Mouse Model Of Colon Cancer Research Articles

Combinational delivery of TLR4 and TLR7/8 agonist enhanced the therapeutic efficacy of immune checkpoint inhibitors to colon tumor.

Immunotherapy is regarded as a potent cancer treatment, with DC vaccines playing a crucial role. Although clinical trials have demonstrated the safety and efficacy of DC vaccines, loading antigens in vitro is challenging, and their therapeutic effects remain unpredictable. Moreover, the diverse subtypes and maturity states of DCs in the body could induce both immune responses and immune tolerance, potentially affecting the vaccine's efficacy. Hence, the optimization of DC vaccines remains imperative. Our study discovered a new therapeutic strategy by using CT26 and MC38 mouse colon cancer models, as well as LLC mouse lung cancer models. The strategy involved the synergistic activation of DCs through intertumoral administration of TLR4 agonist high-mobility group nucleosome binding protein 1 (HMGN1) and TLR7/8 agonist (R848/resiquimod), combined with intraperitoneal administration of TNFR2 immunosuppressant antibody. The experimental results indicated that the combined use of HMGN1, R848, and α-TNFR2 had no effect on LLC cold tumors. However, it was effective in eradicating CT26 and MC38 colon cancer and inducing long-term immune memory. The combination of these three drugs altered the TME and promoted an increase in anti-tumor immune components. This may provide a promising new treatment strategy for colon cancer.

The Impact of Atmospheric Cadmium Exposure on Colon Cancer and the Invasiveness of Intestinal Stents in the Cancerous Colon.

Inhalation exposure to carcinogenic metals such as cadmium (Cd) is a significant global health concern linked to various cancers. However, the precise carcinogenic mechanism underlying inhalation exposure remains elusive. In this study, CT26 mouse colon cancer (CC) cells were implanted into BALB/c mice to establish CC mouse models. Some of the CC mice were implanted with intestinal stents. The mice were exposed to atomized oxygen and nitrogen (O2/N2) gas containing Cd. Atmospheric Cd intensified inflammation in CC cells and heightened Nicotinamide Adenine Dinucleotide Phosphate (NADPH) Oxidase 1 (NOX1) activity, which is an indirect measurement of increased reactive oxygen species (ROS) production. This escalated ROS production triggered abnormal Wnt protein secretion, activated the Wnt/β-catenin signaling pathway, and stimulated CC cell proliferation. No discernible body weight effect was seen in the CC mice, possibly due to the later-stage tumor weight gain, which masked the changes in body weight. Cd facilitated colon tumor restructuring and cell migration at the later stage. The implantation of intestinal stents inhibited the expression of Superoxide Dismutase 1 (SOD1) in the colon tumors of the CC mice, with no evident effects on the expression levels of NOX1, SOD2, and Catalase (CAT) enzymes. Elevated ROS levels, indirectly reflected by enzyme activity, did not substantially impact the Wnt/β-catenin signaling pathway and even contributed to slowing its imbalance. Stent implantation eased the inflammation occurring in colon tumors by reducing CC cell proliferation but it induced discomfort in the mice, leading to a reduction in food intake and weight. Cd partially fosters CC tumorigenesis via the ROS-mediated Wnt/β-catenin signaling pathway. The effect of Cd on the invasive effect of intestinal stents in the cancerous colon is not significant.

Onward Spread from Liver Metastases Is a Major Cause of Multi-Organ Metastasis in a Mouse Model of Metastatic Colon Cancer.

Colorectal cancer metastasizes predominantly to the liver but also to the lungs and the peritoneum. The presence of extra-hepatic metastases limits curative (surgical) treatment options and is associated with very poor survival. The mechanisms governing multi-organ metastasis formation are incompletely understood. Here, we tested the hypothesis that the site of tumor growth influences extra-hepatic metastasis formation. To this end, we implanted murine colon cancer organoids into the primary tumor site (i.e., the caecum) and into the primary metastasis site (i.e., the liver) in immunocompetent mice. The organoid-initiated liver tumors were significantly more efficient in seeding distant metastases compared to tumors of the same origin growing in the caecum (intra-hepatic: 51 vs. 40%, p = 0.001; peritoneal cavity: 51% vs. 33%, p = 0.001; lungs: 30% vs. 7%, p = 0.017). The enhanced metastatic capacity of the liver tumors was associated with the formation of 'hotspots' of vitronectin-positive blood vessels surrounded by macrophages. RNA sequencing analysis of clinical samples showed a high expression of vitronectin in liver metastases, along with signatures reflecting hypoxia, angiogenesis, coagulation, and macrophages. We conclude that 'onward spread' from liver metastases is facilitated by liver-specific microenvironmental signals that cause the formation of macrophage-associated vascular hotspots. The therapeutic targeting of these signals may help to contain the disease within the liver and prevent onward spread.

