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BACKGROUND: We previously showed that total tumor resection enhances metastatic growth in a syngeneic metastatic mouse model of neuroblastoma. In this study, we further investigated which surgical factors contributed most to metastatic growth. METHODS: Tumor cells derived from MYCN transgenic mice were subcutaneously injected into wild-type mice. Mice were randomly assigned to receive partial resection (PR group), subcutaneous implantation of a sponge (Sp group), or observation (Obs group). The lymph node metastasis volume and the frequency of lung metastasis were compared 14 days after assignment by measuring C-reactive protein (CRP) and interleukin-6 (IL-6) levels. RESULTS: The lymph node metastasis volume in the Sp group was larger than in the Obs group (148.4 [standard deviation {SD}: 209.5] vs. 10.2 [SD 12.8] mm3). The frequency of lung metastasis was greater in the Sp group than in the PR group (11.9 [SD 12.2] vs. 6.6 [SD 4.0] counts/slide). The CRP level in the Sp group was higher than in the PR group (2.3 [SD 0.5] vs. 1.5 [SD 0.4] µg/mL), and the IL-6 level in the Sp group was higher than in the PR or Obs groups (28.4 [SD 34.5] vs. 12.4 [SD 19.0] vs. 5.4 [SD 8.1] pg/mL). CONCLUSION: Metastatic growth may be enhanced by systemic inflammation.
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Proteína C-Reativa , Modelos Animais de Doenças , Inflamação , Neoplasias Pulmonares , Neuroblastoma , Animais , Neuroblastoma/patologia , Camundongos , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Proteína C-Reativa/metabolismo , Inflamação/patologia , Interleucina-6 , Metástase Linfática , Camundongos TransgênicosRESUMO
Basal-like breast cancer (BLBC) is the most aggressive and heterogeneous breast cancer (BC) subtype. Conventional chemotherapies represent next to surgery the most frequently employed treatment options. Unfortunately, resistant tumor phenotypes often develop, resulting in therapeutic failure. To identify the early events occurring upon the first drug application and initiating chemotherapy resistance in BLBC, we leveraged the WAP-T syngeneic mammary carcinoma mouse model and we developed a strategy combining magnetic-activated cell sorting (MACS)-based tumor cell enrichment with high-throughput transcriptome analyses. We discovered that chemotherapy induced a massive gene expression reprogramming toward stemness acquisition to tolerate and survive the cytotoxic treatment in vitro and in vivo. Retransplantation experiments revealed that one single cycle of cytotoxic drug combination therapy (Cyclophosphamide, Adriamycin and 5-Fluorouracil) suffices to induce resistant tumor cell phenotypes in vivo. We identified Axl and its ligand Pros1 as highly induced genes driving cancer stem cell (CSC) properties upon chemotherapy in vivo and in vitro. Furthermore, from our analysis of BLBC patient datasets, we found that AXL expression is also strongly correlated with CSC-gene signatures, a poor response to conventional therapies and worse survival outcomes in those patients. Finally, we demonstrate that AXL inhibition sensitized BLBC-cells to cytotoxic treatment in vitro. Together, our data support AXL as a promising therapeutic target to optimize the efficiency of conventional cytotoxic therapies in BLBC.
