The first pilot study of expanded newborn screening for inborn errors of metabolism and survey of related knowledge and opinions of health care professionals in Hong Kong.

INTRODUCTION: Newborn screening is important for early diagnosis and effective treatment of inborn errors of metabolism (IEM). In response to a 2008 coroners' report of a 14-year-old boy who died of an undiagnosed IEM, the OPathPaed service model was proposed. In the present study, we investigated the feasibility of the OPathPaed model for delivering expanded newborn screening in Hong Kong. In addition, health care professionals were surveyed on their knowledge and opinions of newborn screening for IEM. METHODS: The present prospective study involving three regional hospitals was conducted in phases, from 1 October 2012 to 31 August 2014. The 10 steps of the OPathPaed model were evaluated: parental education, consent, sampling, sample dispatch, dried blood spot preparation and testing, reporting, recall and counselling, confirmation test, treatment and monitoring, and cost-benefit analysis. A fully automated online extraction system for dried blood spot analysis was also evaluated. A questionnaire was distributed to 430 health care professionals by convenience sampling. RESULTS: In total, 2440 neonates were recruited for newborn screening; no true-positive cases were found. Completed questionnaires were received from 210 respondents. Health care professionals supported implementation of an expanded newborn screening for IEM. In addition, there is a substantial need of more education for health care professionals. The majority of respondents supported implementing the expanded newborn screening for IEM immediately or within 3 years. CONCLUSION: The feasibility of OPathPaed model has been confirmed. It is significant and timely that when this pilot study was completed, a government-led initiative to study the feasibility of newborn screening for IEM in the public health care system on a larger scale was announced in the Hong Kong Special Administrative Region Chief Executive Policy Address of 2015.

Clinical utility of late-night and post-overnight dexamethasone suppression salivary cortisone for the investigation of Cushing's syndrome.

INTRODUCTION: There is a pressing need to identify diagnostic testing for Cushing's syndrome that can be achieved with ease and at low cost. This study aimed to explore the usefulness of late-night and post-overnight 1-mg dexamethasone suppression salivary cortisone, as measured by liquid chromatography-tandem mass spectrometry, for investigation of hypercortisolism. METHODS: Salivary cortisone data of subjects were investigated according to a pre-specified protocol. Subjects were classified as having 'hypercortisolism' or 'eucortisolism' on the basis of histological or biochemical criteria. Receiver operating characteristic curves were drawn to identify the cut-off values and study their performance characteristics. We measured 24-hour urinary free cortisol; late-night salivary cortisol and cortisone; and post-overnight 1-mg dexamethasone suppression serum cortisol, and salivary cortisol and cortisone. Saliva and urine samples were assayed by liquid chromatography-tandem mass spectrometry. RESULTS: In this study, 21 subjects were classified as having hypercortisolism and 78 as having eucortisolism. A late-night salivary cortisone cut-off of 13.50 nmol/L had a sensitivity of 94.7% and a specificity of 87.2%. After taking 1-mg dexamethasone the night before, a salivary cortisol cut-off of 0.85 nmol/L had a sensitivity of 76.2% and a specificity of 96.2%; a salivary cortisone cut-off of 7.45 nmol/L had a sensitivity of 85.7% and a specificity of 94.9%, while a salivary cortisone cut-off of 3.25 nmol/L had a sensitivity of 95.2% and a specificity of 79.5%. Many salivary cortisol samples were below the detection limit of liquid chromatography-tandem mass spectrometry. In comparison with salivary cortisol, salivary cortisone had a better correlation with total serum cortisol and better diagnostic performance following dexamethasone suppression. CONCLUSIONS: Both late-night and post-overnight dexamethasone suppression salivary cortisone levels are of diagnostic value in the investigation of hypercortisolism.

Aetiological bases of 46,XY disorders of sex development in the Hong Kong Chinese population.

