Search Results (216)

Vascular mimicry (VM) refers to the formation of vessel-like structures by tumor cells independent of endothelial cells. These VM channels connect to the host’s vascular network and are associated with aggressive tumors and poor patient prognosis. Most VM research has been conducted on melanoma, relying on patient-derived and mouse cell lines. In other solid tumors, VM studies rely on human cell lines, which have certain limitations for in vivo studies. Specifically, most in vivo VM research involving human cells uses subcutaneous mouse models that fail to recapitulate organ-specific tumor microenvironments. As the microenvironment is an essential driver of tumor vascularization, including VM, murine cell lines could facilitate VM investigations in syngeneic mouse models. Here, we present CT26 and KPC, well-characterized murine colorectal and pancreatic cancer cell lines, as cell models for VM investigations. Using in vitro cell-based assays, we demonstrate that CT26 and KPC undergo VM, a cell-intrinsic process that is enhanced by serum deprivation and exposure to hypoxia and is independent of tumor-secreted growth factors. Additionally, we demonstrate the importance of YAP/TAZ signaling in VM formation, as inhibition at non-cytotoxic concentrations attenuated VM formation. Remarkably, CA3, the most potent of the two inhibitors, significantly reduced cell proliferation in both cell lines at the IC₅₀ concentration. This reduction in cell proliferation was associated with the induction of apoptosis in CT26 cells and changes in the cell cycle in both CT26 and KPC cells. Finally, dual YAP/TAZ knockdown in both cell lines significantly abrogated VM formation, validating our initial findings using inhibitors. These results show that CT26 and KPC cells undergo VM, and given their extensive use in cancer research, can be used to investigate VM in vivo using syngeneic models. Full article

Background: Patients with systemic sclerosis have a significantly increased incidence of developing various solid malignancies within a few years of systemic sclerosis onset, but the mechanism of tumor promotion is not well understood. The tight skin (TSK) mouse has been a valuable model for investigating systemic sclerosis-related pathologies due to increased extracellular matrix deposition, fibrosis in connective tissues, and altered immune cell activation. Despite the role of extracellular matrix and fibrosis in cancer progression, the potential of the TSK mouse as a model for cancer studies is unexplored. Methods: To investigate the impact of the altered microenvironment in TSK mice on cancer progression, we compared the tumor-forming capabilities (by subcutaneous and intraperitoneal injection) in TSK mice and WT mice using syngeneic breast cancer, melanoma, and ovarian cancer cell lines. We used bulk and single-cell RNA sequencing to characterize these tumors and identify the changes in the TSK microenvironment that promote cancer formation. Results: In all three cancer types, TSK mice exhibited more invasive subcutaneous tumors in comparison to WT controls, underscoring the role of the TSK subcutaneous microenvironment in promoting cancer progression. Furthermore, the heightened invasiveness of ovarian tumors implanted intraperitoneally suggests that the peritoneal microenvironment in TSK mice also promotes tumor progression. Single-cell RNA sequencing analyses of subcutaneous tumors from TSK and WT mice revealed tumor-specific changes in the composition and phenotype of various cell populations. The most consistent alteration in TSK mice included a higher neutrophil-to-lymphocyte ratio and an enrichment in profibrotic subpopulations of myofibroblasts and macrophages. Conclusions: Our research unveils the TSK mouse as a valuable model for studying the intricate connections between systemic sclerosis and cancer Full article

