antibodies blocking pd1  (Bio X Cell)

Journal: bioRxiv

Article Title: Tissue Injury and Biomaterial Treatment Modulate Tumor Growth and Response to Immunotherapy

doi: 10.64898/2026.02.02.703323

Figure Lengend Snippet: (A) Experimental scheme consisting of bilateral VML surgery performed on the hindlimb quadriceps muscles, followed by unilateral subcutaneous inoculation of syngeneic cancer cells (CT26 or B16F10) on the right flank. Survival criteria comprised of tumor volume >1500 mm 3 or severe involuting ulceration. (B) CT26 and (C) B16F10 tumor growth kinetics of uninjured and VML-injured mice (n=8-10). (D) Experimental scheme introducing ICB therapy to the concurrent VML injury-tumor model. ICB therapy consisted of either combination αPD1/αCTLA4 (5 mg/kg each) or delayed αPD1 monotherapy (5 mg/kg) delivered via intraperitoneal injection every 3 days for 4 total doses. (E) CT26 tumor growth and survival curves of uninjured and VML injured mice treated with αPD1/αCTLA4 or (F) delayed αPD1 monotherapy. (G) B16F10 tumor growth and survival curves of uninjured and VML injured mice treated with αPD1/αCTLA4. (Statistics) Tumor growth curves: mean±SEM. Bar graphs: mean±SD (displaying earliest survival timepoint). Normally distributed data (Shapiro-Wilk test, α=0.05) was analyzed using an unpaired two-tailed student t-test ( B ); otherwise, a non-parametric two-tailed Mann-Whitney test was used ( C, F, G ). Results representative of at least 2 independent experiments ( B, C ). Survival: Kaplan-Meier curve with Log-Rank Mantel-Cox test ( E-G ). Survival results combined from 2 independent experiments with separate results presented in supplemental figure 2 ( E-G ). NS: Not significant p>0.05, * p<0.05, ** p<0.01, *** p<0.001. CR: Complete Responder.

Article Snippet: ICB therapy consisted of monoclonal antibodies blocking PD1 (clone RPM1-14, BioXCell BP0146) and CTLA4 (clone 9H10, BioXCell BP0131) delivered via intraperitoneal (IP) injection at 5mg/kg body weight each prepared in sterile dilution buffer ( InVivo Pure pH 7.0, BioXCell IP0070).

Techniques: Muscles, Injection, Two Tailed Test, MANN-WHITNEY

Journal: bioRxiv

Article Title: Tissue Injury and Biomaterial Treatment Modulate Tumor Growth and Response to Immunotherapy

doi: 10.64898/2026.02.02.703323

Figure Lengend Snippet: VML Injury Impacts Anti-Tumor CD8 + T Cell Response. (A) Flow cytometric profiling of CT26 tumor-infiltrating CD8 + T cells, (B) PD1 expression on CD8 + T cells, and (C) CT26 tumor-specific CD8 + T cells marked by AH1-loaded MHC class I tetramer in uninjured and VML injured mice. (D) Single-cell RNA-sequencing (scRNAseq) of CT26 tumor-infiltrating T cells from uninjured and VML injured mice. UMAP depicting main T cell clusters (e.g., CD8, CD4, Treg, unconventional). (E) T cell UMAP separated by tumor-reactive AH1-Tetramer + CD8 + T cells (top) and other T cells (bottom) colored by cells from uninjured (gray) and VML injured (green) mice. (F) Differential gene expression and (G, H) GSEA using Hallmark or (I) curated list of T cell-related pathways with top leading edge genes comparing tumor-reactive AH1-Tetramer + CD8 + T cells of uninjured and VML injured mice. (J) Intracellular cytokine staining following ex vivo stimulation of IFNγ-producing effector cell types in CT26 tumors of uninjured and VML injured mice. (Statistics) Bar graphs: mean±SD. Normally distributed data (Shapiro-Wilk test, α=0.05) was analyzed using an unpaired two-tailed student t-test ( A-C, J ). Results representative of at least 2 independent experiments ( A-C ). NS: Not significant p>0.05, * p<0.05, ** p<0.01. UMAP: Uniform manifold approximation and projection. GSEA: Gene set enrichment analysis.

