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Ma J, Al Moussawi K, Lou H, Chan HF, Wang Y, Chadwick J, Phetsouphanh C, Slee EA, Zhong S, Leissing TM, Roth A, Qin X, Chen S, Yin J, Ratnayaka I, Hu Y, Louphrasitthiphol P, Taylor L, Bettencourt PJG, Muers M, Greaves DR, McShane H, Goldin R, Soilleux EJ, Coleman ML, Ratcliffe PJ, Lu X. Deficiency of factor-inhibiting HIF creates a tumor-promoting immune microenvironment. Proc Natl Acad Sci U S A 2024; 121:e2309957121. [PMID: 38422022 PMCID: PMC10927516 DOI: 10.1073/pnas.2309957121] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2023] [Accepted: 01/03/2024] [Indexed: 03/02/2024] Open
Abstract
Hypoxia signaling influences tumor development through both cell-intrinsic and -extrinsic pathways. Inhibiting hypoxia-inducible factor (HIF) function has recently been approved as a cancer treatment strategy. Hence, it is important to understand how regulators of HIF may affect tumor growth under physiological conditions. Here we report that in aging mice factor-inhibiting HIF (FIH), one of the most studied negative regulators of HIF, is a haploinsufficient suppressor of spontaneous B cell lymphomas, particular pulmonary B cell lymphomas. FIH deficiency alters immune composition in aged mice and creates a tumor-supportive immune environment demonstrated in syngeneic mouse tumor models. Mechanistically, FIH-defective myeloid cells acquire tumor-supportive properties in response to signals secreted by cancer cells or produced in the tumor microenvironment with enhanced arginase expression and cytokine-directed migration. Together, these data demonstrate that under physiological conditions, FIH plays a key role in maintaining immune homeostasis and can suppress tumorigenesis through a cell-extrinsic pathway.
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Affiliation(s)
- Jingyi Ma
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
- Ministry of Health Holdings, Singapore099253, Singapore
| | - Khatoun Al Moussawi
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Hantao Lou
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Hok Fung Chan
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Yihua Wang
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
- Biological Sciences, Faculty of Environmental and Life Sciences, University of Southampton, SouthamptonSO17 1BJ, United Kingdom
| | - Joseph Chadwick
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Chansavath Phetsouphanh
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
- The Kirby Institute, University of New South Wales, Kensington, NSW2052, Australia
| | - Elizabeth A. Slee
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Shan Zhong
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Thomas M. Leissing
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Andrew Roth
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
- Department of Molecular Oncology, BC Cancer, Vancouver, BCV5Z 4E6, Canada
- Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BCV6T 1Z7, Canada
- Department of Computer Science, University of British Columbia, Vancouver, BCV6T 1Z4, Canada
| | - Xiao Qin
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
- Department of Oncology, Faculty of Medical Sciences, University College London, LondonWC1E 6BT, United Kingdom
| | - Shuo Chen
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Jie Yin
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Indrika Ratnayaka
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Yang Hu
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Pakavarin Louphrasitthiphol
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Lewis Taylor
- Sir William Dunn School of Pathology, University of Oxford, OxfordOX1 3RE, United Kingdom
| | - Paulo J. G. Bettencourt
- The Jenner Institute, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
- Center for Interdisciplinary Research in Health, Faculty of Medicine, Universidade Católica Portuguesa, Lisbon1649-023, Portugal
| | - Mary Muers
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - David R. Greaves
- Sir William Dunn School of Pathology, University of Oxford, OxfordOX1 3RE, United Kingdom
| | - Helen McShane
- The Jenner Institute, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Robert Goldin
- Department of Metabolism, Digestion and Reproduction, Faculty of Medicine, Imperial College London, LondonW2 1NY, United Kingdom
| | | | - Mathew L. Coleman
- Institute of Cancer and Genomic Sciences, University of Birmingham, BirminghamB15 2TT, United Kingdom
| | - Peter J. Ratcliffe
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
| | - Xin Lu
- Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, OxfordOX3 7DQ, United Kingdom
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Neospora caninum inhibits tumor development by activating the immune response and destroying tumor cells in a B16F10 melanoma model. Parasit Vectors 2022; 15:332. [PMID: 36138417 PMCID: PMC9503190 DOI: 10.1186/s13071-022-05456-8] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2022] [Accepted: 08/29/2022] [Indexed: 11/22/2022] Open
Abstract
Background Melanoma is a malignant tumor with a high mortality rate. Some microorganisms have been shown to activate the immune system and limit cancer progression. The objective of this study is to evaluate the anti-melanoma effect of Neospora caninum, a livestock pathogen with no pathogenic activity in humans. Methods Neospora caninum tachyzoites were inoculated into a C57BL/6 mouse melanoma model by intratumoral and distal subcutaneous injections. Tumor volumes were measured, and cell death areas were visualized by hematoxylin and eosin staining and quantified. Apoptosis in cell cultures and whole tumors was detected by propidium iodide (PI) and TUNEL staining, respectively. Cytokine and tumor-associated factor levels in tumors and spleens were detected by real-time quantitative polymerase chain reaction. Infiltration of macrophages and CD8+ T cells in the tumor microenvironment (TME) were detected by immunohistochemistry with anti-CD68 and anti-CD8 antibodies, respectively. Finally, 16S rRNA sequencing of mice cecal contents was performed to evaluate the effect of N. caninum on gut microbial diversity. Results Intratumoral and distal subcutaneous injections of N. caninum resulted in significant inhibition of tumor growth (P < 0.001), and more than 50% of tumor cells were dead without signs of apoptosis. Neospora caninum treatment significantly increased the mRNA expression levels of IL-12, IFN-γ, IL-2, IL-10, TNF-α, and PD-L1 in the TME, and IL-12 and IFN-γ in the spleen of tumor-bearing mice (P < 0.05). An increase in the infiltration of CD8+ T cells and macrophages in the TME was observed with these cytokine changes. Neospora caninum also restored the abundance of gut microbiota Lactobacillus, Lachnospiraceae, Adlercreutzia, and Prevotellaceae associated with tumor growth, but the changes were not significant. Conclusion Neospora caninum inhibits B16F10 melanoma by activating potent immune responses and directly destroying the cancer cells. The stable, non-toxic, and efficacious properties of N. caninum demonstrate the potential for its use as a cancer treatment. Graphical Abstract ![]()
Supplementary Information The online version contains supplementary material available at 10.1186/s13071-022-05456-8.