Pancreatic ductal adenocarcinoma (PDA) exhibits diverse molecular aberrancies that contribute to its aggressive behaviour and poor patient survival. P-21-activated kinase 1 (PAK1) and PAK4 drive the tumorigenesis of PDA. However, their roles in tumour vasculature and the impact on immune response are unclear. This study aims to investigate the effects of PAK1 and PAK4 on tumour vasculature, immune cell infiltration, and the connection between using PAK1-knockout (KO), PAK4 KO, and wild-type (WT) PDA cells in cell-based and mouse experiments. Tumour tissues isolated from a syngeneic mouse model were immuno-stained to determine the changes in tumour vasculature and immune cell infiltration/activation, followed by a proteomic study to assess biological processes involved. PAK1KO or PAK4KO suppressed tumour growth by reducing angiogenesis while enhancing vascular normalisation, enhanced the infiltration/activation of T-cells and dendritic cells associated with upregulation of ICAM-1 and VCAM-1 in the tumour microenvironment, and stimulated vascular immune crosstalk via an ICAM-1-mediated mechanism. This was supported by proteomic profiles indicating the regulation of endothelial cell and leukocyte trans-endothelial migration in PAK1- or PAK4-knockout tumours. In conclusion, PAK1KO or PAK4KO enhanced tumour vascular normalisation while reducing angiogenesis, stimulating immune cell infiltration and activation to suppress tumour growth. Full article

Background/Objectives: Head and neck squamous cell carcinoma (HNSCC) is the seventh most prevalent cancer worldwide. Despite intensive treatments, the prognosis is unfavorable. Recently, immunotherapy has emerged as a novel therapeutic strategy, and several immune-checkpoint blockade blockers provide clinical benefits to patients. However, the response rates of these antibodies are limited, and there is a pressing need to increase the efficacy of immunotherapy for HNSCC patients. Epigenetic treatment is emerging as a promising combination approach able to change immune-related gene signatures in tumors and potentially increase the efficacy of immunotherapy. In this study, we sought to elucidate further immune-related gene signatures altered through epigenetic treatment and explored whether epigenetic drugs can increase the efficacy of anti PD-L1 treatment in HNSCC. Methods: At first, we treated six HNSCC cell lines with 5-azacytidine and romidepsin and analyzed gene expression patterns by microarray and TaqMan arrays analysis. We then explored the therapeutic efficacy of epigenetic treatment with an anti PD-L1 antibody in a syngeneic mouse model. Results: Our microarray analysis revealed the differential expression of immune-related genes in cell lines treated with epigenetic drugs, as compared to untreated controls. Most importantly, these array analyses showed a significant change in the transcription of some immune related-and biologically relevant genes, such as HLA-DRA, HMOX1, IFI6, IL12A, IRF7, NFKB2, RPL3L, STAT1, STAT3, CSF1, CSF2, FAS, OASL, and PD-L1, after epigenetic treatment. Furthermore, the combination of epigenetic treatment with an anti PD-L1 antibody significantly suppressed tumor growth in a syngeneic mouse model. In vivo tumors treated with epigenetic drugs expressed higher STAT1, STAT3, and PD-L1 compared to untreated tumors. Increased PD-L1 expression is postulated to increase the efficacy of anti PD-L1 treatment. Conclusions: Our results highlight the importance of a combinational strategy employing both epigenetic and immunotherapy in HNSCC. Full article

Background/Objectives: Subcutaneous tumor models are widely used in colorectal cancer (CRC) research due to their experimental accessibility; however, the spatial organization and regulatory mechanisms of their tumor microenvironment remain poorly understood. Methods: Here, we applied spatial transcriptomics to systematically characterize spatial heterogeneity within MC38 subcutaneous tumors in a syngeneic mouse model. Results: We identified two spatially distinct tumor zones, partitioned by cancer-associated fibroblasts (CAFs), that differ markedly in cellular composition, oncogenic signaling, immune infiltration, and metabolic states. One zone exhibited features of TGF-β-driven extracellular matrix remodeling, immune exclusion, and hyperproliferative metabolism, while the other was enriched for immunosuppressive macrophages, metabolic reprogramming via PPAR and AMPK pathways, and high-risk cell populations. Spatially resolved cell–cell communication networks further revealed zone-specific ligand–receptor interactions—such as ANGPTL4–SDC2 and PROS1–AXL—that underpin stromal remodeling and immune evasion and are associated with patient prognosis. Conclusions: Collectively, our study uncovers how region-specific cellular ecosystems and intercellular crosstalk establish prognostically divergent niches within subcutaneous CRC tumors, offering insights into spatially guided therapeutic strategies. Full article

