E rythropoietin -stimulating agents resistance and new anemia therapies

Erythropoietin-stimulating agents resistance and new anemia therapies

Narrative Review

Fellow 潘恆之 /VS 鄭昌錡

Outline

Introduction

EPO resistance

New anemia therapies

Introduction

• Definition of anemia in CKD ： Hgb <13.5 g/dl for male; Hgb <12.0 g/dl for female

• Erythropoietin-stimulating agents ：Recombinant human erythropoietin was introduced as

a treatment for the anemia associated with chronic kidney disease (CKD) since 1989.

Erythropoietin therapy rendered many patients free of blood transfusions with dramatic benefits on quality of life (particularly physical capacity) and other physiologic effects of increasing hemoglobin levels from ~6 g/dL up to approximately 11-12 g/dL.

• Several studies showed that partial correction of anemia (to hemoglobin levels in the range of 10-12 g/dL) was a safer strategy, reducing the risk of increased arterial and venous thromboembolism and other possible harmful effects in CKD patients.

Introduction

Rajiv Agarwal, Clin J Am Soc Nephrol , 2010 (5): 1340–1346

HbEPO doseerythropoiesis

1. CV risk2. Mortality3. Stroke4. Vascular access

thrombosis

Hb > 13 g/dL

Naturally Occurring Higher Hemoglobin Concentration Does Not Increase Mortality among Hemodialysis Patients

Goodkin D A et al. JASN 2011;22:358-365

Among 29,796 HD patients in 12 nations

545 Endogenous EPO patients were included in each model.

Adjusted risk of mortality does not differ significantly for Endogenous EPO patients compared with Other patients(

RR 0.94 (95% CI, 0.72 to 1.22

RR, 0.98; 95% CI, 0.80 to 1.19

RR, 0.81; 95% CI, 0.66 to 0.98

Cumulative all- cause mortality among 428 CKD 1~2 patient aged 86 at baseline, by tertile of erythropoietin level at baseline (lowest = 3.4– 8.6 IU/L, middle = 8.7–12.3 IU/L, highest = 12.4– 103.0 IU/L).

• Higher concentration of endogenous EPO => fatal outcome ↑• Excessive EPO synthesis => off-target biological consequences ↑ ??• Limitations: undiagnosed hypoxemia? Impaired bone marrow response? EPO resistance?

Effect of erythropoietin levels on mortality in old age: the Leiden 85-plus study den Elzen W P et al. CMAJ 2010;182:1953-1958

Association of Mean Weekly Epoetin Alfa Dose with Mortality Risk in a Retrospective Cohort Study of Medicare Hemodialysis Patients

Am J Nephrol 2011;34:298–308

The cohort included 137,918 HD patients. Mean age was 63.2 years

Relative hazards of death over mean EPO dose per week during 3-month exposure period, in patient- months with mean hemoglobin

The distribution of mean EPO dose per week is displayed in the shaded area, and the referent dose is shown by the solid vertical line.

Uremic toxins/Oxidative stress/ Inflammation Nutrition deficiency Heavy metal Hematologic disorders Angiotensin-modulating agents EPO inhibitors

Pattern of resistance to erythropoietin-stimulating agents in chronic kidney disease

Oluwatoyin F. Bamgbola, Kidney international (2011) 80, 464-474

Review

• Erythropoietin-stimulating agents (ESA) resistance ：

Hb < 11g/dL over 3 month despite of EPO dose > 400 IU/kg/wk or 20000 IU/wk (≥1.0μg/ kg for darbepoietin)

UREMIC TOXINS/OXIDATIVE

STRESS/INFLAMMATION

Uremic toxins

• The mechanism is uncertain

• There is fair correlation of urea nitrogen clearance with improved cytokine profiles (IL-6, CRP) and lower requirement for ESA.

• Uremia may be related to： Poor bone marrow response to ESA Accelerated turn over rate of RBC from altering erythrocyte morphology by inducing outward expression of the phosphatidyl-serine content of its inner membrane

Oxidative stress

Haase V H Am J Physiol Renal Physiol 2010;299:F1-F13

✗ *Oxidative stress downregulates the generation of hypoxic inducible factor (HIF) protein.

