Title: Investigation of the metabolic fate of tyrosine and tryptophan in patients with alkaptonuria pre- and post treatment with nitisinone
Funder: Health Education England

Abstract: Alkaptonuria (AKU) is a rare inherited metabolic disease (OMIM reference 203500) that is inherited in an autosomal recessive fashion. It has a reported prevalence of 1 in 250,000, although some countries such as Slovakia have a higher prevalence rate of 1 in 19,000 (1-3). AKU is a disorder of tyrosine metabolism, whereby degradation of tyrosine is blocked at the level of homogentisic acid (HGA) due to congenital lack of the enzyme homogentisate dioxygenase (HGD; EC 1.13.11.5) (4). Until recently, medical management of AKU has been palliative relying largely on low protein diet, analgesia and arthroplasty (5). A potential HGA lowering therapy has now become available in the form of nitisinone, an orally administered drug. Nitisinone is an enzyme inhibitor that reduces the accumulation of HGA. One of the metabolic consequences of treatment with nitisinone is the accumulation of tyrosine. This has been seen in patients treated at the National AKU Centre (NAC) in Liverpool. This is also a documented consequence of nitisinone treatment in Hereditary Tyrosinaemia-1 (HT-1), a lethal inherited metabolic disease. It is believed that the supra-physiological concentrations of tyrosine may be responsible for the cognitive impairment observed in HT-1 (6-7).Aims (1) Elucidate the metabolic fate of tyrosine pre- and post-treatment with nitisinone, with specific emphasis on the catecholamine metabolic pathway. (2) Examine whether altered tyrosine metabolism influences the metabolism of tryptophan, specifically its metabolism to serotonin.Plan The metabolic fate of tyrosine and tryptophan will be examined using supervised semi-quantitative high-resolution mass spectrometry (LC-TOF-MS) and quantitative targeted liquid-chromatography tandem mass spectrometry (LC-MS/MS). Urine and serum samples collected during the routine care provided to AKU patients at the NAC in Liverpool and during the DevelopAKUre clinical trial will be analysed. This will be done in three stages: 1.Samples collected from AKU patients treated with nitisinone on an open label basis (2mg daily) collected in the NAC - the metabolic fate of tyrosine and tryptophan will be examined in AKU patients recruited to the NAC (n=60). 24-hour urine collections, and serum samples collected once per year over five years will be analysed to assess the impact of nitisinone treatment on the metabolism of tyrosine and tryptophan. 2. Samples collected in SONIA-1: nitisinone dosing study. (a) Samples collected pre-treatment with nitisinone - the metabolic fate of tyrosine and tryptophan will be examined in 40 patients with AKU. 24-hour urine collections and serum samples (collected hourly for 24 hours) will be analysed to determine the excretory profile of tyrosine and tryptophan metabolites pre-treatment with nitisinone. (b) This will involve the analysis of serum samples from patients (n=40) randomised to take placebo, 1mg, 2mg, 4mg and 8mg once daily for 4-weeks. This will establish the impact of the different doses of nitisinone on tyrosine and tryptophan metabolism. 3. Samples collected in SONIA-2: Long term evaluation (4-years) of nitisinone dose chosen in SONIA-1 (10mg) versus placebo of patients with ochronosis (n=140). This will enable the long term impact of nitisinone treatment on tyrosine and tryptophan metabolism to be assessed in a large cohort. Potential Benefits to Patients and NHS Understanding the metabolic fate of tyrosine in AKU patients pre- and post-nitisinone is vital to fully understand the long-term safety of nitisinone. The data generated in this study will be used to support an application for marketing authorisation of nitisinone. The marketing of nitisinone will be life changing to AKU patients as it is expected to significantly reduce the morbidity associated with AKU. The wider health implications of this project are that is will provide insight into the metbolic fate of tyrosine and tryptophan in HT-1

Lay abstract: ObjectivesTo study the metabolic fate of the amino acids tyrosine and tryptophan in patients with alkaptonuria (AKU) pre- and post-treatment with the orphan drug nitisinone. Design To examine the metabolism of tyrosine and tryptophan using a laboratory technique called mass spectrometry. This can measure the pattern of metabolites and quantify them. Expected outcomeTo improve the understanding of tyrosine and tryptophan metabolism in AKU patients treated with nitisinone. This is necessary to fully understand the long-term safety of nitisinone. Anticipated benefits To provide scientific support for an application for marketing authorisation of nitisinone for the treatment of AKU

Research activity codes: 6.1 PHARMACEUTICALS
Health categories: METABOLIC AND ENDOCRINE