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D T Curiel

Publications and source records attributed to D T Curiel.

At least 55 records · Page 3Linked to original sources
PubMed2001

Human immunodeficiency virus type 1-mediated syncytium formation is compatible with adenovirus replication and facilitates efficient dispersion of viral gene products and de novo-synthesized virus particles.

Conditionally replicative adenovirus (CRAd) vectors are designed for specific oncolytic replication in tumor tissues with concomitant sparing of normal cells. As such, CRAds offer an unprecedented level of anticancer potential for malignancies that have been refractory to previous cancer gene therapy interventions. CRAd efficacy may, however, be compromised by inefficient dispersion of the replicating vector within the tumor tissue. To address this issue, we evaluated the utility of a fusogenic membrane glycoprotein (FMG), which induces the fusion of neighboring cellular membranes to form multinucleated syncytia. We hypothesized that the FMG-mediated syncytia would facilitate dispersion of the adenovirus (Ad) gene products and viral progeny. To test this, human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins, which induce syncytia in the presence of CD4+ target cells, were expressed by an Ad (Ad5HIVenv) in permissive (CD4-positive) and nonpermissive (CD4-negative) cell lines. After validating this Ad-FMG model, the efficiency of Ad replication in the presence or absence of syncytia was evaluated. The results demonstrated that syncytium formation was compatible with Ad replication and dramatically increased the dispersion of virus gene products within the cytoplasm of the syncytia as well as viral particles in the nuclei of the syncytial mass. Moreover, progeny virions were released more efficiently from syncytia compared with nonsyncytial cells. These data demonstrate the utility of FMGs as a dispersion agent and suggest that FMGs can improve the efficacy of CRAd gene therapy.

Adenoviruses, Human↗
PubMed2001

Heat shock and heat shock protein 70i enhance the oncolytic effect of replicative adenovirus.

Replication-competent viruses are currently being evaluated for their cancer cell-killing properties. These vectors are designed to induce tumor regression after selective viral propagation within the tumor. However, replication-competent viruses have not resulted heretofore in complete tumor eradication in the clinical setting. Recently, heat shock has been reported to partially alleviate replication restriction on an avian adenovirus (Ad) in a human lung cancer cell line. Therefore, we hypothesized that heat shock and overexpression of heat shock protein (hsp) would support the oncolytic effect of a replication-competent human Ad. To this end, we tested the oncolytic and burst kinetics of a replication-competent Ad after exposure to heat shock or to inducible hsp 70 overexpression by a replication-deficient Ad (Adhsp 70i). Heat-shock resulted in augmentation of Ad burst and oncolysis while decreasing total intracellular Ad DNA. Overexpression of hsp 70i also enhanced Ad-mediated oncolysis but did not decrease intracellular Ad DNA levels. We conclude that heat shock and Adhsp 70i enhance the Ad cell-killing potential via distinct mechanisms. A potential therapeutic implication would be the use of local hyperthermia to augment oncolysis by increasing the burst of replication-competent Ad. The role of hsp in Ad-mediated oncolysis should be additionally explored.

Adenoviruses, Human↗
PubMed2001

A midkine promoter-based conditionally replicative adenovirus for treatment of pediatric solid tumors and bone marrow tumor purging.

The treatment of advanced neuroblastoma (NB) or Ewing's sarcoma (ES) is one of the major challenges in pediatric oncology. Both malignancies are refractory to conventional therapies and have an extremely poor prognosis. High-dose myeloablative radiochemotherapy with autologous bone marrow or peripheral blood stem cell rescue is one of the most aggressive treatments attempted for these diseases but is often undermined by residual tumor cells contaminating the graft. Thus, in this approach, purging of tumor cells from the graft is key to the prevention of relapse after transplantation. We investigated a novel approach to eliminate tumor cells from the bone marrow or peripheral blood stem cell graft without causing stem cell damage through the use of a conditionally replicative adenovirus (Ad). ES and NB are sensitive to Ad infection, and advanced NBs express a high level of the growth/differentiation factor midkine (MK). We confirmed in this study that ES cell lines (SK-ES-1 and RD-ES) are also sensitive to Ad infection and express high levels of MK. In contrast, CD34+ stem cells are refractory to Ad infection and express very little MK. A conditionally replicative Ad in which the expression of E1 is controlled by the MK promoter achieved good levels of viral replication in NB or ES and induced remarkable tumor cell killing. On the other hand, this virus caused no damage to CD34+ cells even after 3 h of infection at a dose of 1000 multiplicity of infection. We concluded that application of this replication-competent Ad to hematopoietic grafts could be a simple but effective procedure to achieve complete tumor cell purging.

