Mammalian cell expression of single-chain Fv (sFv) antibody proteins and their C-terminal fusions with interleukin-2 and other effector domains.
The production of several single-chain Fv (sFv) antibody proteins was examined by three modes of mammalian cell expression. Our primary model was the 741F8 anti-c-erbB-2 sFv, assembled as either the VH-VL or VL-VH, and expressed alone, with C-terminal cysteine for dimerization, or as fusion proteins with carboxyl-terminal effector domains, including interleukin-2, the B domain of staphylococcal protein A, the S-peptide of ribonuclease S, or hexa-histidine metal chelate peptide. Constructs were expressed and secreted transiently in 293 cells and stably in CHO or Sp2/0 cell lines, the latter yielding up to 10 mg per liter. Single-chain constructs of MOPC 315 myeloma and 26-10 monoclonal antibodies were also expressed, as were hybrids comprising unrelated VH and VL regions. Our results suggest that mammalian expression is a practical and valuable complement to the bacterial expression of single-chain antibodies.
Dorai
H
H
Creative BioMolecules, Inc., Hopkinton, MA 01748.
McCartney
J E
JE
Hudziak
R M
RM
Tai
M S
MS
Laminet
A A
AA
Houston
L L
LL
Huston
J S
JS
Oppermann
H
H
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D001483:Base Sequence; D015153:Blotting, Western; D016466:CHO Cells; D002460:Cell Line; D002846:Chromatography, Affinity; D006224:Cricetinae; D017931:DNA Primers; D066246:ErbB Receptors / Q000276:immunology*; D015870:Gene Expression; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000302:isolation & purification; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000302:isolation & purification; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D010954:Plasmacytoma; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000276:immunology*; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D015183:Restriction Mapping; D014162:Transfection; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D017931:DNA Primers; D066246:ErbB Receptors; D007128:Immunoglobulin Fragments; D007376:Interleukin-2; D046911:Macromolecular Substances; D011518:Proto-Oncogene Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C055966:immunoglobulin Fv