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J G Levy

Publications and source records attributed to J G Levy.

At least 91 records · Page 5Linked to original sources
PubMed1988

Expression of a leukemia-associated antigen (CAMAL) in four myeloid leukemia cell lines.

We have previously described the detection and partial characterization of a common myelogenous leukemia-associated antigen (CAMAL), in CGL and ANLL patients. Both polyclonal and monoclonal (CAMAL-1) antibodies have been raised to p70 (CAMAL) and have been shown to react with both p70 and myeloid leukemia cell preparations. p70 (CAMAL) has been shown to be a monomeric protein of Mr 70,000 and pI 7.2 and was also detectable in the myeloid leukemia cell lines HL60, KG1, K562 and U937, but not in the lymphocytic cell lines Molt-4, Hut-78 and CEM by immunoprecipitation from iodinated cell samples. Using [35S] methionine-labeled cell lines and immunoprecipitation, we have demonstrated the constitutive expression of p70 as well as a major component at p58 and a number at lower molecular weights in the myeloid leukemia cell lines HL60, KG1, K562 and U937, but not in the lymphocytic leukemia cell lines Molt-4, Hut-78 and CEM. The implications of these observations are discussed.

Antibodies, Monoclonal↗
PubMed1988

CAMAL: a new prognostic marker for remission in acute nonlymphoblastic leukemia.

A leukemia-associated antigen, CAMAL, shown to be present in bone marrow (BM) cells from patients with acute nonlymphoblastic leukemia (ANLL) even during remission, has been examined using indirect immunoperoxidase and a monoclonal antibody, CAMAL-1. We are ending the third year of an ongoing blind study designed to monitor the number of BM cells expressing the CAMAL protein (CAMAL BM value) in ANLL patients over the course of their disease. Results thus far have revealed that the CAMAL BM value in individual patients often changed significantly post-chemotherapy. This change appeared to be a useful prognostic marker in many instances. ANLL patients at initial presentation whose CAMAL BM values increased or stayed the same post-chemotherapy had significantly (p less than 0.025) shorter remission lengths (x = 6.8 months, n = 24) than those whose CAMAL BM values decreased (x = 19.2 months, n = 10). There are indications that increasing CAMAL BM values during remission occur prior to clinical relapse.

Acute Disease↗
PubMed1987

A monoclonal antibody raised to tumor-specific T cell-derived suppressor factors also recognizes T suppressor inducer factors of the 4-hydroxy-3-nitrophenyl acetyl hapten suppressor network.

A monoclonal antibody (mAb), B16G, was raised from BALB/c mice immunized with affinity-purified T suppressor factors (TsF) specific for the murine mastocytoma P815. This mAb was found to bind to polyclonal TsF isolated from the spleens of tumor-bearing animals, and to the TsF released from a P815-specific T cell hybridoma. In this study, B16G was tested for its reactivity with TsF produced in the 4-hydroxy-3-nitrophenyl acetyl hapten system. The factors from three types of suppressor T cell hybridomas, each representing the immortalized analogues of the inducer T suppressor cell (Ts1), transducer suppressor cell (Ts2), and effector suppressor cell (Ts3) network populations, were tested. B16G was found to be reactive with two sources of TsF1 as assayed by enzyme-linked immunosorbent assay and delayed-type hypersensitivity bioassay. By contrast, TsF2 and TsF3 were nonreactive with B16G. These results indicate that B16G recognizes class-specific suppressor factor determinants, and that the transducer/effector factors of the network are apparently serologically distinct. Because the B16G mAb fails to recognize 4-hydroxy-3-nitro-phenyl acetyl-specific TsF3 that share idiotype-related determinants with TsF1 yet binds to TsF1 molecules that have interacted with antigen, the binding is apparently independent of the site of antigen recognition. Additionally, the results show that the tumor-specific TsF1 raised in one suppressor system share serologic determinants with anti-hapten TsF1 raised in another.

Animals↗
PubMed1987

Isolation of an antigen-specific T suppressor factor that suppresses the in vivo response of DBA/2 mice to ferredoxin.

