A total of 67 pediatric patients affected by haemoglobinopaties underwent matched related HSCT at our Institute between June 2005 and Nov 2008. Children were referred to our center by Mediterranean and Middle East Countries. Cost of treatment was covered by a cooperation program funded by the Mediterranean Institute of Haematology http://www.fondazioneime.org. Among these, 6 patients (9%) had evidence of severe CSA related neurotoxicity requiring CSA discontinuation. The underlying disease was beta thalassemia major (n = 5) or sickle cell disease (SCD, n = 1). Patients' characteristics are shown in table 1. According to the risk classes described by Lucarelli11, 3 thalassemia patients were classified as class II and 2 as class III. None of the patients had preexisting neurologic deficits. Baseline neurological involvement was excluded in the SCD patient by MRI and EEG. Conditioning regimen is summarized in table 2. All patients received seizures prophylaxis with oral clonazepam (0.05 mg/kg/day) starting 24 hours before the first busulfan dose to 24 hours after the last dose.

GvHD prophylaxis consisted of CSA, short course of iv Methotrexate (MTX) 10 mg/m2 on d 1, 3 and 6 and metilprednisolone 0.5 mg/kg from d -1 until d 25. ATG was added in 3 patients.

CSA was used orally at the dose of 10 mg/kg/day from day -2. The dose was adjusted to a trough plasma level of 150-250 ng/mL with twice weekly monitoring. In the absence of GvHD, CSA was tapered from d 60 until discontinuation at 1 year.

Monitoring for CSA related side effects consisted of daily examination, 6 hourly blood pressure and daily blood routine biochemistry including magnesium till day 30, followed by 3 times weekly till day 60. Arterial hypertension above 75° percentile for age and weight was treated with oral calcium antagonists. Fluid overload was treated with iv furosemide. Magnesium was supplemented for plasma levels below 0.8 mmol/Lt. Children developing CSA neurotoxicity underwent neuro-radiological (MRI scan) and electrophysiological studies (EEG). In case of epileptic signs, EEG was repeated after 24 hours and then on regular basis in order to monitor the outcome of the anticonvulsive therapy. CSA-related toxicity was evaluated using the National Cancer Institute common toxicity criteria (NCI/CTC). Clinical management was lead by the HSCT specialists on the HSCT ward with the collaboration of intensivists, neurologists, neurophysiologists and neuroradiologists.

All six patients developing neurotoxicity presented prodromes 1-2 days before overt neurotoxicity. Prodromes were arterial hypertension (n = 5), headache (n = 2), visual disturbances (n = 1) or vomiting (n = 1). Symptoms of CSA neurological toxicity were characterized by: generalized seizures (grade 3-4 neurotoxicity) (n = 4), endocranial hypertension evidenced by impaired level of consciousness (grade 3 confusion) associated to bradycardia, arterial hypertension and cerebral vomiting (n = 1), and blurred vision (grade 3 visual toxicity) (n = 1). The median day of appearance was 11 days after HSCT (range +1 to +40). When neurological symptoms appeared, CSA plasma levels were in range in all patients (median 178, range 69-250 ng/ml). Clinical and laboratory findings are summarized in Table 3. None of the patients had evidence of thrombotic microangiopathy. None of the clinical pictures were suggestive for viral encephalitis, as evidenced by apyrexia, normal CRP, negative molecular virological screening on plasma, non suggestive MRI and EEG pictures.

Neuroradiological and electrophysiological studies were performed within 24 hours from neurological symptoms appearance. A CT scan performed in patient #5 because MRI was not immediately available showed a minimal subdural haemorrhagic layer compatible with post epileptic trauma. MRI was performed in all patients. In 5/6 patients, MRI showed a symmetric involvement of the posterior portions of the cerebral hemispheres, while one case showed abnormal signals hyperintense in T2 and Flair images in the cerebellar hemispheres, involving predominantly the white matter but also the cortex (figure 1, 2). In 5 out of the 6 patients studied, a moderate gyral swelling was present. On diffusion weighted images these lesions had an increase diffusion (hyperintens sygnal on apparent diffusion coefficient maps) consistent with vasogenic oedema.

In five out of 6 children, it was possible to perform an EEG at the time of the neurological symptoms onset. EEG of three out of five patients showed a status epilepticus (Figure 3, 4, 5). The other two patients showed, respectively, a focal delta activity and focal epileptiform discharges. EEGs were repeated every 2-3 days in the acute phase in order to adjust the anticonvulsive therapy.

On occurrence of neurotoxicity, CSA was immediately withdrawn. After 2 days of calcineurin-inhibitor washout, tacrolimus alone (n = 3) or tacrolimus and mycophenolate mofetile (n = 3) (according to the individual GvHD risk) were introduced with tacrolimus level monitoring. Neurological symptoms resolved within 2-3 days following CSA withdrawal and did not re-occur during tacrolimus treatment. Intravenous lorazepam was immediately administered at seizures unset, followed by anticonvulsive treatment with phenitoyn. Patient #1 (SCD) required association with phenobarbital to control generalized seizures (table 3). Intravenous mannitol was used successfully in patient #2 and #6 to treat confusion, vomiting, headache and blurred vision related to endocranial hypertension. Anticonvulsive therapy was tapered and stopped after normalization of EEG findings. In patient #6 phenitoyn was withdrawn after 3 weeks of treatment because of the onset of a severe hepatic-toxicity (elevation of liver enzymes and bilirubin) in the presence of a liver biopsy suggestive of drug-related injury. Liver function normalized following phenitoyn discontinuation. Anticonvulsive therapy was continued with clonazepam only, without seizure recurrence.

Outcome

Neuroradiological monitoring was performed by MRI every 7-14 days until complete remission of the diffuse signal alterations, which occurred after a median of 15 days (range 12-90 days) from CSA withdrawal. Non epileptiform EEG abnormalities were still detectable in all patients in the 15 days after the onset of symptoms. The complete normalization of EEG occurred after a median of 120 days (range 60-140 days). Five patients are alive and symptoms-free at a median follow up of 1005 days (range 320-1065) without neurological sequelae, while one patient died on day 60 for grade IV GvHD, without neurological symptoms.