Article Snippet: ICB therapy consisted of monoclonal antibodies blocking PD1 (clone RPM1-14, BioXCell BP0146) and CTLA4 (clone 9H10, BioXCell BP0131) delivered via intraperitoneal (IP) injection at 5mg/kg body weight each prepared in sterile dilution buffer ( InVivo Pure pH 7.0, BioXCell IP0070).

Techniques: Expressing, Single Cell, RNA Sequencing, Gene Expression, Staining, Ex Vivo, Two Tailed Test

Journal: bioRxiv

Article Title: Tissue Injury and Biomaterial Treatment Modulate Tumor Growth and Response to Immunotherapy

doi: 10.64898/2026.02.02.703323

Figure Lengend Snippet: Treatment of Injury with Pro-Regenerative Biologics Impacts Cancer Progression. (A) CT26 tumor growth kinetics of uninjured, untreated VML-injured, and extracellular matrix (ECM) scaffold-treated VML-injured mice (n=6-8). (B) Experiment compilation of CT26 tumor volume fold changes (normalized to the uninjured group within each experiment) of untreated and ECM-treated VML-injured groups at day 14 post-inoculation (n=10 independent experiments). (C) B16F10 tumor growth kinetics of uninjured, untreated VML-injured, and ECM-treated VML-injured mice (n=8). (D) CT26 tumor growth kinetics of uninjured, untreated VML-injured, and Schistosoma mansoni regenerative soluble egg antigen (rSEA)-treated VML-injured mice (n=8). (E) Survival curves of untreated and ECM-treated VML-injured mice bearing CT26 tumors treated with delayed αPD1 monotherapy or (F) bearing B16F10 tumors treated with αPD1/αCTLA4. (G) Schematic of patient selection criteria for retrospective cohort study using TriNetX database. (H) Overall survival curve for breast cancer patients treated with ICB therapy and complete mastectomy (CPT 19303), with versus without biological implant placement (CPT 15777), on or up to 3 months after ICB therapy. (I) Summary table of distribution of common cancer treatments (e.g., chemotherapy, radiation) between both patient cohorts. (Statistics) Tumor growth curves: mean±SEM. Bar graphs: mean±SD (earliest survival timepoint or experiment endpoint). Experiment Compilation: Each datapoint represents an independent experiment, with fold change represented as geometric mean±geometric SD on a Log2 scale ( B ). Data was analyzed using an ordinary one-way ANOVA with Tukey’s multiple comparisons test ( A-D ). Survival: Kaplan-Meier curve with Log-Rank Mantel-Cox test ( E, F, I ). Survival results combined from 2 independent experiments with separate results presented in supplemental figures ( E, F ). Survival experiments were conducted in tandem with uninjured controls, thereby untreated VML data are the same as in and ( E, F ). Categorical data in clinical cancer treatment table was analyzed using Fisher’s exact test ( I ). NS: Not significant p>0.05, * p<0.05, ** p<0.01. CR: Complete Responder.

Article Snippet: ICB therapy consisted of monoclonal antibodies blocking PD1 (clone RPM1-14, BioXCell BP0146) and CTLA4 (clone 9H10, BioXCell BP0131) delivered via intraperitoneal (IP) injection at 5mg/kg body weight each prepared in sterile dilution buffer ( InVivo Pure pH 7.0, BioXCell IP0070).

