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Biomedical subjects

R A Robins

Publications and source records attributed to R A Robins.

At least 37 records · Page 2Linked to original sources
PubMed2000

A neoadjuvant clinical trial in colorectal cancer patients of the human anti-idiotypic antibody 105AD7, which mimics CD55.

Thirty-five patients received 105AD7 human anti-idiotype vaccination prior to surgery for colorectal carcinoma. Patients were immunized before and also received one to two immunizations after surgical resection of their colorectal cancer. The vaccine was well tolerated with no associated toxicity. Lymphocytic infiltration within the resected tumors was quantified by immunohistochemistry and image analysis. Enhanced infiltration of helper T cells (CD4) and natural killer (NK) cells (CD56) were observed in the tumors from immunized patients when compared with tumors from stage, grade, site, age, and sex matched unimmunized patients. NK activity was increased in the blood, peaking 7-10 days post immunization and then dropping rapidly and correlating with NK extravasation within the tumor. Comparison of the amino acid sequences of 105AD7 anti-idiotype and the antigen it mimics, CD55, has predicted that patients with HLA-DR1, HLA-DR3, and HLA-DR7 haplotypes should show helper T cell responses following 105AD7 vaccination. Eighty-three percent of patients expressing these haplotypes responded to 105AD7, whereas 88% of patients who failed to express these haplotypes were nonresponders. With a median follow-up of 4 years (range, 2.5-6 years) 65% of patients remained disease free. This trial shows that 105AD7 stimulates antitumor inflammatory responses allowing extravasation within tumor deposits of both helper T cells and NK cells. This represents a way of evaluating immune responses in patients both within the blood and at the tumor site. The study confirms that immunization with a human anti-idiotypic antibody results in immune responses in 83% of patients with a permissive haplotype.

Antibodies, Anti-Idiotypic↗
PubMed1999

Co-cultivation of CD4+ and CD8+ human T-cells leads to the appearance of CD4 cells expressing CD8 through de novo synthesis of the CD8 alpha-subunit.

The appearance of a population of dual-staining CD4+CD8+ cells in human T-lymphocyte cultures has been reported by various authors, including our own observation that they are always seen in simple phytohaemagglutinin-stimulated cultures from several different donors. The purpose of the present study was to investigate factors involved in the dual-staining (DS) phenotype, and to clarify some apparent inconsistencies between published observations. Our findings can be summarised as follows. 1. A population of DS CD4+CD8+ cells always appears in PHA-stimulated T-cell cultures if they contain both CD4 and CD8 subsets. The incidence of DS cells is related to PHA concentration, but other factors are involved since DS cells are not seen in PHA-stimulated cultures of purified CD4+ or CD8+ cells. Stimulation with PHA is not a prerequisite since very similar results are seen with ConA. 2. Direct physical contact between CD4+ and CD8+ cells is required for the appearance of the DS phenotype; soluble factors alone, including IL-4, appear nor to be responsible. 3. The DS phenotype in these conditions is always CD4+ cells weakly expressing CD8 and is a consequence of de novo synthesis of the CD8alpha molecule by the CD4+ cells.

Apoptosis↗
PubMed1999

Increased activation of lymphocytes infiltrating primary colorectal cancers following immunisation with the anti-idiotypic monoclonal antibody 105AD7.

BACKGROUND: The anti-idiotypic monoclonal antibody 105AD7 mimics the tumour associated antigen 791Tgp72, expressed on 70-80% of colorectal cancers. Phase I studies have shown that the vaccine is non-toxic, and a number of patients have been immunised prior to resection of their primary tumours. AIMS: To assess lymphocyte activation at the tumour site by measuring expression of the alpha subunit of the interleukin 2 receptor (CD25). METHODS: Nineteen patients with primary colorectal cancer were immunised with varying doses of 105AD7 prior to resection of their primary tumours. Samples of normal bowel and tumour edge/centre from 16 patients were available for immunohistochemical staining with a monoclonal antibody against CD25. Samples from a matched control group were also stained. Fresh tumours from 14 immunised patients and 31 unimmunised control patients were disaggregated, and the lymphocytes obtained labelled for CD25. Samples were analysed blindly by flow cytometry. RESULTS: Median infiltration of lymphocytes expressing CD25, measured immunohistochemically, was higher in trial patients, as was the ratio of tumour to normal bowel infiltration. Flow cytometric analysis of fresh tumour from immunised patients showed a significantly higher percentage of lymphocytes expressing CD25 tumour infiltrating lymphocytes than their matched and unmatched controls. DISCUSSION: The alpha subunit of the interleukin 2 receptor is increased on tumour infiltrating lymphocytes, in patients immunised with the colorectal cancer vaccine 105AD7. This suggests a population of activated lymphocytes capable of targeting 791Tgp72 expressing tumour cells, such as circulating micrometastases. 105AD7 may have a role as adjuvant therapy in early stage disease.

