1. Indications for Successful Iron Overload Treatment and Monitoring: Other Anaemias Adlette Inati, MD Head, Division of Paediatric Haematology-Oncology Medical Director, Children's Center for Cancer and Blood Diseases Rafik Hariri University Hospital Beirut, Lebanon

2. Diamond Blackfan Anaemia A rare red cell aplasia commonly associated with physical anomalies and rarely with cancer predisposition 1,2 Diagnosis: based on clinical grounds, presence of hypoproductive normocytic or macrocytic anaemia, elevated erythrocyte ADA activity, and mutations of small (RPS19, RPS24, and RPS17) and large (Rpl35) ribosomal subunit genes 1-5 Treatment: corticosteroids, blood transfusion, and bone marrow transplantation 1,2,6,7 1. Halperin DS, Freedman MH. Am J Pediatr Hematol Oncol. 1989;11:380-394. 2. DBAR registry of NA. www.dbar.org.www.dbar.org 3. Draptchinskaia N, et al. Nat Genet. 1999;21:169-175. 4. Choesmel V, et al. Hum Mol Genet. 2008;17:1253-1263. 5. Farrar JE, et al. Blood. 2008;112:1582-1592. 6. Ball SE, et al. Br J Haematol. 1996;94:645-653. 7. Willig TN, et al. Pediatr Res. 1999;46:553-561.

3. 1. Kutler DI, et al. Blood. 2003;101:1249-1256. 2. Fanconi Anemia Research Fund. About Fanconi anemia. 2007. http://www.fanconi.org/aboutfa/FA.htm. 3. Janov AJ, et al. Medicine. 1996;75:77-78. Fanconi’s Anaemia An autosomal recessive bone marrow failure syndrome associated with physical anomalies and a risk for malignancy 1,2 Diagnosis: Based on clinical background, presence of chromosomal breaks and radial chromosomes and Fanconi’s anaemia genes (13 identified so far) 2 Therapy: Supportive, androgens, stem cell transplantation, and treatment of malignancies 2 Median survival: 38 years and can be extraordinarily variable 3

4. Sideroblastic Anaemia A heterogeneous group of disorders characterized by microcytic hypochromic anaemia and ringed sideroblasts in the bone marrow 1 3 forms: hereditary, acquired, and idiopathic 1 Reported mutations: erythroid-specific 5-aminolevulinate synthase (ALAS2) gene, ABC7 transporter gene, SLC19A2 gene, and PUS1 gene 1,2 Diagnosis: based on the presence of hypochromic microcytic anaemia, increased serum transferrin saturation, transferrin and serum ferritin levels, decreased free erythrocyte protoporphyrin level (in X-linked forms), ringed sideroblasts at the late, nondividing erythroblast stage (diagnostic hallmark), and increased iron in bone marrow macrophages 2 Therapy: vitamin B6 (pyridoxine) replacement, removal of the offending agent, red blood cell transfusions and chelation, treatment of malignancies 1 1. Iron Disorders Institute. Sideroblastic anemia. Available at: http://www.irondisorders.org/Disorders/Sideroblastic.asp 2. Camaschella C. Br J Haematol. 2008;143;27-38. 3. Bottomley S. Clinical aspects, diagnosis, and treatment of the sideroblastic anemias. May 2007. Clinical aspects, diagnosis, and treatment of the sideroblastic anemias. In: UpToDate, Rose BD, ed, UpToDate, Waltham, MA, 2007.

