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H J Federoff

Publications and source records attributed to H J Federoff.

At least 37 records · Page 2Linked to original sources
PubMed2001

Functional interaction between fluorodeoxyuridine-induced cellular alterations and replication of a ribonucleotide reductase-negative herpes simplex virus.

G207 is an oncolytic herpes simplex virus (HSV) which is attenuated by inactivation of viral ribonucleotide reductase (RR) and deletion of both gamma(1)34.5 genes. The cellular counterparts that can functionally substitute for viral RR and the carboxyl-terminal domain of ICP34.5 are cellular RR and the corresponding homologous domain of the growth arrest and DNA damage protein 34 (GADD34), respectively. Because the thymidylate synthetase (TS) inhibitor fluorodeoxyuridine (FUdR) can alter expression of cellular RR and GADD34, we examined the effect of FUdR on G207 bioactivity with the hypothesis that FUdR-induced cellular changes will alter viral proliferation and cytotoxicity. Replication of wild-type HSV-1 was impaired in the presence of 10 nM FUdR, whereas G207 demonstrated increased replication under the same conditions. Combined use of FUdR and G207 resulted in synergistic cytotoxicity. FUdR exposure caused elevation of RR activity at 10 and 100 nM, whereas GADD34 was induced only at 100 nM. The effect of enhanced viral replication by FUdR was suppressed by hydroxyurea, a known inhibitor of RR. These results demonstrate that the growth advantage of G207 in FUdR-treated cells is primarily based on an RR-dependent mechanism. Although our findings show that TS inhibition impairs viral replication, the FUdR-induced RR elevation may overcome this disadvantage, resulting in enhanced replication of G207. These data provide the cellular basis for the combined use of RR-negative HSV mutants and TS inhibitors in the treatment of cancer.

Animals↗
PubMed2001

Efflux of glutathione conjugate of monochlorobimane from striatal and cortical neurons.

Evidence for the presence of a novel transporter in primary cultures of rat striatal neurons and mouse cortical neurons similar in function to the multidrug resistance-associated protein (MRP1) is presented. Functional activity was assessed by efflux studies with the glutathione conjugate of monochlorobimane (B-SG). The glutathione transferase-catalyzed formation of B-SG in rat striatal neurons and mouse cortical neurons was inhibited by ethacrynic acid. The efflux of B-SG from rat striatal neurons and mouse cortical neurons was lower at 20 degrees C than at 37 degrees C and was lower in cells with reduced ATP concentrations compared with cells with constitutive ATP concentrations. In addition, the efflux of B-SG was inhibited by MK-571 in both rat striatal and mouse cortical neurons and by probenecid in rat striatal neurons, but not in mouse cortical neurons. Verapamil did not inhibit B-SG efflux in either rat striatal or mouse cortical neurons. Although functionally similar to MRP1, Western blot analysis with commercially available antibodies directed against human and mouse MRP1 failed to show MRP1-like protein in either whole-cell homogenates of rat striatal neurons or mouse cortical neurons, indicating that the described neuronal transporter differs in structure from human or mouse MRP1 or lacks epitopes in common with MRP1.

ATP-Binding Cassette Transporters↗
PubMed2001

HSV vector-mediated gene delivery to the central nervous system.

The efficient and targeted transfer of genes is the goal of gene therapy. In the central nervous system (CNS), this is challenging due in part to the exquisite anatomy of the brain. Herpes simplex virus (HSV) vectors are particularly amenable to CNS therapies as they are capable of transducing a variety of cells, have a large transgene capacity and can exist as either oncolytic or non-immunogenic vectors. The versatility of this vector platform and its potential molecular therapeutic use in two CNS disorders, Alzheimer's disease and malignant brain tumors, will be discussed.

Alzheimer Disease↗
PubMed2000

Gene-experience interaction alters the cholinergic septohippocampal pathway of mice.