Aged↗
PubMed1997

Peripheral blood lymphocyte binding to a soluble FITC-fibronectin conjugate.

Fibronectin receptors are critical to lymphocyte-mediated inflammation, and binding between immobilised plasma fibronectin and peripheral blood lymphocytes (pbl) is enhanced in patients with inflammatory diseases. Such enhancement could be due to an overall increase in receptor capacity and/or be a feature of a subset of cells. It could also depend on the initial binding and/or on events that follow receptor occupancy (e.g., cell spreading). To address these questions, we used fluorescein isothiocyanate-conjugated soluble fibronectin (FITC-FN) in a flow cytometric assay of lymphocyte-fibronectin binding. EDTA-inhibitable adhesiveness to FITC-FN is enhanced in lymphocytes cultured with Interleukin 2 and in fresh peripheral blood lymphocytes (pbl) from untreated patients with acute exacerbations of chronic inflammatory diseases. The enhanced binding was noted on both CD4+ and CD8+ subsets, and decreased with anti-inflammatory therapy.

CD4-Positive T-Lymphocytes↗
PubMed1997

Interleukin-8 and inducible nitric oxide synthase mRNA levels in inflammatory bowel disease at first presentation.

Interleukin-8 (IL-8) and nitric oxide (NO) may be important mediators in the pathogenesis of chronic idiopathic inflammatory bowel disease (CIIBD), but their roles in disease activity in ulcerative colitis (UC) and Crohn's disease (CD) are uncertain. The aim of this study was to measure mRNA for IL-8 and inducible NO synthase (iNOS) in small mucosal biopsies from untreated patients at first presentation and to relate these measurements to the histological levels of polymorph infiltration graded on a ten-point scale. For this purpose, a sensitive enzyme-linked oligonucleotide chemiluminescent assay (ELOCA) was developed to quantitate reverse transcription-polymerase chain reaction (RT-PCR) products amplified from RNA from paired biopsy samples. The levels of IL-8 and iNOS mRNAs were calculated as ratios of the RT-PCR products to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) RT-PCR product. In UC patients, median values of IL-8/GAPDH and iNOS/GAPDH were significantly elevated compared with controls and CD. However, in both UC and CD, the IL-8/GAPDH and iNOS/GAPDH ratios correlated significantly with polymorph infiltration. ELOCA enabled quantitation of multiple mRNAs in small mucosal biopsies from untreated patients with CIIBD and supported a role for IL-8 and iNOS in acute inflammation in both UC and CD.

Adult↗
PubMed1997

Low doses of 105AD7 cancer vaccine preferentially stimulate anti-tumor T-cell immunity.

Clinical studies with the human anti-idiotypic antibody 105AD7 have clearly shown that 791Tgp72 is a good target antigen for cell-mediated immunity. No antibody-related toxicity was observed in any of the 135 patients entered into phase I/II clinical trials of 105AD7, whereas both helper and cytotoxic T-cell responses were induced. The helper responses were exemplified by induction of interleukin-2 (IL-2), antigen-specific blastogenesis, and enhanced natural killer (NK) activity. Anti-tumor cytotoxicity was measured directly and was supported by activation of circulating CD8 cells. In this study, it is shown that a 100-microgram injection of 105AD7 was more effective than the 200-microgram dose. Enhanced IL-2 production was observed following 15/19 injections of 100 micrograms of 105AD7 whereas only 4/11 injections of 200 micrograms of 105AD7 induced responses (p < 0.02). Similarly, time to progression was significantly (p < 0.05) slower (median 6 m) in patients injected with 100 micrograms than patients receiving the higher dose, suggesting that 100 micrograms or lower may be the optimal dose. The standard dose for hepatitis vaccination is 10 micrograms. In vitro blastogenesis assays on naive donors have shown that a dose of 105AD7, which is either too high or too low, fails to activate T cells. The optimal dose in vitro is 10 ng.

