Nitisinone Treatment Affects Biomarkers of Bone and Cartilage Remodelling in Alkaptonuria Patients.

Nitisinone has been approved for treatment of alkaptonuria (AKU). Non-invasive biomarkers of joint tissue remodelling could aid in understanding the molecular changes in AKU pathogenesis and how these can be affected by treatment. Serological and urinary biomarkers of type I collagen and II collagen in AKU were investigated in patients enrolled in the randomized SONIA 2 (NCT01916382) clinical study at baseline and yearly until the end of the study (Year 4). The trajectories of the biomarkers over time were observed. After treatment with nitisinone, the biomarkers of type I collagen remodelling increased at Year 1 (19% and 40% increase in CTX-I and PRO-C1, respectively), which was potentially reflected in the higher degree of mobility seen following treatment. The biomarkers of type II collagen remodelling decreased over time in the nitisinone group: C2M showed a 9.7% decline at Year 1, and levels then remained stable over the following visits; CTX-II showed a 26% decline at Year 3 and 4 in the nitisinone-treated patients. Nitisinone treatment induced changes in biomarkers of bone and cartilage remodelling. These biomarkers can aid patient management and deepen our knowledge of the molecular mechanisms of this rare disease.

Revisiting Quantification of Phenylalanine/Tyrosine Flux in the Ochronotic Pathway during Long-Term Nitisinone Treatment of Alkaptonuria.

Changes in the phenylalanine (PHE)/tyrosine (TYR) pathway metabolites before and during homogentisic acid (HGA)-lowering by nitisinone in the Suitability of Nitisinone in Alkaptonuria (AKU) 2 (SONIA 2) study enabled the magnitude of the flux in the pathway to be examined. SONIA 2 was a 48-month randomised, open-label, evaluator-blinded, parallel-group study performed in the UK, France and Slovakia recruiting patients with confirmed AKU to receive either 10 mg nitisinone or no treatment. Site visits were performed at 3 months and yearly thereafter. Results from history, photographs of eyes/ears, whole body scintigraphy, echocardiography and abdomen/pelvis ultrasonography were combined to produce the Alkaptonuria Severity Score Index (cAKUSSI). PHE, TYR, hydroxyphenylpyruvate (HPPA), hydroxyphenyllactate (HPLA) and HGA metabolites were analysed by liquid chromatography/tandem mass spectrometry in 24 h urine and serum samples collected before and during nitisinone. Serum metabolites were corrected for total body water (TBW), and the sum of 24 h urine plus total body water metabolites of PHE, TYR, HPPA, HPLA and HGA were determined. The sum of urine metabolites (PHE, TYR, HPPA, HPLA and HGA) were similar pre- and peri-nitisinone. The sum of TBW metabolites and sum TBW + URINE metabolites were significantly higher peri-nitisinone (p < 0.001 for both) compared with pre-nitisinone baseline. Significantly higher concentrations of metabolites from the tyrosine metabolic pathway were observed during treatment with nitisinone. Arguments for unmasking of the ochronotic pathway and biliary elimination of HGA are put forward.

Analysis of the Phenotype Differences in Siblings with Alkaptonuria.

Alkaptonuria (AKU) is a rare autosomal recessive disorder caused by mutations within a gene coding for homogentisate 1,2-dioxygenase (HGD). To date, 251 different variants of this gene have been reported. The metabolic disorder in AKU leads to the accumulation of homogentisic acid (HGA), resulting in ochronosis (pigmentation of the connective tissues) and severe ochronotic spondylo-arthropathy, which usually manifests in the mid-thirties. An earlier genotype−phenotype correlation study showed no differences in serum HGA levels, absolute urinary excretion of HGA, or in the clinical symptoms between patients carrying HGD variants leading to 1% or >30% residual HGD activity. Still, as reported previously, the variance of the excretion of the HGA was smaller within affected siblings that share a common genotype. The present study is the first ever to systematically analyze the baseline clinical data of 24 AKU sibling pairs/groups collected in the SONIA 2 (Suitability Of Nitisinone In Alkaptonuria 2) study to evaluate phenotypical differences between patients carrying the same HGD genetic variants. We show that even between siblings there was considerable variability in the disease severity. This indicates that some other yet unidentified genetic, biomechanical, or environmental modifying factors may contribute to accelerated pigmentation and connective tissue damage observed in some patients.