Techniques: Selection

Journal: Nature Communications

Article Title: Cytokine-armed pyroptosis induces antitumor immunity against diverse types of tumors

doi: 10.1038/s41467-024-55083-3

Figure Lengend Snippet: a In vitro screening of IFNγ cotransfected with GSDMD variants. HEK293T cells were transfected with IFNγ and GSDMD variants and LDH release, PI uptake, and ELISA assays were conducted. b Experimental scheme. Subcutaneously growing tumors were electroporated with 20 μg IFNγ ( n = 8) or empty vector-coding plasmid followed by electroporation of an equal amount of NT GSDMD plasmid after 2 days ( n = 9). The same NT GSDMD group as in Fig. is shown. Kaplan-Meier curve ( c ) and tumor volume changes ( d ) present the NT GSDMD + IFNγ treatment efficacy compared to the NT GSDMD treatment. e – j Anti-PD1 but not anti-CTLA4 restores antitumorigenic activity of fully deconstructed inflammasome (NT GSDMD + IL-1β + IL-18). Mice received i.p. 200 μg anti-PD1 ( n = 6) ( e ) or anti-CTLA4 ( n = 4) antibodies ( h ) with appropriate isotype controls on the same day as IL-1β + IL-18 or empty vector control electroporation, and every 3rd day until remission or euthanization. Survival ( f , i ) and tumor volume ( g , j ) were documented over time. Results shown in ( a ) are mean ± SEM representative of three independent experiments. Time in ( c , d , f , g , i , j ) is determined as days after the treatment, and a log-rank test was conducted for survival data analysis ( c , f , i ). Source data are provided as a file. Figure 7b created in BioRender. Hafner Bratkovic, I. (2024) https://BioRender.com/z97l709 ; Fig. 7e created in BioRender. Hafner Bratkovic, I. (2024) https://BioRender.com/a77b958 ; Fig. 7h created in BioRender. Hafner Bratkovic, I. (2024) https://BioRender.com/n94v358 .

Article Snippet: Mice were administered i.p. 200 μg of PD1 blocking antibodies (anti-mouse CD279, clone RMP1-14, product no. P362, Leinco Technologies) or rat IgG2a (clone 1-1, product no. I-1177, Leinco Technologies) for the isotype control, or CTLA4 blocking antibodies (anti-mouse CTLA4, clone 9D9, C2855, Leinco Technologies) or mouse IgG2b isotype control (clone MCP-11, product no. I-119, Leinco Technologies), the same day as the first treatment and again every 3 days until mice were euthanized or no palpable tumor was detected.

Techniques: In Vitro, Transfection, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Electroporation, Activity Assay, Control

Journal: bioRxiv

Article Title: Collagen modification remodels the sarcoma tumor microenvironment and promotes resistance to immune checkpoint inhibition

doi: 10.1101/2024.06.28.601055

Figure Lengend Snippet: A . Representative longitudinal cytolysis curves of shScr or shPLOD2 human STS-109 UPS cells co-cultured with CART-TnMUC1 cells from 1 independent human donor. Measurements indicate percent target (UPS) cell cytolysis. B . Quantification of data from A . Statistics are not shown because n < 3 (n = 2). C . Growth curves of subcutaneous (flank) syngeneic tumors in C57BL/6 mice treated with minoxidil (or vehicle control), alone or in combination with ɑ-Pd1 (or isotype control) antibodies. Growth curves were fit with non-linear regression (exponential fit) models; pairwise curve fitting comparisons (extra sum-of-squares F test) are shown in the table at right. D . Kaplan-Meier survival curves of KP mice treated with minoxidil, alone or in combination with α-Pd1 checkpoint therapy. Statistics are not shown because data from the control and α-Pd1-alone groups are from a historical cohort first reported in.

Article Snippet: In immune checkpoint studies, 200 μg of anti-Pd1 monoclonal blocking antibody (BE0146, BioXCell) or isotype control antibody (BE0089, BioXCell) was administered I.P. every three days once tumors became palpable.