Adult↗
PubMed1997

Correlation between CD29 density on CD8+ lymphocytes and serum IgG in systemic lupus erythematosus.

The purpose of this paper is to establish whether there is increased lymphocyte adhesion molecule density in systemic lupus erythematosus (SLE), which could alter the migration pathways and activation thresholds of lymphocytes and thus contribute to the pathogenesis of the disease. We analysed the CD11a, CD29 and CD2 bound antibody molecule (bam) density on the CD4+ and CD8+ CAMhigh (primed) lymphocytes of 28 SLE patients (8 active and 20 inactive by BILAG), using reproducible flow cytometric measurements, standardized with fluorescent beads and antibodies of known fluorescein: protein ratios. In a second patient cohort (17 patients), we investigated whether CD29 density on CD8+ cells correlated with measures of humoral (serum IgG) or cellular (urine neopterin) activation. In the first cohort, 36% of patients had elevated CD29 (beta 1 integrin) density on CD8+ cells. In the second cohort, CD29 density on CD8+ cells was found to be closely associated with total plasma IgG (r = 0.71, P = 0.001), but not with urine neopterin, disease activity (BILAG) or drug treatment. We conclude that CD29 on CD8+ cells is associated with B cell activation in SLE.

Antigens, CD↗
PubMed1996

Peripheral blood lymphocyte adhesion molecule deployment in the immune response.

Using precise and reproducible flow cytometric measurements, the surface densities of the cell adhesion molecules (CAMs) CD29, CD2 and CD11a were studied on the CAMhigh (primed) subsets of peripheral blood CD4+ and CD8+ lymphocytes in 56 healthy subjects; 18 patients with acute bacterial infections, 19 with acute viral infections and 18 with chronic inflammatory conditions. By Mann-Whitney analysis, with significant P values adjusted for multiple comparisons to less than 0.0007, patients with viral infections were found to have increased CD11a on CAMhigh cells (an increase in median values of 13.7% for CD4+ lymphocytes and 15.8% for CD8+ lymphocytes); patients with chronic conditions have increased CD29 on CD8+CAMhigh lymphocytes (an increase in the median of 19%); and patients with bacterial infections have increased CD2 on CD8+CAMhigh cells (an increase in the median of 8%). There were marked individual increases in CD29 density: eight (15%) patients had CD29 gender-adjusted density on CD4+ cells greater than the control mean + 3 standard deviations (SD). CD29 densities on CD8+ cells were elevated to greater than control mean + 3 SD in 12 (22%) patients. By multiple regression analysis CD11a density on CD8+ and CD4+ cells and CD2 density on CD4+ cells were found to be associated with HLA-DR expression, but not with CD25 expression. Using standardized intercepts the authors demonstrated that there are very few circulating CDllahighCD25+ cells, suggesting that these are rapidly extravasated. This study demonstrates that in disease, lymphocyte adhesion molecules are not deployed in concert and there are characteristic deployment patterns for different types of immune response.

Acute Disease↗
PubMed1996

Peripheral blood based T cell-containing and T cell-depleted culture systems for human IgE synthesis: the role of T cells.

