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Biomedical subjects

D L Morton

Publications and source records attributed to D L Morton.

At least 163 records · Page 9Linked to original sources
PubMed1991

Lymphatic drainage of skin to a sentinel lymph node in a feline model.

To determine the feasibility of selective lymphadenectomy, the authors developed a feline model to identify and determine the utility of mapping dyes for this purpose. Adult cats were injected intradermally with a variety of mapping substances to determine whether the anatomic site of injection had a predictable pattern of drainage to a particular lymph node. Isosulfan blue provided the optimal mapping material. Injection of isosulfan blue intradermally into the skin of the medial thigh consistently led to coloration of the central lymph node, whereas intradermal abdominal wall injections and intradermal lateral thigh injections resulted in coloration of the lateral lymph node. Intradermal injections into skin about the perineum resulted in coloration of the most medial lymph node only. The feline model proved to a useful model to examine the utility of mapping dyes and to demonstrate dermal lymphatics. The predictable pattern of drainage of the skin in this feline model supports the feasibility of selective lymphadenectomy.

Animals↗
PubMed1991

Role of gangliosides in active immunotherapy with melanoma vaccine.

Among various tumor associated cell surface antigens, gangliosides, the glycosphingolipids that contain sialic acids, offer a variety of epitopes, some of which are preferentially expressed on melanoma cells. These surface components of the bilayered lipid membrane of tumor cells are the targets of active immunotherapy with melanoma vaccine. Purified gangliosides in aqueous solution form micelles and, at high density, form lactones. Their antigenic expression (physical conformation and orientation) on the cell surface is governed by the nature of the sphingosine and the fatty acids they contain. Evidence is accruing to show that the nature of the fatty acid moiety of gangliosides differs in normal and neoplastic cells. Gangliosides per se are not immunogenic and require extrinsic adjuvanticity. Preparation of a melanoma cell vaccine for active immunotherapy requires an understanding of the ganglioside profile of melanoma, the ganglioside-associated heterogeneity of melanoma, and the role of shed melanoma gangliosides in the immunosuppression of cell mediated and humoral immunity. In addition, the role of some of the anti-ganglioside antibodies in the elimination of shed gangliosides, the cytotoxic killing of tumor cells, as well as in the down-regulation of lymphocyte functions must be considered in the formulation of vaccine. Different strategies for augmenting the immunogenicity of melanoma associated gangliosides with melanoma vaccine are evaluated.

Clinical Trials as Topic↗
PubMed1990

Suppressor cell activity in a randomized trial of patients receiving active specific immunotherapy with melanoma cell vaccine and low dosages of cyclophosphamide.

Previous studies have shown that melanoma patients develop an immune response to cell surface melanoma-associated antigens. The presence of this antibody response to cell surface antigens has been correlated with a better clinical outcome when melanoma patients are treated with an allogeneic melanoma cell vaccine (MCV) as an active immunotherapy protocol. It was hypothesized that the inability to consistently induce or enhance existing immune responses to melanoma-associated antigens was related to the downregulation by suppressor cells. Patients received treatments of MCV 3 times in a 4-week interval and then every fourth week. The biological response modifier cyclophosphamide (CYP) is an immunomodulator of suppressor T-cell function. In this study we set out to determine whether CYP given prior to MCV could reduce suppressor cell activity during vaccination. In a randomized trial stage II and III melanoma patients (n = 41) were given MCV alone or in conjunction with CYP at dosages of 300, 150, or 75 mg/m2. CYP was given 3 days prior to each MCV treatment. Suppressor cell activity in patients was monitored by a concanavalin A suppressor assay using peripheral blood lymphocytes from serial phlebotomies during a 12-week period of treatment. In each trial group there were patients who had major reduction in suppressor cell activity (greater than 50%). Overall, the greatest reduction in suppressor cell activity occurred in patients receiving 300 mg/m2 CYP compared to the other CYP dosages or MCV alone. For the first two treatments at all CYP dosages there was a greater number of patients showing reduced suppressor cell activity compared to later treatments. In a comparison of patients receiving MCV alone to MCV + CYP 300 mg/m2 phenotypic analysis of lymphocyte subsets showed significant (P = 0.03) reduction in the CD8+CD11B+ (suppressor) cells of the latter group. These studies suggest that CYP can be used at low dosages in conjunction with MCV to reduce suppressor cell activity.

