A mild skeletal phenotype with overlapping features of Miller syndrome and functional characterisation of two new variants of human dihydroorotate dehydrogenase

Dihydroorotate dehydrogenase (DHODH) catalyzes the fourth enzymatic reaction of the pyrimidine biosynthesis pathway. Miller syndrome, also known as postaxial acrofacial dysostosis, is caused by biallelic pathogenic variants in <i>DHODH</i>. We present a patient with a relatively mild skeletal phenotype carrying a novel variant of unknown significance in <i>DHODH</i>: c.829G > A, p.(D277N), in combination with a known variant, c.403C > T, p.(R135C). We functionally characterized the <i>DHODH</i> variant D277N in comparison to a very recently reported, but functionally uncharacterized variant P43L, that was found in a patient with more pronounced Miller syndrome features. Because both cases share the same <i>DHODH</i> variant R135C, we aimed to study the effect on enzyme activity of the two variants D277N and P43L to determine pathogenicity and possibly a genotype-phenotype relationship on the R135C background. We found a significant reduction in enzyme activity for both variants. The variant P43L showed a more pronounced loss of function in all assays compatible with other pathogenic variants reported in Miller, whereas the D277N variant showed milder changes that could reflect the mild phenotypic features in our patient. Yet due to a lack of a known threshold of residual enzyme activity to determine pathogenicity, this needs to be confirmed in further studies.