Inborn errors of metabolism and hepatic disease

Inborn errors of metabolism (IEM) have many clinical manifestations and often prove difficult to diagnose. Hepatic disease is a common feature of many IEM, and may exist in isolation or as part of a wider clinical constellation of multisystem pathology. In developed societies there a changing trend in the patterns of diagnosis of cholestatic liver disease in infants, with up to 20-30% of such cases now being recognised as due to metabolic/genetic conditions (Stormon et al 2001). In Asia, the lack of the availability of the appropriate diagnostic tests may lead to IEM not being recognised as the cause of liver disease. We tested the hypothesis that some Asian children with liver disease, of unknown cause, may have an inborn error of metabolism which could be diagnosed from a blood spot collected onto a filter paper card. This sample is logistically easy to transport to a tertiary laboratory with the capabilities of performing the appropriate analysis. Our aims were: (1). To determine from a literature review which inborn errors of metabolism, which can present with liver disease, can be diagnosed from a blood spot on filter paper. (2). To establish the appropriate tests in a central laboratory. The tests chosen would be those to which the Asian physicians do not have local access. (3). To determine the logistics of transporting blood spots on filter papers into Australia. (4). To liaise with one paediatric gastroenterologist in Asia to arrange collection of the filter paper blood spots from appropriate patients. (5). The ultimate aim is to establish a diagnostic service, which is economical and logistically easy to supplement the diagnostic tests available to the Asian physicians in their own countries. (6). To provide a free and rapid diagnostic screening service for IEMs known to cause hepatic disease and to which our Indonesian colleagues had no access. From a single newborn screening card blood spot, we used electron spray ionisation tandem mass spectrometry (ES-MS:MS) techniques to screen for defects in the metabolism of fatty acid oxidation, amino acids, organic acids, the urea cycle, and peroxisomal biogenesis. SLC25A13 mutation testing was also performed from the sample newborn screening card blood spot. Tandem mass spectrometry techniques provide a rapid, cheap, and efficient method to screen for numerous IEM. ES-MS:MS has been widely implemented as a method for extended newborn screening in many countries around the world, including Australia. The ES-MS:MS techniques used in neoborn screening programes to screen for specific IEM will be utilised in this study. Citrullineamia type II (CTLN2) and neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) are relatively newly discovered clinical phenotypes of defective urea cycle functioning. They are secondary to mutations in the citrin gene (SLC25A13). Nine mutations in the SLC25A13 have previously been documented in the medical literature, mostly in Japanese patients. No group has screened Indonesian patients for SLC25A13 mutations. Defects in citrin metabolism have recently been recognised in other South East Asian countries, therefore we offered mutation analysis for five of the nine SLC25A13 mutations, SLC25A13:c.851_851delGTAT, SLC25A13:c.1177+1G>A, SLC25A13:c.1638ins23, SLC25A13:c.674C>A, SLC25A13:c.1311+1G>A , via polymerase chain reaction (PCR) techniques from the same newborn screening sample collected for ES-MS:MS testing. We have enrolled 13 Indonesian patients and one local Queensland patient with liver disease into the study. No specific IEM were identified during the EI-MS:MS arm of the study, however 46% of the Indonesian population enrolled in the study displayed an elevated propionylcarnitine (C3) level which can be associated with vitamin B12 deficiency. SLC25A13 PCR testing failed to identify any homozygous or heterozygous patients for the five mutations examined in this study. The absent findings of any specific IEM in the group is disappointing but not necessarily unexpected given the small number of the study cohort. IEM are pan-ethnic and are known to exist in our target populations. SLC25A13 mutations have previously been identified primarily in Asian populations. Our failure to uncover a specific IEM is more likely a reflection of the small number of patients enrolled in the study rather than a deficiency in the screening methods employed in the study, but may indicate that IEM are a less common cause of liver disease in Indonesian children than in Australia. Although this was a small study, it does, however, open the door for future collorborative research opportunities including: (1) enrolment of greater patient numbers from several Asian countries; (2) the broadening of range of disorders screening by including transferrin isoforms to detect congenital disorders of glycosylation, bile acids to detect disorders of bile acid synthesis, succinyl acetone to detect tyrosinaemia 1, alpha 1-antitrypsin activity with DNA confirmation for alpha 1- antitrypsin deficiency, galactose-1-phosphate for galactosaemia (in infants with neonatal onset) and lysosomal enzyme measurements to detect the lysosomal storage disorders and (3) testing the SLC25A13 mutation carrier frequency rates in heathly Indonesian population as well as mutation detection projects in patients suspected of having CTLN2 and NICCD. In the search for a patient diagnosis, negative results are still highly valuable, especially when the testing procedures provided by this study are not available locally for our Indonesian colleagues. The study has demonstrated that it is feasible and logistically easy to offer a service to our Asian neighbours to screen for the possibility of IEM in children with liver disease of unknown aetiology