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Choudhary HB, Mandlik SK, Mandlik DS. Role of p53 suppression in the pathogenesis of hepatocellular carcinoma. World J Gastrointest Pathophysiol 2023; 14:46-70. [PMID: 37304923 PMCID: PMC10251250 DOI: 10.4291/wjgp.v14.i3.46] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/27/2023] [Revised: 05/19/2023] [Accepted: 05/31/2023] [Indexed: 06/01/2023] Open
Abstract
In the world, hepatocellular carcinoma (HCC) is among the top 10 most prevalent malignancies. HCC formation has indeed been linked to numerous etiological factors, including alcohol usage, hepatitis viruses and liver cirrhosis. Among the most prevalent defects in a wide range of tumours, notably HCC, is the silencing of the p53 tumour suppressor gene. The control of the cell cycle and the preservation of gene function are both critically important functions of p53. In order to pinpoint the core mechanisms of HCC and find more efficient treatments, molecular research employing HCC tissues has been the main focus. Stimulated p53 triggers necessary reactions that achieve cell cycle arrest, genetic stability, DNA repair and the elimination of DNA-damaged cells’ responses to biological stressors (like oncogenes or DNA damage). To the contrary hand, the oncogene protein of the murine double minute 2 (MDM2) is a significant biological inhibitor of p53. MDM2 causes p53 protein degradation, which in turn adversely controls p53 function. Despite carrying wt-p53, the majority of HCCs show abnormalities in the p53-expressed apoptotic pathway. High p53 in-vivo expression might have two clinical impacts on HCC: (1) Increased levels of exogenous p53 protein cause tumour cells to undergo apoptosis by preventing cell growth through a number of biological pathways; and (2) Exogenous p53 makes HCC susceptible to various anticancer drugs. This review describes the functions and primary mechanisms of p53 in pathological mechanism, chemoresistance and therapeutic mechanisms of HCC.
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Affiliation(s)
- Heena B Choudhary
- Department of Pharmacology, BVDU, Poona College of Pharmacy, Pune 411038, Maharashtra, India
| | - Satish K Mandlik
- Department of Pharmaceutics, BVDU, Poona College of Pharmacy, Pune 411038, Maharashtra, India
| | - Deepa S Mandlik
- Department of Pharmacology, BVDU, Poona College of Pharmacy, Pune 411038, Maharashtra, India
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Yun CO, Hong J, Yoon AR. Current clinical landscape of oncolytic viruses as novel cancer immunotherapeutic and recent preclinical advancements. Front Immunol 2022; 13:953410. [PMID: 36091031 PMCID: PMC9458317 DOI: 10.3389/fimmu.2022.953410] [Citation(s) in RCA: 30] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2022] [Accepted: 08/03/2022] [Indexed: 12/12/2022] Open
Abstract
Oncolytic viruses (OVs) have been gaining attention in the pharmaceutical industry as a novel immunotherapeutic and therapeutic adjuvant due to their ability to induce and boost antitumor immunity through multiple mechanisms. First, intrinsic mechanisms of OVs that enable exploitation of the host immune system (e.g., evading immune detection) can nullify the immune escape mechanism of tumors. Second, many types of OVs have been shown to cause direct lysis of tumor cells, resulting in an induction of tumor-specific T cell response mediated by release of tumor-associated antigens and danger signal molecules. Third, armed OV-expressing immune stimulatory therapeutic genes could be highly expressed in tumor tissues to further improve antitumor immunity. Last, these OVs can inflame cold tumors and their microenvironment to be more immunologically favorable for other immunotherapeutics. Due to these unique characteristics, OVs have been tested as an adjuvant of choice in a variety of therapeutics. In light of these promising attributes of OVs in the immune-oncology field, the present review will examine OVs in clinical development and discuss various strategies that are being explored in preclinical stages for the next generation of OVs that are optimized for immunotherapy applications.
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Affiliation(s)
- Chae-Ok Yun
- Department of Bioengineering, College of Engineering, Hanyang University, Seoul, South Korea
- Institute of Nano Science and Technology (INST), Hanyang University, Seoul, South Korea
- Hanyang Institute of Bioscience and Biotechnology (HY-IBB), Hanyang University, Seoul, South Korea
- GeneMedicine CO., Ltd., Seoul, South Korea
| | | | - A-Rum Yoon
- Department of Bioengineering, College of Engineering, Hanyang University, Seoul, South Korea
- Institute of Nano Science and Technology (INST), Hanyang University, Seoul, South Korea
- Hanyang Institute of Bioscience and Biotechnology (HY-IBB), Hanyang University, Seoul, South Korea
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Almuzaini N, Moore M, Robert-Guroff M, Thomas MA. Disruption of NBS1/MRN Complex Formation by E4orf3 Supports NF-κB That Licenses E1B55K-Deleted Adenovirus-Infected Cells to Accumulate DNA>4n. Microbiol Spectr 2022; 10:e0188121. [PMID: 35019694 PMCID: PMC8754114 DOI: 10.1128/spectrum.01881-21] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2021] [Accepted: 12/14/2021] [Indexed: 01/16/2023] Open
Abstract
Cells increase their DNA content greater than the G2/M (DNA > 4n) phases along the path to cancer. The signals that support this increase in DNA content remain poorly understood. Cells infected with adenovirus (Ad) similarly develop DNA > 4n and share a need to bypass the DNA damage response (DDR) signals that trigger cell cycle arrest, and/or cell death. Ads with deletion in early region 1B55K (ΔE1B Ad) are oncolytic agents that are currently being explored for use in vaccine delivery. Interestingly, they promote higher levels of DNA > 4n than Ads that contain E1B55K. Existing in these and almost all Ads that are being explored for clinical use, is early region 4 (E4). The Ad E4 open reading frame 3 (E4orf3) is a viral oncogene that interferes with the ability of cells to respond to DNA damage by disrupting MRN complex formation. Our study reveals that E4orf3 is required for the enhanced fraction of ΔE1B Ad-infected cells with DNA > 4n. For that reason, we explored signaling events mediated by E4orf3. We found that in ΔE1B Ad-infected cells, E4orf3, as reported by others, isolates NBS1 in nuclear dots and tracks. This allows for elevated levels of phosphorylated ATM that is linked to transcriptionally active NF-κB. Pharmacological inhibition of NF-κB reduced the fraction of ΔE1B Ad-infected cells with DNA > 4n while pharmacological inhibition of ATM reduced the levels of nuclear NF-κB and the fraction of ΔE1B Ad-infected cells with DNA > 4n and increased the fraction of dead or dying cells with fragmented DNA. This ability of E4orf3 to disrupt MRN complex formation that allows cells to bypass the cell cycle, evade death, and accumulate DNA > 4n, may be linked to its oncogenic potential. IMPORTANCE Genome instability, a hallmark of cancer, exists as part of a cycle that leads to DNA damage and DNA > 4n that further enhances genome instability. Ad E4orf3 is a viral oncogene. Here, we describe E4orf3 mediated signaling events that support DNA > 4n in ΔE1B Ad-infected cells. These signaling events may be linked to the oncogenic potential of E4orf3 and may provide a basis for how some cells survive with DNA > 4n.
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Affiliation(s)
- Nujud Almuzaini
- Department of Biology, College of Arts and Sciences, Howard University, Washington, D.C., USA
| | - Madison Moore
- Department of Biology, College of Arts and Sciences, Howard University, Washington, D.C., USA
| | - Marjorie Robert-Guroff
- Section on Immune Biology of Retroviral Infection, Vaccine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA
| | - Michael A. Thomas
- Department of Biology, College of Arts and Sciences, Howard University, Washington, D.C., USA
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Li GL, Qian H. Transcriptome using Illumina sequencing reveals the traits of spermatogenesis and developing testes in Eriocheir sinensis. PLoS One 2017; 12:e0172478. [PMID: 28212420 PMCID: PMC5315355 DOI: 10.1371/journal.pone.0172478] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2016] [Accepted: 02/05/2017] [Indexed: 11/19/2022] Open
Abstract
Chinese mitten crab (Eriocheir sinensis) has the spermatozoa with typical aflagellate, decondensed chromatin, cup-shaped nuclei, and radial arms. However, the mechanism of spermatogenesis during which the specific spermatozoa are generated in this species is yet unclear. Here, the transcriptome of developing testis in E. sinensis was analyzed using the ways of RNA-seq and bioinformatics analysis to identify candidate genes potentially involved in development of testis and spermatogenesis. The Illumina HiSeq2500 sequencing of three replicons of samples produced a total of 145.19 M clean reads representing with a total of 21.34 Gb bases and 45.48% GC content. 56.30% clean reads were mapped to the draft genome of E. sinensis. The assembly of the transcriptome yielded contigs of 5691802 sequences and unigenes of 406527 sequences. Total 24246 and 40793 transcripts were annotated using Swissprot and Nr database, respectively. There were 48213 (70.31%) and 7858 (46.25%) transcripts with identity of more than 99 matching to mature testis unigenes in the databases of Nr and EST, respectively. The analytic results of KOG, GO and KEGG showed wide potential molecular functions of transcripts in the developing testes. KEGG analysis of unigenes yielded total 9422 predicted genes. Those predicted genes were involved in total 216 KEGG pathways related to the physiological activities of developing testis. 1975 predicted genes were involved in cellular and subcellular structural alteration of male germ cells. There were important roles of some pathways in the processes of morphological and structural biogenesis pertaining to testis development and spermatogenesis. Other 583 unigenes encoding the genetic and epigenetic factors also be found, which might contribute to the decondensation and stability of decondensed nuclei in the spermatozoa. These predicted events provide a view of the potential molecular mechanisms of development of testis and spermatogenesis in E. sinensis.
