[Show abstract][Hide abstract] ABSTRACT: We previously identified the mouse and human Glipr1 and GLIPR1/RTVP-1 genes, respectively, as direct p53 targets with proapoptotic activities in various cancer cell lines, including prostate cancer (PCa). Intratumoral injection of an adenoviral vector capable of efficient transduction and expression of Glipr1 (AdGlipr1) yielded promising therapeutic results in an orthotopic, metastatic mouse model of PCa. AdGlipr1-transduced macrophages (Mφ/Glipr1) generated greater surface expression of CD40, CD80 and major histocompatibility complex class II molecules and greater production of interleukin 12 (IL-12) and IL-6 in vitro than control macrophages did. Mechanistic analysis indicated that increased production of IL-12 in Mφ/Glipr1 depends on activation of the p38 signaling cascade. Mφ/Glipr1 injected into orthotopic 178-2BMA tumors in vivo resulted in significantly suppressed prostate tumor growth and spontaneous lung metastases and longer survival relative to those observed in control-treated mice. Furthermore, these preclinical data indicate the generation of systemic natural killer cell activity and tumor-specific cytotoxic T lymphocyte responses. Trafficking studies confirmed that intratumorally injected Mφ/Glipr1 could migrate to draining lymph nodes. Overall, our data suggest that this novel gene-modified cell approach is an effective treatment avenue that induces antitumor immune responses in preclinical studies.
[Show abstract][Hide abstract] ABSTRACT: This study presents the control method for a new hybrid high-voltage dc (HVDC) connection for large wind farms with doubly fed induction generators (DFIGs). The hybrid HVDC system consists of a line-commutated converter plus a static synchronous compensator (STATCOM) on the rectifier side and a pulse-width modulation (PWM) current source inverter (CSI) on the inverter side. The overall control strategy is divided into three parts, where the STATCOM subsystem supports the island bus voltage; the rectifier subsystem delivers the active power generated by the wind farm; and the CSI subsystem injects the active power - as well as reactive power, if necessary - to the grid. Taking into account the non-linear and coupling characteristics of each subsystem, a series of control schemes is proposed: for the STATCOM subsystem, a state feedback linearisation and decoupling method is employed; for the rectifier subsystem, a double-loop controller is designed and its inner loop is based on the inverse-system control techniques; for the CSI subsystem, independent active and reactive power control scheme is proposed. Through Simulink/SimPowerSystems simulations on a representative hybrid HVDC system, the startup process is demonstrated and system performances under both normal and fault conditions are investigated.
IET Renewable Power Generation 02/2011; 5(1-5):36 - 47. DOI:10.1049/iet-rpg.2009.0171 · 1.90 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Adenoviral vector delivery of the Herpes simplex virus thymidine kinase (HSV-tk) gene in combination with the prodrug ganciclovir (GCV) has been tested in phase I clinical trials for prostate cancer and found to exhibit a satisfactory toxicity profile. We have developed additional adenoviral vectors with differing promoters to optimize the expression profile and in the present study evaluate the potential systemic toxicity of these vectors. Four recombinant adenoviral vectors that express the HSV-tk gene were generated using three different promoters: CMV (leftward orientation); RSV (both rightward and leftward orientation); and the mouse caveolin-1 (cav-1) promoter (leftward orientation). Efficacy was determined in vitro by cytotoxicity assays in a mouse prostate cancer cell line, RM-9, and in vivo by treating orthotopic tumors. Potential toxicity was evaluated from liver histology and apoptotic cell counts and enzyme levels in the serum following intravenous adenoviral vector injection. Although there were differences in HSV-tk expression at the protein level among the four vectors there were no significant differences in in-vitro cytotoxicity studies with GCV or in vivo in tumor growth suppression of an orthotopic mouse prostate cancer model in GCV treated mice. Intravenous delivery of high doses of all adenoviral vectors lead to abnormalities in liver function as measured by specific serum markers and histological evaluation of liver tissue and increased levels of apoptosis in the liver. These abnormalities were most prevalent with the vector containing the CMV promoter and the rightward oriented RSV promoter. They were least prevalent in the vector regulated by the cav-1 promoter. Upregulation of specific chemokines, MIP-2 and MIP-1beta was correlated with apoptotic counts. Our results demonstrate that comprehensive toxicological analysis of adenoviral vectors provides internally consistent information that can differentiate vectors with comparable efficacy based on toxicity. In these studies vectors with the cav-1 promoter-driven and leftward RSV-driven HSV-tk gene demonstrated minimal toxicities with cytotoxic effectiveness comparable to more toxic vectors. Our studies further suggest that promoter selection can influence the toxic effects of an adenoviral gene therapy vector.