Inflammation

* Pro-inflammatory cytokines such as IL-1, IL-6, INF-γ, TNF could disrupt iron recycling and absorption

DMT 1, divalent methyl transporter-1

CKD pro-inflammatory

cytokines: IL-6

Process of physiological iron recycling involving macrophages and enterocytes


Erythropoietin receptor activation and intracellular signal transduction

SOCS, suppressor of cytokine signaling; STAT-5 signal transducer and activator of transcription-5

Pro-inflammatory cytokines

(+)

Inflammation* Pro-inflammatory cytokines promote EPO resistence by activation of suppressor of cytokine signaling and inhibition of nuclear factor κB


Inflammation

• Failed allograft – nephrectomy is a reasonable approach for transplant rejection with persistent elevation of inflammatory indices and intractable anemia.

• Dialysis catheters – additional sources of oxidative inflammation are the use of per-cath and synthetic grafts as vascular access in HD subjects.

NUTRITION DEFICIENCY

1. Iron2. Folate3. Vitamin C

Nutrients deficiency

• Iron Principally due to poor diet, frequent blood sampling

and high frequency of infectionAnnual loss of 2g of elemental Fe among HD patientDiagnosis: serum iron < 100 mg/dl, TSAT <20%,

and serum Ferritin < 100mg/dl ~ at least 2 indicesTreatment: 1. Iron therapy 2. Vit-C – mobilize iron from RES


• Folate Principally due to poor gastrointestinal

absorption, poor diet, water-soluble nutrient loss, and high catabolic status

An essential ingredient for nucleotide synthesis, DNA repair, and re-methylation of homocysteine

Folic acid deficiency produces oxidative vascular injury by potentiating uremic inhibition of homocysteine catabolism


• Vitamin CA cofactor for several enzymatic metabolismPromotes GI absorption of iron and enhances its

mobilization from RES.Increases Hb synthesis by facilitating

incorporation of Fe into protoporphyrinAn anti-oxidative free oxygen scavenger,

downregulates hepatic synthesis of cytokinesGreater loss of Vit-C in HD compared with PD

HEAVY METAL1. Aluminum2. Cadmium3. Lead4. Mercury

Heavy metal

• Bone marrow function is often impaired in individuals with heavy metal poisoning

• Aluminum toxicity – P-binder, dialysate, poor control of pharmaceutical standards

• Cadmium, lead, mercury – industrial pollution, some folk remedies of Indian and Middle Eastern origins

Heavy metal

• Lead toxicity is more likely to occur in patient with CKD due to： Iron and calcium deficiencies => GI absorption Uremia may increase mobilization of lead from bone tissue stores

• Lead toxicity causes anemia by precluding incorporation of iron into a protoporphyrin ring for heme synthesis

HEMATOLOGIC DISORDERS

Hematological disorders

• The mechanism of EPO resistance includes bone marrow infarction, hemolysis, hypersplenism, and ineffective erythropoiesis.

• Common chronic hemolytic conditions are auto-immune diseases, sickle cell disease, thalassemia, hereditary spherocytosis, glucose 6-phosphate dehydrogenase deficiency.

Hematological disorders

• Autoimmune hemolytic anemia occurs in 5-10% of patient with SLE. It is frequently associated with renal or neurological involvement.

• As reticulocytosis is common in patients on ESA therapy, early diagnosis of hemolytic events may be missed.

• It should be suspected when there is a progressive increase in EPO requirement while there is rapidly decreasing Hb concentration with megakaryocytic cell line and serum titer of anti-DS DNA.

ANGIOTENSIN-MODULATING

AGENTS

Angiotensin-modulating agents

• The influlence is controversial.• A prototype study showed there is a higher EPO

requirement in dialysis hypertensive patients who were treated with ACEI/ARB compared with CCB.

• ACEI suppresses the enzymatic degradation of N-acetyl-seryl-aspartyl- lysyl-proline (AcSDKP), a naturally occurring inhibitor of erythropoiesis.

