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Frontiers in Immunology 2022In recent years, there has been an emphasis on harnessing the immune system for therapeutic interventions. Adoptive cell therapies (ACT) have emerged as an effective... (Review)
Review
In recent years, there has been an emphasis on harnessing the immune system for therapeutic interventions. Adoptive cell therapies (ACT) have emerged as an effective option for B-cell derived hematological malignancies. Despite remarkable successes with ACT, immune dysregulation and the leukemia microenvironment can critically alter clinical responses. Therefore, preclinical modeling can contribute to the advancement of ACT for leukemias. Human xenografts, the current mainstay of ACT models, cannot evaluate the impact of the immunosuppressive leukemia microenvironment on adoptively transferred cells. Syngeneic mouse models utilize murine tumor models and implant them into immunocompetent mice. This provides an alternative model, reducing the need for complicated breeding strategies while maintaining a matched immune system, stromal compartment, and leukemia burden. Syngeneic models that evaluate ACT have analyzed the complexity of cytotoxic T lymphocytes, T cell receptor transgenics, and chimeric antigen receptors. This review examines the immunosuppressive features of the leukemia microenvironment, discusses how preclinical modeling helps predict ACT associated toxicities and dysfunction, and explores publications that have employed syngeneic modeling in ACT studies for the improvement of therapy for leukemias.
Topics: Animals; Humans; Immunosuppressive Agents; Immunotherapy, Adoptive; Leukemia; Mice; Receptors, Chimeric Antigen; T-Lymphocytes, Cytotoxic; Tumor Microenvironment
PubMed: 35401520
DOI: 10.3389/fimmu.2022.867103 -
Hepatology (Baltimore, Md.) May 2023The treatment of hepatocellular carcinoma (HCC) has been transformed by the use of immune checkpoint inhibitors. However, most patients with HCC do not benefit from...
BACKGROUND AND AIMS
The treatment of hepatocellular carcinoma (HCC) has been transformed by the use of immune checkpoint inhibitors. However, most patients with HCC do not benefit from treatment with immunotherapy. There is an urgent need to understand the mechanisms that underlie response or resistance to immunotherapy for patients with HCC. The use of syngeneic mouse models that closely recapitulate the heterogeneity of human HCC will provide opportunities to examine the complex interactions between cancer cells and nonmalignant cells in the tumor microenvironment.
APPROACH AND RESULTS
We leverage a multifaceted approach that includes imaging mass cytometry and suspension cytometry by time of flight to profile the tumor microenvironments of the Hep53.4, Hepa 1-6, RIL-175, and TIBx (derivative of TIB-75) syngeneic mouse HCC models. The immune tumor microenvironments vary across these four models, and various immunosuppressive pathways exist at baseline in orthotopic liver tumors derived from these models. For instance, TIBx, which is resistant to anti-programmed cell death protein 1 therapy, contains a high proportion of "M2-like" tumor-associated macrophages with the potential to diminish antitumor immunity. Investigation of The Cancer Genome Atlas reveals that the baseline immunologic profiles of Hep53.4, RIL-175, and TIBx are broadly representative of human HCCs; however, Hepa 1-6 does not recapitulate the immune tumor microenvironment of the vast majority of human HCCs.
CONCLUSIONS
There is a wide diversity in the immune tumor microenvironments in preclinical models and in human HCC, highlighting the need to use multiple syngeneic HCC models to improve the understanding of how to treat HCC through immune modulation.
Topics: Animals; Humans; Mice; Carcinoma, Hepatocellular; Cell Line, Tumor; Immunotherapy; Liver Neoplasms; Tumor Microenvironment; Programmed Cell Death 1 Receptor
PubMed: 35941803
DOI: 10.1002/hep.32707 -
Molecular Metabolism Dec 2022Adipose tissue is the largest endocrine organ. When activated by cancer cells, adipocytes secrete adipocytokines and release fatty acids, which are then transferred to...
OBJECTIVE
Adipose tissue is the largest endocrine organ. When activated by cancer cells, adipocytes secrete adipocytokines and release fatty acids, which are then transferred to cancer cells and used for structural and biochemical support. How this metabolic symbiosis between cancer cells and adipocytes affects skeletal muscle and thermogenesis during cancer cachexia is unknown. Cancer cachexia is a multiorgan syndrome and how the communication between tissues is established has yet to be determined. We investigated adipose tissue secretory factors and explored their role in crosstalk of adipocytes, muscle, and tumor during pancreatic cancer cachexia.
