Journal: bioRxiv

Article Title: Lung-targeted cytokine-coding RNA-lipoplexes induce T and NK cell-mediated anti-tumor immune response

doi: 10.64898/2026.05.06.723126

Figure Lengend Snippet: ( A-D ) Luc–encoding RNA formulated as RNA-LPX was administered i.v. to non-tumor bearing (A, B; n=3) or CT26 metastases-bearing (C, D) BALB/c mice (n=3-9). ( A ) BLI of total body, ( B ) normalized organ signal from explanted organs. Data were analyzed by ordinary one-way ANOVA and Tukey’s test for multiple comparisons. ( C ) Kinetics of BLI of the lung signal in metastases-bearing mice, significance was determined by mixed-effects analysis with Geisser-Greenhouse correction and Tukey’s test for multiple comparisons. ( D ) Luc RNA, luc protein and CD31 (PECAM-1) were detected via RNAscope and immunohistochemistry on consecutive sections 1, 6 or 24 hours post injection, scale bar: 100 µm, tumor tissue is indicated with dashed line. ( E ) Cytokine quantification in lung after the indicated time points. ( F ) Cytokine fold increase in lung normalized to spleen at 6 hours after injection. ( G ) FDG positron emission tomography (PET) imaging 24 h after cytokine RNA mix injection into naïve mice, ( H ) ex vivo measurement of FDG uptake. Significance was determined using a two-tailed t-test for unpaired samples.

Article Snippet: Colon Carcinoma cell line CT26 was purchased from ATCC (CT26: CRL-2638, lot no. 58494154, female).

Techniques: RNAscope, Immunohistochemistry, Injection, Positron Emission Tomography, Imaging, Ex Vivo, Two Tailed Test

Journal: bioRxiv

Article Title: Lung-targeted cytokine-coding RNA-lipoplexes induce T and NK cell-mediated anti-tumor immune response

doi: 10.64898/2026.05.06.723126

Figure Lengend Snippet: ( A ) Experimental design: BALB/c mice (n=15 per group) were injected i.v. with CT26 tumor cells; subsequently, cytokine RNA mix or irrelevant RNA was administered i.v. as RNA-LPX twice per week for a total of 8 injections from d3 to d27. Survivor mice from the cytokine RNA mix-treated group (n=5) compared to naïve BALB/c mice (n=10) were re-challenged with CT26 tumor cells. ( B ) Survival after the initial i.v. CT26 tumor cell inoculation. ( C ) Survival after re-challenge. Significance was determined via Mantel-Cox logrank. ( D-F ) BALB/c mice (n=5 per group) were injected i.v. with CT26 tumor cells, RNA-LPX was administered i.v. at d3, d6 and d10 p.t.i., mice were sacrificed at d11, lungs were collected and analyzed via flow cytometry. Significance for pairwise comparisons was determined by unpaired two-tailed t-test. Flow cytometric analysis of ( D ) tumor burden, ( E ) CD8 + T and NK cells and ( F ) CD4 + Foxp3 + CD25 + T reg . ( G ) Functional analysis: after tumor cell inoculation, mice were treated with RNA-LPX at d3, d6, d10 and d13 p.t.i., mice were sacrificed at d14 for an ex vivo stimulation assay with PMA/Ionomycin ( H ) qRT-PCR was done to determine fold-change expression of cytokine RNA mix-treated normalized to irrelevant RNA LPX-treated lung samples (n=5 per group indicated in rows) for the indicated cytokines and chemokines.

Article Snippet: Colon Carcinoma cell line CT26 was purchased from ATCC (CT26: CRL-2638, lot no. 58494154, female).

Techniques: Injection, Flow Cytometry, Two Tailed Test, Functional Assay, Ex Vivo, Quantitative RT-PCR, Expressing

Journal: bioRxiv

Article Title: Lung-targeted cytokine-coding RNA-lipoplexes induce T and NK cell-mediated anti-tumor immune response

doi: 10.64898/2026.05.06.723126

Figure Lengend Snippet: Experimental design: BALB/c mice (n=5 per group) were injected i.v. with CT26 tumor cells; cytokine RNA mix or irrelevant RNA was administered i.v. at d3, d6 and d10 post tumor cell injection, mice were sacrificed at d11, lungs were collected and pooled at same ratios before sorting of CD45 + cells. CD45 + cells were subjected to scRNAseq. ( A,B ) Uniform manifold approximation and projection (UMAP) of 22 assigned clusters ( A ) and cell type frequencies ( B ) of CD45 + cells isolated from the lungs of cytokine RNA mix-treated versus irrelevant RNA-treated mice. ( C ) Effector function visualized as bubble plot. ( D ) Selected differentially expressed genes in cytokine RNA mix-treated versus irrelevant RNA-treated samples are shown as average log2 fold change. Note only significantly expressed values (p ≤ 0.05) are shown in colouring (blue = downregulated, red = upregulated), non-significant values are set to “0”/white.

