Iron Overload and Iron Chelation: The Inside Story

Iron Overload and Iron Chelation:

The Inside Story

Jerry L. Spivak, MD

Professor of Medicine and Oncology

Johns Hopkins University School of Medicine

Baltimore, Maryland

Iron as a Prosthetic Group

• Oxygen transport - Hemoglobin, myoglobin

• Cell proliferation - Ribonucleotide reductase

• Electron transport -

Flavoproteins

• Respiratory enzymes - Cytochromes

• Oxidases - Catalase

Body Iron Stores (

♂

)

Hemoglobin 2.5 gm

Myoglobin/heme and nonheme 0.4 gm

enzymes

Ferritin/hemosiderin 1.0 gm(

2/1ratio

)

Transferrin 0.005 gm

“Tales From the Crypt”

Iron Absorption and the Mucosal Iron Block

Duodenum

Sugars,

amino acids

and Vitamin C

_Fe++

Plasma transferrin

Enterocyte precursor

(Macrophage)

Noniron-loaded

FPN

Enterocyte precursor

(Macrophage)

Iron-loaded

_DMT1

Hephaestin

FPN (

Hepcidin

)

Ferritin

Fe++ Fe+++

Other cells

Iron Balance in Adults

Gastrointestinal Absorption

1-2 mg/day

Physiologic daily iron loss

1-2 mg/day

Plasma transferrin

4 mg

Storage Iron

Liver cells and Macrophages

1000 mg

Functional iron

Bone marrow Red cell hemoglobin

Myoglobin Cytochromes

2500 mg

Spinach ,whole grains such as buckwheat and amaranth, other vegetables such as

chard and rhubarb, as well as beans and nuts, all contain significant levels of oxalic acid, which binds with iron, inhibiting its absorption. Soy beans contain phytic acid, which also bind iron. Tea and coffee contain tannins, which block iron absorption. Clay and

heavy metals also inhibit iron absorption.

Iron Absorption Enhancers

•Meat/fish/poultry

•Vitamin C-rich fruits: oranges, cantaloupe, strawberries, grapefruit

•Vegetables: broccoli, brussel sprouts, tomato, tomato juice, potato, green & red peppers

•White wine

Natural Modifiers of Iron Absorption

Iron Turnover in the Anemia of Chronic Disease

Gastrointestinal Absorption

<1 mg/day

Physiologic daily iron loss

1 mg/day

Plasma transferrin

2 mg

Storage Iron

Liver cells and RES

1500 mg

Functional iron

Bone marrow Red cell hemoglobin

Myoglobin Cytochromes

2000 mg

Hepcidin Hepcidin

Semin Liver Dis 31:280, 2011

Hepcidin Regulation*

Preservation of the MCHC at the Expense of the MCV

Iron Regulatory Proteins

• HFE

• Tfr-2

• Hemojuvelin

(HJV)

• Hepcidin

• DMT1

• Ferroportin (FPN)

• Tfr-1

• Hephaestin and

ceruloplasmin

• Senses cellular iron uptake

• Senses cellular iron uptake

• Upregulates Hepcidin (with Tfr-2

and HFE)

• Downregulates Ferroportin

• Imports GI iron

• Exports intracellular iron to Tf

• Receptor for Tf-bound iron

• Iron oxidases (cellular and

circulating)