METHODS
We used a pancreatic cancer cachexia mouse model generated by syngenic implantation of pancreatic ductal adenocarcinoma (PDAC) cells (KPC) intraperitoneally into C57BL/6 mice and Lcn2-knockout mice. For in vitro studies, adipocytes (3T3-L1 and primary adipocytes), cachectic cancer cells (Panc0203), non-cachectic cancer cells (Du145 cells), and skeletal muscle cells (C2C12 myoblasts) were used.
RESULTS
To identify molecules involved in the crosstalk of adipose tissue with muscle and tumors, we treated 3T3-L1 adipocytes with conditioned medium (CM) from cancer cells. Upon screening the secretomes from PDAC-induced adipocytes, several adipocytokines were identified, including lipocalin 2 (Lcn2). We investigated Lcn2 as a potential mediator of cachexia induced by adipocytes in response to PDAC. During tumor progression, mice exhibited a decline in body weight gain, which was accompanied by loss of adipose and muscle tissues. Tumor-harboring mice developed drastic hypothermia because of a dramatic loss of fat in brown adipose tissue (BAT) and suppression of the thermogenesis pathway. We inhibited Lcn2 with an anti-Lcn2 antibody neutralization or genomic ablation in mice. Lcn2 deficiency significantly improved body temperature in tumor-bearing mice, which was supported by the increased expression of Ucp1 and β3-adrenergic receptor in BAT. In addition, Lcn2 inhibition abrogated the loss of fat and muscle in tumor-bearing mice. In contrast to tumor-bearing WT mice, the corresponding Lcn2-knockout mice showed reduced ATGL expression in iWAT and decreased the expression of muscle atrophy molecular markers MuRF-1 and Fbx32.
CONCLUSIONS
This study showed that Lcn2 is causally involved in the dysregulation of adipose tissue-muscle-tumor crosstalk during pancreatic cancer cachexia. Therapeutic targets that suppress Lcn2 may minimize the progression of cachexia.
Topics: Animals; Mice; Adipocytes; Adipokines; Adipose Tissue, Brown; Cachexia; Hypothermia; Lipocalin-2; Mice, Inbred C57BL; Mice, Knockout; Muscle, Skeletal; Pancreatic Neoplasms
PubMed: 36243318
DOI: 10.1016/j.molmet.2022.101612 -
JCI Insight Jun 2022In many solid cancers, tumor-associated macrophages (TAM) represent the predominant myeloid cell population. Antigen (Ag) cross-presentation leading to tumor Ag-directed...
In many solid cancers, tumor-associated macrophages (TAM) represent the predominant myeloid cell population. Antigen (Ag) cross-presentation leading to tumor Ag-directed cytotoxic CD8+ T cell responses is crucial for antitumor immunity. However, the role of recruited monocyte-derived macrophages, including TAM, as potential cross-presenting cells is not well understood. Here, we show that primary human as well as mouse CD206+ macrophages are effective in functional cross-presentation of soluble self-Ag and non-self-Ag, including tumor-associated Ag (TAA), as well as viral Ag. To confirm the presence of cross-presenting TAM in vivo, we performed phenotypic and functional analysis of TAM from B16-F10 and CT26 syngeneic tumor models and have identified CD11b+F4/80hiCD206+ TAM to effectively cross-present TAA. We show that CD11b+CD206+ TAM represent the dominant tumor-infiltrating myeloid cell population, expressing a unique cell surface repertoire, promoting Ag cross-presentation and Ag-specific CD8+ T cell activation comparable with cross-presenting CLEC9A+ DCs (cDC1). The presence of cross-presenting CD206+ TAM is associated with reduced tumor burden in mouse syngeneic tumor models and with improved overall survival in cutaneous melanoma patients. Therefore, the demonstration of effective Ag cross-presentation capabilities of CD206+ TAM, including their clinical relevance, expands our understanding of TAM phenotypic diversity and functional versatility.
