In vivo magnetic resonance imaging of transgene expression.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
E A Chiocca
Publications and source records attributed to E A Chiocca.
Cytotoxicity, apoptosis, and viral replication in tumor cells treated with oncolytic ribonucleotide reductase-defective herpes simplex type 1 virus (hrR3) combined with ionizing radiation.
The viral ribonucleotide reductase (rR)-defective herpes simplex type-1 (HSV-1) virus (hrR3) has been shown previously to preferentially replicate in and kill tumor cells. This selectivity is associated with tumor cell up-regulation of mammalian rR. Ionizing radiation (IR) is currently used in the therapy of many malignancies, including glioblastoma, cervical carcinoma, and pancreatic carcinoma. IR has been shown to up-regulate mammalian rR in tumor cells and appears to increase the efficacy of at least one non-rR-deleted HSV-1 strain in an in vivo tumor model. Here, we test the hypothesis that a single therapeutic radiation fraction will increase the replication and toxicity of hrR3 for malignant cell lines in vitro. PANC-1 pancreatic carcinoma, U-87 glioblastoma, and CaSki cervical carcinoma cell lines were treated with varying doses of IR and subsequently infected with hrR3 or KOS (wild-type HSV-1 strain). Cell survival was then measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay and trypan blue exclusion cytometry. At 72 hours posttreatment, irradiation with 2 Gy reduced survival from 100% to 76% in noninfected cells, from 61% to 48% in KOS-infected cells, and from 39% to 27% in hrR3-infected PANC-1 cells. As such, analysis of variance indicated that the toxicity of the two modalities was additive. Similar additivity was seen in U-87 MG and CaSki cells. Absolute survival of hrR3-infected or KOS-infected PANC-1 cells decreased as a function of time after treatment (24-72 hours) and multiplicity of infection (MOI) (0.05-5.0). However, the relative decrease in survival with the addition of IR to hrR3 or KOS in PANC-1 cells was not markedly affected by altering MOI (0.05-5.0), time (24-72 hours), radiation dose (2-20 Gy), or cell culture conditions (confluent/growth arrested). We used fluorescence-activated cell sorter analysis with the cationic lipophilic dye DiOC6 to quantify a reduction in mitochondrial membrane potential that'is associated with apoptosis. Fluorescence-activated cell sorter analysis indicated increased apoptosis in both hrR3- and IR-treated cells at 48-72 hours, with hrR3 alone producing the most induction. Viral yields from PANC-1 cells after irradiation and infection were examined. No significant differences were seen between irradiated and nonirradiated cells in viral replication, with hrR3 producing single-step titers of 3.1 +/- 0.9 x 10(5) and 4.0 +/- 1.2 x 10(5) plaque-forming units/mL in nonirradiated and irradiated cells. Thus, complementary toxicity was seen between IR and hrR3 or KOS, regardless of cell type, time, MOI, IR dose, or culture conditions, without evidence of augmented apoptosis or viral replication.
Apoptosis↗
Primary lymphoma of peripheral nerve: report of four cases.
Lymphoma presenting as a solitary tumor of peripheral nerve is exceedingly rare, with only six previously reported cases. The authors describe an additional four cases of primary lymphoma of peripheral nerve involving the sciatic nerve (two cases), the radial nerve, and the sympathetic chain and spinal nerve. The patients were two men and two women with an average age of 55.5 years. All tumors were high-grade B-cell lymphomas. Two patients experienced relapse of disease with involvement of other nervous system sites and died of lymphoma. One patient is alive with stable local disease at 57 months. The fourth patient is alive with no evidence of disease at 54 months. Expression of neural cell adhesion molecule (CD56) has been reported to correlate with an increased incidence of central nervous system involvement in peripheral T-cell lymphoma; all their cases were CD56 negative. Recent reports indicate a high proportion of primary brain lymphomas show loss of CDKN2A/p16 gene expression. Therefore, CDKN2A/p16 was evaluated in their patients both by polymerase chain reaction and by immunohistochemistry for the p16 protein. The authors found homozygous deletion of the CDKN2A/p16 gene in one of three patients studied, confirmed immunohistochemically by absent staining for p16. The fourth patient showed absent staining for p16, suggesting inactivation of the gene in this case as well. The two patients with p16 loss both died of lymphoma, whereas the two patients with normal p16 expression are alive. Primary lymphoma of peripheral nerve is a rare neoplasm, usually of large B-cell type, has a variable prognosis, and appears to have less consistent loss of p16 expression than primary central nervous system lymphoma.
