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Biomedical subjects

V Rousseau

Publications and source records attributed to V Rousseau.

31 records · Page 2Linked to original sources
PubMed1995

Antiviral activity of autocrine interferon-beta requires the presence of a functional interferon type I receptor.

We and others have previously observed that the antiviral effects of autocrine interferon (IFN)-alpha/beta activity cannot be abolished by neutralizing antibodies, even when present to a large excess. This raises the possibility that the major part of autocrine activity is triggered intracellularly, possibly bypassing the transmembrane IFN-alpha/beta receptor. To examine this possibility, cells derived from IFN-alpha/beta Ro/o knockout mice lacking a functional IFN-alpha/beta receptor were stably transformed with pHMB-KbMuIFN beta or pMFG-MuIFN beta plasmids encoding a constitutively expressed murine IFN-beta gene. Four different clones were isolated and examined for resistance to a retrovirus, MFG-LacZ, and to Semliki Forest virus. Despite the production of autocrine IFN-beta at levels inducing high antiviral resistance in control cells, none of the clones displayed antiviral resistance. Thus, despite its failure to be neutralized by potent antiserum, the antiviral activity of autocrine IFN-beta takes place via the transmembrane IFN-alpha/beta receptor, and no additional pathway is involved.

3T3 Cells↗
PubMed1994

Blocking of retroviral infection at a step prior to reverse transcription in cells transformed to constitutively express interferon beta.

We are developing methods for somatic-cell gene therapy directed against infection with human immunodeficiency virus, by enhancing antiviral resistance of target cells through the constitutive production of autocrine interferon (IFN). Using the human IFN-beta coding sequence under the constitutive low-expression control of a 0.6-kb murine H-2Kb promoter-fragment, we have constructed a retroviral vector, HMB-KbHuIFN beta, and have transformed cells of the T98G human neuroblastoma line, the U-937 human promonocytic line, and the CEM human lymphocytic line. These human IFN-beta-transformed cell populations have acquired a low, constitutive production of human IFN, while replicating at a rate similar to that of untransformed cells and of cells transformed with the control vector carrying a human IFN-beta sequence encoding an inactive, mutated protein. In the three different cell populations tested, transformation with the HMB-KbHuIFN beta vector resulted in a 1.3-2.3 log10 reduction in the number of cells infected with a defective amphotropic MFG-LaZ retrovirus. A kinetic study of the fate of the MFG-LacZ retrovirus in the culture medium and intracellularly immediately after exposure of the cells to virus revealed a significant reduction of the appearance of intracellular virus in human IFN-beta-transformed cells. A similar effect was obtained by treating untransformed T98G, U-937, and CEM cells with exogenous human IFN-beta. The blocking effect of autocrine or exogenous human IFN-beta on viral entry was not limited to virus specific for the amphotropic receptor but was also obtained in murine IFN-beta-treated NIH 3T3 mouse fibroblasts infected with an ecotropic MFG-LacZ retrovirus. Infection of human IFN-beta-transformed CEM cells with human immunodeficiency virus type 1 gave comparable results. Immediately following exposure of the cells to human immunodeficiency virus, a kinetic study of the fate of the virus failed to reveal the appearance of intracellular virus and showed that the majority of the input virus remained in the extracellular medium. We conclude that low autocrine IFN-beta synthesis, or exposure of cells to exogenous IFN-beta, prevents virus from getting inside the cells, regardless of the virus receptor involved.

Animals↗
PubMed1994

French multicentre trial comparing Casodex (ICI 176,334) monotherapy with castration plus nilutamide in metastatic prostate cancer: a preliminary report.

This trial compares Casodex (ICI 176,334) monotherapy with the combination of castration (medical or surgical) plus nilutamide. The trial is now closed to entry, 270 patients having been recruited from 32 French centres. As it is too early to present efficacy data, only patient characteristics and interim tolerability data appear in this paper. In the combined treatment group, interstitial pneumonitis (4.5%) was observed, leading to withdrawal from the trial. Other adverse events leading to withdrawal included dyspnoea and ocular problems. There was also 1 case of hepatitis in this treatment group. In the Casodex treatment group, only 6 patients (as compared with 13 in the combined treatment group) withdrew from the trial because of adverse events. As expected with this group, the adverse events were mainly pharmacological effects of an anti-androgen as monotherapy. In the majority of patients, the effects of gynaecomastia and breast tenderness did not result in withdrawal.

Aged↗
PubMed1994

Stable antiviral expression (SAVE) as an approach to somatic cell gene therapy directed against HIV infection.

We are developing methods for somatic cell gene therapy directed against infection with human immunodeficiency virus by enhancing the antiviral resistance of target cells through the constitutive production of interferon-beta. Cells that have been transformed by plasmids or retroviral vectors carrying the human interferon-beta gene placed under the expression control of a murine H2Kb promoter fragment become resistant to HIV infection. Part of this enhanced resistance is due to inhibition of virus entry into the transformed cells, a hitherto unreported mechanism of interferon action.

