Jonathan C. Cohen

University of Texas SouthwesternMedical Center

A Short History of PCSK9From Discovery to Clinical Trials

Disclosures

PfizerRegeneron

MerckRocheEli Lilly

Speaker/Consultant:

Timeline: From Discovery to Clinical Trials2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Discovery: PCSK9 Mutations Cause Dominant LDL

Segregation of Chr 1p34.1−p32

Pedigree of family HC92

Nat Genet, 2003 (Boileau)

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

S127R D374Y

PCSK9 Mutations Cause Dominant LDL

Nat Genet, 2003 (Boileau)Proc Natl Acad Sci, 2003 (Seidah)

Autocatalytic cleavage

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Overexpression of PCSK9 Eliminates LDLRs in Liver and Increases LDL-C

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

J Biol Chem, 2004 (Horton);Proc Natl Acad Sci , 2004 (Breslow); J Biol Chem, 2004 (Seidah)

LDL

LDL

LDL

Gain of Function (GOF) mutations LDL

Loss of Function (LOF) mutation LDLHypothesis

- S127R- D374Y

J Biol Chem, 2004 (Horton);Proc Natl Acad Sci , 2004 (Breslow); J Biol Chem, 2004 (Seidah)

Normal Function of PCSK9 to Promote Degradation of LDLRs

LDL receptorLiver

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

PCSK9

LDL

LDL LDL

<5%

Dallas Heart Studyn = 3,557

50% African-AmericanLDL-C

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

N - - C

African-Americans2%

Do LOF Mutations in PCSK9 Lower LDL?

LOF Mutations in PCSK9 Lower LDL-C

European-Americans3%

African-Americans2%

LDL : 21% 40%

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

N - - C

Plasma Cholesterol Levels are Reduced in PCSK9 KO Mice

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

CHD Deaths(per 1000 / 10 y)

PCSK9 Mutations: A Tool to Test Role of LDL-Cholesterol in CHD Risk

Does Higher LDL-C Alone ExplainDifference between Shanghai and MRFIT?

Mendelian Randomization

= No DNA sequence variant

= DNA sequence variant that alters LDL-C level

VSCompare CHD in

LOF Mutations in PCSK9 Lower CHD

LDL: 28%

African-Americans

CHD:(over 15 y)

88%

Lancet, 2007 (McPherson)NEJM, 2008 (Katherisan)

15%

European-Americans

46%

ARIC Study (NIH): Eric Boerwinkle

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

LDL-Lowering and Reduction in CHD

Reduction in LDL-C

Reduction in CHD

-10%

-100%

-50%

-10% -20% -30%0

Linsel-Nitchke et al. PlosOne, 2008Willer et al. Nat Gen, 2008;

APOBSORT1

LDLR

Statins - 5 y

PCSK9

PCSK9

R46L

Y142XC679X

Phenotype(LDL level) Genotype(s)

Static Integrated • chronic exposure• single measurement

0 y

80 y

Genotypes Capture Another Dimension: Time

Cumulative LDL-C: mg/dL X years

PCSK9: From Genes to Public HealthA little too little

Year

LDL

Goa

l

100

140

'87 '93 '0160

'04

NCEP Guidelines

A little too late

40 50 60 70

Ris

k R

educ

tion

(%)

Age at Onset of Rx (years)

30

?

A Healthy Woman with No PCSK9

PCSK9in blood

Age (y): 51

32

53

6 3

10

Healthy, college graduate & aerobic instructor (now 38 y)

Zimbabwe: 21 y woman pregnant woman. LDL=16 mg/dL Hooper et al., Atherosclerosis 193:445

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Discovery

Target Validation3 Years

PCSK9: New Target for LDL-Lowering & CHD Prevention

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

PCSK9: Target for LDL-Lowering

STATINS SREBP2(transcription

factor) LDLR

PCSK9

LDL• Efficacy • Safety (?)• Mechanism of action ?

