Codon optimization and replacement of TCR constant regions with murine sequences.

<p>A) Schematic representation of the three constructs generated for the expression of TCR-5 and derivatives. LTR: long terminal repeat, sd: splice donor, sa: splice acceptor, ψ: retrovirus encapsidation signal, MC: mouse TCR constant region, 2A: linker peptide. B) Analysis of expression of TCR-5 variants by tetramer staining. OKT3-stimulated lymphocytes were transduced twice with the corresponding TCR-expressing vector and stained with anti-CD3, anti-CD8 and SSX2_41-49 tetramers one week after transduction. Representative results from three independent experiments. Values in parentheses represent the mean fluorescence intensity of tetramer staining within the CD8 T cell population. C) ⁵¹Cr-release assay for the evaluation of antigen-specific cytolysis induced by TCR-5-transduced lymphocytes after four-hour coculture with the indicated target cells. Percentage of lysis depicted for each target cell line at different effector:target ratios is the average of duplicates in a representative experiment of three independent experiments. UT: untransduced T cells used as negative control of unspecific lysis, WT: Wild-type TCR, Co Op: copon-optimized TCR, MCR: codon-optimized TCR with mouse constant region.