Spatial learning requires the septohippocampal pathway. The interaction of learning experience with gene products to modulate the function of a pathway may underlie use-dependent plasticity. The regulated release of nerve growth factor (NGF) from hippocampal cultures and hippocampus, as well as its actions on cholinergic septal neurons, suggest it as a candidate protein to interact with a learning experience. A method was used to evaluate NGF gene-experience interaction on the septohippocampal neural circuitry in mice. The method permits brain region-specific expression of a new gene by using a two-component approach: a virus vector directing expression of cre recombinase; and transgenic mice carrying genomic recombination substrates rendered transcriptionally inactive by a "floxed" stop cassette. Cre recombinase vector delivery into transgenic mouse hippocampus resulted in recombination in 30% of infected cells and the expression of a new gene in those cells. To examine the interaction of the NGF gene and experience, adult mice carrying a NGF transgene with a floxed stop cassette (NGFXAT) received a cre recombinase vector to produce localized unilateral hippocampal NGF gene expression, so-called "activated" mice. Activated and control nonactivated NGFXAT mice were subjected to different experiences: repeated spatial learning, repeated rote performance, or standard vivarium housing. Latency, the time to complete the learning task, declined in the repeated spatial learning groups. The measurement of interaction between NGF gene expression and experience on the septohippocampal circuitry was assessed by counting retrogradely labeled basal forebrain cholinergic neurons projecting to the hippocampal site of NGF gene activation. Comparison of all NGF activated groups revealed a graded effect of experience on the septohippocampal pathway, with the largest change occurring in activated mice provided with repeated learning experience. These data demonstrate that plasticity of the adult spatial learning circuitry can be robustly modulated by experience-dependent interactions with a specific hippocampal gene product.

Animals↗
PubMed2000

Enhanced learning in mice parallels vector-mediated nerve growth factor expression in hippocampus.

Spatial learning requires the integrity of the nerve growth factor (NGF)-responsive septohippocampal pathway. Loss of a single NGF allele at the mouse NGF locus (heterozygous null, ngf(+/-)) reduces septohippocampal NGF levels and NGF-regulated cholinergic neurotransmitter enzymes and results in spatial learning deficits in adult animals. A herpes simplex virus (HSV) amplicon vector was utilized to locally deliver NGF to the hippocampus of mice heterozygous and wild type (ngf(+/+)) at the NGF gene locus. NGF gene transfer produced transient increases in NGF protein levels and choline acetyltransferase activity in both ngf(+/-) and ngf(+/+) mice. However, spatial learning capability was improved only in ngf(+/-) mice. In aggregate, these findings suggest that amplicon-directed expression of NGF in subjects with baseline septohippocampal dysfunction can correct spatial learning deficits.

Animals↗
PubMed2000

Discordance between expression and genome transfer titering of HSV amplicon vectors: recommendation for standardized enumeration.

Herpes simplex virus-derived amplicon vectors are well suited to the development of gene-based therapy for neurodegenerative diseases. The plasmid-based amplicon vector system allows for facile introduction of transcription units, possesses the potential for carrying gene inserts up to approximately 130 kb in length, and can be packaged into infectious virus devoid of contaminating cytotoxic helper virus. For accurate assessments to be made regarding vector comparison and improvements in vector design, a standard for titering prepared virus stocks must be established. At present, packaged amplicon vectors are routinely titered using reporter gene expression units to quantitate numbers of infectious amplicon virions. The strength of the promoter, sensitivity of detection of the gene product, and choice of titering cell type can greatly influence the apparent numbers of infectious virus particles. This is especially evident when comparisons are made between two amplicon vectors that possess different promoters. To this end, we have developed a new titering method based on a real-time quantitative PCR technique that allows for enumeration of transducing particles. This new approach ensures that amplicon comparison experiments are initiated with equivalent transduction units, thus allowing for a fair assessment of expression and therapeutic efficacy differences.

3T3 Cells↗
PubMed2000

Overexpression of GluR6 in rat hippocampus produces seizures and spontaneous nonsynaptic bursting in vitro.