Radiological evolution of spinal disease in alkaptonuria and the effect of nitisinone.

OBJECTIVES: Ochronotic spondyloarthropathy represents one of the main clinical manifestations of alkaptonuria (AKU); however, prospective data and description of the effect of nitisinone treatment are lacking. METHODS: Patients with AKU aged 25 years or older were randomly assigned to receive either oral nitisinone 10 mg/day (N=69) or no treatment (N=69). Spine radiographs were recorded yearly at baseline, 12, 24, 36 and 48 months, and the images were scored for the presence of intervertebral space narrowing, soft tissue calcifications, vacuum phenomena, osteophytes/hyperostosis and spinal fusion in the cervical, thoracic and lumbosacral segment at each of the time points. RESULTS: At baseline, narrowing of the intervertebral spaces, the presence of osteophytes/hyperostosis and calcifications were the three most frequent radiographic features in AKU. The rate of progression of the five main features during the 4 years, ranked from the highest to lowest was as follows: intervertebral spaces narrowing, calcifications, vacuum phenomena, osteophytes/hyperostosis and fusions. The rate of progression did not differ between the treated and untreated groups in any of the five radiographic parameters except for a slower rate of progression (sum of all five features) in the treatment group compared with the control group (0.45 (1.11) nitisinone vs 0.74 (1.11) controls, p=0.049) in the thoracic segment. CONCLUSION: The present study shows a relatively slow but significant worsening of radiographic features in patients with AKU over 4 years. Our results demonstrate a modest beneficial effect of 10 mg/day of nitisinone on the slowly progressing spondylosis in AKU during the relatively limited follow-up time. TRIAL REGISTRATION NUMBER: NCT01916382.

Comparing the Phenylalanine/Tyrosine Pathway and Related Factors between Keratopathy and No-Keratopathy Groups as Well as between Genders in Alkaptonuria during Nitisinone Treatment.

Nitisinone (NIT) causes tyrosinaemia and corneal keratopathy (KP), especially in men. However, the adaptation within the phenylalanine (PHE)/tyrosine (TYR) catabolic pathway during KP is not understood. The objective of this study is to assess potential differences in the PHE/TYR pathway during KP and the influence of gender in NIT-induced tyrosinaemia in alkaptonuria (AKU). Samples of serum and 24 h urine collected from patients treated with NIT during a 4-year randomized study in NIT vs. no-treatment controls (SONIA 2; Suitability Of Nitisinone In Alkaptonuria 2; EudraCT no. 2013-001633-41) at months 3 (V2), 12 (V3), 24 (V4), 36 (V5) and 48 (V6) were included in these analyses. Homogentisic acid (HGA), TYR, PHE, hydroxyphenylpyruvate (HPPA), hydroxyphenyllactate (HPLA) and sNIT were analysed at all time-points in serum and urine in the NIT-group. All statistical analyses were post hoc. Keratopathy occurred in 10 out of 69 AKU patients, eight of them male. Thirty-five sampling points (serum and 24 h urine) were analysed in patients experiencing KP and 272 in those with no-KP (NKP) during NIT therapy. The KP group had a lower HPLA/TYR ratio and a higher TYR/PHE ratio compared with the NKP group (p < 0.05 for both). There were 24, 45, 100 and 207 sampling points (serum and 24 h urine) in the NIT group which were pre-NIT female, pre-NIT male, NIT female and NIT male, respectively. The PHE/TYR ratio and the HPLA/TYR ratio were lower in males (p < 0.001 and p < 0.01, respectively). In the KP group and in the male group during NIT therapy, adaptive responses to minimise TYR formation were impaired compared to NKP group and females, respectively.

Improving the clinical accuracy and flexibility of the Alkaptonuria severity score index.