3T3 Cells↗
PubMed1993

Development of methods for somatic cell gene therapy directed against viral diseases, using retroviral vectors carrying the murine or human interferon-beta coding sequence: establishment of the antiviral state in human cells.

We are developing methods for somatic cell gene therapy directed against chronic and fatal virus infections, such as acquired immunodeficiency (AIDS), by transforming cells with a constitutively expressed interferon (IFN) coding sequence. Previous work from our laboratory has shown that stable antiviral expression (SAVE) can be obtained in murine BALB/c 3T3 cells and human U937 cells transformed with plasmids carrying either the murine or the human IFN-beta coding sequence placed under the expression control of a 0.6-kb Xho II-Nru I promoter region of the murine H-2Kb major histocompatibility complex (MHC) gene (Macé et al., 1991; Seif et al., 1991). In the present paper, we report the construction of murine (Mu) and human (Hu) IFN-beta-expressing retroviral vectors (pMPZen-MuIFN beta, pHMB-KbMuIFN beta) and the problems encountered. Because of the murine origin of commonly used packaging cells and the species specificity of IFN, it was evident that placing the murine IFN-beta sequence under constitutive expression control could result in the production of Mu IFN in the murine packaging system, and thereby lead to decreased vector production and also to enhanced resistance of target cells. Using a packaging cell line that releases a beta-galactosidase-expressing vector, we show that, as expected, Mu IFN-alpha/beta decreases vector production of murine packaging cells and also inhibits the transformation of target NIH-3T3 cells with this vector, but the presence of anti-Mu IFN antibodies rescues the viral titer of the packaging cells and restores the sensitivity of target cells to virus transformation. However, the same antibody treatment is unable to rescue the viral titer of psi-2 packaging cells producing autocrine Mu IFN-beta encoded by the pMPZen-MuIFN beta and pHMB-KbMuIFN beta vectors. Because of the species specificity of IFN, this problem is circumvented with the pMFG-HuIFN beta vector carrying the human IFN-beta sequence. In spite of the production of Hu IFN, murine psi-CRIP packaging cells are able to release retroviral vectors expressing Hu IFN-beta, and these amphotropic vectors can transform human MRC-5 cells and confer to these cells an enhanced resistance to vesicular stomatitis virus (VSV) infection.

3T3 Cells↗
PubMed1990

Enhanced in vitro cytotoxicity of 1,3-bis-(2-chloroethyl)-1-nitrosourea or buthionine sulfoximine combined with a reactive oxygen-generating enzyme immunotoxin.

The glutathione inhibitor drugs, 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and buthionine sulfoximine (BSO), were tested in vitro in order to assess their cytotoxic effectiveness when combined with an enzyme immunotoxin (eIT) composed of a T-cell reactive monoclonal antibody (mAb) 097 coupled to the reactive oxygen-generating enzyme, glucose oxidase (GO) (EC 1.1.3.4). As targets of this eIT we used mature human T-cells or leukemia cells that expressed the 097 epitope. We found that treatment of the cells with subtoxic amounts of mixtures of both a drug and the 097 eIT markedly potentiated cytotoxicity compared to either drug or eIT alone.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1989

Immunotoxins containing glucose oxidase and lactoperoxidase with tumoricidal properties: in vitro killing effectiveness in a mouse plasmacytoma cell model.

We have tested the tumoricidal potency of enzyme immunotoxins constructed of antibodies conjugated to glucose oxidase and to lactoperoxidase. Murine plasmacytoma cells were targeted in vitro with the use of affinity-purified rabbit anti-plasmacytoma membrane antibodies (conjugated to glucose oxidase or lactoperoxidase) or rabbit serum raised against plasmacytoma microsome membranes followed by goat anti-rabbit immunoglobulin conjugates (to glucose oxidase or lactoperoxidase). Cytotoxicity was generated subsequently by incubation of the washed cells in a medium supplemented with glucose and sodium iodide, which were the substrates of these enzymes. This resulted in the presumed metabolic release of highly toxic reduced oxygen species and iodinated derivatives. Targeting of tumor cells with both conjugates, as opposed to one of them alone, produced a synergistic killing effect. The gain of specific versus unspecific cytotoxicity was upwards of 10,000-fold. The killing rates were elevated (t10 values less than 30 min) and linear over time. The resultant reduction in tumor cell viability was in the order of 5 to 6 logs after only 20 to 90 min of incubation in the glucose/NaI medium. Cytotoxicity was enhanced by the gamma-glutamyl cysteine synthetase inhibitor buthionine-S,R-sulfoximine and by the glutathione reductase inhibitor 1,3-bis(2-chloroethyl)-1-nitrosourea, while catalase was inhibitory. The results suggest that these enzyme immunotoxins may be suitable for the ex vivo purging of autologous bone marrow grafts.

