Generation of helper-free amphotropic retroviruses that transduce a dominant-acting, methotrexate-resistant dihydrofolate reductase gene.

Miller, A D; Law, Ming-Fan; Verma, Inder M.

doi:10.1128/mcb.5.3.431

Cited by 329 publications

(173 citation statements)

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“…Two recombinant retroviruses were used and their construction is detailed elsewhere (8,(22)(23)(24). The LSNL virus encodes a neomyein resistance (neoO gene for use as a selection marker (22), and the LSNL-HuIL2R virus encodes, in addition, the cDNA ofthe human p55 receptor protein (8).…”

Section: Insertion Of Human Il2 Receptor Into Murine Cell Linesmentioning

confidence: 99%

“…A Pstl-Xbal fragment of a cDNA clone containing the entire p55 receptor coding sequence (25)(26)(27) but lacking the polyadenylation signal was ligated into the LSNL vector using BamHl linkers (8). Both retroviruses were packaged by the \|/2 cell line (23,24). Following infection of FD.C/1 cells with virus, eells were selected for neomyein resistanee by culture in medium supplemented with 100 U/ml [L3 and 2 mg/ml G418, a neomyein analogue.…”

Section: Insertion Of Human Il2 Receptor Into Murine Cell Linesmentioning

confidence: 99%

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Expression of functional human interleukin‐2 receptors in murine interleukin‐3‐dependent cells

et al. 1988

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Summary A murine recombinant retrovirus containing the cDNA encoding the human p55 interleukin-2 (IL2)-binding protein was used to insert this gene into a murine interleukin-3 (IL3)-dependentcell line, FD.C/1. Virus-infected cells, maintained in medium supplemented with IL3, expressedhumanp55on the cell surface and readily adapted to growth using human IL2. In the presence of human IL2. the synthesis ofthe endogenous murine p55 binding protein was induced in FD.C/1 cells, making it difficult to determine whether the human p55 protein was actively involved in the process of growth signal transduction. A cloned cell line, FD.huIL2R-2, was identified which grew in the presence of human IL2 but which had lost the ability to synthesize murine p55 protein. Growth of this clone was inhibited by the monoclonal antibody 2A3 which specifically blocked binding of human 1L2 to the human p55 binding protein. Analysis of restriction enzyme digests of FD.huIL2R-2 cell DNA revealed that a rearrangement of a murine p55 gene had occurred, implying that virus infection had resulted in the integration of retroviral DNA at a site close to or within a murine p55 gene. 1 f IL2 signal transduction involves binding to a surface heterodimeric receptor for IL2, it is argued that FD.huIL2R-2 cells contain an IL2 receptor complex of murine p70 and human p55 IL2-binding proteins. Alternatively, it is possible that integration of human p55 DNA into a site close to a murine p55 gene may lead to a hybrid p55 IL2-binding protein. If FD.huIL2R-2 cells express murine p70 IL2-binding protein as part ofthe receptor complex, the inability of cells to grow in murine IL2 implies that IL2 binding to p70 protein alone is insufficient for a growth signal in these cells. FD.huIL2R-2 cells grow at rates similar in IL3-or human IL2-dependent states. It is likely therefore that the biochemical pathways that control each of these lymphokine-dependent growth states are ver>' similar.

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Section: Insertion Of Human Il2 Receptor Into Murine Cell Linesmentioning

confidence: 99%

Section: Insertion Of Human Il2 Receptor Into Murine Cell Linesmentioning

confidence: 99%

Expression of functional human interleukin‐2 receptors in murine interleukin‐3‐dependent cells

et al. 1988

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“…The development of recombinant retroviruses as gene transfer vehicles (3,38) has provided a system which combines a high efficiency of integration, due to stable insertion of the proviral sequence into the host cell genome as a part of the replicative cycle, with the predictability of integrated proviral structures (3,49). Retroviral constructions designed for the transfer and expression of gene sequences (7,10,11,20,24,31,32,47,52,53) have been reported, and several of these have been introduced into bone marrow stem cells of mice (9,10,20,30,52,53). In addition, the use of amphotropic packaging systems pro-tems.…”

