Showing papers in "Human Gene Therapy in 1990"
TL;DR: A retroviral vector recently has been used to mark tumor infiltrating lymphocytes in patients with melanoma to follow the persistence of these cells following infusion into patients.
Abstract: Retroviral vectors promote the efficient transfer of genes into a variety of cell types from many animal species. An important contribution to their utility was the development of retrovir...
449 citations
TL;DR: The experiments reported here document the feasibility of using adenovirus for the direct delivery in vivo of a gene to restore an impaired metabolism and observe the relatively long-term presence of the transferred gene.
Abstract: Mutant mice of the Spf-ash strain have an inherited defect in ornithine transcarbamylase (OTC) protein synthesis, and were used to ascertain the potential of recombinant adenoviruses for i...
402 citations
TL;DR: The results of this study demonstrate that chemosensitivity genes can enhance the efficacy of treatment in hosts who subsequently develop a neoplasm and imply the merits of exploring an alternative version of the strategy in which somatic insertion of chemos sensitivity genes in mosaic fashion is used prophylactically to enhance the prospect that a subsequent tumor will respond to therapy.
Abstract: The dose limitations imposed on cancer chemotherapeutic agents by their lack of selectivity can, in theory, be circumvented by a strategy entailing the prophylactic insertion into hosts of drug-sensitivity genes that are acquired or expressed in some but not all cells. This strategy predicts that neoplasms arising from drug-sensitive cells might be safely treatable with tumor-eradicative drug doses because the presence of a modicum of drug-insensitive stem cells will protect vital tissues from lethal depopulation. To test this prediction, lymphomas were induced with Abelson leukemia virus in mice bearing a herpes simplex virus thymidine kinase (HSV-TK) transgene selectively expressed in lymphoid cells. Of 12 transgenic mice treated with the HSV-TK-specific substrate ganciclovir (GCV), 11 exhibited complete tumor regressions; 5 of these mice remained tumor-free over observation periods that exceeded 100 days. Among the lymphomas that recurred, most appeared to represent mutant subpopulations that ...
226 citations
TL;DR: A small number of patients diagnosed with severe combined immunodeficiency with adenosine deaminase deficiency also have a history of cytokine release syndrome, suggesting that these patients are at higher risk of developing cytokine-receptor-based diseases.
Abstract: SCIENTIFIC ABSTRACT Severe combined immunodeficiency (SCID) due to deficiency of the purine metabolic enzyme adenosine deaminase (ADA) is a fatal childhood immunodeficiency disease. Immune reconsti...
220 citations
TL;DR: Once very safe retrov virus vector-helper cell systems are constructed and in use, safety considerations should not hold up further human trials of retrovirus vectors.
Abstract: Somatic gene therapy of human disease with retrovirus vectors is a new technology with potentially important medical benefits. Although it involves recombinant DNA technologies and modified retroviruses, proper design of the vectors and delivery systems removes most potential foreseen risks. Furthermore, even in the very remote possibility that there is a nontherapeutic biological effect of the treatment, it is unlikely to be a harmful one. Thus, once very safe retrovirus vector–helper cell systems are constructed and in use, safety considerations should not hold up further human trials of retrovirus vectors.
159 citations
TL;DR: In the 5 animals studied (combined mean follow-up of 25.7 months), clinical illness has not been identified at any time and murine amphotropic retroviruses do not appear to pose an acute health risk.
Abstract: The in vivo fate of amphotropic murine leukemia retrovirus was studied in five rhesus monkeys. Retrovirus infused intravenously into 3 normal animals and 1 immunosuppressed animal was cleared rapidly from the circulation and subsequent viremia has not been detected (mean follow-up of 27.4 months). A fifth monkey was immunosuppressed and transplanted with virus-producing autologous fibroblasts in addition to an intraperitoneal injection of virus. This animal was viremic for 2 days and its lymph node cells and peripheral blood mononuclear cells were shown to be producing virus for up to 22 days post-inoculation, but subsequently has been negative after 17.0 months of analysis. In the 5 animals studied (combined mean follow-up of 25.7 months), clinical illness has not been identified at any time. Therefore, murine amphotropic retroviruses do not appear to pose an acute health risk.
155 citations
TL;DR: The results suggest that techniques that produce high rates of gene transfer into long-lived human HPC will facilitate studies to quantitate expression of exogenous genes in hematopoietic cells and may be applicable to clinical gene therapy.