Adenoviridae↗
PubMed2001

Targeting oncolytic adenoviral agents to the epidermal growth factor pathway with a secretory fusion molecule.

Cancer gene therapy with conditionally replicating adenoviruses is a powerful way of overcoming low tumor transduction. However, one of the main remaining obstacles is the highly variable level of the coxsackie-adenovirus receptor expression on human primary cancers. In contrast, the epidermal growth factor receptor (EGFR) is overexpressed in various tumor types, and its expression correlates with metastatic behavior and poor prognosis. We constructed an adenovirus expressing a secretory adaptor capable of retargeting adenovirus to EGFR, resulting in a more than 150-fold increase in gene transfer. A replication-competent dual-virus system secreting the adaptor displayed increased oncolytic potency in vitro and therapeutic gain in vivo. This approach could translate into increased efficacy and specificity in the treatment of EGFR overexpressing human cancers.

Adenoviridae↗
PubMed2001

Cytoprotection of pancreatic islets before and soon after transplantation by gene transfer of the anti-apoptotic Bcl-2 gene.

Isolated pancreatic islet transplantation is a promising alternative to conventional insulin-dependent diabetes treatment but is not yet a practical clinical therapy. In the first few days after pancreatic islet transplantation, substantial donor pancreatic islet dysfunction and apoptosis commonly occur. Islet apoptosis has been documented after extracellular matrix disruption and exposure to proinflammatory cytokines, and during hypoxia before islet revascularization and rejection. These studies show that targeting the apoptosis pathway by adenoviral-mediated gene transfer of the anti-apoptotic Bcl-2 gene exerts a major cytoprotective effect on isolated macaque pancreatic islets. Bcl-2 transfection ex vivo protects these islets from apoptosis induced by disruption of the islet extracellular matrix during pancreatic digestion. Additionally, overexpression of Bcl-2 confers long-term, stable protection and maintenance of functional islet mass after transplantation of macaque islets into diabetic severe combined immunodeficency mice. Notably, genetic modification of pancreatic islets also reduced the islet mass required to achieve stable euglycemia. Ex vivo gene transfer of anti-apoptotic genes has potential as a therapeutic approach to both minimize loss of functional islet mass after transplant and reduce the high donor islet requirement currently needed for successful stable reversal of insulin-dependent diabetes.

Adenoviridae↗
PubMed2001

Overcoming multi-drug resistance using an intracellular anti-MDR1 sFv.

We made an intracellular single-chain variable fragment (sFv) from the C219 monoclonal antibody that recognized the intracellular domain of the multidrug resistance (MDR) gene product, P-glycoprotein (P-gp). Immuno-cytochemistry using the FITC conjugated anti-C-myc tag antibody showed that the sFv protein was expressed in the cytoplasm of the cells. Although transfection of the sFv did not result in the down-regulation of P-gp expression in P-gp positive MDR cells as determined by flow cytometry analysis, Adriamycin (ADM) uptake and Rhodamine123 (Rh123) retention were increased by the C219 intra-cellular sFv transfection. The transfected cells exhibited a higher sensitivity to ADM using a 10-day colony formation assay. The conventional 3-day MTT assay showed the drug resistant tendency in C219 sFv transfected cell we tested. The growth rate of C219 sFv transfected cells was delayed in all non-MDR and MDR cells that might be the reason why C219 transfected cells exhibited the drug resistant tendency in the MTT assay. Despite this unexpected effect of C219 sFv on growth rate, our data suggest that the intra-cellular sFv technique could knockout MDR functionally and may offer a means of increasing the effectiveness of tumor chemotherapy.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

Fiber knob modifications overcome low, heterogeneous expression of the coxsackievirus-adenovirus receptor that limits adenovirus gene transfer and oncolysis for human rhabdomyosarcoma cells.