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Antineoplásicos , Carcinoma , Camundongos , Animais , Antineoplásicos/farmacologia , Transdução de Sinais , Ciclofosfamida/farmacologia , Células-Tronco Neoplásicas/metabolismo , Carcinoma/metabolismo , Linhagem Celular TumoralRESUMO
BACKGROUND: Infection with high-risk human papillomavirus (HPV) strains is one of the risk factors for the development of oral squamous cell carcinoma (OSCC). Some patients with HPV-positive OSCC have a better prognosis and respond better to various treatment modalities, including radiotherapy or immunotherapy. However, since HPV can only infect human cells, there are only a few immunocompetent mouse models available that enable immunological studies. Therefore, the aim of our study was to develop a transplantable immunocompetent mouse model of HPV-positive OSCC and characterize it in vitro and in vivo. METHODS: Two monoclonal HPV-positive OSCC mouse cell lines were established by inducing the expression of HPV-16 oncogenes E6 and E7 in the MOC1 OSCC cell line using retroviral transduction. After confirming stable expression of HPV-16 E6 and E7 with quantitative real-time PCR and immunofluorescence staining, the cell lines were further characterized in vitro using proliferation assay, wound healing assay, clonogenic assay and RNA sequencing. In addition, tumor models were characterized in vivo in C57Bl/6NCrl mice in terms of their histological properties, tumor growth kinetics, and radiosensitivity. Furthermore, immunofluorescence staining of blood vessels, hypoxic areas, proliferating cells and immune cells was performed to characterize the tumor microenvironment of all three tumor models. RESULTS: Characterization of the resulting MOC1-HPV cell lines and tumor models confirmed stable expression of HPV-16 oncogenes and differences in cell morphology, in vitro migration capacity, and tumor microenvironment characteristics. Although the cell lines did not differ in their intrinsic radiosensitivity, one of the HPV-positive tumor models, MOC1-HPV K1, showed a significantly longer growth delay after irradiation with a single dose of 15 Gy compared to parental MOC1 tumors. Consistent with this, MOC1-HPV K1 tumors had a lower percentage of hypoxic tumor area and a higher percentage of proliferating cells. Characteristics of the newly developed HPV-positive OSCC tumor models correlate with the transcriptomic profile of MOC1-HPV cell lines. CONCLUSIONS: In conclusion, we developed and characterized a novel immunocompetent mouse model of HPV-positive OSCC that exhibits increased radiosensitivity and enables studies of immune-based treatment approaches in HPV-positive OSCC.
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Carcinoma de Células Escamosas , Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Infecções por Papillomavirus , Humanos , Animais , Camundongos , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Neoplasias Bucais/genética , Neoplasias Bucais/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço , Infecções por Papillomavirus/complicações , Microambiente TumoralRESUMO
Stimulator of interferon genes (STING) agonists have shown potent anti-tumor efficacy in various mouse tumor models and have the potential to overcome resistance to immune checkpoint inhibitors (ICI) by linking the innate and acquired immune systems. First-generation STING agonists are administered intratumorally; however, a systemic delivery route would greatly expand the clinical use of STING agonists. Biochemical and cell-based experiments, as well as syngeneic mouse efficacy models, were used to demonstrate the anti-tumoral activity of ALG-031048, a novel STING agonist. In vitro, ALG-031048 is highly stable in plasma and liver microsomes and is resistant to degradation via phosphodiesterases. The high stability in biological matrices translated to good cellular potency in a HEK 293 STING R232 reporter assay, efficient activation and maturation of primary human dendritic cells and monocytes, as well as long-lasting, antigen-specific anti-tumor activity in up to 90% of animals in the CT26 mouse colon carcinoma model. Significant reductions in tumor growth were observed in two syngeneic mouse tumor models following subcutaneous administration. Combinations of ALG-031048 and ICIs further enhanced the in vivo anti-tumor activity. This initial demonstration of anti-tumor activity after systemic administration of ALG-031048 warrants further investigation, while the combination of systemically administered ALG-031048 with ICIs offers an attractive approach to overcome key limitations of ICIs in the clinic.
Assuntos
Neoplasias do Colo , Neoplasias , Camundongos , Animais , Humanos , Células HEK293 , Neoplasias/patologia , Neoplasias do Colo/tratamento farmacológico , Modelos Animais de Doenças , Imunoterapia , Microambiente TumoralRESUMO
Cold atmospheric plasma (CAP) may have applications in treating various types of malignant tumors. This study assessed the anticancer effects of CAP using melanoma and colon cancer cell lines. CAP treatment significantly reduced the in vitro viability of melanoma and colon cancer cell lines and had a negligible effect on the viability of normal human melanocytes. Additionally, CAP and epidermal growth factor receptor (EGFR) inhibitor had an additive anticancer effect in a CAP-resistant melanoma cell line. Reactive oxygen and nitrogen species known to be generated by CAP enhanced the anticancer effects of CAP and EGFR inhibitors. The in vivo anticancer activities of CAP were evaluated by testing its effects against syngeneic tumors induced in mice by melanoma and colon cancer cells. CAP treatment reduced tumor volume and weight in both cancer models, with the extent of tumor reduction dependent on the duration and number of CAP treatments. Histologic examination also revealed the tumoricidal effects of CAP in both tumor models. In conclusion, CAP inhibits the growth of mouse melanoma and colon cancer cell lines in vitro and shows tumoricidal effects against mouse models of melanoma and colon cancer in vivo.
