Study of early detection of patients with inherited metabolic disorders characterized clinically by severe ketoacidosis.

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Episodes of ketoacidosis are rather common in childhood, but we sometimes come across severe patients. There is a possibility that inherited metabolic disorders like organic acidemias are concealed in such patients. Many organic acidemias can be readily detected by GC/MS.Recently, ketone body catabolic disoredrs, which are closely related to organic acidemias, have also been recognized. The ketone body disorders include 3 types of diseases, 3-ketothiolase deficiency, cytosolic acetoacetyl-CoA thiolase (CT) deficiency or succinyl-CoA : 3-ketoacid CoA transferase (SCOT) deficiency. The latter two diseases may be difficult in diagnosis by GC/MS alone, because its metabolic profile is often nonspecific. Further, accurate assessment of CT activity in cultured cells is difficult at the present time. We developed a system to precisely diagnose the diseases associated with severe ketoacidosis.1) Development of the system of automated GC/MS data profiling and disease detection : This automated system enabled to profile urinary organic acids and chemically diagnose organic acidemias, which are often associated with severe ketoacidosis. We confirmed the usefulness of this system by testing urine samples from patients with previously known diseases.2) Development of simple and practical assay method for the detection of CT deficiency : We used cultured lymphoblasts and developed a practical assay method of CT,separating from mitochondrial thiolases or SCOT,both related to ketone body catabolism, using rapid digitonine separation of cytosol and organelle fractions. The efficiency of separation were confirmed by immunoblotting and immunotitration experiments.