Most IRS+ T-cell clones from patients with PNH bear IRS of the activating type. T-cell clones expressing IRS were derived from 3 different patients with PNH (PNH5, PNH6, PNH7) and from 5 healthy donors (HD3, HD8, HD9, HD15, HD19). (A) Each clone was analyzed in redirected killing assay using P815 target cells in the presence of anti-CLIR or anti-KIR mAbs at (1) an E/T ratio of 20:1 to identify T-cell clones bearing inhibiting IRS isoforms (▪) and (2) an E/T ratio of 2:1 to identify T-cell clones bearing activating IRS isoforms (▪). (B) Cytolysis by T-cell clones obtained from patients with PNH upon the engagement of activating IRS. Cytolytic activity of T-cell clones derived from patients with PNH was analyzed in a 4-hour redirected killing assay in the absence (none) or in the presence of anti-CLIR or anti-KIR mAb or anti-CD3 mAb. Anti-CD54 mAb was used as an isotype-matched control mAb. The clone A15.25 (PNH5) was CLIR+KIR2DS1+ (CD158h+) and B25.10 (PNH7) was CLIR+KIR2DS2+ (CD158j+) while the clones from PNH6 A12.3 expressed only CLIR (CD94) and the clone C25.2 was CLIR-KIR2DS1+. All the clones were selected for the expression of activating isoforms of either CLIR and/or KIR. These clones are representative of 65 clones analyzed. Results are expressed as % of 51 Cr release, mean of triplicate samples; bars indicate standard deviation of triplicate samples.