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Neoplasias do Colo , Melanoma , Gases em Plasma , Humanos , Animais , Camundongos , Gases em Plasma/farmacologia , Gases em Plasma/uso terapêutico , Linhagem Celular Tumoral , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Neoplasias do Colo/tratamento farmacológico , Receptores ErbBRESUMO
BACKGROUND: Human epidermal growth factor receptor-2 (HER2)-targeted therapies prolong survival in HER2-positive breast cancer patients. Benefit stems primarily from improved control of systemic disease, but up to 50% of patients progress to incurable brain metastases due to acquired resistance and/or limited permeability of inhibitors across the blood-brain barrier. Neratinib, a potent irreversible pan-tyrosine kinase inhibitor, prolongs disease-free survival in the extended adjuvant setting, and several trials evaluating its efficacy alone or combination with other inhibitors in early and advanced HER2-positive breast cancer patients are ongoing. However, its efficacy as a first-line therapy against HER2-positive breast cancer brain metastasis has not been fully explored, in part due to the lack of relevant pre-clinical models that faithfully recapitulate this disease. Here, we describe the development and characterisation of a novel syngeneic model of spontaneous HER2-positive breast cancer brain metastasis (TBCP-1) and its use to evaluate the efficacy and mechanism of action of neratinib. METHODS: TBCP-1 cells were derived from a spontaneous BALB/C mouse mammary tumour and characterised for hormone receptors and HER2 expression by flow cytometry, immunoblotting and immunohistochemistry. Neratinib was evaluated in vitro and in vivo in the metastatic and neoadjuvant setting. Its mechanism of action was examined by transcriptomic profiling, function inhibition assays and immunoblotting. RESULTS: TBCP-1 cells naturally express high levels of HER2 but lack expression of hormone receptors. TBCP-1 tumours maintain a HER2-positive phenotype in vivo and give rise to a high incidence of spontaneous and experimental metastases in the brain and other organs. Cell proliferation/viability in vitro is inhibited by neratinib and by other HER2 inhibitors, but not by anti-oestrogens, indicating phenotypic and functional similarities to human HER2-positive breast cancer. Mechanistically, neratinib promotes a non-apoptotic form of cell death termed ferroptosis. Importantly, metastasis assays demonstrate that neratinib potently inhibits tumour growth and metastasis, including to the brain, and prolongs survival, particularly when used as a neoadjuvant therapy. CONCLUSIONS: The TBCP-1 model recapitulates the spontaneous spread of HER2-positive breast cancer to the brain seen in patients and provides a unique tool to identify novel therapeutics and biomarkers. Neratinib-induced ferroptosis provides new opportunities for therapeutic intervention. Further evaluation of neratinib neoadjuvant therapy is warranted.
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Neoplasias Encefálicas/prevenção & controle , Neoplasias Encefálicas/secundário , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ferroptose/efeitos dos fármacos , Quinolinas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Biologia Computacional/métodos , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Imuno-Histoquímica , Isoenxertos , Camundongos , Terapia de Alvo Molecular , Terapia Neoadjuvante , Quinolinas/uso terapêutico , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismoRESUMO
Compared to normal tissues, many cancer cells overexpress autotaxin (ATX). This secreted enzyme produces extracellular lysophosphatidate, which signals through 6 GPCRs to drive cancer progression. Our previous work showed that ATX inhibition decreases 4T1 breast tumor growth in BALB/c mice by 60% for about 11 d. However, 4T1 cells do not produce significant ATX. Instead, the ATX is produced by adjacent mammary adipose tissue. We investigated the molecular basis of this interaction in human and mouse breast tumors. Inflammatory mediators secreted by breast cancer cells increased ATX production in adipose tissue. The increased lysophosphatidate signaling further increased inflammatory mediator production in adipose tissue and tumors. Blocking ATX activity in mice bearing 4T1 tumors with 10 mg/kg/d ONO-8430506 (a competitive ATX inhibitor, IC90 = 100 nM; Ono Pharma Co., Ltd., Osaka, Japan) broke this vicious inflammatory cycle by decreasing 20 inflammatory mediators by 1.5-8-fold in cancer-inflamed adipose tissue. There was no significant decrease in inflammatory mediator levels in fat pads that did not bear tumors. ONO-8430506 also decreased plasma TNF-α and G-CSF cytokine levels by >70% and leukocyte infiltration in breast tumors and adjacent adipose tissue by >50%. Hence, blocking tumor-driven inflammation by ATX inhibition is effective in decreasing tumor growth in breast cancers where the cancer cells express negligible ATX.