Small molecules that disrupt RAD54-BLM interaction hamper tumor proliferation in colon cancer chemoresistance models.

RAD54 and BLM helicase play pivotal roles during homologous recombination repair (HRR) to ensure genome maintenance. BLM amino acids (aa 181-212) interact with RAD54 and enhance its chromatin remodeling activity. Functionally, this interaction heightens HRR, leading to a decrease in residual DNA damage in colon cancer cells. This contributes to chemoresistance in colon cancer cells against cisplatin, camptothecin, and oxaliplatin, eventually promoting tumorigenesis in preclinical colon cancer mouse models. ChIP-Seq analysis and validation revealed increased BLM and RAD54 corecruitment on the MRP2 promoter in camptothecin-resistant colon cancer cells, leading to BLM-dependent enhancement of RAD54-mediated chromatin remodeling. We screened the Prestwick small-molecule library, with the intent to revert camptothecin- and oxaliplatin-induced chemoresistance by disrupting the RAD54-BLM interaction. Three FDA/European Medicines Agency-approved candidates were identified that could disrupt this interaction. These drugs bound to RAD54, altered its conformation, and abrogated RAD54-BLM-dependent chromatin remodeling on G5E4 and MRP2 arrays. Notably, the small molecules also reduced HRR efficiency in resistant lines, diminished anchorage-independent growth, and hampered the proliferation of tumors generated using camptothecin- and oxaliplatin-resistant colon cancer cells in both xenograft and syngeneic mouse models in BLM-dependent manner. Therefore, the 3 identified small molecules can serve as possible viable candidates for adjunct therapy in colon cancer treatment.

Magnetic resonance imaging-based approaches for detecting the efficacy of combining therapy following VEGFR-2 and PD-1 blockade in a colon cancer model

BackgroundAngiogenesis inhibitors have been identified to improve the efficacy of immunotherapy in recent studies. However, the delayed therapeutic effect of immunotherapy poses challenges in treatment planning. Therefore, this study aims to explore the potential of non-invasive imaging techniques, specifically intravoxel-incoherent-motion diffusion-weighted imaging (IVIM-DWI) and blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI), in detecting the anti-tumor response to the combination therapy involving immune checkpoint blockade therapy and anti-angiogenesis therapy in a tumor-bearing animal model.MethodsThe C57BL/6 mice were implanted with murine MC-38 cells to establish colon cancer xenograft model, and randomly divided into the control group, anti-PD-1 therapy group, and combination therapy group (VEGFR-2 inhibitor combined with anti-PD-1 antibody treatment). All mice were imaged before and, on the 3rd, 6th, 9th, and 12th day after administration, and pathological examinations were conducted at the same time points.ResultsThe combination therapy group effectively suppressed tumor growth, exhibiting a significantly higher tumor inhibition rate of 69.96% compared to the anti-PD-1 group (56.71%). The f value and D* value of IVIM-DWI exhibit advantages in reflecting tumor angiogenesis. The D* value showed the highest correlation with CD31 (r = 0.702, P = 0.001), and the f value demonstrated the closest correlation with vessel maturity (r = 0.693, P = 0.001). While the BOLD-MRI parameter, R2* value, shows the highest correlation with Hif-1α(r = 0.778, P < 0.001), indicating the capability of BOLD-MRI to evaluate tumor hypoxia. In addition, the D value of IVIM-DWI is closely related to tumor cell proliferation, apoptosis, and infiltration of lymphocytes. The D value was highly correlated with Ki-67 (r = − 0.792, P < 0.001), TUNEL (r = 0.910, P < 0.001) and CD8a (r = 0.918, P < 0.001).ConclusionsThe combination of VEGFR-2 inhibitors with PD-1 immunotherapy shows a synergistic anti-tumor effect on the mouse colon cancer model. IVIM-DWI and BOLD-MRI are expected to be used as non-invasive approaches to provide imaging-based evidence for tumor response detection and efficacy evaluation.

