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Cytotherapy ; 7(1): 46-56, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16040383

RESUMO

BACKGROUND: GM-CSF-secreting, allogeneic cell-based cancer vaccines have shown promise for the treatment of a variety of solid tumors. We have now applied this approach to breast cancer. The aim of these studies was to optimize expansion parameters, qualify the manufacturing process, and establish expected outcomes for cGMP-compliant manufacturing of two GM-CSF-secreting breast tumor cell lines. METHODS: The variables affecting the efficiency of expanding and formulating two allogeneic GM-CSF-secreting cell lines, 2T47D-V and 3SKBR3-7, were systematically evaluated. Production criteria investigated included alternative cell culture vessels (flasks vs. cell factories), centrifugation time and speed variables for large volume cell concentration, cell seeding density, the minimal concentration of FBS required for maximal cell expansion, and the dose and timing of irradiation in relation to cryopreservation. RESULTS: These studies demonstrate that, in comparison with standard 150-cm2 tissue culture flasks, Nunc 10-Stack Cell Factories are a more efficient and practical cell culture vessel for vaccine cell line manufacture. Centrifugation optimization studies using the COBE 2991 Cell Processor established that a speed of 2000 r.p.m. (450 g) for 2 min reliably concentrated the cells while maintaining acceptable viability and bioactivity. Radiation studies established that lethal irradiation prior to cryopreservation does not compromise the quality of the product, as measured by post-thaw cell viability and GM-CSF cell line-specific secretion levels. Finally, studies aimed at optimizing the production of one vaccine cell line, 3SKBR3-7, demonstrated that seeding the cells at a higher density and maintaining them in half the initial concentration of FBS maximized the yield of bioactive cells, resulting in significant cost savings. DISCUSSION: A manufacturing process that simultaneously maximizes cell yield, minimizes cell manipulation and maintains vaccine cell potency is critical for producing cell-based cancer vaccines in an academic setting. These studies define a feasible, reproducible and cost-effective methodology for production of a GM-CSF-secreting breast cancer vaccine that is cGMP compliant.


Assuntos
Neoplasias da Mama/terapia , Vacinas Anticâncer/síntese química , Carcinoma/terapia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/síntese química , Centros Médicos Acadêmicos/economia , Centros Médicos Acadêmicos/métodos , Centros Médicos Acadêmicos/normas , Neoplasias da Mama/imunologia , Vacinas Anticâncer/economia , Vacinas Anticâncer/efeitos da radiação , Carcinoma/imunologia , Técnicas de Cultura de Células/economia , Técnicas de Cultura de Células/métodos , Linhagem Celular Tumoral , Análise Custo-Benefício , Criopreservação/métodos , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/uso terapêutico , Fidelidade a Diretrizes , Humanos , Laboratórios/economia , Laboratórios/normas , Doses de Radiação , Transplante Homólogo/economia , Transplante Homólogo/imunologia , Transplante Homólogo/métodos
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