OBJECTIVE: Disorders of sex development are due to congenital defects in chromosomal, gonadal, or anatomical sex development. The objective of this study was to determine the aetiology of this group of disorders in the Hong Kong Chinese population. SETTING: Five public hospitals in Hong Kong. PATIENTS: Patients with 46,XY disorders of sex development under the care of paediatric endocrinologists between July 2009 and June 2011. MAIN OUTCOME MEASURES: Measurement of serum gonadotropins, adrenal and testicular hormones, and urinary steroid profiling. Mutational analysis of genes involved in sexual differentiation by direct DNA sequencing and multiplex ligation-dependent probe amplification. RESULTS: Overall, 64 patients were recruited for the study. Their age at presentation ranged from birth to 17 years. The majority presented with ambiguous external genitalia including micropenis and severe hypospadias. A few presented with delayed puberty and primary amenorrhea. Baseline and post-human chorionic gonadotropin-stimulated testosterone and dihydrotestosterone levels were not discriminatory in patients with or without AR gene mutations. Of the patients, 22 had a confirmed genetic disease, with 11 having 5α-reductase 2 deficiency, seven with androgen insensitivity syndrome, one each with cholesterol side-chain cleavage enzyme deficiency, Frasier syndrome, NR5A1-related sex reversal, and persistent Müllerian duct syndrome. CONCLUSIONS: Our findings suggest that 5α-reductase 2 deficiency and androgen insensitivity syndrome are possibly the two most common causes of 46,XY disorders of sex development in the Hong Kong Chinese population. Since hormonal findings can be unreliable, mutational analysis of the SRD5A2 and AR genes should be considered the first-line tests for these patients.

Late presentation of simple virilising 21-hydroxylase deficiency in a Chinese woman with Turner's syndrome.

Classical congenital adrenal hyperplasia due to 21-hydroxylase deficiency is a well-known disorder of sexual development (previously known as ambiguous genitalia) in genotypic female neonates. We report on a 66-year-old Chinese, brought up as male, with a simple virilising form of congenital adrenal hyperplasia associated with Turner's syndrome (karyotype 45,X/47,XXX/46,XX). His late presentation was recognised due to his exceptionally short stature and persistent sexual ambiguity. His condition was only brought to medical attention as he developed a huge abdominal mass, which later turned out to be a benign ovarian mucinous cyst. It is therefore important to look out for co-existing congenital adrenal hyperplasia in patients with Turner's syndrome and virilisation, after the presence of Y chromosome material has been excluded.

Urinary steroid profiling in the diagnosis of congenital adrenal hyperplasia and disorders of sex development: experience of a urinary steroid referral centre in Hong Kong.

BACKGROUND: Deficiency in any one of the steroidogenic enzymes may result in congenital adrenal hyperplasia (CAH) and disorders of sex development (DSD). Urinary steroid profiling (USP) can quantify metabolites of all relevant steroids simultaneously in a single analysis and has established clinical applications in the investigation and diagnosis in these disorders. PATIENTS AND METHODS: A retrospective review was performed on all the samples sent to the Chemical Pathology Laboratory, Queen Elizabeth Hospital, Hong Kong, for the investigation of suspected disorders in steroid metabolism by USP between 2003 and 2011. RESULTS: 432 patients had urine samples sent to our laboratory for USP for the investigation of CAH and DSD in the review period. USP showed diagnostic pattern of 21-hydroxylase deficiency (n=21), 5α-reductase 2 deficiency (n=12), 17α-hydroxylase deficiency (n=3), isolated 17,20-lyase deficiency (n=1), 11ß-hydroxylase deficiency (n=1) and P450 oxidoreductase deficiency (n=1). CONCLUSIONS: 21-hydroxylase deficiency is the most common form of CAH while 5α-reductase 2 deficiency is the most common cause of 46,XY DSD in our population. USP is a useful tool in the investigation and diagnosis of CAH and DSD due to different steroidogenesis defects and should be included as a first-line endocrine investigation in this group of patients.

Hypertriglyceridaemia-induced pancreatitis: a contributory role of capecitabine?

Capecitabine is an orally administered pro-drug of 5-fluorouracil that confers superior disease-free survival and presumably has a more favourable side-effect profile. Here we report on a patient who developed acute necrotising pancreatitis and very high triglyceride levels as well as hand-foot syndrome after receiving capecitabine for colonic cancer. Increased awareness of this potential side-effect and close monitoring of lipid levels may be warranted, especially in patients who have other conditions predisposing them to severe secondary hyperlipidaemia when using this drug.

Hyperammonaemic encephalopathy in an adult patient with citrin deficiency associated with a novel mutation.

