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Construction of stable producer cells to make high‐titer lentiviral vectors for dendritic cell‐based vaccination
Author(s) -
Lee ChiLin,
Chou Michael,
Dai Bingbing,
Xiao Liang,
Wang Pin
Publication year - 2012
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.24413
Subject(s) - viral vector , dendritic cell , transfection , biology , virology , cell culture , antigen , transduction (biophysics) , sindbis virus , cross presentation , titer , virus , microbiology and biotechnology , cd8 , immunology , recombinant dna , gene , biochemistry , rna , genetics , mhc class i
Lentiviral vectors (LVs) enveloped with an engineered Sindbis virus glycoprotein can specifically bind to dendritic cells (DCs) through the surface receptor DC‐SIGN and induce antigen expression, thus providing an efficient method for delivering DC‐directed vaccines. In this study, we constructed a stable producer line (LV‐MGFP) for synthesizing DC‐SIGN‐targeted HIV‐1‐based LVs (DC‐LVs) encoding green fluorescent protein (GFP) by a concatemeric array transfection technique. We demonstrated that the established stable clones could routinely produce vector supernatants with titers above 10 7 transduction units per milliliter (TU/mL) during a continuous 3‐month cell passage. The producer cells were also capable of generating similar titers of DC‐LVs in serum‐free medium. Moreover, the addition of 1‐deoxymannojirimycin (DMJ) enabled the producer cells to manufacture DC‐LVs with both improved titers and enhanced potency to evoke antigen‐specific CD8 + T cell responses in mice. The stable lines could accommodate the replacement of the internal murine stem cell virus (MSCV) promoter with the human ubiquitin‐C (Ubi) promoter in the lentiviral backbone. The resulting DC‐LVs bearing Ubi exhibited the enhanced potency to elicit vaccine‐specific immunity. Based on accumulated evidence, our studies support the application of this production method in manufacturing DC‐LVs for preclinical and clinical testing of novel DC‐based immunization. Biotechnol. Bioeng. 2012; 109:1551–1560. © 2011 Wiley Periodicals, Inc.