What’s Up with Bleeding2012

• Michael Recht, MD, PhD• Director, The Hemophilia Center• Oregon Health & Science University

Disclosures

• Research funding directly to OHSU from– NovoNordisk– Baxter– Biogen Idec– Pfizer

Agenda

• Bio-engineering of coagulation proteins– Chemical modification– Fusion to protein conjugates

• Alternative hemostatic approaches• Gene transfer

History of hemophilia

• The existence of hemophilia has been known for centuries—first mentioned in the Talmud ~2000 years ago– “If she circumcised her first child and he died, and

a second one also died, she must not circumcise her third child”

History of hemophilia therapy

• Prior to the 1940’s-supportive care and whole blood transfusion– Low concentration of factors VIII and IX– Individuals suffered significant pain and morbidity– Average lifespan 27 years

• 1964-Judith Graham Pool described method to produce cryoprecipitate– Rich in FVIII and fibrinogen– Beginning of home infusions– Average lifespan 40 years

History of hemophilia therapy

• 1970’s-plasma-derived factor concentrates– Pools of 20,000+ donors– Made school, work, and travel possible– Average lifespan 60 years

However….

• Hepatitis B and C were known to be in plasma supply• Thought to be an “acceptable” risk in light of drastic

improvement in quality of life• First individual with hemophilia that died from HIV

infection reported in 1982– Only retrospectively was it discovered that plasma-

derived factor was vector– HIV was isolated in 1984– Heat treatment of plasma-derived factor became

standard practice in 1985

Recombinant factor VIII

• FVIII gene sequenced and cloned in 1984• First clinical trial of recombinant FVIII

concentrate reported in 1990• First recombinant FVIII concentrate marketed

for clinical use in 1992

Recombinant factor IX

• Human FIX gene was cloned in 1982• Minor differences in post-translational

sulfation between recombinant FIX and plasma derived FIX

• 30% lower in vivo recovery of recombinant FIX• Became available for clinical use in 1994

Engineering of coagulation proteins

• Impetus derives from clinical limitations of current therapies– Require IV access– Must be given repeatedly due to short half-lives– Associated with development of inhibitory antibodies

• 30% of patients with hemophilia A• <5% of patients with hemophilia B (though associated with

anaphylaxis)

– Available therapy for those with inhibitors is suboptimal

Rational design of new products

• Increase half-life• Improve ease of delivery• Reduce immunogenicity• Increase potency

Wish list

Characteristic

Relevance to all hemostatic therapies

Relevance to bioengineered

coagulation factorsLeast invasive mode of administration x x

Least requirement for dose manipulation x x

Maximal half-life x

Lowest immunogenicity x

Highest tolerability x x

Lowest thrombogenicity x

Lowest cost over a lifetime x x

Fogarty, Hematology 2011

Chemical modification

PEGylation

• Covalent conjugation of polyethylene glycol (PEG) to a therapeutic protein– Creates “cloud” around protein– Shields from exposure to:

• Proteolytic enzymes• Clearance receptors• Immune effector cells

• Effective at prolonging half-life of many other biopharmaceuticals– Interferon– Asparaginase– G-CSF

PEGylation concerns

• PEG polymers may interfere with the peptide interaction with substrates, reducing activity

• Randomly PEGylated B-domain-deleted recombinant factor VIII

Interaction of FVIII with vWF

Conjugate Specific activity Protein able to bind vWF

rFVIII 15 u/mg 98%

Conjugate #1 7.0 u/mg 43%

Conjugate #2 5.3 u/mg 26%

Rostin, Bioconjugate Chem, 2000

Site-directed PEGylation

Mei, Blood, 2010

Glyco-PEGylation

• Recombinant factor VIIa (Ghosh, J Thromb Haemost, 2009)

– Increased factor X cleavage at lower concentrations

– Reduced interaction with phospholipids– NN7128-three phase 1/2 clinical trials complete

• Recombinant factor IX – Equivalent potency to recombinant factor IX– Better duration of hemostasis after bleeding

challenge

Glyco-PEGylation

• Recombinant factor IX – Equivalent potency to

recombinant factor IX– Better duration of

hemostasis after bleeding challenge in mice

– Half-life prolonged to 93 hours

Negrier, Blood, 2011

Liposomal PEGylation (PEGLip)• Liposomes are small vesicles

consisting of a phospholipid bilayer surrounding aqueous interior

• PEGylation of the liposome decreases clearance

• No difference in clotting or other laboratory parameters between study drug and standard recombinant factor VIII concentrate.

