Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: High-affinity oligoclonal TCRs define effective adoptive T cell therapy targeting mutant KRAS-G12D

doi: 10.1073/pnas.1921964117

Figure Lengend Snippet: KRAS-G12D–specific TCRs display high affinities that inversely correlate with in vivo persistence. (A–D) Binding of TCR9a (A), 9b (B), 9c (C), and 9d (D) to captured HLA-C*08:02–KRAS-G12D-9-mer at the indicated nanomolar concentrations determined by surface plasmon resonance. Dissociation constants were determined by kinetic curve fitting. Data are representative of two independent experiments. (E) Binding of TCR10 to captured HLA-C*08:02–KRAS-G12D-10-mer at the indicated micromolar concentrations determined by SPR. Data are representative of three independent experiments. (F) Equilibrium binding and affinity (steady state) of TCR10 to HLA-C*08:02–KRAS-G12D-10-mer and KRAS-G12D-9-mer. Data are representative of three independent experiments. (G and H) Correlation of TCR affinity (KA) with TCR frequency in the infusion product used to treat patient 4095 (G) and in the periphery of patient 4095, 9 mo after T cell transfer (H). TCR frequencies are from ref. 17.

Article Snippet: To investigate the impact of TCR–pHLA binding affinity on the success of KRAS-G12D–specific immunotherapy, we measured the solution binding affinities of KRAS-G12D–specific TCRs to their cognate HLA-C by surface plasmon resonance (SPR) using BIAcore ( ).

Techniques: In Vivo, Binding Assay, SPR Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: High-affinity oligoclonal TCRs define effective adoptive T cell therapy targeting mutant KRAS-G12D

doi: 10.1073/pnas.1921964117

Figure Lengend Snippet: T cell recognition of the KRAS-G12D nonamer via CDR2β. (A) TCR9d CDR2β interactions with p7 Lys of the KRAS-G12D-9-mer. CDR2β, red; HLA-C, gray; KRAS-G12D-9-mer, green. (B) TCR9d CDR3α interactions with HLA-C*08:02 and the KRAS-G12D-9-mer. CDR3α, turquoise; CDR3β, orange; HLA-C, gray; KRAS-G12D-9-mer, green.(C) Frequency of TCR+ Jurkat T cells expressing CD69 after incubation with 221-C*08:02-ICP47 cells loaded with KRAS-G12D-9-mer peptides with the indicated amino acid substitutions. Amino acids identical to the KRAS sequence are indicated with “–.” Peptides were tested from 1,000 to 1 nM, shown here at 10 nM; data are a mean of three independent experiments. Statistical significance was assessed by one-way ANOVA with Dunnett’s multiple comparison test (****P < 0.0001). (D) TCR9a/d CDR3β interactions with HLA-C*08:02 Arg-69. TCR9a-CDR3β, blue; TCR9d-CDR3β, orange; HLA-C, gray; KRAS-G12D-9-mer, green. (E and F) Binding of TCR9a-CDR3α Q98A (E) and TCR9a-CDR2β YE48,49AA (F) to captured HLA-C*08:02–KRAS-G12D-9-mer at the indicated nanomolar concentrations determined by SPR. Dissociation constants were determined by kinetic curve fitting. Data are representative of two independent experiments.

Article Snippet: To investigate the impact of TCR–pHLA binding affinity on the success of KRAS-G12D–specific immunotherapy, we measured the solution binding affinities of KRAS-G12D–specific TCRs to their cognate HLA-C by surface plasmon resonance (SPR) using BIAcore ( ).

Techniques: Expressing, Incubation, Sequencing, Comparison, Binding Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: High-affinity oligoclonal TCRs define effective adoptive T cell therapy targeting mutant KRAS-G12D

doi: 10.1073/pnas.1921964117

Figure Lengend Snippet: T cell recognition of the KRAS-G12D decamer via an altered peptide conformation. (A) Cartoon (Top) and stick (Bottom) models of the KRAS-G12D-10-mer in two conformations. TCR-free conformation, blue; the conformation in complex with TCR10, black. (B) Interactions of TCR10 with the KRAS-G12D-10-mer in TCR-bound (Left) conformation. H bonds are between Tyr-97 of CDR3α and the carbonyl of Gly (p4) and between the amide of CDR3β Gly-97 and the carbonyl of Val (p5). The salt bridge was between CDR3β Asp-95 and Lys (p7). CDR3α, purple; CDR3β, red; HLA-C, gray; KRAS-G12D-10-mer, black. (Right) Modeling of the TCR10 interaction with the KRAS-G12D-10-mer in the TCR-free conformation. KRAS-G12D-10-mer, blue. (C) Frequency of TCR+ Jurkat T cells expressing CD69 after incubation with 221-C*08:02-ICP47 cells loaded with KRAS-G12D-10-mer peptides with the indicated amino acid substitutions. Amino acids identical to the KRAS sequence are indicated with “–.” Peptides were tested from 1,000 to 1 nM, shown here at 10 nM; data are a mean of three independent experiments. Statistical significance was assessed by one-way ANOVA with Dunnett’s multiple comparison test (****P < 0.0001). (D and E) Binding of WT and mutant TCR10 with indicated amino acid substitutions to captured HLA-C*08:02–KRAS-G12D-10-mer at 5 μM, determined by SPR. Representative of two independent experiments (D) and summary (E).

Article Snippet: To investigate the impact of TCR–pHLA binding affinity on the success of KRAS-G12D–specific immunotherapy, we measured the solution binding affinities of KRAS-G12D–specific TCRs to their cognate HLA-C by surface plasmon resonance (SPR) using BIAcore ( ).

Techniques: Expressing, Incubation, Sequencing, Comparison, Binding Assay, Mutagenesis

Journal: bioRxiv

Article Title: Neutralizing Antibodies Isolated by a site-directed Screening have Potent Protection on SARS-CoV-2 Infection

doi: 10.1101/2020.05.03.074914

Figure Lengend Snippet: SPR to measure the binding affinity of SARS-CoV-2-RBD-his protein to captured antibodies.

Article Snippet: Surface plasmon resonance (SPR) was performed by GenScript (GenScript, Nanjing) to measure the affinity of the antibody.

Techniques: Binding Assay