Biomedical subjects

D T Curiel

Publications and source records attributed to D T Curiel.

At least 19 recordsLinked to original sources

PubMed2006

Evolving gene therapy approaches for osteosarcoma using viral vectors: review.

This review begins with an introduction to the malignant bone tumor, osteosarcoma [OS] and then moves to a discussion of the commonly used vectors for gene transfer. We first briefly highlight non-viral vectors including polymeric and liposomal delivery systems but concentrate predominantly on the 5 leading viral vectors used in cancer gene therapy, specifically retroviruses, adeno-associated viruses, herpes viruses and lentiviruses with the most detailed analysis reserved for adenoviruses. The 3 main strategies for gene therapy in osteosarcoma are next summarized. As part of this review, the several prodrug-converting enzymes utilized in OS suicide gene therapy are examined. The text then turns to a discussion of adenovirus-mediated gene transfer and the need for tumor targeting via transductional or transcriptional approaches. Because of practical problems with use of replication-incompetent viruses in achieving complete tumor kill in vivo, virotherapy utilizing replication competent viruses has come to the fore. This topic is, thus, next reviewed which allows for a natural transition to a discussion of armed therapeutic viruses many of which are conditionally replicating adenoviruses carrying transgenes with established anti-tumor efficacy. We recognize that several other issues have arisen which hamper progress in the field of cancer gene therapy. We, therefore, review viral-induced toxicity in the host and vector delivery issues which have been found to potentially influence safety. We end with a brief perspective including commenting on animal models used in examining delivery strategies for osteosarcoma gene therapy. The challenges remaining are touched upon most especially the need to deal with pulmonary metastatic disease from OS.

Bone Neoplasms↗

PubMed2006

Directing adenovirus across the blood-brain barrier via melanotransferrin (P97) transcytosis pathway in an in vitro model.

Adenovirus serotype 5 (Ad5) is widely used in the development of gene therapy protocols. However, current gene therapy strategies involving brain are mostly based on intra-cranial injection. A major obstacle for systemically administered vectors to infect brain tissue is the blood-brain barrier (BBB). One strategy to cross the BBB is transcytosis, a transcellular transport process that shuttles a molecule from one side of the cell to the other side. Recently, melanotransferrin (MTf)/P97 was found to be able to cross the BBB and accumulate in brain. We thus hypothesize that re-directing Ad5 vectors to the MTf transcytosis pathway may facilitate Ad5 vectors to cross the BBB. To test this hypothesis, we constructed a bi-specific adaptor protein containing the extracellular domain of the coxsackie-adenovirus receptor (CAR) and the full-length melanotransferrin (sCAR-MTf), and investigated its ability to re-direct Ad5 vectors to the MTf transcytosis pathway. We found this adaptor protein could re-direct Ad5 to the MTf transcytosis pathway in an in vitro BBB model, and the transcytosed Ad5 viral particles retained their native infectivity. The sCAR-MTf-mediated Ad5 transcytosis was temperature- and dose dependent. In addition, we examined the directionality of sCAR-MTf-mediated Ad5 transcytosis, and found the efficiency of apical-to-basal transcytosis was much higher than that of basal-to-apical direction, supporting a role of this strategy in transporting Ad5 vectors towards the brain. Taken together, our study demonstrated that re-directing Ad5 to the MTf transcytosis pathway could facilitate gene delivery across the BBB.

Adenoviridae↗

PubMed2006

Vascular endothelial growth factor promoter-based conditionally replicative adenoviruses for pan-carcinoma application.