We hypothesized that overexpression of specific glutamate receptors within the hippocampus would induce seizures and the associated cellular changes seen in temporal lobe epilepsy (TLE). The GluR6 kainate receptor was overexpressed by injecting rat hippocampi with HSVGluR6, a viral vector transducing fully edited GluR6. These animals experienced limbic seizures approximately 4 h following the injection. Control animals injected with HSVlac, a vector expressing beta-galactosidase, did not have seizures. Recordings from hippocampal CA1 pyramidal cells were performed 12 to 48 h and 1 week to 1 month postinjection. We observed nonsynaptic Na(+)-mediated bursting in 77.5% of cells 12 to 48 h following injection of HSVGluR6 but not HSVlac. The synaptic responses were normal in both groups. However, the physiological properties of cells from HSVGluR6-injected hippocampi changed over time. Two weeks following HSVGluR6 injection, synaptic bursts could be evoked, but intrinsic bursting became rare. These changes persisted for at least 1 month. We postulate that this transition from intrinsic to synaptic hyperexcitability may be important in the development of TLE.

Action Potentials↗
PubMed2000

Repeated acquisition and performance chamber for mice: a paradigm for assessment of spatial learning and memory.

Molecular genetic manipulation of the mouse offers the possibility of elucidating the function of individual gene products in neural systems underlying learning and memory. Many extant learning paradigms for mice rely on negative reinforcement, involve simple problems that are relatively rapidly acquired and thus preclude time-course assessment, and may impose the need to undertake additional experiments to determine the extent to which noncognitive behaviors influence the measures of learning. To overcome such limitations, a multiple schedule of repeated acquisition and performance was behaviorally engineered to assess learning vs rote performance within-behavioral test session and within-subject utilizing an apparatus modified from the rat (the repeated acquisition and performance chamber; RAPC). The multiple schedule required mice to learn a new sequence of door openings leading to saccharin availability in the learning component during each session, while the sequence of door openings for the performance component remained constant across sessions. The learning and performance components alternated over the course of each test session, with different auditory stimuli signaling which component was currently in effect. To validate this paradigm, learning vs performance was evaluated in two inbred strains of mice: C57BL/6J and 129/SvJ. The hippocampal dependence of this measure was examined in lesioned C57BL/6J mice. Both strains exhibited longer latencies and higher errors in the learning compared to the performance component and evidenced declines in both measures across the trials of each session, consistent with an acquisition phenomenon. These same measures showed little or no evidence of change in the performance component. Whereas three trials per session were utilized with C57BL/65 mice in each component, behavior of 129/SvJ mice could only be sustained for two trials per component per session, demonstrating differences in testing capabilities between these two strains under these experimental conditions and thus precluding the ability to make systematic strain comparisons of learning capabilities. Hippocampal lesions in C57BL/6J mice resulted in substantially longer latencies and increased errors in the learning but not the performance component, demonstrating the importance of this region to spatial learning as measured in the RAPC. In aggregate, this positive reinforcement-based operant paradigm to evaluate murine spatial learning detects strain differences and hippocampal dependence and permits explicit differentiation of the impact of noncognitive contributions to learning measures on a within-subject, within-session basis.

Animals↗
PubMed1999

Gene therapy in the inner ear. Mechanisms and clinical implications.

The application of gene therapy to the inner ear is an emerging field of study. Most studies report the expression of marker genes (e.g., galactosidase) within the tissues of the cochlea. The first biologic response of an inner ear tissue (i.e., auditory neurons) to transduction by a gene therapy vector expressing a therapeutic gene (a herpes amplicon vector containing a BDNF gene) was observed in spiral explants obtained from early postnatal rat cochleae. This study was important because it demonstrated the feasibility of a gene augmentation approach to treat traumatized cochleae. Long-term expression of transduced or transfected genes in cochlear tissues have been obtained with adenovirus, adeno-associated virus, and herpes amplicon vectors. The herpes amplicon vector (i.e., HSVbdnflac) that evoked a biologic response in vitro has also been successfully used to support the survival of auditory neurons in vivo following loss of the auditory hair cells (i.e., loss of trophic factor). Gene therapy has been successfully applied to the cochlea of a laboratory animal, and future studies will define the types of vectors and therapeutic genes that will work best for the treatment of inner ear diseases in the clinic.

