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Transthyretin (TTR) is a plasma homotetrameric protein implicated in fatal systemic amyloidoses. TTR tetramer dissociation precedes pathological TTR aggregation. Native state stabilizers are promising drugs to treat TTR amyloidoses. Here we repurpose tolcapone, an FDA-approved molecule for Parkinson's disease, as a potent TTR aggregation inhibitor. Tolcapone binds specifically to TTR in human plasma, stabilizes the native tetramer in vivo in mice and humans and inhibits TTR cytotoxicity. Crystal structures of tolcapone bound to wild-type TTR and to the V122I cardiomyopathy-associated variant show that it docks better into the TTR T4 pocket than tafamidis, so far the only drug on the market to treat TTR amyloidoses. These data indicate that tolcapone, already in clinical trials for familial amyloid polyneuropathy, is a strong candidate for therapeutic intervention in these diseases, including those affecting the central nervous system, for which no small-molecule therapy exists.

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Background: Iodide organification defects are associated with mutations in the dual oxidase 2 (DUOX2) gene and are characterized by a positive perchlorate discharge test. These mutations produce a congenital goitrous hypothyroidism, usually transmitted in an autosomal recessive mode. Methods: We studied the complete coding sequence of the human DUOX2 gene by single-strand conformational polymorphism (SSCP) analysis of DNA from 17 unrelated patients with iodide organification defects. Samples showing an aberrant pattern were directly sequenced. All mutations were validated by SSCP analysis. Finally, the effect of a splicing mutation was studied by construction of minigenes.

Thyroid peroxidase (TPO) defects, typically transmitted as autosomal recessive traits, result in hypothyroid goiters with failure to convert iodide into organic iodine. We analyzed the TPO gene in 14 unrelated patients with clinical evidence of iodide organification defects. Seven of the affected individuals harbored mutations in the TPO gene; one was compound heterozygous, the others were simply heterozygous for TPO mutations. Five novel mutations have been identified, one of which was found to be a single nucleotide deletion, while the other four were single nucleotide substitutions. A frameshift mutation c.387delC was detected in exon 5 which leads to an early termination signal in exon 7 (p.N129fsX208). Two missense mutations were identified in exon 8. The first, a c.920A>C transversion that results in a p.N307T substitution, was found in two patients. The second, a c

We report three patients with congenital hypothyroidism with goitre caused by two compound heterozygous mutations, p.C164Y/p.L234fsX237 and p.R277X/p.A2215D, and one homozygous mutation, p.R277X, in the TG gene. To our knowledge this is the first report of the presence of a nucleotide insertion mutation in the TG gene.

Paramyxoviruses share the essential RNA polymerase complex components, namely, the polymerase (L), phosphoprotein (P), and nucleoprotein (N). Human respiratory syncytial virus (RSV) P is the smallest polypeptide among the family, sharing a coiled coil tetramerization domain, which disruption renders the virus inactive. We show that unfolding of P displays a first transition with low cooperativity but substantial loss of α-helix content and accessibility to hydrophobic sites, indicative of loose chain packing and fluctuating tertiary structure, typical of molten globules. The lack of unfolding baseline indicates a native state in conformational exchange and metastable at 20 °C. The second transition starts from a true intermediate state, with only the tetramerization domain remaining folded. The tetramerization domain undergoes a two-state dissociation/unfolding reaction (37.3 kcal mol(-1)). The M(2-1) transcription antiterminator, unique to RSV and Metapneumovirus, forms a nonglobular P:M(2-1) complex with a 1:1 stoichiometry and a K(D) of 8.1 nM determined by fluorescence anisotropy, far from the strikingly coincident dissociation range of P and M(2-1) tetramers (10(-28) M(3)). The M(2-1) binding region has been previously mapped to the N-terminal module of P, strongly suggesting the latter as the metastable molten globule domain. Folding, oligomerization, and assembly events between proteins and with RNA are coupled in the RNA polymerase complex. Quantitative assessment of the hierarchy of these interactions and their mechanisms contribute to the general understanding of RNA replication and transcription in Paramyxoviruses. In particular, the unique P-M(2-1) interface present in RSV provides a valuable antiviral target for this worldwide spread human pathogen.

BackgroundTherapies to interrupt progression of early COVID-19 remain elusive. Among them, convalescent plasma in hospitalized patients was unsuccessful, perhaps because antibody should be administered earlier. We advanced plasma infusions to the first 72 hours of symptoms to arrest COVID-19 progression.MethodsA randomized, double-blind, placebo-controlled trial of convalescent plasma with high IgG titers against SARS-CoV2 in elderly subjects within 72 hours of mild COVID-19 symptoms. The primary endpoint was severe respiratory disease defined as a respiratory rate ≥30 and/or an O2 sat<93% in room air. The study was interrupted at 76% of its projected sample size, because cases in the region decreased considerably and steady enrollment of study subjects became virtually impossible.Results160 patients underwent randomization. In the intention-to-treat analysis (ITT), 13/80(16.2%) patients receiving plasma vs. 25/80(31.2%) receiving placebo experienced severe respiratory disease [RR(95%CI)= 0.52(0.29,0.94); p=0.026)] with an RRR=48%.A modified ITT analysis, excluding six subjects who experienced the primary endpoint before infusion, showed a larger effect size [RR(95%CI) = 0.40(0.20, 0.81), p=0.007]. High- and low-titer donor analyses, based on a median IgG titer=1:3,200, evidenced a dose-dependent response with an RRR=73.3% for recipients of high-titer plasma (p=0.016) and a number needed to treat (NNT)=4.4. All secondary endpoints exhibited trends towards protection. No solicited adverse events were observed.ConclusionsEarly administration of high-titer convalescent plasma against SARS-CoV2 to mildly ill infected seniors reduced COVID-19 progression. This safe, inexpensive, outpatient intervention facilitates access to treatment from industrialized to LMIC, can decompress demands on hospitals, and may contribute to save lives.Funded by The Bill & Melinda Gates Foundation and The Fundación INFANT Pandemic Fund. Registered in the Dirección de Sangre y Medicina Transfusional del Ministerio de Salud (PAEPCC19), Plataforma PRIISA (1421), and clinicaltrials.gov (NCT04479163).All authors have completed the ICMJE uniform disclosure form at www.icmje.org/coi_disclosure.pdf and declare: no support from any organization for the submitted work; RL, GPM, DW and FPP are investigators in a phase 3 SARS CoV2 trial from Pfizer; no other relationships or activities that could appear to have influenced the submitted work.