Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.5.4.1 (cytosine deaminase)
 747 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Gene therapy of cancer is a novel approach with the potential to selectively eradicate tumour cells, whilst sparing normal tissue from damage. In particular, gene-directed enzyme prodrug therapy (GDEPT) is based on the delivery of a gene that encodes an enzyme which is non-toxic per se, but is able to convert a prodrug into a potent cytotoxin. Several GDEPT systems have been investigated so far, demonstrating effectiveness in both tissue culture and animal models. Based on these encouraging results, phase I/II clinical trials have been performed and are still ongoing. The aim of this review is to summarise the progress made in the design and application of GDEPT strategies. The most widely used enzyme/prodrug combinations already in clinical trials (e.g., herpes simplex 1 virus thymidine kinase/ganciclovir and cytosine deaminase/5-fluorocytosine), as well as novel approaches (carboxypeptidase G2/CMDA, horseradish peroxidase/indole-3-acetic acid) are described, with a particular attention to translational research and early clinical results.
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PMID:Gene directed enzyme/prodrug therapy of cancer: historical appraisal and future prospectives. 1124 46

The potential of gene therapy to treat premalignant disease or recurrent cancer has not been investigated. The goal of the present investigation was to explore the efficacy of pro-drug-mediated, suicide gene therapy as a strategy to treat incipient neoplasia in stratified squamous epithelium. To test this strategy, a tissue model of premalignancy was generated by mixing normal human keratinocytes (NHK) that express the bacterial cytosine deaminase gene (CD) with premalignant keratinocytes which have been genetically marked with the bacterial gene for beta-galactosidase (II-4-beta-gal) in skin-like organotypic cultures. Preliminary studies in monolayer cultures demonstrated that CD-transduced NHK (NHK/CD) efficiently expressed the transgene and deaminated the pro-drug 5-fluorocytosine (5FC) to the toxic product 5-fluorouracil (5FU). The capacity of NHK/CD to kill II-4-beta-gal cells through bystander effect was assayed in both submerged culture and in the organotypic model of premalignancy. In submerged cultures, it was found that CD-mediated killing of II-4-beta-gal cells did not require cell-cell contact and that the LD(50) of 5FC for efficient bystander killing of II-4-beta-gal was 0.5 mM. When this concentration of pro-drug was used in organotypic cultures, a significant number of dysplastic II-4-beta-gal cells were eliminated from the tissue. Bystander killing of II-4-beta-gal cells was related to the number of NHK/CD present. These findings demonstrated that potentially malignant keratinocytes could be eliminated from a dysplastic tissue through activation of pro-drug and killing of adjacent cells through the bystander effect. By establishing an in vitro model to eliminate premalignant cells using suicide gene therapy, these studies provide a new approach for the treatment of incipient cancer as it develops, thereby preventing invasive disease.
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PMID:Suicide gene therapy for premalignant disease: a new strategy for the treatment of intraepithelial neoplasia. 1131 95

A 2.4-kb truncated L-plastin promoter was inserted either 5' to the LacZ gene (Ad-Lp-LacZ) or 5' to the cytosine deaminase (CD) gene (Ad-Lp-CD) in a replication-incompetent adenoviral vector backbone. Infectivity and cytotoxicity experiments with the LacZ and CD vectors suggested that the L-plastin promoter-driven transcriptional units were expressed at much higher levels in explants of ovarian cancer cells from patients and in established ovarian or bladder cancer cell lines than they were in normal peritoneal mesothelial cells from surgical specimens, in organ cultures of normal ovarian cells, or in the established CCD minimal deviation fibroblast cell line. Control experiments showed that this difference was not attributable to the lack of infectivity of the normal peritoneal cells, the normal ovarian cells, or the minimal deviation CCD fibroblast cell line, because these cells showed expression of the LacZ reporter gene when exposed to the replication-incompetent adenoviral vector carrying the cytomegalovirus (CMV)-driven LacZ gene (Ad-CMV-LacZ). The Ovcar-5 and Skov-3 ovarian cancer cell lines exposed to the Ad-Lp-CD adenoviral vector were much more sensitive to the prodrug 5-fluorocytosine (5FC), which is converted from the 5FC prodrug into the toxic chemical 5-fluorouracil, than was the CCD minimal deviation fibroblast cell line after exposure to the same vector. A mouse xenograft model was used to show that the Ad-Lp-CD vector/5FC system could prevent engraftment of ovarian cancer cells in nude mice. Finally, injection of the Ad-Lp-CD vector into s.c. tumor nodules generated a greater reduction of the size of the tumor nodules than did injection of the Ad-CMV-LacZ vectors into tumor nodules. The Ad-Lp-CD vectors were as suppressive to tumor growth as the Ad-CMV-CD vectors. These results suggest that an adenoviral vector carrying the CD gene controlled by the L-plastin promoter (Ad-Lp-CD) may be of potential value for the i.p. therapy of ovarian cancer.
Cancer Res 2001 Jun 01
PMID:The use of the L-plastin promoter for adenoviral-mediated, tumor-specific gene expression in ovarian and bladder cancer cell lines. 1138 68