Treatment of advanced lung cancer is one of the major challenges in current medicine because of the high morbidity and mortality of the disease. Advanced stage lung cancer is refractory to conventional therapies and has an extremely poor prognosis. Thus, new therapeutic approaches are needed. Lung tumor formation depends on angiogenesis in which the vascular endothelial growth factor (VEGF) produced by cancer cells plays a pivotal role. Neutralizing VEGF with a soluble VEGF receptor suppresses tumor growth; however, the anticancer effect with this therapy is weakened after the intratumoral vascular network is completed. In this study, we turned the expression of VEGF by tumors to therapeutic advantage using a conditionally replication-competent adenovirus (CRAd) in which the expression of E1 is controlled by the human VEGF promoter. This virus achieved good levels of viral replication in lung cancer cells and induced a substantial anticancer effect in vitro and in vivo. As a further enhancement, the cancer cell killing effect was improved with tropism modification of the virus to express the knob domain of Ad3, which improved infectivity for cancer cells. These VEGF promoter-based CRAds also showed a significant cell killing effect for various types of cancer lines other than lung cancer. Conversely, the VEGF promoter has low activity in normal tissues, and the CRAd caused no damage to normal bronchial epithelial cells. Since tumor-associated angiogenesis via VEGF signalling is common in many types of cancers, these CRAds may be applicable to a wide range of tumors. We concluded that VEGF promoter-based CRAds have the potential to be an effective strategy for cancer treatment.

Adenoviridae↗

PubMed2006

Transductional targeting of adenovirus vectors for gene therapy.

Cancer gene therapy approaches will derive considerable benefit from adenovirus (Ad) vectors capable of self-directed localization to neoplastic disease or immunomodulatory targets in vivo. The ablation of native Ad tropism coupled with active targeting modalities has demonstrated that innate gene delivery efficiency may be retained while circumventing Ad dependence on its primary cellular receptor, the coxsackie and Ad receptor. Herein, we describe advances in Ad targeting that are predicated on a fundamental understanding of vector/cell interplay. Further, we propose strategies by which existing paradigms, such as nanotechnology, may be combined with Ad vectors to form advanced delivery vehicles with multiple functions.

Adenoviridae↗

PubMed2006

Gene therapy for carcinoma of the breast.

In view of the limited success of available treatment modalities for breast cancer, alternative and complementary strategies need to be developed. The delineation of the molecular basis of breast cancer provides the possibility of specific intervention by gene therapy through the introduction of genetic material for therapeutic purposes. In this regard, several gene therapy approaches for carcinoma of the breast have been developed. These approaches can be divided into six broad categories: (1) mutation compensation, (2) molecular chemotherapy, (3) proapoptotic gene therapy, (4) antiangiogenic gene therapy, (5) genetic immunopotentiation, and (6) genetic modulation of resistance/sensitivity. Clinical trials for breast cancer have been initiated to evaluate safety, toxicity, and efficacy. Combined modality therapy with gene therapy and chemotherapy or radiation therapy has shown promising results. It is expected that as new therapeutic targets and approaches are identified and advances in vector design are realized, gene therapy will play an increasing role in clinical breast cancer treatment.

Animals↗

PubMed2006

An ex vivo human model system to evaluate specificity of replicating and non-replicating gene therapy agents.

BACKGROUND: Inefficiency, aspecificity and toxicity of gene transfer vectors hamper gene therapy from showing its full potential. On this basis significant research currently focuses on developing vectors with improved infection and/or expression profiles. Screening assays with validity to the clinical context to determine improved characteristics of such agents are not readily available since this requires a close relationship to the human situation. We present a clinically relevant tissue slice technology to preclinically test improved vector characteristics. METHODS: Slices were prepared from rat, mouse and human liver samples and from tumor tissue. Specificity of gene expression and replication was determined by infecting target and non-target tissue slices with transcriptionally retargeted adenoviruses and oncolytic viruses. RESULTS: Using rat liver slices, we demonstrate efficient knob-mediated adenoviral infectivity. A favorable tumor-on/liver-off profile, resembling in vitro and mouse in vivo data, was shown for a tumor-specific transcriptionally retargeted adenovirus by infecting slices prepared from tumor or liver tissue. Similar liver-off data were found for mouse, rat and human samples (over 3-log lower activity of the tumor-specific promoter compared to cytomegalovirus (CMV)). More importantly, we show that this technology when applied to human livers is a powerful tool to determine aspecific replication of oncolytic viruses in liver tissue. A 2- to 6-log reduction in viral replication was observed for a tumor-specific oncolytic virus compared to the wild-type adenovirus. CONCLUSIONS: The precision-cut tissue slice technology is a powerful method to test specificity and efficiency of gene transfer as well as of viral replication using human tissue.

Adenoviridae↗

PubMed2006

Suppressor of cytokine signaling-1 expression by infectivity-enhanced adenoviral vector inhibits IL-6-dependent proliferation of multiple myeloma cells.

