Alzheimer’s Disease Drug Discovery:

NMEs vs. NCEs, Pros and Cons of Each

Allen B. Reitz, Ph.D., CEO: areitz@fc-cdci.com

3805 Old Easton Rd., Doylestown, PA 18902

3700 Horizon Dr., King of Prussia, PA 19406

14th Annual ADDF Drug Discovery for Neurodegeneration

Section IV. I Wish I Knew…

20 April 2020

www.alsbiopharma.com, www.fc-cdci.com

FDA-Approved Drugs to Treat AD

Cummings, J.; Lee, G. Ritter, A.; Zhong, K. Alzheimer’s disease drug development pipeline:

2018. Alzheimer’s & Dementia: Trans. Res. & Clin. Interventions 2018, 4, 195-214.

alzheimer’s association

35 Phase III trials for AD ongoing as of 2018

O

N

Me

HO

H OMe

Me

Me

NH2

Me2NO

O

N

Me

Et

Me

N Bn

O

MeO

MeO

memantine, 2003memantine - donepezil 2014

donepezil, 1996

rivastigmine, 1997

galantamine, 2001

Acetylcholinesterase Inhibitors

NMDA Antagonist

FCCDC and the Drug Discovery Process

FCCDC

• Early-stage value-added probe and drug discovery

• Focus on unmet medical need, orphan indications, IP

creation, and innovative pharmacology

Target

ValidationHit to Lead

Lead

OptimizationPhase IIIPhase IIPhase I

Pre-Clinical

Development

Start Hits Lead PCD FIH Drug

FIH = day on which the first clinical

dose is administered to any patient

New Chemical Entities vs. New Molecular Entities

Food, Drug and Cosmetic Act 21 CFR 314.108

New Molecular Entity (NME)

A drug that contains an active moiety that has never been approved by the FDA

Can also contain an active moiety that has been approved by the FDA

Three years of exclusivity post approval

May be able to use the clinical data of others to support the NDA without a license

New Chemical Entity (NCE)

A drug that contains no active moiety that has been approved by the FDA

Five years of exclusivity post approval

Active moiety

“The molecule or ion, excluding those appended portions of the molecule that

cause the drug to be an ester, salt . . . , or other noncovalent derivative . . ..”

Esters as Prodrugs: NMEs vs. NCEs

NO N

N

N

Me

O

ORF

O

N

Me

RO

H OMe O

Me

HO

H

Me

F

F

H

Me

OR

OS

F

NCEs, R = H

NMEs, R = C(O)R'

Galantamine Paliperidone Fluticasone

Memogain(GLM-1062, R' = Ph)

Paliperidone palmitate

(R' = (CH2)14Me)

Fluticasone furoate(R' = 2-furanyl)

Fox Chase Chemical Diversity Center, Inc.

Small-molecule drug discovery company (2008)

First SBIR with the Fox Chase Cancer Center (2009)

>11,700 New Chemical Entities (NCEs) prepared

Currently: ~20 chemists, 5 pharmacologists

Two locations in PA: Doylestown and King of Prussia

Two compounds in human clinical trials

Have had >20 Phase NIH/DOD/foundation grants

Have had 10 Phase 2 SBIR or STTR awards

Two commercial exits from Phase 2 SBIRs

Current Collaborators

FCCDC Technical and Administrative Staff

Most have worked in big pharma in the Philadelphia area

About half with Ph.D. degrees

Experience in different therapeutic areas against diverse molecular targets

Inventors on >200 issued U.S. patents

Troriluzole Product Development Timeline

2008 2009 2010 2011 2012 2013 2014 2015 2016 2017

Founding of FCCDCNovember 5, 2008

Rutgers mtg:

riluzole for melanomaNovember 16, 2009

Phase 2 SBIR:

“Riluzole Prodrugs for Melanoma and ALS”

June 19, 2013

Phase I FIH

July 6, 2016

1st Prep of Troriluzole

(FC-4157)March 26, 2015

Today

Founding of ALS Biopharma:

review of riluzole for ALS

Phase 1 SBIR:

“Riluzole Prodrugs for Melanoma and ALS”

November 2, 2011

May 28, 2009Biohaven Pharmaceuticals

Funding and Agreement

August 10 2015

Initial Preparation to First-in-Human (FIH) in 15 months, 11 days!