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Tecido Adiposo/enzimologia , Neoplasias da Mama/enzimologia , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Glândulas Mamárias Animais/enzimologia , Glândulas Mamárias Humanas/enzimologia , Neoplasias Mamárias Experimentais/enzimologia , Proteínas de Neoplasias/biossíntese , Diester Fosfórico Hidrolases/biossíntese , Tecido Adiposo/patologia , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Feminino , Humanos , Glândulas Mamárias Animais/patologia , Glândulas Mamárias Humanas/patologia , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Neoplasias/genética , Diester Fosfórico Hidrolases/genéticaRESUMO
Although many preclinical studies have implicated ß3 integrin receptors (αvß3 and αIIbß3) in cancer progression, ß3 inhibitors have shown only modest efficacy in patients with advanced solid tumours. The limited efficacy of ß3 inhibitors in patients could arise from our incomplete understanding of the precise function of ß3 integrin and, consequently, inappropriate clinical application. Data from animal studies are conflicting and indicate heterogeneity with respect to the relative contributions of ß3-expressing tumour and stromal cell populations in different cancers. Here we aimed to clarify the function and relative contributions to metastasis of tumour versus stromal ß3 integrin in clinically relevant models of spontaneous breast cancer metastasis, with particular emphasis on bone metastasis. We show that stable down-regulation of tumour ß3 integrin dramatically impairs spontaneous (but not experimental) metastasis to bone and lung without affecting primary tumour growth in the mammary gland. Unexpectedly, and in contrast to subcutaneous tumours, orthotopic tumour vascularity, growth and spontaneous metastasis were not altered in mice null for ß3 integrin. Tumour ß3 integrin promoted migration, protease expression and trans-endothelial migration in vitro and increased vascular dissemination in vivo, but was not necessary for bone colonization in experimental metastasis assays. We conclude that tumour, rather than stromal, ß3 expression is essential and is required early for efficient spontaneous breast cancer metastasis to bone and soft tissues. Accordingly, differential gene expression analysis in cohorts of breast cancer patients showed a strong association between high ß3 expression, early metastasis and shorter disease-free survival in patients with oestrogen receptor-negative tumours. We propose that ß3 inhibitors may be more efficacious if used in a neoadjuvant setting, rather than after metastases are established.
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Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/secundário , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Integrina beta3/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Células Estromais/metabolismo , Animais , Neoplasias Ósseas/genética , Neoplasias Ósseas/prevenção & controle , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Movimento Celular , Intervalo Livre de Doença , Regulação para Baixo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Integrina beta3/genética , Neoplasias Mamárias Experimentais/genética , Camundongos Endogâmicos BALB C , Camundongos Knockout , Invasividade Neoplásica , Transdução de Sinais , Células Estromais/patologia , Fatores de Tempo , Transfecção , Carga TumoralRESUMO
Autotaxin is a secreted enzyme that produces most extracellular lysophosphatidate, which stimulates 6 G-protein-coupled receptors. Lysophosphatidate promotes cancer cell survival, growth, migration, invasion, metastasis, and resistance to chemotherapy and radiotherapy. The present work investigated whether inhibiting autotaxin could decrease breast tumor growth and metastasis. We used a new autotaxin inhibitor (ONO-8430506; IC90=100 nM), which decreased plasma autotaxin activity by >60% and concentrations of unsaturated lysophosphatidates by >75% for 24 h compared with vehicle-treated mice. The effects of ONO-8430506 on tumor growth were determined in a syngeneic orthotopic mouse model of breast cancer following injection of 20,000 BALB/c mouse 4T1 or 4T1-12B cancer cells. We show for the first time that inhibiting autotaxin decreases initial tumor growth and subsequent lung metastatic nodules both by 60% compared with vehicle-treated mice. Significantly, 4T1 cells express negligible autotaxin compared with the mammary fat pad. Autotaxin activity in the fat pad of nontreated mice was increased 2-fold by tumor growth. Our results emphasize the importance of tumor interaction with its environment and the role of autotaxin in promoting breast cancer growth and metastasis. We also established that autotaxin inhibition could provide a novel therapeutic approach to blocking the adverse effects of lysophosphatidate in cancer.