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Chen S, Ma S, Wang H, Shao X, Ding B, Guo Z, Chen X, Wang Y. Unraveling the mechanism of alkaloids from Sophora alopecuroides Linn combined with immune checkpoint blockade in the treatment of non-small cell lung cancer based on systems pharmacology. Bioorg Med Chem 2022; 64:116724. [PMID: 35468537 DOI: 10.1016/j.bmc.2022.116724] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2022] [Revised: 03/07/2022] [Accepted: 03/22/2022] [Indexed: 11/26/2022]
Abstract
Quinolizidine alkaloids, as essential active ingredients extracted from Sophora alopecuroides Linn (SAL), have been proven to be pharmacologically active in a variety of cancers including non-small cell lung cancer (NSCLC). However, whether these alkaloids have substantial benefits in combination with immune checkpoint blockade (ICB) for the treatment of NSCLC is unknown. Here, we explore the potential of these alkaloids in combination with ICB therapy based on a systems pharmacology and bioinformatics approach. We found that 37 alkaloids in SAL have highly similar characteristics in the molecular skeleton, pharmacological properties, and targets. The expression of targets of these alkaloids are significantly correlated with the infiltration level of tumor infiltrating lymphocytes and the expression levels of multiple immune checkpoints in NSCLC. They share similar molecular mechanisms in antitumor immunity. Sophocarpine (Sop) is one of the most representative constituents of these alkaloids. We demonstrated that the Sop promotes PD-L1 expression to improve the effects of PD-L1 blockade treatment via the ADORA1-ATF3 axis. In conclusion, our study identified these alkaloids as promising candidates for the treatment of NSCLC, either alone or in combination with ICB, with potential value for drug development and may provide a promising strategy for improving the survival of NSCLC patients.
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Affiliation(s)
- Sen Chen
- Center of Bioinformatics, College of Life Science, Northwest A & F University, Yangling, Shaanxi 712100, China
| | - Shuangxin Ma
- Center of Bioinformatics, College of Life Science, Northwest A & F University, Yangling, Shaanxi 712100, China
| | - Haiqing Wang
- Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, School of Life Sciences, Northwest University, Xi'an, China
| | - Xuexue Shao
- Key Laboratory of Xinjiang Phytomedicine Resource and Utilization, Ministry of Education, Shihezi University, Shihezi, Xinjiang 832002, China
| | - Bojiao Ding
- Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, School of Life Sciences, Northwest University, Xi'an, China
| | - Zihu Guo
- Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, School of Life Sciences, Northwest University, Xi'an, China
| | - Xuetong Chen
- Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, School of Life Sciences, Northwest University, Xi'an, China.
| | - Yonghua Wang
- Center of Bioinformatics, College of Life Science, Northwest A & F University, Yangling, Shaanxi 712100, China; Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, School of Life Sciences, Northwest University, Xi'an, China.
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De Luca R, Neri D. Potentiation of PD-L1 blockade with a potency-matched dual cytokine-antibody fusion protein leads to cancer eradication in BALB/c-derived tumors but not in other mouse strains. Cancer Immunol Immunother 2018; 67:1381-1391. [PMID: 29971465 DOI: 10.1007/s00262-018-2194-0] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2017] [Accepted: 06/29/2018] [Indexed: 01/20/2023]
Abstract
We have recently described a novel therapeutic antibody product (IL2-F8-TNFmut), featuring the simultaneous fusion of murine IL2 and of a TNF mutant with scFv(F8), an antibody specific to the alternatively-spliced extra domain A of fibronectin (EDA). Here, we report on the in vivo characterization of the anti-cancer activity of IL2-F8-TNFmut in four immunocompetent murine models of cancer, CT26, WEHI-164, F9 teratocarcinoma and Lewis lung carcinoma (LLC), using the product alone or in combination with a monoclonal antibody specific to murine PD-L1. All four models exhibited a strong expression of EDA-fibronectin, which was confined to vascular structures for F9 tumors, while the other three malignancies exhibited a more stromal pattern of staining. A complete and long-lasting tumor eradication of CT26 and WEHI-164 tumors was observed in BALB/c mice when IL2-F8-TNFmut was used in combination with PD-L1 blockade. The combination treatment led to improved tumor growth inhibition in 129/SvEv mice bearing murine teratocarcinoma or in C57BL/6 mice bearing murine LLC, but those cancer cures were difficult to achieve in those models. A microscopic analysis of tumor sections, obtained 24 h after pharmacological treatment, revealed that the PD-L1 antibody had homogenously reached tumor cells in vivo and that the combination of PD-L1 blockade with IL2-F8-TNFmut stimulated an influx of NK cells and of T cells into the neoplastic mass. These data indicate that potency-matched dual-cytokine fusion proteins may be ideally suited to potentiate the therapeutic activity of immune check-point inhibitors.
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Affiliation(s)
- Roberto De Luca
- Department of Chemistry and Applied Biosciences, Swiss Federal Institute of Technology (ETH Zürich), Vladimir-Prelog-Weg 4, 8093, Zurich, Switzerland
| | - Dario Neri
- Department of Chemistry and Applied Biosciences, Swiss Federal Institute of Technology (ETH Zürich), Vladimir-Prelog-Weg 4, 8093, Zurich, Switzerland.
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Pan WL, Ng TB. A dimeric Phaseolus coccineus lectin with anti-oxidative, anti-proliferative and cytokine-inducing activities. Int J Biol Macromol 2015; 81:960-6. [PMID: 26410813 DOI: 10.1016/j.ijbiomac.2015.09.034] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2015] [Revised: 09/17/2015] [Accepted: 09/18/2015] [Indexed: 12/27/2022]
Abstract
In the present study, a dimeric glucosamine binding lectin, designated as CHL, was purified from Phaseolus coccineus L. var. albonanus Bailey through three chromatographic steps. The molecular weight of CHL was approximately 66kDa. Its hemagglutinating activity toward rabbit erythrocytes was dependent on carbohydrates, especially glucosamine, and was stable at temperatures between 20 and 70°C, and at pH between 1 and 13. Intriguingly, further characterization showed that CHL served as a potent antioxidant to prevent erythrocytes from haemolysis induced by 2'-azobis (2-amidinopropane) dihydrochloride (AAPH) in a dose-dependent manner. Moreover, it exerted antitumor activity against human nasopharyngeal carcinoma CNE1 cells, hepatoma HepG2 cells, and breast cancer MCF7 cells but was devoid of antifungal activity. In addition, the CHL could bring about a significant dose-dependent increase in the production of mRNAs of pro-inflammatory cytokines including interferon-gamma and interleukin-2. These results suggest the potential therapeutic utility of CHL.
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Affiliation(s)
- Wen Liang Pan
- School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, China
| | - Tzi Bun Ng
- School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong, China.