• There was a trend towards fewer bleeding episodes 14 days after infusion of PEGLip-recFVIII concentrate

• Pulled from further trials

Powell, J Thromb Haemost, 2008

Long term use of PEGylated proteins

• Accumulation of PEG polymers has been documented in some preclinical studies

• Whether accumulation will be evident in those receiving these medications over a life-time is unknown

• In other compounds, anti-PEG antibodies have been demonstrated

Fusion to protein conjugates

rFVIII:Fc-IgG fusion (rFVIIIFc)

• IgG molecule has a prolonged t1/2 secondary to continual recycling• Fc domain of IgG is a “natural” molecule with no known toxicity• rFVIIIFc is a single molecule of rFVIII covalently linked to human IgG1

Fc domain• Compared to full length rFVIII, rFVIIIFc had a 1.7-fold longer t1/2

Powell, Blood, 2012 preprint

rFIX:Fc-IgG fusion (rFIXFc)

• Same technology as used in rFVIIIFc

• Compared to rFIX, t1/2 was extended from 18 hours to 60 hours

Shapiro, Blood, 2012 preprint

Prophylaxis schedule with rFIXFc

Shapiro, Blood, 2012 preprint

Alternative hemostatic approaches

• Non-peptide molecules may provide opportunities to enhance hemostasis in bleeding disorders patients

• These molecules have the potential for oral or subcutaneous administration

Aptamers

• Ribosomal or deoxyribosomal oligonucleic acids that can be produced to any length or 3D conformation

• Allow binding to an unlimited array of physiologic targets

ARC19499

• Aptamer antagonist of tissue factor pathway inhibitor

• Improved thrombin generation in plasma from subjects with FVIII deficiency

• Improved clot times as measured by thromboelastography in monkeys depleted of FVIII

Waters, Blood, 2009

Fucoidans

• Also known as non-anticoagulant-sulfated polysaccharides (NASPs)

• Heterogeneously sized anionic compounds that derive from marine plants (brown seaweed)

• Have demonstrated pharmacologic activity:– Anti-cancer– Anti-inflammatory– Anti-angiogenesis– Anti-coagulant

AV513

• Fucoidan with anti-TFPI activity

• Fed to hemophilia A dogs:– Improved clotting time as

measured by TEG– Improved cuticle bleeding

time

• Phase 1 trial of BAX513 in healthy subjects is underway

Prasad, Blood, 2008

Read-through of nonsense mutations

• Nonsense mutations (premature termination codons [PCTs]) account for 11% of known mutations in severe hemophilia

• First described in 1964, new compounds are being developed to promote ribosomal read-through of PCTs

• Has been shown to be safe and effective in restoring the cystic fibrosis and Duchenne muscular dystrophy proteins

Ataluren (PCT124)

• No major adverse events reported in CF or Duchenne MD

• Currently phase 2 trial of 28 day treatment cycle with Ataluren in hemophilia A and B is recruiting patients

AAV-vector mediated gene transfer in hemophilia B

• AAV8 vector– Packaged as complementary dimers within single

virionmediate transgene expression at higher levels than single-stranded AAV vectors

– AAV8 has lower seroprevalence that previous AAV vectorsdecreased humoral immunity

– Since AAV8 has tropism for liver, able to administer via peripheral vein

Nathwani, NEJM, 2011

Study design

Inclusion Criteria• 18-60 years of age• Factor IX activity < 1%• No history of inhibitor or

anaphylaxis to FIX• No immunity to AAV8

Dose level cohorts• Cohort 1: 2 x 1011 vg/kg• Cohort 2: 6 x 1011 vg/kg• Cohort 3: 2 x 1012 vg/kg

Results

• AAV-mediated expression of FIX at 2-11% of baseline levels was observed in all participants

• Four of the six discontinued FIX prophylaxis• In the other two, time between prophylactic

infusions has increased• Both participants receiving highest dose had

asymptomatic elevations of AST treated with short course of prednisone

Results

Nathwani, NEJM, 2011

Conclusions

• Peripheral vein infusion of AAV8 vector resulted in FIX expression at levels sufficient to improve bleeding phenotype with few side effects

• Immune-mediated clearance of AAV-transduced hepatocytes remains a concern

Benefits of correcting hemostatic defect in hemophilia

Reduced mortality and morbidity

Reduced joint bleeding and hemophilic arthropathy

Improved joint health, mobility, and physical activity

Improved patient perception of disease state

Improved quality of life

Improved tolerability of antithrombotic medications

Improved tolerability of invasive procedures

Fogarty, Hematology 2011

Benefits of novel therapeutics

• Prophylaxis– Half of US treatment centers do not follow the

advice of MASAC of NHF to start three times weekly prophylaxis in those with severe hemophilia

– Requirement of frequent infusions and need for central venous access in children most common reason given

– Fewer infusions and non-intravenous alternatives may improve adherence and long term outcome

Benefits of novel therapeutics

• Joint health– Early three times weekly prophylaxis has been

demonstrated to protect the joints of children with severe hemophilia A

– Follow-up to the initial study is ongoing, but preliminary data indicate that this protective effect continues into adolesence

– Decreasing infusion frequency may increase adherence

Benefits of novel therapeutics

• Quality of life– QoL increases in children with severe hemophilia

A on prophylaxis compared to those receiving factor on-demand

– The potential impact of improved hemostasis with new products is substantial• Decreased missed work or school• Decreased pain

Questions?

Conclusions

• Great strides have been made in the treatment of hemophilia– Understanding of the inherent limitations of currently

available therapies– Unmet clinical needs of the affected population

• Novel agents hold promise for:– Improved safety and efficacy– Improved joint health– Improved quality of life– Increased tolerability

Contact information

• The Hemophilia Center at OHSU– 503-494-8716 (office)– 503-494-0714 (fax)– rechtm@ohsu.edu