Biomedical subjects

M G Wing

Publications and source records attributed to M G Wing.

26 records · Page 2Linked to original sources

PubMed1992

Oligodendrocytes lack glycolipid anchored proteins which protect them against complement lysis. Restoration of resistance to lysis by incorporation of CD59.

Rat oligodendrocytes, which activate the classical pathway of complement in the absence of antibody, are highly sensitive in a reactive lysis assay using human C5b6 and EDTA serum. Oligodendrocytes may be relatively deficient in glycolipid-linked complement regulatory protein(s), since digestion with phosphatidylinositol-specific phospholipase C (PI-PLC) failed to increase their sensitivity to serum, whereas complement-insensitive astrocytes, when treated with PI-PLC, became strikingly sensitive. To test the hypothesis that oligodendrocytes lack terminal complement regulatory molecule(s), human erythrocyte CD59, a recently described complement regulatory protein, was purified to homogeneity. The biological activity of the preparation was confirmed by reincorporating the protein into guinea-pig erythrocytes through its glycolipid anchor, which resulted in dose-dependent protection against human C5b6 and EDTA serum. Incorporation of 10(5) molecules of human CD59 into rat oligodendrocytes resulted in good protection against homologous human complement (76%), and significant protection against rat complement homologous to the cell (36%). Protection could be reversed using an antibody to CD59.

Animals↗

PubMed1992

The targeting of T-helper cells and tumourcidal macrophages to a B-cell lymphoma using a PPD-monoclonal antibody heteroconjugate.

This paper describes a T-cell targeting strategy based on the use of an antigen-monoclonal antibody heteroconjugate. A rat anti-idiotypic monoclonal antibody specific for a murine B-cell lymphoma was conjugated to the purified protein derivative (PPD) of tuberculin. This construct selectively delivered up to 4.5 x 10(4) molecules of PPD onto each tumour cell. Targeted PPD was internalized for endosomal processing and was presented in association with the I-A class II restriction element to PPD-reactive T-helper (Th) cells. Activated Th cells were demonstrated to proliferate and secrete significant levels of tumour necrosis factor (TNF). Such lymphokine secretion was observed at a PPD concentration as low as 1 ng/ml. Despite the secretion of TNF, the B-cell lymphoma was found to be resistant to autonomous Th-mediated cytotoxicity. Targeted Th cells did, however, activate tumourcidal macrophages that subsequently mediated significant tumour cytostasis. Based on this observation, it is proposed that the targeting system described may be exploited as the basis for a future immunotherapeutic strategy.

Animals↗

PubMed1990

An improved method for the detection of cell surface antigens in samples of low viability using flow cytometry.

A high non-specific background fluorescence signal was observed when cell surface antigen analysis was carried out using flow cytometry on a cell sample which contained a high proportion of dead and dying cells. To overcome this problem it was necessary to analyse the cells in three stages. First the intact cells were identified by their forward (FWD) and 90 degree scatter profile. These cells were gated-on, then analysed on the basis of their FWD scatter and propidium iodide (PI) signal, allowing the dead PI positive cells to be gated out. The PI negative cells were then displayed using their 90 degree scatter and fluorescence signals following staining with the irrelevant antibody control. This revealed a population of dead cells, which despite being PI negative, were non-specifically binding antibody molecules. Such multiparameter analysis permitted the successful analysis of cell surface antigens in preparations of low viability by gating out the high background fluorescence associated with dead PI positive and negative cells.

Animals↗

PubMed1990

A novel strategy for targeting CD4+ PPD-reactive T cells against tumour cells using PPD monoclonal antibody heteroconjugates.

We have constructed PPD monoclonal antibody heteroconjugates specific for a tumour-associated antigen of C57BL/6 melanomas or for human complement component C3d fixed de novo to murine fibrosarcoma cells (MC6A). The ability of our heteroconjugates to target CD4+ PPD-reactive T cells against the appropriate tumour targets was then determined in vitro. Heteroconjugate-treated B16-F10 and MC6A tumour targets were both able to present PPD to the specific T cells, resulting in activation and concomitant lymphokine secretion. Secreted lymphokines were then demonstrated to cause significant tumour cytolysis and cytostasis in vitro. Preliminary experiments in vivo suggest that this targeting system may provide the basis for a future immunotherapeutic strategy.

Animals↗

PubMed1990

Cytostasis of different tumours by a murine PPD-reactive CD4+ T lymphocyte clone is mediated by interferon-gamma and tumour necrosis factor alone or synergistically.

