anti cd8  (Bio X Cell)

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Kaplan-Meier curve of GBM patients on standard of care (SOC; red, median=12.4 months) or combinations of SCO with metformin (blue, media=13 months) or DPP4 inhibitors (DPP4i, median=15.9 months) based on the SEER dataset. (B) DPP4 expression from immune lineages as determined by the scRNA-seq of 13 brain tumour patients (3 Grade II; 1 Grade III-IV; 9 GBM). Mean fluorescence intensity (MFI) of DPP-4 from (C) tumor-infiltrating and (D) splenic immune populations. n=5; *** p<0.001 by two-way ANOVA. (E) Representative gating strategy used to define effector, progenitor exhausted (PEX), or terminally exhausted (TEX) populations. (F) DPP-4 MFI from effector, PEX, and TEX populations. n=8; ** p<0.01 by two-way ANOVA. (G) Exhausted CD8 + T cells from GBM and tumor-adjacent tissue were defined as: CD45 + CD11b - CD3 + PD1 + TIGIT + . DPP-4 MFI from TOX + and TOX - population. n=4; * p<0.05 by paired t-test.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Expressing, Fluorescence

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Gating strategy to identify immune populations from healthy human PBMCs. (B) DPP-4 geometric MFI in memory CD8 + T cells from healthy donors. n=4; * p<0.05, ** p<0.01, *** p<0.001 by ordinary one-way ANOVA. (C) Diagram showing how naïve CD45.1 + p14 CD8 + T cells were adoptively transferred into recipient wild-type CD45.2 + mice. Recipient mice were infected with 2 x 10 5 plaque forming units (PFU) of LCMV Armstrong intraperitoneally (i.p.) or with 4 x 10 6 LCMV clone 13 intravenously (i.v.) the day after. Peripheral blood was collected on day 16 and 21 to confirm infection, and spleens collected on day 35 for endpoint immune analysis . (D) DPP-4 MFI from splenic CD8 + T cells infected with two different LCMV strains to promote effector, activated, exhausted, and memory differentiation. Clone 13, n=11, Amstrong, n=4; * p<0.05, **** p<0.0001, ns=not significant by one-way ANOVA. (E) Splenic CD8 + T cells from OT-I transgenic mice were activated with 10 ng OVA for two days or repetitively stimulated for five days to drive exhaustion. (F) Gating strategy used to define exhausted CD8 + T cells within human high-grade gliomas and surrounding tissue.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Infection, Transgenic Assay

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Representative histogram depicting CFSE dilution of mouse CD8 + T cells activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of 100 μM sitagliptin, saxagliptin, or vehicle control for 4 days. (B) The percentage of proliferating mouse CD8 + T cells treated with sitagliptin vs. control. n=4; ** p<0.01 by paired t-test. (C) Representative zebra plots depicting Granzyme B and IFNγ production from OT-I T cells on Day 5. (D) The percentage of OT-I T cells expressing high levels of Granzyme B or IFNγ. n=4; * p<0.05 by paired t-test. (E) Representative zebra plot showing 7-AAD levels in SB28 and ovalbumin-expressing SB28 (SB28-OVA) cells co-cultured with OT-I CD8 + T cells at a 1:1 ratio. (F) The frequency of lysed wild-type SB28 or SB28-OVA cells cultured with sitagliptin- or vehicle-treated OT-I T cells. n=5; * p<0.05, ns=not significant by paired t-test.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Control, Expressing, Cell Culture

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Mouse CD8 + T cells activated with anti-CD3/CD28 plus 100IU IL-2 in the presence of 100 μM sitagliptin or vehicle control for 4 days were intracellularly stained with Ki67 to determine proliferative capacity. n=4; *** p<0.001 by paired t-test. (B) Human CD8 + T cells isolated from healthy PBMCs were expanded with anti-CD3/CD28 and 100IU IL-2 for 6 days, and CFSE dilution determined by flow cytometry. n=5; * p<0.05 by paired t-test . (C) Schematics of the killing assay. Splenic CD8 + T cells from transgenic OT-I mice were isolated and activated with 10 ng OVA for 2 days, co-cultured at 1:1 E:T with either OVA- or empty vector-expressing cancer cells for six hours and then stained with apoptotic marker 7-AAD to determine cancer cell death by flow cytometry. (D) Mouse CD8 + T cells activated with anti-CD3/CD28 plus 100 IU IL-2 in the presence of 100 μM sitagliptin or vehicle control for 4 days were stained with exhaustion marker, anti-PD-1 and anti-2B4, to determine exhaustion post treatment. n=5; * p<0.05 by paired t-test. (E) Schematics of the LCMV experiment. Naïve CD45.1 + p14 CD8 + T cells were adoptively transferred into recipient wild-type CD45.2 + mice, and then recipient mice were infected with 4 x 10 6 PFU LCMV clone 13 intravenously (i.v.) the day after. Recipient mice were treated intraperitoneally with 20 mg/kg sitagliptin daily for 5 continues doses, 2-day break, and 5 more doses for a total of 10 doses. Mice were then observed daily for indications of endpoint. (F) Splenic CD8 + T cells isolated from (E) at endpoint and stained with exhaustion marker, anti-PD-1 and anti-2B4, to determine sitagliptin induced effect in LCMV Clone 13 model. n=4; * p<0.05 by t-test. (G) Frequency of grandparents for TEX, PEX, and effector populations was determined for OT-I CD8 + T cells repetitively stimulated with 10 ng OVA for 5 days. n=7; * p<0.05, ns=not significant by paired t-test.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Control, Staining, Isolation, Flow Cytometry, Transgenic Assay, Cell Culture, Plasmid Preparation, Expressing, Marker, Infection

