David Hart Dec 13, 2006

HEME CH 3 -

Bonkovsky ASH Education Book December 2005

Hentze, Muckenthaler & Andrews Cell, Vol 117, , April 30, 2004 Hepcidin

Lecture Outline Heme function Heme synthesis and regulation Iron metabolism Porphyrias Heme degradation

Disorders of Heme Synthesis X-linked Sideroblastic Anemia Lead Poisoning Iron Deficiency Anemia The Porphyrias

Porphyrias Inherited defects in heme synthesis –Accumulation and excretion of porphyrins –Pattern depends on which enzyme affected –Decreased heme synthesis derepresses hepatic  ALAS Most are Autosomal Dominant Erythropoietic, Hepatic or Mixed Acute and Chronic –Acute: Neurovisceral attacks Porphyrin accumulation: Photosensitivity –Formation of reactive oxygen species –Damage tissues, Release lysosomal enzymes

Fuseli Three Witches Tate Gillray 1791 Weird Sisters Tate

Very Rare Recessive Porphyria Lead Poisoning ALA-D Porphyria X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia

Acute Hepatic PBG and  ALA (Neurotoxic) Accumulate in Urine PBG in Urine: Diagnostic Screen Neurovisceral Attacks No Photosensitivity with AIP Hydroxymethylbilane Synthase Lead Poisoning X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia ALA-D Porphyria

Acute Porphyrias Clinically indistinguishable (Locus Heterogeneity) PBG in Urine: Diagnostic Screen Agents which induce cytochrome P450 –Drugs, Alcohol, Hormones Precipitated by fasting, treated with glucose After puberty; more in women Begin with minor behavioral changes Proceeds to autonomic and sensomotoric neuropathy; Convulsions Pain: Back, Extremities, Abdomen Hypertension and Tachycardia Arrhythmias; cardiac arrest

Acute Hepatic Erythropoietic RECESSIVE Porphyrin accumulation: Photosensitivity Porphyrins are Fluorescent compounds Formation of reactive oxygen species, Activate Complement Lead Poisoning Hydroxymethylbilane Synthase ALA-D Porphyria X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia

Clin Med 2005:5 Dr. Meyer-Betz 1912

GM Murphy, Dermatologic Therapy, March 2003 CEP

NEJM 9/7/2006

mccaskey4.home.mindspring.com

Erythropoietic Acute Hepatic Chronic Lead Poisoning Hydroxymethylbilane Synthase ALA-D Porphyria X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia

Porphyria Cutanea Tarda (PCT) Most common Porphyria –80% sporadic Hepatic and Erythropoietic Photosensitivity Uroporphyrin accumulates in Urine –Red-Brown in natural light Clinical expression in 4th - 5th decade Decrease in UROD activity by Iron-dependent mechanism –Alcohol, viruses, drugs, hormones –HFE Hemochromatosis Venesection, Chloroquine

Autosomal Dominant PCT (Hepatoerythropoietic Porphyria) Hepatic UROD activity < 50% during symptoms Additional decrease from reversible inactivation C282Y HFE causes earlier onset

PCT;

Erythropoietic Acute Hepatic Acute Hepatic Photosensitivity (Unlike AIP) Lead Poisoning Hydroxymethylbilane Synthase Chronic ALA-D Porphyria X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia

medlib.med.utah.edu Normal Liver

Granular, Dark Reddish Brown Surface of Liver in Hemochromatosis

Lecha, Herrero, Ozalla, Dermatologic Therapy, March 2003 Hepatic Porphyria

Acute Hepatic Erythropoietic Acute Hepatic Acute Hepatic Photosensitivity Lead Poisoning Hydroxymethylbilane Synthase Chronic ALA-D Porphyria X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia

Hair Analysis Lancet July 2005

Lancet July 23-29, 2005 King George III ( ) Likely diagnosis of Variegate Porphyria –Proposed 1969 based on family tree Lock of hair showed high lead –Widespread use in his era Extremely high levels of arsenic –Likely secondary to medications

Lancet July 2005 Color of Urine “Alicante Wine”

Introduction of Fe 2+ into PPIX Occurs spontaneously, but Enhanced by FERROCHELATASE An enzyme which is inhibited by LEAD

Acute Hepatic Acute Hepatic Erythropoietic Acute Hepatic Erythropoietic Photosensitivity Lead Poisoning Hydroxymethylbilane Synthase Chronic ALA-D Porphyria X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia

Erythropoietic Protoporphyria Presentation in early childhood Burning, stinging pain with sunlight Subsequent skin changes Expression requires low-expression allele in trans –10% of population of France and UK –IVS3-48 alternative splice acceptor –With AD mutation FECH 35% of normal –Homozygosity does not cause disease Beta carotene: free radical scavenger

GM Murphy, Dermatologic Therapy, March 2003 EPP

GM Murphy, Dermatologic Therapy, March 2003 EPP

Acute Hepatic Acute Hepatic Erythropoietic Acute Hepatic Erythropoietic Photosensitivity Lead Poisoning No Photosensitivity With Lead Lead Poisoning Hydroxymethylbilane Synthase Chronic X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia ALA-D Porphyria

