Extended Data Fig. 2: Development and in vitro testing of RIPR-PD1.
From: Immune receptor inhibition through enforced phosphatase recruitment
a, Coomassie-stained SDS–PAGE of SEC-purified RIPR-PD1(nivo). b, c, Curve fitting of recorded resonance units at steady state for RIPR-PD1 binding to immobilized CD45 (b) and PD-1 (c). Data shown for RIPR-PD1 concentrations ranging from 4 μM to 35 nM (b), and from 8 μM to 17.5 nM (c). Data are representative of 2 independent experiments. d, e, Fraction of CD25+CD69+ (d) and IL-2 (e) expression for wild-type Jurkat T cells stimulated with 2 μg ml−1 of plate-bound OKT3 and treated with nivolumab or RIPR-PD1 at the concentrations indicated for 24 h. f–i, Quantification of CD25 (f) or CD69 (g) expression and fraction of CD25+CD69+ (h) for Jurkat T cells transduced with wild-type PD-1 (shown in Extended Data Fig. 1b) and IL-2 secretion (i) for SKW-3 T cells stimulated with 2 μg ml−1 of plate-bound OKT3 and treated with nivolumab or RIPR-PD1 at the indicated concentrations for 24 h. j, Quantification of IL-2 secretion by TCR transduced SKW-T cells after stimulation for 24 h with cognate peptide in the presence or absence of PD-L1 and nivolumab or RIPR-PD1 at the indicated concentrations. k, l, IFNγ (k) and IL-2 (l) secretion by indicated CAR T cells (or mock untransduced T cells) after incubation with tumour target cells in the presence of increasing concentrations of RIPR-PD1. In d–l, data are mean ± s.d. from n = 3 biological replicates representative of 3 independent experiments.