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Mouse CD8 + T cells were activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of 100 μM sitagliptin, or vehicle control for 4 days. Top differentially up-regulated pathways in sitagliptin-treated CD8 + T cells based on genes expressed >2-fold with p<0.05. n=6. Mouse CD8 + T cells were activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of 100 μM sitagliptin, or vehicle control for 4 days and changes in cellular bioenergetics were determined with a Seahorse assay. (B) Oxygen consumption rate (OCR) and (C) Maximal respiration (top) and reserve capacity (bottom) of sitagliptin- vs. vehicle-treated CD8 + T cells. n=6; *** p<0.001, **** p<0.0001 by t-test. (D) Mouse CD8 + T cells were activated for 2 days and stained with the mitochondrial membrane potential sensitive dye TMRM at 100 nM for 30 minutes. TMRM MFI of vehicle- vs sitagliptin-treated CD8 + T cells. n=5; **p<0.01 by paired t-test. (E) Proton efflux rate (PER) and (F) extracellular acidification rate (ECAR) of CD8 + T cells from B, C. n=6. (G) CD8 + T cells activated for two days were incubated with the glucose mimetic, 2-NBDG (50 μM) for 30 min. The percentage of T cells with 2-NBDG uptake was determined by flow cytometry. n=4; ** p<0.01 by paired t-test.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Control, Staining, Membrane, Incubation, Flow Cytometry

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Volcano plot of differentially regulated genes (1799 down and 762 up) upon sitagliptin treatment for 4 days. (B) Top differentially down-regulated pathways in sitagliptin-treated CD8 + T cells based on genes expressed >2-fold with p<0.05. n=6. (C) Splenic mouse CD8 + T cells were activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of 100 μM sitagliptin or vehicle control for 2 days and then stained with 20 nM of the mitochondrial permeable stain, Mitotracker green, for 30 minutes to quantify mitochondria mass. n=4; ns=not significant by paired t-test. (D) Splenic mouse CD8 + T cells were activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of 100 μM sitagliptin or vehicle control overnight. Cells were collected and lysed with RIPA buffer containing phosphatase/protease inhibitors, and 20 μM of protein were loaded on an SDS-Page for protein resolution, and western blotting analysis. (E) Splenic mouse CD8 + T cells were activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of 100 μM sitagliptin or vehicle control for 2 days, and then ATP levels were determined with CellTiter-Glo Luminescent Cell Viability Assay. n=4; ** p<0.01 by paired t-test. (F) CD8 + T cells activated for two days were incubated with the superoxide indicator, MitoSOX Deep Red, at 500 nM for 60 minutes. The percentage of T cells positive for superoxide accumulation was determined by flow cytometry. n=4; * p<0.05 by paired t-test. (G) CD8 + T cells from OT-I mice were isolated and stimulated with 10 ng OVA repetitively for 5 days to elicit exhaustion. On Day 6, cells were incubated with Mitotracker and TMRM. TMRM MFI was normalized to mitochondrial mass. n=4; * p<0.05 by paired t-test.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Control, Staining, SDS Page, Western Blot, Cell Viability Assay, Incubation, Flow Cytometry, Isolation

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Targeted metabolomics was performed with mouse CD8 + T cells activated for two days with 100 μM sitagliptin or vehicle control. Top enriched metabolic pathways were determined by >1.3-fold increase using Metaboanalyst, n=4. (B) T cell receptor (TCR) signaling-focused phosphoproteomics was performed with CD8 + T cells activated overnight. Top protein hits that were upregulated or phosphorylated >2-fold in sitagliptin- vs vehicle-treated CD8 + T cells. (C) Graphical abstract of how GAD1 feeds glutamate to the TCA cycle. (D) Representative histograms depicting CFSE dilution of CD8 + T cells expanded in the presence of sitagliptin, the GAD1 inhibitor, L-Allylglycine (100 μM), or combination treatment for 6 days. (E) Fold change in the frequency of proliferating CD8 + T cells normalized to background activation. n=4; * p<0.05, ** p<0.01, *** p<0.001 by one–way ANOVA. (F) OCR, and (G) reserve capacity of CD8 + T cells after 4 days of treatment with sitagliptin, L-Allylglycine (100 μM), or combination treatment, n=6; ** p<0.01, **** p<0.0001 by ordinary one–way ANOVA.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Control, Phospho-proteomics, Activation Assay