Adenovirus E1 Proteins↗
PubMed1999

Hypoxia-inducible factor-1alpha mediates hypoxia-induced delayed neuronal death that involves p53.

Hypoxia-induced delayed neuronal death is known to require de novo gene expression; however, the molecular mediators that are involved remain undefined. The transcription factor hypoxia-inducible factor-1alpha (HIF-1alpha), in addition to promoting the expression of adaptive genes under conditions of hypoxia, has been implicated as being a necessary component in p53-mediated cell death in tumors. Using herpes amplicon-mediated gene transfer in cortical neuronal cultures, we demonstrate that delivery of a dominant-negative form of HIF-1alpha (HIFdn), capable of disrupting hypoxia-dependent transcription, reduces delayed neuronal death that follows hypoxic stress. In contrast, hypoxia-resistant p53-null primary cultures are not protected by HIFdn expression. These data indicate that, in hypoxic neurons, HIF-1alpha and p53 conspire to promote a pathological sequence resulting in cell death.

Animals↗
PubMed1999

Selective infection and cytolysis of human head and neck squamous cell carcinoma with sparing of normal mucosa by a cytotoxic herpes simplex virus type 1 (G207).

This study evaluates inhibition of human squamous cell carcinomas (SCCs) by a replication-competent multimutated herpes simplex virus type 1 (G207). Infectivity and cytotoxicity of the G207 virus were evaluated in vitro in seven human SCC cell lines. In vivo effects of the G207 virus on human tumor xenografts in an athymic rat model were then investigated by injecting established tumors with 1 x 10(7) virus particles and monitoring tumor growth. In addition, oral cavity tumors in immunocompetent hamster were infected with the G207 virus by selective intraarterial perfusion and the tumor response was monitored. In vitro studies demonstrated infection rates, measured 24 hr after exposure, exceeding 40% at an MOI of 2 in five of seven human SCC cell lines. Cytotoxic effects, as measured by percent cell death on day 5, exceeded 90% in five of seven SCC cell lines. In vivo inhibition of tumor growth in an athymic rat model was seen (p < 0.005) and in two of the cell lines a complete clinical response was seen in 12 of 14 tumors. In the hamster model, selective intraarterial perfusion with G207 virus showed selective infection of the tumor cells, with sparing of the adjacent normal mucosa, which leading to significant suppression of tumor growth (p < 0.005). The G207 virus displayed efficient and selective cytotoxicity and tumor growth inhibition against human SCC and may prove useful as a therapeutic agent for head and neck SCC.

Animals↗
PubMed1999

Paraquat elicited neurobehavioral syndrome caused by dopaminergic neuron loss.

The herbicide paraquat, bearing structural similarity to the known dopaminergic neurotoxicant MPTP, has been suggested as a potential etiologic factor in Parkinson's disease. Consideration of paraquat as a candidate neurotoxicant requires demonstration that systemic delivery produces substantia nigra dopaminergic neuron loss and the attendant neurobehavioral syndrome reflecting depletion of dopamine terminals within the striatum. To address these issues paraquat was administered systemically into adult C57 bl/6 mice, ambulatory behavior monitored, substantia nigra dopamine neuron number and striatal dopamine terminal density quantified. The data indicate that paraquat like MPTP elicits a dose-dependent decrease in substantia nigra dopaminergic neurons assessed by a Fluoro-gold prelabeling method, a decline in striatal dopamine nerve terminal density assessed by measurement of tyrosine hydroxylase immunoreactivity; and neurobehavioral syndrome characterized by reduced ambulatory activity. Taken together, these data suggest that systemically absorbed paraquat crosses the blood-brain barrier to cause destruction of dopamine neurons in the substantia nigra, consequent reduction of dopaminergic innervation of the striatum and a neurobehavioral syndrome similar to the well characterized and bona fide dopaminergic toxin MPTP.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1999

Focal hippocampal gain of NGF function elicits specific septal cholinergic reorganization.

