Journal: bioRxiv

Article Title: Mono-ADP-ribosylation-driven immunosuppression and cross-resistance to therapy through cancer cell intrinsic and extrinsic mechanisms

doi: 10.64898/2026.06.01.729331

Figure Lengend Snippet: (A) Flow cytometry analysis of the frequency (% of live CD45 + cells) of CD4 + T cells, CD8 + T cells, and NK1.1 + or CD49b + cells in the spleens from C57BL6 or Balb/c mice bearing subcutaneous tumors described in . (B) Gating strategy for immune profiling of tumor and splenic tissues. (C) Flow cytometry analysis of the frequency (% of live CD45 + cells) and numbers of tumor-associated macrophages (TAMs, CD11b + F4/80 + ), polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs, CD11b + Ly6G + ), and monocytic myeloid-derived suppressor cells (M-MDSCs, CD11b + Ly6C + ), in subcutaneous MC38 or CT26 tumors described in and . (D) Flow cytometry analysis of the frequency and number of CD4 + T cells and NK cells in subcutaneous MC38 or CT26 tumors described in and . (E) Flow cytometry analysis of the percentage of IFN-γ + , Granzyme B + , and perforin + OT-1 T cells in the co-culture system described in (n = 3). (F) Flow cytometry analysis of MFI of H2-KbDb on splenocytes isolated from spleens of WT or Parp11 −/− mice (n = 3). (G) Flow cytometry analysis of MFI of H2-KbDb on MC38 cells pretreated with adenosine (100 μM), epinephrine (10 μM), GLP-1 ( – ) (100 nM), or vehicle for 4 h (n = 3). Data are presented as fold change relative to the vehicle group. Each dot represents one biological replicate. Data are presented as mean ± SEM. Statistical significance was determined using two-tailed unpaired Student’s t test (A, C–G).

Article Snippet: OT-1 donor mice (C57BL6-Tg (TcraTcrb)1100Mjb/J, RRID: IMSR_JAX:003831) were purchased from The Jackson Laboratory.

Techniques: Flow Cytometry, Derivative Assay, Co-Culture Assay, Isolation, Two Tailed Test

Journal: bioRxiv

Article Title: Mono-ADP-ribosylation-driven immunosuppression and cross-resistance to therapy through cancer cell intrinsic and extrinsic mechanisms

doi: 10.64898/2026.06.01.729331

Figure Lengend Snippet: (A) Flow cytometry analysis of the frequency (% of live CD45 + cells) and the number (per gram of tumor tissue) of CD8 + T cells from subcutaneous MC38 or CT26 tumors overexpressing wild-type PARP11, PARP11 HY mutant, or EV control in C57BL6 (n = 5) or Balb/c (n = 7) mice, as described in and . (B) Flow cytometry analysis of the percentage of IFN-γ + and Granzyme B + CD8 + T cells isolated from subcutaneous MC38 or CT26 tumors overexpressing wild-type PARP11, PARP11 HY mutant, or EV control in C57BL6 (n = 5) or Balb/c (n = 7) mice, as described in and . (C) Killing of MC38-OVA-luc cells pre-treated or not with ITK-7 (1 μM, 48 h) or vehicle, followed by adenosine (100 μM), or epinephrine (10 μM), or GLP-1 (100 nM) before co-culture with OT-1 CTLs for 12 h (OT-1: MC38-OVA-luc = 5:1; n = 3). (D) Killing of MC38-OVA-luc cells or B16F10-OVA-luc cells pretreated with ITK-7 (1 μM, 48 h) or vehicle prior to co-culture with OT-1 CTLs for 12 h (OT-1: MC38-OVA-luc = 5:1; OT-1: B16F10-OVA-luc = 5:1; n = 3). (E) Flow cytometry analysis of mean fluorescence intensity (MFI) of H2-Kd on malignant (CD45 − Podoplanin + ) cells from subcutaneous CT26 tumors overexpressing wild-type PARP11, PARP11 HY mutant, or EV control in Balb/c mice (n = 7), as described in . (F) Flow cytometry analysis of MFI of H2-KbDb on malignant (CD45 − EpCAM + ) cells from subcutaneous MC38 tumors overexpressing wild-type PARP11, PARP11 HY mutant, or EV control in C57BL6 mice (n = 5), as described in . (G) Flow cytometry analysis of MFI of H2-KbDb on malignant (CD45 − EpCAM + ) cells from subcutaneous MC38 tumors expressing sgParp11 or NTC in C57BL6 mice (n = 5), as described in . (H) Flow cytometry analysis of MFI of H2-Kd on malignant (CD45 − PDPN + ) cells from subcutaneous CT26 tumors treated as described in (n = 8). Each dot represents one biological replicate. Data are presented as mean ± SEM. Statistical significance was determined using two-tailed unpaired Student’s t test (A–H).

Article Snippet: OT-1 donor mice (C57BL6-Tg (TcraTcrb)1100Mjb/J, RRID: IMSR_JAX:003831) were purchased from The Jackson Laboratory.

Techniques: Flow Cytometry, Mutagenesis, Control, Isolation, Co-Culture Assay, Fluorescence, Expressing, Two Tailed Test

Journal: bioRxiv

Article Title: Mono-ADP-ribosylation-driven immunosuppression and cross-resistance to therapy through cancer cell intrinsic and extrinsic mechanisms

doi: 10.64898/2026.06.01.729331

Figure Lengend Snippet: (A) MC38-OVA cells with endogenous Parp11 knockout were reconstituted with wild-type PARP11, PARP11 HY mutant, or EV control, followed by overexpression of H2-Kb and β2-microglobulin to restore equivalent MHC-I surface levels. Killing of these cells by co-cultured OT-1 CTLs (12 h, OT-1: MC38-OVA = 1:1) was assessed by flow cytometry analysis of the frequency of dying 7-AAD + MC38-OVA cells (n = 3 per group). (B) Flow cytometry analysis of the frequency (% of live CD45 + cells) of CD8 + T cells in subcutaneous MC38 tumors in C57BL6 mice (n = 8). MC38 cells overexpressing wild-type PARP11 or EV control were further transduced with H2-Kb and β2-microglobulin to equalize tumor cell surface MHC-I levels. (C) Flow cytometry analysis of the percentage of Granzyme B + , IFN-γ + , and Perforin + CD8 + T cells isolated from subcutaneous MC38 tumors in C57BL6 mice treated as described in (B). (D) Tumor growth curves of subcutaneous MC38 tumors in C57BL6 mice (n = 8 per group) treated as described in (B). (E) Representative tumor images and tumor weights of subcutaneous MC38 tumors from C57BL6 mice (n = 8 per group) treated as described in (B). Each dot represents one biological replicate. Data are presented as mean ± SEM. Statistical significance was determined using two-tailed unpaired Student’s t test for (A–C) and (E), and one-way ANOVA followed by Tukey’s multiple-comparisons test for (D).

Article Snippet: OT-1 donor mice (C57BL6-Tg (TcraTcrb)1100Mjb/J, RRID: IMSR_JAX:003831) were purchased from The Jackson Laboratory.

Techniques: Knock-Out, Mutagenesis, Control, Over Expression, Cell Culture, Flow Cytometry, Transduction, Isolation, Two Tailed Test