Methyl jasmonate is a promising tool against colon cancer

J Ávila-Román1, E Talero1, C de los Reyes3, E Zubía3, V Motilva1, S García-Mauriño2. 1University of Seville, Pharmacology 41012, Spain, 2University of Seville, Plant Biology and Ecology 41012, Spain, 3Universiy of Cádiz, Organic Chemistry 11510, Spain

Background. Oxylipins are metabolites produced by the oxidative transformation of unsaturated fatty acids. In mammals, oxylipins are formed mainly via the arachidonic (C20) acid cascade, and play a major role in inflammatory processes and, in general, in stress responses to infection, allergy and exposure to xenobiotics. In plants, phyto-oxylipins are mostly produced from polyunsaturated octadeca(e)noic (C18) fatty acids. The family of jasmonates includes phyto-oxylipins with main roles in plant responses to stress. Jasmonic acid (JA), methyl jasmonate (MJ) and some of their synthetic derivatives, exhibit anti-cancer activity in vitro and in vivo against cancer cells of various histological origins. In this regard, four mechanisms of action have been proposed to explain the anti-cancer activity of these compounds: 1) ATP depletion in cancer cells, 2) induction of re-differentiation via mitogen-activated protein kinase (MAPK) activity, 3) induction of apoptosis via reactive oxygen species (ROS) and pro-apoptotic proteins of the Bcl-2 family, and 4) inhibition of the phosphorylation and subsequent translocation into the nucleus of NF-κB-p65.

MJ has recently attracted attention as a hopeful antitumoral compound because of its highly specific pro-apoptotic properties in a wide range of malignancies. However, the high doses (mM) required to achieve a therapeutic benefit have limited its clinical development. The combination of jasmonates and various chemotherapeutic agents opens a new therapeutic prospective for these compounds.

Objectives. To study the in vitro cytotoxicity of MJ, either alone or in combination with 5-fluorouracil (5-FU), against the human adenocarcinoma colon cell line HT-29, and to clarify the mechanism(s) of action responsible for the effects.

Material and Methods. HT-29 cells were grown in McCoy´s 5A without glutamine supplemented with 10% FBS and Strep/Pen 1x, at 37ºC and in a 5% CO2 atmosphere. The growth inhibitory effect of MJ and 5-FU was evaluated with the sulforhodamine (SRB) assay. The cultures were treated for 48h with 0.5 to 5 mM 5-FU and 0.5 mM MJ. The effect on cell cycle arrest was tested for MJ at 3 and 6 mM for 24h. Statistical significance of the data was evaluated with Student´s t test.

Results. The in vitro cytotoxicity of MJ and 5-FU, either independently or in combination, was measured with the SRB method. The half maximal inhibitory concentration (IC50) of MJ was around 3 mM after 48h of exposure, being the range according with other cell lines. The pre-treatment (1h) with 0.5 mM MJ reduced the IC50 of 5-FU from 5 to 2.5 mM, thus resulting in an important enhancement of the cytotoxic activity of 5-FU. Cell cycle and apoptosis were monitorized by flow cytometry. The results showed that the treatment with MJ resulted in an S-G2/M arrest and the induction of apoptosis in HT-29 cells (Table.1).

Conclusions. The cytotoxic effect of MJ on HT-29 cells, showing IC50 in the milimolar range as published for other human cancer cell lines, is reported for the first time. MJ demonstrated the ability to produce cell cycle arrest and to induce apoptosis in these cells. More importantly, we show that MJ is able to enhance the cytotoxicity of 5-FU against HT-29 cells decreasing by a half the 5-FU amount necessary to achieve the same outcome. This improvement of 5-FU effects may allow reduction of the administered dose, decrease of unwanted side effects, and reduction of the likelihood that the tumor will display resistance to the combined therapy. This could be used as an alternative treatment option either alone or, principally, in combination with other chemotherapeutic drugs.

Table 1. Fase cells percentage (*p<0,05). The results were performed in duplicate separated assays.