We report on an adult patient with citrin deficiency in Hong Kong, in whom a novel mutation was identified. The patient presented with recurrent hyperammonaemic encephalopathy due to impairment of the liver urea cycle enzyme argininosuccinate synthetase. This autosomal recessive condition is also characterised by interesting food preferences, notably aversion to carbohydrates and craving for protein-rich and/or lipid-rich foods, as well as neuropsychiatric symptoms. Plasma amino acid analysis is very useful in revealing urea cycle disorders, and mutational analysis of the SLC25A13 gene can confirm the diagnosis.

Enzyme replacement therapy for mucopolysaccharidosis VI (Maroteaux-Lamy syndrome): experience in Hong Kong.

Mucopolysaccharidosis VI (Maroteaux-Lamy syndrome) is a very rare inherited lysosomal storage disease. We evaluated the efficacy and safety of weekly infusions of recombinant human arylsulfatase B as enzyme replacement therapy for two patients in whom this condition was advanced. The primary outcome variables were the distance walked in a 6-minute walk test, forced vital capacity, and ejection fraction. The secondary outcome variables were the number of stairs climbed in a 3-minute stair climbing test, joint mobility, urinary glycosaminoglycan excretion, auto-continuous positive airway pressure study and liver size. After 24 weeks of treatment, patient A walked 40 m (36%) and patient B walked 66 m (58%) more in the walk test than at baseline. After 48 weeks, in patient A the corresponding improvements were 142 m (129%) in the walk test and 33 stairs (60%) in the 3-minute stair climbing test, and in patient B the respective improvements were 198 m (174%) and 77 stairs (140%). There was a significant decline in urinary glycosaminoglycan excretion and improvement in range of motion of joints in both patients. The auto-continuous positive airway pressure study revealed improvements in patient A, while other efficacy variables remained static. There were no drug-related adverse events or allergic reactions reported during and after the infusions of recombinant human arylsulfatase B. Recombinant human arylsulfatase B significantly improves endurance and reduces urinary glycosaminoglycan excretion. The drug is generally safe and well tolerated.

Molecular analysis of congenital adrenal hyperplasia due to 21-hydroxylase deficiency in Hong Kong Chinese patients.

BACKGROUND: Congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency (21OHD) is an autosomal recessive disorder due to mutation in the CYP21A2 gene. OBJECTIVE: To elucidate the genetic basis of 21-hydroxylase-deficient CAH in Hong Kong Chinese patients. PATIENTS AND METHODS: Mutational analysis of the CYP21A2 gene was performed on 35 Hong Kong Chinese patients with 21OHD using direct DNA sequencing and multiplex ligation-dependent probe amplification (MLPA). RESULTS: The genetic findings of 21 male and 14 female patients are the following: c.293-13A/C>G (intron 2 splice site; 20 alleles), p.I172N (13), p.R356W (7), p.Q318X (4). A total of 20 mutant alleles contained gross deletion/conversion of all or part of the CYP21A2 gene. A novel mutation, c.1367delA (p.D456fs), was detected in one patient. One patient had only a heterozygous mutation detected. Out of 35 patients, 16 would have been incorrectly genotyped if either DNA sequencing or MLPA alone was used for molecular analysis. CONCLUSIONS: The frequency of various mutations in the studied patients differs from those reported in other Asian populations. Gross deletion/conversion accounts for nearly one-third of the genetic defects. Therefore, laboratories must include methods for detecting point mutations as well as gross deletions/conversions to avoid misinterpretation of genotype. Genotyping has increasingly been proven to be a useful tool for supplementing, if not replacing, hormonal profiling for the diagnosis of 21OHD.

The reference interval of thyroid-stimulating hormone in Hong Kong Chinese.

AIM: The upper reference limit of thyroid-stimulating hormone (TSH) is critical for defining patients with subclinical hypothyroidism, a condition which carries a higher risk of progression to overt hypothyroidism and adverse cardiovascular events. Yet, there is a lack of consensus on its absolute value, and data in non-pregnant adult Chinese are lacking. METHODS: Apparently healthy and drug-free local adult Chinese were recruited by completing health questionnaires. Their serum samples were tested for TSH, free thyroxine (FT4), thyroglobulin antibody and thyroid peroxidase antibody levels. After excluding subjects with thyroid antibodies, the TSH level was log-transformed, and the reference limits were defined as mean ± 1.96SD. The 2.5th and 97.5th percentiles of FT4 were also calculated. RESULTS: Serum samples from 212 subjects were used in this study. 51 subjects were seropositive to thyroglobulin antibody, 31 were seropositive to thyroid peroxidase antibody, and 27 were seropositive to both. The reference intervals after excluding subjects seropositive to thyroid antibodies were: TSH: 0.68-3.70 mIU/l; FT4: 13.5-21.3 pmol/l (male) and 12.6-19.7 pmol/l (female). Including subjects with thyroid antibodies only minimally changed the reference intervals of these hormones. CONCLUSION: The authors have set up the reference interval of TSH for the local population, and their findings also suggest that the importance of excluding subjects with thyroid antibodies in the reference population should not be overemphasised. Moreover, the international authorities should consider recommending percentile-equivalent action limits instead of an absolute cut-off on TSH for categorisation of different types of thyroid dysfunction.

