•required for life hemoglobin cytochromes DNA repair catalase

•iron fortification

•excess dangerous oxidative stress ‘food’ for invaders cancer heart disease

•give blood?

Fe++ Fe+++

Iron

too littletoo little anemiaanemiamuch of the 20th century spent trying get iron into

people

too muchtoo much cancercancer??

since mid-1970s, growing concerns about too much

iron

William Crosby(1914 - 2005)

•WWII veteran - Bronze star

•pioneer of modern hematology

•1951: established hematology and oncology specialties at Walter Reed Army Hospital

•over 450 publications

““The safety of iron-fortified food”The safety of iron-fortified food”JAMA 1978.JAMA 1978.

Biological Rationale

•Iron as food to a cancer cell iron is an essential nutrient animal experiments

•Iron and oxidative stress DNA damage iron as radiosensitizer

CC CC

CCCC

C

C

N

Normal cell Cancer cellCancer

O + H O2 2 2

•OHFeFe++++

FeFe++++

FeFe++++

carcinogencarcinogen foodfood

++++FeFe

CFeFe++++

DNNA

Stevens & Kalkwarf, Stevens & Kalkwarf, Environ Environ Health PerspecHealth Perspec, 87:291–300, , 87:291–300, 19901990

Animal Experiments

•Breast diet - Thompson HJ, Carcinogenesis, 12:111, 1991 diet - Hann H, Cancer Res, 48:4168, 1988

•Lung diet - Yano T, Cancer Letters, 76:57, 1994

•Liver injection - Smith AG, Carcinogenesis, 11:437, 1990

•Colon diet - Siegers CP, Cancer Letters, 65:245, 1992

Relevant Iron

•High iron content of blood and interstitial fluid in a form that can be utilized by cancer cells; e.g., high transferrin saturation

•High intracellular iron content in a form that can engage in pro-oxidant reactions; e.g., with DNA, ferritin, and non-specific iron complexes such as ATP•Fe++

Stevens et al., Stevens et al., N Engl J MedN Engl J Med, 319:1047-52, 1988, 319:1047-52, 1988

Potentially Reactive Iron in Cells

•Ferritin Whiting et al., Cancer Res, 41:1628, 1981 Nelson & Stevens, Cell Prolif, 25:579, 1992

•DNA-iron complexes Loeb et al., PNAS, 85:3918, 1988 Imlay & Linn, Science, 240:1302, 1988

•Other complexes, e.g., ADP•Fe++

Floyd, Arch Biochem Biophys, 225:263, 1983

Graf et al., J Biol Chem, 259:3620, 1984

Syllogism:Syllogism:

IF: IF: high body iron increases cancer riskhigh body iron increases cancer risk

THEN: THEN: cancer should be associated with cancer should be associated with high serum ferritin and transferrin high serum ferritin and transferrin saturation and low TIBC and serum saturation and low TIBC and serum transferrin prior to development of diseasetransferrin prior to development of disease

IN GENERAL: IN GENERAL: high body iron is reflected in high body iron is reflected in high serum ferritin and transferrin high serum ferritin and transferrin saturation and low TIBC and serum saturation and low TIBC and serum transferrintransferrin

EpidemiologyEpidemiology

Blumberg:‘Australia antigen’ led to discovery of HBV

Gajdusek:Kuru disease in New Guinea led to discovery of Prions

R. Palmer Beasley

Taiwan studies of HBV

• study begun in 1975• male government workers aged 40 to 59 initially• funded by NIH and by Taiwan National Science Council

co-factors: why do some HBV carriers get PHC and not others?

Cancer Risk in NHANES IMen and Women Combined

Stevens et al., Stevens et al., NEJMNEJM, 319:1047, 1988; , 319:1047, 1988; Int J CancerInt J Cancer, 56:364, 1994, 56:364, 1994

Cancer MortalityMen and Women Separately

Moderate Iron Excess

U.S. and Finland

Knekt et al., Knekt et al., Int J CancerInt J Cancer, 56:379, 1994, 56:379, 1994

real?real?

Breast Cancer Study from Hiroshima

•Nested case-control design

•Interactions among 3 factors: Estradiol (Kabuto et al., CEBP,

9:575,2000) Radiation dose (Land et al., JNCI, 65:353,

1980) Iron (Stevens et al., Radiat Res,

153:844, 2000)

•Known risk factors

dropped at 8:15 in the morning, August 6, 1945 - clear skies, a nice day in Hiroshima

exploded 1 minute later at about 500 meters above the city

3 km circle

Nested Case-control Study of Body Iron and Breast Cancer

Rick in Japanese Atomic Bomb Survivors

•do women with higher body iron have greater risk of breast cancer?

•are women with higher body iron more sensitive to radiation for causing breast cancer?

•what is ‘body iron’?