• Stimulation of erythroid cellular proliferation by angiotensin binding of its type II surface receptor is inhibited by ARB

=> therapeutic use of ARB in post-transplant erythrocytosis.

CIRCULATING EPO INHIBITORS

Circulating EPO inhibitors

• Circulating EPO inhibitors may result in pure red cell aplasia.

• Pure red cell aplasia should be suspected： 1. Received ESA therapy for 44 weeks 2. Rapid decrease in Hb mass (> 0.5 g/dl per week), 3. Reduction in absolute reticulocyte count <10, 000/ ml and/or 1U of RBC transffusion per week 4. Leucocytes and platelets are normal. 5. The more common causes of EPO-resistant anemia should be excluded.

Circulating EPO inhibitors

• Diagnosis ： 1. Absence of erythroid precursors on bone marrow sample and low EPO content of the serum. 2. The serum sample inhibits growth of erythroid colonies in a bone marrow culture. 3. Radioimmunoassay identifies circulating neutralizing anti-EPO IgG in serum• Treatment ： 1. Discontinuation of rhEPO 2. Steroid and/or calcineurin inhibitor 3. Plasma exchange and/or allograft transplantation

SECONDARY HYPERPARATHYROIDISM

AND 1,25-VITAMIN D DEFICIENCY

Secondary hyperparathyroidism

• There is a higher prevalence of anemia and greater EPO requirement HD subjects who are in the upper 50th percentile of intact parathyroid hormone.

• As a proof of causal relationship, surgical parathyroidectomy led to an improved control of anemia and a lower need for ESA

• Nevertheless, there is no evidence for a direct inhibition of erythropoiesis by excessive PTH.

1,25-vitamin D deficiency

• Vit-D may have synergistic effect on ESA control of anemia

• Calcitriol treatment in subjects with uremic bone disease increases the proliferation of erythroid precursors.

• Lower EPO requirement among subjects with BB gene of vitamin D receptor genotype compared with those with the Bb/bb gene.

Etiology of ESA resistance

Risk factor Mechanism of ESA resistance Therapeutic intervention

Uremic toxins EPO synthesis / erythroid response

Longer effective dialysis

Oxidative stress Downregulation of HIF Vit E and Vit C

Inflammation Cytokines: IL-1, IL-6, TNF-α Avoid sepsis and malnutrition

Iron deficiency Hemoglobin synthesis Replenish iron/ blood loss

Hyperparathyroidism/ Vit D deficiency

Vitamin D synergism (erythropoiesis) Low P diet/ 1,25 (OH)2 vit D

Aluminum toxicity Aluminum bone disease Avoid aluminum intake

Hemolysis HbSS/G6PDD/AIHA Treat underlying diesease

Drug: angiotensin-modulating agents

Erythroid ANG II receptors/ endogenous EPO inhibitor, AcSDKP

Dose of ACEI/ARB

Potential Pharmacological intervention in EPO resistance

• Anti-inflammatory agents• Nutritional supplements• EPO-mimetic peptide• Endogenous induction of EPO

EPO-mimetic peptide HIF Stabilization Hepcidin Modulation GATA-2 Inhibitors EPO gene therapy

New anemia therapies: Translating Novel Strategies From Bench to Bedside

Iain C. Macdougall, Am J Kidney Dis. 2012;59(3):444-451

Narrative Review

EPO-MIMETIC PEPTIDE

EPO-mimetic peptide

• The concept that a peptide could activate the erythropoietin receptor and stimulate erythropoiesis was described first by Wrighton et al in 1996 in Science.

• EMP-1 (erythropoietin-mimetic peptide 1) was able to stimulate cellular proliferation of erythroid cells in a dose-dependent manner and also increase reticulocyte counts in animal models.

• However, EMP-1 had low affinity for the erythropoietin receptor and low biological activity.

EPO-mimetic peptide

• Peginesatide is a dimeric peptide joined with a spacer linker to a pegylation chain to enhance its metabolic stability in vivo.

• No structural homology between peginesatide and erythropoietin. Antibodies against erythropoietin do not cross-react with peginesatide, and vice versa.