Animals↗
PubMed1989

An immunotoxin system intended for bone marrow purging composed of glucose oxidase and lactoperoxidase coupled to monoclonal antibody 097.

The effectiveness of an antibody-enzyme immunotoxin (eIT) was investigated on human T cells. This enzyme immunotoxin contained glucose oxidase (GO) and lactoperoxidase (LPO) chemically coupled to the pan-leukocyte-specific mouse monoclonal antibody (MoAb) 097 (097-GO and 097-LPO). Human peripheral blood mononuclear cells or tumor cells were suspended in a mixture of 097-GO and 097-LPO for 30 min, and then for 2 h with glucose and NaI. The effectiveness of this eIT system was indicated by the almost complete reduction of T cell viability, as estimated by a phytohemagglutinin induced proliferation assay (99.4% +/- 0.31 depletion, mean +/- SEM of 15 experiments). The specificity of the cytotoxicity reaction was indicated by the lack of cytotoxicity of control irrelevant MoAb conjugates to T cells (1.9% +/- 4.17 of T cell depletion, eight experiments). The growth of human bone marrow myeloid progenitors (CFU-GM) was not affected by the conjugates even by increasing 100-fold the optimal cytotoxic dose. T cells were susceptible to the conjugates in the presence of up to 90% of erythrocytes. This eIT system may thus represent a new alternative immunospecific procedure for allograft and/or autograft purging, and appears to effectively replace complement-mediated methods of T cell depletion.

Antibodies, Monoclonal↗
PubMed1988

A new strain-specific cross-reactive idiotope with possible regulatory function expressed on BALB/c anti-alpha (1-3) dextran antibodies.

We obtained a mAb, B5, defining a major crossreactive Id, the B5 Id, having an in vivo regulatory function. BALB/c mice immunized against the bacterial dextran Ag B1355S produce lambda 1-bearing anti-alpha (1-3) glucosidic linkage of dextran B1355S (DEX) antibodies. Of these antibodies 30% expressed the B5 Id determinant. As was observed for the IdX-Id, B5 idiotope expression was also linked to the BALB/c H chain allotype. The antibody family expressing the B5 Id included IdX+ as well as IdX- defined monoclonal anti-alpha (1-3) DEX antibodies. Injection into BALB/c mice of mAb B5 conjugated to keyhole limpet hemocyanin led to enhancement of serum anti-alpha (1-3) DEX antibody synthesis, and to a strong increase in B5+ Ig lacking dextran-binding activity. IdX+ serum Ig were also increased in these mice. Injection of this mAb into allotype congenic nonresponder C.B20 mice induced a significant serum lambda 1-bearing anti-alpha (1-3) dextran antibody response (5 to 50 micrograms/ml). However, in A/J nonresponder mice, a similar treatment induced only the dextran-nonbinding component, which reached high levels (approximately 7 mg/ml) in the sera of these mice. In the majority of these A/J mice, we detected a specific anti-(4-hydroxy-5-iodo-3-nitrophenyl)acetyl antibody response characterized by B5 Id-positive antibodies. This observation can be explained by an in vivo regulatory idiotope-mediated cross-regulation.

Adjuvants, Immunologic↗
PubMed1986

Monoclonal anti-alpha (1----3) dextran antibodies of Igha BALB/c and Ighb C.B20 mice display striking similarities.

Allotype Ighb congenic C.B20 mice when immunized with dextran B1355S are unable to produce anti-alpha (1----3) dextran antibodies that express the VH-associated cross-reactive IdX idiotype. This intrastrain-specific idiotype is normally associated only with the anti-dextran response of Igha mice of which BALB/c is a prototype strain. In this study we have obtained monoclonal hybridoma antibodies specific for the alpha (1----3) glucosidic linkage of dextran from C.B20 mice that were presensitized with rabbit anti-IdX antibodies. These antibodies display the light chain isotype distribution, the H chain amino terminal sequence, share VH-associated IdX idiotypic determinants, and finally the similar fine specificity for dextrans observed for anti-alpha (1----3) dextran antibodies of BALB/c mice.

Amino Acid Sequence↗
PubMed1986

In-vitro cytolysis of myeloma tumor cells with glucose oxidase and lactoperoxidase antibody conjugates.

We have assessed the tumoricidal potential of enzyme-antibody conjugates on murine myeloma cells. Conjugates of glucose oxidase (EC 1.1.3.4) and lactoperoxidase (EC 1.11.1.7) were specifically targeted on the NSO tumor cells. Optimal conditions for tumor cell killing, as assayed by [51Cr] release required the binding of both antibody conjugates to the cell membrane. This is followed by washing and incubation in medium containing glucose and 0.1 mM iodide. Under these conditions 90% of the incorporated [51Cr] labeled is released from the cells, and NSO clonogenicity is reduced by a factor greater than 5 logs by 2 h of incubation.

Animals↗