Iron overload No anemia

Iron overload anemia

Essential Factors in Erythropoiesis and the Effect of

Cancer, Inflammation or Infection

•

Intensity of the stimulus

•

Functional capacity of the

bone marrow

•

Available nutrients

•

Red cell survival

•

Erythropoietin production is

suppressed by cytokines and

iron overload

•

Erythroid progenitor cell

proliferation is suppressed by

cytokines and erythropoietin

lack

•

Iron is sequestered and its

absorption is inhibited by

hepcidin

•

Red cell survival is reduced

and blood loss is increased due

to diagnostic testing

Role of Iron Sequestration in the Anemia of Chronic Disease

•

There is no impairment of utilization of absorbed iron

•

There is no impairment of plasma transferrin iron uptake by erythroid cells

•

Reduced transferrin receptor expression and decreased iron utilization are

primarily consequences of EPO deficiency

•

Iron therapy cannot correct the anemia of chronic disease in the absence

of tissue iron deficiency

•

Pharmacologic concentrations of EPO can correct the anemia of chronic

disease but not iron deficiency anemia

•

Correction of the anemia of chronic disease with EPO can occur without a

change in the serum iron abnormalities

•

Correction of the anemia of chronic disease with EPO is associated with

mobilization of iron stores and sometimes iron deficiency

Hypoxic Regulation of Erythropoietin Production

Serum Epo (mU/ml) Time (days) –2 0 2 4 6 8 10 12 0 1600 1400 1200 800 600 400 200 1000 Hgb, g/dL 10 9 7 6 5 8 sEpo, mU/mL Hb, g/dL Blood 91:2139, 1998

Effect of Iron Administration on the Serum Erythropoietin Level

J Clin Invest 110:1042, 2002

Hepcidin Expression is Subordinate to Tissue Hypoxia

Increased storage iron

Serum Immunoreactive Erythropoietin in Iron

Deficiency Anemia

Time (months)

Hemoglobin

gm

%

B J Haematol 94:288, 1996

Body Iron Homeostasis

• After intake, iron is normally sequestered in complexes:

– Serum transferrin

• Iron transport protein in blood/extracellular fluid

• Capacity can be exceeded resulting in Nontransferrin-Bound Iron

(NTBI)

– NTBI is the most toxic form of iron

– Ferritin

• Binds intracellular iron

• High levels in the serum reflect iron overload but can be affected by

NASH, inflammation (Still’s disease and cancer) infection

(hematophagocytosis, hepatitis)

• A transferrin saturation < 45 % with a high serum ferritin (>400

ng/mL) is characteristic of inflammation or liver disease, not iron

overload, where the transferrin saturation is always > 50 % and often

> 95 %

Iron excess as a Toxin

Increased transferrin saturation (>50 %) leads to deposition of iron in

nonerythroid tissues such as the heart, liver and pancreas leading to:

Congestive heart failure

Hepatic fibrosis

Diabetes mellitus and other endocrinopathies

Increased susceptibility to infection

Increased transferrin saturation leads to the accumulation of

nontransferrin-bound iron (NBTI), labile or bound to other proteins, and

free radical formation.

The generation of free hydroxyl radicals causes tissue damage

through oxidative reactions with proteins, lipids and nuclei acids.

Int J Hematol 76:219, 2002

Measures of Body Iron Content

• Serum iron

• Serum transferrin

• Serum ferritin

• Liver biopsy

• Liver iron by MRI

• Cardiac MRI (T2*)

• Bone marrow aspirate

• Diurnal variation

,

affected by diet

inflammation and infection

• Affected by nutrition, liver disease,

inflammation, infection and FPN and

ceruloplasmin mutations

• An acute phase reactant, specific only

if low. Correlation with LIC = 0.63

• Defines iron storage site and liver

histology. May not correlate with

cardiac iron burden

• May not correlate with cardiac iron

status

• Correlates with cardiac function

• Considered the “gold standard” but

is invasive and not always technically

adequate

Correlation between Plasma Ferritin (Pl Fer) and Hepatic Iron Concentration

Am J Hematol 42:81, 1993

R = 0.63 Months of chelation therapy Pl Fer

LIC

World J Gastroenterol 12:5866, 2006

Hereditary (HFE) Hemochromatosis

• An autosomal recessive disorder due to a C282Y mutation and rarely an H63D mutation in Northern Europeans (0.3-1.2 % prevalence) with variable penetrance • Serum ferritin is elevated in 84 % of men and 65 % of female C282Y homozygotes • Serum ferritin is > 1000 µg in 37 % of men and 3% of female C282Y homozygotes • If baseline ferritin is < 1000 µg , < 50 % of men and 20 % of females exceeded 1000

µg after 12 years

• If baseline ferritin is < 1000 µg at age 55, < 15 % progressed to > 1000 µg in 12 years

• Iron overload was present in 28 % of men and 1 % of women at age 65.