=> potential therapeutic use for pure red cell aplasia

Iain C. Macdougall, Am J Kidney Dis. 2012;59(3):444-

451

Peginesatide Phase 3 Clinical Trials Overview


HIF STABILIZATION

Regulation of hypoxia inducible factor (HIF) activity

HIF stabilization


HIF Stabilization

• Prolyl hydroxylase inhibitors Advantages –

1. Orally active 2. Modulate other genes involved in erythropoiesis in addition to the EPO gene.

Disadvantages – 1. Upregulate several hundred other hypoxia-sensitive genes, including those involved in glucose regulation, angiogenesis, such as VEGF (vascular endothelial growth factor) => enhance tumor growth or proliferative diabetic retinopathy ??

HIF Stabilization

• The first-generation HIF stabilizer molecule (prolyl hydroxylase inhibitors) -- FG 2216 => Patients developed abnormal liver enzyme test results, and one developed fatal hepatic necrosis in phase 2 clinical trial

• The second-generation HIF stabilizer molecule – FG 4592 => Significantly increase Hct and decrease serum Hepcidin levels => This is now in phase 2 clinical trial.

HEPCIDIN MODULATION

Hepcidin Modulation

* Hepcidin inhibits ferroportin, which controls iron efflux from duodenal enterocytes, hepatocytes, and macrophages

* Uremia is a chronic inflammatory state. Dialysis patients have much higher serum hepcidin levels.

Babitt JL, Molecular mechansms of hepcidin regulation: implications for the anemia of CKD. Am J Kidney Dis. 2010 (55):726-741

Hepcidin Modulation

• Strategies：Monoclonal antibody against hepcidin (NOX-H94) has

been shown the effect on inhibition of IL-6 induced anemia in mouse models.

Inhibition of the hepcidin production by using antisense oligonucleotides or sliencing messenger RNA transcribed from the hepcidin gene(HAMP)

• None of the strategies have been subjected to clinical trials.

=> Hepcidin has antimicrobial properties. Inhibition of hepcidin might exacerbate the risk of infection ?

GATA-2 INHIBITORS

• GATA-2 inhibitors: K-7174 and K-11706.


GATA-2inhibitor

Imagawa S. Negative regulation of the

erythropoietin gene expression by the GATA transcription

factors. Blood. 1997(89):1430-1439

GATA-2 Inhibitors

GATA-2 Inhibitors• K-11706 was found to

evoke greater hypoxic induction compared with K-7174, possibly through stimulation of HIF-1 binding activity in addition to GATA inhibition.

• Potential role for an orally administered GATA inhibitor in the treatment of anemia.

Imagawa S. Negative regulation of the erythropoietin gene expression by the GATA transcription factors. Blood. 1997(89):1430-1439

ERYTHROPOIETIN GENE THERAPY

Erythropoietin Gene Therapy

• In 2005, a group of Israeli scientists developed a functional delivery system for the EPO gene using skin cells on SCID mice (using adenovirus vector).

• The mice responded by producing increased levels of erythropoietin, and this was associated with an increase in hematocrit. No such effect was seen with the vector alone.

Erythropoietin Gene Therapy

Erythropietin Gene Therapy

• In 2010, a small group of patients with CKD in Israel have taken part in a proof-of-concept phase 1-2 clinical trial of this delivery system for the EPO gene.

• All patients showed increased erythropoietin production, with most showing sustained elevation of hemoglobin levels (the primary end point) in the target range of 10-12 g/dL for 6-12 months without receiving additional erythropoietin injections.

CONCLUSION

Conclusion

• It took us nearly 20 years to realize the limitations of ESAs and the potential for harm if used too aggressively.

• This review summarizes our current knowledge about a variety of new strategies for stimulating erythropoiesis.

• They will need to be subjected to the same degree of scientific investigation as the existing ESAs, and it may be many years before the true efficacy-safety balance of these novel scientific strategies is realized.

Clinical evaluation of resistance to erythropoietin stimulating agent in chronic kidney disease


Al toxicityPb toxicity

Cu deficiencyHypothyroidism

Thanks For You Attention !!

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E rythropoietin -stimulating agents resistance and new anemia therapies