The presence of severe hypoxia and necrosis in solid tumors offers the potential to apply an anaerobic bacterial enzyme/prodrug approach in cancer treatment. In this context the apathogenic C. acetobutylicum was genetically engineered to express and secrete E. coli cytosine deaminase (CDase). Considerable levels of functional cytosine deaminase were detected in lysates and supernatants of recombinant C acetobutylicum cultures. After administration of the recombinant Clostridium to rhabdomyosarcoma bearing rats used as a model, cytosine deaminase could be detected at the tumor site. Moreover, following administration of the vascular targeting agent combretastatin A-4 phosphate significantly increased levels of cytosine deaminase were detected at the tumor site as a consequence of enlarged tumor necrosis and subsequently improved growth of C. acetobutylicum. The results provide evidence for the potential application of Clostrisdium-based therapeutic protein transfer to tumors in anticancer therapy.
Cancer Gene Ther 2001 Apr
PMID:Specific targeting of cytosine deaminase to solid tumors by engineered Clostridium acetobutylicum. 1139 82

Liver metastasis is the most serious event for physicians and surgeons treating patients with colorectal cancer. Gene therapy is expected to become a novel strategy to prevent liver metastasis. Four types of clinical studies are currently underway: 1) suicide-gene therapy with the cytosine deaminase gene; 2) immune gene therapy with cytokine (inter leukin-2) or major histocompatibility complex class I gene HLA-B7; 3) tumor suppressor gene p53 therapy; and 4) lysis of p53 mutant cancer cells with E1B55k-deleted adenovirus (Onyx-015). Basic research provided several candidates for the liver metastasis-associated genes, including MMP7, DCC, CDC25B, E-cadherin, CD44, vascular endothelial growth factor, etc. There is an alternative approach to liver metastasis, which attempts to introduce a specific gene such as cytosine deaminase and TIMP-2 into the hepatocytes but not into the tumor itself. This concept is based on results showing that hepatocytes can incorporate genes more readily than cancer cells can. Recently, mutant virus therapy has been developed, which includes Onyx-015, adenovirus dl922-947, and mutant-type herpes simplex virus. These mutant types of virus specifically proliferate in the cancer cells and result in their lysis. In the future, development of gene delivery systems that are powerful and specific to cancer type is essential.
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PMID:[Future scope for gene therapy for liver metastasis of colon cancer]. 1139 6

Infection of tumor cells by herpes simplex virus 1 (HSV-1) results in cell destruction and production of progeny virion in a process referred to as viral oncolysis. In this study, an HSV-1 mutant (HSV1yCD) was engineered such that the viral ribonucleotide reductase gene is disrupted by sequences encoding yeast cytosine deaminase, which efficiently metabolizes the prodrug 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU). HSV1yCD-infected cells convert 5-FC to 5-FU, which enhances cytotoxicity without significantly reducing viral replication and oncolysis. Oncolysis by a replicating HSV-1 mutant combined with therapeutic transgene delivery represents a new paradigm; HSV1yCD-infected cells are destroyed by viral replication, and uninfected cells are subjected to bystander killing from both progeny virion and extracellular diffusion of 5-FU. In contrast, HSV1yCD-mediated bioactivation of another prodrug, ganciclovir, impairs viral replication. HSV1yCD administered into the portal venous system replicates preferentially in liver metastases rather than normal liver. The anti-neoplastic activity of HSV1yCD combined with systemic 5-FC administration is greater than that achieved with HSV-1 replication alone. Combination oncolysis and prodrug bioactivation leads to significant prolongation of survival in mice with diffuse liver metastases.
Cancer Res 2001 Jul 15
PMID:Multimodality therapy with a replication-conditional herpes simplex virus 1 mutant that expresses yeast cytosine deaminase for intratumoral conversion of 5-fluorocytosine to 5-fluorouracil. 1145 90