Anti-OX40 Antibody Combined with HBc VLPs Delays Tumor Growth in a Mouse Colon Cancer Model

ObjectiveCombination immunotherapy strategies targeting OX40, a co-stimulatory molecule that can enhance antitumor immunity by modulating the proliferation, differentiation, and effector function of tumor-infiltrating T cells, have attracted much attention for their excellent therapeutic effects. In this study, we aimed to evaluate the antitumor efficacy of combined anti-OX40 and hepatitis B core virus-like particles (HBc VLPs) therapy using a mouse colon cancer model. MethodsHumanized B-hOX40 mice were injected subcutaneously with MC38 colon tumor cells and treated with HBc VLPs+anti-hOX40 antibody. Tumor growth was monitored. Flow cytometric analysis was performed to evaluate the populations of T cell subsets in the tumors. ResultsThe combination of anti-OX40 with HBc VLPs resulted in a significant delay in tumor growth, suggesting that a potent antitumor immunity was induced by the combination therapy. Further studies revealed that HBc VLPs+anti-OX40 treatment induced a significant increase in effector T cells (Teffs) and a significant decrease in regulatory T cells (Tregs) in the tumor microenvironment (TME), which accounted for the synergistic antitumor effect of anti-OX40 in combination with HBc VLPs. ConclusionCombination therapy of anti-hOX40 and HBc VLPs provides synergistic antitumor activity in colon cancer-bearing mice, which may represent a potential design strategy for cancer immunotherapy.

Musashi-2 potentiates colorectal cancer immune infiltration by regulating the post-translational modifications of HMGB1 to promote DCs maturation and migration

Post-translational modifications (PTMs) of the non-histone protein high-mobility group protein B1 (HMGB1) are involved in modulating inflammation and immune responses. Recent studies have implicated that the RNA-binding protein (RBP) Musashi-2 (MSI2) regulates multiple critical biological metabolic and immunoregulatory functions. However, the precise role of MSI2 in regulating PTMs and tumor immunity in colorectal cancer (CRC) remains unclear. Here, we present data indicating that MSI2 potentiates CRC immunopathology in colitis-associated colon cancer (CAC) mouse models, cell lines and clinical specimens, specifically via HMGB1-mediated dendritic cell (DC) maturation and migration, further contributes to the infiltration of CD4+ and CD8+ T cells and inflammatory responses. Under stress conditions, MSI2 can exacerbate the production, nucleocytoplasmic transport and extracellular release of damage-associated molecular patterns (DAMPs)-HMGB1 in CRC cells. Mechanistically, MSI2 mainly enhances the disulfide HMGB1 production and protein translation via direct binding to nucleotides 1403–1409 in the HMGB1 3′ UTR, and interacts with the cytoplasmic acetyltransferase P300 to upregulate its expression, further promoting the acetylation of K29 residue in HMGB1, thus leading to K29-HMGB1 nucleocytoplasmic translocation and extracellular release. Furthermore, blocking HMGB1 activity with glycyrrhizic acid (Gly) attenuates MSI2-mediated immunopathology and immune infiltration in CRC in vitro and in vivo. Collectively, this study suggests that MSI2 may improve the prognosis of CRC patients by reprogramming the tumor immune microenvironment (TIME) through HMGB1-mediated PTMs, which might be a novel therapeutic option for CRC immunotherapy.

Inhibitory Effect of Ginsenoside Rg3 Combined With 5-Fluorouracil on Tumor Angiogenesis and Tumor Growth of Colon Cancer in Mice: An Experimental Study