Article Snippet: Colon Carcinoma cell line CT26 was purchased from ATCC (CT26: CRL-2638, lot no. 58494154, female).

Techniques: Injection, Isolation

Journal: bioRxiv

Article Title: Lung-targeted cytokine-coding RNA-lipoplexes induce T and NK cell-mediated anti-tumor immune response

doi: 10.64898/2026.05.06.723126

Figure Lengend Snippet: ( A ) Experimental design for ( B-C; CT26 tumor model) and ( E; CT26 B2M k.o. tumor model); n=15 BALB/c mice per group. Depletion or blocking antibody treatment was started 2 days prior to RNA-LPX treatment to ensure depletion before treatment start. ( B, C ) Survival according to termination criteria. ( D ) Experimental design and survival in CT26B2M k.o. or CT26gp70 k.o. tumor cells i.v. tumor model, n=15 mice per group. ( E ) BALB/c mice injected i.v. with CT26B2M k.o and depletion/neutralization antibodies were applied as shown in ( A ). Note that data in ( E ) were generated within the same experiment, irrelevant RNA + isotype mix as well as cytokine mix RNA + isotype mix refers to the same groups in all 3 plots. Survival was analyzed via Mantel-Cox logrank test. For ( B ) and ( C ), groups 1 and 2 were furthermore compared via logrank test with emphasis on early and late differences, ( B ) ## Logrank test with emphasis on late differences ((rho=0, lambda=1): group 1 vs 2: p=0,00235; ( C ) # Logrank test with emphasis on early differences (rho=1, lambda=0): group 1 vs 2: p=0,0378.

Article Snippet: Colon Carcinoma cell line CT26 was purchased from ATCC (CT26: CRL-2638, lot no. 58494154, female).

Techniques: Blocking Assay, Injection, Neutralization, Generated

Journal: Frontiers in Immunology

Article Title: STAT6 inhibition of M2 macrophages suppresses tumor growth by modulating the tumor microenvironment in colon cancer model

doi: 10.3389/fimmu.2026.1733991

Figure Lengend Snippet: TAMpep-IP suppresses tumor growth in the colon cancer model. (A) BALB/c mice were subcutaneously inoculated with CT26 colon carcinoma cells (3 × 10 5 cells per mouse). Starting on day 7 post-inoculation, TAMpep-IP (400 nmol/kg) was administered subcutaneously every three days for a total of seven doses. (B) Representative images of tumors excised at the experimental endpoint (day 25) showed visibly reduced tumor size in the TAMpep-IP–treated group compared to control. (C) Tumor volumes were measured every 3 days following tumor implantation. Mice treated with TAMpep-IP exhibited significantly reduced tumor growth relative to the control group (control: n = 6; TAMpep-IP: n = 6). (D) Tumor proliferation was evaluated by immunohistochemical staining of Ki-67 in tumor sections. Quantitative analysis showed a significantly lower proportion of Ki-67 + proliferating cells in TAMpep-IP–treated tumors. Representative immunohistochemistry images were acquired at ×100 magnification. Scale bar = 1000 μm. All data are presented as mean ± SEM. *p<0.05, ***p<0.001.

Article Snippet: The murine colon carcinoma cell line CT26 (KCLB 80009; Korean Cell Line Bank, Seoul, Korea) was cultured in Dulbecco’s modified Eagle’s medium (DMEM) (D5671; Welgene, Gyeongsangbuk, Korea) supplemented with 10% heat-inactivated FBS, 100 U/mL penicillin, and 1% penicillin–streptomycin.