We have shown that a murine CD4+ PPD-reactive T lymphocyte clone was weakly cytotoxic towards the syngeneic tumour B16 melanoma and MC6A fibrosarcoma which had been coated with PPD using a monoclonal antibody-PPD heteroconjugate. Cell-free supernatants produced by PPD-stimulated T lymphocyte clones were however highly cytostatic for the two tumour targets when assayed over 48-72 h. In this study we have demonstrated good titres of tumour necrosis factor (TNF) and interferon-gamma (IFN-gamma) in the supernatants, which accounted for their observed cytostatic activity on the tumour targets. The high level of cytostasis seen with the B16 melanoma using the supernatants could be attributed to their sensitivity to the cytostatic activity of IFN-gamma; the lower levels of cytostasis seen with the IFN-gamma-resistant MC6A target was the result of IFN-gamma increasing the sensitivity of this target to TNF. Antibodies to IFN-gamma were able to neutralize the majority of the cytostatic activity of the supernatants on both targets, consistent with the role demonstrated for this lymphokine.

Animals↗

PubMed1988

Comparison of excision versus cryosurgery of an HSV-2-induced fibrosarcoma. I. Survival, extent of metastatic disease and host immunocompetence following surgery.

Cryosurgery and excision were used to treat primary tumours of HSV-2-transformed hamster tumour sublines, and post-operative survival and the extent of metastatic disease were compared in the two groups. An inferior prognosis was observed following cryosurgery although the extent of metastatic disease was similar in both groups. Using this model it would appear that cryosurgery enhances the development of micrometastases rather than affecting the number of cells shed from the primary tumour during surgery. To investigate the underlying causes of the decrease in survival following cryosurgery, in vitro assays were used to monitor host immunocompetence following surgery. The results showed that whilst natural killer cell cytotoxicity was only marginally depressed, mitogen responsiveness and lymphocyte participation in a mixed lymphocyte reaction were severely reduced 3-7 days post-cryosurgery. In parallel with immunosuppression, extensive cell proliferation in the spleen of cryosurgically treated tumour-bearing animals was observed. Histological examination of the spleen demonstrated the presence of large numbers of transformed cells which correlated with the loss of mitogen responsiveness and the ability to participate in a mixed lymphocyte reaction. Further studies (manuscript submitted for publication) have demonstrated that spleen cells from animals whose tumour is treated by cryosurgery are capable of suppressing immunocompetence in vitro, implying they have a role in the uncontrolled growth of micrometastases in vivo.

Animals↗

PubMed1988

Characterisation of suppressor cells generated following cryosurgery of an HSV-2-induced fibrosarcoma.

Cryosurgery of a primary HSV-2-induced hamster fibrosarcoma resulted in the generation of a population of suppressor cells. These cells were detectable in the spleen 1-10 days post-cryosurgery by their ability to suppress the proliferation of immunocompetent splenic T-lymphocytes following exposure to concanavalin A (Con A). The spleens of tumour-bearing (t.b.) animals which received cryosurgery 3 days previously displayed gross splenomegaly due to the generation of large numbers of highly proliferative erythroblasts. The erythroblast cells were unlikely to be the source of suppression since time course studies have demonstrated the presence of suppressor cells before and after their appearance in the spleen. The erythroblasts therefore probably reflected a response by the host to regenerate the erythrocytes lost during surgery and their presence was independent of the appearance of suppressor cells. Characterisation of the suppressor cell has revealed it to be non-adherent and esterase negative making it unlikely to be of macrophage (MO) lineage. This was confirmed by the ability of splenic MOs from day 3 t.b. cryosurgery-treated animals to completely restore Con A-dependent T-lymphocyte proliferation following MO depletion. As nylonwool column-eluted cells are able to suppress Con A-dependent T-lymphocyte proliferation, it seemed unlikely that B-lymphocytes play a role in cryosurgery-induced immunosuppression. These findings suggest that cryosurgery of a t.b. animal results in the generation of a population of T-lymphocytes capable of suppressing Con A-dependent T-lymphocyte proliferation, and infers that these cells contribute to the inferior prognosis following cryosurgery as compared to excision of a metastatic tumour.

Animals↗

PubMedDate not supplied

In vitro preclinical lead optimisation technologies (PLOTs) in pharmaceutical development.

The explosion of genuine high throughput technologies has allowed large compound libraries to be screened with ever-increasing biological specificity, exacerbating the problem of lead candidate selection for subsequent drug development. To avoid creating a bottleneck, compounds identified from the high throughput screens undergo lead optimisation by employing medium-throughput screen which permits ranking in terms of their basic absorption, distribution, metabolism, excretion (ADME) and toxicological properties. The historical role of the CRO in the drug discovery/development continuum has been to perform efficacy and toxicology studies, simply to support the regulatory submission of lead candidates. This situation is, however, changing with the development of preclinical lead optimisation technologies facilitating the selection of leading candidates, thereby bridging the gap between high throughput efficacy screens and conventional safety assessment programmes.

Drug Approval↗