title: Production of anti TNF-α antibodies in eukaryotic cells using different combinations of vectors carrying heavy and light chains.
abstract: Tumor necrosis factor-α (TNF-α) plays a key role in rheumatoid arthritis and some other autoimmune diseases. Therapy with anti-TNF-α recombinant antibodies (Ab) appears to be highly effective. Production of new hyper-producing eukaryotic cell lines can decrease the treatment cost, which currently is very high. However, due to the complexity of protein transcription, translation, processing, and secretion in mammalian cells, the stages at which antibody expression is affected are still poorly determined. The aim of this work was to compare the productivity of two cell lines developed in CHO DG44 cells, deficient in dihydrofolate reductase, transfected with vectors carrying either heavy (H) or light (L) chains of chimeric antibody under different combinations of selective elements. Both H and L chains were cloned either in pOptiVEC or pcDNA3.3 vectors and different combinations were used to produce HL and LH cell lines. We have shown that Ab production has been low and comparable between HL and LH cells until selection on methotrexate (MTX) when LH but not HL cells have responded with 3.5 times increased productivity. Flow cytometry analysis has demonstrated that intracellular concentration of full size Abs in LH cells was 5.6 times higher than in HL ones due to higher amount of H chain synthesis. No differences in viability between HL and LH cells have been found. We have concluded that the expression of H chain in the pOptiVEC vector, which is responsible for MTX resistance, has led to the suppression of H chain synthesis and limitation in full Ab assembly.
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, RAS, GSP-7, Miklukho-Maklaya, 16/10, 117997, Moscow, Russia, dbalabashin@gmail.com.
authors.forename: Dmitriy
authors.initials: D
authors.lastname: Balabashin
authors.forename: Elena
authors.initials: E
authors.lastname: Kovalenko
authors.forename: Viktoria
authors.initials: V
authors.lastname: Toporova
authors.forename: Teimur
authors.initials: T
authors.lastname: Aliev
authors.forename: Anna
authors.initials: A
authors.lastname: Panina
authors.forename: Elena
authors.initials: E
authors.lastname: Svirshchevskaya
authors.forename: Dmitry
authors.initials: D
authors.lastname: Dolgikh
authors.forename: Mikhail
authors.initials: M
authors.lastname: Kirpichnikov