Autologous Dendritic Cell Vaccine Production

AGS-003 was manufactured at a centralized GMP compliant facility (Argos Therapeutics, Durham, NC). Following screening and consent, autologous tumor total RNA was isolated from nephrectomy or metastasectomy tissue samples and messenger RNA was amplified using RT/PCR and in vitro transcription technologies as previously described [47 (link)]. CD40L RNA was manufactured using in vitro transcription and a post-transcriptional capping method [48 (link)]. Patients had leukapheresis at the clinical site’s donor center using a COBE Spectra® Leukapheresis System (Gambro BCT, Lakewood, CO). Monocytes were cultured in AIM-V media with 800 U/mL granulocyte macrophage-colony stimulating factor (Berlex) and 1000 U/mL IL-4 (R&D Systems) to generate immature DCs that were then matured using 20 ng/mL tumor necrosis factor alpha (TNF α) (R&D Systems)/1000 U/mL IFN-γ (InterMune)/1 μg/mL prostaglandin E2 (Sigma). Mature DCs were electroporated with the amplified tumor RNA and CD40L RNA using a post-maturation electroporation protocol [10 (link)].
The final AGS-003 product was formulated as 1.4 × 10⁷ DC/0.7 mL in 80% autologous plasma, 10% dextrose (50% w/v) (Hospira), and 10% DMSO (Sigma) and cryopreserved in liquid nitrogen vapor phase. Thawed samples of final product were assessed for sterility, mycoplasma, endotoxin, and viability prior to release for clinical use.