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Affiliation(s)
- Gen-Liang Li
- Youjiang Medical University for Nationalities, Baise, Guangxi, China
| | - Hui Qian
- Youjiang Medical University for Nationalities, Baise, Guangxi, China
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5
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Optimal Control Model of Tumor Treatment with Oncolytic Virus and MEK Inhibitor. BIOMED RESEARCH INTERNATIONAL 2016; 2016:5621313. [PMID: 28097139 PMCID: PMC5210284 DOI: 10.1155/2016/5621313] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/21/2016] [Accepted: 11/27/2016] [Indexed: 11/28/2022]
Abstract
Tumors are a serious threat to human health. The oncolytic virus is a kind of tumor killer virus which can infect and lyse cancer cells and spread through the tumor, while leaving normal cells largely unharmed. Mathematical models can help us to understand the tumor-virus dynamics and find better treatment strategies. This paper gives a new mathematical model of tumor therapy with oncolytic virus and MEK inhibitor. Stable analysis was given. Because mitogen-activated protein kinase (MEK) can not only lead to greater oncolytic virus infection into cancer cells, but also limit the replication of the virus, in order to provide the best dosage of MEK inhibitors and balance the positive and negative effect of the inhibitors, we put forward an optimal control problem of the inhibitor. The optimal strategies are given by theory and simulation.
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Tookman LA, Browne AK, Connell CM, Bridge G, Ingemarsdotter CK, Dowson S, Shibata A, Lockley M, Martin SA, McNeish IA. RAD51 and BRCA2 Enhance Oncolytic Adenovirus Type 5 Activity in Ovarian Cancer. Mol Cancer Res 2016; 14:44-55. [PMID: 26452665 PMCID: PMC4716290 DOI: 10.1158/1541-7786.mcr-15-0188-t] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/27/2015] [Accepted: 09/30/2015] [Indexed: 12/16/2022]
Abstract
UNLABELLED Homologous recombination (HR) function is critically important in high-grade serous ovarian cancer (HGSOC). HGSOC with intact HR has a worse prognosis and is less likely to respond to platinum chemotherapy and PARP inhibitors. Oncolytic adenovirus, a novel therapy for human malignancies, stimulates a potent DNA damage response that influences overall antitumor activity. Here, the importance of HR was investigated by determining the efficacy of adenovirus type 5 (Ad5) vectors in ovarian cancer. Using matched BRCA2-mutant and wild-type HGSOC cells, it was demonstrated that intact HR function promotes viral DNA replication and augments overall efficacy, without influencing viral DNA processing. These data were confirmed in a wider panel of HR competent and defective ovarian cancer lines. Mechanistically, both BRCA2 and RAD51 localize to viral replication centers within the infected cell nucleus and that RAD51 localization occurs independently of BRCA2. In addition, a direct interaction was identified between RAD51 and adenovirus E2 DNA binding protein. Finally, using functional assays of HR competence, despite inducing degradation of MRE11, Ad5 infection does not alter cellular ability to repair DNA double-strand break damage via HR. These data reveal that Ad5 redistributes critical HR components to viral replication centers and enhances cytotoxicity. IMPLICATIONS Oncolytic adenoviral therapy may be most clinically relevant in tumors with intact HR function.
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Affiliation(s)
- Laura A Tookman
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Ashley K Browne
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Claire M Connell
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Gemma Bridge
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Carin K Ingemarsdotter
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Suzanne Dowson
- Institute of Cancer Sciences, University of Glasgow, Glasgow, United Kingdom
| | - Atsushi Shibata
- Advanced Scientific Research Leaders Development Unit, Gunma University, Maebashi, Gunma, Japan
| | - Michelle Lockley
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Sarah A Martin
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
| | - Iain A McNeish
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom. Institute of Cancer Sciences, University of Glasgow, Glasgow, United Kingdom.
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Yamauchi S, Kawamura K, Okamoto S, Morinaga T, Jiang Y, Shingyoji M, Sekine I, Kubo S, Tada Y, Tatsumi K, Shimada H, Hiroshima K, Tagawa M. Replication-competent adenoviruses with the type 35-derived fiber-knob region achieve reactive oxygen species-dependent cytotoxicity and produce greater toxicity than those with the type 5-derived region in pancreatic carcinoma. Apoptosis 2015; 20:1587-98. [PMID: 26373551 DOI: 10.1007/s10495-015-1171-8] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Pancreatic carcinoma is relatively resistant to chemotherapy and cell death induced by replication of adenoviruses (Ad) can be one of the therapeutic options. Transduction efficacy of conventional type 5 Ad (Ad5) is however low and the cytotoxic mechanism by replication-competent Ad was not well understood. We constructed replication-competent Ad5 of which the E1A promoter region was replaced with a transcriptional regulatory region of the midkine, the survivin or the cyclooxygenase-2 gene, all of which were expressed at a high level in human tumors. We also prepared replication-competent Ad5 that were activated with the same region but had the type 35 Ad-derived fiber-knob region (AdF35) to convert the major cellular receptor for Ad infection from the coxsackie adenovirus receptor to CD46 molecules. Replication-competent AdF35 that were activated with the exogenous region produced cytotoxic effects on human pancreatic carcinoma cells greater than the corresponding Ad5 bearing with the same regulatory region. Cells infected with the AdF35 showed cytopathic effects and increased sub-G1 fractions. Caspase-9, less significantly caspase-8 and poly (ADP-ribose) polymerase, but not caspase-3 was cleaved and expression of molecules involved in autophagy and caspase-independent cell death pathways remained unchanged. Nevertheless, H2A histone family member X molecules were phosphorylated, and N-acetyl-L-cystein, an inhibitor for reactive oxygen species, suppressed the AdF35-mediated cytotoxicity. These data indicated a novel mechanism of Ad-mediated cell death and suggest a possible clinical application of the fiber-knob modified Ad.
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Affiliation(s)
- Suguru Yamauchi
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Kiyoko Kawamura
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
| | - Shinya Okamoto
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Takao Morinaga
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Yuanyuan Jiang
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | | | - Ikuo Sekine
- Division of Respirology, Chiba Cancer Center, Chiba, Japan
| | - Shuji Kubo
- Department of Genetics, Hyogo College of Medicine, Nishinomiya, Japan
| | - Yuji Tada
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Koichiro Tatsumi
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Hideaki Shimada
- Department of Surgery, School of Medicine, Toho University, Tokyo, Japan
| | - Kenzo Hiroshima
- Department of Pathology, Tokyo Women's Medical University Yachiyo Medical Center, Yachiyo, Japan
| | - Masatoshi Tagawa
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan.
- Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan.
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Synergistic cytotoxicity against human tumor cell lines by oncolytic adenovirus dl1520 (ONYX-015) and melphalan. TUMORI JOURNAL 2015; 102:31-9. [PMID: 26429639 DOI: 10.5301/tj.5000438] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 09/04/2015] [Indexed: 11/20/2022]
Abstract
AIMS AND BACKGROUND In light of the need for more selective anticancer therapy, much work has been directed at developing compounds or biological agents that target functions specific to cancer cells. To this end, numerous viruses have been engineered to exploit the dependence of cancer cells on particular anomalies that contribute to their rogue proliferative activity, such as dysfunctional p53, overactive mitogenic signaling, or a defective interferon response. The oncolytic human adenovirus dl1520 (ONYX-015) was engineered to propagate specifically in p53-deficient tumors, which comprise over half of all tumors. Based on successes in clinical trials, the full potential of dl1520 and other oncolytic viruses may be even better realized by using them in combination with conventional chemotherapy drugs. METHODS As a model system in which to test this potential, representative cell lines from 2 common cancer types, oral squamous cell carcinoma (HN-5a) and colon adenocarcinoma (HT-29), were chosen, as well as platinum-drug-resistant variants of each. RESULTS Following preliminary screening of virus and drug combinations, dl1520 and melphalan were found to synergistically inhibit proliferation of all the cancer cell lines. Melphalan pretreatment or cotreatment with dl1520 enhanced inhibition of proliferation by dl1520 by up to 60% and increased apoptosis by up to 25%. The tight-junction protein CAR (coxsackie and adenovirus receptor), via which adenovirus enters cells, was not upregulated by treatment with melphalan, suggesting that other mechanisms contribute to synergy. CONCLUSIONS The synergy between melphalan and dl1520 suggests that tumor-selective cell killing by oncolytic viruses may be augmented by combining with cytotoxic drugs.
