1. Future directions for CKD anaemia
Iain C Macdougall BSc, MD, FRCP
Consultant Nephrologist and Honorary Senior Lecturer
Renal Unit, King’s College Hospital, London, UK

2. Disclosure Honoraria, lecture fees, grants from:- Amgen Ortho Biotech
Roche
Affymax
Shire 2

3. 1

4. Lancet, 9th September 2006 1

5. Outline of presentation
Overview of erythropoiesis in CKD in 2007
Cellular and molecular mechanisms relevant to anaemia
Future therapies for stimulating erythropoiesis
Future developments in the management of CKD anaemia 2

6. Outline of presentation
Overview of erythropoiesis in CKD in 2007
Cellular and molecular mechanisms relevant to anaemia
Future therapies for stimulating erythropoiesis
Future developments in the management of CKD anaemia 2

8. Erythropoiesis in CKD Erythropoietin Iron SCF, GM-CSF, IL-3
SCF, IL-1, IL-3,
IL-6, IL-11
About 8 Days
Pluripotent Stem Cell
Burst-Forming Unit-Erythroid Cells (BFU-E)
Colony-Forming Unit-Erythroid Cells (CFU-E)
Proerythro- blasts
Erythro- blasts
Reticulocytes
RBCs
Papayannopoulou T, et al. In: Hoffman R, et al., ed. Hematology: Basic Principles and Practice. 4th ed. 2005;

9. Erythropoiesis in CKD EPO Iron + + ─ ─ Pro-inflammatory cytokines
 Fas Ag
Apoptosis ─ +
Iron
 Fe absorption
 Fe transport
 Fe availability
(EPO-R, Tf, TfR, Ferriportin, DMT-1) ─
 EPO production
Pro-inflammatory
cytokines
(IL-1, TNFα, IL-6, IFNγ)
hepcidin

10. Outline of presentation
Overview of erythropoiesis in CKD in 2007
Cellular and molecular mechanisms relevant to anaemia
Future therapies for stimulating erythropoiesis
Future developments in the management of CKD anaemia 2

11. rHuEPO stimulates erythropoiesis by activating EPO receptors
Activation
membrane
Signal Transduction
Growth and Differentiation

12. Activation of EPO receptors results in initiation of intracellular signalling pathways
Jak2
rHuEPO
Jak2 P
Jak2 P
Jak2 P
membrane
Conformational change
Phosphorylation of JAK2, a tyrosine kinase
Phosphorylation of EPO receptor
Initiation of intracellular signalling pathways

13. EPO receptor activation leads to activation of genes that promote erythropoiesis
membrane P P
Jak2
Jak2 P P
STAT 5 P
nucleus
Gene Activation
Survival, differentiation, proliferation, and maturation of RBC progenitors and precursors

14. EPO receptor activation leads to activation of genes that promote erythropoiesis
membrane P P
Jak2
Jak2 P P
SCH
SOS
GRB
STAT 5 P
MAPK
PI-3-Kinase
nucleus
Gene Activation
Survival, differentiation, proliferation, and maturation of RBC progenitors and precursors

15. All ESAs have the same signalling pathway
Investigational
EPO, rHuEPO
Darbepoetin alfa
C.E.R.A.
Peg-rHuEPO
EPO dimer
EPO mimetic peptide
Jak2 P
Jak2 P
Jak2 P
Jak2 P
Jak2 P
membrane
Signal Transduction
Survival, differentiation, proliferation, and maturation of RBC progenitors and precursors
Gene Activation

16. Termination of EPO-receptor signalling
JAK 2
JAK 2 P
Haemopoietic cell phosphatase P P P
Internalisation and degradation