This study aimed to clarify the alterations in gene expression in metastatic renal cell carcinoma (mRCC) during disease progression and in response to treatment with immune checkpoint inhibitors using a syngeneic mouse mRCC model. RENCA cells were orthotopically implanted in BALB/c mice. Mice received first-line treatment with cabozantinib, anti-PD-1 antibody, or a combination. Tumor progression was monitored using serial micro-computed tomography. Lung metastasis samples were collected, and RNA sequencing was performed. Mice with apparent disease progression received second-line treatment with axitinib, everolimus, or lenvatinib after combination therapy. The median overall survival was 28, 34, 34, and 49 days in untreated mice and those treated with cabozantinib, anti-PD-1, or their combination, respectively (p < 0.05). RNA sequencing revealed upregulation of the fibroblast growth factor pathway in lung metastases after monotherapy, whereas mTOR pathway activation was observed only after combination therapy. Treatment-specific gene expression changes occur in mRCC, suggesting that the optimal target for sequential therapy in mRCC varies depending on prior treatment. Full article

Immunotherapy targeting the immune functions of the tumor microenvironment (TME) is beneficial for colorectal cancer; however, the response rate is poor. Ciprofloxacin is a fluoroquinolone-class antibiotic that is used to treat bacterial infections. The purpose of this study is to assess the mechanism of ciprofloxacin that enhances anti-PD1 in colorectal cancer. We found that ciprofloxacin induced cytosolic DNA, including single-stranded and double-stranded DNA, formation in mouse CT26 colorectal adenocarcinoma cells. Molecules in DNA-sensing signaling such as cGAS, STING, and IFNβ mRNA and protein expression were elicited after ciprofloxacin treatment in CT26 cells. STING siRNA abrogated the cGAS-STING pathway activation by ciprofloxacin. In vivo, ciprofloxacin exhibited a synergistic effect with anti-PD1 to suppress tumor growth in a CT26 syngeneic animal model without biological toxicity. The examination of TME revealed that ciprofloxacin, alone and in combination therapy, induced M1 and red pulp macrophage production in the spleen. In tumors, M1 and M2 macrophage levels were increased by ciprofloxacin, and CD8⁺ T cell granzyme B expression was increased after combination therapy. STING showed the highest expression in tumor specimens after combination treatment. Ciprofloxacin may enhance the anti-PD1 efficacy and modulate the TME through the cGAS-STING pathway. Full article

Background/Objectives: In this study, a mouse model of sutured endothelial keratoplasty was established and compared with a traditional penetrating keratoplasty (PKP) model in both syngeneic (BALB/c) and allogeneic (C57/BL6) patterns. Methods: For the endothelial keratoplasty (EK) model, chimeric donor tissues consisting of BALB/c epithelium-stroma combined with either syngeneic (BALB/c) or allogeneic (C57/BL6) stroma-endothelium were transplanted into BALB/c mice. Graft transparency, gene expression, and mRNA levels in the transplanted tissues were assessed using polymerase chain reaction (PCR) and quantitative real-time reverse transcription PCR (qRT-PCR) to evaluate inflammatory status. Results: Allogeneic PKP had a higher opacity score than syngeneic PKP. In contrast, syngeneic EK mice had similar opacity scores to those of allogeneic EK mice. Upregulation of CXCR3, the receptor for CXCL10, was demonstrated by qRT-PCR in allogeneic PKP mice but not in allogeneic EK mice. Conclusions: Comparison between the syngeneic and allogeneic PKP groups revealed differences in CXCR3 mRNA expression, suggesting that CXCR3 could be a potential biomarker for rejection in the PKP mouse model. Additionally, the EK model did not show CXCR3 upregulation despite the opaque cornea due to nonspecific inflammation. Therefore, our mouse model was considered to be a successfully established EK model. Full article