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Ma G, Zhong B, Okamoto S, Jiang Y, Kawamura K, Liu H, Li Q, Shingyoji M, Sekine I, Tada Y, Tatsumi K, Shimada H, Hiroshima K, Tagawa M. A combinatory use of adenoviruses expressing melanoma differentiation-associated gene-7 and replication-competent adenoviruses produces synergistic effects on pancreatic carcinoma cells. Tumour Biol 2015; 36:8137-45. [PMID: 25990458 DOI: 10.1007/s13277-015-3555-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2015] [Accepted: 05/11/2015] [Indexed: 11/30/2022] Open
Abstract
Type 5 adenoviruses expressing mda-7 gene (Ad-mda-7) induced cell death in various kinds of human tumors, but pancreatic carcinoma cells were relatively resistant to Ad-mda-7-mediated cytotoxicity. We then examined whether infection of Ad-mda-7 together with replication-competent Ad produced combinatory cytotoxic effects. We prepared replication-competent Ad, defective of the E1B55kDa gene or activated by a transcriptional regulatory region of the midkine or the survivin gene of which the expression was up-regulated in human tumors. Type 5 Ad bearing the exogenous regulatory region were further modified by replacing the fiber-knob region with that of type 35 Ad. Pancreatic carcinoma cells were infected with replication-incompetent Ad-mda-7 and the replication-competent Ad. Combinatory effects were examined with the CalcuSyn software and cell cycle analyses. Ad-mda-7 and the replication-competent Ad achieved cytotoxicity to pancreatic carcinoma. A combinatory use of Ad-mda-7 and either Ad defective of the E1B55kDa gene or Ad activated by the regulatory region produced synergistic cytotoxic effects. Cell cycle analyses demonstrated that the combination increased sub-G1 populations. These data collectively suggest that expression of MDA-7 augments cytotoxicity of replication-competent Ad and achieves adjuvant effects on Ad-mediated cell death.
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Affiliation(s)
- Guangyu Ma
- Department of Hematology, The Fourth Hospital of Hebei Medical University, Shijiazhuang, China
| | - Boya Zhong
- Department of Hematology, The Fourth Hospital of Hebei Medical University, Shijiazhuang, China
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Shinya Okamoto
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Yuanyuan Jiang
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
- Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Kiyoko Kawamura
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
| | - Hongdan Liu
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan
| | - Quanhai Li
- Department of Immunology, Hebei Medical University, Shijiazhuang, China
- Cell Therapy Center, The First Hospital of Hebei Medical University, Shijiazhuang, China
| | - Masato Shingyoji
- Department of Thoracic Diseases, Chiba Cancer Center, Chiba, Japan
| | - Ikuo Sekine
- Department of Thoracic Diseases, Chiba Cancer Center, Chiba, Japan
| | - Yuji Tada
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Koichiro Tatsumi
- Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Hideaki Shimada
- Department of Surgery, School of Medicine, Toho University, Tokyo, Japan
| | - Kenzo Hiroshima
- Department of Pathology, Tokyo Women's Medical University Yachiyo Medical Center, Yachiyo, Japan
| | - Masatoshi Tagawa
- Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, 666-2 Nitona, Chuo-ku, Chiba, 260-8717, Japan.
- Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan.
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10
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Takei Y, Okamoto S, Kawamura K, Jiang Y, Morinaga T, Shingyoji M, Sekine I, Kubo S, Tada Y, Tatsumi K, Shimada H, Hiroshima K, Yamaguchi N, Tagawa M. Expression of p53 synergistically augments caspases-mediated apoptosis induced by replication-competent adenoviruses in pancreatic carcinoma cells. Cancer Gene Ther 2015; 22:445-53. [PMID: 26251031 DOI: 10.1038/cgt.2015.33] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/21/2014] [Revised: 07/02/2015] [Accepted: 07/03/2015] [Indexed: 12/16/2022]
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11
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Cytotoxic effects of replication-competent adenoviruses on human esophageal carcinoma are enhanced by forced p53 expression. BMC Cancer 2015; 15:464. [PMID: 26059686 PMCID: PMC4460641 DOI: 10.1186/s12885-015-1482-8] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2014] [Accepted: 06/02/2015] [Indexed: 12/05/2022] Open
Abstract
Background Improvement of transduction and augmentation of cytotoxicity are crucial for adenoviruses (Ad)-mediated gene therapy for cancer. Down-regulated expression of type 5 Ad (Ad5) receptors on human tumors hampered Ad-mediated transduction. Furthermore, a role of the p53 pathways in cytotoxicity mediated by replication-competent Ad remained uncharacterized. Methods We constructed replication-competent Ad5 of which the E1 region genes were activated by a transcriptional regulatory region of the midkine or the survivin gene, which is expressed preferentially in human tumors. We also prepared replication-competent Ad5 which were regulated by the same region but had a fiber-knob region derived from serotype 35 (AdF35). We examined the cytotoxicity of these Ad and a possible combinatory use of the replication-competent AdF35 and Ad5 expressing the wild-type p53 gene (Ad5/p53) in esophageal carcinoma cells. Expression levels of molecules involved in cell death, anti-tumor effects in vivo and production of viral progenies were also investigated. Results Replication-competent AdF35 in general achieved greater cytotoxic effects to esophageal carcinoma cells than the corresponding replication-competent Ad5. Infection with the AdF35 induced cleavages of caspases and increased sub-G1 fractions, but did not activate the autophagy pathway. Transduction with Ad5/p53 in combination with the replication-competent AdF35 further enhanced the cytotoxicity in a synergistic manner. We also demonstrated the combinatory effects in an animal model. Transduction with Ad5/p53 however suppressed production of replication-competent AdF35 progenies, but the combination augmented Ad5/p53-mediated p53 expression levels and the downstream pathways. Conclusions Combination of replication-competent AdF35 and Ad5/p53 achieved synergistic cytotoxicity due to enhanced p53-mediated apoptotic pathways. Electronic supplementary material The online version of this article (doi:10.1186/s12885-015-1482-8) contains supplementary material, which is available to authorized users.
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12
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Turner RL, Groitl P, Dobner T, Ornelles DA. Adenovirus replaces mitotic checkpoint controls. J Virol 2015; 89:5083-96. [PMID: 25694601 PMCID: PMC4403466 DOI: 10.1128/jvi.00213-15] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2015] [Accepted: 02/17/2015] [Indexed: 12/23/2022] Open
Abstract
UNLABELLED Infection with adenovirus triggers the cellular DNA damage response, elements of which include cell death and cell cycle arrest. Early adenoviral proteins, including the E1B-55K and E4orf3 proteins, inhibit signaling in response to DNA damage. A fraction of cells infected with an adenovirus mutant unable to express the E1B-55K and E4orf3 genes appeared to arrest in a mitotic-like state. Cells infected early in G1 of the cell cycle were predisposed to arrest in this state at late times of infection. This arrested state, which displays hallmarks of mitotic catastrophe, was prevented by expression of either the E1B-55K or the E4orf3 genes. However, E1B-55K mutant virus-infected cells became trapped in a mitotic-like state in the presence of the microtubule poison colcemid, suggesting that the two viral proteins restrict entry into mitosis or facilitate exit from mitosis in order to prevent infected cells from arresting in mitosis. The E1B-55K protein appeared to prevent inappropriate entry into mitosis through its interaction with the cellular tumor suppressor protein p53. The E4orf3 protein facilitated exit from mitosis by possibly mislocalizing and functionally inactivating cyclin B1. When expressed in noninfected cells, E4orf3 overcame the mitotic arrest caused by the degradation-resistant R42A cyclin B1 variant. IMPORTANCE Cells that are infected with adenovirus type 5 early in G1 of the cell cycle are predisposed to arrest in a mitotic-like state in a p53-dependent manner. The adenoviral E1B-55K protein prevents entry into mitosis. This newly described activity for the E1B-55K protein appears to depend on the interaction between the E1B-55K protein and the tumor suppressor p53. The adenoviral E4orf3 protein facilitates exit from mitosis, possibly by altering the intracellular distribution of cyclin B1. By preventing entry into mitosis and by promoting exit from mitosis, these adenoviral proteins act to prevent the infected cell from arresting in a mitotic-like state.
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Affiliation(s)
- Roberta L Turner
- Department of Microbiology and Immunology, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Peter Groitl
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, Hamburg, Germany
| | - Thomas Dobner
- Heinrich Pette Institute, Leibniz Institute for Experimental Virology, Hamburg, Germany
| | - David A Ornelles
- Department of Microbiology and Immunology, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
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Turner RL, Wilkinson JC, Ornelles DA. E1B and E4 oncoproteins of adenovirus antagonize the effect of apoptosis inducing factor. Virology 2014; 456-457:205-19. [PMID: 24889240 DOI: 10.1016/j.virol.2014.03.010] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2014] [Revised: 02/28/2014] [Accepted: 03/10/2014] [Indexed: 01/03/2023]
Abstract
Adenovirus inundates the productively infected cell with linear, double-stranded DNA and an abundance of single-stranded DNA. The cellular response to this stimulus is antagonized by the adenoviral E1B and E4 early genes. A mutant group C adenovirus that fails to express the E1B-55K and E4orf3 genes is unable to suppress the DNA-damage response. Cells infected with this double-mutant virus display significant morphological heterogeneity at late times of infection and frequently contain fragmented nuclei. Nuclear fragmentation was due to the translocation of apoptosis inducing factor (AIF) from the mitochondria into the nucleus. The release of AIF was dependent on active poly(ADP-ribose) polymerase-1 (PARP-1), which appeared to be activated by viral DNA replication. Nuclear fragmentation did not occur in AIF-deficient cells or in cells treated with a PARP-1 inhibitor. The E1B-55K or E4orf3 proteins independently prevented nuclear fragmentation subsequent to PARP-1 activation, possibly by altering the intracellular distribution of PAR-modified proteins.