Ambiguous genitalia, impaired steroidogenesis, and Antley-Bixler syndrome in a patient with P450 oxidoreductase deficiency.

Cytochrome P450 oxidoreductase deficiency is a recently established autosomal recessive disease characterised by ambiguous genitalia, impaired steroidogenesis, and skeletal malformations, referred to as Antley-Bixler syndrome. Clinical manifestations in affected patients are highly variable. We report on a girl with P450 oxidoreductase deficiency who presented with virilisation at birth. There was transient maternal virilisation during pregnancy as well. She was initially diagnosed with congenital adrenal hyperplasia caused by 21-hydroxylase deficiency and/or aromatase deficiency. At 1 year of age, skeletal abnormalities suggestive of Antley-Bixler syndrome were detected. Molecular analysis of the fibroblast growth factor receptor 2 (FGFR2) gene was normal but POR gene analysis showed that she was homozygous for an R457H missense mutation. The diagnosis, P450 oxidoreductase deficiency, was confirmed. Results of her endocrine studies and urinary steroid profiling are also presented.

Use of urinary steroid profiling for diagnosing and monitoring adrenocortical tumours.

It has been suggested that urinary steroid profiling may be used to provide information aiding the diagnosis and monitoring of adrenocortical carcinoma. Nonetheless, the abnormal patterns suggestive of adrenal malignancy are not well defined. We retrospectively studied the urinary steroid profiles of five patients with adrenocortical carcinoma at presentation and at follow-up, and compared these results with those from 76 patients with benign adrenocortical adenoma and 172 healthy controls. Three abnormal patterns of urinary steroid excretion were identified in patients with adrenocortical carcinoma at presentation and/or follow-up of residual disease: (1) hypersecretion in multiple steroid axes; (2) excretion of unusual metabolites, notably 5-pregnene-3alpha,16alpha,20alpha-triol, 5-pregnene-3beta,16alpha,20alpha-triol, and neonatal steroid metabolites in the post-neonatal period; (3) increase of tetrahydro-11-deoxycortisol relative to total cortisol metabolites. These preliminary findings offer ways in which urinary steroid profiling performed using gas chromatography-mass spectrometry can be helpful in the diagnosis and monitoring of adrenocortical carcinoma.

Diagnosis of 5alpha-reductase 2 deficiency: a local experience.

5Alpha-reductase 2 deficiency is an autosomal recessive disorder characterised by lack of masculinisation in XY individuals due to failure to convert testosterone to dihydrotestosterone, the bioactive androgen. Traditionally, the testosterone-to-dihydrotestosterone ratio is used to diagnose this condition but interpreting these results is not always straightforward, thus they may be inconclusive. On the contrary, urinary steroid profiling unambiguously demonstrates a significantly reduced excretion of 5alpha-reduced steroid metabolites compared to their 5beta counterparts. This analytical technique can also simultaneously confirm or rule out other causes of ambiguous genitalia due to steroidogenic defects. Making a DNA-based diagnosis by studying the SRD5A2 gene has become increasingly popular. Here, we report six Chinese patients from different families who were all diagnosed with 5alpha-reductase 2 deficiency based on urinary steroid profile findings and mutational analysis of the SRD5A2 gene. R227Q was the most commonly identified mutation in these patients. Management of sexual development disorders is also discussed.

A rare cause of nephrotic syndrome: lipoprotein glomerulopathy.