To evaluate the inhibitory effect of ginsenoside Rg3 combined with 5-fluorouracil (5-FU) on tumor angiogenesis and tumor growth in colon cancer in mice. CT26 mouse model of colon cancer was established and the mice were randomly assigned to the control group, the ginsenoside Rg3 group, the 5-FU group, and the Rg3 combined with 5-FU group. The 5-FU group was injected intraperitoneally at the dose of 20 mg/kg, 0.2 mL/animal, and once a day for 10 days. Treatment for the Rg3 group was given at the dose of 20 mg/kg, 0.2 mL/animal, and once a day for 21 days via gastric gavage. The dose and the mode of treatment for the Rg3+5-FU combination group were the same as those for the 5-FU and the Rg3 group. The control group was intraperitoneally injected with 0.2 mL/d of normal saline for 10 days. The expression of vascular endothelial growth factor (VEGF) and CD31 and the microvascular density (MVD) of the tumor tissues were examined by immunohistochemistry. The blood flow signals and tumor necrosis were examined by color Doppler flow imaging (CDFI). The quality of life, survival rate, tumor volume, tumor mass, and tumor inhibition rate of the mice were monitored. After 21 days of treatment, the tumor volume and the tumor mass of all treatment groups were significantly decreased compared with those the control group, with the combination treatment group exhibiting the most significant decrease. The tumor inhibition rates of the Rg3 group, the 5-FU group, and the combination group were 29.96%, 68.78%, and 73.42%, respectively. Rg3 treatment alone had inhibitory effect on tumor growth to a certain degree, while 5-FU treatment alone or 5-FU combined with Rg3 had a stronger inhibitory effect on tumor growth. The tumor inhibition rate of the combination group was higher than that of the 5-FU group, but the difference was not statistically significant (P>0.05). Color Doppler ultrasound showed that there were multiple localized and large tumor necrotic areas that were obvious and observable in the Rg3 group and the combination group, and that there were only small tumor necrotic areas in the 5-FU group and the control group. The tumor necrosis rate of the combination group was (55.63±3.12)%, which was significantly higher than those of the other groups (P<0.05). CDFI examination of the blood flow inside of the tumor of the mice showed that the blood flow signals in the combination group were mostly grade 0-Ⅰ, and that the blood flow signals in the control group were the most abundant, being mostly grade Ⅱ-Ⅲ. The abundance of the blood flow signals in the Rg3 and 5-FU groups were between those of the control group and the combination group. Compared with those of the control group, the expression levels of MVD and VEGF in the tumor tissues of the Rg3 group, the 5-FU group, and the combination group were significantly decreased, with the combination group showing the most significant decrease (P<0.05). HE staining results indicated that there was significant tumor necrosis in mice in the control group and that there were more blood vessels. In contrast, in the tumor of the Rg3 group and the 5-FU group, there were fewer blood vessels and necrotic gaps appeared within the tumors. In the combination group, the tumor tissues had the fewest blood vessels and rope-like necrosis was observed. The mice started dying on the 18th day after treatment started, and all the mice in the control group died on the 42nd day. By this time, there were 3, 5, and 7 mice still alive in the Rg3 group, the 5-FU group, and the combination group, respectively, presenting a survival rate of 30%, 50%, and 70%, respectively. All mice in all the groups died on day 60 after treatment started. Ginsenoside Rg3 combined with 5-FU can significantly inhibit tumor angiogenesis and tumor growth of colon cancer in mice and improve the survival and quality of life of tumor-bearing mice.

A phase I clinical trial of riluzole in combination with mFOLFOX6 and bevacizumab in treating patients with metastatic colorectal cancer.

101 Background: Colorectal cancer (CRC) is the second leading cause of cancer-related mortality in the US. 5-fluorouracil (5-FU) based chemotherapies in combination with targeted agents remain the standard of care in patients with metastatic or locally advanced disease. It is essential to develop new treatment strategies in metastatic CRC patients with microsatellite stable disease, in whom immune checkpoint blockade is ineffective. Our preclinical studies showed riluzole, an oral medicine for amyotrophic lateral sclerosis, can increase intratumoral CD8+ T cells and suppress tumor growth in a syngeneic colon cancer mouse model. Riluzole-mediated tumor growth inhibition depends on the presence of CD8+ T cells and activation of cGAS/STING/IFNβ signaling. Methods: This was a single-arm phase I trial of riluzole in combination with mFOLFOX6/bevacizumab for patients with metastatic CRC. The dose of riluzole started from 50 mg twice daily with dose escalation to 100 mg twice daily or dose de-escalation to 50 mg once daily. Patients received riluzole for 16 weeks in combination with mFOLOFX6/bevacizumab for 8 cycles. Then patients either continued mFOLFOX/bevacizumab or switched therapy. The primary objective was adverse events (AE). The secondary objectives were objective response rate (ORR), disease control rate (DCR), and duration of response (DoR). Results: Fourteen patients were enrolled, and twelve patients were evaluable. All patients received FOLFOX in the past; two patients received 2 lines of chemotherapies; one patient received 3 lines and eight patients had ≥ 4 lines before the study. Five patients (41.7%) had disease resistance to FOLFOX. During the study, two patients received only 2 cycles of treatment due to poor PS. Two patients received 7 cycles, and nine patients completed 8 cycles of treatment. The common grade 3 &amp; 4 AEs included neutropenia (46.2%), lymphopenia (30.8%), and abdominal pain (15.4%). No complete response was observed; two patients obtained partial response; nine patients had stable disease, and only one patient had progressive disease. The ORR was 16.7%, and DCR was 91.7%. The median DoR was 5.1 (95% CI 3.2-7.0) months. The maximum tolerated dose for riluzole is 100 mg twice daily. Conclusions: Our study showed that riluzole plus mFOLFOX6/bevacizumab is well tolerated in patients with metastatic CRC and may have clinical activity in patients whose disease is resistant to FOLFOX. Further study is necessary to confirm the immunomodulation effect of riluzole. Clinical trial information: NCT04761614 .

