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Biomedical subjects

D T Curiel

Publications and source records attributed to D T Curiel.

At least 181 records · Page 10Linked to original sources
PubMed1997

Targeted gene delivery to Kaposi's sarcoma cells via the fibroblast growth factor receptor.

Kaposi's sarcoma (KS) is a major AIDS-related malignancy associated with significant morbidity and mortality. Current chemotherapeutic regimens are associated with a dismal prognosis. In an effort to develop a new approach to KS treatment, we devised a gene therapy-based adenovirus retargeting schema that redirects the adenovirus to fibroblast growth factor receptors endogenously present on the cell surface of KS cells. By using a bifunctional conjugate consisting of a blocking antiadenoviral knob Fab linked to basic fibroblast growth factor, FGF2, the gene transduction of KS cells was enhanced 7.7-44 fold; recombinant adenoviruses encoding either the firefly luciferase reporter gene, or the herpes simplex thymidine kinase gene, demonstrated quantitative enhancement of expression in the KS cell lines. In this regard, two KS cell lines that were previously refractory to native adenovirus transduction could be successfully transduced by the addition of the conjugate. This study thus addresses the utility of adenoviral retargeting to the FGF receptor in KS cells that are ordinarily transduction refractory to standardized approaches and allows practical development of gene therapy approaches for the treatment of human KS.

Acquired Immunodeficiency Syndrome↗
PubMed1997

A novel gene therapy strategy for elimination of prostate carcinoma cells from human bone marrow.

We report a novel means to purge bone marrow of a specific subset of prostate carcinoma cells based on transductional and genetic selectivity. Using both adenovirus-polylysine-DNA complexes and E1A/B-deleted replication-deficient adenoviruses, we have demonstrated a transductional preference of these vectors for the prostate carcinoma cell lines DU 145, LNCaP, and PC-3 over primary human bone marrow cells and the leukemia cell line KG-1. We have also shown a genetic selectivity of an anti-erbB-2 intracellular single-chain antibody (sFv) encoding adenovirus, Ad21, for the erbB-2-positive prostate carcinoma cell lines DU 145 and LNCaP. Delivery of Ad21 resulted in cytotoxicity to the DU 145 and LNCaP, but not PC-3, cell lines and reduced the clonogenic capacity of DU 145 cells cultured alone or mixed with various ratios of irradiated human bone marrow. Finally, quantitative, competitive reverse transcription polymerase chain reaction (QC-RT-PCR) analysis demonstrated that Ad21 could effectively reduce DU 145 and erbB-2-positive primary prostate tumor contamination in bone marrow cultures. Delivery of Ad21 had no effect on the ability of progenitor cells to form colonies. These results suggest that an anti-erbB-2 sFv-encoding adenoviral vector is efficacious for removal of erbB-2-positive prostate carcinoma cells from human bone marrow, and demonstrates a novel method for ex vivo genetic purge of malignant cells from bone marrow for autologous bone marrow transplantation (ABMT) therapy.

Adenoviridae↗
PubMed1997

Adenoviral vector infection of the human exocrine pancreas.

OBJECTIVE: To determine if a viable cadaveric pancreas might be used to study viral transfection efficacy in a manner precisely mimicking in vivo human studies. DESIGN: Ex vivo gene transfer to an intact human pancreatic duct. SETTING: Molecular biology laboratory and organ procurement center. INTERVENTION: The recombinant adenoviral vector that contains the Escherichia coli beta-galactosidase (LacZ) gene driven by the human cytomegalovirus promoter, ie, AdCMVLacZ, was used to transfect the epithelial cells of the pancreatic ductal system. A human pancreas (150 g wt/wt) procured for transplantation, but subsequently found unsuitable, was used for the study. The splenic, superior mesenteric arteries and portal vein were cannulated and perfused in a heat-controlled organ procurement perfusion system. A segment of vascularized, perfused distal pancreatic duct was isolated with a balloon occlusion catheter. The recombinant adenoviral vector AdCMVLacZ was introduced into the lumen of the distal segment of the pancreatic duct and incubated for 6 hours at 25 degrees C. The proximal segment of the pancreatic duct was not exposed to the vector and served as control tissue. Tissue was harvested and processed for evaluation of beta-galactosidase activity. RESULTS: Adenoviral vector-infected pancreatic ducts exhibited intense blue staining, indicative of reporter gene expression in the epithelial cells of the pancreatic duct. The phenotype of these cells was confirmed by immunohistochemical studies using anti-annexin III polyclonal antibody. Control tissue not exposed to the adenoviral vector was subjected to an identical analysis and did not reveal evidence of expression of the reporter gene. CONCLUSIONS: This study demonstrates the first successful transfection of epithelial cells of the pancreatic duct from normal human pancreas with a recombinant adenovirus. This system will provide not only information on the efficacy of transfection but also a novel gene therapeutic approach to target pancreatic ductal adenocarcinoma.