Antineoplastic Combined Chemotherapy Protocols↗
Multicolumn infusion of gene therapy cells into human brain tumors: technical report.
OBJECTIVE: Effective gene therapy for brain tumors may require saturation of the tumors with tumoricidal doses of the therapeutic gene. Safe, precise, and efficient delivery of gene therapy vectors is required. Most reported cases of and published protocols for gene therapy for brain tumors involve freehand injection of retroviral vector-producing cells (VPCs) into the brain. Major disadvantages of this method include the inaccuracy of hand-guided needle placement and limited control of injection parameters. These factors can result in failure to deliver the viral vectors to specifically targeted sites within the brain, extensive tissue disruption resulting from excessively forceful injection, and reflux of the injectate along the needle tract. METHODS: We describe a novel stereotactic strategy for saturating tumor volumes with tumoricidal doses of gene therapy vectors and a new, more precise method of infusing VPCs. With our new instrument, the multicolumn stereotactic infusion system, needle placement is stereotactically guided and both VPC infusion and needle withdrawal are mechanically controlled. RESULTS: This technique, which has been used effectively for six patients, permits precise deposition of columns of VPCs throughout the targeted tumor volume. CONCLUSION: This technique should facilitate saturation of tumors with tumoricidal doses of gene therapy vectors and should improve the results of gene therapy protocols that rely on intraparenchymal injection for delivery.
Biopsy↗
Complement depletion facilitates the infection of multiple brain tumors by an intravascular, replication-conditional herpes simplex virus mutant.
Intravascular routes of administration can provide a means to target gene- and virus-based therapies to multiple tumor foci located within an organ, such as the brain. However, we demonstrate here that rodent plasma inhibits cell transduction by replication-conditional (oncolytic) herpes simplex viruses (HSV), replication-defective HSV, and adenovirus vectors. In vitro depletion of complement with mild heat treatment or in vivo depletion by treatment of athymic rats with cobra venom factor (CVF) partially reverses this effect. Without CVF, inhibition of cell infection by HSV is observed at plasma dilution as high as 1:32, while plasma from CVF-treated animals displays anti-HSV activity at lower dilutions (1:8). When applied to the therapy of intracerebral brain tumors, in vivo complement depletion facilitates the initial infection (assayed at the 2-day time point) by an intra-arterial replication-conditional HSV of tumor cells, located within three separate and distinct human glioma masses. However, at the 4-day time point, no propagation of HSV from initially infected tumor cells could be observed. Previously, we have shown that the immunosuppressive agent, cyclophosphamide (CPA), facilitates the in vivo propagation of an oncolytic HSV, delivered intravascularly, within infected multiple intracerebral masses, by inhibition of both innate and elicited anti-HSV neutralizing antibody response (K. Ikeda et al., Nat. Med. 5:881-889, 1999). In this study, we thus show that the addition of CPA to the CVF treatment results in a significant increase in viral propagation within infected tumors, measured at the 4-day time period. The concerted action of CVF and CPA significantly increases the life span of athymic rodents harboring three separate and large glioma xenografts after treatment with intravascular, oncolytic HSV. Southern analysis of viral genomes analyzed by PCR reveals the presence of the oncolytic virus in the brains, livers, spleens, kidneys, and intestine of treated animals, although none of these tissues displays evidence of HSV-mediated gene expression. In light of clinical trials of oncolytic HSV for malignant brain tumors, these findings suggest that antitumor efficacy may be limited by the host innate and elicited humoral responses.