Topics: Animals; Antigens, Neoplasm; Cross-Priming; Humans; Melanoma; Mice; Skin Neoplasms; Tumor-Associated Macrophages
PubMed: 35503656
DOI: 10.1172/jci.insight.155022 -
Biology Methods & Protocols 2022While subcutaneous tumor models remain the standard for studying drug efficacy , these tumors rarely metastasize and lack physiological relevance due to differences in...
While subcutaneous tumor models remain the standard for studying drug efficacy , these tumors rarely metastasize and lack physiological relevance due to differences in the tumor microenvironment, vascularization, immune landscape, and physiological cues associated with the organ of interest. Orthotopic tumors, grown from the organ corresponding with the cancer type, provide a more translational approach to study disease progression and drug efficacy. Utilization of a syngeneic mouse model allows for a complete immune landscape, key for adaptive immunotherapy studies. MC38 and CT26 cells are commonly used murine colorectal cancer cell lines with clinically relevant mutations. While CT26 cells have been orthotopically implanted with high fidelity, successful engraftment of orthotopic MC38 tumors varies greatly between studies. Thus, we have developed a detailed protocol for MC38 orthotopic tumor inoculation via intracecal injection. Nine C57BL/6 mice were injected with 2 × 10 cells into the cecal wall and sacrificed after 7 weeks. Survival after surgery was 100%, and one mouse died before the 7-week study end point from tumor burden and metastatic spread. We observed a successful tumor engraftment rate of 67%. Half of mice presenting with tumors were found to have macroscopic metastatic lesions in clinically relevant foci, including the mesenteric lymph nodes, liver, and peritoneum. These mice also presented with very large tumors and an enlarged spleen. The other half of the mice presented with small, localized tumors that did not metastasize. Herein, we describe tips specific for the intracecal injection of MC38 cells to improve the engraftment rate consistency in this model.
PubMed: 36225595
DOI: 10.1093/biomethods/bpac024 -
Cell Reports Nov 2018Tumor ecosystems are composed of multiple cell types that communicate by ligand-receptor interactions. Targeting ligand-receptor interactions (for instance, with immune...
Tumor ecosystems are composed of multiple cell types that communicate by ligand-receptor interactions. Targeting ligand-receptor interactions (for instance, with immune checkpoint inhibitors) can provide significant benefits for patients. However, our knowledge of which interactions occur in a tumor and how these interactions affect outcome is still limited. We present an approach to characterize communication by ligand-receptor interactions across all cell types in a microenvironment using single-cell RNA sequencing. We apply this approach to identify and compare the ligand-receptor interactions present in six syngeneic mouse tumor models. To identify interactions potentially associated with outcome, we regress interactions against phenotypic measurements of tumor growth rate. In addition, we quantify ligand-receptor interactions between T cell subsets and their relation to immune infiltration using a publicly available human melanoma dataset. Overall, this approach provides a tool for studying cell-cell interactions, their variability across tumors, and their relationship to outcome.
Topics: Animals; Cell Communication; Cell Line, Tumor; Disease Models, Animal; Ligands; Melanoma; Mice; Neoplasm Metastasis; Neoplasms; Phenotype; Receptors, Cell Surface; Sequence Analysis, RNA; Single-Cell Analysis; Tumor Microenvironment
PubMed: 30404002
DOI: 10.1016/j.celrep.2018.10.047 -
Journal For Immunotherapy of Cancer Feb 2023The immunosuppressive tumor microenvironment (TME) is a major barrier to the efficacy of chimeric antigen receptor T cells (CAR-T cells) in glioblastoma (GBM)....
INTRODUCTION
The immunosuppressive tumor microenvironment (TME) is a major barrier to the efficacy of chimeric antigen receptor T cells (CAR-T cells) in glioblastoma (GBM). Transgenic expression of IL15 is one attractive strategy to modulate the TME. However, at present, it is unclear if IL15 could be used to directly target myeloid-derived suppressor cells (MDSCs), a major cellular component of the GBM TME. Here, we explored if MDSC express IL15Rα and the feasibility of exploiting its expression as an immunotherapeutic target.
