Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 5 de 5
Filtrar
Mais filtros











Base de dados
Intervalo de ano de publicação
1.
MAbs ; 14(1): 2013594, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35000569

RESUMO

The ongoing SARS-CoV-2 coronavirus pandemic of 2020-2021 underscores the need for manufacturing platforms that can rapidly produce monoclonal antibody (mAb) therapies. As reported here, a platform based on Nicotiana benthamiana produced mAb therapeutics with high batch-to-batch reproducibility and flexibility, enabling production of 19 different mAbs of sufficient purity and safety for clinical application(s). With a single manufacturing run, impurities were effectively removed for a representative mAb product (the ZMapp component c4G7). Our results show for the first time the reproducibility of the platform for production of multiple batches of clinical-grade mAb, manufactured under current Good Manufacturing Practices, from Nicotiana benthamiana. The flexibility of the system was confirmed by the results of release testing of 19 different mAbs generated with the platform. The process from plant infection to product can be completed within 10 days. Therefore, with a constant supply of plants, response to the outbreak of an infectious disease could be initiated within a matter of weeks. Thus, these data demonstrated that this platform represents a reproducible, flexible system for rapid production of mAb therapeutics to support clinical development.


Assuntos
Anticorpos Monoclonais , Anticorpos Antivirais , COVID-19/imunologia , Nicotiana , Plantas Geneticamente Modificadas , SARS-CoV-2/imunologia , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/química , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/biossíntese , Anticorpos Antivirais/química , Anticorpos Antivirais/genética , Anticorpos Antivirais/imunologia , Humanos , Plantas Geneticamente Modificadas/química , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/imunologia , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Nicotiana/química , Nicotiana/genética , Nicotiana/crescimento & desenvolvimento , Nicotiana/imunologia , Tratamento Farmacológico da COVID-19
2.
Proc Natl Acad Sci U S A ; 118(48)2021 11 30.
Artigo em Inglês | MEDLINE | ID: mdl-34815336

RESUMO

Nonhormonal products for on-demand contraception are a global health technology gap; this unmet need motivated us to pursue the use of sperm-binding monoclonal antibodies to enable effective on-demand contraception. Here, using the cGMP-compliant Nicotiana-expression system, we produced an ultrapotent sperm-binding IgG antibody possessing 6 Fab arms per molecule that bind a well-established contraceptive antigen target, CD52g. We term this hexavalent antibody "Fab-IgG-Fab" (FIF). The Nicotiana-produced FIF had at least 10-fold greater sperm-agglutination potency and kinetics than the parent IgG, while preserving Fc-mediated trapping of individual spermatozoa in mucus. We formulated the Nicotiana-produced FIF into a polyvinyl alcohol-based water-soluble contraceptive film and evaluated its potency in reducing progressively motile sperm in the sheep vagina. Two minutes after vaginal instillation of human semen, no progressively motile sperm were recovered from the vaginas of sheep receiving FIF Film. Our work supports the potential of multivalent contraceptive antibodies to provide safe, effective, on-demand nonhormonal contraception.


Assuntos
Anticorpos Monoclonais/farmacologia , Anticoncepção/métodos , Espermatozoides/imunologia , Administração Intravaginal , Animais , Anticorpos/imunologia , Anticoncepcionais/farmacologia , Feminino , Humanos , Fragmentos Fab das Imunoglobulinas/farmacologia , Imunoglobulina G/farmacologia , Masculino , Modelos Animais , Ovinos , Motilidade dos Espermatozoides
3.
EBioMedicine ; 69: 103478, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34256345

RESUMO

BACKGROUND: Approximately 40% of human pregnancies are unintended, indicating a need for more acceptable effective contraception methods. New antibody production systems make it possible to manufacture reagent-grade human monoclonal antibodies (mAbs) for clinical use. We used the Nicotiana platform to produce a human antisperm mAb and tested its efficacy for on-demand topical contraception. METHODS: Heavy and light chain variable region DNA sequences of a human IgM antisperm antibody derived from an infertile woman were inserted with human IgG1 constant region sequences into an agrobacterium and transfected into Nicotiana benthamiana. The product, an IgG1 mAb ["Human Contraception Antibody" (HCA)], was purified on Protein A columns, and QC was performed using the LabChip GXII Touch protein characterization system and SEC-HPLC. HCA was tested for antigen specificity by immunofluorescence and western blot assays, antisperm activity by sperm agglutination and complement dependent sperm immobilization assays, and safety in a human vaginal tissue (EpiVaginal™) model. FINDINGS: HCA was obtained at concentrations ranging from 0.4 to 4 mg/ml and consisted of > 90% IgG monomers. The mAb specifically reacted with a glycan epitope on CD52g, a glycoprotein produced in the male reproductive tract and found in abundance on sperm. HCA potently agglutinated sperm under a variety of relevant physiological conditions at concentrations ≥ 6.25 µg/ml, and mediated complement-dependent sperm immobilization at concentrations ≥ 1 µg/ml. HCA and its immune complexes did not induce inflammation in EpiVaginal™ tissue. INTERPRETATION: HCA, an IgG1 mAb with potent sperm agglutination and immobilization activity and a good safety profile, is a promising candidate for female contraception. FUNDING: This research was supported by grants R01 HD095630 and P50HD096957 from the National Institutes of Health.


