Fig. 1: PD-(L)1 blockade is associated with increased intratumoral collagen deposition.
a In vivo volume measurements at indicated time points for 344SQ subcutaneous tumors implanted in syngeneic WT mice treated weekly with anti-PD-L1 (200 µg/mouse) or isotype control (200 µg/mouse). Treatment start time denoted by blue (1-week post-implantation) or green (3-week post-implantation) arrow; n = 5 mice per treatment group. Statistics calculated using a one-way repeated mixed-effects model (REML) with *p = 0.0371. b Heatmap of RPPA profile showing statistically significant (FDR < 0.05) differentially expressed proteins in 344SQ subcutaneous tumors treated weekly with anti-PD-L1 or isotype control, starting 1-week post-implantation in mice. Tumor samples were collected for RPPA at the endpoint of experiment in (a), ~7 weeks, when tumors developed resistance. c Heatmap of statistically significant (FDR < 0.05) differentially expressed mRNAs related to collagen genes in 344SQ tumors treated weekly with anti-PD-L1 or isotype control, starting after one week of implantation. Tumors samples and RNA profiling data were obtained from a prior study8 at treatment endpoint, ~8 weeks, when tumors developed resistance. d Representative Masson’s trichrome stains and quantification of percent collagen area per field of untreated 344SQ tumors collected at 1 or 3 weeks post-implantation in mice; n = 5 tumors per time point and four microscopy fields per tumor sample were analyzed. Scale bars, 50 µm. Statistics calculated using two-sided student’s t-test. e Representative trichrome stains and quantification of percent collagen area per field of 344SQ tumors treated weekly with IgG isotype control or anti-PD-L1, starting after 1 week of implantation. Tumors were analyzed at endpoint of the experiment in (a); n = 5 tumors per treatment group with 34 total microscopy fields analyzed for IgG Ctrl and 52-total fields analyzed for anti-PD-L1 groups. Scale bars, 50 µm. Statistics calculated using a two-sided student’s t-test. f Left: Representative H&E stains and SHG microscopy, including SHG quantification of percent collagen area per field of lung tumor tissues from KrasG12D;p53-/- (KP) mice treated weekly with anti-PD-1, -PD-L1 or isotype control for 12–14 weeks; n = 5 lung tissues per treatment group with 45 (isotype Ctrl), 15 (anti-PD-L1), and 35 (anti-PD-1) total microscopy fields analyzed across all tissues in respective groups. Scale bars, 100 µm. Data presented as mean +/− SD. Statistics calculated using one-way ANOVA post hoc Tukey for multiple comparisons. *P < 0.05.