• Iron overload with C282Y/H63D is rare without other risk factors such as liver disease

• C282Y homozygosity doubles the colon cancer risk in everyone and the breast cancer risk in women’

• H63D homozygosity triples the hereditary nonpolyposis colon cancer risk • Environmental cofactors are alcohol, hepatic steatosis and viral hepatitis • Non-citrus fruits are protective

• An elevated transferrin saturation (> 45 %) is the earliest clue

• Phlebotomy should start if the ferritin is if there is evidence of iron overload to achieve a ferritin of 50 µg

Major Complications of Iron Overload

• Cirrhosis

• Hepatic fibrosis (reversible with phlebotomy) • Hepatocellular cancer • Diabetes mellitus • Arthritis • Cardiomyopathy • Hypogonadism • Hypothyroidism

• Pituitary-adrenal axis impairment

• Increased susceptibility to infection (MDS)

• Increased risk of leukemic transformation (MDS) • Impaired survival post BMT

Circulation 30: 698,1964

Lancet 334:27, 1989

Am J Hematol 84:29, 2009

Survival in MDS According to Diagnosis (WHO criteria)

Overall survival

(HR = 1.91; p < 0.001)

Leukaemia-free survival (HR = 1.84; p = 0.001)

Survival of MDS patients by transfusion

dependence (N = 467)

180

Cumulative proportion surviving

Transfusion independent Transfusion dependent 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 0 20 40 60 80 100 120 140 160

Cumulative proportion surviving

0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 0 20 40 60 80 100 120 140 160 180

Survival time (months) Survival time (months)

Transfusion independent Transfusion dependent

Malcovati L, et al. J Clin Oncol. 2005;23:7594-603. HR = hazard ratio.

Survival of MDS patients by severity of

transfusion requirement

Survival time (months)

0.2 0.4 0.6 0.8 1.0

Survival time (months)

0.0 0.2 0.4 0.6 0.8 1.0 0 20 40 60 80 100 120 140 160 180 Cumulative survival 0 U pRBC/4 weeks 1 U pRBC/4 weeks 2 U pRBC/4 weeks 3 U pRBC/4 weeks 4 U pRBC/4 weeks

Malcovati L, et al. Haematologica. 2006;91:1588-90.

0 U pRBC/4 weeks 1 U pRBC/4 weeks 2 U pRBC/4 weeks 3 U pRBC/4 weeks 4 U pRBC/4 weeks Cumulative survival

pRBC = packed red blood cells.

Overall survival (HR = 1.36; p < 0.001) Leukaemia-free survival (HR = 1.40; p < 0.001) 180 0 20 40 60 80 100 120 140 160 0.0

RA/RARS/5q−

(HR = 1.42; p < 0.001)

RCMD/RCMD-RS (HR = 1.33; p = 0.07)

Malcovati L, et al. Haematologica. 2006;91:1588-90.

Overall survival of transfusion-dependent patients by

serum ferritin level

180

Cumulative proportion surviving

0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 0 20 40 60 80 100 120 140 160

Cumulative proportion surviving

0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 0 20 40 60 80 100 120 140 160 180

Survival time (months) Survival time (months)

Serum ferritin 1,000 µg/L 1,500 µg/L 2,000 µg/L 2,500 µg/L Serum ferritin 1,000 µg/L 1,500 µg/L 2,000 µg/L 2,500 µg/L

RA = refractory anaemia; RARS = RA with ringed sideroblasts; RCMD = refractory cytopenia with multilineage dysplasia;

Blood 109:4586, 2007

Prevalence of comorbidities in

transfusion-dependent MDS

Transfused MDS patients have a higher prevalence of cardiac events, diabetes mellitus, dyspnoea, and hepatic and infectious diseases than non-transfused MDS patients

Goldberg SL, et al. J Clin Oncol. 2010;28:2847-52.

82.4 44.4 62.9 1.0 81.0 14.6 67.1 37.1 40.4 0.7 55.7 6.2 0 50 100 Cardiac events 2003–2005 Diabetes 2003–2005 Dyspnoea 2003–2005 Hepatic events 2003–2005 Infectious complications 2003–2005 Fungal infection 2003–2005 Patients (%) With transfusion (n = 205) Without transfusion (n = 307)

Probability of non-leukemic death in

transfusion dependent MDS patients

51 31 8 8 2 0 25 50 75 100 Cardiac failure

Infection Haemorrhage Hepatic cirrhosis Other P e rcen ta g e N = 467 p = 0.01

Current Guidelines for Iron Chelation in MDS Patients Organization Transfusion Status (units of blood) Serum Ferritin (ng/mL) Life Expectancy/ MDS Risk Score Italian Hematology Society > 50 units – > 6 months UK Hematology Society