BACKGROUND: Comparable T cell-containing and T cell-depleted culture systems for human IgE synthesis are currently not available. OBJECTIVE: This has prompted us to develop peripheral blood mononuclear cell (PBMC) based culture systems for human IgE synthesis in the presence and absence of T cells. METHODS: In this paper we describe simplified conditions for in vitro synthesis of high levels of IgE by human peripheral blood B cells, both in T cell-containing cultures and in anti-CD40 stimulated T cell-depleted cultures. RESULTS: T cell-depleted cultures released approximately 20 times more IgE [range 410-2220 ng/mliter (mean 1270 ng/mliter); based on six experiments using cells from three donors] than did T cell-containing cultures [range 23-105 ng/mliter (mean 58 ng/mliter); based on 15 experiments using cells from three donors]. Reconstitution experiments were performed to investigate the role of T cells on IgE synthesis. Adding T cells back to the anti-CD40 stimulated T cell-depleted cultures resulted in a dose-dependent inhibition of IgE production. In the absence of anti-CD40 low numbers of T cells stimulated, while high numbers suppressed, IgE production: the optimal ratio of T cells to non-T cells for maximal IgE production was found to be 1:1. At this ratio, irradiated (non-replicating) T cells supported a much greater IgE synthesis than did non-irradiated T cells. CONCLUSION: The development of these systems provides directly comparable T cell-containing and T cell-depleted cultures for human IgE synthesis from peripheral blood, allowing further study of the role of T cells in IgE regulation. These systems will also be of use for determining whether potential modulators of IgE synthesis act on the T cells or on other cell types.

Cells, Cultured↗
PubMed1995

Induction of cellular immune responses by a murine monoclonal anti-idiotypic antibody recognizing the 791Tgp72 antigen expressed on colorectal, gastric and ovarian human tumours.

There is accumulating evidence that cellular rather than antibody responses are more effective for tumour rejection. It is therefore important to screen anti-idiotypic (anti-id) antibodies for their ability to stimulate anti-tumour T-cell responses. The human anti-id monoclonal antibody (MAb) 105AD7 stimulated both delayed-type hypersensitivity (DTH) responses in animals and antigen-specific blastogenesis and IL-2 induction in advanced cancer patients. It may not be necessary to use human anti-id antibodies as murine anti-id antibodies, which elicit DTH responses against immunodominant human T-cell epitopes and may be just as useful in the clinic. We have therefore produced a murine anti-id antibody to the same MAb as was used to generate the human anti-id antibody and screened it for its ability to generate cellular anti-tumour immune responses. Low-dose immunization with the murine anti-id MAb NCRC60, which recognises the paratope of the anti-791Tgp72 MAb 791T/36, induced DTH responses to 791Tgp72-expressing tumour cells but not to antigen-negative cells. DTH responses with no detectable antibody responses were induced with 5 micrograms of anti-id NCRC60 without adjuvant. Addition of either complete Freund's adjuvant or Quil A did not enhance DTH responses. However, when the anti-id NCRC60 was linked to KLH and injected in the presence of Freund's adjuvant anti-anti-id antibodies and anti-791Tgp72 antibodies were induced. NCRC60 anti-id was also capable in vitro of priming human T cells from cancer patients to proliferate in response to secondary stimulation with 791Tgp72-expressing tumour cells, suggesting that it may have therapeutic potential in cancer patients.

Animals↗
PubMed1995

A flow cytometric method to measure shape change of human neutrophils.

1. We report a flow cytometric method in which changes in forward angle light scatter are shown to correlate with microscopically evaluated shape change in stimulated human neutrophils. Neutrophil movement and chemotaxis is conventionally measured using Boyden chambers, which is a laborious and exacting technique. Microscopic scoring of neutrophil shape change has been shown to correlate well with Boyden chamber measurements, and although less laborious, still requires manual counting. 2. We now show that measurement of forward angle light scatter in a benchtop flow cytometer correlates closely with microscopic evaluation of neutrophil shape change in dose-response stimulation experiments with leukotriene B4, N-formyl-methionine-leucine-phenylalanine or interleukin-8. The relationship between shape change and increased forward angle light scatter was confirmed using the fluorescence-activated cell sorter to separate partially stimulated neutrophils, followed by reanalysis by flow cytometry and microscopic examination. 3. This flow cytometric method provides a convenient, rapid and objective measure of neutrophil responses to external stimuli.

Cell Size↗
PubMed1994

Enhanced cell-mediated tumor killing in patients immunized with human monoclonal antiidiotypic antibody 105AD7.