5. 1. Wickramasinghe SN. Curr Opin Hematol. 2000;7:71-78. 2. Iolascon A, et al. Blood. 2001;98:1258-1260. 3. Mehaffey MG, et al. Blood. 2001;98:2681-2688. 4. Dgany O, et al. Am J Genet. 2002;71:1467-1474. 5. Goede JS, et al. Ann Hematol. 2006;85:591-595. 6. Wickramasinghe SN, Wood WG. Br J Haematol. 2005;131:431-446. 7. Heimpel H. Ann Hematol. 2004;83:613-621. Congenital Dyserythropoietic Anaemia A rare group of macrocytic anaemias characterized by ineffective erythropoiesis, dysplastic erythroblast changes, and substantial iron overload (even in nontransfused patients) 1 Other manifestations: splenomegaly, cholelithiasis, and skeletal anomalies 1,2 Diagnosis: based on presence of haemolytic anaemia, low reticulocytes, binucleated normoblasts on a blood smear, nuclear abnormalities in erythroid precursors (pathognomonic) 1-3, mutations in the CDAN I gene (for type 1 4-6 ) and in GATA-1 gene 3 Therapy: splenectomy (for type II), interferon α (for type I), red cell transfusion, and iron chelation for those with iron overload 1,2,7

6. Dyskeratosis Congenita A rare bone marrow failure syndrome characterized by abnormal skin pigmentation, nail dystrophy, and oral leukoplakia 1-5 Other manifestations: pulmonary abnormalities, cellular immunodeficiency, and cancer predisposition 1-5 Inheritance: X-linked (most common and most severe), autosomal dominant, and autosomal recessive 1 Diagnosis: must be considered in any child or adult with bone marrow failure, acute myelogenous leukaemia or myelodysplastic syndrome, skin and oral changes, negative mitomycin C and diepoxybutane tests 1-5, and TERC and DKC1 genes 1,2 Therapy: transfusions, androgens, growth factors, and stem cell transplantation 1 Principal cause (71%) of early mortality: attributed to bone marrow failure 3 1. Dokal I. Br J Haematol. 2000;110:768-779. 2. Knight S, et al. Br J Haematol. 1998;103;990-996. 3. Sílvia Pimenta de Carvalho et al. An bras Dermatol. Rio de Janeiro, 78:579-586, set./out. 2003. 4. Drachtman RA, Alter BP. Dermatol Clin.1995;13:33-39. 5. Kraemer KH. In Freedberg IM, et al, eds. Fitzpatrick´s Dermatology in General Medicine. McGraw-Hill; 1999.

7. Iron Overload in Other Anaemias An inevitable consequence of chronic red cell transfusion therapy regardless of the underlying cause of anaemia Can be seen even in patients who have not been transfused and may be local or systemic Local mitochondrial iron overload is present in all sideroblastic anaemias, whereas systemic iron overload occurs only in forms due to primary or secondary ALAS2 deficiency Renders affected patients at increased risk for developing co- morbidities with a resultant negative impact on survival Requires effective chelation therapy No guidelines exist for managing iron overload in nonthalassaemics and specifically rare anaemias, and treatment is based on experience in thalassaemia Camaschella C. Br J Haematol. 2008;143:27-38.

8. Iron overload can occur after 10–20 transfusions 1 blood unit contains 200 mg iron Iron chelation therapy should be considered when a patient has received approximately 20 units (about 100 mL/kg) of pRBCs or when serum ferritin is >1000 µg/L Transfusion Therapy Results in Iron Overload Porter JB. Br J Haematol. 2001;115:239-252. Cappellini MD, et al. Blood. 2006;107:3455-3462. Transfusional iron intake = volume of packed RBCs (mL) x 1.08 Volume of packed RBCs (mL) = volume of blood (mL) x haematocrit (%) Example: 285 mL blood transfused x 65% haematocrit = 185 mL RBCs x 1.08 mg iron/mL RBCs = 200 mg iron

9. Iron Overload Monitoring Many tools are available for assessing iron overload, and combining these tools allows more accurate assessment Serial ferritin measures—most practical Liver iron concentration (biopsy/MRI/SQUID) Cardiac iron load by MRI

10. The Challenge of Iron Chelation A Question of Balance Too much ironToo much chelator Uncoordinated iron Free-radical generation Organ damage Growth failure Organ failure Cardiac death Uncoordinated chelator Inhibition of metalloenzymes Neurotoxicity Growth failure Bone marrow toxicity Graphic courtesy of Dr. J. Porter.