Self-assembled fructo-oligosaccharide conjugated ferulic acid microparticle: Anticancer, anti-inflammatory and immunomodulatory effects

Fructo-oligosaccharide ferulic acid conjugate was synthesized, which self-assembles into disc shaped amorphous microparticles (FA FOS I), that could be used for targeted delivery to the colon. It is accessed for its ability to ameliorate colorectal cancer and inflammation. The microparticle exhibited selective cytotoxicity and apoptosis induction in colon cancer cells. The microparticle arrested the cell cycle in the G0-G1 phase. In-vivo anti-colorectal cancer activity were evaluated in colitis associated colon cancer mice model. There was a reduction in 60.83% of tumour lesions and 48.27% reduction in the malignant cells. The immunohistochemistry (IHC) revealed the suppression of inflammation and related inflammatory cytokines in the colon. The inhibition of cell proliferation, up-regulation of tumour suppressor protein and induction of apoptosis in the malignant cells were observed. The evaluation of immune modulation by FA FOS I microparticle revealed a 9% increase in WBC count and an increase in the infiltration of T-helper cells and cytotoxic T-cells into the tumour microenvironment. There was a considerable decrease in the expression of immune suppressing PD-L1 by the tumour cells on treatment with FA FOS I microparticle. All these data implicate better efficacy of the FA FOS I microparticle delivery to colon and amelioration of colitis associated colorectal cancer through multimodal route of inflammation reduction, apoptosis induction and positive modulation of immune system in the tumour microenvironment. FA FOS I was proposed to be a candidate drug which is physiologically and metabolically stable under gastro-intestinal condition and potentiating its effect in the target site large intestine; without being absorbed or transported to other tissue nor causing any adverse cytotoxicity to the neighbouring normal tissue.

Non-classical Monocytes Enhance the Efficacy of Immune Checkpoint Inhibitors on Colon Cancer in a Syngeneic Mouse Model.

The response rate to immune checkpoint inhibitors (ICIs) is approximately 10%-30% and only in a few cancer types. In the present study, we determined whether non-classical monocytes (NCMs) could enhance ICI efficacy in colon cancer using a syngeneic mouse model. The MC38 C57BL/6 mouse colon cancer model was used. Cells collected from the bone marrow of C57BL/6 mice were cultured, and NCMs were fractionated by cell sorting and administered via the tail veins to the mice implanted with MC38 cells. The anti-mouse PD-L1 antibody was administered three times, and tumor volume and overall survival were observed. More tumors were eradicated and more complete response occurred, after cotreatment with ICIs and NCMs than after treatment with ICIs alone. Moreover, no efficacy was observed when NCMs were administered alone. NCMs enhance ICI efficacy. The underlying mechanisms and clinical applications will be studied in the future.

Anticancer effect of hUC-MSC-derived exosome-mediated delivery of PMO-miR-146b-5p in colorectal cancer

Antisense oligonucleotide (ASO) is a novel therapeutic platform for targeted cancer therapy. Previously, we have demonstrated that miR-146b-5p plays an important role in colorectal cancer progression. However, a safe and effective strategy for delivery of an ASO to its targeted RNA remains as a major hurdle in translational advances. Human umbilical cord mesenchymal cell (hUC-MSC)–derived exosomes were used as vehicles to deliver an anti-miR-146b-5p ASO (PMO-146b). PMO-146b was assembled onto the surface of exosomes (e) through covalent conjugation to an anchor peptide CP05 (P) that recognized an exosomal surface marker, CD63, forming a complex named ePPMO-146b. After ePPMO-146b treatment, cell proliferation, uptake ability, and migration assays were performed, and epithelial-mesenchymal transition progression was evaluated in vitro. A mouse xenograft model was used to determine the antitumor effect and distribution of ePPMO-146b in vivo. ePPMO-146b was taken up by SW620 cells and effectively inhibited cell proliferation and migration. The conjugate also exerted antitumor efficacy in a xenograft mouse model of colon cancer by systematic administration, where PPMO-146b was enriched in tumor tissue. Our study highlights the potential of hUC-MSC-derived exosomes anchored with PPMO-146b as a novel safe and effective approach for PMO backboned ASO delivery.Graphical Schematic illustration of the preparation of an exosomal anchor peptide (CP05)-PMO that conjugately binds to exosomes from hUC-MSCs (ePPMO-146b) and the antitumor effect of ePPMO-146b in CRC, which occurs through the inhibition of Smad signaling and epithelial–mesenchymal transition.