Lipoprotein glomerulopathy is a rare kidney disease in which lipoprotein thrombi are seen in the glomerular capillaries. Most of these patients are found in Japan and East Asian countries. The presenting symptoms include proteinuria, an abnormal plasma lipoprotein profile that resembles type III hyperlipoproteinaemia, and a marked increase in serum apolipoprotein E concentration. Previous studies have suggested that lipoprotein glomerulopathy might be related to APOE gene mutation. No effective therapeutic regimen has been established for lipoprotein glomerulopathy. We report the first case of biopsy-proven lipoprotein glomerulopathy in Hong Kong in a patient who presented with nephrotic syndrome and dyslipidaemia. DNA analysis revealed apolipoprotein E Kyoto together with a novel apolipoprotein E mutation, apolipoprotein E (Asp230Tyr) Hong Kong. There was significant improvement in the clinical parameters and resolution of symptoms after the introduction of statins. Further studies will be needed to clarify the role of apolipoprotein E Hong Kong and its interaction with apolipoprotein E Kyoto in the pathogenesis of lipoprotein glomerulopathy.

Reference intervals of urinary steroid metabolites using gas chromatography-mass spectrometry in Chinese adults.

BACKGROUND: Urinary steroid profiling by GC or GC-MS are established clinical tools to complement other biochemical tests in the diagnosis and investigation of a wide range of adrenocortical disorders, but normative data on adults using the more specific GC-MS are lacking. Our objective was to set up the reference intervals of commonly detected urinary steroid metabolites as well as marker metabolites seen in disease states. METHOD: Apparently healthy adult Chinese males and females were recruited by completing health questionnaires. A 24-h urine specimen was collected from all the participants for urinary steroid profiling by GC-MS in cyclic scan mode. The analyzer was calibrated by using authentic steroid standards. Statistical methods recommended by the National Committee for Clinical Laboratory Standards were followed for setting up the reference intervals of various steroid metabolites. After outliers were excluded, the data were tested for the necessity to partition into sex-, menopausal status- and age-specific reference intervals. RESULTS: 83 males and 89 females were recruited for the study. Necessity to partition into sex-specific reference intervals was demonstrated for almost all steroid metabolites. Menopausal status and age also had a significant impact on steroid metabolite excretion, making separate reference intervals necessary. CONCLUSIONS: We have set up the normative data on the levels of urinary steroid metabolite excretion in Chinese adults for future reference in patient management and research in steroid metabolism.

Reference intervals of common serum analytes of Hong Kong Chinese.

AIM: Setting up reference intervals from the local service populations is one of the major responsibilities of clinical laboratories. Yet, this task is difficult to achieve because it is costly and time consuming when compared with validating reference intervals from assay manufacturers. METHODS: Following the recommendations of the International Federation of Clinical Chemistry, healthy local Chinese adults were recruited to set up reference intervals for common serum analytes. Statistical methods recommended by the National Committee for Clinical Laboratory Standards were used for defining the reference limits. RESULTS: Data from 335 subjects were analysed. The reference intervals set up were broadly similar to those provided by the assay manufacturer, except for sodium and potassium. Glomerular filtration rate was estimated by the modification of diet in renal disease equation, with or without modification for Chinese. Body mass index had a significant impact on serum urate and alanine aminotransferase levels. CONCLUSION: Reference intervals of common serum analytes have been set up for the local Chinese population. A good example of quality laboratory service has also been set up to provide clinicians with reliable reference intervals that they can confidently rely on for the diagnosis and management of patients.

Cryoglobulinaemia: clinical and laboratory perspectives.

Cryoglobulins are immunoglobulins that precipitate in the serum upon cooling to below core body temperature and re-dissolve at higher temperatures. Cryoglobulinaemia may be life-threatening. The three types of cryoglobulinaemia are associated with a wide spectrum of haematological, autoimmune, and chronic infectious diseases, especially hepatitis C infection. Our laboratory has received 378 requests for cryoglobulin testing over the past 5 years, with a detection rate of 4.8% in the 271 patients involved. Twelve per cent of the specimens were not processed due to being at an inappropriate temperature on arrival at the laboratory. Clinicians should be aware of temperature requirements when requesting cryoglobulin testing in suspected cases, and for all relevant protein tests in patients with cryoglobulinaemia. Handling specimens at inappropriate temperatures in the pre-analytical and analytical phases of the investigation might lead to cryoprecipitation and therefore false-negative results. The potential pitfalls encountered with specimen handling, analysis, and result interpretation are discussed in detail.