Joan S. Henthorn

Neonatal screening for haemoglobinopathies : the results of a 10-year programme in an English Health Region

Neonatal identification of sickle cell disease can significantly reduce mortality and morbidity during the first 5 years of life. During a 10‐year period, 414 801 neonates were screened by isoelectric focusing. The most common variants detected were haemoglobins S, C, D and E. Two hundred and fifty of the samples tested were homozygotes or compound heterozygotes, and 6554 samples were heterozygotes for the common variants. The gene frequencies in the population tested were calculated from this data for the most common variants. They were: S, 0·0057; C, 0·0014; DPunjab(Los Angeles), 0·0007; E, 0·0005. Additionally, 16 babies had beta thalassaemia major and 405 had rarer variants, of which six had never previously been described. Knowledge of the distribution of these inherited diseases is useful in healthcare planning and appropriate allocation of resources, while counselling targeted at appropriate couples enables informed parental choice and may prevent disease.

Neonatal screening for sickle cell disorders.

The aim of a neonatal screening programme is to identify those individuals within a population who have a specific serious disorder, the outcome of which can be ameliorated by early intervention. Implementation of neonatal screening requires a complex framework comprising information, education and consent of parents, sampling, transport, laboratory testing (both first line and confirmatory), issue of reports and counselling of families with affected children. In order to work effectively, an integrated approach must be developed, with agreed standards and responsibilities at every stage and in-built audit. Implementation of a programme requires education, co-ordination and funding. Both isoelectric focusing and high-performance liquid chromatography have high specificity and sensitivity, and are the two main laboratory techniques for haemoglobinopathy screening currently suitable for routine use. Laboratory centralization to achieve costeffectiveness and technical expertise is essential. No programme will achieve its aim without appropriate high quality counselling and subsequent clinical care. This poses problems of capacity, education and funding for most of the health systems. However, it is indisputable that an effective neonatal screening programme for sickle cell disorders will reduce morbidity and save the lives of children with sickle cell disease. The impending implementation by 2004 of a ‘new national linked antenatal and neonatal screening programme for haemoglobinopathy and sickle cell disease’ (SCD) (The NHS Plan, 2000, Command Paper 4818) raises several issues regarding its execution in England. The positions of Wales, Northern Ireland and Scotland regarding this issue have yet to be defined. Screening programmes aim to identify those individuals within a population who have a specific disorder for which intervention, such as medical treatment, education or counselling, can improve the natural course of the disease. Several authors have suggested criteria, which should be applied in considering whether population screening for a specific condition should be instituted (Wilson & Jungner, 1968; Lappe et al, 1972). The case for neonatal screening for sickle cell disorders is based on the advantages gained by infants and children with SCD who have been diagnosed early and thus commenced on prophylactic penicillin (Gaston et al, 1986; Davies et al, 2000; Riddington & Owusu-Ofori, 2002), administered antipneumococcal vaccination (Davies et al, 2000; Lees et al, 2000), parents taught to detect acute splenic sequestration (Sickle Cell Disease Guideline Panel, 1993) and given comprehensive care. Despite the fact that early diagnosis of babies with b-thalassaemia major carries few advantages, it also allows genetic counselling for parents with affected newborns (Laird et al, 1996). In 1993, the Standing Medical Advisory Committee recommended that, as a minimum, screening for haemoglobinopathies in the UK should be targeted, apart from in areas where 15% or more of the population was at risk of conceiving a child with SCD, where it should be universal (Standing Medical Advisory Committee, 1993). In the same year, the Sickle Cell Disease Guideline Panel in the USA recommended universal screening for SCD because of the ethical and practical difficulties raised by targeting (Sickle Cell Disease Guideline Panel, 1993). Subsequently, reports to the NHS Health Technology Assessment Programme on screening for haemoglobinopathies (Davies et al, 2000) recommended universal screening for haemoglobinopathies in the UK. This was incorporated into the NHS National Plan (The NHS Plan, 2000, Command Paper 4818): ‘a linked neonatal and antenatal screening programme for haemoglobinopathies’. This programme is now in the early phase of implementation, developing standards, guidelines, framework and accrediting laboratories (http://www.kcl-phs.org.uk/haemscreeening). Such a programme involves a large number of health professionals in different sectors of the health care system, as outlined below. It is important to define roles, standards and responsibilities, all of which are aided by audit and feedback.

Organisation and cost‐effectiveness of antenatal haemoglobinopathy screening and follow up in a community‐based programme

Objective To consider the organisation cost and effectiveness, of universal, community‐based antenatal screening for the haemoglobinopathies, and to estimate the cost‐effectiveness of programmes at different levels of prevalence and mix of haemoglobinopathy traits.

Haematological and clinical aspects of sickle cell disease in Britain

Publisher Summary This chapter focuses on the hematological and clinical aspects of sickle cell disease (SCD) in Britain. SCD was considered to be rare and of little significance in Britain. The great majority of cases in the United Kingdom are British born of West Indian or African extraction and are permanently domiciled in Britain. About 5% are temporarily resident in the United Kingdom as students, visitors, diplomats, and businessmen from Africa, and some 10% are their children. Most cases are under the age of 30, and over 20% live in the boroughs of Brent and Lambeth in London and in Birmingham, the remainder resides in other London boroughs, the Midlands, and Mersey side. In Brent, almost all 26 pregnancies reviewed in 1984 suffered some SCD complication. These included a maternal death from stroke at 39 weeks, a stillbirth following a fulminant abdominal sickling crisis at 29 weeks, splenic infarction in the second and third trimesters and rapidly progressive aseptic necrosis of the hip and sickle retinopathy. An important feature was the postpartum cluster of pulmonary thromboembolic events in patients with SC which had been abolished by the use of prophylactic twice daily subcutaneous heparin from 36 weeks to 6 weeks postpartum. Adequate hypertransfusion can prevent some of these complications if started sufficiently early in pregnancy.

