ABSTRACTS

BIOLOGY

GROWTH FACTORS/ CELL SIGNALING

001

INHIBITION OF BOMBESIN STIMULATED MITOGENESIS IN A SMALL-CELL LUNG CANCER CELL LINE BY DEMETHOXYVIRIDIN; A PUTATIVE INHIBITOR OF RECEPTOR-MEDIATED PHOSPHOLIPASE D ACTIVATION. Jane A. Plumb, R. Milroy and S.B. Kaye, CRC Department of Medical Oncology, University of Glasgow, Beatson Laboratories, Garscube Estate, Bearsden, Glasgow G61 lBD, U.K.

Most oncogene products are components of intracellular signal transduction pathways and transfection of cells by various oncogenes has been shown to subvert the regulation of pathways controlling second messenger concentrations. This suggests that these pathways contain sites that could serve as targets for novel anti-cancer agents. Following mitogen stimulation protein kinase C is activated by a sustained elevation of diacylglycerol (DAG). This DAG is derived from the breakdown of phosphatidylcholine to phosphatidic acid (PA) by phospholipase D (PLD) and subsequent conversion of PA to DAG by phosphatidate phosphohydrolase. Since PA may be an important mediator of the mitogenic signal we have identified PLD as a novel target for drug development. In human neutrophils the fungal metabolite wortmannin and the structural analogue demethoxyviridin (DMV) inhibits IMet-Leu-Phe-stimulated PLD activation.

We have shown that DMV inhibits the growth of bombesin stimulated Swiss 3T3 fibtoblasts with an IDso of 1uM. Growth of the human small-cell lung cancer cell line NCI-H69 can be stimulated by bombesin. When grown in medium containing foetal calf serum the IDso for DMV was 2uM. However, bombesin stimulated mitogenesis was inhibited by 0.2uM DMV. In the fibroblast cell line DMV was only a weak inhibitor of PLD but was a potent inhibitor of phospholipase Az(PLAz, ICsc 1nM). Furthermore, we have shown that DMV inhibits tyrosine phosphorylation induced by bombesin in Swiss 3T3 cells and also inhibits autophosphorylation of the EGF receptor in A431 calls. These results indicate that DMV is a potent inhibitor of growth factor stimulated mitogenesis. Although inhibition of PLD was not observed DMV may act through inhibition of PLAz and/or receptor tyrosine kinase activity.

002

TRANSFORMING GROWTH FACTORu(TGFu~ IS RELEASED BY A SMALL CELL LUNG CANCER (SCLCI CELL LINE AND MAY ACT AS AN AUTOCRINE GROWTH FACTOR. L. Damstrup, M. Spang-Thomsen, R. Graves-Deal, P.J. Dempsey, R.J. Coffey, H. Skovgaard Poulsen, Dept. Med. & Cell Biology, Vanderbilt University, Nashville, TN, Institute of Pathological Anatomy, University of Copenhagen and Dept. Oncology, Rigshospitalet, National University Hospital, Denmark.

We have previously demonstrated that 11121 SCLC cell lines in our panel express epidermal growth factor receptor (EGFR). EGF and TGFo binds to the EGFR and we have examined the mitogenic effect of exogenous TGFa and EGF to the EGFR positive SCLC cell line CPH 54A. No significant mitogenic effect was seen with administration of either recombinant human peptides under serum-free or 10% serum-containing conditions, as determined by 3H-thymidine incorporation. In the presence of the EGFR monoclonal antibody, 528, a minor inhibition of mitogenicity was found. To determine whether this insensitivity was due to endogenous TGFo production, levels of TGFo protein in conditioned media was determined by a sensitive and specific TGFo RIA, in the presence and absence of 528. Confluent CPH 54A cells released 5.4 ng TGFu/lO” cells, whereas this increased to 6.8 ng/lO’ cells in the presence of 528. Another experiment showed that 80% of the produced TGFo was found in the medium and 20% in the cell lysate.

Our results indicates that in CPH 54A TGFu has an active role possible in an autocrine manner. Further exploration regarding additional SCLC cell lines and other TGFu family members will be presented.