The B Cell Lymphoma-2 (BCL-2) family proteins are central regulators of apoptosis, and their dysregulation is frequently associated with cancer progression and resistance to therapy. While small molecules like venetoclax have shown promise, nucleic acid-based therapeutics targeting BCL-2 remain underexplored. Here, we report a novel in vitro evolution strategy to generate trans-acting RNA-cleaving DNAzymes targeting natural BCL-2 mRNA without requiring covalent substrate-linking. Using a 50-base region of BCL-2 mRNA as a selection target, we evolved several DNAzymes that demonstrate significant RNA cleavage activity. These DNAzymes downregulated BCL-2 expression, induced apoptosis, and reduced cell viability in HepG2 and MCF-7 cancer cells. In vivo, our novel DNAzymes significantly suppressed tumor growth in a syngeneic mouse breast cancer model, with efficacy comparable to 5-Fluorouracil. This study presents a proof of concept for a novel strategy to evolve functional DNAzymes against native mRNA sequences and highlights their potential as gene-silencing tools in cancer therapy. Future studies will explore the therapeutic potential of these findings in cancer patients. Additionally, investigating the underlying molecular mechanisms in more complex cancer models will further validate the observed effects. Full article

Background/Objectives: Reprogramming of the cellular metabolism is a hallmark of cancer, offering therapeutic opportunities to target cancer cell vulnerabilities for therapeutic purposes. 3-Bromopyruvate (3BP) is a small alkylating agent that functions as an anti-metabolite, targeting key substrates in cancer metabolism and demonstrating antitumor activity across multiple cancer types. However, unformulated 3BP is associated with significant toxicity. This study investigates the efficacy of KAT/3BP, a clinical derivative of 3BP currently in phase 1 trials for hepatocellular carcinoma, in preclinical lymphoma models. Results: In vitro, KAT/3BP exhibited cytotoxic activity across 12 lymphoma cell lines—including diffuse large B-cell lymphoma and mantle cell lymphoma—with a median IC₅₀ of 3.7 μM. It also remained effective against lymphoma cell lines with acquired resistance to FDA-approved therapies. In vivo, treatment with KAT/3BP led to reduced tumor size in a syngeneic mouse model, with the combination of oral and intratumoral administration showing the greatest efficacy. Furthermore, KAT/3BP demonstrated synergistic activity when combined with standard lymphoma therapies such as bendamustine and R-CHOP. Conclusions: Our findings highlight the potential of KAT/3BP as a novel therapeutic option, either as a single agent or in combination regimens, for treating lymphomas. Full article

Background: Breast cancer (BC) is the second most common cancer among women in the United States. It has been estimated that one in eight women will be diagnosed with breast cancer in her lifetime. Various BC risk factors, such as age, physical inactivity, and smoking, play a substantial role in BC occurrence and development. Early life dietary intervention with plant-based bioactive compounds has been studied for its potential role in BC prevention. Sulforaphane (SFN), an isothiocyanate, is an antioxidant and anti-inflammatory agent extracted from broccoli sprouts (BSp) and other plants. Dietary supplementation of SFN suppresses tumor growth by inducing protective epigenetic changes and inhibiting cancer cell proliferation. Inulin, as a dietary fiber, has been studied for alleviating GI discomfort and weight loss by promoting the growth of beneficial bacteria in the gut. Objective: Early-life combinatorial treatment with both phytochemical SFN and potential prebiotic agent inulin at lower and safer dosages may confer more efficacious and beneficial effects in BC prevention. Methods: Transgenic mice representing estrogen receptor-negative BC were fed 26% (w/w) BSp and 2% (w/v) inulin supplemented in food and water, respectively. Results: The combinatorial treatment inhibited tumor growth, increased tumor onset latency, and synergistically reduced tumor weight. Gut microbial composition was analyzed between groups, where Ruminococcus, Muribaculaceae, and Faecalibaculum significantly increased, while Blautia, Turicibacter, and Clostridium sensu stricto 1 significantly decreased in the combinatorial group compared with the control group. Furthermore, combinatorial treatment induced a protective epigenetic effect by inhibiting histone deacetylases (HDACs) and DNA methyltransferases (DNMTs). Intermediates in the AKT/PI3K/MTOR pathway were significantly suppressed by the combinatorial treatment, including PI3K p85, p-AKT, p-PI3K p55, MTOR, and NF-κB. Cell cycle arrest and programmed cell death were induced by the combinatorial treatment via elevating the expression of cleaved-caspase 3 and 7 and inhibiting the expressions of CDK2 and CDK4, respectively. Orally administering F. rodentium attenuated tumor growth and induced apoptosis in a syngeneic triple-negative breast cancer (TNBC) mouse model. Conclusions: Overall, the findings suggest that early-life dietary combinatorial treatment contributed to BC prevention and may be a potential epigenetic therapy that serves as an adjunct to other traditional neoadjuvant therapies. Full article