Nested Case-control Study

THE PAST THE PRESENT

save: biological samples, medical

info., questionnaire

Population

Cohort

cases

controls

assay samples

calculate odds ratio

sample from risk set

find and identify

Syllogism:Syllogism:

IF: IF: high body iron increases breast high body iron increases breast cancer riskcancer risk

THEN: THEN: women with breast cancer should women with breast cancer should have higher serum ferritin than women have higher serum ferritin than women without breast cancerwithout breast cancer

IN GENERAL: IN GENERAL: high body iron is reflected high body iron is reflected in high serum ferritin in high serum ferritin

EpidemiologyEpidemiology

Life Span Study

94,000 survivors and 27,000 unexposed (out of city on 94,000 survivors and 27,000 unexposed (out of city on August 6)August 6)

Adult Health Study

20,000 enrolled in AHS for annual physical 20,000 enrolled in AHS for annual physical exams; exams;

blood samples taken every two years beginning blood samples taken every two years beginning in 1969in 1969

Risk Setcohort study of breast cancer and body iron (serum ferritin) begun in 1970 which saved blood sample from 10,000 females will do nested case-control study of cases up to 2001

9,350 still in cohort 7/15/84 - but only 52 cancer-free

women aged 57

on 7/15/1984, a 57 yo woman is diagnosed

select 2 or 3 of the 52 randomly as control

1970 2001

10,000 8,300 6,100

year

persons in cohort

1990

Risk Set

•retrieve the blood sample from storage for the 57 year old case and her control

•assay sample for ferritin

•repeat this process for all cases which occur up until 2001

•analyze data – Logistic regression

back to genetic iron

overload conditions

Editorial, Nature Genetics, August, 1996

At a small meeting in the Wasatch mountains in the late 1970s, there was discussion of an iron-overload disease gene associated with an HLA region on chromosome 6.

“After months of further discussion, Botstein, Davis, Skolnick and Ray White proposed the framework for a comprehensive genetic map, culminating in 1980 in their classic publication.”

Hemochromatosis “…is now recognized as one of the most common single-gene hereditary disorders [among EuroAmericans]…with a remarkable carrier frequency of 1 in 10”

Hereditary Hemochromatosis

•HH results in morbid iron overload

•HH afflicts 1 in 200-400 individuals of Northern European descent

•1 in ~10 are carriers (i.e., heterozygotes)

•HFE - chromosome 6p

•two missense alterations, homozygous in 83% of 178 HH patients

Feder et al., Feder et al., Nature GeneticsNature Genetics, 13:399-408, 1996, 13:399-408, 1996

Cancer in Hemochromatosis

•Bradbear et al. (JNCI, 1985) 208 subjects 16 liver cancers - 0.07 expected 4 colorectal cancers - 1.8

expected

•Niederau et al. (NEJM, 1985) 163 subjects 16 liver cancers - 0.07 expected 7 “other” cancers - 5.3 expected

Hemochromatosis(based on pedigree HLA linkage)

transferrin saturation

normal heterozygotes homozygotesnormal heterozygotes homozygotes

males 32.8 % 42.6 %males 32.8 % 42.6 % 93.0 %93.0 %

females 30.2 % 40.1 %females 30.2 % 40.1 % 83.7 %83.7 %

Bulaj et al., Bulaj et al., N Engl J MedN Engl J Med, 335:1799-805, 1996, 335:1799-805, 1996

conventional wisdom:this can’t be a problem

conventional wisdom:this is a problem

Iron Overload in Africa

(Interaction between a Gene and Dietary Iron Content)

•traditional beer brewed in iron pots (Zimbabwe)

•propensity for iron-overload from beer drinking follows Mendelian pattern, however:

•not HLA-linked (i.e., not HFE)

•estimate that 3 to 4% homozygotes, ~30% heterozygotes

Gordeuk et al., Gordeuk et al., N Engl J MedN Engl J Med, 326:95, 1992, 326:95, 1992

Ferroportin

•chromosome 2q32

•Q248H polymorphism associated with elevated ferritin in Africans and in African Americans

•also associated with elevated serum C-Reactive Protein

Rivers CA, et al., Rivers CA, et al., Blood Cells, Mol DisBlood Cells, Mol Dis, 38:247, 2007, 38:247, 2007McNamara L, et al. McNamara L, et al. J Gastro HepatolJ Gastro Hepatol, 20:1855, 2005, 20:1855, 2005

what about iron in cancer prognosis?

Iron and breast cancer prognosis

•publically available microarray data on 674 breast cancer cases

•divided into ‘test’ and ‘training’ groups

•‘fooled around’ in the ‘training’ group to find best predictor of 5-year survival

•16 gene ‘Iron Regulatory Gene Signature’

•applied to ‘test’ group

•strongly associated with survival after adjustment for other known prognostic indicators

cites the epidemiology

Frank Torti

Suzy Torti