Alkaptonuria (AKU) is a rare genetic disorder where oxidised homogentisic acid accumulates in connective tissues, leading to multisystem disease. The clinical evaluation Alkaptonuria Severity Score Index (cAKUSSI) is a composite score that assesses the extent of AKU disease. However, some components assess similar disease features, are difficult to measure reliably or cannot be measured in resource-limited environments. cAKUSSI data from the 4-year SONIA 2 randomised controlled trial, which investigated nitisinone treatment in adults with AKU, were analysed (N = 125). Potentially biased or low-information cAKUSSI measurements were identified using clinical and statistical input to create a revised AKUSSI for use in AKU research (cAKUSSI 2.0). Additionally, resource-intensive measurements were removed to explore a flexible AKUSSI (flex-AKUSSI) for use in low-resource environments. Revised scores were compared to cAKUSSI in terms of correlation and how they capture disease progression and treatment response. Eight measurements were removed from the cAKUSSI to create the cAKUSSI 2.0, which performed comparably to the cAKUSSI in measuring disease extent, progression and treatment response. When removing resource-intensive measurements except for osteoarticular disease, the flex-AKUSSI was highly correlated with the cAKUSSI, indicating that they quantified disease extent similarly. However, when osteoarticular disease (measured using scans) was removed, the corresponding flex-AKUSSI underestimated disease progression and overestimated treatment response compared to the cAKUSSI. Clinicians may use the cAKUSSI 2.0 to reduce time, effort and patient burden. Clinicians in resource-limited environments may find value in computing a flex-AKUSSI score, offering potential for future global registries to expand knowledge about AKU.

Effects of a protein-restricted diet on body weight and serum tyrosine concentrations in patients with alkaptonuria.

In an open-label, controlled study of nitisinone in alkaptonuria (SONIA 2), patients were advised to lower dietary protein intake to reduce serum tyrosine (s-Tyr) levels and the risk of keratopathy. A body weight increase was observed in the nitisinone-treated patients but not in the control group. To investigate the effectiveness and consequence of protein restriction in patients with alkaptonuria, a post-hoc analysis of SONIA 2 was performed. One hundred and thirty-eight patients were randomised (nitisinone: n = 69, controls: n = 69). Comparison of baseline and Month 12 data on 24-h urinary excretion of HGA (u-HGA24) and urea (u-urea24, used as an approximate protein intake measure), tyrosine and body weight were performed using paired t tests. Comparisons of data between groups were made using 2-sample t tests. We found that u-urea24 decreased more in nitisinone-treated than controls. The study centre with lowest average s-Tyr and u-urea24 (nitisinone arm) at Month 12 also had lowest keratopathy incidence (3.1%), while the centre with highest values showed the highest (14.6%). S-Tyr was generally high in those with keratopathy, but those without keratopathy had similar elevated values. A similar pattern across centres was seen for body weight changes, with a statistically significant weight increase in nitisinone-treated patients at centres with lower u-urea24 values. Therefore, in nitisinone-treated patients, protein restriction led to increased body weight but may also have lowered the risk of developing keratopathies. If introduced, a protein-restricted diet should be supervised by a dietician and, when appropriate, include amino acid supplements deficient in tyrosine and phenylalanine, to avoid malnutrition and undesired weight increase.

Clinical development innovation in rare diseases: lessons learned and best practices from the DevelopAKUre consortium.

New opportunities have arisen for development of therapies for rare diseases with the increased focus and progress in the field. However, standardised framework integrating individual initiatives has not been formed. We present lessons learned and best practice from a collaborative success case in developing a treatment for a rare genetic disease. Our unique consortium model incorporated several of the identified developments under one project, DevelopAKUre, truly bringing together academia, industry and patient organisations in clinical drug development. We found that the equal partnership between all parties in our consortium was a key success factor creating a momentum based on a strong organisational culture where all partners had high engagement and taking ownership of the entire programme. With an agreed mutual objective, this provided synergies through connecting the strengths of the individual parties. Another key success factor was the central role of the patient organisation within the management team, and their unique study participants' advocacy role securing the understanding and meeting the needs of the clinical study participants in real-time. This resulted in an accelerated enrolment into the clinical studies with a high retention rate allowing for delivery of the programme with significantly improved timelines. Our project was partly funded through an external EU research grant, enabling our model with equal partnership. Further attention within the community should be given to establishing a functional framework where sustainable funding and risk sharing between private and public organisations allow for our model to be replicated.