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Ren G, Tian G, Liu Y, He J, Gao X, Yu Y, Liu X, Zhang X, Sun T, Liu S, Yin J, Li D. Recombinant Newcastle Disease Virus Encoding IL-12 and/or IL-2 as Potential Candidate for Hepatoma Carcinoma Therapy. Technol Cancer Res Treat 2015; 15:NP83-94. [PMID: 26303327 DOI: 10.1177/1533034615601521] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2015] [Accepted: 07/01/2015] [Indexed: 11/15/2022] Open
Abstract
Interleukins as immunomodulators are promising therapeutic agents for cancer therapy. Previous studies showed that there was an improved antitumor immunity in tumor-bearing mice using recombinant Newcastle disease virus carrying for interleukin-2. Interleukin-12 is a promising antitumor cytokine too. So we investigated and compared the antitumor effect of genetically engineered Newcastle disease virus strains expressing both interleukin-12 and/or interleukin-2 (rClone30-interleukin-2, rClone30-interleukin-12, and rClone30-interleukin-12-interleukin-2). In vitro studies showed that rClone30s could efficiently infect tumor cells and express interleukin-12 and/or interleukin-2. 3-(4,5-Dimethylthiazol-2-y)-2,5-diphenyl-tetrazolium bromide results showed rClone30s possessed strong cytotoxic activities against multiple tumor cell lines (U251, HepG2, A549, and Hela). Animal studies showed that rClone30-interleukin-12-interleukin-2 was more effective in inhibition of murine hepatoma carcinoma tumors, with the mean tumor volume (day 14) of 141.70 mm(3) comparing 165.67 mm(3) of rClone30-interleukin-12 group, 210.47 mm(3) of rClone30-interleukin-2 group, 574.70 mm(3) of rClone30 group, and 1206.83 mm(3) of phosphate-buffered saline group. Moreover, the rClone30-interleukin-12-interleukin-2 treated mice secreted more interferon γ (333.518 pg/mL) and its downstream cytokine interferon-γ induced protein 10 (16.006 pg/mL) in tumor than the rClone30-interleukin-12 group (interferon γ: 257.548 pg/mL; interferon-γ induced protein 10: 13.601 pg/mL), rClone30-interleukin2 group (interferon γ: 124.601 pg/mL; interferon-γ induced protein 10: 9.779 pg/mL), or rClone30 group (interferon γ: 48.630 pg/mL; interferon-γ induced protein 10:1.650 pg/mL). For the survival study, rClone30-interleukin12-interleukin2 increased the survival rate (12 of 16) of the tumor-bearing mice versus 11 of 16 in rClone30-interleukin-12 group, 10 of 16 in rClone30-interleukin-2 group, 7 of 16 in Clone30 group, and 0/16 in phosphate-buffered saline group, respectively. To determine whether the mice treated with recombinant virus developed protective immune response, the mice were rechallenged with the same tumor cells. The results showed that viral-treated mice were significantly protected from rechallenge. These results suggest that expressing both interleukin-2 and/or interleukin-12 could be ideal approaches to enhance the antitumor ability of Newcastle disease virus, and rClone30-interleukin-12-interleukin-2 is slightly superior over rClone30-interleukin-12 and rClone30-interleukin-2 alone.
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Affiliation(s)
- Guiping Ren
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China Key Laboratory of Agricultural Biological Functional Gene, Northeast Agricultural University, Harbin, China
| | - Guiyou Tian
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Yunye Liu
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Jinjiao He
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Xinyu Gao
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Yinhang Yu
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Xin Liu
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Xu Zhang
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Tian Sun
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Shuangqing Liu
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Jiechao Yin
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China
| | - Deshan Li
- College of Life Science, Northeast Agricultural University, Xiangfang District, Harbin, China Key Laboratory of Agricultural Biological Functional Gene, Northeast Agricultural University, Harbin, China
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A new Phaseolus vulgaris lectin induces selective toxicity on human liver carcinoma Hep G2 cells. Arch Toxicol 2011; 85:1551-63. [PMID: 21445585 DOI: 10.1007/s00204-011-0698-x] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/19/2011] [Accepted: 03/14/2011] [Indexed: 01/10/2023]
Abstract
We describe here the purification and characterization of a new Phaseolus vulgaris lectin that exhibits selective toxicity to human hepatoma Hep G2 cells and lacks significant toxicity on normal liver WRL 68 cells. This polygalacturonic acid-specific lectin (termed BTKL) was purified from seeds of P. vulgaris cv. Blue tiger king by liquid chromatography techniques. The 60-kDa dimeric lectin showed strong and broad-spectrum hemagglutinating activity toward human, rabbit, rat, and mouse erythrocytes. Bioinformatic analysis unveils substantial N-terminal sequence similarity of BTKL to other Phaseolus lectins. Among a number of tumor cells tested, BTKL exhibits potent anti-Hep G2 activity which is associated with (1) induction of DNA fragmentation, (2) production of apoptotic bodies and chromatin condensation, (3) triggering of cell apoptosis and necrosis, and (4) depolarization of mitochondrial membrane (low ΔΨm). Furthermore, BTKL could induce inducible nitric oxide synthase (iNOS) expression and subsequent nitric oxide production in vitro in mouse macrophages, which may contribute to its antitumor activity. In addition, BTKL could bring about a significant dose-dependent increase in the production of mRNAs of proinflammatory cytokines including interleukin-1 beta, interleukin-2, tumor necrosis factor alpha, and interferon-gamma. In sum, the antitumor activity and mechanism of BTKL provided here suggest that it has potential therapeutic value for human liver cancer.
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Tagalakis AD, Grosse SM, Meng QH, Mustapa MFM, Kwok A, Salehi SE, Tabor AB, Hailes HC, Hart SL. Integrin-targeted nanocomplexes for tumour specific delivery and therapy by systemic administration. Biomaterials 2010; 32:1370-6. [PMID: 21074847 DOI: 10.1016/j.biomaterials.2010.10.037] [Citation(s) in RCA: 49] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2010] [Accepted: 10/15/2010] [Indexed: 11/30/2022]
Abstract
Nanoparticle formulations offer opportunities for tumour delivery of therapeutic reagents. The Receptor-Targeted Nanocomplex (RTN) formulation consists of a PEGylated, endosomally-cleavable lipid and an RGD integrin-targeting, endosomally-cleavable peptide. Nancomplexes self-assemble on mixing with plasmid DNA to produce nanoparticles of about 100 nm. The environmentally-sensitive linkers promote intracellular disassembly and release of the DNA. RTNs carrying luciferase genes were administered intravenously to mice carrying subcutaneous neuroblastoma tumours. Luciferase expression was much higher in tumours than in liver, spleen and lungs while plasmid biodistribution studies supported the expression data. Transfection in tumours was enhanced two-fold by integrin-targeting peptides compared to non-targeted nanocomplexes. RTNs containing the interleukin-2 (IL-2) and IL-12 genes were administered intravenously with seven doses at 48 h intervals and tumour growth monitored. Tumours from treated animals were approximately 75% smaller on day 11 compared with RTNs containing control plasmids with one third of treated mice surviving long-term. Extensive leukocyte infiltration, decreased vascularization and increased necrotic areas were observed in the tumours from IL2/IL12 treated animals. Splenocytes from re-challenged mice displayed enhanced IL-2 production following Neuro-2A co-culture, which, combined with infiltration studies, suggested a cytotoxic T cell-mediated9 tumour-rejection process. The integrin-targeted RTN formulation may have broader applications in the further development of cancer therapeutics.