Acute Hepatic Acute Hepatic Erythropoietic Acute Hepatic Erythropoietic Photosensitivity Iron Deficiency “Free” Erythrocyte PPIX accumulates in Lead Poisoning and Iron Deficiency Lead Poisoning Hydroxymethylbilane Synthase Chronic X-linked Sideroblastic Anemia X-linked Sideroblastic Anemia ALA-D Porphyria

Porphyrias: Genetics / Epigenetics 5 out of 7 are Low-penetrance Autosomal Dominant Most mutations are restricted to one family Rare Homozygotes very severe No dominant negative mutants described 50% residual activity is normally sufficient

Molecular basis of “low penetrance” Genotype/phenotype correlations Increased demand –Fasting (low Glucose) –Cell, August 26, 2005 Low expression allele in trans Iron / HFE hemochromatosis can directly inhibit enzymes Other epigenetic phenomena

Treatment Medical Support during acute attacks Treatment for pain and vomiting Glucose infusion until Hemin available Intravenous Hemin –Decreases synthesis of  ALAS Avoid Sunlight  -carotene, a free-radical scavenger Chronic transfusion for Erythropoietic

Degradation of Heme At end of their 120 day lifespan, red blood cells are taken up and degraded by the reticuloendothelial (RE) system (liver and spleen) 85% heme for degradation from RBC 15% immature RBC, cytochromes from extraerythroid tissues

N NHN HN H 3 C- -CH=CH 2 -CH 3 CH 2 COOH CH 2 COOH CH 3 Fe 2+ CH=CH 2 HEME

N NHN HN H 3 C- -CH=CH 2 -CH 3 CH 2 COOH CH 2 COOH CH 3 Fe 2+ CH=CH 2 MACROPHAGE takes up HEME Heme Oxygenase is Inducible by a variety of agents Inhibited by Tin Protoporphyrin

N NHN HN H 3 C- -CH=CH 2 -CH 3 CH 2 COOH CH 2 COOH CH 3 CH=CH 2 MACROPHAGE Heme Oxygenase Step 1 NADPH O 2 Fe 3+ OH

N NHN HN H 3 C- -CH=CH 2 -CH 3 CH 2 COOH CH 2 COOH CH 3 CH=CH 2 MACROPHAGE Heme Oxygenase Step 2 Fe 3+ CO Released OO BILIVERDIN HO also has Cytoprotective effects

J Cell Mol Med 2006

NHNH CH V M N PM NHNH MP NHNH O VM O BILIVERDIN

NHNH CH V M N PM NHNH MP NHNH O VM O NHNH V M NHNH CH 2 PM NHNH CH MP NHNH O VM O BILIVERDIN BILIVERDIN REDUCTASE + NADPH BILIRUBIN

ALBUMIN Low Albumin Anionic Drugs: Salicylates Sulfonamides BILIRUBINALBUMIN Unbound Bilirubin Can enter CNS Cause Kernicterus in Neonate Bilirubin Released from Macrophage, binds to Albumin in the Plasma

BILIRUBINALBUMIN LIVER BILIRUBIN LIGANDIN CONJUGATION with 2 Molecules Glucuronic Acid from UDP-glucuronic acid Bilirubin Glucuronyltransferase Bilirubin Diglucuronide BILE Active Transport

BILIRUBIN: Unconjugated (Indirect) LIVER CONJUGATION Bilirubin Glucuronyltransferase Bilirubin Diglucuronide: Conjugated (Direct) Bilirubin VAN DEN BERGH COLORMETRIC REACTION TOTAL BILIRUBIN: Soluble in Methanol Less Soluble in Aqueous Solution, reacts more slowly More Soluble, reacts more quickly

GALL BLADDER Active Transport Bilirubin Diglucuronide LIVER Bilirubin Diglucuronide UROBILINOGENSTERCOBILIN KIDNEY BILE INTESTINAL BACTERIA Urobilinogen Enterohepatic Circulation UROBILIN Bilirubin

GALL BLADDER Active Transport Bilirubin Diglucuronide LIVER Bilirubin Diglucuronide UROBILINOGENSTERCOBILIN KIDNEY BILE INTESTINAL BACTERIA Urobilinogen Enterohepatic Circulation UROBILIN BILIRUBIN HEMOLYSIS: Unconjugated (Indirect) Hyperbilirubinemia

GALL BLADDER Active Transport Bilirubin DG LIVER Bilirubin Diglucuronide UROBILINOGENSTERCOBILIN KIDNEY BILE INTESTINAL BACTERIA Urobilinogen Enterohepatic Circulation UROBILIN BILIRUBIN NEONATAL JAUNDICE: Unconjugated Hyperbilirubinemia

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GALL BLADDER Active Transport Bilirubin Diglucuronide LIVER KIDNEY BILE INTESTINAL BACTERIA UROBILIN Bilirubin OBSTRUCTIVE JAUNDICE Direct (Conjugated) Bilirubin

GALL BLADDER Active Transport Bilirubin DG LIVER Bilirubin Diglucuronide UROBILINOGEN STERCOBILIN KIDNEY BILE INTESTINAL BACTERIA Urobilinogen Dark Urine BILIRUBIN HEPATOCELLULAR JAUNDICE: Unconjugated Hyperbilirubinemia ENTEROHEPATIC CIRCULATION

Carotenemia