A T cell hybridoma (Fd11) has been produced from B10.D2 mice that secretes a putative antigen-specific T suppressor factor (TsF). The TsF is isolable from culture supernatants of Fd11 by affinity purification over columns containing either a monoclonal antibody (B16G) shown previously to be capable of binding murine TsF or ferredoxin (Fd), the nominal antigen to which the Fd11 TsF binds. Specificity of the Fd11 TsF for Fd was established by comparing it to another TsF isolated by us (A10 TsF) in a sandwich ELISA, and by demonstrating the specific reactivity to Fd of the hybridoma in calcium flux studies. The Fd11 affinity-purified TsF was shown to contain two major unique components with m.w. in the region of 80,000 and 35,000 when run on reducing polyacrylamide gels in the presence of sodium dodecyl sulphate. Specific immunosuppressive properties of Fd11 were demonstrated when Fd11 TsF (10 micrograms) was injected i.v. into Fd-primed syngeneic mice at the time of antigen boost. Fd11 TsF specifically and significantly diminished the secondary antibody response to Fd in DBA/2 mice.

Aminoquinolines↗
PubMed1987

Isolation of a T-cell clone that reacts with both antigen and anti-idiotype: evidence for anti-idiotype as internal image for antigen at the T-cell level.

T-cell lines were derived from ferredoxin nonresponder B10.D2 mice that share an idiotype expressed by a monoclonal antibody (Fd-B2) with specificity for one of the two major antigenic determinants (the C determinant) of the antigen. The T-cell line and T-cell clones derived from it release interleukin 2 not only in the presence of anti-Fd-B2 idiotype antibody but in the presence of ferredoxin. The line was shown to be major histocompatibility complex-restricted in that it would respond to the anti-idiotype and antigen only in the context of presentation by cells of the H-2d haplotype. This observation also establishes that the nonresponder status of H-2d animals cannot be attributed to a lesion at the level of antigen presentation. Analysis of the fine specificity of one idiotypic clone showed that it responded only to the anti-idiotype or products of the antigen containing the C determinant, since enzymatically degraded peptides devoid of this determinant did not stimulate these cells. Furthermore, it was found that presentation of both the antigen and the anti-idiotype to the specific clone could be blocked by the Fd-B2 monoclonal antibody.

Animals↗
PubMed1987

Production and characterization of high affinity monoclonal antibodies to cyclic anti-depressant molecules.

A procedure is reported by which high levels of the tricyclic molecule desipramine was modified and conjugated at high density to the carrier molecules keyhole limpet hemocyanin and bovine serum albumin so that these could be used as immunogens in Balb/c mice. Such conjugates generated immune responses with high levels of antibody with specificity for the tricyclic. B cell hybridomas generated by fusion of immune Balb/c splenocytes to NS-1 cells which secreted monoclonal antibodies with specificity for the tricyclic were selected in a standard ELISA. In this report, we show that the binding constants of these monoclonal antibodies with various haptens can be assessed accurately by measuring fluorescence polarization, that a high degree of cross-reactivity between the monoclonals and various tricyclics exists, and that this procedure can be used to generate monoclonal antibodies of high binding constants.

Animals↗
PubMed1987

Antigen-specific T-cell hyporesponsiveness in MRL congenic mice can be explained by two independent cellular defects.

MRL-+, MRL-lpr and B6-lpr have been shown to be useful models in studying systemic lupus erythematosus. MRL-lpr and B6-lpr differ from their congenic counterparts by the presence and expression of the homozygous recessive lymphoproliferation (lpr) gene. One manifestation of this gene is a massive T-cell proliferation that results in a generalized lymphadenopathy in older animals. A paradox that has developed out of the work utilizing the congenic mice is that the gene responsible for lymphoproliferation also appears to be responsible for the inability of T cells to respond to proliferative signals in vitro. In this paper we investigate the basis for this hyporesponsiveness in antigen-induced activation of proliferation and antibody synthesis. We have demonstrated that spleen cells from both MRL-+ and MRL-lpr mice gave minimal stimulation in a one-way mixed lymphocyte reaction against allogeneic T cells. These findings were extended to include antigen-specific proliferation involving antigen that must be processed and presented to responder lymphocytes in a H-2 restricted manner. Thus, MRL-+ and MRL-lpr spleen cells pulsed with ferredoxin also failed to stimulate ferredoxin-primed T cells from B10.Br animals in vitro. We then investigated whether any T-cell defect(s) was also contributing to this proliferative hyporesponsiveness. T lymphocytes from the spleen of MRL-+, 2-month-old MRL-lpr, and 6-month-old MRL-lpr were tested in a one-way mixed lymphocyte reaction. It was found that only the MRL-+ T cells gave responses approaching normal, suggesting lpr gene involvement in T-cell non-responsiveness. This was confirmed by the demonstration of an age-onset T-cell proliferative hyporesponsiveness in B6-lpr mice. This lpr gene-linked non-responsiveness was also shown to extend to T-cell helper function in a positive allogeneic effect assay. We can conclude from these studies that antigenic nonresponsiveness in MRL congenic mice can be explained by two defects: the failure of antigen-presenting cells in MRL-+ and MRL-lpr mice to provide the necessary signal(s) to immunocompetent T cells, this defect not being associated with the lpr gene, and the lpr gene controlled outgrowth of a unique T-cell population that cannot respond in our assay systems.