Adenoviridae↗
Oncolytic viruses as novel anticancer agents: turning one scourge against another.
Although the use of viruses as oncolytic agents is an historic concept, the use of genetically modified viruses to selectively target tumour cells is relatively novel and recent. The ability of viruses to efficiently infect and lyse cells, combined with the potential augmentation of this effect by progeny viruses throughout the tumour provide justification for exploitation of these agents in cancer therapy. Before application to humans, though, issues related to tumour cell selectivity, lack of toxicity to normal tissues and the effect of the antiviral immune response, will have to be clarified. The more commonly used oncolytic viruses are based on mutant strains of herpes simplex virus, adenovirus and reovirus. The tumour selectivity of each of these strains is discussed, particularly the complementation of the viral defect by cellular pathways involved in tumourigenesis. The combination of oncolytic viruses with radiation, chemotherapy and gene therapy is also reviewed. Further study of the interaction of viral proteins with cellular pathways involved in cell cycle control will provide the rationale for viral mutants with increased selectivity for tumour cells.
Adenoviridae↗
Advances in neurofibromatosis 2 (NF2): a workshop report.
Explore the source record for details and available documents.
Adult↗
Solitary sciatic nerve lymphoma as an initial manifestation of diffuse neurolymphomatosis. Case report and review of the literature.
Solitary peripheral nerve lymphomas are exceedingly rare primary manifestations of diffuse peripheral nervous system or central nervous system (CNS) lymphomatosis. A 52-year-old man presented with progressive weakness in gastrocnemius and anterior tibial muscle function, which was associated with radiating pain in the right leg. Magnetic resonance imaging studies revealed a solitary fusiform tumor, extending from the sciatic nerve, at the level of the lesser trochanter of the femur, into the posterior tibial nerve below the popliteal fossa. Intraoperative gross examination found that the tumor diffusely expanded the nerve, but did not extend from or into surrounding muscle or tendons. The final histological diagnosis was a solitary extranodal lymphoma (Burkittlike high-grade B-cell lymphoma). Postoperative staging did not reveal evidence of lymphomatous involvement of other organs, but additional chemo- and radiotherapies were administered. Four months after the surgical biopsy, the patient presented with a right facial nerve palsy. The results of cytological examination of cerebrospinal fluid were positive for the presence of atypical lymphocytes, which was consistent with apparently progressive neurolymphomatosis; however, the results of radiological studies were negative for systemic progression. The patient underwent intrathecal chemotherapy followed by systemic myelosuppressive chemotherapy with bone marrow rescue, but died of respiratory failure while still receiving treatment. Postmortem examination revealed extensive lymphomatosis in the peripheral nerves and spinal nerve roots without evidence of cranial nerve, CNS, or other organ system involvement. The aggressive biological characteristics of these tumors, their management, and pertinent literature are reviewed.
Diagnosis, Differential↗
Thymidine kinase activation of ganciclovir in recurrent malignant gliomas: a gene-marking and neuropathological study.
OBJECT: The gene therapy paradigm of intratumoral activation of ganciclovir (GCV) following transduction of tumor cells by retroviral vectors bearing the thymidine kinase (tk) gene has produced dramatic remissions of malignant gliomas in animal models. In human trials, although the technique has been deemed safe, little antitumor effect has been demonstrated. To evaluate the basis of this inefficacy in human gliomas, the authors conducted a gene-marking trial involving neuropathological and biochemical studies of treated tumor specimens. METHODS: Five patients with malignant recurrent gliomas underwent stereotactic biopsy sampling and intratumoral implantation procedures with three aliquots of 10(6) vector-producing cells (VPCs) in columns. After 5 days, the tumor was resected and the tumor bed reimplanted with VPCs, and a course of GCV was given. Patients received clinical and radiological follow up for 6 months. Tumor specimens were analyzed neuropathologically and for tk gene expression by anti-TK immunohistochemistry and TK enzymatic activity. Four patients tolerated the treatment well but experienced tumor progression. The other developed an abscess after the second operation and died. Increased TK enzymatic activity was demonstrated in the one tumor specimen analyzed. Immunohistochemical evidence of tk gene expression was limited to VPCs. Transduction of tumor cells was not seen. Viable tumor cells were seen near VPCs containing TK. The lymphocytic immune response was mild. CONCLUSIONS: Except for the risk of infection inherent in reoperation, this tk-GCV paradigm was both feasible and safe. Pathological studies indicated that limited dissemination of VPCs and vector from the infusion site and failure to transduce tumor cells with the tk gene are major barriers to efficacy.