Prostate Cancer and Prostatic Diseases 02/2002; 5(4):316-25. DOI:10.1038/sj.pcan.4500610 · 3.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Caveolin-1, a structural component of caveolae, is overexpressed in metastatic and androgen-resistant prostate cancer and highly expressed in tumor-associated endothelial cells. The mouse cav-1 promoter was cloned and placed upstream of the HSV-tk gene in an adenoviral vector (Adcav-1tk) and compared with a cytomegalovirus (CMV) or Rous sarcoma virus (RSV) promoter-driven HSV-tk, AdCMVtk and AdRSVtk vectors, respectively. Mouse and human prostate cancer cells and mouse endothelial cells were infected with Adcav-1tk, AdCMVtk or control vectors without the HSV-tk gene (Adcav-1 and AdCMV) and subsequently treated with ganciclovir (GCV). GCV-mediated in vitro cytotoxicity induced by the Adcav-1tk vector was comparable to that for AdCMVtk in multiple mouse and human prostate cancer cell lines. To evaluate the activity of Adcav-1tk in vivo, orthotopic mouse prostate cancer tumors were generated with RM-9 cells and injected in situ with Adcav-1tk, AdCMVtk, AdRSVtk, or AdCMVbetagal (control) and treated with GCV. All three HSV-tk transducing vectors produced statistically significant reductions in wet weight and increased apoptotic indices compared with the control vector. However, only Adcav-1tk produced significant necrosis, and only Adcav-1tk and AdRSVtk caused significant decreases in microvessel density. In conclusion, Adcav-1tk demonstrated efficacy in vitro and in vivo in preclinical models of prostate cancer. Our results suggest that the cav-1 promoter may have unique benefits in targeting gene therapy to prostate cancer and its associated vasculature.
Clinical Cancer Research 01/2002; 7(12):4272-9. DOI:10.1016/S1051-0443(00)70200-4 · 8.72 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Previously, we demonstrated that up-regulation of caveolin-1 (cav-1) was associated with prostate cancer metastasis, biochemical recurrence after radical prostatectomy, and androgen insensitivity. The objective of this study was to characterize the regulation of cav-1 by testosterone (T) and to test the effects of cav-1 on prostate cancer cell survival/clonal growth and metastatic activities. Our results demonstrated that T up-regulated cav-1 protein levels in part through transcriptional regulation and significantly enhanced survival of prostate cancer cell lines ABAC3 and LNCaP after serum starvation (>40% and >60% increased viability, respectively) and in an extended clonogenic assay (approximately 4-fold and 6-fold increase in colonies, respectively). Importantly, antisense cav-1 inhibited the survival effects of T in these assay systems. Modest but not high levels of adenoviral vector-mediated cav-1 expression alone also significantly increased viability (>40%) and clonal growth (10-fold increase in colonies) after serum starvation. Analysis of spontaneous metastasis in stably transfected antisense cav-1 mouse prostate cancer cell clones demonstrated reduction of spontaneous lymph node metastasis incidence (13%), spontaneous lymph node metastasis volume (46%), and experimental lung metastasis incidence (40%) compared with vector control cell clones. Surgical castration further reduced spontaneous lymph node metastasis incidence and volume (18% and 28%, respectively) in antisense cancer cell clones, but not in vector control clones. Our studies demonstrate that cav-1 is a downstream effector of T-mediated prostate cancer cell survival/clonal growth and that modest levels of cav-1 can independently promote prostate cancer cell survival/clonal growth and metastatic activities.
Cancer Research 07/2001; 61(11):4386-92. · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The development of effective treatments for prostate cancer is thwarted by the natural history of the disease. The biological and clinical potential of most individual cancers is uncertain. In many cases the disease will not progress to clinical significance but experimental and clinical studies indicate that prostate cancer can and may metastasis early in the course of the disease from relatively small foci (i.e., not necessarily the largest or index cancer). Localised prostate cancer is potentially curable with localised therapies (radical prostatectomy or irradiation therapy). However, there are no curative therapies for metastatic prostate cancer. Gene therapy, especially those approaches with an immunomodulatory component, may provide additional therapeutic options with the potential to affect both localised and systemic disease. We have pioneered the development and application of in situ gene therapy protocols using adenoviral vectors to transduce specific genes that generate cytotoxic activity and/or a systemic antitumour immune response. In addition we have completed initial studies that demonstrate the therapeutic potential of adenoviral vector-mediated gene modified cell-based vaccines. Our review discusses preclinical studies focused on the development of immunostimulatory in situ gene therapy approaches that hopefully will provide novel and effective treatments for localised and metastatic prostate cancer.