METHODS
RNA-seq, RT-qPCR, and flow cytometry were used to determine IL15Rα expression in paired peripheral and tumor-infiltrating immune cells of GBM patients and two syngeneic murine GBM models. We generated murine T cells expressing IL13Rα2-CARs and secretory IL15 (CAR.IL15s) or IL13Rα2-CARs in which IL15 was fused to the CAR to serve as an IL15Rα-targeting moiety (CAR.IL15f), and characterized their effector function in vitro and in syngeneic IL13Rα2+glioma models.
RESULTS
IL15Rα was preferentially expressed in myeloid, B, and dendritic cells in patients' and syngeneic GBMs. In vitro, CAR.IL15s and CAR.IL15f T cells depleted MDSC and decreased their secretion of immunosuppressive molecules with CAR.IL15f T cells being more efficacious. Similarly, CAR.IL15f T cells significantly improved the survival of mice in two GBM models. TME analysis showed that treatment with CAR.IL15f T cells resulted in higher frequencies of CD8+T cells, NK, and B cells, but a decrease in CD11b+cells in tumors compared with therapy with CAR T cells.
CONCLUSIONS
We demonstrate that MDSC of the glioma TME express IL15Ra and that these cells can be targeted with secretory IL15 or an IL15Rα-targeting moiety incorporated into the CAR. Thus, IL15-modified CAR T cells act as a dual targeting agent against tumor cells and MDSC in GBM, warranting their future evaluation in early-phase clinical studies.
Topics: Animals; Mice; Glioblastoma; Glioma; Interleukin-13 Receptor alpha2 Subunit; Interleukin-15; Myeloid-Derived Suppressor Cells; Tumor Microenvironment; T-Lymphocytes
PubMed: 36759014
DOI: 10.1136/jitc-2022-006239 -
BMC Cancer Nov 2021Immune checkpoint blockade (ICB) therapies have changed the paradigm of cancer therapies. However, anti-tumor response of the ICB is insufficient for many patients and...
BACKGROUND
Immune checkpoint blockade (ICB) therapies have changed the paradigm of cancer therapies. However, anti-tumor response of the ICB is insufficient for many patients and limited to specific tumor types. Despite many preclinical and clinical studies to understand the mechanism of anti-tumor efficacy of ICB, the mechanism is not completely understood. Harnessing preclinical tumor models is one way to understand the mechanism of treatment response.
METHODS
In order to delineate the mechanisms of anti-tumor activity of ICB in preclinical syngeneic tumor models, we selected two syngeneic murine colorectal cancer models based on in vivo screening for sensitivity with anti-PD-1 therapy. We performed tumor-immune profiling of the two models to identify the potential mechanism for anti-PD-1 response.
RESULTS
We performed in vivo screening for anti-PD-1 therapy across 23 syngeneic tumor models and found that CT-26 and Colon 26, which are murine colorectal carcinoma derived from BALB/c mice, showed different sensitivity to anti-PD-1. CT-26 tumor mice were more sensitive to the anti-PD-1 antibody than Colon 26, while both models show similarly sensitivity to anti-CTLA4 antibody. Immune-profiling showed that CT-26 tumor tissue was infiltrated with more immune cells than Colon 26. Genomic/transcriptomic analyses highlighted thatWnt pathway was one of the potential differences between CT-26 and Colon 26, showing Wnt activity was higher in Colon 26 than CT-26. .
CONCLUSIONS
CT-26 and Colon 26 syngeneic tumor models showed different sensitivity to anti-PD-1 therapy, although both tumor cells are murine colorectal carcinoma cell lines from BALB/c strain. By characterizing the mouse cells lines and tumor-immune context in the tumor tissues with comprehensive analysis approaches, we found that CT-26 showed "hot tumor" profile with more infiltrated immune cells than Colon 26. Further pathway analyses enable us to propose a hypothesis that Wnt pathway could be one of the major factors to differentiate CT-26 from Colon 26 model and link to anti-PD-1 response. Our approach to focus on preclinical tumor models with similar genetic background but different sensitivity to anti-PD-1 therapy would contribute to illustrating the potential mechanism of anti-PD-1 response and to generating a novel concept to synergize current anti-PD-1 therapies for cancer patients.