Assuntos
Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Mamárias Experimentais/tratamento farmacológico , Inibidores de Fosfodiesterase/uso terapêutico , Diester Fosfórico Hidrolases/efeitos dos fármacos , Animais , Carbolinas/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Neoplasias Pulmonares/secundário , Lisofosfolipídeos/antagonistas & inibidores , Lisofosfolipídeos/farmacologia , Neoplasias Mamárias Experimentais/patologia , CamundongosRESUMO
Background: This study aimed to evaluate the effects of continuous administration of xylitol (a commonly used dental prebiotic) via a subcutaneous osmotic minipump in a B16F10 syngeneic mouse model. Methods: The B16F10 syngeneic model consisted of 6-8-week-old C57BL/6 male mice subcutaneously injected with five × 105 B16F10 cells suspended in 100 µl PBS in the right flank. The mice were randomly assigned to two groups: Group 1 was the treatment group with 10% xylitol-loaded pumps (n=10), while Group 2 was the control group with saline-loaded pumps (n=10). Alzet minipumps were implanted subcutaneously in the left flank of B16F10-injected mice once more than 50% of all mice developed palpable tumors. After pump implantation surgery, the mice were monitored daily and weighed 2-3x/week. Tumor sizes were measured with calipers 2-3x/week, and all mice were euthanized when their tumors became too large (20 mm on any axis or 2,000 mm3). The excised tumors were weighed and cut in half, with one half sent for histology and the other for metabolomic analysis. Results: The xylitol-treated group survived substantially longer than the control group. The tumor size was reduced by approximately 35% by volume. Histological sections of xylitol treat mice suggested reduced infiltration and angiogenesis, which is consistent with previous studies. The metabolomic analysis demonstrates that xylitol reduces the tumor production of histamine, NADP+, ATP, and glutathione from the tumor, thereby improving the host immune response with ROS reactive oxygen species. Conclusions: The results of this study suggest that xylitol has potential as an adjunct to oncological treatment and is being further investigated in comparison to monoclonal antibody therapy (Opdualag).
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BACKGROUND: Preclinical models are invaluable for studies of head and neck cancer. There is growing interest in the use of orthotopic syngeneic models, wherein cell lines are injected into the oral cavity of immunocompetent mice. In this brief report, we describe injection of mouse oral cancer 1 (MOC1) cells into the buccal mucosa and illustrate the tumor growth pattern, lymph node response, and changes in the tumor immune microenvironment over time. METHODS: MOC1 cells were injected into the buccal mucosa of C57BL6 mice. Animals were sacrificed at 7, 14, 21, or 27 days. Tumors and lymph nodes were analyzed by flow cytometry. RESULTS: All mice developed tumors by day 7 and required euthanasia for tumor burden and/or weight loss by day 27. Lymph node mapping showed that these tumors reliably drain to a submandibular lymph node. The proportion of intratumoral CD8+ T cells decreased over time, while neutrophilic myeloid cells increased dramatically. Growth of orthotopic MOC2 and MOC22 also showed similar growth patterns versus published data in flank tumors. CONCLUSIONS: When used orthotopically in the buccal mucosa, the MOC1 model induces a robust lymph node response and distinct pattern of immune cell infiltration, with peak immune infiltration by day 14.
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Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Animais , Camundongos , Mucosa Bucal , Camundongos Endogâmicos C57BL , Microambiente Tumoral , Linhagem Celular TumoralRESUMO
BACKGROUND/AIM: We and others have previously shown that cell fusion plays an important role in cancer metastasis. Color coding of cancer and stromal cells with spectrally-distinct fluorescent proteins is a powerful tool, as pioneered by our laboratory to detect cell fusion. We have previously reported color-coded cell fusion between cancer cells and stromal cells in metastatic sites by using color-coded EL4 murine lymphoma cells and host mice expressing spectrally-distinct fluorescent proteins. Cell fusion occurred between cancer cells or, between cancer cells and normal cells, such as macrophages, fibroblasts, and mesenchymal stem cells. In the present study, the aim was to morphologically classify the fusion-hybrid cells observed in the primary tumor and multiple metastases EL4 formed from cells expressing red fluorescent protein (RFP) in transgenic mice expressing green fluorescent protein (GFP), in a syngeneic model. MATERIALS AND METHODS: RFP-expressing EL4 murine lymphoma cells were cultured in vitro. EL4-RFP cells were harvested and injected intraperitoneally into immunocompetent transgenic C57/BL6-GFP mice to establish a syngeneic model. Two weeks later, mice were sacrificed and each organ was harvested, cultured, and observed using confocal microscopy. RESULTS: EL4 intraperitoneal tumors (primary) and metastases in the lung, liver, blood, and bone marrow were formed. All tumors were harvested and cultured. In all specimens, RFP-EL4 cells, GFP-stromal cells, and fused yellow-fluorescent hybrid cells were observed. The fused hybrid cells showed various morphologies. Immune cell-like round-shaped yellow-fluorescent fused cells had a tendency to decrease with time in liver metastases and circulating blood. In contrast fibroblast-like spindle-shaped yellow-fluorescent fused cells increased in the intraperitoneal primary tumor, lung metastases, and bone marrow. CONCLUSION: Cell fusion between EL4-RFP cells and GFP stromal cells occurred in primary tumors and all metastatic sites. The morphology of the fused hybrid cells varied in the primary and metastatic sites. The present results suggest that fused cancer and stromal hybrid cells of varying morphology may play an important role in cancer progression.