Techniques: Cell Culture, Control

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: ( A ) Sham surgery or SLNB was performed on C57Bl6 mice (day -10) prior to orthotopic tumor implantation (day 1). Mice were treated by rat IgG control or anti-PD1 antibody (day 7) three times (every 2 days for B16F10 or every 3 days for E0771, 10mg/kg). Tumor volume ( left & middle ) and overall survival ( right ) were plotted for mice bearing orthotopic B16F10 ( top ) or E0771 ( bottom ) tumors, respectively. ( B ) To evaluate the impact of more aggressive surgery, sham surgery or CLND was performed followed by tumor implantation and subsequent anti-PD1 inhibition using the same regimen described above. Tumor volume ( left ) and overall survival ( right ) were plotted for mice bearing B16F10 ( top ) or E0771 ( bottom ) tumors, respectively. ( C ) In contrast with pre-emptive approach, impact of post-tumor implantation LN dissection was also assessed. After tumor implantation (day 1), sham surgery, SLNB or CLND was performed on C57Bl6 mice (day 6) right before anti-PD1 treatment (day 7). Tumor volume ( left ) and overall survival ( right ) were plotted for mice bearing B16F10 ( top ) and E0771 ( bottom ) tumors. ( D ) Frequency of complete responders from pre-emptive and post-tumor TDLN removal studies were plotted. Fisher’s exact test was used for statistical analysis. ( E ) B16F10 ( top ) or E0771 ( bottom ) tumor response to anti-PD1 treatment was tested in Lta tm1Dc mice. ICB response of wildtype (WT) sham mice from ( A ) SLNB study was plotted in dashed lines for comparison. Tumor volume ( left ) and overall survival ( right ) were plotted. In all animal studies, “(n)” indicates number of mice used. Log-rank test with Bonferroni test was used for statistical analysis of survival data. * p <0.05, **** p <0.0001.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation, Inhibition, Dissection, Comparison

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: ( A ) For pre-emptive setting, sham surgery or CLND was performed on C57Bl6 mice (day -10) prior to E0771 tumor implantation to dermis (day 1). Mice were treated by anti-PD1 inhibition (day 7) for three times at 10mg/kg. Tumor volume ( left ) and overall survival ( right ) were plotted. ( B ) For post-tumor setting, after E0771 tumor implantation to dermis (day 1), sham surgery, SLNB or CLND was performed on C57Bl6 mice (day 6) right before anti-PD1 treatment. Tumor volume ( left ) and overall survival ( right ) were plotted. In all animal studies, “(n)” indicates the number of mice used. Log-rank test with Bonferroni test was used for statistical analysis of survival data. * p <0.05.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation, Inhibition

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: To assess the tumor-specific T cell response, B16F10 tumors overexpressing Ovabulmin257-264 (OVA) were implanted to sham and CLND mice followed by anti-PD1 inhibition. ( A ) In ipsilateral and contralateral LNs as well as ( B ) spleens, CD8 T cell expansion is shown by immunofluorescence images. ( C ) Additionally, OVA-specific CD8 T cells in ipsilateral, contralateral and cervical LNs, spleens and blood were determined by tetramer staining using flow cytometry analysis. Gating approach and representative density plots from ipsilateral and contralateral LNs are shown. ( D ) Abundance of OVA-specific CD8 T cells (CD8+, Tetramer+) is plotted for various LNs, blood and spleens collected from sham and CLND mice; each dot represents data from a single mouse. ( E ) OVA-specific CD8 T cell response in ipsilateral and contralateral LNs is shown in an adjuvant setting where primary tumor resection was performed with or without CLND before anti-PD1 inhibition. ( F ) Similar to E , Kaede mice bearing B16F10-OVA tumors were treated by adjuvant anti-PD1 inhibition, followed by tumor rechallenge to primary site. Once recurrent tumor formed, contralateral inguinal LNs were photoconverted prior to tissue harvesting for flow analysis. Abundance of Kaede red+ tumor infiltrating OVA-specific CD8 T cells is plotted. ( G ) To assess antigen-presenting cell-mediated T cell response after LN dissection, BMDCs were pulsed with OVA and LPS for 1 day, and injected to the footpad of mice that underwent sham surgery or ILND. OVA-specific CD8 T cells in ipsilateral and contralateral LNs are plotted. In this figure, fluorescence signal quantification is calculated by the occupied area, while DAPI is used to normalize. Two tailed student t test (two conditions) and one way ANOVA with Tukey post analysis (more than two conditions) were used for statistical analysis. * p <0.05, ** p <0.01, **** p <0.0001.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Inhibition, Immunofluorescence, Staining, Flow Cytometry, Adjuvant, Dissection, Injection, Fluorescence, Two Tailed Test