Efficacy and safety of once-daily nitisinone for patients with alkaptonuria (SONIA 2): an international, multicentre, open-label, randomised controlled trial.

BACKGROUND: Alkaptonuria is a rare, genetic, multisystem disease characterised by the accumulation of homogentisic acid (HGA). No HGA-lowering therapy has been approved to date. The aim of SONIA 2 was to investigate the efficacy and safety of once-daily nitisinone for reducing HGA excretion in patients with alkaptonuria and to evaluate whether nitisinone has a clinical benefit. METHODS: SONIA 2 was a 4-year, open-label, evaluator-blind, randomised, no treatment controlled, parallel-group study done at three sites in the UK, France, and Slovakia. Patients aged 25 years or older with confirmed alkaptonuria and any clinical disease manifestations were randomly assigned (1:1) to receive either oral nitisinone 10 mg daily or no treatment. Patients could not be masked to treatment due to colour changes in the urine, but the study was evaluator-blinded as far as possible. The primary endpoint was daily urinary HGA excretion (u-HGA24) after 12 months. Clinical evaluation Alkaptonuria Severity Score Index (cAKUSSI) score was assessed at 12, 24, 36, and 48 months. Efficacy variables were analysed in all randomly assigned patients with a valid u-HGA24 measurement at baseline. Safety variables were analysed in all randomly assigned patients. The study was registered at ClinicalTrials.gov (NCT01916382). FINDINGS: Between May 7, 2014, and Feb 16, 2015, 139 patients were screened, of whom 138 were included in the study, with 69 patients randomly assigned to each group. 55 patients in the nitisinone group and 53 in the control group completed the study. u-HGA24 at 12 months was significantly decreased by 99·7% in the nitisinone group compared with the control group (adjusted geometric mean ratio of nitisinone/control 0·003 [95% CI 0·003 to 0·004], p<0·0001). At 48 months, the increase in cAKUSSI score from baseline was significantly lower in the nitisinone group compared with the control group (adjusted mean difference -8·6 points [-16·0 to -1·2], p=0·023). 400 adverse events occurred in 59 (86%) patients in the nitisinone group and 284 events occurred in 57 (83%) patients in the control group. No treatment-related deaths occurred. INTERPRETATION: Nitisinone 10 mg daily was well tolerated and effective in reducing urinary excretion of HGA. Nitisinone decreased ochronosis and improved clinical signs, indicating a slower disease progression. FUNDING: European Commission Seventh Framework Programme.

Homogentisic acid is not only eliminated by glomerular filtration and tubular secretion but also produced in the kidney in alkaptonuria.

The clinical effects of alkaptonuria (AKU) are delayed and ageing influences disease progression. Morbidity of AKU is secondary to high circulating homogentisic acid (HGA) and ochronosis. It is not known whether HGA is produced by or processed in the kidney in AKU. Data from AKU patients from four studies were merged to form a single AKU group. A control group of non-AKU subjects was generated by merging data from two non-AKU studies. Data were used to derive renal clearance and fractional excretion (FE) ratios for creatinine, HGA, phenylalanine (PHE) and tyrosine (TYR) using standard calculations, for comparison between the AKU and the control groups. There were 225 AKU patients in the AKU group and 52 in the non-AKU control group. Circulating HGA increased with age (P < 0.001), and was significantly associated with decreased HGA clearance (CLHGA ) (P < 0.001) and FEHGA (P < 0.001). CLHGA and FEHGA were increased beyond the theoretical maximum renal plasma flow, confirming renal production and emphasising the greater contribution of net tubular secretion than glomerular filtration to renal elimination of HGA. The kidneys are crucial to elimination of HGA. Elimination of HGA is impaired with age resulting in worsening disease over time. The kidney is an important site for production of HGA. Tubular secretion of HGA contributes more to elimination of HGA in AKU than glomerular filtration.

Homogentisate 1,2-dioxygenase (HGD) gene variants, their analysis and genotype-phenotype correlations in the largest cohort of patients with AKU.