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Fusão Celular , Modelos Animais de Doenças , Proteínas Luminescentes , Linfoma , Camundongos Transgênicos , Proteína Vermelha Fluorescente , Células Estromais , Animais , Camundongos , Células Estromais/patologia , Células Estromais/metabolismo , Linhagem Celular Tumoral , Linfoma/patologia , Linfoma/genética , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Metástase Neoplásica , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células Híbridas/patologiaRESUMO
Purpose: The mortality in patients with MYCN-amplified high-risk neuroblastoma remains greater than 50% despite advances in multimodal therapy. Novel therapies are urgently needed that requires preclinical evaluation in appropriate mice models. Combinatorial treatment with high-dose radiotherapy (HDRT) and immunotherapy has emerged as an effective treatment option in a variety of cancers. Current models of neuroblastoma do not recapitulate the anatomic and immune environment in which multimodal therapies can be effectively tested, and there is a need for an appropriate syngeneic neuroblastoma mice model to study interaction of immunotherapy with host immune cells. Here, we develop a novel syngeneic mouse model of MYCN-amplified neuroblastoma and report the relevance and opportunities of this model to study radiotherapy and immunotherapy. Materials and methods: A syngeneic allograft tumor model was developed using the murine neuroblastoma cell line 9464D derived a tumor from TH-MYCN transgenic mouse. Tumors were generated by transplanting 1 mm3 portions of 9464D flank tumors into the left kidney of C57Bl/6 mice. We investigated the effect of combining HDRT with anti-PD1 antibody on tumor growth and tumor microenvironment. HDRT (8 Gy x 3) was delivered by the small animal radiation research platform (SARRP). Tumor growth was monitored by ultrasound. To assess the effect on immune cells tumors sections were co-imuunostained for six biomarkers using the Vectra multispectral imaging platform. Results: Tumor growth was uniform and confined to the kidney in 100% of transplanted tumors. HDRT was largely restricted to the tumor region with minimal scattered out-of-field dose. Combinatorial treatment with HDRT and PD-1 blockade significantly inhibited tumor growth and prolonged mice survival. We observed augmented T-lymphocyte infiltration, especially CD3+CD8+ lymphocytes, in tumors of mice which received combination treatment. Conclusion: We have developed a novel syngeneic mouse model of MYCN amplified high-risk neuroblastoma. We have utilized this model to show that combining immunotherapy with HDRT inhibits tumor growth and prolongs mice survival.
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Cancer-associated mesothelial cells (CAMCs) in the tumor microenvironment are thought to promote growth and immune evasion. We find that, in mouse and human ovarian tumors, cancer cells express anti-Müllerian hormone (AMH) while CAMCs express its receptor AMHR2, suggesting a paracrine axis. Factors secreted by cancer cells induce AMHR2 expression during their reprogramming into CAMCs in mouse and human in vitro models. Overexpression of AMHR2 in the Met5a mesothelial cell line is sufficient to induce expression of immunosuppressive cytokines and growth factors that stimulate ovarian cancer cell growth in an AMH-dependent way. Finally, syngeneic cancer cells implanted in transgenic mice with Amhr2-/- CAMCs grow significantly slower than in wild-type hosts. The cytokine profile of Amhr2-/- tumor-bearing mice is altered and their tumors express less immune checkpoint markers programmed-cell-death 1 (PD1) and cytotoxic T lymphocyte-associated protein 4 (CTLA4). Taken together, these data suggest that the AMH/AMHR2 axis plays a critical role in regulating the pro-tumoral function of CAMCs in ovarian cancer.