Adult↗
PubMed1990

Characterization of a 90-100 kDa tumor-associated antigen in the sera of melanoma patients.

Using allogeneic antibody, we previously described a high-molecular-weight glycoprotein in the urine of 68% of melanoma patients. This glycoprotein has been termed urinary-tumor-associated antigen (U-TAA). A murine monoclonal antibody (MAb) specific for U-TAA (ADI-40F4) has been developed. By the use of ADI-40F4, U-TAA was detected in serum samples from 63% (33/52) of stage II and stage III melanoma patients, but from only 5% (1/20) of normal controls. This report describes the physical and immunochemical properties of U-TAA in the serum. The antigen elutes from a DEAE-Sephacel column in association with IgG in the void volume and as free antigen in a second peak. The molecular mass of the free antigen is 590-620 kDa and it sediments in the region of 28-29% sucrose by density gradient ultracentrifugation. Free antigen has an isoelectric point of 6.1. This high molecular weight antigen is composed of smaller subunits linked by reducible bonds. The ADI-40F4 reactive epitope resides on a 90-100 kDa subunit. These results provide evidence that U-TAA which is produced by melanoma cells in vitro is present in the circulation of melanoma patients.

Antibodies, Monoclonal↗
PubMed1990

Lymphocyte subset alterations in nodes regional to human melanoma.

The lymphocyte subpopulations in tumor-draining lymph nodes of melanoma patients were determined using two-color flow cytometry. Data were analyzed according to: (a) the staging of the melanoma; (b) whether or not the nodes contained tumor; and (c) their distance from the primary tumor. Compared with Stage I patients (without metastasis), uninvolved nodes of stage II patients (with nodal metastases) had a significant decrease in helper/inducer (CD4+) T-cells (P less than 0.001), with a corresponding increase in cytotoxic/suppressor (CD8+) cells (P less than 0.001) and Leu 19+ natural killer (CD56+) cells (P less than 0.01). In some patients the presence of tumor within a node was associated with a large decrease in CD3+ total T-cells, whereas in others tumor involvement had little influence on lymphocyte phenotypes. When analyzed by distance from the primary tumor, nodes closest to tumor in Stage I patients contained a smaller percentage of CD19+ B-cells. In Stage II, tumor-free nodes nearest to tumor showed an increase in CD19+ cells, but statistical significance was not reached. CD56+ natural killer cells increased progressively in nodes near tumor and were more numerous in Stage II uninvolved nodes compared with Stage I nodes. Alterations in phenotypically defined lymph node lymphocytes occur in nodes regional to melanoma as the disease progresses, as growth of metastases occurs, and in tumor-free nodes nearest to tumor. These alterations may be essential to the establishment and progression of metastases.

Adult↗
PubMed1990

Detection and isolation of antigen-specific immune complexes from sera of melanoma patients.

Detection of antigen-specific circulating immune complexes (IC) in sera from cancer patients provides an approach for defining tumor antigens that induce a host immune response and could prove useful for purposes of immunoprognosis. In the present study, a sandwich ELISA was developed to detect antigen-specific IC in cancer patients, utilizing a murine monoclonal antibody designated MAb JSI. The MAb was produced by cell fusion using standard hybridoma technology following immunization with a partially purified fetal antigen that had been isolated from the spent culture medium of a melanoma cell line. The partially purified antigen appeared to be broadly expressed on melanomas, sarcomas, and carcinomas. The MAb was mass-produced in pristine-primed mice. MAb JSI reacted with the cultured melanoma cell M14 but not the autologous lymphoblastoid cell L14. Following purification by ammonium sulfate precipitation, the MAb was immobilized on polystyrene plates and utilized to capture antigen-specific immune complexes from sera of melanoma patients which were detected with anti-human globulins. The second antibody in the sandwich was shown to be endogenous human IgG. Antigen-specific immune complexes were present in melanoma patients but only infrequently in sera from normal individuals and patients with active autoimmune disease. Antigen-specific immune complexes detected in melanoma sera were isolated by affinity chromatography utilizing the MAb JSI. The assay described in this report is simple and reproducible, without plate effect (p = 0.97) or time effect (p = 0.34) and could provide a useful new approach to examine the role of antigen-specific circulating immune complex analysis in cancer patients.