Multiple myeloma (MM) accounts for 10% of hematological malignant disorders. Its refractory nature indicates the necessity of developing novel therapeutic modalities. Since interleukin 6 (IL-6) is one of the major growth factors for MM cells, we expressed suppressor of cytokine signaling-1 (SOCS-1), one of the blockades of IL-6 receptor downstream signaling, to suppress the proliferation of MM cells. Because MM cells are resistant to conventional adenoviral vector infection, we utilized infectivity-enhanced adenoviral vectors with an RGD4C motif in the adenoviral fiber-knob region (RGD-modified vector). In infectivity analysis, RGD-modified vectors were superior to unmodified controls in the majority of the MM cell lines tested. The overexpression of SOCS-1 using infectivity-enhanced adenoviral vectors achieved growth suppression in IL-6-dependent MM cells, but not in the IL-6-independent cells. IL-6-induced STAT3 phosphorylation was suppressed in IL-6-dependent cells, indicating that the signal transduction cascade of the IL-6 receptor signaling was blocked. In aggregate, SOCS-1 overexpression with RGD-modified adenoviral vectors achieved the antiproliferative effect in IL-6-dependent MM cells. These results provide an initial proof-of-principle of the anticancer effect of SOCS-1 expression vector as well as a promise for the future development of therapeutic modality for MM based on this vector.

Adenoviridae↗

PubMed2006

The carcinoma-specific epithelial glycoprotein-2 promoter controls efficient and selective gene expression in an adenoviral context.

Adenoviral vectors are widely used in cancer gene therapy. After systemic administration however, the majority of the virus homes to the liver and the expressed transgene may cause hepatotoxicity. To restrict transgene expression to tumor cells, tumor- or tissue-specific promoters are utilized. The tumor antigen epithelial glycoprotein-2 (EGP-2), also known as Ep-CAM, is expressed in many cancers from different epithelial origins. In this study, the EGP-2 promoter was shown to restrict the expression of luciferase and thymidine kinase in an adenoviral context in different cell lines. In vivo, the EGP-2 promoter mediated efficient expression of luciferase in tumors but showed a 3-log lower activity in liver tissue when compared with the cytomegalovirus (CMV) promoter. Similarly, the EGP-2 promoter mediated specific cell killing after ganciclovir treatment in EGP-2-positive cells. Moreover, in vivo, this treatment regiment did not cause any rise in the liver enzymes aspartate aminotransferase (ASAT) and alanine aminotransferase (ALAT), demonstrating absence of liver toxicity. In contrast, CMV-mediated expression of thymidine kinase in combination with ganciclovir treatment resulted in high ASAT and ALAT values. This study demonstrates the value of the EGP-2 promoter to restrict transgene expression to a broad range of tumor types, thereby preventing liver toxicity.

Adenoviridae↗

PubMed2006

Employment of liver tissue slice analysis to assay hepatotoxicity linked to replicative and nonreplicative adenoviral agents.

Whereas virotherapy has emerged as a novel and promising approach for neoplastic diseases, appropriate model systems have hampered preclinical evaluation of candidate conditionally replicative adenovirus agents (CRAds) with respect to liver toxicity. This is due to the inability of human viral agents to cross species. We have recently shown the human liver tissue slice model to be a facile means to validate adenoviral replication. On this basis, we sought to determine whether our ex vivo liver tissue slice model could be used to assess CRAd-mediated liver toxicity. We analyzed and compared the toxicity of a conditionally replicative adenovirus (AdDelta24) to that of a replication incompetent adenovirus (Adnull [E1-]) in mouse and human liver tissue slices. To accomplish this, we examined the hepatic apoptosis expression profile by DNA microarray analyses, and compared these results to extracellular release of aminotransferase enzymes, along with direct evidence of apoptosis by caspase-3 immunhistochemical staining and TUNEL assays. Human and mouse liver tissue slices demonstrated a marked increase in extracellular release of aminotransferase enzymes on infection with AdDelta24 compared to Adnull. AdDelta24-mediated liver toxicity was further demonstrated by apoptosis induction, as detected by caspase-3 immunohistochemical staining, TUNEL assay and microarray analysis. In conclusion, concordance of CRAd-mediated apoptosis in both the human and the mouse liver tissue slice models was demonstrated, despite the limited replication ability of CRAds in mouse liver slices. The results of this study, defining the CRAd-mediated apoptosis gene expression profiles in human and mouse liver, may lay a foundation for preclinical liver toxicity analysis of CRAd agents.

