Anti-tumour potential of a gallic acid-containing phenolic fraction from Oenothera biennis

Abstract

A phenolic fraction purified form defatted seeds of Oenothera biennis promoted selective apoptosis of human and mouse bone marrow-derived cell lines following first-order kinetics through a caspase-dependent pathway. In non-leukemia tumour cell lines, such as human colon carcinoma CaCo₂ cells and mouse fibrosarcoma WEHI164 cells, this fraction inhibited ³H-thymidine incorporation but not cell death or cell cycle arrest. Human peripheral blood mononuclear cells showed low sensitivity to treatment. Single bolus injection of the phenolic fraction could delay the growth of established myeloma tumours in syngeneic animals. HPLC and mass spectrometry analysis revealed that the fraction contains gallic acid. However, the biological activity of the fraction differs from the activity of this phenol and hence it should be attributed to other co-purified molecules which remain still unidentified.

Introduction

Plants are a well-known source of molecules with pharmacologic activity. A huge number of secondary metabolites of plant origin are now being investigated in pre-clinical and clinical settings as potential or actual anti-tumour drugs. Among these are vincristin and vinblastin, two alkaloids extracted from Catharantus roseus, etoposide, a glucoside from Podophyllum peltatum, and paclitaxel, a diterpen extracted from Taxus brevifolia [1]. Phenolics are widespread in the vegetal kingdom and most of these molecules have anti-inflammatory properties [2]. Other phenolics, such as gallic acid (3,4,5-trihydroxicbenzoic acid) and its derivatives, show selective cytotoxicity against a variety of tumour cells with a higher activity than that shown against normal cells [3], [4], [5], [6]. These compounds may serve per se as anti-tumour drugs or as the molecular basis for the synthesis of new drugs by targeted chemical modifications.

Oenothera biennis (known as evening primrose) is a herbal plant. The edible oil from its seeds has been shown to have several pharmacologic effects [7] though studies on the onset of possible side-effects deriving from its consumption are still scanty. Recent work has demonstrated specific anti-tumour activity of an ethanol extract from evening primrose defatted seeds [7]. This extract can induce apoptosis in tumour cells through a caspases-independent pathway triggered by increased levels of intracellular peroxides which in turn promote translocation of apoptosis-inducing factor (AIF) from mitochondria to the cell nucleus [8], [9]. In addition, in tumour cells the extract can also inhibit the phosphorylation of the retinoblastoma tumour suppressor protein thus promoting inhibition of DNA synthesis and cell cycle arrest [7]. Though promising, these studies do not address the chemical composition of the evening primrose extract that is likely to contain a mixture of ethanol-soluble molecules, among which phenolics. As a consequence, a positive identification of the bioactive molecule(s) is still missing. In addition, to our knowledge no information is available on the possible anti-tumour effects of phenolics from evening primrose.

We have studied the anti-tumour potential of phenolics extracted from defatted seeds of evening primrose using either leukemia and non-leukemia human and mouse cell lines. Human peripheral blood mononuclear cells (PBMC) were also used as normal cells. An HPLC-purified fraction showed potent and selective cytotoxic effects against bone marrow-derived tumour cells in vitro and in vivo, where it delayed the growth of established tumours. Attempts were made by mass spectrometry to further characterize the molecular content of this phenolic fraction.