Tumor cells that express a fusion gene of Escherichia coli cytosine deaminase (CD) and herpes simplex virus type 1 thymidine kinase (TK) sequences activate and are subsequently killed by the nontoxic prodrugs 5-fluorocytosine and ganciclovir. We have previously developed a recombinant adenovirus containing the CD-TK fusion gene controlled by the human inducible heat shock protein 70 promoter so that heat at 41 degrees C for 1 hour induces therapeutic gene expression. This adenovirus effectively transduces heat-inducible expression of the CD-TK gene into human prostate carcinoma cells. However, because a limited number of cells in a tumor can actually be infected, we created a replicating adenoviral vector to increase CD-TK gene expression. This vector is a replication-competent, E1B-attenuated adenoviral vector containing the hsp70 promoter-driven CD-TK gene (Ad.E1A(+)HS-CDTK). When human prostate adenocarcinoma DU-145 cells (mutant p53) were infected with the virus at a multiplicity of infection (MOI) of 1 or 10, the viral replication was detected within 2 days at both MOIs. Similar results were observed in human colorectal carcinoma CX-1 cells. When DU-145 cells were infected with the virus at an MOI of 10, incubated for 24 hours, heated at 41 degrees C for 4 hours, and then harvested 20 hours later, Western blot analysis demonstrated that this virus successfully produced viral E1A proteins and heat shock stimulated the CD-TK gene expression by 12.3-fold. In addition, Ad.E1A(+)HS-CDTK effectively suppressed cell proliferation by viral cytopathic effect). Unlike with a replication-incompetent virus (Ad.HS-CDTK), the cytopathic effect of the virus and cytotoxicity in the presence of the prodrugs were still observed even at low MOI (MOI=1.0).
Cancer Gene Ther 2001 Jun
PMID:Replicating adenoviral vector-mediated transfer of a heat-inducible double suicide gene for gene therapy. 1149 59

Tumor-specific gene delivery is crucial to achieving successful effects in suicide gene therapy. Carcinoembryonic antigen (CEA) promoter has been widely used for this purpose, but the expression level of tumor-specific promoters such as CEA promoter is generally low. In the previous study, we used the Cre/loxP system and showed that LacZ expression by the CEA promoter was remarkably enhanced and maintained its specificity using the Cre/loxP regulation system. In this study, the Cre/loxP system was first applied to augmentation of selective expression of the cytosine deaminase (CD) gene as a suicide gene therapy in CEA-producing cells. The double infection with AxCEANCre expressing Cre recombinase under the control of the CEA promoter and AxCALNLCD expressing the CD gene under the control of the CAG promoter by the Cre switching system rendered CEA-producing tumor cells 13-fold more sensitive to 5-fluorocytosine (5-FC) compared with the single infection with AxCEACD expressing CD gene driven by the CEA promoter. The therapeutic efficacy of the enhanced CD/5-FC suicide gene therapy was evaluated in orthotopic implantation models of human gastric carcinoma. Adenovirus vectors (1 x 10(9) plaque-forming units) were administered i.p. into mice three times, and then 5-FC was administered i.p. for the next 10 days. Tumor volume and weight in mice treated with AxCEANCre and AxCALNLCD/5-FC were significantly reduced as compared with those in mice treated not only with Mock (AxCALacZ) but also with AxCEACD/5-FC (P < 0.0001). This beneficial effect on tumor burden was also reflected in the overall survival. The survival periods of the mice treated with AxCEANCre and AxCALNLCD/5-FC were longer than those of mice treated with Mock or AxCEACD/5-FC (P < 0.01). These results suggested that application of the Cre/loxP system could provide a new approach for enhanced selective suicide gene therapy of CD/5-FC for the treatment of advanced gastric carcinoma.
Cancer Res 2001 Aug 15
PMID:Carcinoembryonic antigen-specific suicide gene therapy of cytosine deaminase/5-fluorocytosine enhanced by the cre/loxP system in the orthotopic gastric carcinoma model. 1150 67