Animals↗
PubMed1987

Preliminary studies on a more effective phototoxic agent than hematoporphyrin.

Phototoxicity of benzoporphyrin derivative (BPD) has been tested in vitro and compared with that of hematoporphyrin (HP). After 1-hour activation with visible light, BPD was 10 times more cytotoxic than HP toward human adherent cell lines: A549 lung cancer, Calu-1 lung carcinoma, and CCD-19Lu normal lung, killing 100% of cells at the concentration of 70 ng/ml. Under the same conditions, BPD was 10-70 times more cytotoxic than HP toward nonadherent cells and cell lines. Tested were human leukemia cell lines HL60, K562, and KG1, normal human lymphocytes, and mouse mastocytoma cell line P815. The concentrations required to kill 100% of cells varied between 10 and 500 ng BPD/ml and between 0.2 and 10 micrograms HP/ml. The difference between the nonadherent cell lines in respect to their sensitivity to phototoxicity of both BPD and HP seemed to be related to the cell sizes, with the smallest cells being the most vulnerable. The most attractive characteristic of BPD in addition to its powerful phototoxicity is its maximum absorption around 700 nm, which is in the range of wavelengths penetrating tissues the best. This characteristic alone could make BPD a drug of choice in cancer photodynamic therapy when the safety of its use is ensured. Preliminary tests in vivo have shown that DBA/2J mice can tolerate a single ip injection of 20-60 micrograms BPD as well as the same dose of HP. The biodistribution and toxicity studies of BPD are under way in our laboratory.

Animals↗
PubMed1986

Characterization of an anti-idiotypic T cell hybridoma involved in the regulation of the immune response to the P815 mastocytoma.

We report the isolation and characterization of a T cell hybridoma (A29) which secretes a factor that exhibits anti-idiotypic and immune-modulating characteristics. The A29 cell line is thought to represent the hybrid analog of the Ts2 suppressor cell population in the cascade regulating the immune response to the P815 tumor in DBA/2 mice. The putative TsF2 molecule is reactive with the monoclonal antibody B16G, shown previously by us to bind a public specificity of T suppressor factors (TsF). A29 TsF also exhibits specific binding to a TsF1 secreted by another T cell hybridoma, A10, which shows specificity for antigen from the P815 tumor (this has been described previously). A29 itself does not exhibit binding to P815 antigens. Affinity-purified material from A29 appears to share characteristics with A10 molecules in that the predominant material has an apparent m.w. of 70,000. Studies with calcium flux of A29 cells showed that they respond significantly and specifically on exposure to A10 TsF stimulus. We showed further that affinity-purified A29 TsF molecules can specifically suppress the in vitro generation of syngeneic CTL to the P815 tumor, and that panning of DBA/2 splenocytes over A29-TsF-coated plates renders cell populations capable of generating a higher in vitro CTL response to P815 than appropriately treated controls.

Animals↗
PubMed1986

Immunization of DBA/2 mice with a T cell hybridoma-derived TsF increases immune resistance to the syngeneic tumors P815 and L1210.