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Affiliation(s)
- Aristides D Tagalakis
- Molecular Immunology Unit, UCL Institute of Child Health, University College London, London, UK
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Fang EF, Wong JH, Bah CSF, Lin P, Tsao SW, Ng TB. Bauhinia variegata var. variegata trypsin inhibitor: From isolation to potential medicinal applications. Biochem Biophys Res Commun 2010; 396:806-11. [DOI: 10.1016/j.bbrc.2010.04.140] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2010] [Accepted: 04/27/2010] [Indexed: 10/19/2022]
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Kim JO, Jung SS, Kim SY, Kim TY, Shin DW, Lee JH, Lee YH. Inhibition of Lewis lung carcinoma growth by Toxoplasma gondii through induction of Th1 immune responses and inhibition of angiogenesis. J Korean Med Sci 2007; 22 Suppl:S38-46. [PMID: 17923753 PMCID: PMC2694397 DOI: 10.3346/jkms.2007.22.s.s38] [Citation(s) in RCA: 58] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/02/2022] Open
Abstract
Toxoplasma gondii is an obligate intracellular protozoan parasite that induces antitumor activity against certain types of cancers. However, little information is available regarding the immunologic mechanisms that regulate these effects. For this purpose, C57BL/6 mice were administered either the T. gondii Me49 strain orally or Lewis lung carcinoma (LLC) cells intramuscularly. Survival rates, tumor size, histopathology, and immune responses were determined for each group, and angiogenesis was evaluated by in vivo Matrigel plug assay. Toxoplasma-infected (TG-injected) mice survived the entire experimental period, whereas cancer cell-bearing (LLC-injected) mice died within six weeks. Mice injected with both T. gondii and cancer cells (TG/LLC-injected group) showed significantly increased survival rates, CD8+ T-cell percentages, IFN-gamma mRNA expression levels, serum IgG2a titers, and CTL responses as compared to the LLC-injected mice. In addition, angiogenesis in the TG/LLC-injected mice was notably inhibited. These effects in TG/LCC-injected mice were similar or were increased by the addition of an adjuvant, Quil-A. However, TG/LLC-injected mice showed decreased percentages of CD4+ and CD8+ T cells, IFN-gamma mRNA expression levels, and serum IgG1 and IgG2a titers as compared to TG-injected mice. Taken together, our results demonstrate that T. gondii infection inhibits tumor growth in the Lewis lung carcinoma mouse model through the induction of Th1 immune responses and antiangiogenic activity.
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MESH Headings
- Animals
- Base Sequence
- CD4-Positive T-Lymphocytes/immunology
- CD8-Positive T-Lymphocytes/immunology
- Carcinoma, Lewis Lung/blood supply
- Carcinoma, Lewis Lung/genetics
- Carcinoma, Lewis Lung/immunology
- Carcinoma, Lewis Lung/therapy
- Cell Line, Tumor
- Cytotoxicity, Immunologic
- DNA Primers/genetics
- Female
- Immunoglobulin G/blood
- Immunotherapy/methods
- In Vitro Techniques
- Interferon-gamma/genetics
- Mice
- Mice, Inbred C57BL
- Neovascularization, Pathologic
- RNA, Messenger/genetics
- RNA, Messenger/metabolism
- Th1 Cells/immunology
- Toxoplasma/immunology
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Affiliation(s)
- Ju-Ock Kim
- Department of Internal Medicine, College of Medicine, Chungnam National University, Daejeon, Korea
| | - Sung-Soo Jung
- Department of Internal Medicine, College of Medicine, Chungnam National University, Daejeon, Korea
| | - Sun-Young Kim
- Department of Internal Medicine, College of Medicine, Chungnam National University, Daejeon, Korea
| | - Tae Yun Kim
- Department of Infection Biology, College of Medicine, Chungnam National University, Daejeon, Korea
| | - Dae-Whan Shin
- Department of Infection Biology, College of Medicine, Chungnam National University, Daejeon, Korea
- Research Institute for Medical Science, Chungnam National University, Daejeon, Korea
| | - Jae-Ho Lee
- Department of Pediatrics, College of Medicine, Chungnam National University, Daejeon, Korea
| | - Young-Ha Lee
- Department of Infection Biology, College of Medicine, Chungnam National University, Daejeon, Korea
- Research Institute for Medical Science, Chungnam National University, Daejeon, Korea
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Zhang J, Wang Q, Zhao D, Cao X. Induction of potent anti-tumor immunity by direct injection of Ad-LIGHT at the site of tumor inoculation. Cytotherapy 2007; 9:386-96. [PMID: 17573614 DOI: 10.1080/14653240701326749] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
BACKGROUND The aim of this study was to observe the therapeutic effects of adenovirus-mediated LIGHT gene transfer in murine B16 melanoma in vivo. METHODS C57BL/6 mice were inoculated subcutaneously with B16 cells to establish the murine melanoma model. The tumor-bearing mice were injected at the site of tumor inoculation with recombinant adenoviral vectors expressing the murine LIGHT gene. The tumor growth and survival period of tumor-bearing mice were observed. The splenic NK and CTL activity were measured in vitro by lactate dehydrogenase (LDH) release assay. The amounts of cytokines were determined with ELISA kits. RESULTS The LIGHT gene could be efficiently transduced into tumor tissue after injection of Ad-LIGHT. Treatment with Ad-LIGHT significantly inhibited the tumor growth and prolonged the survival period of the tumor-bearing mice. The splenic NK and CTL activity of the mice was also enhanced after LIGHT gene transfer. The production of IL-2 and IFN-gamma from lymphocytes derived from mice treated with Ad-LIGHT was increased significantly compared with control groups. DISCUSSION Our results indicate that local expression of the LIGHT gene can induce potent anti-tumor immunity and may be a promising treatment strategy for melanoma.
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Affiliation(s)
- J Zhang
- Department of Medical Microbiology and Immunology, Medical School, Shaoxing University, Shaoxing, P. R. China.
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12
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Alves A, Vibert E, Trajcevski S, Solly S, Fabre M, Soubrane O, Qian C, Prieto J, Klatzmann D, Panis Y. Adjuvant interleukin-12 gene therapy for the management of colorectal liver metastases. Cancer Gene Ther 2005; 11:782-9. [PMID: 15472716 DOI: 10.1038/sj.cgt.7700760] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
In humans, no efficient treatment exists not only against multifocal liver metastases (LM) but also against recurrent microscopic liver metastases within the liver remnant following curative liver resection. Furthermore, in nonmultifocal LM, partial liver resection could be performed, but in more than 50% of the patients, tumor recurrence within liver remnant is observed, partly due to the growth of dormant cancer cells in the setting of postoperative host immune dysfunction. We investigated the therapeutic potential of interleukin-12 (IL-12) immuno-gene therapies in these experimental models under total vascular exclusion (TVE) of the liver. In rats with multiple LM of DHDK12 colon cancer cells, we observed a significant reduction in tumor volume after retroviral-mediated gene transfer of either herpes simplex virus thymidine kinase (HSV1-TK) and ganciclovir (GCV) administration, or IL-12. Combined treatment with HSV1-TK/GCV and IL-12 resulted in improved tumor volume reduction and even survival. In rats with recurrent microscopic DHDK12 LM established after partial liver resection, we observed significantly decreased recurrent tumor volumes and increased survival after retroviral-mediated IL-12 gene transfer. In both settings, immunohistological analysis revealed that IL-12 immuno-gene therapy was accompanied by an infiltration of CD8+ T lymphocytes within the tumors. Altogether, our results suggest that IL-12 adjuvant gene therapy could improve the management of patients with either resectable or unresectable LM.