Adult↗
An oncolytic herpes simplex virus type 1 selectively destroys diffuse liver metastases from colon carcinoma.
Viruses used for gene therapy are usually genetically modified to deliver therapeutic transgenes and prevent viral replication. In contrast, replication-competent viruses may be used for cancer therapy because replication of some viruses within cancer cells can result in their destruction (oncolysis). Viral ribonucleotide reductase expression is defective in the HSV1 mutant hrR3. Cellular ribonucleotide reductase, which is scarce in normal liver and abundant in liver metastases, can substitute for its viral counterpart to allow hrR3 replication in infected cells. Two or three log orders more of hrR3 virions are produced from infection of colon carcinoma cells than from infection of normal hepatocytes in viral replication assays. This viral replication is oncolytic. A single intravascular administration of hrR3 into immune-competent mice bearing diffuse liver metastases dramatically reduces tumor burden. hrR3-mediated tumor inhibition is equivalent in immune-competent and immune-incompetent mice, suggesting that viral oncolysis and not the host immune response is the primary mechanism of tumor destruction. HSV1-mediated oncolysis of diffuse liver metastases is effective in mice preimmunized against HSV1. These results indicate that replication-competent HSV1 mutants hold significant promise as cancer therapeutic agents. Yoon, S. S., Nakamura, H., Carroll, N. M., Bode, B. P., Chiocca, E. A., Tanabe, K. K. An oncolytic herpes simplex virus type 1 selectively destroys diffuse liver metastases from colon carcinoma.
Animals↗
[Herpes simplex type-1 virus-based vectors for gene therapy].
Recent advances in recombinant nucleic acid technologies and the understanding of the molecular biology of diseases provide the basis for gene therapy's ability to impact on the clinical practice of medicine. Herpes simplex virus type 1 (HSV-1) has features that make it suitable as a vector for gene therapy. These properties include: 1) wide host range and high efficiency of gene transfer; 2) large transgene capacity which is provided by deletion of genes unnecessary for viral replication; 3) unique ability of entering a state of latency in neurons; 4) specific oncolytic effect of some mutants by deletion of certain early genes. Three types of vectors designed for gene therapy have been developed from this virus, termed replication-defective vectors, replication-conditional vectors and amplicon vectors. In this review, we describe the recent advances in HSV-1-based vector systems and their applications for gene therapy.
Animals↗
Multimodal cancer treatment mediated by a replicating oncolytic virus that delivers the oxazaphosphorine/rat cytochrome P450 2B1 and ganciclovir/herpes simplex virus thymidine kinase gene therapies.
Multimodal therapy is generally more effective than single-agent treatment for cancer. rRp450 is an engineered herpes simplex viral mutant that replicates in and kills tumor cells in a relatively selective fashion. It also expresses, in infected cells, the cyclophosphamide (CPA)-sensitive rat cytochrome P450 2B1 (CYP2B1) and the ganciclovir (GCV)-sensitive herpes simplex virus thymidine kinase (HSV-TK) transgenes. We show that cultured rat 9L and human U87deltaEGFR glioma cells, infected and lysed by rRp450, also exhibit supra-additive sensitivity to both CPA and GCV, as determined by Chou-Talalay synergy analysis. DNA cross-linking, assayed by ethidium bromide fluorescence, was significantly inhibited in the presence of GCV, suggesting that interactions between the CPA/CYP2B1 and GCV/HSV-TK gene therapies occurred at the level of DNA repair. In vivo, regression of 9L s.c. tumor volumes in athymic mice was achieved only by the multimodal treatment allowed by rRp450 viral oncolysis combined with CPA/CYP2B1 and GCV/HSV-TK gene therapies, whereas all other treatment combinations produced only tumor growth retardation.