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Affiliation(s)
- Roberta L Turner
- Department of Microbiology and Immunology, Wake Forest School of Medicine, Winston-Salem, NC 27157, United States
| | - John C Wilkinson
- Department of Biochemistry, Wake Forest School of Medicine, Winston-Salem, NC 27157, United States.
| | - David A Ornelles
- Department of Microbiology and Immunology, Wake Forest School of Medicine, Winston-Salem, NC 27157, United States.
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Thomas MA, Song R, Demberg T, Vargas-Inchaustegui DA, Venzon D, Robert-Guroff M. Effects of the deletion of early region 4 (E4) open reading frame 1 (orf1), orf1-2, orf1-3 and orf1-4 on virus-host cell interaction, transgene expression, and immunogenicity of replicating adenovirus HIV vaccine vectors. PLoS One 2013; 8:e76344. [PMID: 24143187 PMCID: PMC3797075 DOI: 10.1371/journal.pone.0076344] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2013] [Accepted: 08/23/2013] [Indexed: 12/03/2022] Open
Abstract
The global health burden engendered by human immunodeficiency virus (HIV)-induced acquired immunodeficiency syndrome (AIDS) is a sobering reminder of the pressing need for a preventative vaccine. In non-human primate models replicating adenovirus (Ad)-HIV/SIV recombinant vaccine vectors have been shown to stimulate potent immune responses culminating in protection against challenge exposures. Nonetheless, an increase in the transgene carrying capacity of these Ad vectors, currently limited to approximately 3000 base pairs, would greatly enhance their utility. Using a replicating, E3-deleted Ad type 5 host range mutant (Ad5 hr) encoding full-length single-chain HIVBaLgp120 linked to the D1 and D2 domains of rhesus macaque CD4 (rhFLSC) we systematically deleted the genes encoding early region 4 open reading frame 1 (E4orf1) through E4orf4. All the Ad-rhFLSC vectors produced similar levels of viral progeny. Cell cycle analysis of infected human and monkey cells revealed no differences in virus-host interaction. The parental and E4-deleted viruses expressed comparable levels of the transgene with kinetics similar to Ad late proteins. Similar levels of cellular immune responses and transgene-specific antibodies were elicited in vaccinated mice. However, differences in recognition of Ad proteins and induced antibody subtypes were observed, suggesting that the E4 gene products might modulate antibody responses by as yet unknown mechanisms. In short, we have improved the transgene carrying capacity by one thousand base pairs while preserving the replicability, levels of transgene expression, and immunogenicity critical to these vaccine vectors. This additional space allows for flexibility in vaccine design that could not be obtained with the current vector and as such should facilitate the goal of improving vaccine efficacy. To the best of our knowledge, this is the first report describing the effects of these E4 deletions on transgene expression and immunogenicity in a replicating Ad vector.
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Affiliation(s)
- Michael A. Thomas
- Section on Immune Biology of Retroviral Infection, Vaccine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Rui Song
- Section on Immune Biology of Retroviral Infection, Vaccine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Thorsten Demberg
- Section on Immune Biology of Retroviral Infection, Vaccine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Diego A. Vargas-Inchaustegui
- Section on Immune Biology of Retroviral Infection, Vaccine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - David Venzon
- Biostatistics and Data Management Section, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Marjorie Robert-Guroff
- Section on Immune Biology of Retroviral Infection, Vaccine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
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Adenovirus E1A oncogene induces rereplication of cellular DNA and alters DNA replication dynamics. J Virol 2013; 87:8767-78. [PMID: 23740993 DOI: 10.1128/jvi.00879-13] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The oncogenic property of the adenovirus (Ad) transforming E1A protein is linked to its capacity to induce cellular DNA synthesis which occurs as a result of its interaction with several host proteins, including pRb and p300/CBP. While the proteins that contribute to the forced induction of cellular DNA synthesis have been intensively studied, the nature of the cellular DNA replication that is induced by E1A in quiescent cells is not well understood. Here we show that E1A expression in quiescent cells leads to massive cellular DNA rereplication in late S phase. Using a single-molecule DNA fiber assay, we studied the cellular DNA replication dynamics in E1A-expressing cells. Our studies show that the DNA replication pattern is dramatically altered in E1A-expressing cells, with increased replicon length, fork velocity, and interorigin distance. The interorigin distance increased by about 3-fold, suggesting that fewer DNA replication origins are used in E1A-expressing cells. These aberrant replication events led to replication stress, as evidenced by the activation of the DNA damage response. In earlier studies, we showed that E1A induces c-Myc as a result of E1A binding to p300. Using an antisense c-Myc to block c-Myc expression, our results indicate that induction of c-Myc in E1A-expressing cells contributes to the induction of host DNA replication. Together, our results suggest that the E1A oncogene-induced cellular DNA replication stress is due to dramatically altered cellular replication events and that E1A-induced c-Myc may contribute to these events.
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E1B-55 kDa-defective adenoviruses activate p53 in mesothelioma and enhance cytotoxicity of anticancer agents. J Thorac Oncol 2013; 7:1850-1857. [PMID: 23154556 DOI: 10.1097/jto.0b013e3182725fa4] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
INTRODUCTION Genetic characterization of malignant mesothelioma shows a homozygous deletion of the INK4A/ARF locus, which results in inactivation of the p53 pathways. METHODS We examined possible antitumor effects of adenoviruses with a deletion of the E1B-55kD gene (Ad-delE1B55) on mesothelioma and investigated combinatory actions with the first-line chemotherapeutic agents. RESULTS Ad-delE1B55 produced cytotoxicity on mesothelioma cells, which was associated with p53 phosphorylation, pRb dephosphorylation, and cleavage of caspases. Ad-delE1B55-infected cells displayed hyperploidy at the cell-cycle analysis and showed enlarged nuclear configurations. Combination of Ad-delE1B55 plus cisplatin or pemetrexed produced antitumor effects in vitro. Furthermore, Ad-delE1B55 and cisplatin showed combinatory effects in an orthotopic animal model. CONCLUSIONS Cell death caused by Ad-delE1B55 is attributable to cell-cycle arrest at M-phase checkpoint followed by activated apoptotic pathways, and combination of the first-line chemotherapeutic agents and the oncolytic adenovirus is a potential therapeutic for mesothelioma.
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Huang X, Wang L, Zhang H, Wang H, Zhao X, Qian G, Hu J, Ge S, Fan X. Therapeutic efficacy by targeting correction of Notch1-induced aberrants in uveal tumors. PLoS One 2012; 7:e44301. [PMID: 22937170 PMCID: PMC3429424 DOI: 10.1371/journal.pone.0044301] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2011] [Accepted: 08/02/2012] [Indexed: 11/19/2022] Open
Abstract
There is a need for more effective treatments for uveal melanoma. The recombinant oncolytic adenovirus H101 replicates specifically in p53-depleted tumor cells, and has been approved for use by the Chinese State Food and Drug Administration. However, this treatment is associated with subsequent remission. Transfection of uveal melanoma cells with a small interfering RNA against Notch1 (siNotch1) effectively suppressed Notch1 expression, resulting in significant cell growth inhibition when combined with H101 treatment. Combined treatment with siNotch1 and H101 (H101-Notch1-siRNA) greatly enhanced apoptosis and cell cycle arrest in vitro as compared to treatment with H101 or siNotch1 alone. For in vivo treatments, the combined treatment of siNotch1 and H101 showed remarkable tumor growth inhibition and prolonged mouse survival in the OCM1 xenograft model. We predict that Notch pathway deregulation could be a feature of uveal melanoma, and could be a therapeutic target, especially if p53 is concurrently targeted.