11. Overview of Iron Chelators PropertyDesferrioxamine 1 Deferiprone 2 Deferasirox 3 RouteSC IV (8–12 h, 5 d/w) Oral 3 times daily Oral once daily Half-life20–30 min3–4 h8–16 h ExcretionUrinary, faecalUrinaryFaecal Adverse effects Local reactions, ophthalmologic, auditory, growth retardation, allergic Gastrointestinal disturbances, agranulocytosis/ neutropaenia, arthralgia, elevated liver enzymes Gastrointestinal disturbances, rash, mild nonprogressive creatinine increase, ophthalmologic, auditory, elevated liver enzymes StatusLicensedNot licensed in USA/Canada Licensed in USA and Europe IndicationsTreatment of chronic iron overload due to transfusion-dependent anaemias ThalassaemiaTreatment of chronic iron overload due to transfusion-dependent anaemias 1. Desferrioxamine [PI]. Novartis Pharma Stein AG. Stein, Switzerland Nov. 2007. 2. Deferiprone Ferriprox [Summary of Product Characteristics] [PI]. Apotex Europe LTD. 1999. 3. Deferasirox [PI]. Novartis Europharm LT. West Sussex, UK. Aug. 2006.

12. Management of Iron Overload To date, management of transfusional iron overload and improved survival have been achieved with parenteral desferrioxamine Around 1/3 to 1/2 of patients are not compliant with desferrioxamine therapy, largely because of the discomfort and demanding nature of the regimen In 2005, the FDA approved deferasirox, an oral tridentate chelator, for the treatment of chronic overload due to transfusional hemosiderosis Deferasirox has been studied in >700 adult and paediatric patients who had transfusion-related iron overload and underlying thalassaemia, sickle cell anaemia, and other anaemias Stumpf JL. Am J Health Syst Pharm. 2007;64:606-616.

13. Retrospective Nationwide Survey of Japanese Patients with Transfusion-Dependent Myelodysplastic Syndrome and Aplastic Anaemia Japanese National Research Group on Idiopathic Bone Marrow Failure Syndromes Investigated relationships between iron overload, chelation practices, and morbidity/mortality in 292 patients with MDS, AA, pure red cell aplasia, myelofibrosis, and other conditions MDS and AA accounted for about 80% of the underlying diseases — MDS: 52.1% — AA: 30.8% — PRCA: 5.1% — MF: 4.5% Abbreviations: AA, aplastic anaemia; MDS, myelodysplastic syndrome; MF, myelofibrosis; PRCA, pure red cell aplasia. Takatoku M, et al. Eur J Haematol. 2007;78:487-494.

14. Retrospective Nationwide Survey of Japanese Patients with Transfusion- Dependent Myelodysplastic Syndrome and Aplastic Anaemia 43% of patients received desferrioxamine therapy but only 8.6% received daily/continuous desferrioxamine Abnormal cardiac and liver function observed in 21.9% (14/64) and 84.6% (11/13) of all patients assessed 75 deaths (25.7%) reported, with cardiac and liver failure noted in 24.0% and 6.7% of cases, respectively, and ferritin levels >1000 ng/mL in 97% of deaths Takatoku M, et al. Eur J Haematol. 2007;78:487-494.

15. Average Changes in Laboratory Values During the Period of Transfusion Dependence in Patients Receiving Desferrioxamine Treatment ParameterIntermittent (once/1.9 wk)Concurrent with TransfusionDaily/Continuous Serum ferritin a,b (ng/mL)+2222.8 (n = 36)+2204.8 (n = 19)-1135.2 (n = 9) SGOT a,c (mU/mL)+28.0 (n = 53)+40.0 (n = 30)-9.2 (n = 10) SGPT (mU/mL)+28.6 (n = 53)+10.3 (n = 30)-28.8 (n = 10) FBS (mg/dL)+31.2 (n = 31)+8.2 (n = 12)-4.8 (n = 5) a Intermittent vs continuous, P <.05. b Continuous vs concurrent, P <.01. c Continuous vs concurrent, P <.05. Daily continuous chelation with desferrioxamine resulted in improved serum ferritin, liver enzymes, and fasting blood sugar Abbreviations: SGOT, serum glutamic oxaldacetic mansaminase; SGPT, serum glutamic pyruvic transaminase; FBS = fasting blood sugar Reprinted from Takatoku M. Eur J Haematol. 2007;78:487-497, with permission from John Wiley & Sons.