Analysis of Variance↗
PubMed1990

Tumor-associated antigen immune complexes. A potential marker of recurrent melanoma.

To determine the potential utility of antigen-specific immune complex analysis, we developed a competitive enzyme-linked immunosorbent assay utilizing polyclonal human antibody to detect tumor-associated antigen-specific immune complexes. Sera from 10 normal volunteers and 19 patients with recurrent melanoma were studied. Patients with recurrent melanoma had a mean +/- SD percent inhibition of 27.6% +/- 29.8% in contrast to normal individuals with a mean value of 8.4% +/- 17.8%. A monoclonal antibody (MAb JSI) was developed following immunization with a partially purified antigen. Utilizing MAb JSI, we developed a "sandwich" enzyme-linked immunosorbent assay and studied sera from 45 normal volunteers and 44 patients with cancer with recurrent melanoma. Results were expressed as a percent maximum binding of a positive control. The mean +/- SD percent maximum binding for normal subjects was 4.9% +/- 7.7% in contrast to sera from patients with melanoma who had a mean of 38.3% +/- 33.3%. Serial analysis of four patients with melanoma with tumor-associated antigen-specific immune complexes demonstrated the presence of tumor-associated antigen-specific immune complexes up to 12 years prior to clinical recurrence.

Adult↗
PubMed1990

The impact of microinvasion on axillary node metastases and survival in patients with intraductal breast cancer.

A rational approach to the local treatment of intraductal breast cancer continues to generate considerable debate. However, the finding of an invasive component in intraductal breast cancer is widely regarded as an appropriate indication for axillary node dissection as part of the local treatment and staging of this disease. Despite this view, the natural history of patients with intraductal breast cancer with foci of microinvasion is poorly defined. Between 1965 and 1988, 41 patients with this pathologic finding of intraductal carcinoma with foci of microinvasion were seen at the UCLA Medical Center. Twenty-three patients presented with mammographic abnormalities, while 17 patients presented with a palpable mass. One patient presented with Paget's disease of the nipple. Thirty-three patients underwent axillary node dissection as part of their local treatment. No lymph node metastases were identified. The median follow-up in 37 patients was 47 months. There have been no local recurrences and no deaths from recurrent breast cancer. Intraductal breast cancer associated with microinvasion appears to be an extremely favorable lesion with minimal risk of nodal metastases.

Adult↗
PubMed1990

Natural history and selective management of in transit melanoma.

Ninety five patients who developed in transit melanoma either as their initial site of recurrent melanoma or following regional node metastases were retrospectively reviewed. In transit melanoma occurred most frequently on the lower extremity and appeared to be associated with deeply invasive primary tumors. The median time to development of in transit melanoma was 16 months. Eighty-two (86%) of these patients have progressed to systemic disease from 2 to 244 months (median 16 months) following the development of in transit melanoma, and 72 (79%) died (median survival 19 months). Survival appears to correlate with the extent of in transit melanoma and with the disease-free interval. These findings suggest that in transit melanoma represents an early manifestation of systemic disease, warranting careful clinical follow-up and perhaps systemic treatment, when effective therapy becomes available. However, some patients will respond to local immunotherapy, surgical excision, and isolated limb perfusion and will enjoy significant length and quality of life. This sequential approach remains the treatment of choice for this manifestation of metastatic melanoma.

Adolescent↗
PubMed1990

Association between allo-immunoreactive and xeno-immunoreactive subunits of a glycoprotein tumor-associated antigen.