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Affiliation(s)
- Arnaud Alves
- Laboratory of Biology and Therapeutic of Immune Diseases, University Pierre and Marie Curie, CNRS UMR7087, Pitié-Salpétrière Hospital, Paris, France
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13
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Suzuki T, Fukuhara T, Tanaka M, Nakamura A, Akiyama K, Sakakibara T, Koinuma D, Kikuchi T, Tazawa R, Maemondo M, Hagiwara K, Saijo Y, Nukiwa T. Vaccination of Dendritic Cells Loaded with Interleukin-12-Secreting Cancer Cells Augments In vivo Antitumor Immunity: Characteristics of Syngeneic and Allogeneic Antigen-Presenting Cell Cancer Hybrid Cells. Clin Cancer Res 2005. [DOI: 10.1158/1078-0432.58.11.1] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Abstract
Cancer immunotherapy by fusion of antigen-presenting cells and tumor cells has been shown to induce potent antitumor immunity. In this study, we characterized syngeneic and allogeneic, murine macrophage/dendritic cell (DC)-cancer fusion cells for the antitumor effects. The results showed the superiority of allogeneic cells as fusion partners in both types of antigen-presenting cells in an in vivo immunotherapy model. A potent induction of tumor-specific CTLs was observed in these immunized conditions. In addition, the immunization with DC-cancer fusion cells was better than that with macrophage-cancer fusion cells. Both syngeneic and allogeneic DC-cancer fusion cells induced higher levels of IFN-γ production than macrophage-cancer fusion cells. Interestingly, allogeneic DC-cancer fusion cells were superior in that they efficiently induced Th1-type cytokines but not the Th2-type cytokines interleukin (IL)-10 and IL-4, whereas syngeneic DC-cancer fusion cells were powerful inducers of both Th1 and Th2 cytokines. These results suggest that allogeneic DCs are suitable as fusion cells in cancer immunotherapy. To further enhance the antitumor immunity in the clinical setting, we prepared DCs fused with IL-12 gene-transferred cancer cells and thus generated IL-12-secreting DC-cancer fusion cells. Immunization with these gene-modified DC-cancer fusion cells was able to elicit a markedly enhanced antitumor effect in the in vivo therapeutic model. This novel IL-12-producing fusion cell vaccine might be one promising intervention for future cancer immunotherapy.
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Affiliation(s)
- Takuji Suzuki
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Tatsuro Fukuhara
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Masashi Tanaka
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Akira Nakamura
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Kenichi Akiyama
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Tomohiro Sakakibara
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Daizo Koinuma
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Toshiaki Kikuchi
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Ryushi Tazawa
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Makoto Maemondo
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
| | - Koichi Hagiwara
- 3Department of Respiratory Medicine, Saitama Medical School, Saitama, Japan
| | - Yasuo Saijo
- 2Department of Molecular Medicine and Gene Transfer Research, Graduate School of Medicine, Tohoku University, Sendai, Japan; and
| | - Toshihiro Nukiwa
- 1Department of Respiratory Oncology and Molecular Medicine, Institute of Development, Aging, and Cancer
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14
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Ebert O, Wilbert D, Buttgereit P, Ziske C, Flieger D, Schmidt-Wolf IGH. Effects of recombinant adenovirus-mediated expression of IL-2 and IL-12 in human B lymphoma cells on co-cultured PBMC. GENETIC VACCINES AND THERAPY 2004; 2:15. [PMID: 15485577 PMCID: PMC526758 DOI: 10.1186/1479-0556-2-15] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/28/2004] [Accepted: 10/14/2004] [Indexed: 11/10/2022]
Abstract
Background Modulation of the immune system by genetically modified lymphoma cell vaccines is of potential therapeutic value in the treatment of B cell lymphoma. However, the anti-tumor effect of any single immunogene transfer has so far been limited. Combination treatment of recombinant IL-2 and IL-12 has been reported to be synergistic for inducing anti-tumor responses in solid tumors but the potential of IL-2/IL-12 gene modified B cell lymphoma cells has not been explored yet. Methods Using three different human B cell lymphoma cell lines and primary samples from patients with B cell neoplasms, expression levels of the coxsackie B-adenovirus receptor (CAR) and alpha (v) integrins were analyzed by fluorescence-activated cell sorter (FACS). Adenoviral transduction efficiencies were determined by GFP expression analysis and IL-2 and IL-12 cytokine production was quantified by enzyme-linked immunosorbent (ELISA) assays. Proliferative activities of peripheral blood mononuclear cells (PBMC) stimulated with either cytokine derived from supernatants of transduced lymphoma cells were measured by cell proliferation (MTT) assays. An EuTDA cytotoxicity assay was used to compare cytotoxic activities of IL-2 and/or IL-12 stimulated PBMC against unmodified lymphoma cells. Results We found that B cell lymphoma cell lines could be transduced with much higher efficiency than primary tumor samples, which appeared to correlate with the expression of CAR. Adenoviral-expressed IL-2 and IL-12 similarly led to dose-dependent increases in proliferation rates of PBMC obtained from healthy donors. IL-2 and/or IL-12 transduced lymphoma cells were co-cultured with PBMC, which were assayed for their cytolytic activity against unmodified lymphoma cells. We found that IL-2 stimulated PBMC elicited a significant anti-tumor effect but not the combined effect of IL-2/IL-12 or IL-12 alone. Conclusion This study demonstrates that the generation of recombinant adenovirus modified lymphoma cell vaccines based on lymphoma cell lines expressing IL-2 and IL-12 cytokine genes is technically feasible, induces increases in proliferation rates and cytotoxic activity of co-cultured PBMC, and warrants further development for the treatment of lymphoma patients in the future.