Parabiosis in Mice

PCSK9 acts extracellularly

J Clin Invest, 2006 (Horton)

PCSK9-Tg WTImmunoblot of liver proteins

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Normal Arh-/-

- PCSK9

+ PCSK9

PCSK9 Promotes LDLR Degradation in Hepatocytes

PCSK9

LDLR

C-terminalDomain

CatalyticDomain

Prodomain

Nat Struc Mol Bio, 2007 (Pfizer); Structure, 2007 (Amgen) ; Proc Natl Acad Sci, 2007 (Novartis); Proc Natl Acad Sci, 2008 (Diesenhofer)

PCSK9 Crystal Structure

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Adapted from Brown and Goldstein

LDL

PCSK9 & LDL Receptor

J Biol Chem, 2004 (Horton)Proc Natl Acad Sci, 2005 (Breslow)

Therapeutic Approaches to Inhibit PCSK9 Action

1. Inhibit enzymatic activity Small molecule

2. Disrupt PCSK9:LDLR interaction Anti-PCSK9 antibody Peptides or small molecule

3. Inhibit PCSK9 synthesis RNAi Anti-sense RNA

mAb(primates)

Proc Natl Acad Sci, 2008 J Lipid Res, 2007

Proc Natl Acad Sci, 2009

ASO(mice)

RNAi(primates)

Inhibiting PCSK9 Lowers LDL-C in Animals….

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Evolocumab (AMG 145) in Patients with Hypercholesterolemia (MENDEL-2)

Randomized, Double-blind, Placebo-controlled, Phase 3 Study

Koren et al. J. Am. Coll. Cardiol. 2014. In press.

Alirocumab (REGN727) + Statin in Pt’s with Hypercholesterolemia

(Randomized, Double-blind, Placebo-controlled, Phase 2 Study)

Roth EM et al. 2012. N Engl J Med. 367:1891-1900

siRNA to Silence PCSK9(Phase I)

Fitzgerald et al. 2014. Lancet. 2014. 383:60-68.

PCSK9: From Discovery to Clinical Trialsin 7 Years

Discovery

Target Validation3 Years

Target Validation

Phase I4 Years

Outcomes

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

JCC Seminars: From Discovery to History in 7 Years

Phase 1“Genetics of

PCSK9”

Phase 2“Mechanism of

Action of PCSK9”

Phase 3“A Short History

Of PCSK9”

2003 2004 2005 2006 2007 2008 20102009 2011 2012 2013

Human Genetics & Translation

• Selection of target - Lessons from lipoproteins

• Leveraging human genetic variation- Target validation- Target safety- ‘Goldilocks’ alleles (low frequency,large effect size)

• Genetics coupled to biological expertise

Is PCSK9 Paradigmatic or Exceptional?

Both.

Exceptional in the narrow sense (few relatively common alleles with large effects on disease).

Paradigmatic in the broad sense (more alleles, lower frequency, moderate effects on disease).

0

40

80

120

LDL‐C TG HDL‐C

ANGPTL3 +/FsANGPTL3 +/+P=0.03

P=0.08

P=0.53

2009: Frameshift Mutations in ANGPTL3 and Lipoprotein Levels in the Dallas Heart Study

mg/dl

2010: Mutations in ANGPTL3 Cause Hypolipidemia*

TG: 51 98 LDL: 73 99HDL: 33 35

TG: 24 22 17 19 LDL: 35 31 29 37HDL: 16 16 22 18

*Musunuru et. al. NEJM

Time

- Male sex- LDL- Diabetes- Smoking- Hypertension- HDL

Complex Diseases Are Not Necessarily So Complex

*Age 50 y

LDL X Time = Cumulative ExposureRx

• Diet• Lipid-lowering meds

University of Texas Southwestern Medical Center

Helen HobbsJay D. Horton

NIH PO1 HL20948 – Year 35D.W. Reynolds FoundationHoward Hughes Medical Institute

FPLC – fasting refeeding

FPLC of Plasma From Angptl8-/- Mice

Reduced Fat Mass in Angptl8-/- Mice

* P < 0.05** P < 0.01No significant differences (indirect calorimetry- 5 d)

• Food intake• O2 consumption• RER• Activity