The search for bioactive compounds in tropical plants to target hormone imbalance associated diseases.

Abstract

Benign prostatic hyperplasia (BPH) and/or prostate cancer (PC) will affect at least 50% of the males once they have reached their fifties. However, despite the range of medical therapies available, effective treatment against BPH and PC still currently remains inadequate for some. The annoying symptoms of BPH are mainly attributed to an enlarged prostate. Therefore, the current treatment strategy is to halt the androgen-dependent growth of the prostate and reduce its size. Several drugs have been employed with variable success to control prostatic growth. However, patients tend to self-medicate over a long period. As a result, this leads to another problem, unpleasant long-term side effects. The treatment of PC in its early stages often warrants disease free survival for about 70-80% of the patients. Despite early aggressive therapy, 20% of the cases, unfortunately, experience disease progression to a state where the cancer no longer responses to therapy. At the moment, well-established medical options for this condition are limited and thus PC is one of the leading causes of cancer-associated deaths in western countries. Evidence has supported the undoubted role of the androgen-signalling pathway in BPH, the pre cancerous prostatic hyperplasia and dysplasia that may progress to PC. The reduction of androgen-dependent prostatic growth has been the rational endocrine therapy for both BPH and PC. However, since the etiology of both diseases is multi-faceted, it is necessary to consider other contributing factors to develop more effective medication. Medicinal plants are considered to be multi-component drugs (they contain numerous phytochemicals) and are thought to display a wide range of beneficial effects. They have been used therapeutically for centuries. Because of their historical place in medicine, they may have a better safety profile than synthetic drugs. The objective of this thesis is to identify tropical medicinal plants, which could be used to target or support treatments for BPH and PC. Twenty herbal plants, with no known to date indications for both diseases, were selected. They were fractionated by using different ethanol (EtOH) concentrations. The initial screen (Chapter 4) aimed to identify plant extracts with the ability to inhibit the proliferation of LNCaP cells, an androgen dependent human prostate cancer cell line. All extracts were tested at a concentration of 30 μg/mL. Four extracts, Api, (70% EtOH Alpinia oxyphylla extract), Aquil (70% EtOH Aquilaria sinensis extract), Astra (aqueous Astragalus membranaceus extract) and P9605 (96% EtOH Piper cubeba extract) were selected for further investigations. Recent research has demonstrated that androgens are not solely responsible for BPH and PC, estrogens, defective apoptosis and inflammation are, for example, also involved. An experimental test system using several methodological approaches was designed to test the above-mentioned extracts. The potential cytotoxicity of the extracts was investigated first to ensure that they did not attenuate LNCaP growth by inducing unspecific cell death. The extracts were also tested on HepG2 cells, a human hepatocarcinoma cell line, to identity any potential induction of liver-toxicity. Anti-androgenic and anti-estrogenic effects were determined by observing if the extracts 1) blocked the production of certain androgens and estrogens, 2) the steroid hormone receptor activation process, and 3) the actions of these sex hormones. The ability to induce apoptosis and the anti-inflammatory properties of the extracts were also tested. The methods employed were validated and synthetic controls were used whenever possible and compared with literature. Api reduced the cellular viability of LNCaP and HepG2 cells at 20-30 μg/mL. It was not further investigated because the apparent reduced LNCap cell growth was most probably attributed to due to its cytotoxicity. The other extracts were non-cytotoxic on both cell lines at 30 μg/mL. Astra inhibited androgen-dependent growth of LNCaP cells, however it did not show significant anti-androgenic, anti-estrogenic and anti-inflammatory properties. Unfortunately, it is beyond the scope of this project to discover its anti-proliferative mode of action. The results of Aquil and P9605 derived from the test system were more promising. P9605 inhibited 5a-reductase type II and aromatase, which were involved in synthesising dihydrotestosterone (DHT) and estradiol respectively. It also antagonised the effects of DHT by several mechanisms. Furthermore, it inhibited the cyclo-oxygenases (COX) and 5-lipooxygenase which are involved in generating inflammatory mediators. Aquil possessed similar properties as P9605, except that it had no effects on the COXs. In conclusion, we have identified some possible mechanisms of 2 tropical plants, Aquilaria sinensis and Piper cubeba, which could potentially be used to prevent/alleviate BPH and/or PC. This is the first time that these plants have shown to possess anti-androgenic and antiestrogenic properties.