Alkaptonuria (AKU) is a rare metabolic disorder caused by a deficient enzyme in the tyrosine degradation pathway, homogentisate 1,2-dioxygenase (HGD). In 172 AKU patients from 39 countries, we identified 28 novel variants of the HGD gene, which include three larger genomic deletions within this gene discovered via self-designed multiplex ligation-dependent probe amplification (MLPA) probes. In addition, using a reporter minigene assay, we provide evidence that three of eight tested variants potentially affecting splicing cause exon skipping or cryptic splice-site activation. Extensive bioinformatics analysis of novel missense variants, and of the entire HGD monomer, confirmed mCSM as an effective computational tool for evaluating possible enzyme inactivation mechanisms. For the first time for AKU, a genotype-phenotype correlation study was performed for the three most frequent HGD variants identified in the Suitability Of Nitisinone in Alkaptonuria 2 (SONIA2) study. We found a small but statistically significant difference in urinary homogentisic acid (HGA) excretion, corrected for dietary protein intake, between variants leading to 1% or >30% residual HGD activity. There was, interestingly, no difference in serum levels or absolute urinary excretion of HGA, or clinical symptoms, indicating that protein intake is more important than differences in HGD variants for the amounts of HGA that accumulate in the body of AKU patients.

Non randomized study on the potential of nitisinone to inhibit cytochrome P450 2C9, 2D6, 2E1 and the organic anion transporters OAT1 and OAT3 in healthy volunteers.

PURPOSE: Nitisinone inhibits the cytochrome P450 (CYP) subfamilies CYP2C9, CYP2D6, and CYP2E1 and the organic anion transporter (OAT) isoforms OAT1 and OAT3 in vitro. Since the effect of nitisinone on these enzymes and transporters in humans is still unknown, the purpose of this study was to evaluate the effect of nitisinone on these CYP subfamilies and OAT isoforms. METHODS: This was an open-label, nonrandomized, two-arm, phase 1 study (EudraCT: 2016-004297-17) in healthy volunteers. The substrates (tolbutamide, metoprolol, and chlorzoxazone for the respective CYPs and furosemide for the OATs) were administered as single doses, before and after 15 days of once daily dosing of 80 mg nitisinone, to determine the AUC∞ ratios ([substrate+nitisinone]/[substrate]). Nitisinone pharmacokinetics, safety, and tolerability were also assessed, and blood and urine were collected to determine substrate and nitisinone concentrations by LC-MS/MS. RESULTS: Thirty-six subjects were enrolled with 18 subjects included in each arm. The least square mean ratio (90% confidence interval) for AUC∞ was 2.31 (2.11-2.53) for tolbutamide, 0.95 (0.88-1.03) for metoprolol, 0.73 (0.67-0.80) for chlorzoxazone, and 1.72 (1.63-1.81) for furosemide. Clinically relevant nitisinone steady-state concentrations were reached after 12 days: mean Cav,ss of 94.08 µM. All treatments were well tolerated, and no safety concerns were identified. CONCLUSIONS: Nitisinone did not affect CYP2D6 activity, was a weak inducer of CYP2E1, and was a weak inhibitor of OAT1 and OAT3. Nitisinone was a moderate inhibitor of CYP2C9, and treatment may therefore result in increased plasma concentrations of comedications metabolized primarily via this enzyme. CLINICAL TRIAL REGISTRY IDENTIFICATION: EudraCT 2016-004297-17.

Open-Label Single-Sequence Crossover Study Evaluating Pharmacokinetics, Efficacy, and Safety of Once-Daily Dosing of Nitisinone in Patients with Hereditary Tyrosinemia Type 1.