Topics: Animals; Base Sequence; Cell Line, Tumor; Colonic Neoplasms; Disease Models, Animal; Female; Gene Expression Profiling; Immune Checkpoint Inhibitors; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Neoplasm Transplantation; Transcriptome; Exome Sequencing; Wnt Signaling Pathway
PubMed: 34774008
DOI: 10.1186/s12885-021-08974-3 -
Cancer Immunology Research Jan 2017Murine syngeneic tumor models are critical to novel immuno-based therapy development, but the molecular and immunologic features of these models are still not clearly...
Murine syngeneic tumor models are critical to novel immuno-based therapy development, but the molecular and immunologic features of these models are still not clearly defined. The translational relevance of differences between the models is not fully understood, impeding appropriate preclinical model selection for target validation, and ultimately hindering drug development. Across a panel of commonly used murine syngeneic tumor models, we showed variable responsiveness to immunotherapies. We used array comparative genomic hybridization, whole-exome sequencing, exon microarray analysis, and flow cytometry to extensively characterize these models, which revealed striking differences that may underlie these contrasting response profiles. We identified strong differential gene expression in immune-related pathways and changes in immune cell-specific genes that suggested differences in tumor immune infiltrates between models. Further investigation using flow cytometry showed differences in both the composition and magnitude of the tumor immune infiltrates, identifying models that harbor "inflamed" and "non-inflamed" tumor immune infiltrate phenotypes. We also found that immunosuppressive cell types predominated in syngeneic mouse tumor models that did not respond to immune-checkpoint blockade, whereas cytotoxic effector immune cells were enriched in responsive models. A cytotoxic cell-rich tumor immune infiltrate has been correlated with increased efficacy of immunotherapies in the clinic, and these differences could underlie the varying response profiles to immunotherapy between the syngeneic models. This characterization highlighted the importance of extensive profiling and will enable investigators to select appropriate models to interrogate the activity of immunotherapies as well as combinations with targeted therapies in vivo Cancer Immunol Res; 5(1); 29-41. ©2016 AACR.
Topics: Animals; Antineoplastic Agents, Immunological; B7-H1 Antigen; CTLA-4 Antigen; Comparative Genomic Hybridization; DNA Copy Number Variations; Disease Models, Animal; Drug Discovery; Drug Evaluation, Preclinical; Drug Synergism; Exome; Gene Expression Regulation, Neoplastic; Genomics; High-Throughput Nucleotide Sequencing; Immunomodulation; Mice; Molecular Targeted Therapy; Mutation; Neoplasms; Signal Transduction; Transcriptome; Tumor Microenvironment
PubMed: 27923825
DOI: 10.1158/2326-6066.CIR-16-0114 -
Biochimica Et Biophysica Acta.... May 2019Cholangiocarcinoma (CCA) is an aggressive biliary tract malignancy with a poor overall prognosis. There is a critical need to develop effective targeted therapies for... (Review)
Review
Cholangiocarcinoma (CCA) is an aggressive biliary tract malignancy with a poor overall prognosis. There is a critical need to develop effective targeted therapies for the treatment of this lethal disease. In an effort to address this challenge, preclinical in vivo studies have become paramount in understanding CCA carcinogenesis, progression, and therapy. Various CCA animal models exist including carcinogen-based models in which animals develop CCA after exposure to a carcinogen, genetically engineered mouse models in which genetic changes are induced in mice leading to CCA, murine syngeneic orthotopic models, as well as xenograft tumors derived from xenotransplantation of CCA cells, organoids, and patient-derived tissue. Each type has distinct advantages as well as shortcomings. In the ideal animal model of CCA, the tumor arises from the biliary tract in an immunocompetent host with a species-matched tumor microenvironment. Such a model would also be time-efficient, recapitulate the genetic and histopathological features of human CCA, and predict therapeutic response in humans. Recently developed biliary tract transduction and orthotopic syngeneic transplant mouse models encompass several of these elements. Herein, we review the different animal models of CCA, their advantages and deficiencies, as well as features which mimic human CCA.
Topics: Animals; Bile Duct Neoplasms; Cholangiocarcinoma; Cholestasis; Disease Models, Animal; Gene Editing; Humans; Xenograft Model Antitumor Assays
PubMed: 29627364
DOI: 10.1016/j.bbadis.2018.03.026