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: ( A ) Sham surgery (SP+ condition) or splenectomy (SP-condition) was performed on mice 14 days prior to CLND. E0771 tumor implantation and IgG or anti-PD1 treatment were conducted using same procedure as before. Tumor volume ( left ) and overall survival ( right ) were plotted. ( B ) E0771 tumor rechallenge was performed using complete ICB responders. Tumor volume was plotted when tumor was palpable (after day 7) and when the study was ended. In all animal studies, “(n)” indicates the number of mice used. Log-rank test with Bonferroni test was used for statistical analysis of survival data. * p <0.05.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: Sham surgery or CLNB was performed on C57Bl6 mice (day -10) prior to orthotopic E0771 tumor implantation (day 1). IgG control or combination of anti-PD1 and anti-CTLA4 antibodies were intradermally delivered to ipsilateral TDLNs or contralateral LNs in sham and CLND conditions, respectively (day 7) for three doses at 5mg/kg. LNs exposed to treatment were harvested for flow cytometry analysis measuring CD45, TCR-beta, CD4, CD8, CD44, PD-1, TCF-1, CD39 and CD62L. Non-tumor bearing naïve mice are used as control. ( A ) LN Volume of naïve mice or mice treated by IgG or ICB combination is plotted. ( B ) Principle component analysis is performed on naïve or treatment-exposed LNs, using the abundance of antigen-experienced T cells (CD44+, PD-1+), memory T cells (CD44+, PD-1+, TCF1+), central memory T cells (CD44+, PD-1+, TCF1+, CD62L+), effector memory T cells (CD44+, PD-1+, TCF1+, CD62L-), stem-like effector memory T cells (CD44+, PD-1+, TCF1+, CD62L-, CD39-) and their CD4/CD8 subsets. ( C ) The percentage of stem-like effector memory T cells to total T cells is plotted. ( D ) Using the same surgery, tumor implantation and treatment procedure, ICB was given intraperitoneally or intradermally to the lymphatic basin of contralateral inguinal LN (day 7) for three times at 5mg/kg in sham and CLND mice. Tumor volume ( left ) and overall survival ( right ) were plotted. ( E ) Timeline for neoadjuvant-adjuvant treatment and tumor rechallenge. ( F ) Representative images of vitiligo (arrows) that developed in mice after neoadjuvant treatment ( left ). Percentage of mice with local, distant or local & distant vitiligo across conditions ( right ). ( G ) Overall survival of mice treated by neoadjuvant ICB with various LN surgery status. ( H ) Tumor-free survival of mice previously treated by neoadjuvant ICB after tumor rechallenge. Naïve mice were used as control. ( I ) Overall survival of mice previously treated by neoadjuvant-adjuvant ICB after tumor rechallenge. Of note, only mice that developed tumors after rechallenge were given adjuvant treatment. In all studies, “(n)” indicates the number of mice used. One way ANOVA with Tukey post analysis (comparing more than two conditions) and Log-rank test with Bonferroni test (survival) were used for statistical analysis. * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation, Flow Cytometry, Adjuvant