Adenoviruses, Human↗
PubMed1997

Transductional efficacy and safety of an intraperitoneally delivered adenovirus encoding an anti-erbB-2 intracellular single-chain antibody for ovarian cancer gene therapy.

We have previously shown that adenoviral-mediated delivery of an anti-erbB-2 intracellular single-chain antibody (sFv) causes specific cytotoxicy in erbB-2-overexpressing ovarian carcinoma cells. Furthermore, intraperitoneal delivery of the anti-erbB-2 sFv enhances survival and reduces tumor burden in a xenograft model of human ovarian carcinoma in SCID mice. These findings have led to an RAC-approved Phase I clinical trial for patients with ovarian cancer. In this report, we show that expression of the anti-erbB-2 sFv could be readily detected in target tumor cells by in situ hybridization methodology. PCR analysis of DNA extracted from various murine tissues demonstrated that the anti-erbB-2 sFv remained localized to the peritoneum. Delivery of the sFv to the non-erbB-2-overexpressing REN mesothelial and Hep G2 hepatocellular carcinoma cell lines was not deleterious to either one, affirming the tumor specificity of this gene therapy strategy. In addition, histopathological analysis of various tissues showed that adenoviral-mediated delivery of the anti-erbB-2 sFv to immunocompetent mice with either primary exposure or previous vector challenge at different doses produced no abnormal changes when compared to untreated animals. These findings suggest that adenoviral-mediated delivery of the anti-erbB-2 sFv in a human context can be effectively assayed, is potentially free of vector-associated toxicity, and retains biologic utility based on tumor specificity.

Adenoviridae↗
PubMed1997

Gene therapy and ovarian cancer: a review.

OBJECTIVE: To provide the obstetrician-gynecologist with the basic concepts of gene therapy, an overview of cancer gene therapy treatment strategies, and a summary of currently approved human gene therapy protocols for ovarian carcinoma. DATA SOURCES: A computerized search of articles published through December 1995 was performed on the MEDLINE data base. Additional sources were identified through cross-referencing. METHODS OF STUDY SELECTION: All identified references were reviewed with particular attention to their relevance to gene therapy for ovarian cancer. TABULATION, INTEGRATION, AND RESULTS: Each reference was reviewed to determine the relevant contribution to the fundamental science of gene therapy. Particular attention was paid to those studies that offered a rational strategy that might contribute to therapy for ovarian malignancy. CONCLUSION: Advances in molecular biology, immunology, and virology have enabled the concept of gene therapy for neoplastic disorders to become a reality. Given the lack of effective conventional therapy, those patients with recurrent or refractory ovarian cancer should be considered for currently approved investigational gene therapy protocols.

Adjuvants, Immunologic↗
PubMed1997

Strategies to accomplish targeted gene delivery to muscle cells employing tropism-modified adenoviral vectors.

We are developing strategies to modify the tropism of adenoviral vectors to accomplish targeted transduction of muscle cells for DMD gene therapy. In one approach, we have introduced targeting ligands into the adenovirus fibre, which mediates the binding of the virus Ad5 to the primary cellular receptor. In order to incorporate these fibre-ligand fusions into recombinant adenoviral vectors, we have employed a method based upon homologous DNA recombination between a fibre-deleted, propagation-defective rescue plasmid and a shuttle plasmid encoding a variant fibre. To date, we have generated an adenoviral vector containing chimeric fibres composed of the tail and shaft domains of adenovirus serotype 5 and the knob domain of serotype 3. This modification altered the receptor recognition profile of the virus containing the fibre chimera. In an alternative approach to the generation of a targeted adenoviral vector, we conjugated folate to the neutralising Fab fragment of an anti-fibre monoclonal antibody. This Fab-folate conjugate was shown to redirect adenoviral infection of target cells via the folate receptor at a high efficiency. These studies suggest that it will be possible to achieve our goal of deriving targeted adenoviral vectors for muscle cell-specific gene delivery in vivo.

Adenoviridae↗
PubMed1997

In vitro and in vivo gene delivery mediated by a synthetic polycationic amino polymer.

A synthetic polyamino polymer with a glucose backbone was used for gene transfer in vitro and in vivo. Gene transfer in vitro to various human carcinoma cell lines was achieved with an efficiency superior to a commercially available cationic liposome preparation. The polymer was resistant to inhibition by serum, which allowed for efficient gene transfer in vivo. Direct Intracranial tumor injection using this reagent resulted in reporter gene expression levels comparable to those achieved by a recombinant adenoviral vector. Thus, this compound represents a new class of agent that may have broad utility for gene transfer and gene therapy applications.