BACKGROUND: Although nitisinone is successfully used to treat hereditary tyrosinemia type 1 (HT-1) with the recommended twice-daily dosing, data describing a long half-life motivate less frequent dosing. Therefore, in agreement with the Pharmacovigilance Risk Assessment Committee at the European Medicines Agency, this study was performed to investigate the switch to once-daily dosing. METHODS: This open-label, non-randomized, single-sequence crossover study evaluated the pharmacokinetics, efficacy, and safety of once-daily compared to twice-daily dosing of nitisinone in patients with HT-1 (NCT02323529). Well-controlled patients of <2, 2 to <12, 12 to <18, and ≥18 years of age who were on twice-daily dosing were eligible for participation. Nitisinone and succinylacetone levels were determined from dry blood spots by tandem mass spectrometry. The primary endpoint was Cmin of nitisinone after ≥4 weeks of treatment on each dosing regimen. Secondary objectives were evaluation of efficacy and safety during each dosing regimen. RESULTS: In total, 19 patients were enrolled and 17 included in the per-protocol analysis set. The mean (SD) nitisinone Cmin decreased by 23%, from 26.4 (10.2) to 21.2 (9.9) µmol/L in dry blood spot samples (not equivalent to plasma concentrations), when patients switched from twice- to once-daily dosing. There was no apparent age- or bodyweight-related trend in the degree of Cmin decrease. No patient had quantifiable succinylacetone levels during the once-daily treatment period, indicating efficacious treatment. All adverse events were mild or moderate and judged unrelated to nitisinone. CONCLUSION: The switch to once-daily treatment with nitisinone appeared efficacious and safe in the treatment of patients with HT-1.

Safety, Pharmacokinetics, and Efficacy of Palifermin in Children and Adolescents with Acute Leukemias Undergoing Myeloablative Therapy and Allogeneic Hematopoietic Stem Cell Transplantation: A Pediatric Blood and Marrow Transplant Consortium Trial.

Currently, effective pharmacologic treatment to reduce severe oral mucositis (OM) resulting from high-dose myeloablative cytotoxic therapy in the pediatric population is not available. Palifermin has been proven to decrease the incidence and duration of severe OM in adults with hematologic malignancies undergoing hematopoietic stem cell transplantation (HSCT). In the pediatric population, however, data on palifermin treatment are limited. A phase I dose-escalation study of palifermin in pediatric patients with acute leukemias undergoing myeloablative HSCT with total body irradiation, etoposide, and cyclophosphamide was performed to determine a safe and tolerable dose and to characterize the pharmacokinetic (PK) profile and efficacy of palifermin. Twenty-seven patients in 3 age groups (1 to 2, 3 to 11, and 12 to 16 years) and 3 dose levels (40, 60, and 80 µg/kg/day) were studied. There were no deaths, dose-limiting toxicities, or treatment-related serious adverse events. Long-term safety outcomes did not differ from what would be expected in this population. PK data showed no differences between the 3 age groups. Exposure did not increase with increase in dose. The maximum severity of OM (WHO grade 4) occurred in 6 patients (22%), none of whom was in the 80-µg/kg/day dosing group. This study showed that all doses were well tolerated and a good safety profile in all 3 pediatric age groups was seen.

Twelve novel HGD gene variants identified in 99 alkaptonuria patients: focus on 'black bone disease' in Italy.

Alkaptonuria (AKU) is an autosomal recessive disorder caused by mutations in homogentisate-1,2-dioxygenase (HGD) gene leading to the deficiency of HGD enzyme activity. The DevelopAKUre project is underway to test nitisinone as a specific treatment to counteract this derangement of the phenylalanine-tyrosine catabolic pathway. We analysed DNA of 40 AKU patients enrolled for SONIA1, the first study in DevelopAKUre, and of 59 other AKU patients sent to our laboratory for molecular diagnostics. We identified 12 novel DNA variants: one was identified in patients from Brazil (c.557T>A), Slovakia (c.500C>T) and France (c.440T>C), three in patients from India (c.469+6T>C, c.650-85A>G, c.158G>A), and six in patients from Italy (c.742A>G, c.614G>A, c.1057A>C, c.752G>A, c.119A>C, c.926G>T). Thus, the total number of potential AKU-causing variants found in 380 patients reported in the HGD mutation database is now 129. Using mCSM and DUET, computational approaches based on the protein 3D structure, the novel missense variants are predicted to affect the activity of the enzyme by three mechanisms: decrease of stability of individual protomers, disruption of protomer-protomer interactions or modification of residues in the region of the active site. We also present an overview of AKU in Italy, where so far about 60 AKU cases are known and DNA analysis has been reported for 34 of them. In this rather small group, 26 different HGD variants affecting function were described, indicating rather high heterogeneity. Twelve of these variants seem to be specific for Italy.

