Stability of porphyrins and porphyrin precursors in urine and plasma samples: implications for sample handling and storage.

The porphyrias are rare disorders of haem biosynthesis. Diagnosis requires demonstrating increased porphyrins or porphyrin precursors in blood, urine and faeces. Patients may only be investigated once, and therefore, understanding the preanalytical factors affecting the reliability of results is crucial. Guidance for sample handling exists, but published evidence regarding the stability of porphyrins and their precursors is limited. The aim of this study was to evaluate the effect of light exposure and different storage temperatures on analyte stability for measurement of urinary aminolaevulinic acid and porphobilinogen, total urine porphyrin and plasma porphyrin. Our results confirm that all samples should be protected from light. Results from samples exposed to light for greater than 4 hours should be interpreted with caution and repeat samples requested. If transported to a specialist laboratory, samples should be stored at 4°C before transport. Transit time at ambient temperatures should be less than 24 hours.

Investigation of the relationship between phenylalanine in venous plasma and capillary blood using volumetric blood collection devices.

Measurement of plasma and dried blood spot (DBS) phenylalanine (Phe) is key to monitoring patients with phenylketonuria (PKU). The relationship between plasma and capillary DBS Phe concentrations has been investigated previously, however, differences in methodology, calibration approach and assumptions about the volume of blood in a DBS sub-punch has complicated this. Volumetric blood collection devices (VBCDs) provide an opportunity to re-evaluate this relationship. Paired venous and capillary samples were collected from patients with PKU (n = 51). Capillary blood was collected onto both conventional newborn screening (NBS) cards and VBCDs. Specimens were analysed by liquid-chromatography tandem mass-spectrometry (LC-MS/MS) using a common calibrator. Use of VBCDs was evaluated qualitatively by patients. Mean bias between plasma and volumetrically collected capillary DBS Phe was -13%. Mean recovery (SD) of Phe from DBS was 89.4% (4.6). VBCDs confirmed that the volume of blood typically assumed to be present in a 3.2 mm sub-punch is over-estimated by 9.7%. Determination of the relationship between plasma and capillary DBS Phe, using a single analytical method, common calibration and VBCDs, demonstrated that once the under-recovery of Phe from DBS has been taken into account, there is no significant difference in the concentration of Phe in plasma and capillary blood. Conversely, comparison of plasma Phe with capillary DBS Phe collected on a NBS card highlighted the limitations of this approach. Introducing VBCDs for the routine monitoring of patients with PKU would provide a simple, acceptable specimen collection technique that ensures consistent sample quality and produces accurate and precise blood Phe results which are interchangeable with plasma Phe.

Update on the diagnosis and management of the autosomal dominant acute hepatic porphyrias.

The autosomal dominant acute hepatic porphyrias (AHPs), acute intermittent porphyria, hereditary coproporphyria (HCP) and variegate porphyria (VP), are low penetrance adult onset disorders caused by partial deficiency of enzymes of haem biosynthesis. All are associated with acute neurovisceral attacks, which are a consequence of the increased hepatic demand for haem triggered by hormones, stress, drugs or systemic infections which leads to upregulation of the pathway and overproduction of haem precursors 5-aminolaevulinic acid (ALA) and porphobilinogen (PBG). Acute episodes are characterised by severe abdominal pain, nausea, vomiting, hyponatraemia, hypertension and tachycardia, behavioural disturbance and can progress to include seizures, peripheral motor neuropathy and posterior reversible encephalopathy syndrome if undiagnosed and untreated. VP and HCP may also present with photocutaneous skin lesions either alone or during acute symptoms. Diagnosis involves demonstrating increased excretion of PBG in urine. Treatment focuses on removing or managing triggers, supportive treatment and suppressing the hepatic haem pathway by administering human haemin. Chronic complications include hypertension, chronic kidney disease and hepatocellular carcinoma. A small proportion of symptomatic patients with AHP progress to repeated acute attacks which require preventative therapy. A new RNA interference therapy has recently been licensed and is likely to become the treatment of choice in this situation.

Performance of laboratory tests used to measure blood phenylalanine for the monitoring of patients with phenylketonuria.