Modulatory effects of supplementation of Lentinula edodes mycelia extract and l-arginine on the therapeutic efficacy of immunogenic chemotherapy in colon cancer-bearing mice.

Some chemotherapeutic drugs can induce cancer cell death and enhance antitumor T-cell immunity in cancer-bearing hosts. Immunomodulatory reagents could augment such chemotherapy-induced effects. We previously reported that oral digestion of Lentinula edodes mycelia (L.E.M.) extract or l-arginine supplementation can augment antitumor T-cell responses in cancer-bearing mice. In this study, the effects of L.E.M. extract with or without l-arginine on the therapeutic efficacy of immunogenic chemotherapy by 5-fluorouracil (5-FU)/oxaliplatin (L-OHP) and/or cyclophosphamide (CP) are examined using two mouse colon cancer models. In MC38 and CT26 cancer models, therapy with 5-FU/L-OHP/CP significantly suppressed tumor growth, and supplementation with L.E.M. extract halved the tumor volumes. However, the modulatory effect of L.E.M. extract was not significant. In the CT26 cancer model, supplementation with L.E.M. extract and l-arginine had no clear effect on tumor growth. In contrast, their addition to chemotherapy halved the tumor volumes, although the effect was not significant. There was no difference in the cytotoxicity of tumor-specific cytotoxic T cells generated from CT26-cured mice treated by chemotherapy alone versus chemotherapy combined with L.E.M. extract/ l-arginine. These results indicate that the antitumor effects of immunogenic chemotherapy were too strong to ascertain the effects of supplementation of L.E.M. extract and l-arginine, but these reagents nonetheless have immunomodulatory effects on the therapeutic efficacy of immunogenic chemotherapy in colon cancer-bearing mice.

Data Mining Suggests That CXCL14 Gene Silencing in Colon Cancer Is Due to Promoter Methylation.

CXCL14 is one of the most evolutionarily conserved members of the chemokine family and is constitutionally expressed in multiple organs, suggesting that it is involved in the homeostasis maintenance of the system. CXCL14 is highly expressed in colon epithelial cells and shows obvious gene silencing in clinical colon cancer samples, suggesting that its silencing is related to the immune escape of cancer cells. In this paper, we analyzed the expression profiles of multiple human clinical colon cancer datasets and mouse colon cancer models to reveal the variation trend of CXCL14 expression during colitis, colon polyps, primary colon cancer, and liver metastases. The relationship between CXCL14 gene silencing and promoter hypermethylation was revealed through the colorectal carcinoma methylation database. The results suggest that CXCL14 is a tumor suppressor gene in colorectal carcinoma which is activated first and then silenced during the process of tumor occurrence and deterioration. Promoter hypermethylation is the main cause of CXCL14 silencing. The methylation level of CXCL14 is correlated with the anatomic site of tumor occurrence, positively correlated with patient age, and associated with prognosis. Reversing the hypermethylation of CXCL14 may be an epigenetic therapy for colon cancer.

HRS mediates tumor immune evasion by regulating proteostasis-associated interferon pathway activation

By sorting receptor tyrosine kinases into endolysosomes, the endosomal sorting complexes required for transport (ESCRTs) are thought to attenuate oncogenic signaling in tumor cells. Paradoxically, ESCRT members are upregulated in tumors. Here, we show that disruption of hepatocyte growth factor-regulated tyrosine kinase substrate (HRS), a pivotal ESCRT component, inhibited tumor growth by promoting CD8+ Tcell infiltration in melanoma and colon cancer mouse models. HRS ablation led to misfolded protein accumulation and triggered endoplasmic reticulum (ER) stress, resulting in the activation of the type I interferon pathway in an inositol-requiring enzyme-1α (IRE1α)/X-box binding protein 1 (XBP1)-dependent manner. HRS was upregulated in tumor cells with high tumor mutational burden (TMB). HRS expression associates with the response to PD-L1/PD-1 blockade therapy in melanoma patients with high TMB tumors. HRS ablation sensitized anti-PD-1 treatment in mouse melanoma models. Our study shows a mechanism by which tumor cells with high TMB evade immune surveillance and suggests HRS as a promising target to improve immunotherapy.