The ability of a tumor-specific T suppressor factor (TsF) isolated from a T cell hybridoma, A10, to act as an immunogen in DBA/2 mice was investigated. The TsF was affinity purified from ascites over an immunoadsorbent column containing a monoclonal antibody (B16G) that has specificity for the TsF molecule, or over columns containing membrane extracts of the P815 mastocytoma (the tumor for which A10 is specific). The specificity control was BW5147 (the fusion partner for A10) membrane extracts treated in the same way as A10. DBA/2 mice were immunized with the affinity-purified material or PBS and were subsequently challenged with either the P815 tumor or the L1210 DBA/2 thymoma. When mice were immunized with material affinity purified over B16G, eluted material from both A10 ascites and BW5147 membrane extracts enhanced resistance to both P815 and L1210 challenge, indicating that B16G was binding immunogenic material derived from both preparations, which exerted a tumor-protective effect. However, when a P815 affinity column was used, protective material was eluted only from A10 ascites, and this bestowed resistance to both P815 and L1210. When irradiated whole cells were used as immunogens, only A10 cells stimulated anti-tumor immunity, and this appeared to be directed specifically to the P815 tumor. The implications of these findings in terms of the potential for immune modulation with anti-suppressor therapy, and the specificity of the B16G monoclonal, are discussed. The demonstration of B16G binding material (TsF) in the membranes (but not the ascites) of the BW5147 line is also of significance to investigators using BW5147 fused suppressor hybridomas.

Animals↗
PubMed1986

Isolation of functional polyclonal TsF from MRL-lpr spleens using monoclonal antibody absorbance.

We have previously described a monoclonal antibody, B16G, which has been found to be specific for T-cell derived suppressor factors (TsF). B16G has been shown to react with T-suppressor cells, TsF in the spleens of normal or tumor-bearing mice, the TsF produced by a tumor-specific T-cell hybridoma, and with polyclonal whole human TsF isolated from tonsilar tissue. This panreactivity inherent to the B16G MAb has made it clear that it recognizes some common, shared epitope of the TsF molecule. In this study we have used B16G as a probe to isolate TsF from the spleens of MRL-lpr mice and compare the activity with these factors isolated from the spleens of an MHC compatible nonautoimmune strain, CBA. We find that equivalent quantities of functional TsF are isolable from both strains and thus, it can be concluded that the associated oligoclonal B-cell activation characteristic of MRL-lpr mice is not due to a polyclonal T-suppressor cell deficit, nor to the ability of TsC in these mice to produce soluble, functional TsFs. The molecular and biochemical characteristics of these TsFs are discussed.

Animals↗
PubMed1985

Abrogation and reconstitution of nonresponsiveness: a correlation with high network connectivity.

A monoclonal antibody (Fd-B2) to ferredoxin, which bears an idiotype scarcely expressed in any of a wide variety of mouse strains, is able to markedly enhance the response to ferredoxin of both high-responder and intermediate-responder strains. A rabbit anti-idiotype serum to Fd-B2 also specifically enhances the response to ferredoxin in such mice. Most remarkably, the treatment of nonresponder T cells by either the idiotype (Fd-B2) plus complement or anti-idiotype antiserum plus complement causes them to be responsive in adoptive transfer experiments. The two responding populations (idiotype-treated and anti-idiotype-treated) can then be combined to reconstitute the nonresponsive state. When the nonresponders are treated with either Fd-B2 idiotype plus complement or anti-idiotype plus complement and subsequently respond, the idiotype of the anti-ferredoxin antibody produced does not bear the Fd-B2 idiotype. We interpret the results as being consistent with a model in which the unresponsive state for ferredoxin is a state of high network connectivity of the ferredoxin-specific T cells.

Animals↗
PubMed1985

An immunocytochemical investigation with monoclonal antibodies to somatostatin.

Four monoclonal antibodies specific for somatostatin have been produced and characterized. These antibodies were used to assess the anatomical relationship of somatostatin-containing cells in the pancreas and gastrointestinal tract of man, baboon and rat with ten other peptide-containing endocrine cells. The peptides investigated were gastrin, cholecystokinin, motilin, secretin, neurotensin, gastric inhibitory polypeptide, gut-glucagon, pancreatic glucagon, pancreatic polypeptide and insulin. The only regions in which somatostatin cells were seen in close contact with another endocrine cell were in the pancreas and the gastric antrum. In the pancreas somatostatin cells were commonly seen in close contact with insulin, glucagon and pancreatic polypeptide cells and infrequent contact was demonstrable with the gastrin-immunoreactive cells in the antrum of both rat and man. In all other cases no evidence was obtained for a close anatomical relationship between somatostatin cells and the other enteroendocrine cells.

Animals↗
PubMed1985

Preliminary characterization of a soluble immunosuppressive molecule from DBA/2 spleen cells using monoclonal antibody immunoadsorbence.

