RADIOSENSITIZATION MEDIATED BY A TRANSFECTED ANTI-ERBB-2 SINGLE-CHAINANTIBODY IN-VITRO AND IN-VIVO

Purpose: The erbB-2 receptor is overexpressed in several human cancers , including ovarian, prostate, and breast. We have developed plasmid a nd adenoviral vectors expressing an anti-erbB-2 single chain antibody (sFv), directed to the endoplasmic reticulum (ER) of target cells, tha t is cytotoxic to tumor cells overexpressing erbB-2 through induction of apoptosis. The anti-erbB3 sFv also sensitizes erbB-2 overexpressing cells to the cytotoxic effects of cisplatin. On this basis, we hypoth esized that human ovarian cancer cells expressing anti-erbB-2 sFv with downregulated erbB-2 product, p185(erbB-2), also would be sensitized to ionizing radiation. Therefore, we designed experiments to test the ability of the anti-erbB-2 sFv tea radiosensitize human ovarian cancer cells in vitro and in vivo. Methods and Materials: To test our hypoth esis, we established subcutaneous (s.c.) tumors in the Banks of nude m ice with SKOV3.ip1 human ovarian cancer cells and SKOV3 cells stably e xpressing the ER directed anti-erbB-2 sFv (SKOV3/pGT21). The tumors we re treated with 10 Gy Co-60, or received no radiation. We then determi ned the regression rate, delay in regrowth, and time to tumor doubling of the tumors treated with radiation in the transfected group and con trols. In addition, SKOV3.ip1 and SKOV3/pGT21 tumors were dissected fr om the irradiated animals and assayed for differences in p185(erbB-2) expression at 12 weeks after irradiation by immunohistochemistry. Furt her, in vitro clonogenic survival assays were performed on the parenta l SKOV3.ip1 and SKOV3/pGT21 cell lines.Results: A statistical analysis of the combined data was done for two in vivo experiments. The analys is of the combined data showed that animals with irradiated tumor SKOV 3/pGT21 had a significantly higher regression rate (p = 0.0055), longe r delay in regrowth (p = 0.0001) and time to tumor doubling (p = 0.000 4), than those animals with tumor SKOV3.ip1 that received radiation. W e observed a similar significant effect for the same parameters in the unirradiated tumor SKOV3/pGT21 compared to unirradiated tumor SKOV3.i p1. Immunohistochemical analysis of the SKOV3/pGT21 tumor cells demons trated focal accumulation of p185(erbB-2) in scattered clumps of cells and less p185(erbB-2) membrane expression than cells of SKOV3.ip1 tum ors. However, SKOV3.ip1 and SKOV3/pGT21 cells had similar in vitro sen sitivity to radiation. Conclusions: These data support the hypothesis that tumors with reduced p185(erbB-2) expression mediated by the anti- erbB-2 sFv are rendered more susceptible in vivo to the cytotoxic effe cts of ionizing radiation than tumors that maintain their normal expre ssion of p185(erbB-2). However, a similar effect was not observed with the same tumor cells in vitro. Thus, as has been described by others (1, 2), in vitro and in vivo results do not always correlate. Therefor e, appropriate assays to assess clinical relevance need to be determin ed for each particular system studied. (C) 1998 Elsevier Science Inc.