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Affiliation(s)
- Xiaolin Huang
- Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Li Wang
- Department of Biochemistry and Molecular Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - He Zhang
- Veterans Affairs Palo Alto Health Care System, Stanford University Medical School, Palo Alto, California, United States of America
- Department of Biochemistry and Molecular Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Haibo Wang
- Veterans Affairs Palo Alto Health Care System, Stanford University Medical School, Palo Alto, California, United States of America
- Department of Biochemistry and Molecular Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Xiaoping Zhao
- Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- Department of Biochemistry and Molecular Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Guanxiang Qian
- Department of Biochemistry and Molecular Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Jifan Hu
- Veterans Affairs Palo Alto Health Care System, Stanford University Medical School, Palo Alto, California, United States of America
| | - Shengfang Ge
- Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- Department of Biochemistry and Molecular Biology, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- * E-mail: (XF); (SG)
| | - Xianqun Fan
- Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
- * E-mail: (XF); (SG)
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18
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Adam V, Ekblad M, Sweeney K, Müller H, Busch KH, Johnsen CT, Kang NR, Lemoine NR, Halldén G. Synergistic and Selective Cancer Cell Killing Mediated by the Oncolytic Adenoviral Mutant AdΔΔ and Dietary Phytochemicals in Prostate Cancer Models. Hum Gene Ther 2012; 23:1003-15. [PMID: 22788991 DOI: 10.1089/hum.2012.046] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
AdΔΔ is an oncolytic adenoviral mutant that has been engineered to selectively target tumors with deregulated cell cycle and apoptosis pathways. AdΔΔ potentiates apoptotic cell death induced by drugs, including mitoxantrone and docetaxel, which are commonly used to treat prostate cancer. Here, we demonstrate that AdΔΔ can also interact synergistically with dietary phytochemicals known to have anti-cancer activities, without incurring the toxic side effects of chemodrugs. Curcumin, genistein, epigallocatechin-gallate, equol, and resveratrol efficiently killed both androgen-receptor positive (22Rv1) and negative cell lines (PC-3, DU145) in combination with adenoviral mutants. Synergistic cell killing was demonstrated with wild-type virus (Ad5) and AdΔΔ in combination with equol and resveratrol. EC(50) values for both phytochemicals and viruses were reduced three- to eightfold in all three combination-treated cell lines. The most potent efficacy was achieved in the cytotoxic drug- and virus-insensitive PC-3 cells, both in vitro and in vivo, while cell killing in normal bronchial epithelial cells was not enhanced. Although equol and resveratrol induced only low levels of apoptosis when administered alone, in combination with wild-type virus or AdΔΔ, the level of apoptotic cell death was significantly increased in PC-3 and DU145 cells. In vivo studies using suboptimal doses of AdΔΔ and equol or resveratrol, showed reduced tumor growth without toxicity to normal tissue. These findings identify novel functions for AdΔΔ and phytochemicals in promoting cancer cell killing and apoptosis, suggesting the use of these natural nontoxic compounds might be a feasible and currently unexploited anti-cancer strategy.
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Affiliation(s)
- Virginie Adam
- Centre for Molecular Oncology, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
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Halldén G, Portella G. Oncolytic virotherapy with modified adenoviruses and novel therapeutic targets. Expert Opin Ther Targets 2012; 16:945-58. [PMID: 22880939 DOI: 10.1517/14728222.2012.712962] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
INTRODUCTION Numerous oncolytic viral mutants derived from a variety of strains have antitumor efficacy with limited or no toxicity to normal tissue. While all modes of administration were determined to be safe in patients with solid cancers refractory to current standard of care, this therapeutic approach requires further improvements to achieve definite efficacy. AREAS COVERED We review the most promising clinical developments with several oncolytic viruses. The focus is on preclinical and clinical findings with replication-selective adenoviral mutants including ONYX-015, H101 and Ad5ΔCR mutants that, to date, are the most studied oncolytic viruses. Cellular pathways reported to play a role in virus-induced cell killing are reviewed as potential targets for the development of more effective combinatorial therapies. EXPERT OPINION The most promising clinical outcomes for metastatic cancers have been reported for oncolytic vaccinia and herpes virus mutants expressing the cytokine GMCSF. However, highly efficacious and selective adenoviral mutants have been developed that interact synergistically with cytotoxic drugs in model systems. We anticipate that by delineating the cellular targets for synergistic cancer cell killing in response to adenoviral mutants and drugs such as apoptosis and autophagy signaling, greatly improved anticancer therapies will result in the near future.
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Affiliation(s)
- Gunnel Halldén
- Queen Mary University of London, Barts Cancer Institute, Centre for Molecular Oncology, London, UK
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20
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Cun B, Song X, Jia R, Zhao X, Wang H, Ge S, Fan X. Combination of oncolytic adenovirus and dacarbazine attenuates antitumor ability against uveal melanoma cells via cell cycle block. Cancer Biol Ther 2012; 13:77-84. [PMID: 22336909 DOI: 10.4161/cbt.13.2.18436] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
Uveal melanoma is the most common primary intraocular malignancy in adults; however, current therapeutic modalities, including chemotherapy, have not been successful. Oncolytic viruses serve as an emerging gene therapy tool for cancer treatment because they specifically kill tumor cells while sparing normal cells. The oncolytic virus H101 has been approved by the Chinese State Food and Drug Administration for the treatment of certain malignancies. Unfortunately, the monotherapy of adenovirus has demonstrated limited efficacy in a clinical setting. Thus, novel treatment strategies in which an oncolytic virus is combined with existing chemicals are advancing toward potential clinical use. In this study, we chose the combination of oncolytic virus H101 and the alkylating agent dacarbazine (DTIC) to treat uveal melanoma cells in vitro. Our results demonstrated that the combination exerted a synergistic antitumor effect without enhanced toxicity to normal cells via a type of cell cycle block other than the induction of apoptosis. Further investigation is warranted to elucidate the specific underlying mechanisms of this co-treatment therapy. Our study suggests the viro-chemo combination therapy is feasible and is a potentially promising approach for the treatment of uveal melanoma.
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Affiliation(s)
- Biyun Cun
- Department of Ophthalmology, Ninth People’s Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China
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21
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The oncolytic adenovirus AdΔΔ enhances selective cancer cell killing in combination with DNA-damaging drugs in pancreatic cancer models. Gene Ther 2011; 18:1157-65. [PMID: 21975464 DOI: 10.1038/gt.2011.141] [Citation(s) in RCA: 44] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
Pancreatic adenocarcinomas are aggressive and frequently develop resistance to all current therapies. Replication-selective adenoviruses can overcome resistance to chemotherapeutics through their sensitizing effects on drug-induced cell killing. We previously found that adenovirus deleted in the anti-apoptotic E1B19K gene enhanced gemcitabine-induced apoptotis. Here we demonstrate that our engineered double-deleted AdΔΔ mutant (deleted in the pRb-binding E1ACR2 region and E1B19K) selectively replicates and enhances cell killing in combination with DNA-damaging cytotoxic drugs in pancreatic cancer cells. Combinations of AdΔΔ with gemcitabine, irinotecan or cisplatin resulted in two- to fourfold decreases in EC(50) (half maximal effective concentration) values and was more efficent than similar combinations with wild-type virus, the dl1520 (ONYX-015) and dl922-947 mutants. AdΔΔ replication was impaired in normal bronchial human epithelial cells and did not sensitize the cells to drugs. Gemcitabine-insensitive AsPC-1, BxPC-3 and PANC-1 cells were efficiently killed by irinotecan in combination with AdΔΔ. Suboptimal doses of AdΔΔ and gemcitabine significantly prolonged time to tumor progression in two human pancreatic tumor xenograft in vivo models, PT45 and SUIT-2. We conclude that AdΔΔ has low toxicity to normal cells while potently sensitizing pancreatic cancer cells to DNA-damaging drugs, and holds promise as an improved therapeutic strategy for pancreatic cancer.
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Cherubini G, Naim V, Caruso P, Burla R, Bogliolo M, Cundari E, Benihoud K, Saggio I, Rosselli F. The FANC pathway is activated by adenovirus infection and promotes viral replication-dependent recombination. Nucleic Acids Res 2011; 39:5459-73. [PMID: 21421559 PMCID: PMC3141233 DOI: 10.1093/nar/gkr084] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Deciphering the crosstalk between a host cell and a virus during infection is important not only to better define viral biology but also to improve our understanding of cellular processes. We identified the FANC pathway as a helper of viral replication and recombination by searching for cellular targets that are modified by adenovirus (Ad) infection and are involved in its outcome. This pathway, which is involved in the DNA damage response and checkpoint control, is altered in Fanconi anaemia, a rare cancer predisposition syndrome. We show here that Ad5 infection activates the FANC pathway independent of the classical DNA damage response. Infection with a non-replicating Ad shows that the presence of viral DNA is not sufficient to induce the monoubiquitination of FANCD2 but still activates the DNA damage response coordinated by phospho-NBS1 and phospho-CHK1. E1A expression alone fails to induce FANCD2 monoubiquitination, indicating that a productive viral infection and/or replication is required for FANC pathway activation. Our data indicate that Ad5 infection induces FANCD2 activation to promote its own replication. Specifically, we show that FANCD2 is involved in the recombination process that accompanies viral DNA replication. This study provides evidence of a DNA damage-independent function of the FANC pathway and identifies a cellular system involved in Ad5 recombination.
