Journal: bioRxiv

Article Title: Serial Thermal Ablation Induces Abscopal Antitumor Immunity and Reveals Targetable CSF1R-Dependent Resistance in Pancreatic Cancer

doi: 10.64898/2026.04.05.713683

Figure Lengend Snippet: A) Representative 20x NK1.1 immunohistochemistry staining images. Scale bars 100 µm. B) NK1.1 staining is significantly increased in 3 RFA contralateral tumors compared to 1 RFA contralateral tumors (***p<0.0001). NK1.1 is significantly increased in 3 RFA-treated tumors compared to 3 RFA contralateral tumors (*p=0.0141). n=10 fields analyzed per tumor. C) Representative 20x immunofluorescence images of CD4 (purple), CD8α (red), GZMB (green), DAPI nuclear stain (blue). D) CD4 + GZMB + cells are not increased in 3 RFA contralateral tumors. E) CD8α + GZMB + cells are significantly increased in the three RFA contralateral tumors compared to 1 RFA contralateral (****p<0.0001) and 3 RFA RFA-treated tumors (**p=0.0046). n=8 fields analyzed per group. F) Schematic of the protocol to evaluate the effects of CD8 depletion on serial RFA-treated and contralateral tumors. Created using BioRender . G) CD8α depletion significantly reduces the %CD8α + cells per live cells in RFA-treated tumors. H) α-CD8α significantly increases the tumor volume of RFA-treated (*p<0.05; ***p<0.001) and I) contralateral tumors after 2 RFA treatments but does not significantly increase the final tumor volumes after the third RFA treatment. Statistics were done using Prism GraphPad software.

Article Snippet: The following primary antibodies were used: CD4 (1:35, ab288724, abcam), CD8α (1:50, MAB116-100, R&D Systems), GZMB (1:100, AF1865, R&D Systems).

Techniques: Immunohistochemistry, Staining, Immunofluorescence, Software

Journal: bioRxiv

Article Title: Serial Thermal Ablation Induces Abscopal Antitumor Immunity and Reveals Targetable CSF1R-Dependent Resistance in Pancreatic Cancer

doi: 10.64898/2026.04.05.713683

Figure Lengend Snippet: A) Experimental design setup. B) Serial thermal ablation using RFA in combination with anti-PD-L1 and Quemli does not significantly reduce the volume of treated tumors. C) However, serial RFA in combination with anti-PD-L1 and Quemli significantly reduces the tumor volume of contralateral tumors compared to serial RFA + IgG+Vehicle (*p<0.05) and compared to serial RFA + Quemli treated alone (****p<0.0001). D) Representative 20x CSF1R immunohistochemistry images. E) Quantification of CSF1R + cells showed an increase in CSF1R + area per field in serial RFA tumors + anti-PD-L1 with or without Quemli compared to serial RFA + vehicle in both RFA-treated and F) contralateral tumors. A two-way Anova was used in Prism GraphPad for statistical comparisons. G) Experimental design setup. H) Serial thermal ablation using RFA in combination with CSF1R inhibition, anti-PD-L1 and Quemli significantly reduces the volume of ablated tumors (**p<0.01; ***p<0.001) and I) contralateral tumors (**p<0.01; ***p<0.001). J) Combination treatment using 3 RFA + anti-PD-L1, Quemli and CSF1R inhibition significantly increases the infiltration of CD8 + T cells in treated (**p<0.01; *p<0.05) and K) contralateral tumors (**p<0.01; *p<0.05) . L) Combination treatment using 3 RFA + anti-PD-L1, Quemli and CSF1R inhibition significantly increases GZMB staining in treated (***p<0.001; **p<0.01) and M) contralateral tumors (*p<0.05; **p<0.01; ***p<0.001). A two-way ANOVA in Prism GraphPad was used for statistical analysis. Scale bars 50 µm.

Article Snippet: The following primary antibodies were used: CD4 (1:35, ab288724, abcam), CD8α (1:50, MAB116-100, R&D Systems), GZMB (1:100, AF1865, R&D Systems).

Techniques: Immunohistochemistry, Inhibition, Staining

Journal: Science Advances

Article Title: A stromal PAI1-tPA axis orchestrates immunosuppression in pancreatic cancer

doi: 10.1126/sciadv.aea6734

Figure Lengend Snippet: ( A to I ) Serpine1 +/+ and Serpine1 −/− mice were orthotopically implanted with pancreatic cancer cells (7940B), and tumors were collected at day 21. [(A) and (B)] Flow cytometry quantification of frequencies of total T cells (CD45 + CD3 + ) (A) and conventional dendritic cells (CD45 + F4/80 − CD11c + MHCII + ) (B) among live cells in orthotopic tumors. (C) Co-IF staining for Arg1 (green), F4/80 (red), and DAPI (blue) in orthotopic tumors, and corresponding quantification ( n = 3 FOV per animal). (D) Quantitative RT-PCR analysis of Arg1 in BMDMs treated with vehicle or recombinant PAI1 (rPAI1) for 20 hours. Each dot represents individual primary BMDM lines. (E) Flow cytometry quantification of frequency of CD8 + T cells (CD45 + CD3 + CD8 + ) among live cells in orthotopic tumors. [(F) and (G)] Flow cytometry quantification of frequencies and absolute numbers of GZMB + CD8 + T cells (F) and Ki67 + CD8 + T cells (G) following a 5-hour ex vivo PMA/ionomycin stimulation of tumor digests. (H) Co-IF staining for GZMB (green), CD8 (red), and DAPI (blue) in orthotopic tumors, and corresponding quantification ( n = 5 FOV per animal). (I) Co-IF staining for Ki67 (red), CD8 (green), and DAPI (blue) in orthotopic tumors, and corresponding quantification ( n = 5 FOV per animal). ( J ) Experimental design for CD8 + T cell depletion. Mice received control immunoglobulin G (IgG) or anti-CD8 antibody twice per week, starting 5 days postimplantation (7940B). Tumors were harvested at day 21. ( K ) Tumor weight of orthotopic tumors with CD8⁺ T cell antibody-mediated depletion. Symbols in (A) to (I) and (K) represent individual mice. Data are means ± SEM. P values were determined by two-tailed unpaired t test [(A), (B), right of (C), (D), (E) to (G), and right of (H)], Mann-Whitney test [left of (C), left of (H), and (I)], and Mann-Whitney test with Holm-Sidak post hoc (K).

