Structural insights into the mechanism of action of a biparatopic anti-HER2 antibody.

Pathways of human epidermal growth factor (EGF) receptors are activated upon ligand-dependent or -independent homo- or heterodimerization and their subsequent transphosphorylation. Overexpression of these receptors positively correlates with transphosphorylation rates and increased tumor growth rates. MEDI4276, an anti-human epidermal growth factor receptor 2 (HER2) biparatopic antibody-drug conjugate, has two paratopes within each antibody arm. One, 39S, is aiming at the HER2 site involved in receptor dimerization and the second, single chain fragment (scFv), mimicking trastuzumab. Here we present the cocrystal structure of the 39S Fab-HER2 complex and, along with biophysical and functional assays, determine the corresponding epitope of MEDI4276 and its underlying mechanism of action. Our results reveal that MEDI4276's uniqueness is based first on the ability of its 39S paratope to block HER2 homo- or heterodimerization and second on its ability to cluster the receptors on the surface of receptor-overexpressing cells.

Immunization of Syrian golden hamsters with F subunit vaccine of human metapneumovirus induces protection against challenge with homologous or heterologous strains.

Human metapneumovirus (hMPV), a newly discovered paramyxovirus, is associated with acute respiratory-tract illness, primarily in young children, individuals with underlying disease and the elderly. Two genetic lineages of hMPV circulate around the world, and viruses from these two lineages demonstrate antigenic differences. The clinical impact of hMPV warrants the development of vaccines. Recombinant soluble fusion (F) proteins of prototype viruses of the two main lineages of hMPV that can be produced in high yields have been constructed. In this study, the antigenicity, immunogenicity and protective efficacy of these soluble F subunit vaccines were evaluated in Syrian golden hamsters (Mesocricetus auratus). Immunization of hamsters with the soluble F proteins, adjuvanted with Specol or iscom matrix, induced high virus-neutralization titres, with higher titres against the homologous than the heterologous virus. The neutralizing antibodies protected from subsequent infection of the lungs with both homologous and heterologous virus. Upon challenge, viral titres in the nasal turbinates of immunized animals were reduced significantly compared with those of PBS-immunized animals. In conclusion, a soluble F subunit vaccine for hMPV that induces cross-protective immunity for infection of the lower respiratory tract in Syrian golden hamsters has been generated.

Toll-like receptor 9-dependent activation by DNA-containing immune complexes is mediated by HMGB1 and RAGE.

Increased concentrations of DNA-containing immune complexes in the serum are associated with systemic autoimmune diseases such as lupus. Stimulation of Toll-like receptor 9 (TLR9) by DNA is important in the activation of plasmacytoid dendritic cells and B cells. Here we show that HMGB1, a nuclear DNA-binding protein released from necrotic cells, was an essential component of DNA-containing immune complexes that stimulated cytokine production through a TLR9-MyD88 pathway involving the multivalent receptor RAGE. Moreover, binding of HMGB1 to class A CpG oligodeoxynucleotides considerably augmented cytokine production by means of TLR9 and RAGE. Our data demonstrate a mechanism by which HMGB1 and RAGE activate plasmacytoid dendritic cells and B cells in response to DNA and contribute to autoimmune pathogenesis.

Isolation and characterization of monoclonal antibodies which neutralize human metapneumovirus in vitro and in vivo.

Human metapneumovirus (hMPV) is a recently described member of the Paramyxoviridae family/Pneumovirinae subfamily and shares many common features with respiratory syncytial virus (RSV), another member of the same subfamily. hMPV causes respiratory tract illnesses that, similar to human RSV, occur predominantly during the winter months and have symptoms that range from mild to severe cough, bronchiolitis, and pneumonia. Like RSV, the hMPV virus can be subdivided into two genetic subgroups, A and B. With RSV, a single monoclonal antibody directed at the fusion (F) protein can prevent severe lower respiratory tract RSV infection. Because of the high level of sequence conservation of the F protein across all the hMPV subgroups, this protein is likely to be the preferred antigenic target for the generation of cross-subgroup neutralizing antibodies. Here we describe the generation of a panel of neutralizing monoclonal antibodies that bind to the hMPV F protein. A subset of these antibodies has the ability to neutralize prototypic strains of both the A and B hMPV subgroups in vitro. Two of these antibodies exhibited high-affinity binding to the F protein and were shown to protect hamsters against infection with hMPV. The data suggest that a monoclonal antibody could be used prophylactically to prevent lower respiratory tract disease caused by hMPV.