Abstract: We have examined the ability of the recombinant hematopoietic growth factors (HGF) interleukin-3 (IL-3), IL-6, and granulocyte-macrophage colony-stimulating factor (GM-CSF) to increase retroviral vector-mediated gene transfer into human hematopoietic progenitor cells (HPC). The efficiency of neo gene transfer by the N2 vector into human HPC was enhanced by preculture with either GM-CSF or IL-3 (but not IL-6) and with each combination of the three factors. The combination of IL-3 plus IL-6 consistently produced significantly higher levels of G418-resistant colonies (50–60%) than any of the other combinations of HGF tested. Following preculture with HGF and transduction by N2, marrow was maintained in long-term bone marrow culture (LTBMC) for 2 months. The levels of G418-resistant HPC remained stable, and no apparent depletion of total HPC content resulted from the prior exposure to highly stimulatory doses of factors. The proliferative status of the HPC, following exposure to the HGF, was measured...
127 citations
TL;DR: Quantification of data obtained from PCR dilution experiment indicates that, under appropriate conditions, amplification is linear with respect to the amount of input DNA, permitting estimations of gene dosage in unknown samples.
Abstract: The polymerase chain reaction (PCR), a widely used new technology, was applied to several aspects of retroviral-mediated gene transfer. Ten oligonucleotide primer pairs were analyzed for their ability to amplify specific regions of a retroviral vector, including the long terminal repeat (LTR), and a NeoR selectable marker gene. By using the appropriate oligonucleotide primers, cells transduced by retroviral vectors could readily be detected and analyzed for deletions in proviral sequences by PCR, without Southern blotting. In combination with a simplified RNA isolation/reverse transcription protocol, an approximate titer of vector producer cell lines could be estimated by PCR in a single day, eliminating the need for time-consuming transductions and biological selection. Quantification of data obtained from PCR dilution experiment indicates that, under appropriate conditions, amplification is linear with respect to the amount of input DNA, permitting estimations of gene dosage in unknown samples....
93 citations
TL;DR: This review attempts to summarize the progress that has been made in the expression of genes introduced into hematopoietic cells and the difficulties still remaining before meaningful application of gene transfer methods can be expected to cure human diseases of bone marrow-derived cells.
Abstract: The use of retroviral vectors allows efficient transfer of genes into a variety of mammalian cells. A focus of research over the past 6 years has been the use of retroviral vectors to effe...
89 citations
TL;DR: Results show that amphotropic vectors are capable of infecting pluripotent hematopoietic stem cells having long-term repopulating ability, and that a variety of promoters allow gene expression following differentiation of these early cells.
Abstract: Three retroviral vectors, containing a human adenosine deaminase (ADA) cDNA linked to either the simian virus 40 (SV40) early promoter, the human cytomegalovirus (CMV) immediate early prom...
58 citations
TL;DR: It is suggested that gene-modified T lymphocytes can survive and circulate for long periods in vivo and can continue to express the introduced genes.
Abstract: Lymphocytes can be readily transduced with retroviral vectors and the gene-modified lymphocytes will stably express the inserted genes in vitro for long periods. As a prelude to studies in...
TL;DR: The history of the ethical debate on human gene therapy is examined and the extended debate has lasted from 1980 to the present, but now adaptation, i.e., a public policy, for somatic cell gene Therapy is emerging.
Abstract: Ethical issues generally evolve through four stages: threshold, open conflict, extended debate, and adaptation. The history of the ethical debate on human gene therapy is examined. The threshold was the Nirenberg appeal in 1967. The open conflict centered around two early controversial cases: those of Rogers and Cline. The extended debate has lasted from 1980 to the present, but now adaptation, i.e., a public policy, for somatic cell gene therapy is emerging.
TL;DR: The results indicate that great care should be given to the transfer of unwanted passengers, like VL30, present in retroviral packaging cell lines like the psi 2 cells, which are currently being used for gene therapy.
Abstract: FTO-2B rat hepatoma cells acquired mouse VL30 retrotransposon(s) when infected with Moloney murine leukemia virus (MoMLV) recombinant retroviruses produced from Ψ2 cells. The VL30 provirus was integrated into the rat genome, expressed at high levels, and its transcription induced 40-fold by dexamethasone. VL30 RNA was detected in hepatoma cells even without selection for the expression of the amino-3′-glycosyl phosphotransferase (neo) gene, which was co-transferred with a MoMLV retrovirus. However, the extent of transfer of the VL30 RNA was inversely related to the titer of the MoMLV recombinant retrovirus. The restriction map analysis of the transferred VL30 provirus was identical to the mouse VL30s of the NVL subfamily which is known to be a significant fraction of the transcriptionally active VL30 subset. Additionally, the regenerating liver from an adult rat, which was infected with a defective MoMLV-derived retrovirus, expressed VL30 RNA. These results indicate that great care should be give...
TL;DR: Analysis of T-cell receptor heterogeneity indicated that the transduced TIL recovered from the tumor biopsy were different from TIL that were kept strictly in vitro and selected in G418, suggesting a simultaneous selection for long-term in vitro growth advantage.
