SUMMARY This is the first report of human gastrointestinal arachidonate and prostanoids measured quantitatively by gas chromatography-mass spectrometry (GC-MS) The methods for gas chromatography-mass spectrometry (GC-MS) were as reported previously.' In brief, the dried extract obtained as described above was dissolved in dichloromethane. An aliquot was taken for GC-MS and further purified by LH20 column chromatography; non-polar impurities were eluted with dichloromethane, and the eicosanoids were eluted with methanol which was then evaporated. Deuterated standards were added, and each dried methanol extract was dissolved in doubledistilled water acidified to pH 3 with hydrochloric acid. Each solution was percolated through an Amberlite column which was then washed with distilled water, and the eicosanoids were eluted with 315 on 11 May 2018 by guest. Protected by copyright.

1The effect of indomethacin on the response of the NC carcinoma to methotrexate has been examined in vivo and in vitro.

Indomethacin reduced the tumour prostaglandin yield, but the biological activity in extracts of tumours from mice given di-me-PGE2 was high. The median survival time was longer in mice receiving indomethacin alone (61 days from tumour transplantation compared with 50 days in controls P<0.02). Di-me-PGE2 alone had little or no effect on survival (median 48 days) but counteracted the increase with indomethacin (di-me-PGE2 +indomethacin, 49 days median survival). There were no obvious effects of the treatments on tumour recurrence at the excision'site, but there was a higher incidence of involved lymph nodes in mice given di-me-PGE2.

Summary Mice transplanted with NC carcinoma were treated with the thromboxane synthetase inhibitor dazmegrel (UK38485) or with nafazatrom (BAY G 6575), a compound that is reported to increase prostacyclin formation. Some experiments included the cytotoxic drugs methotrexate and melphalan. The tumours were excised under anaesthesia on day 14 or day 21 after transplanation, and weighed; some were extracted for prostanoids which were measured by radioimmunoassay. Mouse survival time was determined up to day 121, and cancer spread was determined by postmortem examination. The survival was increased by methotrexate and melphalan but not by the other drugs. Nafazatrom-treated mice tended to have lighter tumours. Although dazmegrel reduced the formation of thromboxane B2 during clotting of blood from normal mice, it did not affect the tumour yields of prostanoids. Nafazatrom had no effect on serum or tumour prostanoids. There were no obvious effects of the treatments on the recurrence of tumour in the excision scar, lung metastasis or spread to lymph nodes.Effects of the arachidonate metabolites thromboxane A2 (TXA2) and prostacyclin (PGI2) on platelet aggregation are well known. The ability of TXA2 produced by platelets to cause their aggregation may normally be balanced by PGI2 which is inhibitory (Moncada & Vane, 1979). Platelet aggregation is thought to be important in the haematogenous spread of some tumours, and Honn (1982) and his colleagues (Honn et al., 1981(Honn et al., , 1983 suggested that circulating tumour cells, or vesicles shed from the primary tumour cells, disrupt the balance between PGI2 and TXA2 in favour of platelet aggregation. Experiments with intravenously injected B-16a melanoma cells showed that thromboxane synthesis inhibitors, PGI2, or nafazatrom (which has various actions including an increase of PGI2 formation), are antimetastatic (Honn, 1982;Honn et al., 1983). A thromboxane synthesis inhibitor also reduced spontaneous metastasis from Lewis lung carcinoma (Honn, 1982 Drugs that inhibit cyclo-oxygenase usually reduce the size of NC tumours, and prolong host survival when given alone or with the cytotoxic drugs methotrexate and melphalan (Bennett et al., 1979(Bennett et al., , 1982. Cyclo-oxygenase inhibitors reduce both thromboxane and prostaglandin formation, but it is not known if the effect on thromboxane production contributes to the effects of indomethacin and flurbiprofen on tumour size and mouse survival.The aims of the present study were: (i) to examine Honn's hypothesis, using nafazatrom or the thromboxane synthetase inhibitor dazmegrel in the mouse NC tumour model, (ii) to measure mouse survival time, tumour weight and prostanoid content and (iii) to study the effect of dazmegrel in combination with cytotoxic drugs. Materials and methodsThe NC carcinoma used in these studies originally arose spontaneously in the mammary region of a WHT/Ht mouse (Hewitt et al., 1976), the same strain used in our experiments. Following local excision of this carcinoma there is a high incidence of l...

Summary WHT/Ht mice were transplanted s.c. with NC carcinoma, and the tumours were excised after 2 weeks. The mice were treated orally throughout the experiments with prednisolone 500pgkg-' or mepacrine 3.6mg kg-1, starting the day after tumour transplantation or, with prednisolone, the day after tumour excision. In some experiments the mice were also treated with the cytotoxic drugs methotrexate 2mg kg 1 and melphalan 1.4mgkg-1. The excised tumours were weighed; some of them, and samples of serum, were extracted for prostanoids which were measured by radioimmunoassay. The chemotherapy lengthened the survival of the mice, but prednisolone or mepacrine had little or no effect on survival, metastasis, the response to chemotherapy, tumour size or the formation of tumour prostanoids.Prostaglandin synthesis inhibitors have been extensively investigated in various murine cancers, and they usually show a beneficial effect (Bennett, 1982;1986). In the NC tumour model the prostaglandin synthesis inhibitors indomethacin or flurbiprofen increase mouse survival and usually reduce tumour size. Survival is even longer when these nonsteroidal anti-inflammatory drugs are used with the cytotoxic drugs methotrexate and melphalan . The effects of the anti-inflammatory drugs seems likely to result from inhibition of prostaglandin synthesis rather than from other properties that the drugs may have (Flower, 1974), since administration of a PGE2 analogue counteracted the effect of indomethacin (Bennett et al., (1985). A similar effect might therefore be expected with other drugs that reduce prostaglandin formation, such as corticosteroids and mepacrine which can inhibit phospholipase activity and so depress the release of prostaglandin precursors (Vane et al., 1982). This action may be particularly relevant to human cancers since prednisolone is frequently used in chemotherapy regimens. We have therefore studied the effects of prednisolone and mepacrine, alone and in combination with the cytotoxic drugs methotrexate and melphalan, in mice with NC tumours. Measurements were made of tumour weight and prostanoid content, the occurrence of metastases, and mouse survival. Materials and methodsThe original NC tumour arose spontaneously in the mammary region of a WHT/Ht mouse (Hewitt et al., 1976) and has since been passaged only in this strain. It has a high incidence of local lymphatic spread, recurrence in the scar following tumour excision, and metastasis mainly to the lungs and mediastinum.On day 0, male or female WHT/Ht mice were injected s.c. into the left flank with -106 NC carcinoma cells in a singlecell suspension from passaged tumours as described previously (Bennett et al., 1979. All tumours were excised at 2 weeks and weighed. The study consisted of 6 separate experiments, each with 9-15 WHT/Ht mice/group. Drugs were given orally in 0.1 ml 50% syrup BP for 5 days (Monday to Friday) each week, and treatment with prednisolone or mepacrine was continued until death or the end of the experiment (day 121). The doses chosen are approx...