Animals↗
PubMed1997

Stable in vivo gene transduction via a novel adenoviral/retroviral chimeric vector.

Gene therapy to correct defective genes requires efficient gene delivery and long-term gene expression. The available vector systems have not allowed the simultaneous achievement of both goals. We have developed a chimeric viral vector system that incorporates favorable aspects of both adenoviral and retroviral vectors. Adenoviral vectors induce target cells to function as transient retroviral producer cells in vivo. The progeny retroviral vector particles are then able to stably transduce neighboring cells. In this system, the nonintegrative adenoviral vector is rendered functionally integrative via the intermediate generation of a retroviral producer cell. The chimeric vectors may allow realization of the requisite goals for specific gene-therapy applications.

Adenoviridae↗
PubMed1997

An intracellular anti-erbB-2 single-chain antibody is specifically cytotoxic to human breast carcinoma cells overexpressing erbB-2.

We previously demonstrated that delivery of a gene encoding an anti-erbB-2 intracellular single-chain antibody (sFv) resulted in down-regulation of cell surface erbB-2 levels and induction of apoptosis in erbB-2 overexpressing ovarian cancer cells. Based upon these findings, we hypothesized that human breast carcinomas overexpressing erbB-2 would be similarly affected by this genetic intervention. We evaluated the phenotypic effects resulting from intracellular expression of the anti-erbB-2 sFv on the human breast cancer cell lines MDA-MB-361, SK-BR-3, BT-474, MCF-7 and MDA-MB-231. Recombinant adenoviruses encoding either a reporter gene (AdCMVLacZ) or the endoplasmic reticulum (ER) directed anti-erbB-2 sFv (Ad21) were delivered to various breast cancer cell lines. Cell viability was determined by a proliferation assay and fluorescent microscopy allowed visualization of apoptotic cells. An erbB-2 ELISA quantified the endogenous erbB-2 levels of each cell line. The anti-erbB-2 sFv-encoding-adenovirus, Ad21, but not the beta-galactosidase encoding adenovirus, AdCMVLacZ, was cytotoxic to > 95% of the tumor cells in the MDA-MB-361 and SK-BR-3 lines, and > 60% of the tumor cells in the BT-474 line. In marked contrast, the MCF-7 and MDA-MB-231 cell lines showed no change in the rate of cell proliferation following this treatment. The cytotoxic effects generated in the first three lines were a consequence of the induction of apoptosis by the anti-erbB-2 sFv. An ELISA specific for erbB-2 showed that the breast cancer cell lines most susceptible to the anti-erbB-2 sFv, MDA-MB-361, SK-BR-3 and BT-474, overexpressed the erbB-2 protein while the cell lines demonstrating no response to the anti-erbB-2 sFv, MCF-7 and MDA-MB-231, expressed the lowest levels of erbB-2. These results demonstrate that targeted killing of erbB-2 overexpressing cells via intracellular knockout can be accomplished in the context of breast carcinoma. Furthermore, erbB-2 levels in breast tumor cells may be predictive of their sensitivity to sFv-mediated killing. The ability to accomplish selective cytotoxicity of breast cancer cell lines overexpressing the erbB-2 tumor marker should allow for derivation of clinical gene therapy strategies for breast cancer utilizing this approach.

Adenoviridae↗
PubMed1997

Immunization with a plasmid expressing pneumococcal surface protein A (PspA) can elicit protection against fatal infection with Streptococcus pneumoniae.

Pneumococcal surface protein A (PspA) is a protectioneliciting protein of Streptococcus pneumoniae. We observed that immunization of BALB/c mice with a plasmid expressing PspA significantly protected the mice from lethal challenge with S. pneumoniae when compared to control mice that received injections of the plasmid vector alone. The plasmid construct expressing PspA has been designated pKSD2601. Mice immunized intramuscularly with pKSD2601 had a mean log of colony-forming units of 2.67 +/- 0.25 pneumococci circulating in their blood at 24 h after challenge as compared with control mice that had a mean log of colony-forming units of 4.95 +/- 0.59. Those mice with lower numbers of pneumococci subsequently survived the challenge. Given the quantitative nature and ultimate end point (ie live versus dead) our mouse model should be useful in working out optimum expression of bacterial genes for DNA immunization.

Animals↗
PubMed1997

Gene therapy strategies for asthma.