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: ( A ) Sham surgery or SLNB was performed on C57Bl6 mice (day -10) prior to orthotopic tumor implantation (day 1). Mice were treated by rat IgG control or anti-PD1 antibody (day 7) three times (every 2 days for B16F10 or every 3 days for E0771, 10mg/kg). Tumor volume ( left & middle ) and overall survival ( right ) were plotted for mice bearing orthotopic B16F10 ( top ) or E0771 ( bottom ) tumors, respectively. ( B ) To evaluate the impact of more aggressive surgery, sham surgery or CLND was performed followed by tumor implantation and subsequent anti-PD1 inhibition using the same regimen described above. Tumor volume ( left ) and overall survival ( right ) were plotted for mice bearing B16F10 ( top ) or E0771 ( bottom ) tumors, respectively. ( C ) In contrast with pre-emptive approach, impact of post-tumor implantation LN dissection was also assessed. After tumor implantation (day 1), sham surgery, SLNB or CLND was performed on C57Bl6 mice (day 6) right before anti-PD1 treatment (day 7). Tumor volume ( left ) and overall survival ( right ) were plotted for mice bearing B16F10 ( top ) and E0771 ( bottom ) tumors. ( D ) Frequency of complete responders from pre-emptive and post-tumor TDLN removal studies were plotted. Fisher’s exact test was used for statistical analysis. ( E ) B16F10 ( top ) or E0771 ( bottom ) tumor response to anti-PD1 treatment was tested in Lta tm1Dc mice. ICB response of wildtype (WT) sham mice from ( A ) SLNB study was plotted in dashed lines for comparison. Tumor volume ( left ) and overall survival ( right ) were plotted. In all animal studies, “(n)” indicates number of mice used. Log-rank test with Bonferroni test was used for statistical analysis of survival data. * p <0.05, **** p <0.0001.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation, Inhibition, Dissection, Comparison

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: ( A ) For pre-emptive setting, sham surgery or CLND was performed on C57Bl6 mice (day -10) prior to E0771 tumor implantation to dermis (day 1). Mice were treated by anti-PD1 inhibition (day 7) for three times at 10mg/kg. Tumor volume ( left ) and overall survival ( right ) were plotted. ( B ) For post-tumor setting, after E0771 tumor implantation to dermis (day 1), sham surgery, SLNB or CLND was performed on C57Bl6 mice (day 6) right before anti-PD1 treatment. Tumor volume ( left ) and overall survival ( right ) were plotted. In all animal studies, “(n)” indicates the number of mice used. Log-rank test with Bonferroni test was used for statistical analysis of survival data. * p <0.05.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation, Inhibition

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: To assess the tumor-specific T cell response, B16F10 tumors overexpressing Ovabulmin257-264 (OVA) were implanted to sham and CLND mice followed by anti-PD1 inhibition. ( A ) In ipsilateral and contralateral LNs as well as ( B ) spleens, CD8 T cell expansion is shown by immunofluorescence images. ( C ) Additionally, OVA-specific CD8 T cells in ipsilateral, contralateral and cervical LNs, spleens and blood were determined by tetramer staining using flow cytometry analysis. Gating approach and representative density plots from ipsilateral and contralateral LNs are shown. ( D ) Abundance of OVA-specific CD8 T cells (CD8+, Tetramer+) is plotted for various LNs, blood and spleens collected from sham and CLND mice; each dot represents data from a single mouse. ( E ) OVA-specific CD8 T cell response in ipsilateral and contralateral LNs is shown in an adjuvant setting where primary tumor resection was performed with or without CLND before anti-PD1 inhibition. ( F ) Similar to E , Kaede mice bearing B16F10-OVA tumors were treated by adjuvant anti-PD1 inhibition, followed by tumor rechallenge to primary site. Once recurrent tumor formed, contralateral inguinal LNs were photoconverted prior to tissue harvesting for flow analysis. Abundance of Kaede red+ tumor infiltrating OVA-specific CD8 T cells is plotted. ( G ) To assess antigen-presenting cell-mediated T cell response after LN dissection, BMDCs were pulsed with OVA and LPS for 1 day, and injected to the footpad of mice that underwent sham surgery or ILND. OVA-specific CD8 T cells in ipsilateral and contralateral LNs are plotted. In this figure, fluorescence signal quantification is calculated by the occupied area, while DAPI is used to normalize. Two tailed student t test (two conditions) and one way ANOVA with Tukey post analysis (more than two conditions) were used for statistical analysis. * p <0.05, ** p <0.01, **** p <0.0001.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Inhibition, Immunofluorescence, Staining, Flow Cytometry, Adjuvant, Dissection, Injection, Fluorescence, Two Tailed Test

