Fc receptor therapies Transfusion residents National CBS 2010

CBS Transfusion Medicine Resident Scientific Session Nov 2, 2010

Recent advances in Fc receptordependent therapies Alan H. Lazarus, PhD

Canadian Blood Services St. Michael’s Hospital University of Toronto

Outline •Mechanisms of platelet destruction in ITP •Recent advances in:

• Fc receptor blockade • IVIg • anti-D • Inhibition of Fc receptor signaling •Conclusions

Immune thrombocytopenic purpura (ITP)

Platelet

An Fc-independent mechanism of potential immune thrombocytopenia-ROS GPIIbIIIa

ROS Zang W et al, Blood 2008 Li Z et al, J Biol Chem 2008 Li Z et al, Blood 2009 Wright JF et al, Br J Haem 1996 Peterson, Blood 2008 Quinine

Fc-independent immune thrombocytopenia-GPIb

Platelet

GPIb

Nieswandt et al Blood 2000 Webster et al Blood 2006 Go et al Haematologica 2007

Carbohydrates important for IgG function?

Fc F(ab’)2

IgG subclass-specific effects of EndoS-mediated hydrolysis of the IgG-associated sugar side chain.

Albert H et al. PNAS 2008;105:15005-15009

©2008 by National Academy of Sciences

A potential alternative mechanism of platelet destruction? Cell Mediated Immunity

Olsson et al, Nature Medicine 2003 Zhang et al, Eur J Haem 2006

An alternative mechanism which may contribute to immune thrombocytopenia?

MK in Bone Marrow

PLT PLT

Decreased platelet production due to antibody and/or T cells inhibiting or destroying MK

Mechanism

Fc receptors Reference

classic anti-IIbIIIa

yes

Harrington Experiment 1951

anti-IIbIIIa (49-66)

no

Zhang, 2008 Li, 2008 Li, 2009

anti-GPIb

no

Nieswandt, 2000 Webster, 2006 Go, 2007

T cells

no

Olsson, 2003 Zhang, 2006 Chow, 2010

Direct effect on MK

?

Chang 2003 McMillan 2004 Houwerzijl 2004

Outline •Mechanisms of platelet destruction in ITP •Recent advances in:

• Fc receptor blockade • IVIg • anti-D • Inhibition of Fc receptor signaling

+other potential Fc receptors for IgG (FCRL’s)

Fc receptor blockade •Antibody specific for FcRIIIa worked in refractory ITP (Clarkson, 1986) •Antibody specific for FcRI worked in ITP (Terjanian, 2000 abstract in Blood) •Humanized antibody for FcRIIIa (GMA161& 3G8) worked but with side effects (Bussel, 2006 & Nakar, 2009 abstracts in Blood)

Fc receptor blockade • Clodronate is effective in a mouse model of passive ITP (Alves-Rosa 2000, 2002; Deng & Balthasar 2005, 2007; Li 2006)

Taken from, www.clodronateliposomes.org

Outline •Mechanisms of platelet destruction in ITP •Recent advances in:

• Fc receptor blockade • IVIg • anti-D • Inhibition of Fc receptor signaling

Intravenous Immunoglobulin

IVIg is IgG

Fc

F(ab’)2

How does IVIg work?

What does IVIg bind to? • • • • • • • •

Other antibodies [Schussler, 1997] Glycolipids [Vuist, 1997] Superantigens [Takei, 1993] HLA antigens [Glotz, 1993] DNA & Phospholipid [Krause, Blank, Shoenfeld. 1998] Fas/FasL [Viard, 1998, Prasad, 1998, Altznauer, 2003] IgE Fc receptor & Tetanus toxoid [Horn, 1999] Galactose disaccharides [Barreau, 2000] • Auto-antigens [reviewed in St-Amour, 2009] • Red Blood cells [Salama, 1984] • ……

Theories of IVIg action • • • • • • • • • •

Fc receptor blockade Inhibitory FcγRIIB expression Regulation of cytokine production Anti-idiotypic antibodies Inhibits complement activation Clearance of pathogenic antibody-FcRn. Apoptosis IVIg possesses anti-inflammatory sugars IVIg forms soluble immune complexes Dendritic cells

Hypothesis Can antibodies which form an immune complex ameliorate ITP?

