RESUMEN
CD38 is the major NAD+-hydrolyzing ecto-enzyme in most mammals. As a type II transmembrane protein, CD38 is also a promising target for the immunotherapy of multiple myeloma (MM). Nanobodies are single immunoglobulin variable domains from heavy chain antibodies that naturally occur in camelids. Using phage display technology, we isolated 13 mouse CD38-specific nanobodies from immunized llamas and produced these as recombinant chimeric mouse IgG2a heavy chain antibodies (hcAbs). Sequence analysis assigned these hcAbs to five distinct families that bind to three non-overlapping epitopes of CD38. Members of families 4 and 5 inhibit the GDPR-cyclase activity of CD38. Members of families 2, 4 and 5 effectively induce complement-dependent cytotoxicity against CD38-expressing tumor cell lines, while all families effectively induce antibody dependent cellular cytotoxicity. Our hcAbs present unique tools to assess cytotoxicity mechanisms of CD38-specific hcAbs in vivo against tumor cells and potential off-target effects on normal cells expressing CD38 in syngeneic mouse tumor models, i.e. in a fully immunocompetent background.
Asunto(s)
ADP-Ribosil Ciclasa 1/inmunología , Anticuerpos Monoclonales de Origen Murino/inmunología , Anticuerpos Antineoplásicos/inmunología , Citotoxicidad Celular Dependiente de Anticuerpos , Cadenas Pesadas de Inmunoglobulina/inmunología , Glicoproteínas de Membrana/inmunología , Neoplasias/inmunología , ADP-Ribosil Ciclasa 1/genética , Animales , Anticuerpos Monoclonales de Origen Murino/genética , Anticuerpos Antineoplásicos/genética , Línea Celular Tumoral , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos BALB C , Ratones NoqueadosRESUMEN
Nanobodies (VHHs) are the single variable immunoglobulin domains of heavy chain antibodies (hcAbs) that naturally occur in alpacas and other camelids. The two variable domains of conventional antibodies typically interact via a hydrophobic interface. In contrast, the corresponding surface area of nanobodies is hydrophilic, rendering these single immunoglobulin domains highly soluble, robust to harsh environments, and exceptionally easy to format into bispecific reagents. In homage to Geoffrey Burnstock, the pioneer of purinergic signaling, we provide a brief history of nanobody-mediated modulation of purinergic signaling, using our nanobodies targeting P2X7 and the NAD+-metabolizing ecto-enzymes CD38 and ARTC2.2 as examples.