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Neoplasias Ovarianas , Hormônios Peptídicos , Feminino , Humanos , Animais , Camundongos , Hormônio Antimülleriano/genética , Neoplasias Ovarianas/genética , Camundongos Transgênicos , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Microambiente TumoralRESUMO
BACKGROUND/AIM: All cancer types so far tested are methionine-addicted. Targeting the methionine addiction of cancer with recombinant methioninase (rMETase) has shown great progress in vitro, in mouse models, and in the clinic. However, administration of rMETase requires multiple doses per day. In the present study, we determined if rMETase-producing Escherichia coli JM109 (E. coli JM109-rMETase) might be an effective anticancer agent when installed into the microbiome. MATERIALS AND METHODS: E. coli JM109-rMETase was administered to a syngeneic model of MC38 colon cancer growing subcutaneously in C57BL/6 mice. JM109-rMETase was administered orally by gavage to the mice twice per day. Tumor size was measured with calipers. RESULTS: The administration of E. coli JM109-rMETase twice a day significantly inhibited MC38 colon-cancer growth. E. coli JM109-rMETase was found in the stool of treated mice, indicating it had entered the microbiome. CONCLUSION: The present study indicates the potential of microbiome-based treatment of cancer targeting methionine addiction.
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Neoplasias do Colo , Microbiota , Animais , Camundongos , Liases de Carbono-Enxofre/farmacologia , Liases de Carbono-Enxofre/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Modelos Animais de Doenças , Escherichia coli , Metionina , Camundongos Endogâmicos C57BL , Camundongos Nus , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêuticoRESUMO
Peripheral T-cell lymphoma (PTCL) represents a rare group of heterogeneous diseases in urgent need of effective treatments. A scarcity of disease-relevant preclinical models hinders research advances. Here, we isolated a novel mouse (m)PTCL by serially transplanting a lymphoma from a germinal center B-cell hyperplasia model (Cγ1-Cre Blimp1fl/fl ) through immune-competent mice. Lymphoma cells were identified as clonal TCRß+ T-helper cells expressing T-follicular helper markers. We also observed coincident B-cell activation and development of a de novo B-cell lymphoma in the model, reminiscent of B-cell activation/lymphomagenesis found in human PTCL. Molecular profiling linked the mPTCL to the high-risk "GATA3" subtype of PTCL, showing GATA3 and Th2 gene expression, PI3K/mTOR pathway enrichment, hyperactivated MYC, and genome instability. Exome sequencing identified a human-relevant oncogenic ß-catenin mutation possibly involved in T-cell lymphomagenesis. Prolonged treatment responses were achieved in vivo by targeting ATR in the DNA damage response (DDR), a result corroborated in PTCL cell lines. This work provides mechanistic insight into the molecular and immunological drivers of T-cell lymphomagenesis and proposes DDR inhibition as an effective and readily translatable therapy in PTCL.
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Dano ao DNA , Fator de Transcrição GATA3 , Linfoma de Células T Periférico , Animais , Linfócitos B/imunologia , Linfócitos B/metabolismo , Fator de Transcrição GATA3/genética , Linfoma de Células T Periférico/genética , Linfoma de Células T Periférico/imunologia , Linfoma de Células T Periférico/metabolismo , Linfoma de Células T Periférico/patologia , Camundongos , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Prostate-specific membrane antigen (PSMA)-targeted radionuclide therapy (RNT) may increase tumor immunogenicity. We aimed at exploiting this effect by combining RNT with immunotherapy in a mouse model of prostate cancer (PC). Methods: C57BL/6-mice bearing syngeneic RM1-PGLS tumors were treated with 225Ac-PSMA617, an anti-PD-1 antibody, or both. Therapeutic efficacy was assessed by tumor volume measurements (CT), time to progression (TTP), and survival. Results: PSMA RNT or anti-PD-1 alone tended to prolong TTP (isotype control, 25 d; anti-PD-1, 33.5 d [P = 0.0153]; RNT, 30 d [P = 0.1038]) and survival (control, 28 d; anti-PD-1, 37 d [P = 0.0098]; RNT, 32 d [P = 0.1018]). Combining PSMA RNT and anti-PD-1 significantly improved disease control compared with either monotherapy. TTP was extended to 47.5 d (P ≤ 0.0199 vs. monotherapies), and survival to 51.5 d (P ≤ 0.0251 vs. monotherapies). Conclusion: PSMA RNT and PD-1 blockade synergistically improve therapeutic outcomes in our PC model, supporting the evaluation of RNT and immunotherapy combinations for PC patients.