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: ( A ) Sham surgery (SP+ condition) or splenectomy (SP-condition) was performed on mice 14 days prior to CLND. E0771 tumor implantation and IgG or anti-PD1 treatment were conducted using same procedure as before. Tumor volume ( left ) and overall survival ( right ) were plotted. ( B ) E0771 tumor rechallenge was performed using complete ICB responders. Tumor volume was plotted when tumor was palpable (after day 7) and when the study was ended. In all animal studies, “(n)” indicates the number of mice used. Log-rank test with Bonferroni test was used for statistical analysis of survival data. * p <0.05.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation

Journal: bioRxiv

Article Title: Cancer immunotherapy responses persist after lymph node resection

doi: 10.1101/2023.09.19.558262

Figure Lengend Snippet: Sham surgery or CLNB was performed on C57Bl6 mice (day -10) prior to orthotopic E0771 tumor implantation (day 1). IgG control or combination of anti-PD1 and anti-CTLA4 antibodies were intradermally delivered to ipsilateral TDLNs or contralateral LNs in sham and CLND conditions, respectively (day 7) for three doses at 5mg/kg. LNs exposed to treatment were harvested for flow cytometry analysis measuring CD45, TCR-beta, CD4, CD8, CD44, PD-1, TCF-1, CD39 and CD62L. Non-tumor bearing naïve mice are used as control. ( A ) LN Volume of naïve mice or mice treated by IgG or ICB combination is plotted. ( B ) Principle component analysis is performed on naïve or treatment-exposed LNs, using the abundance of antigen-experienced T cells (CD44+, PD-1+), memory T cells (CD44+, PD-1+, TCF1+), central memory T cells (CD44+, PD-1+, TCF1+, CD62L+), effector memory T cells (CD44+, PD-1+, TCF1+, CD62L-), stem-like effector memory T cells (CD44+, PD-1+, TCF1+, CD62L-, CD39-) and their CD4/CD8 subsets. ( C ) The percentage of stem-like effector memory T cells to total T cells is plotted. ( D ) Using the same surgery, tumor implantation and treatment procedure, ICB was given intraperitoneally or intradermally to the lymphatic basin of contralateral inguinal LN (day 7) for three times at 5mg/kg in sham and CLND mice. Tumor volume ( left ) and overall survival ( right ) were plotted. ( E ) Timeline for neoadjuvant-adjuvant treatment and tumor rechallenge. ( F ) Representative images of vitiligo (arrows) that developed in mice after neoadjuvant treatment ( left ). Percentage of mice with local, distant or local & distant vitiligo across conditions ( right ). ( G ) Overall survival of mice treated by neoadjuvant ICB with various LN surgery status. ( H ) Tumor-free survival of mice previously treated by neoadjuvant ICB after tumor rechallenge. Naïve mice were used as control. ( I ) Overall survival of mice previously treated by neoadjuvant-adjuvant ICB after tumor rechallenge. Of note, only mice that developed tumors after rechallenge were given adjuvant treatment. In all studies, “(n)” indicates the number of mice used. One way ANOVA with Tukey post analysis (comparing more than two conditions) and Log-rank test with Bonferroni test (survival) were used for statistical analysis. * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001.

Article Snippet: After 7 days, anti-PD1 blocking antibody (Bio X cell, Cat# BP0146) at 10mg/kg was given intraperitoneally or anti-PD1 and anti-CTLA4 (Bio X cell, Cat# BP0131) blocking antibodies each at 5mg/kg were given intraperitoneally or intradermally.

Techniques: Tumor Implantation, Flow Cytometry, Adjuvant