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: (A) Top differentially altered pathways in DPP4 high vs. DPP4 low human GBM-infiltrating CD8 + T cells. Pathways with an adjusted p-value < 0.05 were considered significantly enriched. (B) Phospho-proteomic array fluorescent signal from splenic CD8 + T cells pooled from 4 biological replicates and activated overnight with anti-CD3/CD28 plus 100 IU/mL IL-2 with 100 μM sitagliptin or vehicle control. (C) Differentially regulated pathways from (B) . (D) Targeted metabolomics was performed with mouse CD8 + T cells activated for two days with 100 μM sitagliptin or vehicle control. Metabolite quantification for GABA shunt pathway intermediates was determined. n=4; * p<0.05, ** p<0.01, **** p<0.0001, ns=not significant by paired t-test. (E) Human CD8 + T cells from healthy human donors were isolated form PBMCs and activated with anti-CD3/CD28 plus 100 IU/mL IL-2 in the presence of sitagliptin (100 μM), the GAD1 inhibitor, L-Allylglycine (100 μM), or combination treatment for 6 days. Cells were then collected, stained with trypan blue, and quantified using an automatic cell counter. n=4; * p<0.05, *** p<0.001 by mixed effect analysis. Human CD8 + T cells from healthy human donors were isolated from PBMCs and expanded with anti-CD3/CD28 plus 100 IU/mL IL-2 for 6 days to reduce donor variability. Cells were then further activated in the presence of sitagliptin (100 μM), L-Allylglycine (200 μM), or combination treatment for four days. Seahorse T-Cell Fitness Assay was performed to determine (F) OCR and (G) reserve capacity. n=6; * p<0.05 by Friedman test.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Control, Isolation, Staining

Journal: bioRxiv

Article Title: Pharmacologic DPP-4 inhibition promotes CD8⁺ T cell metabolic fitness to enhance anti-tumor activity

doi: 10.64898/2026.03.31.715681

Figure Lengend Snippet: Mouse CAR T-cells targeting B7-H3 were cultured with cancer cells expressing B7-H3 or empty vector at (A) 2:1 effector:target (E:T) ratio collected after 3-4 days, and co-cultured with fresh cancer cells until CAR T-cells stopped persisting in culture, or (B) at decreasing E:T ratios during the 3 rd stimulation to determine cancer cell cytotoxicity with MTS reagent after 72-hour co-culture. Heatmap demonstrating the killing capacity of either (C) sitagliptin- or (D) vehicle-treated B7-H3 CAR T-cells at reducing E:T ratio across different restimulations. (E) Normalized flux signal on day 35 post-intracranial injection n=4; * p<0.05 by t-test. (F) Diagram depicting the mechanism by which DPP-4 inhibition promotes metabolic/transcriptomic rewiring to enhance the effector function of CD8 + T cells.

Article Snippet: Mouse CD8 + T-cells were isolated from freshly harvested spleens of C57BL/6 mice using CD8a (Ly-2) MicroBeads (Miltenyi #130-114-044), following manufacturer’s instructions.

Techniques: Cell Culture, Expressing, Plasmid Preparation, Co-Culture Assay, Injection, Inhibition

Journal: Breast Cancer : Targets and Therapy

Article Title: The Correlation Between CD8+ Tumor-Infiltrating Lymphocytes and the Efficacy of Neoadjuvant Therapy in Breast Cancer

doi: 10.2147/BCTT.S533799

Figure Lengend Snippet: Immunohistochemistry slides of CD8+ TILs. ( A ) Baseline CD8+ TILs under ×200 magnification. Black arrows indicate iCD8+ TILs, and red arrows indicate sCD8+ TILs. Both intratumoral and stromal CD8+ TILs are diffusely distributed, with sCD8+ TILs being more abundant than iCD8+ TILs. ( B ) Baseline CD8+ TILs under ×400 magnification, showing positive staining of the cell membrane and cytoplasm (indicated by arrows).

Article Snippet: The CD8 rabbit anti-human polyclonal antibodies and secondary antibodies were sourced from Beijing Zhongshan Golden Bridge Biotechnology Co., Ltd., and the immunohistochemistry staining kit was procured from Fuzhou Maixin Biotechnology Development Co., Ltd. Formalin-fixed, paraffin-embedded tissue sections were prepared according to standard procedures, including sectioning, deparaffinization, and rehydration.

Techniques: Immunohistochemistry, Staining, Membrane