16. Study 0108—Phase II Single-Arm Trial  -Thalassaemia and Other Anaemias 1-year trial designed to evaluate the efficacy of deferasirox in 184 regularly transfused patients –85 patients with β-thalassaemia –99 patients with other anaemias (MDS 47, DBA 30, rare anaemias 22) –Aged 3–81 years Patients treated with deferasirox for 1 year, and dosage determined by baseline liver iron concentration assessed by liver biopsy or SQUID A total of 152 patients (82.6%) completed 1 year of treatment Porter J, et al. Eur J Haematol. 2008;80:168-176.

17. Mean Changes in LIC and Ferritin by Disease Group and Dose Abbreviations: DBA, Diamond-Blackfan anaemia; LIC, liver iron concentration; MDS, myelodysplastic syndrome. Reprinted from Porter J, et al. Eur J Haematol. 2008;80:168-176, with permission from John Wiley & Sons.

18. Relative Response of Patients with MDS and Other Transfusion-Dependent Anaemias to Deferasirox—1-Year Prospective Study Iron Excretion Across Dose and Disease Groups Abbreviations: DBA, Diamond-Blackfan anaemia; MDS, myelodysplastic syndrome. Reprinted from Porter J, et al. Eur J Haematol. 2008;80:168-176, with permission from John Wiley & Sons.

19. Relative Response of Patients with MDS and Other Transfusion-Dependent Anaemias to Deferasirox—1-Year Prospective Study Iron-overloaded patients with myelodysplastic syndrome, Diamond-Blackfan anaemia, and other rare anaemias responded to deferasirox in a dose-dependent manner as did patients with  -thalassaemia, with respect to both efficacy and safety parameters Deferasirox was effective and generally well tolerated, resulting in a clinically relevant reduction in overall body iron burden across a broad range of anaemia types Changes in serum ferritin and liver iron concentration correlated, supporting the use of serial serum ferritin measures for monitoring deferasirox therapy Porter J, et al. Eur J Haematol. 2008;80:168-176.

20. Relative Response of Patients with MDS and Other Transfusion-Dependent Anaemias to Deferasirox—1-Year Prospective Study Patients with Diamond-Blackfan anaemia who had the highest average transfusional iron intake, showed the smallest reductions in liver iron concentration (LIC) while patients with myelodysplastic syndrome, who had the lowest mean iron intake, showed the largest dose-dependent reductions in LIC If the differences in transfusional iron loading rate are accounted for, the response to chelation with deferasirox is similar across the different types of transfusion-dependent anaemia studied The most common drug-related adverse events were mild, eg, transient gastrointestinal disturbances, skin rash, and mild, nonprogressive serum creatinine increases Porter J, et al. Eur J Haematol. 2008;80:168-176.

21. Conclusions Patients with rare anaemias often require chronic red blood cell transfusion Iron overload in such patients can occur before transfusion therapy is initiated. This is due to ineffective erythropoiesis and/or increased gastrointestinal iron absorption The most important cause of iron overload in other anaemias remains chronic transfusion therapy Patients with rare anaemias are underscreened and undertreated for their iron overload

22. Conclusions Emerging clinical data indicate that transfusion- dependent patients with rare anaemias are at risk for the consequences of iron overload, including progressive damage to the liver, heart, and endocrine organs Iron overloaded patients, regardless of the underlying disease, should receive iron chelation therapy as early as possible to prevent organ damage The availability of an oral iron chelator may improve compliance in patients noncompliant with the difficult desferrioxamine regimen Identification and registration of patients in an international registry and initiation of collaborative trials are needed in order to lay guidelines for transfusion and iron overload treatment in rare anaemias