The influence of NGF on the neuroanatomic substrate(s) subserving learning and memory was probed by somatic mosaic analysis. NGF XAT mice underwent unilateral hippocampal delivery of virus vector expressing cre recombinase to produce focal NGF gain of function mosaics. Activated mice expressing increased NGF, but not control mice transduced with the lacZ gene showed reorganization of the septohippocampal projection assessed by retrograde labeling with Fluorogold (FG). Mice mosaic for NGF function demonstrated in ipsilateral medial septum and diagonal band significant increases in FG-labeled cell bodies (94%), ChAT-labeled cells (55%), double-positive cells (190%) and increased somal size of double-positive cells (56%) than did non-activated mice. These results indicate that reorganization of the cholinergic septal input to a specific hippocampal region is promoted by gain of NGF function.

Animals↗
PubMed1999

Eradication of pre-established lymphoma using herpes simplex virus amplicon vectors.

Herpes simplex virus amplicon vectors expressing RANTES (HSVrantes) and the T-cell costimulatory ligand B7.1 (HSVB7.1) were studied for their ability to elicit a tumor-specific T-cell response in a murine lymphoma model. HSVB7.1- and HSVrantes-transduced EL4 cells expressed high levels of B7.1 and RANTES as analyzed by flow cytometry and enzyme-linked immunosorbent assay, respectively. Inoculation of ex vivo HSVB7.1 transduced cells in syngeneic mice resulted in regression of both transduced cells and nontransduced cells inoculated contralaterally. Direct intratumoral injection of HSVB7.1 and/or HSVrantes alone or in combination into established EL4 tumors led to complete tumor regression in injected tumors as well as in nontransduced contralaterally implanted tumor, whereas control tumors or tumors injected with HSVlac expressing beta-galactosidase did not regress. Maximal protection was achieved with combined injection of HSVB7.1 and HSVrantes; mice showing tumor regression were resistant to rechallenge with parental EL4 cells, and tumor cell-specific cytolytic T-cell activity was observed in mice demonstrating regression. HSV amplicon-mediated delivery of immune effector molecules may represent a useful strategy for immunotherapy in the setting of pre-existing tumor.

Animals↗
PubMed1999

BCL-2 transduction, using a herpes simplex virus amplicon, protects hippocampal neurons from transient global ischemia.

Transient global ischemia results in selective neuronal damage of hippocampal CA1 neurons. Five minutes of bilateral common carotid artery occlusion, in the Mongolian gerbil, effectively restricts forebrain blood flow, resulting in a delayed neuronal death of CA1 pyramidal cells. While there is a delay of approximately 72 h in the appearance of cell death, markers related to the mechanism of ischemic death become apparent well before neurons die. Ischemia-induced increases in the cell-death-promoting protein, bax, may disrupt the bcl-2/bax ratio necessary for normal neuronal functioning and thus promote transient ischemic death. In order to locally maintain this critical bcl-2/bax ratio and thus protect CA1 neurons from delayed neuronal death, a herpes simplex viral (HSV) vector was used to selectively introduce human bcl-2, under the control of the herpes IE 4/5 promoter, into the CA1 region of the gerbil hippocampus. Twenty-four hours prior to ischemia surgery, 1 microl of HSVbcl-2 was infused unilaterally into the CA1 region at a rate of 2 nl/min. Seventy-two hours after ischemia the animals were sacrificed and processed using Nissl, silver degeneration, and immunohistochemical (anti-human bcl-2) staining. Immunohistochemistry demonstrated both glial and neuronal bcl-2 expression around the HSVbcl-2 infusion site. The evaluation following silver degeneration staining indicated a further degeneration of CA1 neurons in the immediate area of the viral vector infusion. This damage seems to be the result of cellular debris associated with the processing of the viral amplicons. Silver degeneration staining is not present in the areas that demonstrate bcl-2 staining. These neurons appear to have been rescued from ischemic damage. This result was confirmed using the Nissl staining. Therefore, by altering the local ratio of bcl-2/bax using the HSVbcl-2 vector one may protect CA1 pyramidal cell from the delayed neuronal death of transient global ischemia.

