RESUMO
Safe and efficient delivery of messenger RNAs for protein replacement therapies offers great promise but remains challenging. In this report, we demonstrate systemic, in vivo, nonviral mRNA delivery through lipid nanoparticles (LNPs) to treat a Factor IX (FIX)-deficient mouse model of hemophilia B. Delivery of human FIX (hFIX) mRNA encapsulated in our LUNAR LNPs results in a rapid pulse of FIX protein (within 4-6 h) that remains stable for up to 4-6 d and is therapeutically effective, like the recombinant human factor IX protein (rhFIX) that is the current standard of care. Extensive cytokine and liver enzyme profiling showed that repeated administration of the mRNA-LUNAR complex does not cause any adverse innate or adaptive immune responses in immune-competent, hemophilic mice. The levels of hFIX protein that were produced also remained consistent during repeated administrations. These results suggest that delivery of long mRNAs is a viable therapeutic alternative for many clotting disorders and for other hepatic diseases where recombinant proteins may be unaffordable or unsuitable.
Assuntos
Portadores de Fármacos/administração & dosagem , Fator IX/farmacocinética , Hemofilia B/terapia , Nanopartículas/administração & dosagem , RNA Mensageiro/farmacocinética , Animais , Colesterol/química , Citocinas/metabolismo , Modelos Animais de Doenças , Composição de Medicamentos/métodos , Avaliação Pré-Clínica de Medicamentos , Fator IX/genética , Fator IX/metabolismo , Feminino , Terapia Genética/métodos , Hemofilia B/genética , Hemofilia B/metabolismo , Hemofilia B/patologia , Humanos , Concentração de Íons de Hidrogênio , Injeções Intravenosas , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Knockout , Fosfatidilcolinas/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacocinéticaRESUMO
We explored an emerging technology to produce anti-Hantaan virus (HTNV) and anti-Puumala virus (PUUV) neutralizing antibodies for use as pre- or post-exposure prophylactics. The technology involves hyperimmunization of transchomosomic bovines (TcB) engineered to express human polyclonal IgG antibodies with HTNV and PUUV DNA vaccines encoding GnGc glycoproteins. For the anti-HTNV product, TcB was hyperimmunized with HTNV DNA plus adjuvant or HTNV DNA formulated using lipid nanoparticles (LNP). The LNP-formulated vaccine yielded fivefold higher neutralizing antibody titers using 10-fold less DNA. Human IgG purified from the LNP-formulated animal (SAB-159), had anti-HTNV neutralizing antibody titers >100,000. SAB-159 was capable of neutralizing pseudovirions with monoclonal antibody escape mutations in Gn and Gc demonstrating neutralization escape resistance. SAB-159 protected hamsters from HTNV infection when administered pre- or post-exposure, and limited HTNV infection in a marmoset model. An LNP-formulated PUUV DNA vaccine generated purified anti-PUUV IgG, SAB-159P, with a neutralizing antibody titer >600,000. As little as 0.33 mg/kg of SAB-159P protected hamsters against PUUV infection for pre-exposure and 10 mg/kg SAB-159P protected PUUV-infected hamsters post-exposure. These data demonstrate that DNA vaccines combined with the TcB-based manufacturing platform can be used to rapidly produce potent, human, polyclonal, escape-resistant anti-HTNV, and anti-PUUV neutralizing antibodies that are protective in animal models.