Hypofractionated Radiotherapy with Prolonged Inter-Fraction Interval (Hypo-Slow RT) to Facilitate Immune Priming of Tumor

Daily fractionated radiotherapy may pose constant stress for immune activation. A mouse model was built to explore whether hypofractionated radiotherapy with prolonged inter-fraction interval (Hypo-Slow Radiotherapy, HSRT) may enable better immune-priming of tumor than daily radiotherapy. A subcutaneous MC38 murine colon cancer mouse model was irradiated at different radiation regimens. The impact of inter-fraction interval and dose per fraction on tumor control, immune mobilization, and synergistic effect with anti-PD-1 immunotherapy was explored. Immune activation was assessed by analyzing CD4+ and CD8+ T cells from peripheral blood, and intratumoral CD4+ and CD8+ T cells by flow cytometry; both were sampled three days after the completion of scheduled irradiation. In a fixed dose per fraction experiment, the 6×5Gy QOD and 6×5Gy QD irradiation schemes resulted in identical tumor control, while the 6×5Gy BIW scheme led to tumor progression; moreover, both QOD and BIW regimens showed ability to activate immune response whereas QD regimen did not. In a fixed biological equivalent dose (BED) experiment, the comparison of different regimens with increased dose per fraction and prolonged inter-fraction interval showed that 12×3Gy QD regimen, HSRT regimens, including 6×5Gy QOD, 4×7Gy BIW, and 2×11Gy QW, led to identical tumor control. Importantly, all HSRT regimens showed significant mobilization of host immunity whereas 12×3Gy QD did not. Both peripheral and intratumor CD4+ and CD8+ cell increase with increased inter-fraction interval and dose per fraction. Finally, all HSRT regimens combined with anti-PD-1 immunotherapy showed enhanced tumor growth delay than any single treatment while 12×3Gy QD regimen did not. This pre-clinical model demonstrated that conventional daily fractionated radiotherapy is not beneficial for host immune activation against tumor. Preliminary results suggested that prolonged inter-fraction interval with increased dose per fraction may be an optional strategy to balance the tumor control and immune activation.

Mismatch repair deficiency is not sufficient to elicit tumor immunogenicity.

DNA mismatch repair deficiency (MMRd) is associated with a high tumor mutational burden (TMB) and sensitivity to immune checkpoint blockade (ICB) therapy. Nevertheless, most MMRd tumors do not durably respond to ICB and critical questions remain about immunosurveillance and TMB in these tumors. In the present study, we developed autochthonous mouse models of MMRd lung and colon cancer. Surprisingly, these models did not display increased T cell infiltration or ICB response, which we showed to be the result of substantial intratumor heterogeneity of mutations. Furthermore, we found that immunosurveillance shapes the clonal architecture but not the overall burden of neoantigens, and T cell responses against subclonal neoantigens are blunted. Finally, we showed that clonal, but not subclonal, neoantigen burden predicts ICB response in clinical trials of MMRd gastric and colorectal cancer. These results provide important context for understanding immune evasion in cancers with a high TMB and have major implications for therapies aimed at increasing TMB.

ILC1-derived IFN-γ regulates macrophage activation in colon cancer

BackgroundTumor-associated macrophages (TAMs) are an important subset of innate immune cells in the tumor microenvironment, and they are pivotal regulators of tumor-promoting inflammation and tumor progression. Evidence has proven that TAM numbers are substantially increased in cancers, and most of these TAMs are polarized toward the alternatively activated M2 phenotype; Thus, these TAMs strongly promote the progression of cancer diseases. Type 1 innate lymphocytes (ILC1s) are present in high numbers in intestinal tissues and are characterized by the expression of the transcription factor T-bet and the secretion of interferon (IFN)-γ, which can promote macrophages to polarize toward the classically activated antitumor M1 phenotype. However, the relationship between these two cell subsets in colon cancer remains unclear.MethodsFlow cytometry was used to determine the percentages of M1-like macrophages, M2-like macrophages and ILC1s in colon cancer tissues and paracancerous healthy colon tissues in the AOM/DSS-induced mouse model of colon cancer. Furthermore, ILC1s were isolated and bone marrow-derived macrophages were generated to analyze the crosstalk that occurred between these cells when cocultured in vitro. Moreover, ILC1s were adoptively transferred or inhibited in vivo to explore the effects of ILC1s on tumor-infiltrating macrophages and tumor growth.ResultsWe found that the percentages of M1-like macrophages and ILC1s were decreased in colon cancer tissues, and these populations were positively correlated. ILC1s promoted the polarization of macrophages toward the classically activated M1-like phenotype in vitro, and this effect could be blocked by an anti-IFN-γ antibody. The in vivo results showed that the administration of the Group 1 innate lymphocyte-blocking anti-NK1.1 antibody decreased the number of M1-like macrophages in the tumor tissues of MC38 tumor-bearing mice and promoted tumor growth, and adoptive transfer of ILC1s inhibited tumors and increased the percentage of M1-like macrophages in MC38 tumor-bearing mice.ConclusionsOur studies preliminarily prove for the first time that ILC1s promote the activation of M1-like macrophages by secreting IFN-γ and inhibit the progression of colon cancer, which may provide insight into immunotherapeutic approaches for colon cancer.