17. All ESAs have the same mechanism of action
Activation of the EPO receptor is the common mechanism by which all ESAs stimulate erythropoiesis
However, ESAs may differ from one another in:
Biophysical characteristics (e.g. molecular weight)
EPO receptor binding affinity
Pharmacokinetic properties (e.g. serum half-life, clearance)
Main Points
• All erythropoiesis stimulating proteins share the same mechanism of action—activation of the EPO receptor.1-6
• While the mechanism of action for all ESPs is the same, these agents may differ from one another in: (a) biophysical characteristics, (b) EPO receptor binding affinity, and (c) pharmacokinetic properties.1-3
• The next series of slides explore this concept by tracing the development of darbepoetin alfa, and the comparative differences of this agent with rHuEPO.
References
Constantinescu SN, Ghaffari S, Lodish H. The erythropoietin receptor: structure, activation, and intracellular signal transduction. Trends in Endocrinology & Metabolism. 1999;10:18-23.
Elliott S, Egrie J, Browne J, et al. Control of rHuEPO biological activity: the role of carbohydrate. Exp Hematol. 2004;32:
Elliott S, Lorenzini T, Asher S, et al. Enhancement of therapeutic protein in vivo activities through glycoengineering. Nat Biotechnol. 2003;21:
Hasselbeck A, Reigner B, Jordan P, et al. Pre-clinical and phase I pharmacokinetic and mode-of-action studies of CERA (continuous erythropoiesis receptor activator), a novel erythropoietic agent with an extended serum half-life. Proc Am Soc Clin Oncol. 2003;22. Abstract 3006.
Qui H, Belanger A, Yoon H-WP, Bunn HF. Homodimerization restores biological activity to an inactive erythropoietin mutant. J Biol Chem. 1998;273;
Wrighton NC, Balasubramanian P, Barbone FP, et al. Increased potency of an erythropoietin peptide mimetic through covalent dimerization. Nature Biotech. 1997;15:

18. Weiss & Goodnough, NEJM, March 2005

20. Outline of presentation
Overview of erythropoiesis in CKD in 2007
Cellular and molecular mechanisms relevant to anaemia
Future therapies for stimulating erythropoiesis
Future developments in the management of CKD anaemia 2

21. Lancet 9th September 2006 1

22. I. EPO-receptor agonists
Protein-based ESA therapy
Epoetin (alfa, beta, delta, omega)
Biosimilar EPOs (epoetin zeta)
Darbepoetin alfa
C.E.R.A. (methoxy polyethylene glycol epoetin beta)
Synthetic erythropoiesis protein (SEP)
EPO fusion proteins
~ EPO–EPO
~ GM–CSF–EPO
~ Fc–EPO
~ CTNO 528
Small molecule ESAs
Peptide-based (e.g. Hematide)
Non-peptide based

23. II. Other mechanisms Prolyl hydroxylase inhibitors (HIF stabilisers)
GATA inhibitors
Haemopoietic cell phosphatase (HCP) inhibitors
EPO gene therapy

24. I. EPO-receptor agonists
Protein-based ESA therapy
Epoetin (alfa, beta, delta, omega)
Biosimilar EPOs (epoetin alfa, epoetin zeta)
Darbepoetin alfa
C.E.R.A. (methoxy polyethylene glycol epoetin beta)
Synthetic erythropoiesis protein (SEP)
EPO fusion proteins
~ EPO–EPO
~ GM–CSF–EPO
~ Fc–EPO
~ CTNO 528
Small molecule ESAs
Peptide-based (e.g. Hematide)
Non-peptide based

25. II. Other mechanisms Prolyl hydroxylase inhibitors (HIF stabilisers)
GATA inhibitors
Haemopoietic cell phosphatase (HCP) inhibitors
EPO gene therapy

26. Multiple generic epoetin alfas are being marketed worldwide
variable quality
variable biological activity
inaccurate labelling
? immunogenicity
PERU
China

27. The “generic” epoetin alfas differ from approved epoetin alfas
Isoelectric Focusing*
The first biosimilar epoetin alfa was approved in Europe
in June 2007 (Sandoz) – others pending

28. C.E.R.A. Continuous Erythropoietin Receptor Activator
Methoxy polyethylene glycol epoetin beta
C.E.R.A.
EPO
long-acting ESA
IV half-life = SC half-life = 130 hrs
?once–monthly dosing