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confidence: 99%

“…Mouse HPRT-BALB/3T3 cells (30), HeLa S3 cells, and amphotropic packaging line PA12 (32), as well as all sublines, were maintained either in Dulbecco modified minimal essential medium or in a 1:1 mixture of Dulbecco modified minimal essential medium and Ham F-12 medium (DMEM-F12). All media were supplemented with 10% fetal calf serum, 2 mM glutamine, 100 pg of streptomycin per ml, and 100 U of penicillin per ml.…”

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Human purine nucleoside phosphorylase and adenosine deaminase: gene transfer into cultured cells and murine hematopoietic stem cells by using recombinant amphotropic retroviruses.

McIvor¹,

Johnson²,

Miller³

et al. 1987

Mol. Cell. Biol.

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Cell lines were established which produced high titers (_ 106 infectious units per ml) of amphotropic, replication-defective recombinant retroviruses which transduced sequences encoding either human purine nucleoside phosphorylase (PNP) or adenosine deaminase (ADA). These viruses also contained a human hypoxanthine phosphoribosyltransferase gene as a selectable marker and a mouse metallothionein promoter (MMP) sequence just upstream from the PNP or ADA genes. Virus structure was maintained through the replication cycle if a short (216-base pair) MMP sequence was used. However, the use of a longer (1,834-base pair) MMP sequence resulted in the deletion of a significant portion of the recombinant virus genome, including the transcriptional regulatory elements of the MMP sequence. Northern analysis indicated a predominance of genome length transcripts in cells infected with deleted virus. The demonstration of substantial human PNP or ADA activity in virus-infected mouse fibroblasts by isozyme analysis suggested that active gene product was translated from either spliced or bicistronic message. The deleted ADA and PNP viruses were introduced into mouse hematopoietic stem cells by cocultivating freshly explanted bone marrow with virus producer cells. The infected marrow cells were injected into irradiated, syngeneic recipient mice, and the presence of integrated ADA or PNP proviral sequences was demonstrated in the DNA of spleen colonies by Southern analysis. Failure of these integrated proviral sequences to express active, human isozyme in spleen colony tissue indicated the existence of some regulatory constraint not active in cultured mouse cells.Purine nucleoside phosphorylase (PNP) and adenosine deaminase (ADA) are enzymes of purine metabolism, the absence of which in humans results in immunodeficiency disease. A severe combined immunodeficiency is associated with the lack of ADA activity (13), and a T-cell immunodeficiency is associated with the lack of PNP activity (12). Both conditions are recessive and autosomally inherited, and afflicted individuals suffer from opportunistic infections that lead to death early in childhood. The metabolic basis of these diseases has been studied intensively (see references 22 and 27 for reviews). Although a number of hypotheses have been proposed to explain the specific toxicity of these conditions to immune cells, there is much evidence that in the absence of one of these enzyme activities, deoxynucleoside substrates accumulate, resulting in unbalanced deoxynucleoside triphosphate pools, feedback inhibition of ribonucleotide reductase, and limited DNA synthesis which does not support an immunoproliferative response (22,27,46).The availability of cloned ADA and PNP coding sequences (14, 48) and progress in gene transfer technology (3,52) have led to the suggestion that the ADA-and PNPassociated immunodeficiencies might be reasonable candidates for initial attempts at human gene therapy (1,33). To * Corresponding author.

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Progress Toward Human Gene Therapy

Morsy¹

1993

JAMA

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Generation of helper-free amphotropic retroviruses that transduce a dominant-acting, methotrexate-resistant dihydrofolate reductase gene.

Cited by 329 publications

References 33 publications

Expression of functional human interleukin‐2 receptors in murine interleukin‐3‐dependent cells

Expression of functional human interleukin‐2 receptors in murine interleukin‐3‐dependent cells

Human purine nucleoside phosphorylase and adenosine deaminase: gene transfer into cultured cells and murine hematopoietic stem cells by using recombinant amphotropic retroviruses.

Progress Toward Human Gene Therapy

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