Adenoviridae↗

PubMed2006

An adenovirus vector with a chimeric fiber incorporating stabilized single chain antibody achieves targeted gene delivery.

Adenovirus (Ad) vectors are of utility for many therapeutic applications. Strategies have been developed to alter adenoviral tropism to achieve a cell-specific gene delivery capacity employing fiber modifications allowing genetic incorporation of targeting motifs. In this regard, single chain antibodies (scFv) represent potentially useful agents to achieve targeted gene transfer. However, the distinct biosynthetic pathways that scFv and Ad capsid proteins are normally routed through have thus far been problematic with respect to scFv incorporation into the Ad capsid. Utilization of stable scFv, which also maintain correct folding and thus functionality under intracellular reducing conditions, could overcome this restriction. We genetically incorporated a stable scFv into a de-knobbed, fibritin-foldon trimerized Ad fiber and demonstrated selective targeting to the cognate epitope expressed on the membrane surface of cells. We have shown that the scFv employed in this study retains functionality and that stabilizing the targeting molecule, per se, is critical to allow retention of antigen recognition in the adenovirus capsid-incorporated context.

Adenoviridae↗

PubMed2006

Infectivity enhancement for adenoviral transduction of canine osteosarcoma cells.

The full realization of conditionally replicative adenoviruses (CRAds) for cancer therapy has been hampered by the limited knowledge of CRAd function in vivo and particularly in an immunocompetent host. To address this issue, we previously proposed a canine adenovirus type 2 (CAV2)-based CRAd for clinical evaluation in canine patients with osteosarcoma (OS). In this study, we evaluated infectivity-enhancement strategies to establish the foundation for designing a potent CAV2 CRAd with effective transduction capacity in dog osteosarcoma cells. The results indicate that the native CAV2 fiber-knob can mediate increased binding, and consequently gene transfer, in both canine osteosarcoma immortalized and primary cell lines relative to previously reported Ad5 infectivity-enhancement strategies. Gene delivery was further enhanced by incorporating a polylysine polypeptide onto the carboxy terminus of the CAV2 knob. This vector demonstrated improved gene delivery in osteosarcoma xenograft tumors. These data provide the rationale for generation of infectivity-enhanced syngeneic CAV2 CRAds for clinical evaluation in a dog osteosarcoma model.

Adenoviridae↗

PubMed2005

Evaluation of tumor-specific promoter activities in melanoma.

Gene therapy is a novel therapy for melanoma. To date, however, there is still no powerful tumor specific promoter (TSP) to restrict the transgene expression in melanoma cells. In order to define a useful TSP for targeting in the context of melanoma gene therapy, four promoters, the cyclooxygenase-2 (Cox-2), alpha-chemokine SDF-1 receptor (CXCR4), epithelial glycoprotein 2 (EGP-2), and survivin, were tested in both established melanoma cell lines and primary melanoma cells. We employed recombinant adenoviral vectors (reAds) each with a candidate TSP (the Cox-2, CXCR4, EGP-2, or survivin), a reporter luciferase gene, and a poly-A signal, all of which were inserted into the E1-deleted region. A reAdGL3Bcytomegalovirus (CMV), containing the CMV promoter and luciferase gene, was used as a positive control to normalize the luciferase activity. Luciferase activity was measured in multiple tumor cell lines and two primary melanoma cell cultures after infection with reAds. Human epithelial melanocytes, HEM, were used as normal control. In contrast to three other promoters, the survivin promoter exhibited the highest activities within both melanoma cell lines and primary melanoma cells, but not in HEMs. Additionally, the survivin promoter exhibited very low activities in major mouse organs including the liver, in vivo. EGP-2 is not active in melanoma; messenger RNA expressions were correlated to promoter activities both in melanoma cell lines and primary cell cultures. Thus, these data suggest that the survivin promoter achieved a 'tumor-on/liver-off' profile, and thus represents a potentially useful tumor-specific promoter with applications for transcriptional targeting of Ad vector-based cancer gene therapy or oncolysis to melanoma.