Abstract: Patients with malignant melanoma have been treated with interleukin-2 (IL-2) and tumor-infiltrating lymphocytes (TIL) marked by retroviral gene transduction. The retroviral vector contained a gene coding for the bacterial enzyme neomycin phosphotransferase, such that transduced TIL expressing the enzyme could survive otherwise toxic concentrations of the neomycin analogue G418. For 1 patient, who exhibited a complete regression of cancer after treatment with TIL, lymphocytes from post-treatment blood and tumor biopsies were cultured in IL-2, and transduced TIL were recovered by G418 selection. Analysis of T-cell receptor heterogeneity indicated that the transduced TIL recovered from the tumor biopsy were different from TIL that were kept strictly in vitro and selected in G418. The selection process required weeks in culture, during which time control cultures changed radically in subset composition, so there was also a simultaneous selection for long-term in vitro growth advantage. It cannot be c...
TL;DR: A retroviral vector (GTN) in which the glucocerebrosidase (GCase) cDNA is driven by the Moloney murine leukemia virus (Mo-MuLV) long terminal repeat (LTR) was tested for transfer efficiency and expression of the GCase gene in long-term reconstituted mice as discussed by the authors.
Abstract: A retroviral vector (GTN) in which the glucocerebrosidase (GCase) cDNA is driven by the Moloney murine leukemia virus (Mo-MuLV) long terminal repeat (LTR) was tested for transfer efficiency and expression of the GCase gene in long-term reconstituted mice. Eleven W/Wv mice were transplanted with unselected GTN-infected bone marrow cells and 10 of these mice were analyzed 3 months later. Seven of these 10 mice (70%) contained the intact proviral genome in bone marrow, spleen, and thymus. Of these 7,3 mice contained a high-copy number of the provirus in all the hematopoietic tissues tested. The mice contained anywhere from one to four proviral integration sites that were the same in all three tissues, indicating that these mice have been repopulated by one or more transduced multipotential hematopoietic stem cells. Five months after transplantation, bone marrow from the eleventh mouse was transplanted into secondary recipient animals. The secondary recipients contained the intact proviral genome in the bone marrow, spleen, thymus, and macrophages 4 months after the secondary transplantation. This further supports the conclusion that hematopoietic stem cells have indeed been targeted. Human GCase RNA was detected in all 7 mice containing the proviral DNA. These results demonstrate expression of the human GCase gene in the progeny of repopulating hematopoietic stem cells of mice following gene transfer.
TL;DR: The public's involvement in the debate over human gene therapy is used to illustrate two points: complex bioethical issues often come to the public's attention because of a scandal, in this case the Cline episode; enlightened discussion and public understanding are difficult to achieve under these conditions.
Abstract: The public's involvement in the debate over human gene therapy is used to illustrate two points. First, complex bioethical issues often come to the public's attention because of a scandal, in this case the Cline episode; enlightened discussion and public understanding are difficult to achieve under these conditions. Second, even with unfavorable circumstances at the outset, the debate can become channeled into politically responsible institutions which then can develop effective public policy. For gene therapy, four significant forums emerged: the President's Commission's Report Splicing Life, the 1982 Congressional Hearings, the OTA Report, and the RAC's Points to Consider document.
TL;DR: The sources and reach of the NIH "Points to Consider" are examined, which are based on normative considerations inherited from two sets of science policy deliberations that took place in the United States during the 1970s: the discussion of research with human subjects and the recombinant DNA debate.
Abstract: In this essay, I examine the sources and reach of the NIH “Points to Consider.” These guidelines are based on normative considerations inherited from two sets of science policy deliberations that took place in the United States during the 1970s: the discussion of research with human subjects and the recombinant DNA debate. The combined lessons of those deliberations provide six criteria by which to evaluate human gene therapy proposals. While these criteria could be used to reject proposals to attempt germ-line gene therapy or enhancement engineering today, they provide no principled basis for publicly proscribing the development of these forms of genetic intervention. Instead, they will ultimately lead us to approach the moral limits of gene therapy as a professional policy question about the goals of medicine, rather than as a social policy question about the public good.
TL;DR: It is argued that an act- oriented ethics is inadequate and that only a virtue-oriented ethics enables us to recognize and resolve the new problems ahead of us in genetic manipulation.
Abstract: The investigation into the specific moral issues of genetic manipulation requires us to determine exactly the new moral issues of genetic manipulation. But even that determination requires us to consider whether the context in which we live and the method of moral reflection which we use is adequate enough to address genetic manipulation. Given the liberalist context in which we live, this paper argues that an act-oriented ethics is inadequate and that only a virtue-oriented ethics enables us to recognize and resolve the new problems ahead of us in genetic manipulation. Moreover, those problems have a common root, that is, that through genetics we will be in danger of objectifying the human subject.