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Affiliation(s)
- Gioia Cherubini
- University Paris-Sud, UMR8200 CNRS, Institute Gustave Roussy, Villejuif, France
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Connell CM, Shibata A, Tookman LA, Archibald KM, Flak MB, Pirlo KJ, Lockley M, Wheatley SP, McNeish IA. Genomic DNA damage and ATR-Chk1 signaling determine oncolytic adenoviral efficacy in human ovarian cancer cells. J Clin Invest 2011; 121:1283-97. [PMID: 21383502 DOI: 10.1172/jci43976] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2010] [Accepted: 01/12/2011] [Indexed: 12/22/2022] Open
Abstract
Oncolytic adenoviruses replicate selectively within and lyse malignant cells. As such, they are being developed as anticancer therapeutics. However, the sensitivity of ovarian cancers to adenovirus cytotoxicity varies greatly, even in cells of similar infectivity. Using both the adenovirus E1A-CR2 deletion mutant dl922-947 and WT adenovirus serotype 5 in a panel of human ovarian cancer cell lines that cover a 3-log range of sensitivity, we observed profound overreplication of genomic DNA only in highly sensitive cell lines. This was associated with the presence of extensive genomic DNA damage. Inhibition of ataxia telangiectasia and Rad3-related checkpoint kinase 1 (ATR-Chk1), but not ataxia telangiectasia mutated (ATM), promoted genomic DNA damage and overreplication in resistant and partially sensitive cells. This was accompanied by increased adenovirus cytotoxicity both in vitro and in vivo in tumor-bearing mice. We also demonstrated that Cdc25A was upregulated in highly sensitive ovarian cancer cell lines after adenovirus infection and was stabilized after loss of Chk1 activity. Knockdown of Cdc25A inhibited virus-induced DNA damage in highly sensitive cells and blocked the effects of Chk1 inhibition in resistant cells. Finally, inhibition of Chk1 decreased homologous recombination repair of virus-induced genomic DNA double-strand breaks. Thus, virus-induced host cell DNA damage signaling and repair are key determinants of oncolytic adenoviral activity, and promoting unscheduled DNA synthesis and/or impeding homologous recombination repair could potentiate the effects of oncolytic adenoviruses in the treatment of ovarian cancer.
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Affiliation(s)
- Claire M Connell
- Centre for Molecular Oncology and Imaging, Barts Cancer Institute, Queen Mary University of London, London, United Kingdom
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Bagheri N, Shiina M, Lauffenburger DA, Korn WM. A dynamical systems model for combinatorial cancer therapy enhances oncolytic adenovirus efficacy by MEK-inhibition. PLoS Comput Biol 2011; 7:e1001085. [PMID: 21379332 PMCID: PMC3040662 DOI: 10.1371/journal.pcbi.1001085] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2010] [Accepted: 01/18/2011] [Indexed: 01/01/2023] Open
Abstract
Oncolytic adenoviruses, such as ONYX-015, have been tested in clinical trials for currently untreatable tumors, but have yet to demonstrate adequate therapeutic efficacy. The extent to which viruses infect targeted cells determines the efficacy of this approach but many tumors down-regulate the Coxsackievirus and Adenovirus Receptor (CAR), rendering them less susceptible to infection. Disrupting MAPK pathway signaling by pharmacological inhibition of MEK up-regulates CAR expression, offering possible enhanced adenovirus infection. MEK inhibition, however, interferes with adenovirus replication due to resulting G1-phase cell cycle arrest. Therefore, enhanced efficacy will depend on treatment protocols that productively balance these competing effects. Predictive understanding of how to attain and enhance therapeutic efficacy of combinatorial treatment is difficult since the effects of MEK inhibitors, in conjunction with adenovirus/cell interactions, are complex nonlinear dynamic processes. We investigated combinatorial treatment strategies using a mathematical model that predicts the impact of MEK inhibition on tumor cell proliferation, ONYX-015 infection, and oncolysis. Specifically, we fit a nonlinear differential equation system to dedicated experimental data and analyzed the resulting simulations for favorable treatment strategies. Simulations predicted enhanced combinatorial therapy when both treatments were applied simultaneously; we successfully validated these predictions in an ensuing explicit test study. Further analysis revealed that a CAR-independent mechanism may be responsible for amplified virus production and cell death. We conclude that integrated computational and experimental analysis of combinatorial therapy provides a useful means to identify treatment/infection protocols that yield clinically significant oncolysis. Enhanced oncolytic therapy has the potential to dramatically improve non-surgical cancer treatment, especially in locally advanced or metastatic cases where treatment options remain limited. Novel cancer treatment strategies are urgently needed since currently available non-surgical methods for most solid malignancies have limited impact on survival rates. We used conditionally replicating adenoviruses as cancer-fighting agents since they are designed to target and lyse cells with specific aberrations, leaving healthy cells undamaged. Highly malignant cells, however, down-regulate the adenovirus receptor, impairing infection and subsequent cell death. We demonstrated that disruption of the MEK pathway (which is frequently activated in cancer) up-regulated this receptor, resulting in enhanced adenovirus entry. Although receptor expression was restored, disruption of signaling interfered with adenovirus replication due to cell cycle arrest, presenting an opposing trade-off. We developed a dynamical systems model to characterize the response of cancer cells to oncolytic adenovirus infection and drug treatment, providing a means to enhance therapeutic efficacy of combination treatment strategies. Our simulations predicted improved therapeutic efficacy when drug treatment and infection occurred simultaneously. We successfully validated predictions and found that a CAR-independent mechanism may be responsible for regulating adenovirus production and cell death. This work demonstrates the utility of modeling for accurate prediction and optimization of combinatorial treatment strategies, serving as a paradigm for improved design of anti-cancer combination therapies.
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Affiliation(s)
- Neda Bagheri
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America
| | - Marisa Shiina
- Division of Gastroenterology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, California, United States of America
| | - Douglas A. Lauffenburger
- Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America
| | - W. Michael Korn
- Division of Gastroenterology and Helen Diller Family Comprehensive Cancer Center, University of California San Francisco, San Francisco, California, United States of America
- * E-mail:
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Ingemarsdotter CK, Baird SK, Connell CM, Öberg D, Halldén G, McNeish IA. Low-dose paclitaxel synergizes with oncolytic adenoviruses via mitotic slippage and apoptosis in ovarian cancer. Oncogene 2010; 29:6051-63. [PMID: 20729921 PMCID: PMC3007619 DOI: 10.1038/onc.2010.335] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
The microtubule-stabilizing drug paclitaxel has activity in relapsed ovarian cancer. dl922-947, an oncolytic adenovirus with a 24-bp deletion in E1A CR2, replicates selectively within and lyses cells with a dysregulated Rb pathway and has efficacy in ovarian cancer. In the aggressive A2780CP xenograft, combination treatment with weekly dl922-947 and paclitaxel has significantly greater efficacy than either treatment alone and can produce complete tumor eradication in some animals. We investigated the mechanisms of paclitaxel's synergy with dl922-947 in ovarian cancer. The host-cell microtubule network is grossly rearranged and stabilized following adenovirus infection, but paclitaxel does not increase this significantly. Paclitaxel does not synergize by increasing infectivity, viral protein expression or virus release. However, destabilizing the microtubule network with nocodazole reduces viral exit, revealing a novel microtubule-dependent pathway for non-lytic adenoviral exit. dl922-947 can override multiple cell cycle checkpoints but induces cell death by a non-apoptotic mechanism. In combination, dl922-947 and low-dose paclitaxel induces aberrant, multipolar mitoses, mitotic slippage and multinucleation, triggering an apoptotic cell death.
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Affiliation(s)
- C K Ingemarsdotter
- Centre for Molecular Oncology and Imaging, Institute of Cancer, Barts and the London School of Medicine, Queen Mary University of London, London, UK
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Combinatory cytotoxic effects produced by E1B-55kDa-deleted adenoviruses and chemotherapeutic agents are dependent on the agents in esophageal carcinoma. Cancer Gene Ther 2010; 17:803-13. [PMID: 20689571 PMCID: PMC2963731 DOI: 10.1038/cgt.2010.37] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/02/2022]
Abstract
We examined possible combinatory antitumor effects of replication-competent type 5 adenoviruses (Ad) lacking E1B-55kDa molecules (Ad-delE1B55) and chemotherapeutic agents in nine human esophageal carcinoma cells. Ad-delE1B55 produced cytotoxic effects on all the carcinoma cells and the cytotoxicity is not directly linked with the p53 status of the tumors or with the infectivity to respective tumors. A combinatory treatment with Ad-delE1B55 and an anticancer agent, 5-fluorouracil (5-FU), mitomycin C or etoposide, produced greater cytotoxic effects than that with either the Ad or the agent. Administration of 5-FU could minimally inhibit the viral replication and a simultaneous treatment with the Ad and 5-FU achieved better cytotoxicity than sequential treatments. We also confirmed the antitumor effects by the combination of Ad-delE1B55 with 5-FU in vivo. Cisplatin, however, did not achieve the combinatory effects in most of the cells tested. These data indicate that the Ad-delE1B55 produce combinatory antitumor effects with a chemotherapeutic agent irrespective of the administration schedule, but the effects depend on an agent in esophageal carcinoma.