A human antiidiotypic monoclonal antibody (105AD7) has been shown to induce antitumor cellular responses in animals and appears to prolong survival in patients with metastatic colorectal cancer without associated toxicity. Proliferative leukocyte responses to the targeted tumor antigen gp72 were observed in these patients and plasma interleukin 2 levels were increased following immunization. Autologous tumor tissue was not available in these patients, so antitumor cytotoxicity could not be measured. This issue has now been addressed in an adjuvant clinical study in primary rectal cancer patients. Six patients with rectal cancer were immunized preoperatively with 105AD7. Peripheral blood lymphocytes taken prior to immunization were tested against tumor cells extracted from biopsies also obtained prior to immunization or from natural killer (NK)-sensitive target cells. Cryopreserved lymphocytes taken before and after tumor immunization, fresh peripheral blood lymphocytes taken immediately prior to surgery, and lymphocytes from tumor-draining lymph nodes were tested against autologous cells from the resected specimen or NK-sensitive target cells. Significant killing of autologous tumor cells, which was not due to NK activity, was seen with cryopreserved lymphocytes or lymph node cells of three patients at 1-2 weeks postimmunization with 105AD7 but not on pretreatment biopsies. Enhanced NK activity was seen 2-3 weeks postimmunization in 3 of 6 patients. These results indicate that 105AD7 human monoclonal antibody immunization enhances cytotoxicity in rectal cancer patients by specific and nonspecific effector mechanisms.

Aged↗
PubMed1994

Clinical outcome of colorectal cancer patients treated with human monoclonal anti-idiotypic antibody.

A human monoclonal anti-idiotypic antibody (105AD7) has been developed which mimics a colorectal-tumour-associated antigen and induces cellular anti-colorectal tumour immune responses in animals. Thirteen patients with advanced colorectal cancer were immunized with 105AD7 and their survival was compared with that of a contemporary group of unimmunized patients with similar disease status. No toxicity related to anti-idiotype immunization was seen. Cellular responses to anti-idiotypic immunization were indicated by lymphocyte proliferation to gp72-positive tumour cells, and production of interleukin-2; anti-tumour antibodies were not detected. Median survival following diagnosis of advanced disease of immunized patients was 12 months, compared with 4 months in unimmunized patients. The improved survival of immunized patients in this study without associated toxicity suggests that 105AD7 immunization may have considerable potential for immunotherapy of colorectal cancer.

Aged↗
PubMed1994

A phase II study of effect of addition of trichosanthin to zidovudine in patients with HIV disease and failing antiretroviral agents.

Patients infected with HIV, including those with AIDS-related complex and AIDS, and failing treatment with antiretroviral agents such as zidovudine, have been evaluated following addition of trichosanthin to the antiretroviral agent regimen. This ribosomal inhibitory protein is specifically cytotoxic for HIV-infected macrophages and lymphocytes. Ninety-three patients were treated with trichosanthin, using a schedule of weekly, then monthly, intravenous injections of 1.2 mg of drug in combination with antiretroviral agents, usually zidovudine. Side effects included myalgias, fevers, mild elevation in liver function tests, and mild-moderate anaphylactic reactions, which respond well to therapy with steroids and/or benedryl. Reversible mental status changes were noted in two patients, both receiving concomitant therapy with ddI. Clinical responses to trichosanthin treatment were monitored primarily by changes in laboratory parameters, particularly levels of CD4+ T lymphocytes. In the total population evaluated for efficacy (85 patients) there was a significant increase in CD4+ cell levels after initiation of trichosanthin therapy. A second analysis performed on 72 patients measured the rate of change of CD4+ cells during therapy, using an "area under the curve" analysis. During therapy there was a median increase of 1.2 cells/mm3/month. In patients in the top 25th percentile, this increase was greater than 8.4 cells/mm3/month. In 59 of the 72 patients, responses could also be monitored by comparing the rate of loss of CD4+ cell levels on antiretroviral agents (zidovudine or ddI) alone, during the year prior to initiation of trichosanthin, to the rate of change when trichosanthin was added to the treatment regimen. During the period before trichosanthin treatment (311 +/- 11.7 days) the median loss of CD4+ cells was 6.91 cells/mm3/month. Addition of trichosanthin to the treatment regimen resulted in a median gain of 1.1 CD4+ cells/mm3/month.

Adolescent↗