Adenoviridae↗

PubMed2005

Noninvasive dual modality in vivo monitoring of the persistence and potency of a tumor targeted conditionally replicating adenovirus.

In clinical trials with cancer patients, the safety of conditionally replicating adenoviruses (CRAds) has been good. However, marginal data are available on the persistence or antitumor efficacy of these agents. The oncolytic potency of CRAds is determined by their capacity for entering target cells. Consequently, we constructed a retargeted CRAd featuring a secreted marker protein, soluble human carcinoembryogenic antigen (hCEA), which can be measured in growth medium or plasma. We found that virus replication closely correlated with hCEA secretion both in vitro and in vivo. Further, antitumor efficacy and the persistence of the virus could be deduced from plasma hCEA levels. Finally, using in vivo bioluminescence imaging, we were able to detect effective tumor cell killing by the virus, which led to enhanced therapeutic efficacy.

Adenocarcinoma↗

PubMed2005

Mesothelin-mediated targeting of adenoviral vectors for ovarian cancer gene therapy.

Adenoviruses (Ads) are efficient gene transfer vehicles, but Ad-mediated gene therapy for ovarian cancer remains limited in vivo by inefficient and nonspecific gene transfer. Mesothelin (MSLN), a cell surface glycoprotein, is overexpressed in ovarian cancer but not in normal tissues except mesothelial cells. Therefore, MSLN is an attractive candidate for transcriptional and transductional targeting in the context of ovarian cancer gene therapy. We evaluated the expression of MSLN mRNA and MSLN surface protein in ovarian cancer cells. Ads containing the MSLN promoter driving reporter gene expression were created and tested in ovarian cancer cell lines and purified ovarian cancer cells isolated from patients. To evaluate transductional targeting, we used an Ad vector containing an Fc-binding domain within the fiber protein, which served as a docking domain for binding with anti-MSLN immunoglobulins. Both RT-PCR and flow cytometry revealed high MSLN gene and protein expression in ovarian cancer cells. The MSLN promoter was activated in ovarian cancer cells, but showed significantly reduced activity in normal control cells. Transductional targeting of Ads via anti-MSLN antibody increased transgene expression in ovarian cancer cells. This report describes the use of MSLN for transcriptional as well as transductional targeting strategies for ovarian cancer gene therapy.

Adenoviridae↗

PubMed2005

Selective induction of tumor-associated antigens in murine pulmonary vasculature using double-targeted adenoviral vectors.

Targeted therapies directed to tumor-associated antigens are being investigated for the treatment of cancer. However, there are few suitable animal models for testing the ability to target these tumor markers. Therefore, we have exploited mice transgenic for the human coxsackie and adenovirus receptor (hCAR) to establish a new model for transient expression of human tumor-associated antigens in the pulmonary vasculature. Systemic administration of Ad in hCAR mice resulted in an increase in transgene expression in the lungs compared to wild-type mice, as determined using a luciferase reporter gene. To reduce transgene expression in the liver, the predominant organ of ectopic Ad localization and transgene expression following systemic administration, we utilized the endothelial-specific flt-1 promoter, which resulted in a further increased lung-to-liver ratio of luciferase expression. Administration of an adenoviral vector encoding the tumor-associated antigen carcinoembryonic antigen (CEA) under transcriptional control of the flt-1 promoter resulted in selective expression of this antigen in the pulmonary vasculature of hCAR mice. Feasibility of targeting to expressed CEA was subsequently demonstrated using adenoviral vectors preincubated with a bifunctional adapter molecule recognizing this tumor-associated antigen, thus demonstrating utility of this transient transgenic animal model.

Adenoviridae↗

PubMed2005

Primary adenovirus-specific cytotoxic T lymphocyte response occurs after viral clearance and liver enzyme elevation.