First prepared Mar. 26, 2015 at the PA Biotech Center

2018 2019

Four clinical trials ongoing:

1. Alzheimer’s Disease

2. Ataxia (SCA)

3. Obsessive Compulsive Disorder

4. Solid Tumors (Melanoma)

Phase 2B SBIR:

“Riluzole Prodrugs for Melanoma and ALS”

August 29, 2018

AD: www.t2protect.org

S

N

F3CO

NH

O N

NH

O

O NH2

Riluzole Therapeutic Indications and MOA

• Therapeutic Indication - FDA approved for Amyotrophic Lateral Sclerosis (ALS)➢ Rilutek® approved in 1995 (Sanofi)

➢ Extends ventilator-dependence, tracheostomy requirement and life by 2-3 months

• Mechanism of action - Upregulates type II excitatory amino acid transporter (EAAT2)

reducing glutamate hyperstimulation and toxicity➢ Reduces synaptic glutamate levels via enhanced uptake in glia cells

• Clinical trials for other indications completed or in progress➢ Reduces synaptic glutamate levels via enhanced uptake in glia cells

N

SNH2

CF3O

Riluzole Drug Related Liabilities

• Compliance:

➢ BID oral dosing in patients that often have difficulty swallowing

➢ Fasting required before dosing

➢ Liver function test abnormalities in 2% of patients

• Exposure/Efficacy

➢ Poor solubility (<0.1 mg/ml, pH 6.0-8.0)

➢ Riluzole metabolized predominantly by CYP1A2:

o Variable enzyme expression and compound levels

o Significant first pass effect in patients with high CYP1A2 levels

Mitigation via Prodrug Design

• Primary metabolic pathway is CYP1A2 oxidation of the exocyclic amine

• First pass effect leading to variable exposure linked to variability in CYP1A2 levels

• Solution: Functionalize exocyclic amine as a cleavable prodrug Increase solubility

➢ Minimize CYP metabolism and the first pass effect

➢ Regenerate riluzole once the prodrug enters the blood stream

➢ Extend riluzole Tmax potentially allowing for once daily dosing

First Generation Riluzole Prodrugs

Cleavage by peptidases, esterases, neutral pH cyclization, or reduction then cyclization

First Generation Prodrug: FC-311

Bioorg. Med. Chem. 2012; 20:5642-5648.

Cleavage in:

Front row:

Simulated gastric fluid ↓

Simulated intestinal fluid ↓

Human serum stability ↑

Mouse serum stability ↑

Human liver microsomes ↓

Back row:

Mouse liver microsomes ↓

Abandoned: Interspecies variability with regard to chemical stability

Second Generation Prodrug: FC-3423

PO Pharmacokinetics in rat (N = 3, 14.6 mg/kg)

FC-3423 converted to riluzole with long half life, high oral bioavailability

1

10

100

1000

0 5 10 15 20 25

Co

nc

en

tra

tio

n(ng/mL)

Time (hr)

FC-3423 ave

Riluzole ave

FC-3423

Mean SD

Cmax

(ng/mL)213 39.0

tmax (hr) 1.80 0.2

t1/2 (hr) 3.8 0.40

AUClast

(hr·ng/mL)2259 305

AUCinf

(hr·ng/mL)2289 32

%F 26

Riluzole (from FC-3423)

Mean SD

Cmax

(ng/mL)1587 150

tmax (hr) 8.00 0

t1/2 (hr) ND ND

AUClast

(hr·ng/mL)20518 1437

AUCinf

(hr·ng/mL)ND ND

%F >100

Dipeptide conjugate of riluzole:

N-t-(Butylgly)sarcosylriluzole

FC-3423 hERG Liability

hERG activity (patch clamp)

Ion ChannelIC50 (mM)

Chantest WuXi

Ikr (hERG) 0.80 0.80

Cav1.2 2.1 -

Nav1.5 4.9 -

Abandoned: Unacceptable levels of hERG activity

Stable in non-plasma fluids and tissues

Stability Half Life (in vitro, min)

SGF SIF

Human

LM

Mouse

LM

Rat

LM

Dog

LM

>> 240 >>240 >60 119 >60 >60

Pharmacadence Absorption Systems

Modest Protein Binding

High Solubility and Permeability

Human

Protein

Binding

(% bound)