In a previous publication a monoclonal antibody (B16G) which appeared to recognize T suppressor cells and a T-suppressor factor (TsF) in the spleens of DBA/2 mice was described. B16G appears to be directed to a public specificity of DBA/2 TsF and therefore has been shown to inhibit a variety of immunological reactions. The present study involves preliminary characterization of the material with which B16G reacts. It was found that the B16G-reactive protein (putative TsF) could be absorbed and eluted specifically from a B16G immunoadsorbent column. Material eluting from the B16G column reacted with B16G in an ELISA and appeared to run as two or more bands of 40-45 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The eluted material was biologically active (i.e., suppressive) in the standard assay (mixed leukocyte reaction of DBA/2 splenocytes with B10.BR targets), and its suppressive activity was abrogated by the addition of B16G to the mixed leukocyte reaction cultures. Sephadex G-150 chromatography of the B16G-reactive material showed that under these conditions, its native molecular mass was between 80-90 kDa, indicating that it might occur as a dimer under natural conditions.

Animals↗
PubMed1985

Demonstration of a leukemia-associated antigen (CAMAL) in peripheral blood leucocytes of bone marrow transplant patients prior to relapse.

We have previously described a monoclonal antibody (CAMAL-1) that reacts in an indirect immunoperoxidase slide test at high frequency with cells of patients with acute nonlymphoblastic leukemia (ANLL), both at presentation and in remission (1). This article reports on a 12-month blind study carried out on peripheral blood leukocytes (PBL) of patients who had received bone marrow transplants for acute leukemias. PBL of patients attending the Royal Marsden Hospital were sent as cytospins to the University of British Columbia for staining and screening. Results of this study showed the following: of the 15 patients who remained in remission during the period of the study, 13 showed no abnormal increase in reactivity with CAMAL-1 (2 patients did show increased levels of reactivity over time); of the four patients relapsing but surviving within this period with ANLL, all showed elevated numbers of cells reactive with CAMAL-1 as long as 3 months prior to relapse (the two relapsing patients who had acute undifferentiated leukemia and acute lymphoblastic leukemia did not show elevations of CAMAL-1-reactive cells); of the 14 patients dying during this time of causes other than leukemia, none had elevated levels of CAMAL-1-reactive cells--and, of 4 patients dying in relapse, all had extremely elevated levels of CAMAL-1-reactive cells as long as 4 months prior to relapse. The implications of these observations are discussed.

Acute Disease↗
PubMed1985

Isolation and characterization of a tumor-specific T suppressor factor from a T cell hybridoma.

In our laboratory we have described a monoclonal antibody, B16G, which has been shown to bind to suppressive T cell factors (TsF) in DBA/2 mice. Therefore, B16G was used as a probe to identify T cell hybridomas secreting putative TsF. Hybridomas were obtained by the fusion of DBA/2 thymocytes stimulated in vivo by P815 tumor membrane extracts with the thymoma BW5147. One such hybridoma, A10, was selected and used for additional studies. From both the supernatants and ascites fluid of this hybrid a factor could be obtained that could specifically bind to both B16G and P815 antigen immunoadsorbent columns, and that scored positively with B16G in an ELISA after elution. Such reactivity could not be obtained from A10 supernatants or ascites absorbed over irrelevant columns, nor was it obtained from supernatants or ascites from other T cell hybrids that had scored B16G nonreactive in the original screening. In vivo studies indicated that affinity-purified A10 material injected into DBA/2J mice enhanced significantly the growth of P815 tumor cells, but not the growth of other DBA/2 syngeneic tumor lines such as L1210 or M-I. Additionally, this material did not inhibit the in vitro mixed leukocyte reaction (MLR) between DBA/2 splenocytes and allogeneic B10.BR target cells (unlike B16G purified material from whole DBA/2 spleens, which has been demonstrated to be suppressive in this type of MLR). Biochemical analysis of this tumor-specific TsF from A10 was undertaken; the native m.w. was found to be in the region of 80,000 and 90,000. Under reducing conditions, affinity-purified A10 TsF was found to resolve in SDS-PAGE as what appeared to be a heterodimer of 45,000 and 43,000. In most preparations, an associated molecule resolving at about 25,000 was observed. The implications of these observations are discussed.

Animals↗