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Ugai H, Wang M, Le LP, Matthews DA, Yamamoto M, Curiel DT. In vitro dynamic visualization analysis of fluorescently labeled minor capsid protein IX and core protein V by simultaneous detection. J Mol Biol 2010; 395:55-78. [PMID: 19853616 PMCID: PMC2787850 DOI: 10.1016/j.jmb.2009.10.034] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2009] [Revised: 09/29/2009] [Accepted: 10/14/2009] [Indexed: 10/20/2022]
Abstract
Oncolytic adenoviruses represent a promising therapeutic medicine for human cancer therapy, but successful translation into human clinical trials requires careful evaluation of their viral characteristics. While the function of adenovirus proteins has been analyzed in detail, the dynamics of adenovirus infection remain largely unknown due to technological constraints that prevent adequate tracking of adenovirus particles after infection. Fluorescence labeling of adenoviral particles is one new strategy designed to directly analyze the dynamic processes of viral infection in virus-host cell interactions. We hypothesized that the double labeling of an adenovirus with fluorescent proteins would allow us to properly analyze intracellular viruses and the fate of viral proteins in a live analysis of an adenovirus as compared to single labeling. Thus, we generated a fluorescently labeled adenovirus with both a red fluorescent minor capsid protein IX (pIX) [pIX monomeric red fluorescent protein 1 (mRFP1)] and a green fluorescent minor core protein V (pV) [pV enhanced green fluorescent protein (EGFP)], resulting in Ad5-IX-mRFP1-E3-V-EGFP. The fluorescent signals for pIX-mRFP1 and pV-EGFP were detected within 10 min in living cells. However, a growth curve analysis of Ad5-IX-mRFP1-E3-V-EGFP showed an approximately 150-fold reduced production of the viral progeny at 48 h postinfection as compared to adenovirus type 5. Interestingly, pIX-mRFP1 and pV-EGFP were initially localized in the cytoplasm and nucleolus, respectively, at 18 h postinfection. These proteins were observed in the nucleus during the late stage of infection, and relocalization of the proteins was observed in an adenoviral-replication-dependent manner. These results indicate that simultaneous detection of adenoviruses using dual-fluorescent proteins is suitable for real-time analysis, including identification of infected cells and monitoring of viral spread, which will be required for a complete evaluation of oncolytic adenoviruses.
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Affiliation(s)
- Hideyo Ugai
- Division of Human Gene Therapy, Department of Medicine, Obstetrics and Gynecology, Pathology, and Surgery, and the Gene Therapy Center, University of Alabama at Birmingham, Birmingham, AL 35294, USA
| | - Minghui Wang
- Division of Human Gene Therapy, Department of Medicine, Obstetrics and Gynecology, Pathology, and Surgery, and the Gene Therapy Center, University of Alabama at Birmingham, Birmingham, AL 35294, USA
| | - Long P. Le
- Massachusetts General Hospital, Pathology Service, 55 Fruit St.-GRJ 249, Boston, MA 02114, USA
| | - David A. Matthews
- Department of Cellular and Molecular Medicine, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK
| | - Masato Yamamoto
- Division of Basic and Translational Research, Department of Surgery, University of Minnesota, Minneapolis, MN 55455, USA
| | - David T. Curiel
- Division of Human Gene Therapy, Department of Medicine, Obstetrics and Gynecology, Pathology, and Surgery, and the Gene Therapy Center, University of Alabama at Birmingham, Birmingham, AL 35294, USA
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Widespread phosphorylation of histone H2AX by species C adenovirus infection requires viral DNA replication. J Virol 2009; 83:5987-98. [PMID: 19321613 DOI: 10.1128/jvi.00091-09] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Adenovirus infection activates cellular DNA damage response and repair pathways. Viral proteins that are synthesized before viral DNA replication prevent recognition of viral genomes as a substrate for DNA repair by targeting members of the sensor complex composed of Mre11/Rad50/NBS1 for degradation and relocalization, as well as targeting the effector protein DNA ligase IV. Despite inactivation of these cellular sensor and effector proteins, infection results in high levels of histone 2AX phosphorylation, or gammaH2AX. Although phosphorylated H2AX is a characteristic marker of double-stranded DNA breaks, this modification was widely distributed throughout the nucleus of infected cells and was coincident with the bulk of cellular DNA. H2AX phosphorylation occurred after the onset of viral DNA replication and after the degradation of Mre11. Experiments with inhibitors of the serine-threonine kinases ataxia telangiectasia mutated (ATM), AT- and Rad3-related (ATR), and DNA protein kinase (DNA-PK), the kinases responsible for H2AX phosphorylation, indicate that H2AX may be phosphorylated by ATR during a wild-type adenovirus infection, with some contribution from ATM and DNA-PK. Viral DNA replication appears to be the stimulus for this phosphorylation event, since infection with a nonreplicating virus did not elicit phosphorylation of H2AX. Infected cells also responded to high levels of input viral DNA by localized phosphorylation of H2AX. These results are consistent with a model in which adenovirus-infected cells sense and respond to both incoming viral DNA and viral DNA replication.
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Prion expression is activated by Adenovirus 5 infection and affects the adenoviral cycle in human cells. Virology 2009; 385:343-50. [PMID: 19138779 DOI: 10.1016/j.virol.2008.12.005] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2008] [Revised: 10/29/2008] [Accepted: 12/04/2008] [Indexed: 01/01/2023]
Abstract
The prion protein is a cell surface glycoprotein whose physiological role remains elusive, while its implication in transmissible spongiform encephalopathies (TSEs) has been demonstrated. Multiple interactions between the prion protein and viruses have been described: viruses can act as co-factors in TSEs and life cycles of different viruses have been found to be controlled by prion modulation. We present data showing that human Adenovirus 5 induces prion expression. Inactivated Adenovirus did not alter prion transcription, while variants encoding for early products did, suggesting that the prion is stimulated by an early adenoviral function. Down-regulation of the prion through RNA interference showed that the prion controls adenovirus replication and expression. These data suggest that the prion protein could play a role in the defense strategy mounted by the host during viral infection, in a cell autonomous manner. These results have implications for the study of the prion protein and of associated TSEs.
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Connell CM, Wheatley SP, McNeish IA. Nuclear survivin abrogates multiple cell cycle checkpoints and enhances viral oncolysis. Cancer Res 2008; 68:7923-31. [PMID: 18829549 DOI: 10.1158/0008-5472.can-08-0817] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Survivin (BIRC5) promotes cell division and survival with roles as chromosomal passenger protein and inhibitor of apoptosis protein (IAP). It is overexpressed in many cancers and is associated with resistance to chemotherapy and radiation. Previously, we showed that expression of survivin within the nucleus of HeLa cells accelerates its degradation and blocks apoptosis inhibition without affecting localization during mitosis. Here, we have investigated the effects of survivin on cell cycle control and potential therapeutic consequences using HeLa and IGROV1 cells expressing wild-type and nuclear-targeted survivin. We show that overexpression of survivin, especially within the nucleus, increases control over G(1)-S checkpoint via increased nuclear accumulation of cyclin D and cyclin-dependent kinase 4 and subsequent pRb phosphorylation. We investigated the influence of survivin on the activity of the E1A CR2-deleted oncolytic adenovirus dl922-947, which depends critically on an aberrant G(1)-S checkpoint. Nuclear expression of survivin augments virus-induced S-phase induction and increases viral protein expression and overall viral replication. There is a consequent increase in antitumor activity both in vitro and in vivo. The increased dl922-947 activity is restricted to malignant cells and is not associated with induction of apoptosis, nor does it rely on the role of survivin as an IAP. In addition, we observe the appearance of a large >or=4N population coincident with multiple mitotic defects in dl922-947-infected cells, both of which are significantly increased by nuclear survivin. This indicates that adenoviral activity is facilitated by abrogation of multiple cell cycle checkpoints and can be enhanced by expression of survivin within the nucleus.
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Affiliation(s)
- Claire M Connell
- Centre for Molecular Oncology and Imaging, Institute of Cancer, Barts and the London School of Medicine and Dentistry, London, United Kingdom
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31
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Ulasov IV, Tyler MA, Rivera AA, Nettlebeck DM, Douglas JT, Lesniak MS. Evaluation of E1A double mutant oncolytic adenovectors in anti-glioma gene therapy. J Med Virol 2008; 80:1595-603. [PMID: 18649343 DOI: 10.1002/jmv.21264] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023]
Abstract
Malignant glioma, in particular glioblastoma multiforme (GBM), represents one of the most devastating cancers currently known and existing treatment regimens do little to change patient prognosis. Conditionally replicating adenoviral vectors (CRAds) represent attractive experimental anti-cancer agents with potential for clinical application. However, early protein products of the wild type adenovirus backbone--such as E1A--limit CRAds' replicative specificity. In this study, we evaluated the oncolytic potency and specificity of CRAds in which p300/CPB and/or pRb binding capacities of E1A were ablated to reduce non-specific replicative cytolysis. In vitro cytopathic assays, quantitative PCR analysis, Western blot, and flow cytometry studies demonstrate the superior anti-glioma efficacy of a double-mutated CRAd, Ad2/24CMV, which harbors mutations that reduce E1A binding to p300/CPB and pRb. When compared to its single-mutated and wild type counterparts, Ad2/24CMV demonstrated attenuated replication and cytotoxicity in representative normal human brain while displaying enhanced replicative cytotoxicity in malignant glioma. These results have implications for the development of double-mutated CRAd vectors for enhanced GBM therapy.