Effects of concomitant use of THC and irinotecan on tumour growth and biochemical markers in a syngeneic mouse model of colon cancer.

Clinical treatment with the antineoplastic drug irinotecan (IRI) is often hindered by side effects that significantly reduce the quality of life of treated patients. Due to the growing public support for products with Δ9-tetrahydrocannabinol (THC), even though relevant scientific literature does not provide clear evidence of their high antitumour potential, some cancer patients take unregistered preparations containing up to 80 % THC. This study was conducted on a syngeneic colorectal cancer mouse model to test the efficiency and safety of concomitant treatment with IRI and THC. Male BALB/c mice subcutaneously injected with CT26 cells were receiving 60 mg/kg of IRI intraperitoneally on day 1 and 5 of treatment and/or 7 mg/kg of THC by gavage a day for 7 days. Treatment responses were evaluated based on changes in body, brain, and liver weight, tumour growth, blood cholinesterase activity, and oxidative stress parameters. Irinotecan's systemic toxicity was evidenced by weight loss and high oxidative stress. The important finding of this study is that combining THC with IRI diminishes IRI efficiency in inhibiting tumour growth. However, further studies, focused on more subtle molecular methods in tumour tissue and analytical analysis of IRI and THC distribution in tumour-bearing mice, are needed to prove our observations.

Jianpi Decoction Combined with Medroxyprogesterone Acetate Alleviates Cancer Cachexia and Prevents Muscle Atrophy by Directly Inhibiting E3 Ubiquitin Ligase.

To provide comprehensive evidence for the anti-cancer cachexia effect of Jianpi Decoction (JP) and to explore its mechanism of anti-cancer cachexia. A mouse model of colon cancer (CT26)-induced cancer cachexia (CC) was used to investigate the anti-CC effect of JP combined with medroxyprogesterone acetate (MPA). Thirty-six mice were equally divided into 6 groups: normal control, CC, MPA (100 mg•kg-1•d-1), MPA + low-dose (20 mg•kg-1•d-1) JP (L-JP), MPA + medium-dose (30 mg•kg-1•d-1) JP (M-JP), and MPA + high-dose (40 mg•kg-1•d-1) JP (H-JP) groups. After successful modeling, the mice were administered by gavage for 11 d. The body weight and tumor volume were measured and recorded every 2 d starting on the 8th day after implantation. The liver, heart, spleen, lung, kidney, tumor and gastrocnemius muscle of mice were collected and weighed. The pathological changes of the tumor was observed, and the cross-sectional area of the gastrocnemius muscle was calculated. The protein expressions of STAT3 and E3 ubiquitinase in the gastrocnemius muscle were measured by Western blot. In addition, an in vitro C2C12 myotube formation model was established to investigate the role of JP in hindering dexamethasone-induced muscle atrophy. In vitro experiments were divided into control, model, and JP serum groups. After 2-d administration, microscopic photographs were taken and myotube diameters were calculated. Western blot was performed to measure the protein expressions of STAT3 and E3 ubiquitinase. JP combined with MPA restored tumor-induced weight loss (P<0.05, vs. CC) and muscle fiber size (P<0.01, vs. CC). Mechanistically, JP reduced the expression of atrophy-related proteins MuRF1 and MAFbx in tumor-induced muscle atrophy in vivo (P<0.05, vs. CC). In addition, JP reduced the expression of atrophy-related proteins MuRF1 and MAFbx and p-STAT3 phosphorylation (P<0.05 or P<0.01 vs. model group) in C2C12 myotubes treated with dexamethasone in vitro. Administration of JP combined with MPA restores tumor-induced cachexia conditions. In addition, the profound effect of JP combined with MPA on tumor-induced cachexia may be due to its inhibition of muscle proteolysis (E3 ubiquitinase system).