Animals↗
Intraarterial delivery of adenovirus vectors and liposome-DNA complexes to experimental brain neoplasms.
This study investigated the intraarterial delivery of genetically engineered replication-deficient adenovirus vectors (AVs) and cationic liposome-plasmid DNA complexes (lipoDNA) to experimental brain tumors. Adenovirus or lipoDNA was injected into the internal carotid artery (ICA) of F344 rats harboring intracerebral 9L gliosarcomas, using bradykinin (BK) to selectively permeabilize the blood-tumor barrier (BTB). Brain and internal organs of the animals were collected 48 hr after vector injection and stained for expression of the marker gene product, beta-galactosidase (beta-Gal). Intracarotid delivery of AV to 9L rat gliosarcoma without BTB disruption resulted in transgene expression in 3-10% of tumor cells distributed throughout the tumor. Virus-mediated expression of beta-gal gene products in this tumor model was particularly high in small foci (< or = 0.5 mm), which had invaded the normal brain tissue surrounding the main tumor mass. In these foci more than 50% of tumor cells were transduced. BK infusion increased the amount of transgene-expressing cells in larger tumor foci to 15-30%. In the brain parenchyma only a few endothelial cells expressed beta-gal owing to AV-mediated gene transfer. Intracarotid delivery of lipoDNA bearing a cytoplasmic expression cassette rendered more than 30% of the tumor cells positive for the marker gene without BTB disruption. The pattern of distribution was in general homogeneous throughout the tumor. BK infusion was able to increase further the number of transduced tumor cells to more than 50%. Although lipoDNA-mediated gene transfer showed increased efficacy as compared with AV-mediated gene transfer, it had less specificity since a larger number of endothelial and glial cells also expressed the transgene. AV and lipoDNA injections, in the absence and presence of BK, also resulted in transduction of peripheral organs. AV showed its known predilection for liver and lung. In the case of lipoDNA, parenchymal organs such as liver, lung, testes, lymphatic nodes, and especially spleen, were transduced. These findings indicate that intracarotid application of AV and lipoDNA vectors can effectively transduce tumor cells in the brain, and that BTB modulation by BK infusion can further increase the number of transgene-expressing tumor cells.
Adenoviridae↗
Oncolytic virus therapy of multiple tumors in the brain requires suppression of innate and elicited antiviral responses.
The occurrence of multiple tumors in an organ heralds a rapidly fatal course. Although intravascular administration may deliver oncolytic viruses/vectors to each of these tumors, its efficiency is impeded by an antiviral activity present in complement-depleted plasma of rodents and humans. Here, this activity was shown to interact with complement in a calcium-dependent fashion, and antibody neutralization studies indicated preimmune IgM has a contributing role. Short-term exposure to cyclophosphamide (CPA) partially suppressed this activity in rodents and humans. At longer time points, cyclophosphamide also abrogated neutralizing antibody responses. Cyclophosphamide treatment of rats with large single or multiple intracerebral tumors substantially increased viral survival and propagation, leading to neoplastic regression.
Animals↗
Cerebrovascular Biology and the various neural barriers: challenges and future directions.
DESPITE MAJOR ADVANCES in neuroscience, potential therapeutic options for the treatment of central nervous system diseases often cannot be optimized secondary to the presence of the blood-brain barrier (BBB). During the next decade of inquiry, it is crucial that basic science and clinical research that is focused on overcoming the BBB, to optimize delivery to the central nervous system, be identified and supported as a priority topic. For this reason, the third international Cerebrovascular Biology and Blood-Brain Barrier Conference was convened in March 1998 in Gleneden Beach, OR. This meeting brought together basic science and clinical researchers from around the world to analyze BBB function and to discuss delivery of effective agents to the central nervous system for treatment of brain disease. This report summarizes the information presented at the meeting and the discussions that ensued. The current state of knowledge, obstacles to further understanding the BBB, and research priorities are identified.