Suitability Of Nitisinone In Alkaptonuria 1 (SONIA 1): an international, multicentre, randomised, open-label, no-treatment controlled, parallel-group, dose-response study to investigate the effect of once daily nitisinone on 24-h urinary homogentisic acid excretion in patients with alkaptonuria after 4 weeks of treatment.

BACKGROUND: Alkaptonuria (AKU) is a serious genetic disease characterised by premature spondyloarthropathy. Homogentisate-lowering therapy is being investigated for AKU. Nitisinone decreases homogentisic acid (HGA) in AKU but the dose-response relationship has not been previously studied. METHODS: Suitability Of Nitisinone In Alkaptonuria 1 (SONIA 1) was an international, multicentre, randomised, open-label, no-treatment controlled, parallel-group, dose-response study. The primary objective was to investigate the effect of different doses of nitisinone once daily on 24-h urinary HGA excretion (u-HGA24) in patients with AKU after 4 weeks of treatment. Forty patients were randomised into five groups of eight patients each, with groups receiving no treatment or 1 mg, 2 mg, 4 mg and 8 mg of nitisinone. FINDINGS: A clear dose-response relationship was observed between nitisinone and the urinary excretion of HGA. At 4 weeks, the adjusted geometric mean u-HGA24 was 31.53 mmol, 3.26 mmol, 1.44 mmol, 0.57 mmol and 0.15 mmol for the no treatment or 1 mg, 2 mg, 4 mg and 8 mg doses, respectively. For the most efficacious dose, 8 mg daily, this corresponds to a mean reduction of u-HGA24 of 98.8% compared with baseline. An increase in tyrosine levels was seen at all doses but the dose-response relationship was less clear than the effect on HGA. Despite tyrosinaemia, there were no safety concerns and no serious adverse events were reported over the 4 weeks of nitisinone therapy. CONCLUSIONS: In this study in patients with AKU, nitisinone therapy decreased urinary HGA excretion to low levels in a dose-dependent manner and was well tolerated within the studied dose range. TRIAL REGISTRATION NUMBER: EudraCT number: 2012-005340-24. Registered at ClinicalTrials.gov: NCTO1828463.

Pharmacokinetic and pharmacodynamic interactions between palifermin and heparin.

Oral mucositis, a severe complication during chemo- and/or radiotherapy, is prevented with palifermin treatment, a recombinant human keratinocyte growth factor (KGF/FGF-7). The FGF family belongs to the larger family of heparin-binding growth factors. Because it has been shown that heparin modulates binding of KGF to the KGF receptor and subsequently affects cellular proliferation induced by the KGF mitogenic signal, it is critical to understand the drug-drug interactions between palifermin and heparin, particularly because of heparin's narrow therapeutic margin. Two studies were performed in healthy subjects to characterize the effect of palifermin on the pharmacodynamics of heparin (activated partial thromboplastin time) and evaluate the impact of heparin on the pharmacokinetics and pharmacodynamics (Ki67 staining of buccal mucosal tissue) of palifermin. Results demonstrated a pronounced pharmacokinetic interaction; heparin coadministration increased the palifermin AUC 4- to 5-fold and decreased its half-life by 40%-45%, suggesting an approximate 70%-80% decrease in palifermin clearance and volume of distribution. These changes in the pharmacokinetics of palifermin during coadministration of heparin, however, did not affect the pharmacodynamic effect of palifermin, or the anticoagulant activity of heparin, and did not lead to increased safety findings. Therefore, these results suggest that dose adjustments for heparin and palifermin are not warranted when administered concurrently.

Relationship Between Serum Concentrations of Nitisinone and Its Effect on Homogentisic Acid and Tyrosine in Patients with Alkaptonuria.