The virus-specific cytotoxic T lymphocyte (CTL) response is a major obstacle to effective delivery of adenovirus gene therapy. However, its relative role in viral clearance, transgene elimination and hepatotoxicity remains unclear. In this paper, we present an analysis of viral clearance and liver toxicity in relation to the induction of the virus-specific CD8 T-cell response revealed by an MHC class I tetramer. A surprisingly high number of tetramer+ CD8 T cells were found in the liver and lung and reached peak values at days 8 and 10, respectively, post-infection. Nearly 100% of these tetramer+ CD8 T cells expressed high levels of granzyme B and IFNgamma. Remarkably, liver viral load and liver enzyme elevation peaked early, at days 2 and 4, respectively, post-infection, before the specific CTL response was detectable. After generation of CTLs, there was only minimal liver damage or further decrease in virus titer. These results indicated that the primary peak response of tetramer+ CTLs does not correlate with the elimination of adenovirus or liver cytotoxic response.

Adenoviridae↗

PubMed2005

A human adenoviral vector with a chimeric fiber from canine adenovirus type 1 results in novel expanded tropism for cancer gene therapy.

The development of novel therapeutic strategies is imperative for the treatment of advanced cancers like ovarian cancer and glioma, which are resistant to most traditional treatment modalities. In this regard, adenoviral (Ad) cancer gene therapy is a promising approach. However, the gene delivery efficiency of human serotype 5 recombinant adenoviruses (Ad5) in cancer gene therapy clinical trials to date has been limited, mainly due to the paucity of the primary Ad5 receptor, the coxsackie and adenovirus receptor (CAR), on human cancer cells. To circumvent CAR deficiency, Ad5 vectors have been retargeted by creating chimeric fibers possessing the knob domains of alternate human Ad serotypes. Recently, more radical modifications based on 'xenotype' knob switching with non-human adenovirus have been exploited. Herein, we present the characterization of a novel vector derived from a recombinant Ad5 vector containing the canine adenovirus serotype 1 (CAV-1) knob (Ad5Luc1-CK1), the tropism of which has not been previously described. We compared the function of this vector with our other chimeric viruses displaying the CAV-2 knob (Ad5Luc1-CK2) and Ad3 knob (Ad5/3Luc1). Our data demonstrate that the CAV-1 knob can alter Ad5 tropism through the use of a CAR-independent entry pathway distinct from that of both Ad5Luc1-CK2 and Ad5/3-Luc1. In fact, the gene transfer efficiency of this novel vector in ovarian cancer cell lines, and more importantly in patient ovarian cancer primary tissue slice samples, was superior relative to all other vectors applied in this study. Thus, CAV-1 knob xenotype gene transfer represents a viable means to achieve enhanced transduction of low-CAR tumors.

Adenoviruses, Canine↗

PubMed2004

Transcriptional targeting of adenoviral vector through the CXCR4 tumor-specific promoter.

Adenoviral vectors are considered to be good gene delivery vectors for cancer gene therapy due to their wide host tissue range and cell cycle-independent infectivity. However, the disadvantages include the lack of specificity for cancer cells and the high liver accumulation in vivo. The human CXCR4 gene is expressed at high levels in many types of cancers, but is repressed in the liver. We explored the CXCR4 promoter as a candidate to restrict adenoviral transgene expression to tumor cells with a low expression in host tissues. The luciferase activities in multiple cancer cell lines infected with recombinant adenovirus reAdGL3BCXCR4 or the control vector reAdGL3BCMV revealed that the CXCR4 promoter exhibited relatively high transcriptional activity in a breast cancer cell line, MDA-MB-361, and two ovarian cancer cell lines, OVCAR-3 and SKOV3. ip1, 65% (P=0.0087), 16.7% (P=0.1) and 20% (P=0.0079) compared to that of the CMV promoter, respectively, and low expression, 4.9 and 0.1%, respectively, in both normal cell lines HFBC and HMEC. In addition, CXCR4 had a low expression of luciferase (0.32%) compared to that of the CMV promoter in mouse liver in vivo. The data also revealed that the CXCR4 promoter was a stronger tumor-specific promoter (TSP) than the Cox-2M promoter in primary melanomas obtained from two patients. The CXCR4 promoter is shown to have a 'tumor-on' and 'liver-off' status in vitro and in vivo, and CXCR4 may prove to be a good candidate TSP for cancer gene therapy approaches for melanoma and breast cancers.

Adenoviridae↗