Vinblastine (VLB) and VIncristine (VCR) are important antitumor agents. Although binding of these alkaloids to spindle proteins (tubulin), resulting in mitotic arrest, has been suggested as the basis for their antimitotic action, the mechanism for their cytotoxic action is not known. We have shown that in drug sensitive lymphoblasts incubated with low VLB concentrations, extensive cytolysis occurs suggesting a cell surface related drug effect. The presence of cel surface tubulin in these cells may account for their drug sensitivity. Similarly, in cells made resistant in vitro to VLB, new glycoproteins appear on the cell surface. The direct relation of these cell surface proteins to the mechanism of drug transport and resistance in a tumor cel population has not been elucidated. Nor is any information available on the relation between these markers and the cell cycle phase related cytotoxicity of VLB/VCR. We now propose to extract, purify and concentrate tubulin and surface marker proteins (100 and 160 K M.W) from drug sensitive and resistant cells. Monoclonal antibodies to these markers will be labeled and used for correlation between markers, drug transport and resistance in individual cells from a population. We will analyze relation between the appearance and content of the markers on individual cells and the cell cycle phase related cytotoxicity of VLB/VCR. Centrifugal elutriation, density gradient separation and fluorescence activated cell sorting will be used to quantitate amount of bound antibodies and to collect selectively enriched populations. Radiolabelled drugs will be used to study drug transport. Multiparameter flow analysis will be used to correlate cell cycle stage and cell surface labelling. Correlation with effects on cell proliferation will be studied in soft agar assays and nude mice xenografts.
| DiCicco, L M; Lazarus, H; Krishan, A (1989) Class I MHC molecules and doxorubicin resistant P388 murine leukemic cells. Immunopharmacol Immunotoxicol 11:583-92 |