Targeting of colorectal liver metastases by regional gene therapy was tested in a clinically relevant syngeneic model. First, the CEA-CD-113 retroviral vector containing the cytosine deaminase gene controlled by the CEA specific tumour cell promoter, was shown in vitro to convert 5-fluorocytosine to 5-fluorouracil, resulting in cancer cell killing with a large bystander effect. Second, 10 days after the establishment of liver metastases, retroviral vectors were delivered to the liver by hepatic artery injection. After 5-fluorocytosine administration for 7 days, most surface metastases disappeared and tumour volumes were suppressed up to 8.2-fold. The results support the development of this approach for patient treatment.
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PMID:Hepatic intra-arterial delivery of a retroviral vector expressing the cytosine deaminase gene, controlled by the CEA promoter and intraperitoneal treatment with 5-fluorocytosine suppresses growth of colorectal liver metastases. 1150 57

An attenuated strain of Salmonella typhimurium, designated VNP20009, was generated by deletion of the msbB and purl genes. When VNP20009 was administered intravenously (IV) to mice bearing spontaneous, syngeneic, or human xenograft tumors, the bacteria accumulated preferentially within the extracellular components of tumors, forming tumor-to-normal tissue ratios exceeding 300-1000 to 1. NVP20009 was administered safely at doses up to 2.5 x 10(9) cfu/kg in monkey toxicology studies. Based on the preclinical data, VNP20009 entered Phase I human clinical trials in November 1999, and has now been administered to >45 patients by IV or direct intratumoral injection. By the intratumoral route, a maximum tolerated dose has not been reached, and dose escalation continues past the current dose level of 4 x 10(7)/m2. Furthermore, VNP20009 persisted in injected tumors for at least 2 weeks in 8/11 patients treated to date. By 30-min IV administration, a maximum tolerated dose (MTD) of 3 X 10(8) cfu/m2 has been established. In all patients treated to date, VNP20009 was not shed in urine or stool. VNP20009 has been further modified by chromosomal insertion of an E. coli cytosine deaminase (CD) gene at the deltamsbB locus which, when expressed, converts 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU). The CD containing VNP20009 was designated TAPET-CD or VNP20029. TAPET-CD had similar efficacy and safety in murine tumor models and similar safety profiles in animal toxicology studies, compared to its parent VNP20009. Specifically, TAPET-CD had a reduced virulence of >10,000 fold, when compared to the wild-type Salmonella strain. It was well-tolerated at doses up to 2 x 10(6) cfu/mouse and 1 X 10(10) cfu/monkey. After an IV or direct tumor injection to tumor-bearing mice, TAPET-CD reached tumor levels as high as 10(8)-10(9) cfu/gm. When compared to the accumulation in liver or spleen, the normal tissues with the greatest colonization of TAPET-CD, tumor-to-normal tissue ratios of TAPET-CD were 300-1000 to 1. TAPET-CD also caused tumor growth inhibition of >90% in several murine tumor models. When 5-FC was administered by intraperitoneal (IP) injection once or 3 times daily to tumor-bearing mice that had been pre-treated with TAPET-CD, high levels of 5-FU (reaching 20-40 microM/g) were detected in the tumor, with low or undetectable 5-FU levels in normal tissues (e.g., spleen, liver, etc.). Furthermore, co-administration of 5-FC and TAPET-CD in 4 different murine tumor models enhanced anti-tumor activity compared to the significant anti-tumor activity of TAPET-CD alone, further confirming the benefit of the inserted CD gene. On the basis of the preclinical data, a Phase I clinical protocol is proposed in which advanced cancer patients will receive TAPET-CD by direct intratumoral injection and 5-FC. TAPET-CD will be administered on day 1. 5-FC will be given orally q8h daily beginning day 4 or when all toxicities of TAPET-CD have resolved to < or = grade 1, and continued for 14 days. Tumor tissues will be sampled to verify TAPET-CD colonization and to measure intratumoral 5-FC and 5-FU concentrations on day 8. A second sample of tumor tissue will be obtained between day 15-17 in selected patients to confirm the persistence of high levels of bacteria in tumor and to obtain a second measurement of 5-FC and 5-FU intra-tumoral concentrations. The TAPET-CD/5-FC treatment cycle will be repeated in appropriate patients on day 29.
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PMID:A phase I trial of genetically modified Salmonella typhimurium expressing cytosine deaminase (TAPET-CD, VNP20029) administered by intratumoral injection in combination with 5-fluorocytosine for patients with advanced or metastatic cancer. Protocol no: CL-017. Version: April 9, 2001. 1152 49


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