We present prenatal diagnosis of Neu-Laxova syndrome with compound heterozygous variants in PHGDH in a fetus presenting increased nuchal translucency (NT) and severe early-onset fetal growth restriction (FGR) of a dichorionic diamniotic (DCDA) twin pregnancy. A 36-year-old, gravida 3, para 1, woman with an adverse pregnancy history of fetus demise, now bearing a DCDA twin pregnancy conceived by IVF-ET, underwent amniocentesis at 17+2 weeks of gestation because of one of the twin presenting increased NT at 13 + 1 weeks of gestation, and karyotype, FISH and SNP revealed no genetic abnormity. Since 22 weeks of gestation, severe FGR was presented in ultrasound in the fetus with increased NT. Fetal ultrasonic cardiogram at 26 + 1 weeks of gestation reported no abnormity. Trio-WES prescribed at 29+4 weeks of gestation revealed compound heterozygous variants in the affected fetus in PHGDH (NM_006623.4): a known missense variant c.488G > A (p.Arg163Gln) (ClinVar; [VCV000139535.8]) inherited from the father and a novel missense variant c.1129G > C (p.Gly377Arg) inherited from the mother. Brain fetal MRI performed at 36 + 6 weeks of gestation showed callosal agenesis, meanwhile, fetal ultrasound at 36 + 6 weeks of gestation re-confirmed microcephaly and micrognathia in the affected fetus. A novel likely pathological missense variant causing NLS: NM_006623.4(PHGDH):c.1129G > C (p.Gly377Arg) was reported. Trio WES shall be prescribed sequentially after a normal SNP result for fetus with early-onset severe FGR to rule out pathological genetic causes and provide guidance for future reproduction.

The purpose of this study was to improve our understanding of severe serine biosynthesis defects through a comprehensive description of prenatal, and postnatal manifestations and the mutational spectrum in a new cohort of 12 unrelated Egyptian Families. Detailed fetal ultrasound examination, postnatal assessment, and whole exome sequencing (WES) were performed in a cohort of 12 fetuses with suspected Neu-Laxova syndrome (NLS), the most severe expression of serine biosynthesis defects. Additionally, a comprehensive review of the literature was conducted by merging the data from all the molecularly-confirmed cases with ours to gain a better understanding of the clinical variability of NLS. Novel clinical manifestations including intrauterine convulsions, hemivertebrae, natal teeth, holoprosencephaly, and rhombencephalosynapsis were observed. Molecular analysis identified 7 and 2 likely disease-causing variants in the PSAT1 and PHGDH genes, respectively. Four of them were novel, including the c.734G>A missense variant in PSAT1, which has been proposed to be a founder variant among Egyptians. The present cohort expands the spectrum of serine biosynthesis disorders. Moreover, it illuminates the role of prenatal exome sequencing in lethal conditions constituting the most severe end of already-known human diseases.

In the brain, the non-essential amino acid L-serine is produced through the phosphorylated pathway (PP) starting from the glycolytic intermediate 3-phosphoglycerate: among the different roles played by this amino acid, it can be converted into D-serine and glycine, the two main co-agonists of NMDA receptors. In humans, the enzymes of the PP, namely phosphoglycerate dehydrogenase (hPHGDH, which catalyzes the first and rate-limiting step of this pathway), 3-phosphoserine aminotransferase, and 3-phosphoserine phosphatase are likely organized in the cytosol as a metabolic assembly (a "serinosome"). The hPHGDH deficiency is a pathological condition biochemically characterized by reduced levels of L-serine in plasma and cerebrospinal fluid and clinically identified by severe neurological impairment. Here, three single-point variants responsible for hPHGDH deficiency and Neu-Laxova syndrome have been studied. Their biochemical characterization shows that V261M, V425M, and V490M substitutions alter either the kinetic (both maximal activity and Km for 3-phosphoglycerate in the physiological direction) and the structural properties (secondary, tertiary, and quaternary structure, favoring aggregation) of hPHGDH. All the three variants have been successfully ectopically expressed in U251 cells, thus the pathological effect is not due to hindered expression level. At the cellular level, mistargeting and aggregation phenomena have been observed in cells transiently expressing the pathological protein variants, as well as a reduced L-serine cellular level. Previous studies demonstrated that the pharmacological supplementation of L-serine in hPHGDH deficiencies could ameliorate some of the related symptoms: our results now suggest the use of additional and alternative therapeutic approaches.

Reduced activity of the enzymes encoded by PHGDH, PSAT1, and PSPH causes a set of ultrarare, autosomal recessive diseases known as serine biosynthesis defects. These diseases present in a broad phenotypic spectrum: at the severe end is Neu-Laxova syndrome, in the intermediate range are infantile serine biosynthesis defects with severe neurological manifestations and growth deficiency, and at the mild end is childhood disease with intellectual disability. However, L-serine supplementation, especially if started early, can ameliorate and in some cases even prevent symptoms. Therefore, knowledge of pathogenic variants can improve clinical outcomes. Here, we use a yeast-based assay to individually measure the functional impact of 1,914 SNV-accessible amino acid substitutions in PSAT. Results of our assay agree well with clinical interpretations and protein structure-function relationships, supporting the inclusion of our data as functional evidence as part of the ACMG variant interpretation guidelines. We use existing ClinVar variants, disease alleles reported in the literature and variants present as homozygotes in the primAD database to define assay ranges that could aid clinical variant interpretation for up to 98% of the tested variants. In addition to measuring the functional impact of individual variants in yeast haploid cells, we also assay pairwise combinations of PSAT1 alleles that recapitulate human genotypes, including compound heterozygotes, in yeast diploids. Results from our diploid assay successfully distinguish the genotypes of affected individuals from those of healthy carriers and agree well with disease severity. Finally, we present a linear model that uses individual allele measurements to predict the biallelic function of ~1.8 million allele combinations corresponding to potential human genotypes. Taken together, our work provides an example of how large-scale functional assays in model systems can be powerfully applied to the study of ultrarare diseases.