TL;DR: If and when germ-line gene therapy is contemplated, Congress will be faced with difficult choices, but will likely take no action to block trials that appear safe and are intended to produce clinical benefit for particular individuals.
Abstract: Congress was the scene of conspicuous debate about human gene therapy during the 1980s. Congressional interest was sparked primarily by concerns about germ-line gene therapy expressed by clerics and public interest groups. The initial debate was provoked by Martin Cline's misadventures in 1980 and rekindled in 1983 by congressional resolution against germ-line intervention sponsored by Senator Mark Hatfield. The first hearing on gene therapy was held upon the release of the President's Commission report Splicing Life, in November, 1982, before a House subcommittee chaired by Congressman Albert Gore, Jr. Representative Gore later requested a report on gene therapy, which was released by the Office of Technology Assessment in December, 1984. He also sponsored the legislation that established the Biomedical Ethics Board and Biomedical Ethics Advisory Committee, Congress's abortive attempt to reestablish a federal bioethics commission. Implications of advances in human genetics, including gene therapy, were to be among the first topics addressed. Congress passed no substantive legislation affecting gene therapy research or clinical trials, but served principally as a national theater for debate. If and when germ-line gene therapy is contemplated, Congress will be faced with difficult choices, but will likely take no action to block trials that appear safe and are intended to produce clinical benefit for particular individuals.
TL;DR: The way in which religious concerns and their spokesman have now entered into discussions within the scientific community is of notable importance.
Abstract: Beginning with critical writings of certain theological ethicists in 1965, religious bodies increasingly have been fostering studies of emerging issues implied by genetic technology. While...
TL;DR: Now is the time for the scientific and medical communities to come together and to cooperate to make human gene therapy a clinically useful procedure.
Abstract: The use of molecular techniques to correct human genetic diseases is a concept that was considered extremely remote by many investigators until quite recently. Several factors were responsible for changing the scientific community's attitude toward gene therapy: the development of recombinant DNA technology including the ability to clone disease-related genes; maturation of scientific and ethical reflection following apparent failures of early human experiments; and the development of efficient techniques for the transfer of genes into mammalian cells. Now is the time for the scientific and medical communities to come together and to cooperate to make human gene therapy a clinically useful procedure.
TL;DR: Two principal recommendations have been made for setting the boundaries between somatic cell versus germ-line correction and the amelioration of disease and the enhancement of traits.
Abstract: Human genetic modification has begun without a clear consensus on where the moral boundary lines should be placed to insure that the technology of human genetic engineering is not abused. Two principal recommendations have been made for setting the boundaries. The first is between somatic cell versus germ-line correction; the second is between the amelioration of disease and the enhancement of traits. Each proposal involves a distinction and a rule. There is a dilemma in that the first case involves a well-grounded distinction but a dubious rule, while the second offers a more favored rule, but a fuzzy distinction.
TL;DR: The four classifications of genetic manipulation (somatic cell gene therapy, germ-line gene Therapy, enhancement genetic engineering, eugenic genetic engineering) are reviewed from an ethical viewpoint.
Abstract: The four classifications of genetic manipulation (somatic cell gene therapy, germ-line gene therapy, enhancement genetic engineering, eugenic genetic engineering) are reviewed from an ethical viewpoint. Immediately, it needs to be recognized that words like "therapy," "enhancement," and "eugenic" already possess an emotional framework, so that careful reasoning is made more difficult. In each of the areas of genetic manipulation, it can be argued that circumstances could be imagined that would either justify or not justify proceeding. Based on our present state of knowledge, the line should be drawn at not carrying out gene therapy of any type except in specific cases that are carefully evaluated in advance. With increased knowledge, the line should be moved to embrace appropriate situations.
TL;DR: The first, and possibly still the most important, document examining in depth the broad ethical implications of human gene therapy was Splicing Life, published in November 1982, and the senior author reflects on the significance of that study from the perspective of today.
Abstract: Overview summaryThe first, and possibly still the most important, document examining in depth the broad ethical implications of human gene therapy was Splicing Life, published in November 1982. The senior author of that study looks back over the past years and reflects on the significance of Splicing Life from the perspective of today.
TL;DR: The hope of human gene therapy and of other clinical applications of the human genome initiative is in danger because of the obstacles to research in reproductive biology and human genetics in the federal sector.
Abstract: A critical analysis is undertaken of the ethical debate on two general classes of research activities that bear on the future of human gene therapy: (i) the human fetus and (ii) the preimplantation human embryo after in vitro fertilization (IVF). The conclusion is that the hope of human gene therapy and of other clinical applications of the human genome initiative is in danger because of the obstacles to research in reproductive biology and human genetics in the federal sector.