Techniques: Control, Tumor Implantation, Immunohistochemical staining, Staining, Immunohistochemistry

Journal: Frontiers in Immunology

Article Title: STAT6 inhibition of M2 macrophages suppresses tumor growth by modulating the tumor microenvironment in colon cancer model

doi: 10.3389/fimmu.2026.1733991

Figure Lengend Snippet: TAMpep-IP reduces M2 macrophages in tumor tissues of colon cancer model. (A) Tumor-infiltrating immune cells were isolated from CT26 tumors in control and TAMpep-IP–treated mice. Flow cytometry was used to identify CD206 + F4/80 + macrophages within the CD45 + CD11b + population. TAMpep-IP significantly decreased the proportion of M2-like tumor-associated macrophages. (B) Quantitative RT-PCR analysis of tumor tissues revealed that TGF-β mRNA expression, a key M2-associated cytokine, was significantly reduced in TAMpep-IP–treated tumors compared to controls. (C) Western blot analysis of tumor showed a marked decrease in CD206 protein levels following TAMpep-IP, indicating effective suppression of M2 macrophage markers. (D) CD206 + macrophages were further visualized by immunohistochemical staining of tumor sections. ImageJ-based quantification confirmed a significant reduction in CD206 + area in TAMpep-IP–treated tumors. Representative immunohistochemistry images were acquired at ×100 magnification. Scale bar = 1000 μm. All data are presented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001.

Article Snippet: The murine colon carcinoma cell line CT26 (KCLB 80009; Korean Cell Line Bank, Seoul, Korea) was cultured in Dulbecco’s modified Eagle’s medium (DMEM) (D5671; Welgene, Gyeongsangbuk, Korea) supplemented with 10% heat-inactivated FBS, 100 U/mL penicillin, and 1% penicillin–streptomycin.

Techniques: Isolation, Control, Flow Cytometry, Quantitative RT-PCR, Expressing, Western Blot, Immunohistochemical staining, Staining, Immunohistochemistry

Journal: Frontiers in Immunology

Article Title: STAT6 inhibition of M2 macrophages suppresses tumor growth by modulating the tumor microenvironment in colon cancer model

doi: 10.3389/fimmu.2026.1733991

Figure Lengend Snippet: TAMpep-IP enhances inflammatory cytokine expression and activated CD8 + T cells in tumor tissues of colon cancer model. (A) Flow cytometry was used to evaluate CD8 + T cell function in CT26 tumor tissues from control and TAMpep-IP-treated mice. TAMpep-IP significantly increased the proportion of activated Granzyme B + CD8 + T cells, while reducing the frequency of exhausted Tim-3 + CD8 + T cells, indicating enhanced cytotoxic T cell activity. (B, C) Confocal immunofluorescence analysis was performed on tumor sections stained with DAPI (nuclei), anti-CD8 (green), anti-Granzyme B (red), and anti-PD-1 (red). Activated CD8 + T cells were identified by co-localization of CD8 and Granzyme B, whereas exhausted CD8 + T cells were identified by co-localization of CD8 and PD-1. Quantification revealed a significant increase in intertumoral CD8 + Granzyme B + T cells and a concomitant decrease in CD8 + PD-1 + exhausted T cells following TAMpep-IP. Representative confocal images were acquired using a 40× objective lens. Scale bar = 20 μm. (D) Quantitative RT-PCR analysis of CT26 tumor tissues showed significantly elevated mRNA levels of pro-inflammatory cytokines TNF-α, IL-1β, and IL-12 in TAMpep-IP–treated mice compared to controls, indicating induction of a pro-inflammatory tumor microenvironment. All data are presented as mean ± SEM. *p<0.05, **p<0.01, ***p<0.001, ****p < 0.0001.

Article Snippet: The murine colon carcinoma cell line CT26 (KCLB 80009; Korean Cell Line Bank, Seoul, Korea) was cultured in Dulbecco’s modified Eagle’s medium (DMEM) (D5671; Welgene, Gyeongsangbuk, Korea) supplemented with 10% heat-inactivated FBS, 100 U/mL penicillin, and 1% penicillin–streptomycin.

Techniques: Expressing, Flow Cytometry, Cell Function Assay, Control, Activity Assay, Immunofluorescence, Staining, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Breast cancer interactions with osteoclasts generate osteoclast-tumor hybrid-like cells through dynamic non-canonical cell fusion and cell-in-cell processes

doi: 10.64898/2026.04.05.716538

Figure Lengend Snippet: a,b, Fluorescent snapshot image for osteoclast-tumor hybrid-like cells formed between RAW264.7-GFP and other solid tumor cell lines. Hybrids containing tumor nuclei derived from (a) B16F10-H2B-mRFP melanoma and hybrids with tumor nuclei derived from (b) CT26-H2B-mRFP colon cancer.

Article Snippet: RAW264.7, 4T1, EO771, B16F10, and CT26 cell lines were obtained from ATCC.

Techniques: Derivative Assay