Exploiting the lytic life cycle of viruses has gained recent attention as an anticancer strategy (oncolysis). To explore the utility of adenovirus (Ad)-mediated oncolysis for rhabdomyosarcoma (RMS), we tested RMS cell lines for Ad gene transduction and infection. RMS cells were variably transduced by Ad. Compared with control cells, RMS cells were less sensitive or even resistant to oncolysis by wild-type virus. RMS cells expressed the Ad internalization receptors, alpha(v) integrins, but had low or undetectable expression of the major attachment receptor, coxsackievirus-Ad receptor (CAR). Mutant Ads with ablated CAR binding exhibited only 5-20% of transgene expression in RMS cells seen with a wild-type vector, suggesting that residual or heterogeneous CAR expression mediated the little transduction that was detectable. Immunohistochemical analysis of archived clinical specimens showed little detectable CAR expression in five embryonal and eight alveolar RMS tumors. Stable transduction of the cDNA for CAR enabled both efficient Ad gene transfer and oncolysis for otherwise resistant RMS cells, suggesting that poor CAR expression is the limiting feature. Gene transfer to RMS cells was increased >2 logs using Ads engineered with modified fiber knobs containing either an integrin-binding RGD peptide or a polylysine peptide in the exposed HI loop. The RGD modification enabled increased oncolysis for RMS cells by a conditionally replicative Ad, Ad delta24RGD, harboring a retinoblastoma-binding mutation in the E1A gene. Thus, the development of replication-competent vectors targeted to cell surface receptors other than CAR is critical to advance the use of Ad for treating RMS.

Adenoviridae↗
PubMed2001

Efficient oncolysis by a replicating adenovirus (ad) in vivo is critically dependent on tumor expression of primary ad receptors.

Replicating adenoviruses (Ads) are designed to replicate in and destroy cancer cells, generating viral progeny that spread within the tumor. To address the importance of the primary cellular receptor for Ads, the coxsackievirus and Ad receptor (CAR), in permitting intratumoral spread of a replicating Ad, we have used a pair of tumor cell lines differing only in the expression of a primary receptor for Ad5. This novel system thus allowed the first direct evaluation of the relationship between the efficacy of a replicating Ad and the primary receptor levels of the host cell without the confounding influence of other variable cellular factors. We demonstrate that the absence of the primary cellular receptor on the tumor cells restricts the oncolytic potency of a replicating Ad both in vitro and in vivo. Based on these findings, it is apparent that the potential therapeutic advantages afforded by viral replication would be negated by poor intratumoral spread of the viral progeny due to the failure to infect neighboring tumor cells. Because a number of studies have reported that primary cancer cells express only low levels of CAR, our results suggest that strategies to redirect Ads to achieve CAR-independent infection will be necessary to realize the full potential of replicating Ads in the clinical setting.

Adenoviruses, Human↗
PubMed2001

Reciprocal regulation of plasmacytoid dendritic cells and monocytes during viral infection.

Reciprocal regulation of opposing functions characterizes biological systems. We now show that adenovirus-infected plasmacytoid dendritic cells (PDC) inhibit monocyte to myeloid dendritic cell (MDC) differentiation and function, and that adenovirus-infected monocytes inhibit PDC type I interferon secretion. Adenovirus-infected PDC secreted IFN-alpha, beta and omega in an 86:2:1 ratio. PDC type I interferons inhibited MDC differentiation and function (reduced IL-12 secretion, IFN-gamma induction, MLR and CD40 expression, and increased CD1a(+)CD14(+) cells). Type I interferon receptor blocking antibody reversed all PDC effects, and recombinant IFN-alpha, beta or omega replicated all effects, except reduced CD40. Adenovirus-infected monocytes suppressed PDC type I interferon secretion, which was reversed with anti-IL-10 neutralizing antibodies. Exogenous IL-10 suppressed PDC type I interferon secretion without reducing PDC viability. Therefore, monocyte IL-10 regulates PDC type I interferon secretion, and PDC type I interferons inhibit MDC differentiation and function. Such reciprocal regulation of potentially opposing influences may help modulate anti-pathogen immunity.

Adenovirus Infections, Human↗
PubMed2001

An immunomagnetic-based method for the purification of ovarian cancer cells from patient-derived ascites.

OBJECTIVE: Primary ovarian cancer cells obtained from fresh tumor have many advantages over established cell lines. Therefore, a procedure for the specific and efficient purification of such neoplastic cells is critical. We report an effective immunomagnetic method for the isolation of tumor cells from the ascitic fluid of patients diagnosed with ovarian adenocarcinoma. METHODS: This procedure incorporates the use of monoclonal antibody (mAb) CC49, which recognizes the tumor-associated glycoprotein 72 (TAG-72). TAG-72 is highly expressed on ovarian tumor cell surfaces with little or no reactivity with normal tissues. Also used in this protocol are immunomagnetic beads, which bind to the CC49 mAb via a secondary antibody. When ovarian cancer cells adhere to the magnetic beads, a magnetic field is used to separate the tumor cells from all other cellular components. RESULTS: Using ascitic fluid from five patients, we found that preparations before purification contained between 38 and 52% neoplastic cells. Using our method, we produced preparations that were between 63 and 96% pure for cancer cells, thus obtaining an average increase in tumor cell enrichment of 86%. CONCLUSION: We, therefore, believe this method is preferable for producing high yields of pure ovarian neoplastic cells. We are now employing this technique in our laboratory to provide a stringent and pure template for our studies on gene transfer to primary ovarian cancer cells.