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Affiliation(s)
- Oliver Ebert
- Medizinische Klinik und Poliklinik I, Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
- Department of Gene and Cell Medicine, Mount Sinai School of Medicine, New York, New York, USA
| | - Dorothee Wilbert
- Medizinische Klinik und Poliklinik I, Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
| | - Peter Buttgereit
- Medizinische Klinik und Poliklinik I, Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
| | - Carsten Ziske
- Medizinische Klinik und Poliklinik I, Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
| | | | - Ingo GH Schmidt-Wolf
- Medizinische Klinik und Poliklinik I, Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
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15
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Alatrash G, Hutson TE, Molto L, Richmond A, Nemec C, Mekhail T, Elson P, Tannenbaum C, Olencki T, Finke J, Bukowski RM. Clinical and immunologic effects of subcutaneously administered interleukin-12 and interferon alfa-2b: phase I trial of patients with metastatic renal cell carcinoma or malignant melanoma. J Clin Oncol 2004; 22:2891-900. [PMID: 15254058 DOI: 10.1200/jco.2004.10.045] [Citation(s) in RCA: 58] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
PURPOSE Interleukin-12 (IL-12) and interferon alfa-2b (IFN-alpha-2b) are pleiotropic cytokines with activity in renal cell carcinoma (RCC) and malignant melanoma (MM) as single agents. Preclinical studies suggest concurrent administration may have synergistic antitumor effects. We conducted a phase I trial of concurrent subcutaneous (SC) administration of IL-12 and IFN-alpha-2b in patients with metastatic RCC or MM to determine toxicity, maximum-tolerated dose, preliminary efficacy, and effects on chemokine/cytokine gene expression in peripheral blood mononuclear cells (PBMCs). PATIENTS AND METHODS Cohorts of three to six patients were treated with escalating doses of IL-12 (dose I, 100 ng/kg; dose II, 300 ng/kg; dose III, 500 ng/kg; dose IV, 500 ng/kg SC) given twice weekly and IFN-alpha-2b (dose I, 1.0 MU/m(2); dose II, 1.0 MU/m(2); dose III, 1.0 MU/m(2); dose IV, 3.0 MU/m(2) SC) three times weekly in 4-week cycles. Effects on gene expression were assessed by reverse transcriptase polymerase chain reaction. RESULTS Twenty-six patients (19 with RCC, seven with MM) were accrued at dose levels I (n = 3), II (n = 3), III (n = 13), and IV (n = 7). Dose-limiting toxicity included grades 3 and 4 hepatotoxicity and neutropenia/leukopenia. Patients received a median of three cycles of treatment. Two patients with RCC and one patient with MM had partial responses. Median survival was 13.8 months. Reverse transcriptase polymerase chain reaction on PBMCs revealed induction of IP-10, Mig, B7.1 (CD80), interleukin-5, and interferon gamma in selected patients. CONCLUSION Concurrent SC administration of IL-12 and IFN-alpha-2b is possible at the dose levels utilized. Recommended doses for phase II trials are 500 ng/kg IL-12 and 1.0 MU/m(2) IFN-alpha-2b. Consistent induction of IP-10 and Mig, as well as variable induction of B7.1, interleukin-5, and interferon gamma expression was noted in PBMCs.
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Affiliation(s)
- Gheath Alatrash
- Experimental Therapeutics Program, Department of Hematology and Medical Oncology, Taussig Cancer Center, The Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, OH 44195, USA
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16
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Nagaraj S, Ziske C, Schmidt-Wolf IGH. Human cytokine-induced killer cells have enhanced in vitro cytolytic activity via non-viral interleukin-2 gene transfer. GENETIC VACCINES AND THERAPY 2004; 2:12. [PMID: 15329148 PMCID: PMC516021 DOI: 10.1186/1479-0556-2-12] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/11/2004] [Accepted: 08/25/2004] [Indexed: 11/21/2022]
Abstract
Modulation of the immune system by genetically modified immunological effector cells is of potential therapeutic value in the treatment of malignancies. Interleukin-2 (IL-2) is a crucial cytokine which induces potent antitumor response. Cytokine-induced killer cells (CIK) have been described as highly efficient cytotoxic effector cells capable of lysing tumor cell targets and are capable of recognizing these cells in a non-MHC restricted fashion. Dendritic cells (DC) are the major antigen presenting cells. This study evaluated the antitumor effect of CIK cells which were non-virally transfected with IL-2 and co-cultured with pulsed and unpulsed DC. Human CIK cells generated from peripheral blood were transfected in vitro with plasmid encoding for the human IL-2. Transfection involved a combination of electrical parameters and a specific solution to deliver plasmid directly to the cell nucleus by using the Nucleofector® electroporation system. Nucleofection resulted in the production of IL-2 with a mean of 478.5 pg/106 cells (range of 107.6–1079.3 pg /106 cells/24 h) compared to mock transfected CIK cells (31 pg/106 cells) (P = 0.05). After co-culturing with DC their functional ability was assessed in vitro by a cytotoxicity assay. On comparison with non-transfected CIK cells co-cultured with DCs (36.5 ± 5.3 %), transfected CIK cells co-cultured with DC had a significantly higher lytic activity of 58.5 ± 3.2% (P = 0.03) against Dan G cells, a human pancreatic carcinoma cell line.
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Affiliation(s)
- Srinivas Nagaraj
- Department of Internal Medicine I, General Internal Medicine Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
| | - Carsten Ziske
- Department of Internal Medicine I, General Internal Medicine Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
| | - Ingo GH Schmidt-Wolf
- Department of Internal Medicine I, General Internal Medicine Rheinische Friedrich-Wilhelms-Universität, Bonn, Germany
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17
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You TG, Wang HS, Yang JH, Qian QJ, Fan RF, Wu MC. Transfection of IL-2 and/or IL-12 genes into spleen in treatment of rat liver cancer. World J Gastroenterol 2004; 10:2190-4. [PMID: 15259063 PMCID: PMC4724966 DOI: 10.3748/wjg.v10.i15.2190] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
AIM: To test the efficacy of gene therapy in rat liver tumor.
METHODS: A retroviral vector GCIL12EIL2PN encoding human IL-2 (hIL-2) and mouse IL-12 (mIL-12) fused gene and its packaging cell were constructed. The packaging cell lines contained of IL-2 and/or IL-12 genes were injected intrasplenically to transfect splenocyte at different time. The therapeutic effect, immune function and toxic effect were evaluated.
RESULTS: The average survival times of the 4 groups using IL genes at days 1, 3, 5 and 7 after tumor implantation were 53.3 ± 3.7, 49.3 ± 4.2, 31.0 ± 2.1 and 24.3 ± 1.4 d respectively in IL-2/IL-12 fused gene group, 25.0 ± 2.5, 23.5 ± 2.0, 18.3 ± 2.4 and 12.0 ± 1.8 d respectively in IL-2 gene treatment group, and 39.0 ± 4.8, 32.0 ± 3.9, 23.0 ± 2.5 and 19.4 ± 2.1 d respectively in IL-12 gene treatment group (P < 0.01, n = 10). In the IL-12/IL-2 fused gene treatment group, 30% of rats treated at days 1 and 3 survived more than 60 d and serum mIL-12 and hIL-2 levels were still high at day 3 after treatment. Compared with IL alone, NK cell activity was strongly stimulated by IL-2/IL-12 gene. Microscopy showed that livers were infiltrated by a number of lymphocytes.
CONCLUSION: IL-2 and/or IL-12 genes injected directly into spleen increase serum IL-2 and IL-12 levels and enhance the NK cell activity, which may inhibit the liver tumor growth. The therapy of fused gene IL-2/IL-12 is of low toxicity and relatively high NK cell activity. Our data suggest that IL-2/IL-12 fused gene may be a safe and efficient gene therapy for liver tumor. The gene therapy should be administrated as early as possible.