Using allogeneic antibody, we previously described a tumor-associated antigen (TAA) in the urine of 68% of melanoma patients. The TAA was purified from urine of a melanoma patient and used as immunogen to develop a murine monoclonal antibody (AD1-40F4) and xenopolyclonal antibodies in a baboon. Sera from melanoma patients treated with whole melanoma cell vaccine were used as the source of human antibody to the glycoprotein antigen. Treatment with 2-mercaptoethanol and separation by sodium dodecyl sulfate/polyacrylamide gel electrophoresis resolved the high-molecular-mass glycoprotein TAA into smaller subunits. Immunoblot analysis indicates that the murine monoclonal antibody (AD1-40F4) recognized a 90-100-kDa subunit of the antigen while human anti-TAA antibodies primarily recognized a 65-kDa subunit in addition to the 90-100-kDa subunit. Baboon polyclonal antibodies recognized the same subunits plus a 120-kDa subunit. Blocking studies indicated that the murine monoclonal and baboon polyclonal antibodies recognized the closely related epitopes on the 90-100-kDa subunit, while human antibodies recognized an epitope entirely distinct from that recognized by the mouse antibody. These results demonstrate the epitope complexity associated with the high-molecular-mass glycoprotein TAA.

Animals↗
PubMed1990

Immunochemical characterization of a tumor-associated antigen defined by a monoclonal antibody.

A murine monoclonal antibody of the subclass IgG2b, designated MAb JSI, was produced utilizing standard hybridoma technology. Female BALB/c mice were immunized with a fetal antigen that had been partially purified from the spent serum-free culture media of a human melanoma cell line. Utilizing MAb JSI, the antigen was isolated from serum of melanoma patients by affinity chromatography utilizing an acid elution and studied following exposure to trypsin, protease, and heat in an enzyme-linked immunosorbent assay (ELISA). The antigenic activity was destroyed following treatment with trypsin and protease as well as by exposure to heat (100 degrees C). By immunoperoxidase staining, MAb JSI reacted with melanoma, carcinoma, and sarcoma cell lines, but not with cell lines derived from normal skin or lungs. Affinity-isolated antigen was subjected to polyacrylamide gel electrophoresis and stained with Coomassie blue. Under dissociating conditions, a band in the 31,000 to 42,700 Da range was identified that was shown to be reactive with MAb JSI in ELISA. The antigenic determinant defined by MAb JSI appears to be a protein, with expression on a number of malignant tissues.

Animals↗
PubMed1990

Hanganutziu-Deicher antigen as a possible target for immunotherapy of melanoma.

Hanganutziu-Deicher (H-D) antigen is classified as a heterophile antigen and chemically defined as a glycoconjugate which contains N-glycolylneuraminic acid. H-D antigens are absent from normal human tissues, but can be expressed on a variety of human malignant cells, including melanoma. Natural anti-H-D antibodies have been detected in man with and without malignancies, but in this study when the level of antibody was compared between healthy adults and patients with melanoma, elevated anti-H-D antibody levels were found more frequently in melanoma patients for both IgM (p = 0.0001) and IgG (p = 0.0001). The present study was designed to evaluate the significance of the H-D antigen-antibody system in melanoma suppression. Sera from melanoma patients containing anti-H-D antibody reacted strongly to H-D antigen expressed on melanoma by means of flow cytometry. In a complement-dependent cytotoxicity assay this antibody killed melanoma cells in vitro. In vivo significance of the antibody was assessed by evaluating the relationship between the antibody levels and the clinical course in patients with stage II melanoma. Antibody levels were measured by enzyme-linked immunosorbent assay using a H-D glycoprotein antigen isolated from bovine erythrocytes. A significantly higher level of IgG (p = 0.0640) and IgM (p = 0.0644) anti-H-D antibody was demonstrated in those patients who were free of disease more than 5 years after surgery than in those who relapsed within 2 years. This study provides a rational basis for immunotherapy targeting H-D antigen in human melanoma.

Antibodies↗
PubMed1990

Differential effects of granulocyte-macrophage colony-stimulating factor and macrophage colony-stimulating factor on tumor necrosis factor and interleukin-1 production in human monocytes.

Human peripheral blood monocytes (PBM) cultured in the presence of 100-5,000 u/ml granulocyte-macrophage colony-stimulating factor (GM-CSF) for 24 hr secreted small quantities of tumor necrosis factor (TNF), but not interleukin-1 (IL-1). The activation of PBM to produce TNF was weak and could be blocked by polyclonal anti-GM-CSF anti-serum. Neither LPS nor IL-2 were synergistic with GM-CSF in the production of TNF or IL-1. IFN gamma alone did not induce either cytokine, but in the presence of GM-CSF it caused a synergistic (100-fold) increase in TNF but not IL-1 production. Macrophage colony-stimulating factor (M-CSF) alone or in combination with LPS, IFN gamma or IL-2 did not stimulate PBM to produce TNF or IL-1 in 24 hr culture.