[Show abstract][Hide abstract] ABSTRACT: Interleukin-12 (IL-12) can elicit potent antitumoral effects that involve the recruitment of specific immune effector cells. We investigated the efficacy of a single injection of a recombinant adenovirus expressing murine IL-12 (AdmIL-12) directly into orthotopic mouse prostate carcinomas generated from a poorly immunogenic cell line (RM-9) derived from the mouse prostate reconstitution system. Significant growth suppression (> 50% reduction of tumor weight) and increased mean survival time (23.4 to 28.9 days) were observed compared with controls. Suppression of pre-established lung metastases was also observed following the injection of AdmIL-12 into the orthotopic tumor. Cytolytic natural killer cell activity was markedly enhanced 1-2 days after virus injection. Immunohistochemical analysis showed significantly elevated intratumoral infiltration of CD4+ and CD8+ T cells 7 days after virus injection. However, splenocyte-derived cytotoxic T lymphocytes were not detected during the 14 days following treatment. Increased numbers of nitric oxide synthase-positive macrophages were seen in the AdmIL-12 treated group 7 days following injection. Systemic inhibition of natural killer cells with antiasialo-GM1 serum led to increased numbers of lung metastases in AdmIL-12-treated orthotopic tumors but did not affect local tumor growth. In this model system the antitumor effects of a single injection of adenovirus-mediated IL-12 appears to be based to a large extent on the activation of nitric oxide synthase in macrophages and possibly T cell activities, whereas the relatively early cytolytic activity of natural killer cells are largely but not exclusively responsible for the antimetastatic effects.
[Show abstract][Hide abstract] ABSTRACT: We introduced the gene for wild-type human p53 or p21, a critical downstream mediator of p53-induced growth suppression, into a p53-deficient mouse prostate cancer cell line using a recombinant adenoviral vector (Ad5CMV-p53 or Ad5CMV-p21). Elevated levels of endogenous mouse p21 mRNA provided evidence for the functional activity of virally transduced p53. Functional activity of viral-transduced p21 was demonstrated through immunoprecipitation of cellular protein extracts, which showed that the viral-transduced p21 associates with cyclin-dependent kinase 2 and was sufficient to down-regulate the activity of the cyclin-dependent kinase by approximately 65%. In vitro growth assays revealed significantly higher growth suppression after Ad5CMV-p21 infection compared to Ad5CMV-p53. In vivo studies in syngeneic male mice with established s.c. prostate tumors demonstrated that the rate of growth and final tumor volume were reduced to a much greater extent in mice that received intratumor injection of Ad5CMV-p21 compared to Ad5CMV-p53. In addition, the survival of host animals bearing tumors that were infected with Ad5CMV-p21, but not Ad5CMV-p53, was significantly extended. These data suggest that Ad5CMV-p21 may be effective as a therapeutic agent for prostate cancer.
Cancer Research 12/1995; 55(22):5151-5. · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recombinant adenovirus Ad5CMV-p53 induced a strong cytocidal effect in Saos-LM2 cells. This cell line, derived from human osteosarcoma Saos-2 cells, has a homozygous deletion of the p53 gene. By using immunocytochemical and Western blot analyses, we demonstrated that Ad5CMV-p53 effectively infected Saos-LM2 cells at multiplicities of infection of 10 to 50 plaque-forming units/cell and mediated a high-level expression of the exogenous wild-type p53 protein in the cells. Growth of the infected cells was greatly suppressed whereas that of mock- or control virus-infected cells was not. Because wild-type p53 induces apoptosis in certain types of cells, we studied DNA fragmentation in situ in the Ad5CMV-p53-infected Saos-LM2 cells by terminal deoxynucleotidyl transferase assays, which yielded positive nuclear staining. Further analysis of the Ad5CMV-p53-infected Saos-LM2 cells by light and electron microscopy demonstrated that the cells underwent the characteristic morphological changes of apoptosis such as plasma-membrane blebbing, nuclear condensation, and fragmentation. These changes were not observed in mock- or control virus-infected cells. Our results on Saos-LM2 cells indicate that apoptosis induced by Ad5CMV-p53 may be one of the mechanisms underlying the cytocidal effect of Ad5CMV-p53.
Cancer Gene Therapy 04/1995; 2(1):9-17. · 2.42 Impact Factor