≥ 25 units – Low/ Int-1

NCCN > 20-30 units > 2500 Low/Int-1 SCT MDS Foundation ≥ 24 units > 1000 > 1year Austrian Hematology Society Transfusion-dependent > 2000 or organ damage 2years/SCT/ Chemotherapy Canadian Hematology Society Transfusion-dependent > 1000 or organ damage Low/Int-1/SCT Int-2 if > 1year life expectancy

Japanese Hematology Society

Blood 34:441, 1969

Iron Promotes the Growth of Candida

PLoS One 6:e23109, 2011

What about higher risk MDS and AML progression?

• Iron is mutagenic in hematopoietic cells and can promote

progression to AML in mice

• NTBI



LPI



ROS*

– ROS damage

X

Mutagenesis

Apoptosis

ROS = reactive oxygen species, NTBI = non-transferrin bound iron

Genomic instability

AML?

●

Chelation induced apoptosis, differentiation & repressed signalling in

AML cells & cell lines in vitro & in vivo

1. Chan LSA, et al. Blood 2010;116:[abstract 122] 2. Eberhard Y, et al. Blood. 2009;114:3064-73. 3. Jiang Y, et al. Leukemia. 2005;19:1239-47. 4. Ohyashiki JH, et al. Cancer Sci. 2009;100:970-7. 5. Callens C, et al. J Exp Med. 2010;207:731-50.

Currently Available Iron Chelation Agents

Deferoxamine Deferiprone Deferasirox

Usual dose (mg/kg/day) 25−60 75 20−30

Route of administration Subcutaneous or intravenous, Oral, three times daily Oral, once daily 8−12 hours, 5 days/week

Half-life 20−30 minutes 3−4 hours 8−16 hours

Route of excretion Urinary and fecal Urinary Fecal

Main adverse effects Local reactions, ocular and Gastrointestinal disturbances, Gastrointestinal disturbances, auditory abnormalities, growth agranulocytosis/neutropenia rash, mild non-progressive retardation, allergic reaction arthralgia, elevated liver enzymes creatinine increase, elevated

liver enzymes

Leuk Res 31(S3):S16, 2007

J Cardio Mag Res 13: 45, 2011

B J Haematol 136:501, 2007

Deferasirox Normalizes LPI in MDS Patients

1. List AF, et al. Blood. 2008;112:[abstract 634]. 2. Gattermann N, et al. Leuk Res. 2010;34:1143-50.

Patients, n 55 38 39 37 34

Patients with baseline LPI ≥ 0.5 μmol/L = 41% Threshold of normal LPI (≤ 0.5 µmol/L)

Mean LPI (µmol/L)

0 0.2 0.4 0.6 0.8 1.0 1.2 BL 3 6 9 12

Months from baseline

p 0.00001*

*Comparison of baseline LPI vs each treatment time point

Br J Haematol 94:288, 1996

ERYTHROID RESPONSE DURING IRON CHELATION

EPIC: reduction in serum ferritin is

associated with improvement in ALT in MDS

• At 12 months, there were significant reductions in

– median serum ferritin (

−

253 µg/L; p = 0.002)

– mean ALT (

−

27.7 ± 37.4 U/L; p < 0.0001)

0 500 1,000 1,500 2,000 2,500 3,000 3 6 9 12 Baseline 3 6 9 12 Time (months) Median serum ferritin (µg/L) Mean AL T (U/L) ALT Serum ferritin ALT

Mean actual deferasirox dose: 19.2 ± 5.4 mg/kg/day 0 10 20 30 40 50 60 70

ALT = alanine transaminase; EPIC = European

Survival in chelated versus non-chelated MDS patients with (a) transfusion requirements <3 PRBC/month and (b) transfusion requirements >3 PRBC/month.

Leukemia Res 34:864, 2010

IPSS = Low

Median: not reached vs 69 months (p < 0.002) Survival distribution function 0.00 0.25 0.50 0.75 1.00

Time from diagnosis to death (months)

0 50 100 150 200 250

Iron chelation (n = 30)

No chelation (n = 15) Survival distribution

function 0.00 0.25 0.50 0.75 1.00

Time from diagnosis to death (months)

0 20 40 60 80 100 120 140

No chelation (n = 29)

IPSS = Int-1

Median: 115 vs 50 months (p < 0.003)

Results were the same regardless of sex and age.