Caco-2 Permeability Solubility

pH 7.4

Aqueous

Buffer

(mg/mL)

Absorption

Potential

Classification

Significant

Efflux

88.2 High No 159

Absorption Systems

However…..

hERG is a K channel associated

with cardiac arrhythmias in vivo

Assay R = H R = CH2Ph

Rat serum riluzole

formation t1/2 (mins.) 120 <60

hERG IC50 (mM) >30 0.54

Addition of a third amino acid:

• Reduce hERG activity

• Promote peptidase substrate activity to release active moiety

Third Generation Prodrug: Troriluzole

R = H, Troriluzole, FC-4157

Mechanism of Oral Absorption

Gly-Sar is a recognition element of the peptide 1 transporter PepT1• Digestive tract transporter of di- and tripeptides

• Similar homology/function between different species of mammals

• Other prodrugs absorbed via PepT1 include valacyclovir

• Troriluzole is transported by PepT1 (Km > 1 mM)

• <2% Troriluzole remaining in rat alimentary tract after 30 mins, PO, 5 mg/kg

N

SNH

N

O Me

O

NH

O

NH2

F3CO

Riluzole Sub-Structure Gly-Gly-Sar PepT1 Recognition Element

Troriluzole (FC-4157, BHV-4157)

Troriluzole: Third Generation Prodrug

Troriluzole

• Troriluzole is a human PepT1 substrate

• Avoids the food effect: no fasting required

• Troriluzole undergoes cleavage after absorption,

has longer half-life than parent

• Expected patent protection to March, 2036

• NCE prodrug of riluzole

• Potential for once daily dosing

• Stable in saliva and GI fluids but quickly

converts to riluzole upon absorption

• High (>90%) oral bioavailability of riluzole

Troriluzole in AD Clinical Trials

The Troriluzole Trial (T2) for Alzheimer’s Disease

T2 Protect AD (www.t2protect.org)

BIOHAVEN'S TRORILUZOLE SUCCESSFULLY ADVANCES PAST INTERIM

FUTILITY ANALYSIS IN PIVOTAL PHASE 2/3 ALZHEIMER'S DISEASE STUDY:

12/06/2019 (www.biohaven.com)

Troriluzole, an oral, once-daily tablet that modulates glutamate, is being studied as a

symptomatic treatment for mild-to-moderate Alzheimer's disease:

The independent Data Safety Monitoring Board (DSMB) completed its review of the

prespecified interim futility analysis and communicated that futility was not

met…because troriluzole demonstrate numerically greater benefit over placebo on at

least one of the two pre-specified criteria at 26 weeks: (i) cognitive function as

measured by the Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-

Cog) or (ii) hippocampal volume as assessed by magnetic resonance imaging

Fox Chase Chemical Diversity Center, Inc.Jay Wrobel, Garry Smith, Haiyan Bian, Jeff Pelletier, Allen Reitz (co-inventors);

Rutgers UniversitySuzie Chen, Brian Wall, Raj Shah, Ann Robinson, Annie Silk, Jim Goydos, Chris Molloy

ALS Biopharma, LLCAllen Reitz, Anonymous Angel Investor

Biohaven Pharmaceutical Holding Company, Ltd. (BHVN)Vlad Coric, Rob Berman, Kim Gentile, Irfan Qureshi

CRO’sAbsorption Systems, Pharmacadence, Davos, Anthem Biosciences, Preclinical GPS

IP SupportBen Blass, David Tener, Ralph Loren, Warren Volles

The Troriluzole Team

TDP-43 Sequence and Function

http://www-personal.umich.edu/~sbarmada/research.html

Limbic-predominant age-related TDP-43

encephalopathy (LATE) is a recently recognized form

of Alzheimer’s disease (AD)

TDP-43 Structure and Cleavage Sites

Kumar-Singh et al. J. Mol. Neurosci. 2011, 45, 561-573.

AlphaScreen TDP-43 – ssDNA/RNA

Cassel, J. A.; Blass, B. E.; Reitz, A. B.; Pawlyk, A. C. Development of a Nonradiometric Assay for Nucleic

Acid Binding to TDP-43 Suitable for High-Throughput Screening Using AlphaScreen® Technology.