Animals↗
PubMed1999

HIF-1alpha and p53 promote hypoxia-induced delayed neuronal death in models of CNS ischemia.

Brain ischemia is a cause of substantial morbidity and mortality during the later decades of life. In light of this, many studies have used in vitro and in vivo models of acute necrosis to test candidate therapeutic agents. More recently, the existence of a genetically programmed component of ischemic death has become widely accepted. We have used molecular genetic approaches to investigate the potential link between hypoxia-induced gene transcription and the delayed death of ischemic neurons. Hypoxia-induced gene expression is an evolutionarily conserved response comprising both transcriptional activation and posttranscriptional and posttranslational stabilization events. Members of the PER-ARNT-SIM (PAS) family of basic helix-loop-helix transcription factors have been shown to regulate hypoxic transcripts in nonneuronal cultured lines. However, evidence for ischemic activation of PAS proteins within the neuronal compartment or possible involvement in neuronal death is lacking. The tumor-suppressor protein p53 is a known transcriptional activator within the central nervous system that is clearly involved in the pathologic response to ischemia. This article will provide data that implicate the coordinate activities of p53 and the PAS protein HIF-1alpha in driving ischemia-induced delayed neuronal death. Background regarding mechanisms of ischemic neuronal death will also be provided with special attention paid to the role of de novo gene expression in promoting this pathologic sequence. The identification of the HIF-1alpha/p53-mediated signaling pathway in neurons highlights a novel target toward which anti-ischemic neuroprotective drug discovery can be applied.

Animals↗
PubMed1999

Oncolytic viral therapy for human colorectal cancer and liver metastases using a multi-mutated herpes simplex virus type-1 (G207).

G207 is a multi-mutated, replication-competent type-1 herpes simplex virus designed to target, infect, and lyse neurological tumors. This study examines the feasibility of using G207 in the treatment of human colorectal cancer and defines the biological determinants of its antitumor efficacy. This virus was tested on five human colorectal cancer cell lines in vitro to determine efficacy of infection and tumor cell kill. These results were correlated to measures of tumor cell proliferation. In vivo testing was performed through direct injections of G207 into xenografts of human colorectal cancer tumors grown in flanks of athymic rats. To evaluate an alternate method of administration, hepatic portal vein infusion of G207 was performed in a syngeneic model of liver metastases in Buffalo rats. Among the five cell lines tested, infection rates ranged between 10% and 90%, which correlated directly with S-phase fraction (8.6%-36.6%) and was proportional to response to G207 therapy in vitro (1%-93%). Direct injection of G207 into nude rat flank tumors suppressed tumor growth significantly vs. control (0.58 +/- 0.60 cm(3) vs. 9.16 +/- 3.70 cm(3), P<0. 0001). In vivo tumor suppression correlated with in vitro effect. In the syngeneic liver tumor model, portal infusion resulted in significant reduction in number of liver nodules (13 +/- 10 nodules in G207-treated livers vs. 80 +/- 30 nodules in control livers, P<0.05). G207 infects and kills human colorectal cancer cells efficiently. In vitro cytotoxicity assay and tumor S-phase fraction can be used to predict response to treatment in vivo. This antineoplastic agent can be delivered effectively by both direct tumor injection and regional vascular infusion. G207 should be investigated further as therapy for colorectal cancer and liver metastases.

Animals↗