Effects of a prostaglandin precursor, gamma-linolenic acid (GLA), on malignant cells in vitro and in vivo.
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Recent studies have shown that the proliferation of various human and murine tumour lines can be inhibited by the addition of gamma-linolenic acid (GLA) to the culture medium. These findings are consistent with the proposal put forward by Horrobin (1980) that malignant cells lack the enzyme/ A 6 desaturase; which is responsible for the conversion of linoleic acid (LA) to GLA. Since GLA is a prostaglandin (PG) precursor/ inadequate conversion of LA to GLA would result in decreased production of PGs/ particularly PGEi/ which has been shown to have an inhibitory effect on cell growth. Provision of GLA to enzyme deficient malignant cells should therefore bypass this blockade/ increase PGET synthesis and thus "normalise malignant cells". This study was performed to examine further the effects of exogenous GLA on growth of malignant cells in vitro and in vivo. Cells of the continuous murine sarcoma (M52B) line and primary cultures of non malignant fibroblasts were used to investigate effects of GLA in vitro. Cultures were exposed to either single or multiple doses of a range of concentrations of GLA. Radioimmunoassay (RIA) was performed to compare the amounts of PGE and PGF released into the medium by GLA treated and control M52B cultures and thus determine whether the addition of GLA in vitro significantly affected production of these PGs. Athymic BALB/c mice and immunocompetent BALB/c and Biozze mice as well as mice of the "Onderstepoort Strain" were used in various in vivo studies. Tumours were induced by the subcutaneous inoculation of approximately 1 x 106 cells of either the M52B line (into immunocompetent and athymic mice) or human breast carcinoma (NUB 1) line (into athymic mice). Take rates and latent periods were recorded. GLA treatment was initiated after tumours were established. In one study the fatty acid in hydrogenated coconut oil (HCO), which contains no PG precursors/ was administered parenterally (100 ug/ml/day) to Biozze mice. Control mice were either untreated or injected with HCO only. In another study, BALB/c mice and mice of the "Onderstepoort Strain" had their diet supplemented with GLA (in the form of EPO) to an extent of 3.5%. Control mice consumed either standard laboratory chow only or, chow supplemented with either 35% sunflower seed oil (SSO) or 35% HCO/ neither of which contain GLA. All diets were supplied ad libitum. Tumour sizes were measured every 48 hours and at the end of each experiment at which time tumours were excised and examined histologically. GLA was found to produce inhibitory and toxic effects on growth of both M52B cells and non malignant fibroblasts in vitro/ although the effect in the latter was observed only with high concentrations of the fatty acid. The inhibition of malignant cell growth was concentration dependant and was positively related to the duration of exposure to the fatty acid. Prior to death/ cells treated with GLA accumulated vii paranuclear granules which were shown histochemically to be lipid in nature. Electron microscopy confirmed the presence of large lipid deposits. Cultured M52B cells treated with GLA also released more PGE and PGF into the medium than did cells not exposed to the fatty acid. However, analysis of results using the Mann Whitney U test showed these differences to be statistically non significant for both PGE and PGF on two tailed tests. In contrast to the inhibition of M52B cell growth observed in vitro, growth of solid M52B sarcomas and NUB 1 carcinoma xenografts in athymic mice was apparently unaffected by administration of dietary GLA. Analysis of data using an unpaired student's t-test showed that the differences in tumour volumes between control and treated groups were not statistically significant either before or at the end of the experiment. While the inhibition of malignant cell growth caused by GLA in vitro was consistent with Horrobin's proposal that malignant cells may lack this PG precursor, whether or not these actions are mediated by the PGs remains obscure. Although an increase in PGE production by M52B cells was observed following GLA treatment, besides this increase being statistically non significant, it was not possible to determine whether this was due to PGE, (as suggested by Horrobin) or PGE2. It is possible that the effect produced in vitro was due to some factor other than raised PGE production, for example a non-specific fat-overload effect or a change in cell membrane fluidity. The lack of effect of GLA on tumour growth in vivo may have been due to inadequate delivery of the fatty acid to the tumour site. However, whatever the mechanism of action of GLA in vitro/ since oral GLA was supplemented to the maximum tolerated extent and produced no effect in immunodeficient mice inyiyo, it would seem that in a similar clinical situation oral doses which would be practical may be ineffective.