Engineering a Rugged Nanoscaffold To Enhance Plug-and-Display Vaccination

Nanoscale organization is crucial to stimulating an immune response. Using self-assembling proteins as multimerization platforms provides a safe and immunogenic system to vaccinate against otherwise weakly immunogenic antigens. Such multimerization platforms are generally based on icosahedral viruses and have led to vaccines given to millions of people. It is unclear whether synthetic protein nanoassemblies would show similar potency. Here we take the computationally designed porous dodecahedral i301 60-mer and rationally engineer this particle, giving a mutated i301 (mi3) with improved particle uniformity and stability. To simplify the conjugation of this nanoparticle, we employ a SpyCatcher fusion of mi3, such that an antigen of interest linked to the SpyTag peptide can spontaneously couple through isopeptide bond formation (Plug-and-Display). SpyCatcher-mi3 expressed solubly to high yields in <i>Escherichia coli</i>, giving more than 10-fold greater yield than a comparable phage-derived icosahedral nanoparticle, SpyCatcher-AP205. SpyCatcher-mi3 nanoparticles showed high stability to temperature, freeze–thaw, lyophilization, and storage over time. We demonstrate approximately 95% efficiency coupling to different transmission-blocking and blood-stage malaria antigens. <i>Plasmodium falciparum</i> CyRPA was conjugated to SpyCatcher-mi3 nanoparticles and elicited a high avidity antibody response, comparable to phage-derived virus-like particles despite their higher valency and RNA cargo. The simple production, precise derivatization, and exceptional ruggedness of this nanoscaffold should facilitate broad application for nanobiotechnology and vaccine development

Phenotype, biochemical features, genotype and treatment outcome of pyridoxine-dependent epilepsy

We report treatment outcome of eleven patients with pyridoxine-dependent epilepsy caused by pathogenic variants in ALDH7A1 (PDE-ALDH7A1). We developed a clinical severity score to compare phenotype with biochemical features, genotype and delays in the initiation of pyridoxine. Clinical severity score included 1) global developmental delay/ intellectual disability; 2) age of seizure onset prior to pyridoxine; 3) current seizures on treatment. Phenotype scored 1-3 = mild; 4-6 = moderate; and 7-9 = severe. Five patients had mild, four patients had moderate, and two patients had severe phenotype. Phenotype ranged from mild to severe in eight patients (no lysine-restricted diet in the infantile period) with more than 10-fold elevated urine or plasma α-AASA levels. Phenotype ranged from mild to moderate in patients with homozygous truncating variants and from moderate to severe in patients with homozygous missense variants. There was no correlation between severity of the phenotype and the degree of α-AASA elevation in urine or genotype. All patients were on pyridoxine, nine patients were on arginine and five patients were on the lysine-restricted diet. 73% of the patients became seizure free on pyridoxine. 25% of the patients had a mild phenotype on pyridoxine monotherapy. Whereas, 100% of the patients, on the lysine-restricted diet initiated within their first 7 months of life, had a mild phenotype. Early initiation of lysine-restricted diet and/or arginine therapy likely improved neurodevelopmental outcome in young patients with PDE-ALDH7A1

Treatment outcome of creatine transporter deficiency: international retrospective cohort study.

To evaluate the outcome of current treatment for creatine transporter (CRTR) deficiency, we developed a clinical severity score and initiated an international treatment registry. An online questionnaire was completed by physicians following patients with CRTR deficiency on a treatment, including creatine and/or arginine, and/or glycine. Clinical severity score included 1) global developmental delay/intellectual disability; 2) seizures; 3) behavioural disorder. Phenotype scored 1-3 = mild; 4-6 = moderate; and 7-9 = severe. We applied the clinical severity score pre- and on-treatment. Seventeen patients, 14 males and 3 females, from 16 families were included. Four patients had severe, 6 patients had moderate, and 7 patients had a mild phenotype. The phenotype ranged from mild to severe in patients diagnosed at or before 2 years of age or older than 6 years of age. The phenotype ranged from mild to severe in patients with mildly elevated urine creatine to creatinine ratio. Fourteen patients were on the combined creatine, arginine and glycine therapy. On the combined treatment with creatine, arginine and glycine, none of the males showed either deterioration or improvements in their clinical severity score, whereas two females showed improvements in the clinical severity score. Creatine monotherapy resulted in deterioration of the clinical severity score in one male. There seems to be no correlation between phenotype and degree of elevation in urine creatine to creatinine ratio, genotype, or age at diagnosis. Combined creatine, arginine and glycine therapy might have stopped disease progression in males and improved phenotype in females