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Affiliation(s)
- Tian-Geng You
- Department of Comprehensive Treatment III, Eastern Hepatobiliary Hospital, Second Military Medical University, Changhai Road 225, Shanghai 200433, China
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18
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O'Donnell MA, Luo Y, Hunter SE, Chen X, Hayes LL, Clinton SK. Interleukin-12 immunotherapy of murine transitional cell carcinoma of the bladder: dose dependent tumor eradication and generation of protective immunity. J Urol 2004; 171:1330-5. [PMID: 14767343 DOI: 10.1097/01.ju.0000109742.88380.a2] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023]
Abstract
PURPOSE The antitumor activity of interleukin (IL)-12 has been demonstrated in a number of tumor models but barely tested in bladder cancer models. We evaluated the antibladder cancer activity of this cytokine in syngeneic mice bearing subcutaneous, metastatic and orthotopic tumors. MATERIALS AND METHODS Mice were implanted subcutaneously, intravenously or orthotopically with syngeneic transitional cell carcinoma (TCC) of the bladder. The tumor bearing mice were then treated with IL-12 locally or systemically and monitored for tumor regression and survival. RESULTS In the subcutaneous model dose dependent suppression of tumorigenesis was observed when IL-12 was administered subcutaneously at a distal site with the MB49 line being more sensitive than MBT-2. IL-12 (10 days) above 50 ng daily was tumor inhibitory, while doses of 500 or 1000 ng daily prolonged survival and cured 70% and 75% of subjects, respectively. Upon re-challenge with parental tumor cells mice previously cured with IL-12 (1000 vs 500 ng daily) exhibited specific protection (70% vs 35% rejection) that was dependent on the earlier dose of cytokine. IL-12 administered intraperitoneally at a dose of 250 ng daily was more potent than subcutaneous administration and complete regression was observed. Metastatic TCC in the lungs and orthotopic tumors in the bladder also favorably responded to systemic or intravesical IL-12 therapy, respectively. Addition of IL-2 to IL-12 therapy increased tumor regression, long-term survival and rejection of re-challenged parental tumor. CONCLUSIONS IL-12 is exceptionally effective for treating murine bladder TCC in subcutaneous, metastatic and orthotopic models. The antibladder cancer activity of this cytokine should be tested in human bladder cancer therapy.
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Affiliation(s)
- Michael A O'Donnell
- Department of Urology, University of Iowa Hospitals and Clinics, Iowa City, Iowa 52242, USA.
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19
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Siapati KE, Barker S, Kinnon C, Michalski A, Anderson R, Brickell P, Thrasher AJ, Hart SL. Improved antitumour immunity in murine neuroblastoma using a combination of IL-2 and IL-12. Br J Cancer 2003; 88:1641-8. [PMID: 12771934 PMCID: PMC2377114 DOI: 10.1038/sj.bjc.6600928] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023] Open
Abstract
Neuroblastoma immunotherapy using cytokine-modified tumour cells has been tested in clinical trials. However, because of the complex nature of antitumour immune responses, a number of therapies may be required for complete tumour eradication and generation of systemic immunity. We report here the improved antitumour effect of two cytokines, interleukin-2 (IL-2) and interleukin-12 (IL-12), when coexpressed by neuroblastoma cell lines. Initially, transfection of human and mouse neuroblastoma cell lines resulted in high expression levels of biologically active IL-2 and IL-12 in vitro. These cytokines when expressed by transfected Neuro-2A cells completely abolished their in vivo tumorigenicity in a syngeneic neuroblastoma model. Vaccination of established tumours with IL-12-producing cells exhibited a clear effect with reduced tumour growth in the presence of IL-2. In vivo depletion studies showed that CD4(+) and CD8(+) T cells mediate the response against cytokine-producing cells. These results suggest that IL-2 and IL-12, when cotransfected in tumour cells, are effective against established disease and provide a promising immunotherapeutic approach for the treatment of neuroblastoma.
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Affiliation(s)
- K E Siapati
- Molecular Immunology Unit, Institute of Child Health, London, UK
| | - S Barker
- Molecular Immunology Unit, Institute of Child Health, London, UK
| | - C Kinnon
- Molecular Immunology Unit, Institute of Child Health, London, UK
| | - A Michalski
- Oncology and Haematology Department, Great Ormond Street Hospital for Children NHS Trust, London, UK
| | - R Anderson
- Department of Haematology, Royal Free Hospital School of Medicine, London, UK
| | - P Brickell
- Oncology and Haematology Department, Great Ormond Street Hospital for Children NHS Trust, London, UK
| | - A J Thrasher
- Molecular Immunology Unit, Institute of Child Health, London, UK
| | - S L Hart
- Molecular Immunology Unit, Institute of Child Health, London, UK
- Molecular Immunology Unit, Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK. E-mail:
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20
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Yockman JW, Maheshwari A, Han SO, Kim SW. Tumor regression by repeated intratumoral delivery of water soluble lipopolymers/p2CMVmIL-12 complexes. J Control Release 2003; 87:177-86. [PMID: 12618034 DOI: 10.1016/s0168-3659(02)00362-0] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
The recruitment of the body's own immune system is amongst the most potent defenses known against cancer. Recent attempts to harness this response have enlisted the use of the immune modulating cytokine, interleukin-12 (IL-12). The objective of this work is to investigate the organ distribution and anti-tumor response in vivo after intratumoral administration of IL-12 expression plasmid complexed with water soluble lipopolymer (WSLP). Formulations of WSLP/p2CMVmIL-12 at N/P mol ratio of 20:1 were prepared in the presence of 5% (w/v) glucose. Organ distribution data following intratumoral injection of CT-26 subcutaneous tumor-bearing BALB/c mice demonstrated enhanced retention of WSLP/p2CMVmIL-12 complexes within the tumor and limited accumulation in other organs for up to 96 h. Tumor-bearing BALB/c mice received either single or repeated intratumoral injections at 4- or 8-day intervals to examine the efficacy of single versus repeated injections on tumor regression and survival. Significant tumor growth inhibition during 4- and 8-day injection trials was observed with maximal survival in mice receiving 4-day injections of WSLP/p2CMVmIL-12 complexes. In conclusion, the water-soluble non-toxic lipopolymer complexed with p2CMVIL-12 showed enhanced transgene expression in vivo, inhibits the rate of tumor growth, and significantly increases survival.
