et al, Cell 2010, 140, 918-34.




q  Target validation already demonstrated

q  Rational design of proposed drug

q  In vitro activity demonstrated

q  Preliminary toxicity performed: Non-hepatotoxic

Content





Pre-clinical evaluation

Partnering Opportunities

•  MS mouse model (MOG injection): Scheduled to be started in January 2016 (Miguel P. Soares Ph.D, European expert in inflammation)

•  Pre-clinical toxicity: Big Pharma or CRO companies (Financial support applications: European commission projects)

•  ADMET properties •  Safety pharmacology –CVS, CNS, RS •  Pharmacodynamics •  Pharmacokinetics •  Acute toxicity: 2 species by 2 routes of administration •  Repeat dose toxicity: rodent and non-rodent; two 14 day studies before human trial •  Carcinogenicity •  Reproductive toxicity: Embryo/foetal development studies – 2 species •  Genotoxicity •  Mutagenesis: Chromosomal abnormality

Pre-clinical evaluation financial support: Public-private partnership •  Innovative Medicines Initiative (IMI): Next call 12 January 2016 (Budget 93M €)

•  3er Health Program: 3 different calls to be open, including preclinical studies projects

•  ERA-NET actions under H2020 program: Public-Private partnerships and cofounded actions

•  Joint Technology Initiatives: Specific program in H2020


Barcelona, 20 de octubre de 2015

Compounds related to 3-alkylamine-1H-indolyl acrylate and their use for the treatment of neurodegenerative diseases

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compounds related to 3-alkylamine-1h-indolyl acrylate and

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