BACKGROUND: Alkaptonuria (AKU) is a serious genetic disease due to a defect in tyrosine metabolism, leading to increased serum levels of homogentisic acid (HGA). Nitisinone decreases HGA in AKU, but the concentration-response relationship has not been previously reported. OBJECTIVES: To determine the relationship between serum concentrations of nitisinone and the effect on both HGA and tyrosine; secondly to determine steady-state pharmacokinetics of nitisinone in AKU patients. METHOD: Thirty-two patients with AKU received either 1, 2, 4, or 8 mg nitisinone daily. Urine and serum HGA and serum tyrosine and nitisinone were measured during 24 h at baseline (before first dose) and after 4 weeks of treatment. RESULTS: Nitisinone pharmacokinetics (area under the curve [AUC] and maximum concentrations [C max]) were dose proportional. The median oral clearance determined in all patients, irrespective of dose, was 3.18 mL/h·kg (range 1.6-6.7).Nitisinone decreased urinary excretion of HGA in a concentration-dependent manner, with a maximum effect seen at average nitisinone concentrations of 3 µmol/L. The association between nitisinone and tyrosine concentrations was less pronounced. Serum levels of HGA at Week 4 were below the limit of quantitation in 65% of samples, which prevented determination of the relationship with nitisinone concentrations. CONCLUSION: Nitisinone exhibits dose-proportional pharmacokinetics in the studied dosage interval. Urinary excretion of HGA decreases in a concentration-dependent manner, while the increase in tyrosine is less clearly related to nitisinone concentrations.

rhBSSL improves growth and LCPUFA absorption in preterm infants fed formula or pasteurized breast milk.

OBJECTIVES: Preterm infants often experience suboptimal growth, which can affect organ development. The aim of this study was to improve growth by treatment with bile salt-stimulated lipase (BSSL), naturally present in breast milk, but lost after pasteurization, and absent in formula. METHODS: Two clinical trials were performed with a predefined analysis of combined data to investigate the effects of recombinant human BSSL (rhBSSL) treatment on growth velocity and fat absorption in preterm infants. The studies were randomized and double-blinded comparing 7-day treatment with rhBSSL and placebo, administered in pasteurized breast milk or formula, using a crossover design. RESULTS: Sixty-three infants were evaluated for safety. At randomization, the mean (standard deviation) weight was 1467 (193) g and mean postmenstrual age was 32.6 (0.5) weeks. Sixty and 46 infants were evaluated for growth velocity and fat absorption, respectively. rhBSSL treatment significantly improved mean growth velocity by 2.93 g · kg · day (P<0.001) compared with placebo (mean 16.86 vs 13.93 g · kg · day) and significantly decreased the risk of suboptimal growth (<15 g · kg · day) (30% vs 52%, P=0.004). rhBSSL significantly increased absorption of the long-chain polyunsaturated fatty acids, docosahexaenoic acid, and arachidonic acid by 5.76% (P=0.013) and 8.55% (P=0.001), respectively, but had no significant effect on total fat absorption. The adverse-event profile was similar to placebo. CONCLUSIONS: In preterm infants fed pasteurized breast milk or formula, 1 week of treatment with rhBSSL was well tolerated and significantly improved growth and long-chain polyunsaturated fatty acid absorption compared to placebo. This publication presents the first data regarding the use of rhBSSL in preterms and the results have led to further clinical studies.

A shorter workday as a means of reducing the occurrence of musculoskeletal disorders.

OBJECTIVES: The study examined the relation between daily workhours and the occurrence of neck-shoulder or back pain in physically demanding care work. METHODS: Unpublished data were obtained from three intervention projects in care institutions. The projects had been conducted independently in Oslo (46 participants, 175 referents before and 158 referents after the intervention), Helsingborg (60 participants, 89 referents) and Stockholm (41 participants, 22 referents) between 1995 and 1998. The intervention was a reduction of daily workhours from > or = 7 to 6 hours (or 30 hours weekly). Full-time salary was retained, and extra personnel were employed to compensate for the reduction in workhours. Data were collected by self-administered questionnaires before and during the intervention periods, lasting from 12 to 22 months. RESULTS: The prevalence of neck-shoulder pain decreased from 40.9% to 25.6% in Oslo and from 57.1% to 39.1% in Helsingborg after 1.5 years with a 6-hour workday; for Stockholm the decrease was from 81.6% to 68.3% after 1 year. No decrease was observed in the reference groups. The prevalence of back pain did not show the same consistent pattern. CONCLUSIONS: The shortening of regular workdays from > or = 7 hours to 6 hours may considerably reduce the prevalence of neck-shoulder pain among persons with physically demanding care work. The potential health benefits should encourage intervention studies also in other occupations with increased risk of work-related musculoskeletal disorders.