Two new Variants with the Same Substitution at Position β122: Hb Bushey [β122(GHS)Phe→Leu] and Hb Casablanca [β65(E9)Lys→Met;β122(GHS)Phe→Leu]

Hb Bushey, found in a Chinese baby and his father, is a new variant with a point mutation leading to the substitution Phe→tLeu at position β122. Hb Casablanca, found in a family from Morocco, is a further example of a hemoglobin variant that carries two abnormalities in the same chain; the first is identical to that of Hb Bushey and the second to that of Hb J-Antakya [β65 (E9)Lys→Met]. Structural abnormalities of both Hbs were determined by protein chemistry methods including electrospray and tandem mass spectrometry. Their stability and oxygen binding properties were found to be identical to those of Hb A. Various mechanisms that may lead to two point mutations in the same chain are reviewed briefly.

Hb Harrow [β118(GH1)Phe→Cys]: A New Neutral Hemoglobin Variant

We report here on a new hemoglobin (Hb) variant found in a newborn baby boy of Indian Gujerati origin, living in the Harrow area of London, England. This variant was observed during a systematic program of neonatal screening for the main hemoglobinopathies performed at the Central Middlesex Hospital in the North Thames (West) Region of London. In this program, the samples are collected from a heelprick onto filter paper when the child is 7-10 days old. A dried blood spot of 0.3 cm diameter is obtained from which Hb is eluted by a 5 mM KCN solution to be analyzed by isoelectrofocusing (IEF).

Hb NIIGATA (β1(NA1)Val®Leu) IN A ROMANIAN INDIVIDUAL RESULTING FROM ANOTHER NUCLEOTIDE SUBSTITUTION THAN THAT FOUND IN THE JAPANESE

We present a new case of Hb Niigata that we named Hb Niigata(C), observed in a woman from Romania, with a mutation different from that described in Japanese (GTG→CTG instead of GTG→TTG). This single nucleotide substitution replaces the valine residue for leucine at codon 1 and causes retention of the N-terminal methionine leading to an elongated β chain. This mutation was without any hematological consequences.

Hb Uxbridge [β20(B2)Val → Gly]: A New Variant with Mild Incre in Oxygen Affinity Found During A Neonatal Screening Program

Hb Uxbridge [beta 20(B2)Val-->Gly] was found in an English family during a neonatal hemoglobinopathy screening program. In both the child and the parent carrying this hemoglobin variant, the red cell parameters were normal. By isoelectrofocusing Hb Uxbridge appeared to have an isoelectric point slightly higher than Hb A but was silent on cellulose acetate and acid gel electrophoresis. Structural modifications affecting position beta 20(B2) have been demonstrated to be responsible for a high oxygen affinity and polycythemia in Hb Olympia (Val-->Met) and Hb Trolhattan (Val-->Glu). In the case of Hb Uxbridge, despite an alteration of the same site, the oxygen binding parameters of the patients hemolysate showed only a mild (ca. 20%) increase in oxygen affinity.

A New Silent β Chain Variant: Hb Hounslow [β80(EF4)Asn→Tyr]

A New Silent β Chain Variant: Hb Hounslow [β80(EF4)Asn→Tyr] We report a new neutral hemoglobin (Hb) variant, found, during neonatal screening, in a child originating from Afghanistan. This variant was revealed by cation exchange and reversed phase high performance liquid chromatography (HPLC), but was silent in electrophoretic methods except for globin chain electrophoresis in the presence of urea and Triton X-100. The structural modification was determined by protein structure studies and the substitution established by tandem mass spectrometry (MS/MS). The mutation, located near to the 2,3-diphosphoglycerate (2,3-DPG) binding site, was without any hematological consequences. The pitfalls presented by the presence of neutral Hb variants as modulator factors of the main hemoglobinopathies is discussed.

Alteration of an intersubunit contact in hemoglobin variants: comparative study of modifications at position α126 Asp (H9)

Four human hemoglobin variants have already been described at position alpha 126 (H9), which is normally occupied by an aspartate: Hb Montefiore (-->Tyr), Hb Tarrant (-->Asn), Hb Fukutomi (-->Val), Hb Sassari (-->His). An additional variant, Hb West One (alpha 126 (H9) Asp-->Gly) is herein described. Aspartate alpha 126 (H9) is involved in a set of hydrogen bonds and salt bridges located at the C-terminal portion of the alpha-chains and of the C-helix of the beta-chains, which are broken in the oxy conformer, providing one of the most important sources of the difference in free energy between the T- and R-state in hemoglobin. A comparative study of four of these alpha 126 Hb variants is presented. An identical degree of alteration of the oxygen binding properties (increased oxygen affinity and decreased cooperativity) was found in all cases, when measured under standard experimental conditions (pH 7.2, 0.1 M NaCl). In contrast, the effect of L345 (a derivative of bezafibrate, which is a specific alpha-chain binding effector) on oxygen binding to Hb differed from one variant to another. When a bulky Tyr or His residue occupied the alpha 126 (H9) position, little effect of L345 was observed. Conversely, when this position was occupied by a residue of smaller size (Gly or Asn), normal heterotropic effects were observed. Molecular graphic modelling indicates that two classes of three-dimensional structure modifications may occur.