J. Biomolecular Screening, 2010, 15, 1099-1106.

• GST tagged TDP-43 1-260

• Biotin-tagged oligo

• Final Assay Conditions15 ng/mL GST-TDP43 (56 kD so 0.27 nM)

0.125 nM bTAR32 (-18 to +14 in the HIV1 LTR)

10 ug/mL GST donor bead

10 ug/mL streptavidin acceptor bead

25 mM Tris pH 7.4

0.1% BSA

0.1% Triton

• Compd 50-0.02 mM, 8 ½-log dilutions

• Assayed in duplicate

• FC-8764 and FC-9685 run as controls

Premise: By considering monomeric TDP-43 as target on a molecular level, we could identify small molecules

that could (1) block nucleic acid binding to TDP-43 and pathological over-expression of stress-related proteins

and return to homeostasis, (2) decrease TDP-43 aggregation, nuclear export, caspase cleavage, and/or

phosphorylation, (3) provide TDP-43 based therapy, and (4) provide PET imaging agents.

Therefore: We developed a screen measuring the ability of exogenous agents to bind to monomeric TDP-43

resulting in the inhibition of nucleic acid binding.

Biotin-

Oligo

GSTtagged-

TDP-43

Streptavidin

Donor BeadsAnti-GSTtag

AlphaScreen

Acceptor Beads

Excite

689 nm

Emission

520-620 nm1O2 (single oxygen)

Screen for TDP-43 Binders

Histogram of Inhibition Values

-100-9

0-8

0-7

0-6

0-5

0-4

0-3

0-2

0-1

0 010203040506070809010011

012

0

0

500

1000

1500

2000

2500

% Inhibition @ 10 uM

Co

mp

ou

nd

s

Scatter Plot of Inhibition Values

0 1000 2000 3000 4000 5000 6000 7000

0

25

50

75

100

125

Sample #

% In

hib

itio

n

7,400 cmpd library ➜ 69 initial hits ➜ potency, counterscreening,

structure/purity confirmation ➜ 27 compounds in 4 chemotypes

Number of

analogs

Structure family

360 4- Aminoquinoline

14 2- Aminoisoquinoline

3 4-Amino-8-naphthyridine

2 4-Amine-7-napthyridine

1 1,2,3,4‐tetrahydroquinolin‐4‐amine

1 5,6,7,8‐tetrahydro‐1,8‐naphthyridin‐4‐amine

X X

X

NR1R2

Class A Hit to Lead Medicinal Chemistry

Initial Aminoquinolines

FC-776

Negative

Control

Cassel, J. A.; McDonnell, M. E.; Velvadapu, V.; Andrianov, V.; Reitz, A. B. The effects of small molecule inhibitors of

nucleic acid binding to TDP-43 on TDP-43 metabolism and function. Biochimie, 2012, 94, 1974-1981.

FC-776 Rescues TDP-43 Drosophila

Daniela Zarnescu, University of Arizona

TDP-43 Acknowledgements

FCCDC:

• Biology: Rick Scott, Katie Freeman, Samantha DeSando, Amy Banks, Kevin McClay

• Chemistry: Marianne Carlsen, Randall Binder, Xiao Jin, Mark McDonnell, Garry

Smith, Camille Remeur, Tom Haimowitz, Venkat Velvadapu

• Computational Chemistry: John Kulp, Usha Viswanathan

External Collaborators:

• Johns Hopkins University: Marty Pomper, Phil Wong

• Drosophila, University of Arizona: Daniela Zarnescu

• IP: Ben Blass, Warren Voles

Funding Acknowledgments

National Institutes of Health

R21 NS072749-01, R44 AG059278-01, R43 CA156781-01, R44 CA156781-02

Commonwealth of Pennsylvania (KIZ tax credit funding)

Qualifying Therapeutic Discovery Credit stimulus funding

Anonymous Angel Investor

Alzheimer’s Drug Discovery Fund (ADDF)

Biohaven Pharmaceuticals

I Wish I Knew….

1. Who the true customers are, and what do they want

2. Which projects and approaches would work out and which won’t

3. How to change the project flowchart and criteria in real time ASAP

4. The right questions to ask, also as early as possible

Lessons Learned

1. Risk mitigation is the most important part of drug discovery research

2. Most projects don’t work out in terms of a therapeutic endpoint

3. Most projects provide guidance on other approaches going forward

4. Working with a balanced portfolio of risk-reward is very important