Assuntos
Dipeptídeos/uso terapêutico , Compostos Heterocíclicos com 1 Anel/uso terapêutico , Inibidores de Checkpoint Imunológico/farmacologia , Imunoterapia/métodos , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/radioterapia , Actínio , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptor de Morte Celular Programada 1/imunologia , Antígeno Prostático Específico , Neoplasias da Próstata/patologiaRESUMO
Epithelial ovarian cancer (EOC) is the leading cause of gynecological cancer-related mortality in the developed world. EOC is a heterogeneous disease represented by several histological and molecular subtypes. Therefore, exploration of relevant preclinical animal models that consider the heterogenic nature of EOC is of great importance for the development of novel therapeutic strategies that can be translated clinically to combat this devastating disease. In this review, we discuss recent progress in the development of preclinical mouse models for EOC study as well as their advantages and limitations.
Assuntos
Antineoplásicos/uso terapêutico , Modelos Animais de Doenças , Neoplasias Ovarianas/tratamento farmacológico , Animais , Avaliação Pré-Clínica de Medicamentos , Feminino , CamundongosRESUMO
BiTE ® (bispecific T-cell engager) molecules exert antitumor activity by binding one arm to CD3 on cytotoxic T-cells and the other arm to a tumor-associated antigen. We generated a fully mouse cross-reactive mesothelin (MSLN)-targeted BiTE molecule that is genetically fused to a Fc-domain for half-life extension, and evaluated biodistribution and tumor targeting of a zirconium-89 (89Zr)-labeled MSLN HLE BiTE molecule in 4T1 breast cancer bearing syngeneic mice with positron emission tomography (PET). Biodistribution of 50 µg 89Zr-MLSN HLE BiTE was studied over time by PET imaging in BALB/c mice and revealed uptake in tumor and lymphoid tissues with an elimination half-life of 63.4 hours. Compared to a non-targeting 89Zr-control HLE BiTE, the 89Zr-MLSN HLE BiTE showed a 2-fold higher tumor uptake and higher uptake in lymphoid tissues. Uptake in the tumor colocalized with mesothelin expression, while uptake in the spleen colocalized with CD3 expression. Evaluation of the effect of protein doses on the biodistribution and tumor targeting of 89Zr-MSLN HLE BiTE revealed for all dose groups that uptake in the spleen was faster than in the tumor (day 1 vs day 5). The lowest dose of 10 µg 89Zr-MSLN HLE BiTE had higher spleen uptake and faster blood clearance compared to higher doses of 50 µg and 200 µg. 89Zr-MSLN HLE BiTE tumor uptake was similar at all doses. Conclusion: The MSLN HLE BiTE showed specific tumor uptake and both arms contributed to the biodistribution profile. These findings support the potential for clinical translation of HLE BiTE molecules.
RESUMO
Somatic mutations in EGFR and KRAS as well as chromosome rearrangements affecting ALK, ROS1, and RET have been identified in human lung adenocarcinoma (LUAD). We here developed organoid-based orthotopic and syngeneic mouse models for studies of the pathogenesis and treatment of LUAD. We isolated EpCAM-positive epithelial cells from mouse lungs and cultured them as organoids to maintain epithelial stem cell properties. These cells were transformed by KRAS(G12V) or EML4-ALK and then transplanted via the trachea into the lungs of the syngeneic mice, where they formed tumors that expressed the lung lineage marker TTF-1 and which closely recapitulated the pathology of human LUAD. Treatment with crizotinib suppressed the growth of tumors formed by the EML4-ALK-expressing lung epithelial cells in a subcutaneous transplantation model. Organoid culture of normal lung epithelial cells resulted in enrichment of EpCAM+SCA-1(Ly6a)+ cells as well as in that of cells expressing another member of the Ly6 protein family, Ly6d, which was found to be required for the growth of the LUAD-initiating cells expressing KRAS(G12V) or EML4-ALK. We also found that a high expression level of LY6D was associated with poor prognosis in human LUAD. Our results thus suggest that LY6D is a potential lung cancer stem cell marker.