Rose C, et al. Leuk Res. [Epub ahead of print 2010 Feb 1].

Iron chelation (n = 23)

GFM: effect of iron chelation therapy on survival in

lower-risk MDS patients

NTBI elevation during allogeneic SCT

Sahlstedt L, et al. Br J Haematol. 2001;113:836-8.

0 20 40 60 80 100 120 140 −14 −7 0 7 14 21 Tr ansferrin saturation (%)

Time from SCT (days)

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1.1 0 20 40 60 80 100 120 140 160 NTBI ( μ mol/L)

C = onset of conditioning regimen.

0 20 40 60 80 100 120 140 −14 −7 0 7 14 21 Transferrin saturation (%)

Lee JW, et al. Bone Marrow Transplant. 2009;44:793-7. SF > 1,000 SF < 1,000 IC p = 0.001 0.2 0.4 0.6 0.8 1.0 0 0 12 24 36 48 60 SF > 1,000 SF < 1,000 IC p = 0.000 0 12 24 36 48 60 0.2 0.4 0.6 0.8 1.0 0 SF > 1,000 SF < 1,000 IC p = 0.003 0 12 24 36 48 60 0.2 0.4 0.6 0.8 1.0 0

Overall survival rate Event-free survival Treatment-related mortality

rate

Months from transplantation Months from transplantation Months from transplantation

SF > 1,000 = patients with serum ferritin ≥ 1,000 µg/L at the time of SCT;

SF < 1,000 = patients with serum ferritin < 1,000 µg/L at the time of SCT, without ICT; IC = patients with serum ferritin decreased to < 1,000 µg/L with ICT before SCT.

Iron Chelation Before SCT Improves Survival

(n = 101)

Adverse event*

Number (%)

Diarrhoea

111 (32.6)

Nausea

45 (13.2)

Vomiting

26 (7.6)

Abdominal pain

26 (7.6)

Upper abdominal pain

25 (7.3)

Rash

23 (6.7)

Constipation

21 (6.2)

Total number

341

*Drug-related as assessed by the investigator.

EPIC Study: Adverse Events with Deferasirox

Side effect n %

Gastrointestinal symptoms (abdominal discomfort, pain, nausea, vomiting, diarrhea) 18 37.5

Granulocytopenia (neutrophils = 0.5–1.0 × 109/L)

5 13.0

Agranulocytosis (neutrophils< 0.5 × 109/L) 2 4.0

Elevation of liver enzymes (>3 × upper normal limits) 9 18.8

Weight gain, fluid retention 2 4.0

Most Frequent Side Effects of Deferiprone Therapy in Myelodysplastic Syndrome Patients

Correlation of Anemia and Survival in the Elderly

Men Women

Haematologica 2011

Haematologica 2011

Probability of Cardiac Death in MDS According to the Hemoglobin Level

EPO/G-CSF Control

Improved Overall Survival in MDS with EPO/G-CSF Therapy

Summary

• Iron overload is associated with impaired survival and an increased risk of leukemic transformation in MDS patients with Low or Int-1 disease

• Risk assessment based on iron stores is imperfect because of poor correlation between the transfusion burden and body iron store measurements

• Iron chelation therapy can reduce total body iron stores and NTBI

• Iron chelation is associated with improved survival as well as an improved response to bone marrow transplantation in MDS patients

• Oral chelation may be more effective than parenteral chelation in reducing

intracellular cardiac iron and combining oral and parenteral chelation may be the most effective strategy

• Anemia per se contributes to iron overload and is also an important cofactor for disease morbidity in iron-overloaded MDS patients

• Current transfusion practice needs to be altered to maintain the hematocrit

commensurate with continuous relief of tissue hypoxia on a gender-specific basis • The threshold for initiating iron chelation should be based on organ dysfunction

Gold is for the mistress, silver for the maid,

copper for the craftsman cunning at his trade

but iron said the Baron sitting in his hall, iron,

cold iron is the master of them all.

Which

of

these

Individuals

is

Tebowing?

A

B

C