Antibodies, Monoclonal↗
PubMed2001

Transcriptional targeting for ovarian cancer gene therapy.

Ovarian carcinoma is a leading cause of cancer death in women. Though advances in conventional therapies have been achieved, long-term survival rates for most patients diagnosed with ovarian cancer are still low. Therefore, novel molecular therapeutic strategies such as gene therapy are being intensively pursued. Such approaches are based on the enormous progress that has been achieved in the elucidation of the molecular foundations of ovarian cancer. In this regard transcriptional control elements (promoters) of genes frequently upregulated or specifically expressed in tumors can be applied in a heterologous context to drive expression of therapeutic genes in targeted gene therapy strategies. This review discusses transcriptional targeting strategies in ovarian cancer gene therapy and gives an overview of tumor-specific promoters (TSPs) that have been applied for this purpose.

Animals↗
PubMed2001

Bax-induced apoptosis as a novel gene therapy approach for carcinoma of the cervix.

OBJECTIVE: The transfer of tumor suppressor genes has been shown to revert the malignant phenotype. In this regard, bax is a pro-apoptotic molecule that also functions as a tumor suppressor. The purpose of this study was to evaluate bax as a gene therapeutic in the context of cervical cancer. METHODS: Efficiency of viral transduction in cervical cancer cell lines and primary cervical cancer cells was evaluated with an adenoviral vector encoding green fluorescent protein and luciferase, respectively. We generated a recombinant adenoviral vector that encodes the bax gene under inducible conditions. To this end, expression of this pro-apoptotic gene was controlled by a Cre-LoxP system. Following infection with the recombinant bax adenovirus, the viability of cervical cancer cell lines and primary cervical cancer cells was evaluated using crystal violet staining and FACS analysis. Apoptotic cell death was monitored using annexin V staining. RESULTS: High levels of viral infection were observed in all cervical cancer cell lines (>85%) and primary cervical cancer cells. Significant cytotoxicity was seen in all cervical cancer cells lines and, more importantly, patient-derived primary cervical cancer cells. Moreover, bax-mediated cell death occurred via an apoptotic pathway. CONCLUSIONS: Our results indicate that a bax recombinant adenoviral vector causes cell death mediated via an apoptotic pathway in multiple cervical cancer cell lines and primary cervical cancer cells. These data suggest that bax may be a candidate for human gene therapy in the setting of cervical carcinoma.

Adenoviridae↗
PubMed2001

Selective transduction of murine myelomonocytic leukemia cells (WEHI-3B) with regular and RGD-adenoviral vectors.

On the basis of the susceptibility of normal myelomonocytic cells to adenoviral vectors, we have studied the possibility of selectively transducing myelomonocytic murine leukemic cells (WEHI-3B) with regular (Reg-Ad) and genetically modified (RGD-Ad) adenoviral vectors. An 8-h incubation of WEHI-3B cells with 100 pfu of Reg-Ad vectors/cell resulted in the whole population becoming positive for transgene expression. Under identical conditions of infection, 20-30% of mouse bone marrow (BM) cells were positive for the transgene. When RGD-Ad vectors were used, a brief exposure (10 min) of WEHI-3B cells to 150 pfu of the virus/cell was enough for 100% of the leukemia cells to become positive for the marker transgene (EGFP). Under these conditions, only 15-20% of BM cells and of primitive hematopoietic progenitors (Lin(-)Sca-1(+) cells) became EGFP(+), indicating an improved selectivity of the vectors for the leukemic cells. The incubation of WEHI-3B but not normal BM cells with soluble fiber protein (FP) inhibited the infection with Reg-Ad. The use of the RGD-Ad bypassed the FP-CAR interaction required for the transduction of WEHI-3B cells with Reg-Ad, suggesting that the abrogation of this requirement accounts for the improved infectivity of these leukemic cells and for the selectivity of RGD-Ad in targeting WEHI-3B leukemia cells.

Adenoviridae↗
PubMed2001

Characterization of the cyclooxygenase-2 promoter in an adenoviral vector and its application for the mitigation of toxicity in suicide gene therapy of gastrointestinal cancers.