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Affiliation(s)
- James W Yockman
- Department of Pharmaceutics and Pharmaceutical Chemistry, Center for Controlled Chemical Delivery, University of Utah, BPRB, Room 205, Salt Lake City, UT 84112, USA
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21
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Bartlett EJ, Cull VS, Brekalo NL, Lenzo JC, James CM. Synergy of type I interferon-A6 and interferon-B naked DNA immunotherapy for cytomegalovirus infection. Immunol Cell Biol 2002; 80:425-35. [PMID: 12225378 DOI: 10.1046/j.1440-1711.2002.01103.x] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Delivery of type I IFN transgenes by naked DNA immunization can protect against cytomegalovirus infection and myocarditis. Here, we investigate IFN transgene expression, antiviral efficacy, and immunomodulation of myocarditis using various treatment regimes in a mouse CMV model. In vivo expression of the IFN transgene was observed in the sera for 35 days post-DNA inoculation. Prophylactic IFN-A6 and IFN-B DNA treatment for 14 days prior to murine cytomegalovirus (MCMV) infection was more efficacious in significantly reducing viral titres, than 2 days prior to or 2 days post-virus infection. Similarly, IFN-A6 DNA treatment commencing 14 days prior to virus infection was superior in suppressing both acute and chronic myocarditis. Furthermore, reduction of autoantibody titres was more pronounced when IFN was administered 14 days prior to viral infection. Combinational IFN gene therapy was assessed for synergy between IFN subtypes. Combination treatment with either IFN-A6/A9 or IFN-A6/B greatly reduced spleen viral titres while IFN-A6/B and IFN-A9/B reduced virus replication in the liver. Only IFN-A6/A9 and IFN-A9/B reduced acute viral myocarditis, whereas IFNA6/B treatment was most efficacious for autoimmune chronic myocarditis. Finally, treatment with IFN-A6 DNA 2 weeks post-MCMV infection proved effective at inhibiting the development of chronic autoimmune myocarditis. These findings suggest that immunomodulation of both antiviral and autoimmune responses by IFN DNA immunization may be an avenue for improved viral immunotherapy.
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Affiliation(s)
- Emmalene J Bartlett
- Division of Veterinary and Biomedical Sciences, Murdoch University, Western Australian Biomedical Research Institute, Murdoch, Australia
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22
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Wigginton JM, Wiltrout RH. IL-12/IL-2 combination cytokine therapy for solid tumours: translation from bench to bedside. Expert Opin Biol Ther 2002; 2:513-24. [PMID: 12079487 DOI: 10.1517/14712598.2.5.513] [Citation(s) in RCA: 43] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Abstract
A broad range of approaches are under active investigation for the biological therapy of cancer, in particular, strategies directed at host immune response potentiation. These efforts have been fuelled by studies demonstrating the presence of an endogenous, but ineffective, host antitumour immune response and a greater understanding of the key factors which regulate this response. These mechanisms involve complex interactions between various effector cell populations, soluble factors and the tumour itself and are determined by the timing and relative intensity of positive and negative autoregulatory pathways, as well as a variety of immunosuppressive effects capable of mediating tumour self-defence. Based on these observations, immunotherapeutic regimens have been developed to potentiate antigen-specific sensitisation of effector cells with tumour vaccines/adjuvants, expand and amplify the number and function of effector cells, and to counteract suppressive pathways engaged by tumour cells themselves. Significant effort has focused on evaluating the use of exogenous cytokines, administered either systemically or locally into the tumour site via gene therapy. Several cytokines have demonstrated unique activity in the preclinical setting, including IL-2 and IFN-alpha -inducing cytokines such as IL12 and IL18. Most notably, later studies have now attempted to build on the clinical efficacy of IL-2 alone, to define combinations of agents with synergistic immunoregulatory and/or antitumour efficacy. Several lines of evidence suggest that IL-12 and IL-2 provide complementary immunoregulatory signals and have now shown that in combination, these two cytokines mediate synergistic antitumour activity in preclinical tumour models. This paper will review existing data regarding mechanisms of interaction between IL-2 and IL-12 in vitro and in preclinial models and describe future opportunities for the investigation of these potentially promising cytokines in the treatment of cancer.
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Affiliation(s)
- Jon M Wigginton
- Investigational Biologics Section, Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, MD, USA.
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Didenko VV, Ngo HN, Minchew C, Baskin DS. Apoptosis of T lymphocytes invading glioblastomas multiforme: a possible tumor defense mechanism. J Neurosurg 2002; 96:580-4. [PMID: 11883844 PMCID: PMC1853267 DOI: 10.3171/jns.2002.96.3.0580] [Citation(s) in RCA: 60] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022]
Abstract
OBJECT The goal of this study was to investigate whether apoptosis occurs in T lymphocytes that invade Fas ligand (FasL)-expressing glioblastomas multiforme (GBMs) and if its induction could be mediated by Fas. METHODS Apoptotic T lymphocytes were detected in GBMs by using detection of cell-type markers combined with active caspase-3 immunohistochemical analysis, a recently introduced apoptosis-specific in situ ligation assay, as well as by examining morphological criteria. Apoptotic T cells expressed Fas and were localized in the vicinity or in direct contact with FasL-expressing tumor cells. The T lymphocytes were undergoing apoptosis in spite of Bcl-2 expression. Expression of Bax was also detected in dying T cells, which can explain the absence of the protective effect of Bcl-2. because Bax inhibits Bcl-2 death-repressor activity. CONCLUSIONS On the basis of the data presented in this paper, the authors suggest that GBM cells that express FasL can induce apoptosis in invading immune cells. This phenomenon may play an important role in these tumors' maintenance of immune privilege and evasion of immune attacks. Awareness of this phenomenon should be helpful for the development of novel strategies for treatment of malignant gliomas.
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Affiliation(s)
- Vladimir V Didenko
- Department of Neurosurgery, Baylor College of Medicine, Houston, Texas, USA
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Strayer DS, Branco F, Landré J, BouHamdan M, Shaheen F, Pomerantz RJ. Combination genetic therapy to inhibit HIV-1. Mol Ther 2002; 5:33-41. [PMID: 11786043 DOI: 10.1006/mthe.2001.0513] [Citation(s) in RCA: 19] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Compared with single agents, combination antilentiviral pharmacotherapy targets multiple HIV-1 functions simultaneously, maximizing efficacy and decreasing chances of escape mutations. Combination genetic therapy could theoretically enhance efficacy similarly, but delivery of even single genes to high percentages of hematopoietic cells or their derivatives has proven problematic. Because of their high efficiency of gene delivery, we tested recombinant SV40-derived vectors (rSV40s) for this purpose. We made six rSV40s, each carrying a different transgene that targeted a different lentiviral function. We tested the ability of these constructs, individually and in double and triple combinations, to protect SupT1 human T lymphoma cells from HIV-1 challenge. Single chain antibodies (SFv) against CXCR4 and against HIV-1 reverse transcriptase (RT) and integrase (IN) were used, as were polymeric TAR decoys (PolyTAR) and a dominant-negative mutant of HIV-1 Rev (RevM10). Immunostaining showed that virtually all doubly treated cells expressed both transgenes. All transgenes individually protected from HIV-1 but, except for anti-CXCR4 SFv, their effectiveness diminished as challenge doses increased from 40 through 2500 tissue culture infectious dose(50) (TCID(50))/10(6) cells. However, all combinations of transgenes protected target cells better than individual transgenes, even from the highest challenge doses. Thus, combination gene therapies may inhibit HIV-1 better than single agents, and rSV40s may facilitate delivery of multigene therapeutics.
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Affiliation(s)
- David S Strayer
- Department of Pathology, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
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