Journal: Nature Communications
Article Title: Effective extracellular payload release and immunomodulatory interactions govern the therapeutic effect of trastuzumab deruxtecan (T-DXd)
doi: 10.1038/s41467-025-58266-8
Figure Lengend Snippet: a , b Au565 cells were assessed for T-DXd ( a ) or T-DM1 ( b ) cytotoxicity in vitro in the presence of CTSL inhibitor (3 µM Z-Phe-Phe-FMK). c Au565 cells overexpressing CTSL or lacking CTSL were assessed for T-DXd cytotoxicity in vitro. d Extracellular CTSL secretion by MDA-MB-231 lines was quantified with ELISA analysis of conditioned media. e Secreted proteins in conditioned media of indicated cell lines were concentrated and quantified for CTSL enzymatic activity in assay buffer. a – e Two-way ANOVA with Tukey’s multiple comparisons test ( n = 3 per group). Nonlinear regression curve fit to calculate IC50 values. f Parental and CTSL-overexpressed MDA-MB-231 cells were implanted in SCID mice, and treated with T-DXd (5 mg/kg) or control PBS. g Control-KO and CTSL-KO MDA-MB-231 cells were implanted in SCID mice, and treated with T-DXd (10 mg/kg) or control PBS. f – h Mixed-effects analysis (REML) with Tukey’s multiple comparisons ( n = 10). Arrows indicate time of treatments administered. h Tumor sizes (mm 3 ) measured at end point for studies using MDA-MB-231 CTSL-KO lines. i Immunohistochemistry verifications of CTSL expression in parental or CTSL-modified MDA-MB-231 tumors. j LC-MS quantification of DXd in MDA-MB-231 parental ( n = 15) versus CTSL-overexpressed tumors ( n = 10), treated with T-DXd (10 mg/kg) for 5 days. k LC-MS quantification of DXd in MDA-MB-231 control-KO versus three different CTSL-KO pools, treated with T-DXd (10 mg/kg) for 3 days ( n = 10 for all groups, except n = 6 for CTSL-KO #2). h , k One-way ANOVA with Tukey’s multiple comparisons. j Two-sided Mann–Whitney test. All data is presented as mean ± SEM with p values indicated.
Article Snippet: Antigen unmasking was performed with antigen retrieval buffer A. Endogenous peroxidases/phosphatases were quenched with BLOXALL blocking solution (Vector), and tissues were blocked with Animal-Free Blocker R.T.U. (Vector).
Techniques: In Vitro, Enzyme-linked Immunosorbent Assay, Activity Assay, Control, Immunohistochemistry, Expressing, Modification, Liquid Chromatography with Mass Spectroscopy, MANN-WHITNEY
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