Analysis of blood phenylalanine is central to the monitoring of patients with phenylketonuria (PKU) and age-related phenylalanine target treatment-ranges (0-12 years; 120-360 µmol/L, and >12 years; 120-600 µmol/L) are recommended in order to prevent adverse neurological outcomes. These target treatment-ranges are based upon plasma phenylalanine concentrations. However, patients are routinely monitored using dried bloodspot (DBS) specimens due to the convenience of collection. Significant differences exist between phenylalanine concentrations in plasma and DBS, with phenylalanine concentrations in DBS specimens analyzed by flow-injection analysis tandem mass spectrometry reported to be 18% to 28% lower than paired plasma concentrations analyzed using ion-exchange chromatography. DBS specimens with phenylalanine concentrations of 360 and 600 µmol/L, at the critical upper-target treatment-range thresholds would be plasma equivalents of 461 and 768 µmol/L, respectively, when a reported difference of 28% is taken into account. Furthermore, analytical test imprecision and bias in conjunction with pre-analytical factors such as volume and quality of blood applied to filter paper collection devices to produce DBS specimens affect the final test results. Reporting of inaccurate patient results when comparing DBS results to target treatment-ranges based on plasma concentrations, together with inter-laboratory imprecision could have a significant impact on patient management resulting in inappropriate dietary change and potentially adverse patient outcomes. This review is intended to provide perspective on the issues related to the measurement of phenylalanine in blood specimens and to provide direction for the future needs of PKU patients to ensure reliable monitoring of metabolic control using the target treatment-ranges.

An Audit of the Use of Gonadorelin Analogues to Prevent Recurrent Acute Symptoms in Patients with Acute Porphyria in the United Kingdom.

Severe recurrent acute attacks of porphyria have traditionally been treated with either prophylactic human haemin or gonadorelin analogues (GnA) in females. Evidence on the most effective treatment for this patient subgroup is lacking. This audit surveyed the use of prophylactic GnA in the UK.Twenty female patients (who experienced between 2 and 45 acute attacks of porphyria requiring hospitalisation and treatment with human haemin prior to GnA prophylaxis) were included in the audit. Data was retrospectively collected based on patient history and case review.Twenty three treatment courses were given lasting a median period of 12 months. Monthly subcutaneous Goserelin was most commonly used. In three patients in whom timing with the menstrual cycle was not considered, an acute attack occurred after initiation of the first dose. The majority of patients experienced oestrogen deficiency symptoms during treatment. Fifty percent of the prescribed courses of GnA resulted in a degree of clinical benefit. This successfully treated group experienced between 3 and 20 acute attacks prior to and between 0 and 6 acute attacks during GnA treatment.The audit revealed large variation in practice in the United Kingdom regarding indication, duration of treatment, specific drug used and management of side effects. In view of the limited treatment options available for this cohort and the mixed outcome successes reported, we believe it is reasonable for porphyria specialists to continue offering GnA treatment to women with severe recurrent debilitating acute attacks of porphyria associated with the menstrual cycle, and we propose best practice guidelines to standardise management.

The cutaneous porphyrias.

The porphyrias are a group of mainly inherited disorders of heme biosynthesis where accumulation of porphyrins and/or porphyrin precursors gives rise to 2 types of clinical presentation: cutaneous photosensitivity and/or acute neurovisceral attacks. The cutaneous porphyrias present with either bullous skin fragility or nonbullous acute photosensitivity. This review discusses the epidemiology, pathogenesis, clinical presentation, laboratory diagnosis, complications, and current approach to porphyria management. Although focusing mainly on their dermatological aspects, the article also covers the management of acute porphyria, which by virtue of its association with variegate porphyria and hereditary coproporphyria, may become the responsibility of the clinical dermatologist.

The impact of local guidelines on the tumour marker requesting patterns of a General Surgery Department.

BACKGROUND: The inappropriate use of tumour markers (TMs) is a common problem. The aim of this audit was to evaluate the impact of local guidelines on the TM requesting patterns of a General Surgery Department. METHODS: CA 125, CA 19-9, CA15-3, CEA, AFP and HCG requests from all hospital surgical locations were audited over two periods of eight months before and after the implementation of local requesting guidelines. RESULTS: Postintervention, total TM requests decreased by 32% while patient requests decreased by 9.8%. Single TM requesting increased and requests for panels containing four or more TMs decreased from 279 to 60 requests (78% reduction). CONCLUSION: Interdepartmental collaboration and the implementation of local guidelines have resulted in a change in requesting behaviour, most notably a reduction in multiple TM panel requests.