30. EPO mimetic peptides
Family of peptides discovered with erythropoietin-mimetic activity
Amino acid sequences completely unrelated to native EPO
Same functional / biological properties as EPO
First one described was EMP-1
A similar EMP pegylated and dimerised to produce HematideTM
HematideTM is about to begin Phase III of clinical development
– stable at room temperature
– antibodies do not cross-react with EPO, x1/month, ?cheaper

31. Hematide in pre-dialysis CKD patients (Phase 2)
Injection 1
Injection 2
Injection 3
Injection 4
Injection 5
Injection 6
(Target Hb Range: 11 – 13 g/dL)
-0.5
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0 4 8 12 16 20 24
Week
Mean Hb Change From Baseline (g/dL)
Starting Dose:
0.025 mg/kg SC
0.050 mg/kg SC
0.075 mg/kg SC
0.050 mg/kg IV
Macdougall et al., ASN, San Diego, Nov

32. 10 CKD patients with Ab+PRCA (Germany, France, UK)
Treated with once-monthly SC Hematide 0.05mg/kg
End-point – Hb > 11 g/dL without RBC transfusions 2

33. HIF stabilizers (Prolyl hydroxylase inhibitors)2 O2 3
PHD
proteasomal
degradation
HIF-1 OH 1
HIF-2 OH
HIF 
HIF-2
HIF-1
HIF target genes - EPO, etc.
HRE
Macdougall & Eckardt, Lancet, 2006

34. HIF stabilizers
Orally-active inhibitors of HIF prolyl hydroxylase have been synthesized (FG-2216; FG FibroGen)
They cause an increase in EPO levels, even in CKD patients
FG-2216 was in phase II of its clinical trial programme
Urquilla P et al.: J Am Soc Nephrol 2004; 15: 546A.
FG-2216 Patients
Placebo Patients
Wiecek A. et al. XLII ERA-EDTA Congress, Istanbul 2005.
BUT
FDA has suspended further development of FG-2216 following the death of a female patient from fulminant hepatic necrosis 2
Urquilla P et al.: J Am Soc Nephrol 2004; 15: 546A

35. Outline of presentation
Overview of erythropoiesis in CKD in 2007
Cellular and molecular mechanisms relevant to anaemia
Future therapies for stimulating erythropoiesis
Future developments in the management of CKD anaemia 2

36. Further initiatives
New iron preparations – ferrumoxytol, Ferrinject, etc.
New trials 2

37. New trials n RCT-db RCT-sb EXTEND Study design Study pop. Follow-up
Goals
End-points
RCT-db
Type 2 DM
CKD
4000
4 years
Hb 13 vs 9
CVS
RCT-sb
> 70 yrs
260
88 wks
Hb 13 vs 9.5
QoL
SF36 vit
EXTEND
Observ.
CKD pts
q2wk vs q4wk DA
PREFERENCE
Prospective
cohort
Several hundred
Compare diff.mode of admin.
ESA mode of admin.
MIRCERA in Renal Tx
Open-label
Renal Tx
1 year
 Hb 2

38. 3896* Number of patients 1000 2000 3000 4000 TREAT1 CHOIR2 CREATE3
Besarab4
3896*
* IVRS 12 October 2007 2

39. Total follow-up (pt-yrs)
TREAT vs. CHOIR
No. of centres
Median (pt-mths)
Total follow-up (pt-yrs)
Events (n)
CHOIR
130 14
1900
222 2

40. Total follow-up (pt-yrs) > double that of CHOIR
TREAT vs. CHOIR
No. of centres
Median (pt-mths)
Total follow-up (pt-yrs)
Events (n)
CHOIR
130 14
1900
222
TREAT
700
16 (already)
(already)
> double that of CHOIR 2

41. Questions we don’t have answers to
Are high doses of EPO bad or is it just that “being hyporesponsive” is bad?
Is giving EPO to “hyporesponsive” patients bad?
Is a Hb of 11–12 g/dl appropriate for all CKD patients?
Does Hb variability/instability/cycling impact on mortality/morbidity, or is it just a marker of outcome? 2