Asthma is a disease which is increasing in frequency and severity despite the incontestable advances in the understanding of its physiology and treatment. New therapeutic strategies are therefore required and at the time when a new treatment, that of gene therapy, is giving hope for the treatment of so far incurable illnesses, it would seem to us to be useful to discuss the possible use of this treatment in severe asthma and in particular in the case of the most severe asthmatic patients requiring continuous oral steroid treatment (referred to as steroid-dependent). These patients are those who require particular medical attention, they often need to be admitted to hospital and they represent over 50% of the total costs associated with asthma. No satisfactory alternative therapy is currently available. In this regard, a variety of gene-based strategies have recently been proposed for inflammatory and immunologic disorders. It is thus the purpose of this review to consider its application to asthma therapy, although there are very few published works that directly bear on gene therapy linked to asthma. After a brief outline of the epidemiology of asthma, its genetics and the available animal models, we will focus on its immunopathology slanting the discussion towards the possibility of a gene-based treatment.

Asthma↗
PubMed1997

Incorporation of beta-globin untranslated regions into a Sindbis virus vector for augmentation of heterologous mRNA expression.

Polynucleotide immunization has been employed as a means of inducing immune responses through the introduction of antigen-encoding DNA. While immunization against specific tumor antigens may be achieved through this strategy, various candidate tumor antigens may not be approached via DNA-based vaccines as they represent transforming oncogenes. As an alternative approach, we have explored the utility of mRNA vectors for polynucleotide immunization. The transient expression achieved by mRNA may provide an efficient and safe system for stimulating immune responses to tumor-specific antigens. Our previous work demonstrated that a self-replicating RNA enhances the magnitude and duration of transgene expression for this application. Here we further modify the vector for optimal use in gene therapy through the incorporation of untranslated regions flanking the encoded transgene. The beta-globin 5' and 3' untranslated regions (UTRs) were inserted directly flanking the luciferase gene in both nonreplicative and replicative RNA constructs. In both cases, elevated and prolonged levels of luciferase expression were detected from the beta-globin UTR-flanked luciferase as compared to luciferase without these sequences. These modifications improve the ability of replicative RNA vectors to produce high, yet transient transgene expression for cancer immunotherapy strategies.

Animals↗
PubMed1997

Gene therapy strategies for AIDS-related malignancies.

AIDS-related malignancies (ARM) include AIDS-defining cancers such as Kaposi's sarcoma, non-Hodgkin's lymphoma and cervical carcinoma. In addition, certain other malignancies are also increased with human immunodeficiency virus (HIV) infection. New antiretroviral agents and better prophylaxis and treatment of HIV-related opportunistic infections are prolonging the lives of HIV-infected individuals. There will thus likely be a continued rise in the incidence and prevalence of ARM in the long term, even if effective antiretroviral and other AIDS-related therapies reduce their appearance in the short term. There are presently no curative regimens for the common ARM, with the possible exception of some lymphomas. Survival is shortened by most, and treatment is often toxic and poorly tolerated. Gene therapies may thus offer a useful adjunct to conventional treatment strategies for selected ARM. Although some gene therapy strategies may work well in the HIV setting, the chronic viral infection, immunodeficient status of the host, the tendency for HIV-infected individuals to have altered drug metabolism and an increased rate of adverse drug reactions will likely present special challenges. This review summarizes the state-of-the-art in the fledgling field of gene therapy for ARM, and explores areas for future research.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Use of a novel cross-linking method to modify adenovirus tropism.

Recombinant adenovirus (Ad) vectors can accomplish efficient in vivo gene transfer and thus are important in the context of a variety of gene therapy approaches. The cellular receptor for the Ad fiber knob is prevalent on a number of normal tissues which undermines the targeting of Ad to specific tumor cells. Therefore, the ablation of native Ad tropism and the introduction of novel Ad tropism are both necessary to target Ad vectors specifically to tumors. In this study, we have developed a flexible method for cross-linking the Fab fragment of a neutralizing anti-knob monoclonal antibody (1D6.14) to a cell receptor ligand. The cross-linking moieties are complementary low molecular weight recognition units, similar in concept to the avidin-biotin system. For proof of concept, we cross-linked 1D6.14 Fab to the basic fibroblast growth factor (FGF2). The Fab and FGF2 conjugates were synthesized and characterized both structurally and functionally. The conjugates were then complexed with an adenovirus vector carrying firefly luciferase (AdCMVLuc) and the resulting complex used to show infection of a number of tumor cell lines expressing FGF receptors. This cross-linking system should provide a rapid and convenient method of conjugating various ligands to the Fab fragment for targeting Ad vectors to different types of tumors.

Adenoviridae↗