Measles virus (MeV) is the most contagious respiratory infectious disease known in humans and the leading cause of vaccine-­‐preventable deaths in children worldwide. An attenuated vaccine is in use, but no therapeutic or prophylactic antibodies or MeV antivirals are currently available. Protection against MeV is associated with the development of neutralizing antibodies (nMoAb) that preferentially recognize the measles virus hemagglutinin protein (MVH). Although a single serotype was described, 24 MeV genotypes have been identified with different geographical and temporal distribution showing characteristic nucleotide and amino acid differences in the several genes. Most of the variability, as determined in 48 MeV primary isolates along a six-­‐month-­‐epidemic outbreak in Madrid caused by one foreign person infected with MeV genotype B3.1, appeared in the MVH gene (up to 0,38%) where we have estimated the rate of nucleotide substitution to be 7,27x10-­‐6 per site per day (95% confidence interval, 3,09x10-­‐6 – 1,23x10-­‐5). The effect of the inter-­‐ and intra-­‐genotype variations in the MVH from MeV genotypes that have circulated in the Madrid region for the last 50 years to date (genotypes A, B3.1, C2, D4.1, D4.4, D6, D7, D8, D9, F, G3, H1), were functionally and structurally characterized by several murine nMoAbs targeting distinct epitopes and nMoAb neutralization-­‐escape mutant viruses. Our data revealed neutralizing genotype-­‐specific epitopes on the MVH and significant differences in the kinetics rates and affinity of the nMoAb to the MVH by Surface Plasmon Resonance that correlate with the neutralization sensitivities of the corresponding virus strain. By means of a labelled MoAb binding competition radioimmunoassay (RIA) to MVH in its native oligomeric structure complexed with MVF by post-­‐measles sera, developed in our laboratory, we have found that the genotype-­‐specific epitopes are immunogenic in natural infection and may elicit long-­‐term B memory. We have identified two potent broadly neutralizing epitopes conserved in every MeV genotype. One is located in one MVH receptor-­‐binding site and the other is at the MVH monomers interface. We have assayed comparative binding of each of MeV receptors CD46, CD150 (SLAM) and nectin-­‐4 to cloned MVH protein from different MeV genotypes from acute measles and brain from deceased Subacute Sclerosing Panencephalitis patients, and found some differences among genotypes cohere with amino-­‐acid sequences in receptor-­‐binding sites. This information could be applied to design better measles vaccines, potent broad-­‐ spectrum prophylactic and therapeutic anti-­‐MeV MoAbs, and to dampen pre-­‐existent MeV-­‐ specific antibodies for MeV-­‐based gene therapy and oncolytic cancer therapies.

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología. Fecha de lectura: 17-07-2014