Assuntos
Anticorpos Monoclonais/imunologia , Antígeno CD52/imunologia , Anticoncepção Imunológica/métodos , Espermatozoides/imunologia , Vacinas Anticoncepcionais/imunologia , Especificidade de Anticorpos , Feminino , Humanos , Masculino
4.
PLoS Med ; 18(2): e1003495, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33534791

RESUMO

BACKGROUND: MB66 film is a multipurpose prevention technology (MPT) product with monoclonal antibodies (mAbs) against HIV-1 (VRC01-N) and HSV-1 and 2 (HSV8-N). The mAbs were produced by transient expression in Nicotiana benthamiana (N). We conducted a Phase I clinical trial to assess the safety, pharmacokinetics (PK), and ex vivo efficacy of single and repeated doses of MB66 when used intravaginally. METHODS AND FINDINGS: The clinical trial enrolled healthy reproductive-aged, sexually abstinent women. In Segment A, 9 women received a single MB66 film which was inserted into the vaginal posterior fornix by a clinician. In Segment B, 29 women were randomly assigned to MB66 (Active) or Placebo film groups and were instructed to insert 1 film vaginally for 7 consecutive days. Visits and clinical sampling occurred predose and at various time points after single and repeated film doses. The primary endpoint was number of adverse events (AEs) Grade 2 or higher related to product use. Secondary endpoints included film dissolution rate, Nugent score (a Gram stain scoring system to diagnose bacterial vaginosis), vaginal pH, post-use survey results, cytokine concentrations in cervicovaginal lavage (CVL) specimens (assessed by Luminex assay), mAb concentrations in vaginal fluid collected from 4 sites (assessed by ELISA), and HIV and HSV neutralization activity of CVL samples ex vivo (assessed by TZM-bl and plaque reduction assay, respectively). The product was generally safe and well tolerated, with no serious AEs recorded in either segment. The AEs in this study were primarily genitourinary in nature with the most commonly reported AE being asymptomatic microscopic hematuria. There were no differences in vaginal pH or Nugent scores or significant increases in levels of proinflammatory cytokines for up to 7 days after film insertion in either segment or between Active and Placebo groups. Acceptability and willingness to use the product were judged to be high by post-use surveys. Concentrations of VRC01-N and HSV8-N in vaginal secretions were assessed over time to generate pharmacokinetic curves. Antibody levels peaked 1 hour postdosing with Active film (median: 35 µg/mL) and remained significantly elevated at 24 hours post first and seventh film (median: 1.8 µg/mL). Correcting for sample dilution (1:20), VRC01-N concentrations ranged from 36 to 700 µg/mL at the 24-hour time point, greater than 100-fold the IC50 for VRC01 (0.32 µg/mL); HSV8-N concentrations ranged from 80 to 601 µg/mL, well above the IC50 of 0.1 µg/m. CVL samples collected 24 hours after MB66 insertion significantly neutralized both HIV-1 and HSV-2 ex vivo. Study limitations include the small size of the study cohort, and the fact that no samples were collected between 24 hours and 7 days for pharmacokinetic evaluation. CONCLUSIONS: Single and repeated intravaginal applications of MB66 film were safe, well tolerated, and acceptable. Concentrations and ex vivo bioactivity of both mAbs in vaginal secretions were significantly elevated and thus could provide protection for at least 24 hours postdose. However, further research is needed to evaluate the efficacy of MB66 film in women at risk for HIV and HSV infection. Additional antibodies could be added to this platform to provide protection against other sexually transmitted infections (STIs) and contraception. TRIAL REGISTRATION: ClinicalTrials.gov NCT02579083.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Anticorpos Anti-HIV/uso terapêutico , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Administração Intravaginal , Adolescente , Adulto , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/efeitos adversos , Anticorpos Amplamente Neutralizantes/metabolismo , Anticorpos Amplamente Neutralizantes/uso terapêutico , Feminino , Anticorpos Anti-HIV/administração & dosagem , Anticorpos Anti-HIV/imunologia , HIV-1/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Profilaxia Pré-Exposição/métodos , Vagina/virologia , Adulto Jovem
5.
Gen Comp Endocrinol ; 136(1): 12-6, 2004 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-14980791

RESUMO

alpha-Melanocyte stimulating hormone (MSH) and adrenocorticotropin (ACTH)1-24, the minimal ACTH sequence required for full activity, differ only by the 10 C-terminal amino acids of ACTH1-24. Interestingly, these ten C-terminal residues have been highly conserved throughout vertebrate evolution. To understand the functional constraints of these 10 amino acids we analyzed the effects of mutating these residues on steroidogenic activity in vivo and in vitro. Alanine substitutions of some of the first four amino acid residues (the basic core residues KKRR, 15-18) greatly reduces ACTH activity in vitro and in vivo; replacement of mutant alanines at residues 15 and 17 with glutamine residues partially restores ACTH activity. Thus, for ACTH receptor binding and activation, the amino acid residues 15-18 are important for their side chains. Surprisingly, conversion of the five C-terminal residues (20-24) to alanines increases ACTH activity in vivo over that of native ACTH. With respect to receptor binding and activity, the last five amino acid residues are important only for the peptide length they contribute; however, with respect to serum stability, their side chains are significant.


Assuntos
Hormônio Adrenocorticotrópico/genética , Evolução Biológica , Glândulas Suprarrenais/metabolismo , Hormônio Adrenocorticotrópico/biossíntese , Sequência de Aminoácidos , Animais , Bovinos , Sequência Conservada , Peptídeo da Parte Intermédia da Adeno-Hipófise Semelhante à Corticotropina , Análise Mutacional de DNA , Peixes , Cobaias , Humanos , Masculino , Hormônios Estimuladores de Melanócitos/biossíntese , Hormônios Estimuladores de Melanócitos/genética , Camundongos , Dados de Sequência Molecular , Fragmentos de Peptídeos/biossíntese , Pró-Opiomelanocortina/biossíntese , Pró-Opiomelanocortina/genética , Radioimunoensaio , Ratos , Xenopus
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA