Identification of new biomarkers of pyridoxine-dependent epilepsy by GC/MS-based urine metabolomics

α-Aminoadipic semialdehyde and its cyclic form (Δ¹-piperideine-6-carboxylate) accumulate in patients with α-aminoadipic semialdehyde dehydrogenase (AASADH; antiquitin; ALDH7A1) deficiency. Δ¹-Piperideine-6-carboxylate is known to react with pyridoxal 5′-phosphate (PLP) to form a Knoevenagel condensation product, resulting in pyridoxine-dependent epilepsy. Despite dramatic clinical improvement following pyridoxine supplementation, many patients still suffer some degree of intellectual disability due to delayed diagnosis. In order to expedite the diagnosis of patients with suspected AASADH deficiency and minimize the delay in treatment, we used gas chromatography-mass spectrometry-based metabolomics to search for potentially diagnostic biomarkers in urine from four patients with ALDH7A1 mutations, and identified Δ²-piperideine-6-carboxylate, 6-oxopipecolate, and pipecolate as candidate biomarkers. In a patient at postnatal day six, but before pyridoxine treatment, Δ²-piperideine-6-carboxylate and pipecolate were present at very high concentrations, indicating that these compounds may be good biomarkers for untreated AASADH deficiency patients. On the other hand, following pyridoxine/PLP treatment, 6-oxopipecolate was shown to be greatly elevated. We suggest that noninvasive urine metabolomics screening for Δ²-piperideine-6-carboxylate, 6-oxopipecolate, and pipecolate will be useful for prompt and reliable diagnosis of AASADH deficiency in patients within any age group. The most appropriate combination among Δ²-piperideine-6-carboxylate, 6-oxopipecolate, and pipecolate as biomarkers for AASADH deficiency patients appears to depend on the age of the patient and whether pyridoxine/PLP supplementation has been implemented. We anticipate that the present bioanalytical information will also be useful to researchers studying glutamate, proline, lysine and ornithine metabolism in mammals and other organisms.