Background/Objectives: Cell-based therapies have become increasingly important in the treatment of cancers and inflammatory diseases; however, therapies utilizing monocyte–macrophage lineage cells remain relatively underexplored. Non-invasive cell tracking allows a better understanding of the fate of such cells, which is essential for leveraging their therapeutic potential. Here, we employed a Zirconium-89 (⁸⁹Zr)-oxine cell labeling method to compare the trafficking of monocytes and macrophages in vivo. Methods: Mouse bone marrow-derived monocytes and macrophages were each labeled with ⁸⁹Zr-oxine and evaluated for their viability, radioactivity retention, chemotaxis, and phagocytic function in vitro. Labeled cells were intravenously administered to healthy mice and to murine models of granuloma and syngeneic tumors. Cell migration was monitored using microPET/CT, while cell recruitment to the lesions was further assessed via ex vivo biodistribution and flow cytometry. Results: Labeled cells exhibited similar survival and proliferation to unlabeled cells for up to 7 days in culture. While both maintained phagocytic function, monocytes showed higher CCL2-driven chemotaxis compared to macrophages. ⁸⁹Zr-oxine PET revealed initial cell accumulation in the lungs, followed by their homing to the liver and spleen within 2–24 h, persisting through the 5-day observation period. Notably, monocytes trafficked to the liver and spleen more rapidly than macrophages. In both inflammation and cancer models, monocytes demonstrated higher accumulation at the lesion sites compared to macrophages. Conclusions: This study demonstrates the usefulness of ⁸⁹Zr-oxine PET in tracking monocyte–macrophage lineage cells, highlighting their distinct migration patterns and providing insights that could advance monocyte-centered cell therapies. Full article

Cancer/testis antigen (CTA) gene products are expressed in most malignant tumours, while under normal conditions their expression is primarily restricted to testicular cells. In this study, we investigated the prophylactic application of a xenogeneic (ram-derived) testicular cell (TC) vaccine for cancer prevention in an experimental animal model. C57BL/6 mice were immunised three times with either xenogeneic (ram) or syngeneic (mouse) formaldehyde-fixed spermatogenic tissue-derived cells. Following vaccination, mice were implanted with live B16 melanoma or LLC carcinoma cells. Tumour-bearing mice were subsequently assessed for survival and immunological parameters indicative of anti-cancer immunity. Xenogeneic vaccination with TCs induced cross-reactive immune responses to both B16 melanoma and LLC carcinoma antigens (Ags), as determined by an MTT ((3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Prophylactic vaccination with xenogeneic TCs (xTCs), but not syngeneic TCs (sTCs), significantly improved survival rates, with 30% of vaccinated mice surviving after LLC carcinoma implantation. The induced immunity was long-lasting as mice implanted with LLC carcinoma cells 3–6 months post-vaccination exhibited prolonged survival. Furthermore, lymphoid cells from surviving vaccinated mice were capable of adoptively transferring anti-cancer immunity to naïve animals, significantly increasing their survival rates upon subsequent LLC carcinoma cell implantation. Vaccinated mice bearing LLC tumours exhibited a reduction in regulatory CD4⁺CD25⁺Foxp3⁺ T cells in the spleen, with no effect observed in the central memory CD4⁺CD44⁺CD62L⁺ T-cell compartment. Moreover, vaccinated mice displayed increased interferon gamma (IFN-γ) levels in the blood, with no significant changes in interleukin-10 (IL-10) levels. Prophylactic vaccination with xenogeneic CTAs effectively induces long-term, stable anti-cancer immunity, demonstrating potential for future immunopreventive strategies. Full article