Blood-Brain Barrier↗
Single-step conversion of cells to retrovirus vector producers with herpes simplex virus-Epstein-Barr virus hybrid amplicons.
We report here on the development and characterization of a novel herpes simplex virus type 1 (HSV-1) amplicon-based vector system which takes advantage of the host range and retention properties of HSV-Epstein-Barr virus (EBV) hybrid amplicons to efficiently convert cells to retrovirus vector producer cells after single-step transduction. The retrovirus genes gag-pol and env (GPE) and retroviral vector sequences were modified to minimize sequence overlap and cloned into an HSV-EBV hybrid amplicon. Retrovirus expression cassettes were used to generate the HSV-EBV-retrovirus hybrid vectors, HERE and HERA, which code for the ecotropic and the amphotropic envelopes, respectively. Retrovirus vector sequences encoding lacZ were cloned downstream from the GPE expression unit. Transfection of 293T/17 cells with amplicon plasmids yielded retrovirus titers between 10(6) and 10(7) transducing units/ml, while infection of the same cells with amplicon vectors generated maximum titers 1 order of magnitude lower. Retrovirus titers were dependent on the extent of transduction by amplicon vectors for the same cell line, but different cell lines displayed varying capacities to produce retrovirus vectors even at the same transduction efficiencies. Infection of human and dog primary gliomas with this system resulted in the production of retrovirus vectors for more than 1 week and the long-term retention and increase in transgene activity over time in these cell populations. Although the efficiency of this system still has to be determined in vivo, many applications are foreseeable for this approach to gene delivery.
3T3 Cells↗
B-myb promoter retargeting of herpes simplex virus gamma34.5 gene-mediated virulence toward tumor and cycling cells.
Deletion of the gamma34.5 gene coding for virulence markedly reduces cytotoxicity mediated by herpes simplex virus type 1 (HSV-1) (J. M. Markert et al., Neurosurgery 32:597-603, 1993; N. S. Markovitz et al. , J. Virol. 71:5560-5569, 1997). To target lytic virulence to tumors, we have created a novel HSV-1 mutant, designated Myb34.5. This viral mutant is characterized by a deletion of the gene for infected cell polypeptide 6 (ICP6; also known as UL39 or ribonucleotide reductase) and of the two endogenous copies of the gamma34.5 gene (RL1) and by reintroduction of one copy of gamma34.5 under control of the E2F-responsive, cellular B-myb promoter. On direct intracerebral inoculation in BALB/c mice, the 50% lethal dose (LD(50)) for Myb34.5 was 2.7 x 10(7) PFU while that for HSVs with mutations in the gamma34.5 gene could not be technically achieved with available viral stocks and it was estimated as >1 x 10(7) PFU. The LD(50) for an HSV with a single defect in ICP6 function was 1.3 x 10(6) PFU. Conversely, Myb34.5's oncolytic efficacy against a variety of human glioma cells in culture and in vivo was enhanced compared to that of HSVs with gamma34.5 mutations, and in fact, it was comparable to that of the wild-type F strain and of viral mutants that possess a wild-type gamma34.5 gene. The characteristic shutoff of host protein synthesis, occurring after infection of human SK-N-SH neuroblastoma cells by gamma34.5 mutant viruses (J. Chou and B. Roizman, Proc. Natl. Acad. Sci. USA 89:3266-3270, 1992), was not present after infection with Myb34.5. There was an increase of almost 3 logarithmic units in the production of progeny virus in arrested fibroblasts compared to that in cycling fibroblasts infected with Myb34.5. These results suggest that transcriptional regulation of gamma34.5 by cell cycle-regulated promoters can be used to target HSV-1 virulence toward tumors while maintaining the desirable neuroattenuated phenotype of a gamma34.5 mutant.
Animals↗
Intra-arterial virus and nonvirus vector-mediated gene transfer to experimental rat brain tumors.
Explore the source record for details and available documents.
Adenoviridae↗