Teeling et al, Blood 2001

Experimental Design Cell-associated: (anti-D like effect)

Soluble:

OVA

A murine model of Passive Immune Thrombocytopenia Ravetch et al (NY) Bleeker et al (Amsterdam)

24h

Balthasar & Deng (Buffalo)

Lemieux & Bazin (Quebec City) Webster et al (Toronto)

FSC

800 600 400

Treatment

Platelet count x 109/L

1000

***

***

***

***

2

3

4

200 0

0

1

anti-platelet antibody

Inhibitory FcγRIIB

Activating FcγR

1000

800

800

Treatment

1000

600 400

600 400

200

200

0

0

0

1

2

anti-platelet antibody

3

4

0

1

Treatment

FcγRIIB-/- mice

***

2

3

anti-platelet antibody

***

4

If IVIg functions via the formation of an immune complex, could these complexes be reacted with cells which could then be adoptively transferred to recipients?

Mix

Wash

Load

Inject

Amelioration of ITP?

Adoptive-transfer of IVIg effects with leukocytes

Siragam et al, Nature Medicine 2006

Proposed model of IVIg action in murine ITP

ITP Siragam et al, J Clin Invest 2005 Park-Min, Immunity 2007 Siragam et al, Nat Med 2006 Tha-In et al, Blood 2007 Ephrem et al, Blood 2008 Aubin et al, Blood 2010

Outline •Mechanisms of platelet destruction in ITP •Recent advances in:

• Fc receptor blockade • IVIg • anti-D • Inhibition of Fc receptor signaling

F c  R IIIA E x p re s s io n

Anti-D-like antibodies reduce the ability to detect activating Fcγ receptors on macrophages 200

150

100

*

*

TER119

M1/69

50

0 Nil

IVIg

30-F1

Song et al, Blood 105:1546-8, 2005

Does anti-D ameliorate ITP via the same mechanism as IVIg?

2 hr IVIG

IL-10

Anti-D

IL-10 MCP-1 IL-6 TNFα

Cooper et al, Br J Haem 2004

Blood 2007

Of the 6 patients in the study who had failed to respond to IVIG and anti-D given as single agents, 5 of them responded when given in combination.

Outline •Mechanisms of platelet destruction in ITP •Recent advances in:

• Fc receptor blockade • IVIg • anti-D • Inhibition of Fc receptor signaling •Conclusions

A new therapeutic intervention based upon the mouse model of ITP?

Platelet phagocytosis in ITP

syk

R406

Braselmann, et al. J Pharmacol Exp Ther, 2006

Inhibition of syk ameliorates murine ITP

Platelet count x 10 9/L

1000 Unmanipulated

800 600 400 200 0

Treatment

Nil

Vehicle

R788 25 mg/kg

R788 40 mg/kg

IVIg 2 g/kg

Podolanczuk, A. et al. Blood 2009;113:3154-3160

syk inhibition in 16 adult refractory ITP patients

Podolanczuk, A. et al. Blood 2009;113:3154-3160 Copyright ©2009 American Society of Hematology. Copyright restrictions may apply.

Conclusions •Multiple potential mechanisms of thrombocytopenia in ITP. •The mechanism of IVIg remains unclear but may work via an Fc receptor-dependent cellular pathway (DC) •Anti-D appears to work via a mechanism distinct from IVIg •Inhibition of Fc receptor signaling or function could be a potential therapy in ITP

Thank you! Andrew R Crow Dr. Zhong-Wei Chai Sara Suppa Xi Chen Joan Legarda Patrick Mott

Dr. John Freedman Dr. Bernadette Garvey Dr. John Semple Dr. Heyu Ni Dr. Valery Leytin Dr. Margaret Rand

Derry Roopenian Jim Bussel Anna Podolanczuk