Effects of co‐existing α‐thalassaemia in sickle cell disease on hydroxycarbamide therapy and circulating nucleic acids

Hydroxycarbamide (HC) is a key treatment option for sickle cell disease (SCD) (Charache et al, 1995; Ferster et al, 2001; Ware & Aygun, 2009; Voskaridou et al, 2010), but not all patients respond clinically, despite compliance (Steinberg et al, 1997). HC response is monitored by increases in fetal haemoglobin, mean corpuscular volume and mean corpuscular haemoglobin. We have previously shown that such increases were muted in SCD patients with co-inherited a thalassaemia (a-SCD) compared to those without (SCD) (Vasavda et al, 2008). Cell-free DNA (cfDNA), a marker of tissue damage, is elevated during sickle acute pain (Vasavda et al, 2007) and reduced with HC therapy (Ulug et al, 2008). It is generally accepted that co-inherited a thalassaemia reduces haemolysis in SCD, subsequent to which the haematocrit is relatively increased with predisposition to vaso-occlusive complications (Ballas, 2001). We hypothesized that an increase in vaso-occlusive events would, in turn, have an effect on cfDNA levels in SCD. Here we describe the effect of co-inherited a thalassaemia on clinical response to HC, and on cfDNA levels. Fifty-two participants were recruited from the specialist haematology clinics at King’s College (n = 38) and Guy’s and St Thomas’ (n = 14) Hospitals, London. All participants from Guy’s and St Thomas’ hospital were treated with HC. The study was approved by the South London REC Office (08/ H0803/185); all participants gave informed, written consent. Inclusion criteria were regular attendance at the sickle cell outpatient clinics, and homozygous sickle cell disease (HbSS) or compound heterozygotes with b thalassaemia (HbSb). Exclusion criteria were: lack of data on a globin genotype, regular blood transfusion and poor compliance (in the treated group). Commencing or terminating HC therapy was a purely clinical decision, and dosing criteria were as previously described (Ulug et al, 2008). For patients treated with HC, clinical data were collected retrospectively for the year prior to starting HC therapy (‘Pre-HC’) and prospectively throughout the study (‘OnHC’). For patients commencing HC during the study, ‘OnHC’ data were collected from 6 months after starting HC therapy. Clinical response was assessed in terms of attendance at accident and emergency, hospital admissions per year, and days spent as an inpatient per year due to sickle-related pain (‘inpatient days’). Blood samples were collected at each ‘steady state’ clinic visit for cfDNA measurement, and plasma cfDNA extracted and quantified as previously described (Vasavda et al, 2007). Clinical data or blood samples were not collected for 3 months after any pro rata blood transfusion. Data was analysed using an unpaired Student’s t-test to compare data between the a-SCD and SCD groups (significance threshold P = 0Æ05). To avoid skewing of data in favour of patients with a greater number of samples and clinical data points, a ‘patient mean’ was calculated and used in further analyses. Clinical response was assessed for those patients treated with HC. Mean ‘on HC’ and ‘pre HC’ laboratory values were obtained, and response calculated as the magnitude change in each parameter; this change represents an intra-patient change in response to HC. An overall mean was then calculated for each group (SCD and a-SCD). For the cfDNA analysis data was not analysed intra-individually: results were stratified according to HC therapy, then further by a thalassaemia status. Laboratory (haematological) data were available for ‘pre-HC’ and ‘on-HC’, as these are routine care tests, whereas cfDNA is a research test, requiring a separate sample. Only a very small number of patients started HC during the study period, so sufficient pre and post intra-individual cfDNA values were not available for analysis. However, we did show in a previous study that HC therapy resulted in a significant change in cfDNA levels (Ulug et al, 2008). Patients with heterozygous (aa/-a) and homozygous (-a/-a) a thalassaemia were analysed as one group. Mean cfDNA levels in the a-SCD and SCD groups were compared separately for HC-treated and untreated patients. 33 patients were on HC therapy with a mean HC dose of 1151 mg (18Æ3 mg/kg/d); 19 were untreated during the study duration. Participants were followed for a mean of 10Æ4 months (range 3Æ6–19Æ2 months). Three participants required blood transfusions during the study (all in the ‘NoHC’ group). Further patient characterization is shown in Table I. Pre-HC data was not available for all participants; numbers used are shown with each result. HC therapy dramatically reduced the total number of hospital admissions and accident and emergency department (A&E) attendances per year in both groups (SCD and a-SCD) (Table I) but the difference between the groups was not significant. However, a-SCD patients had a significantly (P = 0Æ02) smaller reduction in the number of inpatient days (3Æ833 d ± 3Æ146 N = 7) compared to the SCD group (19Æ08 ± 3Æ932. N = 12) (Fig 1A); changes in haematological indices reflected our previous findings (Table I). correspondence