RESUMO
Cell line development (CLD) represents a complex but highly critical process during the development of a biological drug. To shed light on this crucial workflow, a team of BioPhorum members (authors) has developed and executed surveys focused on the activities and effort involved in a typical CLD campaign. An average of 27 members from different companies that participate in the BioPhorum CLD working group answered surveys covering three distinguishable stages of a standard CLD process: (1) Pre-transfection, including vector design and construction; (2) Transfection, spanning the initial introduction of vector into cells and subsequent selection and analysis of the pools; and (3) Single Cell Cloning and Lead Clone Selection, comprising methods of isolating single cells and confirming clonal origin, subsequent expansion and screening processes, and methods for identifying and banking lead clones. The surveys were very extensive, including a total of 341 questions split between antibody and complex molecule CLD processes. In this survey review, the authors interpret and highlight responses for antibody development and, where relevant, contrast complex molecule development challenges to provide a comprehensive industry perspective on the typical time and effort required to develop a CHO production cell line.
Assuntos
Cricetulus , Animais , Células CHO , Indústria Farmacêutica , Células Clonais , Transfecção/métodos , HumanosRESUMO
Patients with α-dystroglycanopathies, a subgroup of rare congenital muscular dystrophies, present with a spectrum of clinical manifestations that includes muscular dystrophy as well as CNS and ocular abnormalities. Although patients with α-dystroglycanopathies are genetically heterogeneous, they share a common defect of aberrant post-translational glycosylation modification of the dystroglycan alpha-subunit, which renders it defective in binding to several extracellular ligands such as laminin-211 in skeletal muscles, agrin in neuromuscular junctions, neurexin in the CNS, and pikachurin in the eye, leading to various symptoms. The genetic heterogeneity associated with the development of α-dystroglycanopathies poses significant challenges to developing a generalized treatment to address the spectrum of genetic defects. Here, we propose the development of a bispecific antibody (biAb) that functions as a surrogate molecular linker to reconnect laminin-211 and the dystroglycan beta-subunit to ameliorate sarcolemmal fragility, a primary pathology in patients with α-dystroglycan-related muscular dystrophies. We show that the treatment of LARGEmyd-3J mice, an α-dystroglycanopathy model, with the biAb improved muscle function and protected muscles from exercise-induced damage. These results demonstrate the viability of a biAb that binds to laminin-211 and dystroglycan simultaneously as a potential treatment for patients with α-dystroglycanopathy.
Assuntos
Anticorpos Biespecíficos/farmacologia , Distroglicanas/metabolismo , Laminina/metabolismo , Síndrome de Walker-Warburg/metabolismo , Animais , Anticorpos Biespecíficos/imunologia , Anticorpos Biespecíficos/metabolismo , Modelos Animais de Doenças , Distroglicanas/imunologia , Expressão Gênica , Humanos , Imuno-Histoquímica , Injeções Intramusculares , Laminina/genética , Laminina/imunologia , Camundongos , Camundongos Knockout , Modelos Biológicos , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Ligação Proteica/efeitos dos fármacos , Domínios e Motivos de Interação entre Proteínas/genética , Sarcolema/efeitos dos fármacos , Sarcolema/metabolismo , Síndrome de Walker-Warburg/tratamento farmacológico , Síndrome de Walker-Warburg/etiologiaRESUMO
Alemtuzumab, a monoclonal antibody directed against human CD52, is used in the treatment of MS. To characterize the impact of anti-CD52 administration, a monoclonal antibody to mouse CD52 (anti-muCD52) was generated and evaluated in EAE mouse models of MS. A single course of anti-muCD52 provided a therapeutic benefit accompanied by a reduction in the frequency of autoreactive T lymphocytes and production of pro-inflammatory cytokines. Examination of the CNS revealed a decrease in infiltrating lymphocytes, demyelination and axonal loss. Electrophysiological assessment showed preservation of axonal conductance in the spinal cord. These findings suggest that anti-CD52 therapy may help preserve CNS integrity.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Antígenos CD/imunologia , Antígenos de Neoplasias/imunologia , Doenças Desmielinizantes/tratamento farmacológico , Doenças Desmielinizantes/imunologia , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/imunologia , Glicoproteínas/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/farmacologia , Axônios/efeitos dos fármacos , Axônios/imunologia , Axônios/patologia , Antígeno CD52 , Doenças Desmielinizantes/patologia , Encefalomielite Autoimune Experimental/patologia , Glicoproteínas/antagonistas & inibidores , Humanos , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dados de Sequência MolecularRESUMO
RATIONALE: The susceptibility of neonates to pulmonary and systemic infection has been associated with the immaturity of both lung structure and the immune system. Surfactant protein (SP) D is a member of the collectin family of innate immune molecules that plays an important role in innate host defense of the lung. OBJECTIVES: We tested whether treatment with recombinant human SP-D influenced the response of the lung and systemic circulation to intratracheally administered Escherichia coli lipopolysaccharides. METHODS: After intratracheal lipopolysaccharide instillation, preterm newborn lambs were treated with surfactant and ventilated for 5 h. MEASUREMENT: Survival rate, physiologic lung function, lung and systemic inflammation, and endotoxin level in plasma were evaluated. MAIN RESULTS: In control lambs, intratracheal lipopolysaccharides caused septic shock and death associated with increased endotoxin in plasma. In contrast, all lambs treated with recombinant human SP-D were physiologically stable and survived. Leakage of lipopolysaccharides from the lungs to the systemic circulation was prevented by intratracheal recombinant human SP-D. Recombinant human SP-D prevented systemic inflammation and decreased the expression of IL-1beta, IL-8, and IL-6 in the spleen and liver. Likewise, recombinant human SP-D decreased IL-1beta and IL-6 in the lung and IL-8 in the plasma. Recombinant human SP-D did not alter pulmonary mechanics following endotoxin exposure. Recombinant human SP-D was readily detected in the lung 5 h after intratracheal instillation. CONCLUSIONS: Intratracheal recombinant human SP-D prevented shock caused by endotoxin released from the lung during ventilation in the premature newborn.