Background/Objectives: Prostate cancer (PCa) cells may acquire radioresistance during radiation therapy (RT), resulting in PCa recurrence. This study was aimed at investigating the radiosensitizing effect of 5-aminolevulinic acid (5-ALA) on radioresistant PCa cells. Methods: Radioresistant PCa cells were developed through successive irradiation of two human PCa cell lines (PC-3 and DU 145) and a murine PCa cell line (Myc-CaP). The radiosensitivity of these PCa cells and the radiosensitizing effect of 5-ALA were evaluated using clonogenic assays. Mitochondrial accumulation of protoporphyrin IX (PpIX) and mitochondrial reactive oxygen species (ROS) were evaluated. A syngeneic mouse model with radioresistant PCa was established, and the immunohistochemistry of cell specimens from PCa patients with local recurrence after primary RT was examined. Results: Radioresistant PCa cells showed lower radiosensitivity compared to parental PCa cells. In radioresistant PCa cells with 5-ALA administration, compared to the group administered irradiation alone, the survival rate after irradiation was significantly reduced by promoting mitochondria-mediated apoptosis caused by increased PpIX accumulation and mitochondrial ROS generation. Similar results were observed in vivo. However, compared with parental PCa cells, radioresistant PCa cells were less affected by the radiosensitizing effect of 5-ALA, owing to decreased PpIX accumulation and mitochondrial ROS production caused by upregulated expression of the drug transporter ABCG2. ABCG2 expression was upregulated in human PCa specimens with post-RT recurrence. Conclusions: 5-ALA enhanced the antitumor effects of RT in radioresistant PCa cells; however, ABCG2 upregulation decreased PpIX accumulation, resulting in a reduced radiosensitizing effect of 5-ALA on radioresistant PCa cells compared with that on parental PCa cells. ABCG2 could be a potential therapeutic target for overcoming radioresistance. Full article

Objectives: Pancreatic cancer remains a therapeutic challenge due to its immunosuppressive microenvironment and treatment resistance. This study aimed to develop a novel recombinant oncolytic vaccinia virus (VVL-GL7) co-expressing granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-7 (IL-7), designed to enhance anti-tumor immunity and synergize with immune checkpoint inhibitors. Methods: VVL-GL7 was constructed through CRISPR/Cas9-mediated knockout of TK and A49 genes, combined with the simultaneous insertion of dual cytokine-encoding cassettes. Anti-tumor efficacy was evaluated in vitro and in vivo using C57BL/6 mouse and Syrian hamster pancreatic cancer models. Comprehensive immune profiling evaluated CD8⁺ T-cell and macrophage infiltration dynamics while simultaneously assessing memory T-cell differentiation patterns using flow cytometry. Preclinical combination studies of VVL-GL7 and the PD-1 immune checkpoint inhibitor were systematically evaluated in a syngeneic pancreatic cancer model. Results: VVL-GL7 exhibited potent oncolytic activity, inducing significant tumor regression in both preclinical models. VVL-GL7 therapy significantly augmented CD8⁺ T-cell and macrophage infiltration within the tumor microenvironment, while concomitantly driving memory T-cell differentiation. The synergistic effects of VVL-GL7 and the PD-1 blockade further improved therapeutic outcomes, resulting in significantly higher tumor remission rates compared to monotherapy and achieving complete tumor regression in pancreatic cancer models. Conclusions: VVL-GL7 reprograms the immunosuppressive tumor microenvironment and synergizes with anti-PD-1 antibodies to overcome resistance in pancreatic cancer. Full article