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Affiliation(s)
- Ilya V Ulasov
- The Brain Tumor Center, The University of Chicago, Chicago, Illinois 60637, USA
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32
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Ascencio-Ibáñez JT, Sozzani R, Lee TJ, Chu TM, Wolfinger RD, Cella R, Hanley-Bowdoin L. Global analysis of Arabidopsis gene expression uncovers a complex array of changes impacting pathogen response and cell cycle during geminivirus infection. PLANT PHYSIOLOGY 2008; 148:436-54. [PMID: 18650403 PMCID: PMC2528102 DOI: 10.1104/pp.108.121038] [Citation(s) in RCA: 372] [Impact Index Per Article: 21.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/23/2008] [Accepted: 07/21/2008] [Indexed: 05/18/2023]
Abstract
Geminiviruses are small DNA viruses that use plant replication machinery to amplify their genomes. Microarray analysis of the Arabidopsis (Arabidopsis thaliana) transcriptome in response to cabbage leaf curl virus (CaLCuV) infection uncovered 5,365 genes (false discovery rate <0.005) differentially expressed in infected rosette leaves at 12 d postinoculation. Data mining revealed that CaLCuV triggers a pathogen response via the salicylic acid pathway and induces expression of genes involved in programmed cell death, genotoxic stress, and DNA repair. CaLCuV also altered expression of cell cycle-associated genes, preferentially activating genes expressed during S and G2 and inhibiting genes active in G1 and M. A limited set of core cell cycle genes associated with cell cycle reentry, late G1, S, and early G2 had increased RNA levels, while core cell cycle genes linked to early G1 and late G2 had reduced transcripts. Fluorescence-activated cell sorting of nuclei from infected leaves revealed a depletion of the 4C population and an increase in 8C, 16C, and 32C nuclei. Infectivity studies of transgenic Arabidopsis showed that overexpression of CYCD3;1 or E2FB, both of which promote the mitotic cell cycle, strongly impaired CaLCuV infection. In contrast, overexpression of E2FA or E2FC, which can facilitate the endocycle, had no apparent effect. These results showed that geminiviruses and RNA viruses interface with the host pathogen response via a common mechanism, and that geminiviruses modulate plant cell cycle status by differentially impacting the CYCD/retinoblastoma-related protein/E2F regulatory network and facilitating progression into the endocycle.
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Affiliation(s)
- José Trinidad Ascencio-Ibáñez
- Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, North Carolina 27695, USA.
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33
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Cardoso FM, Kato SEM, Huang W, Flint SJ, Gonzalez RA. An early function of the adenoviral E1B 55 kDa protein is required for the nuclear relocalization of the cellular p53 protein in adenovirus-infected normal human cells. Virology 2008; 378:339-46. [PMID: 18632130 DOI: 10.1016/j.virol.2008.06.016] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2008] [Revised: 06/03/2008] [Accepted: 06/06/2008] [Indexed: 10/21/2022]
Abstract
It is well established that the human subgroup C adenovirus type 5 (Ad5) E1B 55 kDa protein can regulate the activity and concentration of the cellular tumor suppressor, p53. However, the contribution(s) of these functions of the E1B protein to viral reproduction remains unclear. To investigate this issue, we examined properties of p53 in normal human cells infected by E1B mutant viruses that display defective entry into the late phase or viral late mRNA export. The steady-state concentrations of p53 were significantly higher in cells infected by the E1B 55 kDa null mutant Hr6 or three mutants carrying small insertions in the E1B 55 kDa protein coding sequence than in Ad5-infected cells. Nevertheless, none of the mutants induced apoptosis in infected cells. Rather, the localization of p53 to E1B containing nuclear sites observed during infection by Ad5 was prevented by mutations that impair interaction of the E1B protein with p53 and/or with the E4 Orf6 protein. These results indicate that the E1B protein fulfills an early function that correlates efficient entry into the late phase with the localization of E1B and p53 in the nucleus of Ad5-infected normal human cells.
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Affiliation(s)
- F M Cardoso
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias, Universidad Autónoma del Estado de Morelos, Cuernavaca 62209, México
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Guo ZS, Thorne SH, Bartlett DL. Oncolytic virotherapy: molecular targets in tumor-selective replication and carrier cell-mediated delivery of oncolytic viruses. Biochim Biophys Acta Rev Cancer 2008; 1785:217-31. [PMID: 18328829 DOI: 10.1016/j.bbcan.2008.02.001] [Citation(s) in RCA: 90] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2007] [Revised: 02/01/2008] [Accepted: 02/04/2008] [Indexed: 01/13/2023]
Abstract
Tremendous advances have been made in developing oncolytic viruses (OVs) in the last few years. By taking advantage of current knowledge in cancer biology and virology, specific OVs have been genetically engineered to target specific molecules or signal transduction pathways in cancer cells in order to achieve efficient and selective replication. The viral infection and amplification eventually induce cancer cells into cell death pathways and elicit host antitumor immune responses to further help eliminate cancer cells. Specifically targeted molecules or signaling pathways (such as RB/E2F/p16, p53, IFN, PKR, EGFR, Ras, Wnt, anti-apoptosis or hypoxia) in cancer cells or tumor microenvironment have been studied and dissected with a variety of OVs such as adenovirus, herpes simplex virus, poxvirus, vesicular stomatitis virus, measles virus, Newcastle disease virus, influenza virus and reovirus, setting the molecular basis for further improvements in the near future. Another exciting new area of research has been the harnessing of naturally tumor-homing cells as carrier cells (or cellular vehicles) to deliver OVs to tumors. The trafficking of these tumor-homing cells (stem cells, immune cells and cancer cells), which support proliferation of the viruses, is mediated by specific chemokines and cell adhesion molecules and we are just beginning to understand the roles of these molecules. Finally, we will highlight some avenues deserving further study in order to achieve the ultimate goals of utilizing various OVs for effective cancer treatment.
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Affiliation(s)
- Z Sheng Guo
- University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
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Eberle J, Fecker LF, Hossini AM, Kurbanov BM, Fechner H. Apoptosis pathways and oncolytic adenoviral vectors: promising targets and tools to overcome therapy resistance of malignant melanoma. Exp Dermatol 2008; 17:1-11. [PMID: 18095940 DOI: 10.1111/j.1600-0625.2007.00655.x] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Abstract
In the last decades melanoma incidence has been increasing worldwide, while mortality remained on a high level. Until now, there is no suitable therapy for metastasized melanoma, which could lead to a significant increase in overall survival. Apoptosis deficiency is supposed to be a critical factor for therapy resistance, and previous work has characterized the basic mechanisms of apoptosis regulation in melanoma. Genes and strategies suitable for efficient induction of apoptosis in melanoma cells were identified, which are based on proapoptotic Bcl-2 proteins (Bcl-x(S), Bcl-x(AK), Bik/Nbk and Bax) as well as on tumor necrosis factor (TNF)-related death ligands (CD95L/Fas ligand and TNF-related apoptosis-inducing ligand, TRAIL). Proapoptotic genes may be employed in improved gene therapeutic strategies, based on conditional oncolytic adenoviral vectors.
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Affiliation(s)
- Jürgen Eberle
- Department of Dermatology and Allergy, Skin Cancer Center, Charité- Universitätsmedizin Berlin, Berlin, Germany.
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36
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Guan YS, La Z, Yang L, He Q, Li P. p53 gene in treatment of hepatic carcinoma: status quo. World J Gastroenterol 2007; 13:985-992. [PMID: 17373730 PMCID: PMC4146884 DOI: 10.3748/wjg.v13.i7.985] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/30/2006] [Revised: 12/12/2006] [Accepted: 01/16/2007] [Indexed: 02/06/2023] Open
Abstract
Hepatocellular carcinoma (HCC) is one of the 10 most common cancers worldwide. There is no ideal treatment for HCC yet and many researchers are trying to improve the effects of treatment by changing therapeutic strategies. As the majority of human cancers seem to exhibit either abnormal p53 gene or disrupted p53 gene activation pathways, intervention to restore wild-type p53 (wt-p53) activities is an attractive anti-cancer therapy including HCC. Abnormalities of p53 are also considered a predisposition factor for hepatocarcinogenesis. p53 is frequently mutated in HCC. Most HCCs have defects in the p53-mediated apoptotic pathway although they carry wt-p53. High expression of p53 in vivo may exert therapeutic effects on HCC in two aspects: (1) High expression of exogenous p53 protein induces apoptosis of tumor cells by inhibiting proliferation of cells through several biologic pathways and (2) Exogenous p53 renders HCC more sensitive to some chemotherapeutic agents. Several approaches have been designed for the treatment of HCC via the p53 pathway by restoring the tumor suppression function from inactivation, rescuing the mutated p53 gene from instability, or delivering therapeutic exogenous p53. Products with p53 status as the target have been studied extensively in vitro and in vivo. This review elaborates some therapeutic mechanisms and advances in using recombinant human adenovirus p53 and oncolytic virus products for the treatment of HCC.
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Affiliation(s)
- Yong-Song Guan
- Department of Radiology and Oncology, West China Hospital of Sichuan University, Chengdu 610041, Sichuan Province, China.
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