Article Snippet: Depletion of CD8 + T cells were performed starting from 5 days following orthotopic injection of 7940B cells by administration of intraperitoneal injections of 200 μg of anti-mouse CD8α mAb (2.43, no. BE0061, BioXCell) or control isotype antibody (LTF-2, no. BE0090, BioXCell) twice per week for the entire course of the experiments.

Techniques: Flow Cytometry, Staining, Quantitative RT-PCR, Recombinant, Ex Vivo, Control, Two Tailed Test, MANN-WHITNEY

Journal: Science Advances

Article Title: A stromal PAI1-tPA axis orchestrates immunosuppression in pancreatic cancer

doi: 10.1126/sciadv.aea6734

Figure Lengend Snippet: ( A ) Experimental outline of murine PDAC model. Plat +/+ and Plat −/− mice were orthotopically implanted with pancreatic cancer cells (7940B), and tumors were collected 21 days postimplantation. ( B ) Tumor weight of orthotopic tumors harvested from Plat +/+ and Plat −/− mice. ( C ) ELISA of tPA in tissue supernatants from pancreata of Plat +/+ mice (no tumor) and orthotopic tumors from Plat +/+ and Plat −/− mice. ( D ) Picrosirius red staining of orthotopic tumors, and corresponding quantification ( n = 5 FOV per animal). ( E ) Co-IF staining for cCasp3 (red), ECAD (green), and DAPI (blue) in orthotopic tumors, and corresponding quantification ( n = 4 FOV per animal). ( F to H ) Flow cytometry quantification of frequencies of total T cells (F), CD8 + T cells (G), and conventional dendritic cells (H) among live cells in orthotopic tumors. ( I ) Co-IF staining for Arg1 (green), F4/80 (red), and DAPI (blue) in orthotopic tumors, and corresponding quantification ( n = 3 FOV per animal). Symbols in (B) to (I) represent individual mice. Data are means ± SEM. P values were determined by Mann-Whitney test [(B) and (F)], two-way ANOVA with Holm-Sidak post hoc (C), and two-tailed unpaired t test [(D), (E), and (G) to (I)].

Article Snippet: Depletion of CD8 + T cells were performed starting from 5 days following orthotopic injection of 7940B cells by administration of intraperitoneal injections of 200 μg of anti-mouse CD8α mAb (2.43, no. BE0061, BioXCell) or control isotype antibody (LTF-2, no. BE0090, BioXCell) twice per week for the entire course of the experiments.

Techniques: Enzyme-linked Immunosorbent Assay, Staining, Flow Cytometry, MANN-WHITNEY, Two Tailed Test

Journal: Science Advances

Article Title: A stromal PAI1-tPA axis orchestrates immunosuppression in pancreatic cancer

doi: 10.1126/sciadv.aea6734

Figure Lengend Snippet: ( A ) Experimental outline of murine PDAC model. WT, Serpine1 −/− , and Serpine1 −/− ; Plat −/− mice were orthotopically implanted with pancreatic cancer cells (7940B), and tumors were collected 21 days postimplantation. ( B ) Tumor weight of orthotopic tumors harvested from WT, Serpine1 −/− , and Serpine1 −/− ; Plat −/− mice. ( C ) IHC staining for CD8 in orthotopic tumors, and corresponding quantification ( n = 3 FOV per animal). ( D ) Co-IF staining for Arg1 (green), F4/80 (red), and DAPI (blue) in orthotopic tumors, and corresponding quantification ( n = 3 FOV per animal). Symbols in (B) to (D) represent individual mice. Data are means ± SEM. P values were determined by one-way ANOVA with Holm-Sidak post hoc [(B) to (D)]. ( E ) Working model. Left: Hypoxia stabilizes HIF proteins, inducing PAI1 expression in pancreatic CAFs. Although tPA levels are elevated in PDAC, PAI1 predominantly inhibits its activity, thereby suppressing antitumor immunity. Middle: Elimination of stromal PAI1 restores tPA activity, enhancing antitumor CD8 + T cell responses, accompanied by alleviation of immunosuppressive TAM phenotypes and increased dendritic cell (DC) infiltration. Stromal PAI1-driven tumor growth depends on CD8 + T cells. Right: Removal of stromal tPA abolishes residual tPA activity, further promoting an immunosuppressive TME and tumor growth.

Article Snippet: Depletion of CD8 + T cells were performed starting from 5 days following orthotopic injection of 7940B cells by administration of intraperitoneal injections of 200 μg of anti-mouse CD8α mAb (2.43, no. BE0061, BioXCell) or control isotype antibody (LTF-2, no. BE0090, BioXCell) twice per week for the entire course of the experiments.

Techniques: Immunohistochemistry, Staining, Expressing, Activity Assay