The application of adenoviral molecular chemotherapy for systemic malignant disease using herpes simplex virus thymidine kinase has been limited by ectopic transgene expression in the liver due to the vector hepatotropism. The aim of this study was to mitigate this hepatotoxicity using the promoter of cyclooxygenase-2, inactive in liver but active in many gastrointestinal cancers. To analyze the specificity of transgene expression driven by cyclooxygenase-2 (cox-2) promoters, promoters of two different lengths were incorporated into adenoviral vectors to drive luciferase expression. The specific cytocidal effect and in vivo toxicity were analyzed with thymidine kinase expression vectors. The specificity of the cox-2 promoter was well preserved in the adenoviral vector. In vivo, the cox-2 promoter (-1432/+59) showed very little activity in the liver but attained high activity, comparable to that of the cytomegalovirus promoter, in cyclooxygenase-2-positive subcutaneous tumors. The cox-2 promoter-driven thymidine kinase-expressing vectors showed a cytocidal effect specifically in cyclooxygenase-2-positive cells. When mice were treated with the thymidine kinase-expressing vector and ganciclovir, the cox-2 promoter successfully mitigated the fatal hepatotoxicity, which was observed with the cytomegalovirus promoter-driven vector. The cox-2 promoter successfully mitigated the adverse effects of adenoviral suicide gene therapy by minimizing transgene expression in the liver.

Adenoviridae↗
PubMed2001

An adenovirus with enhanced infectivity mediates molecular chemotherapy of ovarian cancer cells and allows imaging of gene expression.

The adenovirus (Ad) is a useful vector for cancer gene therapy due to its unparalleled gene transfer efficiency to dividing and quiescent cells. Primary cancer cells, however, often have highly variable or low levels of the requisite coxsackie-adenovirus receptor (CAR). Also, assessment of gene transfer and vector persistence has been logistically difficult in human clinical trials. We describe here two novel bicistronic adenoviral (Ad) vectors, AdTKSSTR and RGDTKSSTR, which contain the herpes simplex virus thymidine kinase gene (TK) for molecular chemotherapy and bystander effect. In addition, the viruses contain the human somatostatin receptor subtype-2 gene (SSTR2), the expression of which can be noninvasively imaged. We enhanced the infectivity of RGDTKSSTR by genetically incorporating the RGD-4C motif into the HI-loop of the fiber. This allows the virus to circumvent CAR deficiency by binding to alpha(v)beta(3) and alpha(v)beta(5) integrins, which are highly expressed on most ovarian cancers. The expanded tropism of RGDTKSSTR results in increased infectivity of purified primary ovarian cancer cells and allows enhanced gene transfer in the presence of malignant ascites containing anti-Ad antibodies. RGDTKSSTR may be a useful agent for treating ovarian cancer in clinical trials.

Adenoviridae↗
PubMed2001

Combined transductional and transcriptional targeting improves the specificity of transgene expression in vivo.

The promise of gene therapy for health care will not be realized until gene delivery systems are capable of achieving efficient, cell-specific gene delivery in vivo. Here we describe an adenoviral system for achieving cell-specific transgene expression in pulmonary endothelium. The combination of transductional targeting to a pulmonary endothelial marker (angiotensin-converting enzyme, ACE) and an endothelial-specific promoter (for vascular endothelial growth factor receptor type 1, flt-1) resulted in a synergistic, 300,000-fold improvement in the selectivity of transgene expression for lung versus the usual site of vector sequestration, the liver. This combined approach should be useful for the design of other gene delivery systems.

Adenoviridae↗
PubMed2001

Stromal-derived factor-1 in human tumors recruits and alters the function of plasmacytoid precursor dendritic cells.

Dendritic-cell (DC) trafficking and function in tumors is poorly characterized, with studies confined to myeloid DCs (DC1s). Tumors inhibit DC1 migration and function, likely hindering specific immunity. The role of plasmacytoid DCs (DC2s) in tumor immunity is unknown. We show here that malignant human ovarian epithelial tumor cells express very high levels of stromal-derived factor-1, which induces DC2 precursor (preDC2) chemotaxis and adhesion/transmigration, upregulates preDC2 very late antigen (VLA)-5, and protects preDC2s from tumor macrophage interleukin-10-induced apoptosis, all through CXC chemokine receptor-4. The VLA-5 ligand vascular-cell adhesion molecule-1 mediated preDC2 adhesion/transmigration. Tumor preDC2s induced significant T-cell interleukin-10 unrelated to preDC2 differentiation or activation state, and this contributed to poor T-cell activation. Myeloid precursor DCs (preDC1s) were not detected. Tumors may weaken immunity by attracting preDC2s and protecting them from the harsh microenvironment, and by altering preDC1 distribution.

Apoptosis↗