Granulocyte-Macrophage Colony-Stimulating Factor↗
PubMed1989

An epitope common to gangliosides O-acetyl-GD3 and GD3 recognized by antibodies in melanoma patients after active specific immunotherapy.

GD3 is a major ganglioside of human melanoma and was shown to be an effective target for passive immunotherapy with murine monoclonal antibodies. It was noted earlier that GD3 neither purified nor in melanoma cell vaccine (MCV), could elicit an antibody response in melanoma patients. In this study, we demonstrate that melanoma patients who received MCV had autoantibodies against a derivative of GD3, O-acetylated GD3 (O-AcGD3), a minor ganglioside expressed on human melanoma cells, and that the antibodies cross-reacted with GD3. Thin layer chromatographic immunostaining revealed that all of the sera containing antibodies against O-AcGD3 also reacted to GD3. None of the other sera responded only to GD3, although the MCV contained 7- to 12-fold higher GD3 than O-AcGD3. Furthermore, the antibody activity was completely abolished by absorption with animal erythrocytes expressing either O-acetyl disialogangliosides or GD3, indicating that the antibodies recognize an epitope commonly shared by GD3 and O-AcGD3. The antibodies bound only to the sialyloligosaccharide moiety but not to the ceramide portion of GD3 after endoglycosylceramidase treatment. The antibodies failed to bind to GD3 after neuraminidase treatment. These results indicate that the sialyloligosaccharides of the gangliosides are important components of the epitope. Periodate oxidation abolished reactivity of the antibodies to GD3 but not that to O-AcGD3, revealing that the glycerol side chain of the sialic acids in both GD3s was an important structure of the epitope. The binding of the antibodies to melanoma cell surface gangliosides was confirmed by an absorption with a GD3- and O-AcGD3-positive melanoma cell line. These results in the light of previous reports on the inability of GD3 to elicit immune response in humans suggest that anti-GD3 antibodies found in the melanoma patients were induced by immunization with O-AcGD3 and O-AcGD3 present in the MCV would serve as an antigen source for GD3-targeted active specific immunotherapy of melanoma.

Antibodies↗
PubMed1989

Genetic analysis of the human tumor necrosis factor alpha/cachectin promoter region in a macrophage cell line.

The 615-bp 5' flanking region of the human TNF-alpha/cachectin gene was isolated and ligated to the luciferase reporter gene. In addition, a series of truncated promoter constructs was generated by exonuclease III digestion. The promoter activity of these constructs was studied in a transient transfection system using the TNF-alpha-producing U937 cell line. Full-length and truncated TNF promoter constructions extending from -615 to -95 bp relative to the transcription start site (TSS) could be induced by phorbol esters. A construct truncated to within 36 bp of the TSS (and within 11 bp of the TATAA box) was inactive. Therefore, the phorbol ester responsive is localized in the TNF/cachectin promoter to a relatively short region proximal to the TATAA box.

Base Sequence↗
PubMed1989

IL-4 down-regulates IL-1 and TNF gene expression in human monocytes.

The effects of IL-4 on IL-1 and TNF gene expression in human peripheral monocytes (PBM) were examined. Highly purified PBM cultured for 24 to 96 h in the presence of IL-4 produce neither IL-1 nor TNF protein. RNA hybridization studies demonstrated that IL-4 does not induce transcription of IL-1 or TNF. Preincubation of PBM with IL-4 did, however, inhibit LPS-induced IL-1 and TNF production in a dose- and time-related fashion. Maximal inhibition occurred after 96 h of incubation with 200 U/ml IL-4. RNA hybridization studies demonstrated that IL-4 suppressed the expression of IL-1 alpha, IL-1 beta, and TNF mRNA. These results suggest IL-4 modulates monocyte production of TNF and IL-1 by down-regulation of gene expression. This unique property of IL-4 may be important in the regulation of the immune response.

Gene Expression Regulation↗