Australian cane toad venom as a potential herbal medicine for prostate cancer therapy: assessment of quality, therapeutic effects and compatibility with traditional Chinese medicine

Jing, Jing (2013). Australian cane toad venom as a potential herbal medicine for prostate cancer therapy: assessment of quality, therapeutic effects and compatibility with traditional Chinese medicine PhD Thesis, School of Pharmacy, The University of Queensland.

       
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Author Jing, Jing
Thesis Title Australian cane toad venom as a potential herbal medicine for prostate cancer therapy: assessment of quality, therapeutic effects and compatibility with traditional Chinese medicine
School, Centre or Institute School of Pharmacy
Institution The University of Queensland
Publication date 2013
Thesis type PhD Thesis
Supervisor Harendra S. Parekh
Ming Wei
Total pages 216
Language eng
Subjects 0301 Analytical Chemistry
1112 Oncology and Carcinogenesis
Formatted abstract
Prostate cancer is the most common malignancy among men worldwide. Conventional treatment options are often accompanied by inevitable and debilitating side-effects and when the primary disease turns metastatic or hormone refractory, treatment options are exhausted and only palliative care remains. Herbal medicine, most notably traditional Chinese medicine (TCM) is however, yielding important breakthroughs in cancer prevention and treatment, and is increasingly used first line in many parts of the developing world. Thus, the broad aims of the studies described herein were to explore potential herbal agents derived from TCM for the treatment of prostate cancer.

In keeping with these aims, in Chapter 1 we first introduced the basic principles of TCM, elaborating on the rationale behind the composition and formulation of herbal remedies as well as their primary modes of action in cancer treatment; this was followed by a review of the analytical methods commonly employed to investigate their diverse chemical profiles. Next, experimental studies on the families of effective anticancer herbal remedies were highlighted along with their potential effects in cancer treatments. Most notably, the‘bufadienolides’, a class of compounds extracted from Asiatic toad venom (Chan Su, a precious and ever-diminishing TCM with tremendous medicinal value) was the focus of discussions, these being well-recognized agents for the treatment of various cancers, especially of the lower body such as the colon and prostate gland.

Being motivated by the historical use of toad venom in TCM, in Chapter 2 we explored the chemical composition of Australian cane toad venom (ACTV), primarily in terms of profiling bufadienolides in collected venom samples. ACTV was collected from cane toads in the greater metropolitan areas of Brisbane which was assayed to profile its quality through the use of both HPLC and HPLC/MS-TOF; with direct comparison made with the well-reported constituents of Asiatic toad-derived venom. We demonstrated that methanolic extracts of ACTV were representative and in addition to possessing characteristic bufadienolides, our highly sensitive detection technique yielded an additional compound, telocinobufotoxin,thus expanding the known chemical diversity of ACTV.

Primary assessment of the cytotoxic effects of ACTV extract was explored next, in Chapter 3. An MTT assay was employed and optimized, with data confirming that ACTV possessed appreciable in vitro cytotoxicity. EC50 values for ACTV were found to be in the nanomolar range in two "classical" prostatic cancer cell lines, namely, PC3 & DU145; this being comparable to cinobufotalin, a representative compound within Chan Su. Additionally,similar concentrations of ACTV lacked significant cytotoxicity when tested in the noncancerous human umbilical vein endothelial cell line.

In Chapter 4, we describe how a novel ACTV-containing three herb recipe was rationally derived, which is expected to enhance the therapeutic effects of ACTV, whilst decreasing its administered dose when applied for prostate cancer treatment. Separately, we also challenged the traditional preparation methods for herbal ‘teas’, comparing the chemicalcomposition of hot versus cold water extraction of the herbs through the use of HPLC profiling. Interestingly, we detected no appreciable difference, at least chemically, when comparing the three-herb decoction prepared by the two extraction methods i.e. hot vs cold water.

In Chapter 5, the principles of TCM were put to the test using the newly derived herbal recipe. We developed an in vivo proof-of-concept biodistribution study to assess whether our herbal mixture could retain its cytotoxicity following oral gavage, while also being detectable in the lower body, and in our case the prostate gland. Here, serum and prostate tissue samples collected from rats following oral gavage of ACTV and the ACTV-based recipe were separately scrutinized using HPLC/MS-TSQ and further screened for their cytotoxicity in PC3 cell lines using an MTT assay; this work being undertaken personally, although in a collaborator’s laboratory in China. Interestingly, although bufadienolides present in ACTV occur in both unconjugated and C-3 conjugated forms, only the unconjugated (metabolized) bufadienolides were detected in rat serum and prostate tissue samples following oral administration. Next, when compared with the control group, PC3 cells incubated with culture medium containing 10% ACTV and herbal recipe-treated serum both showed comparable growth inhibition rates, indicating that the in vivo biotransformation of herbal constituents did not abolish their cytotoxic profile.

Finally, in Chapter 6 we surmise the overall findings of the systematic research presented in this thesis, reflecting on the viability of ACTV and the herbal recipe for adoption as a possible future (adjunct) therapy for prostate cancer, as well as some limitations stemming from this work. In closing we discuss the outcomes from our initial hypotheses, while providing some direction and insights on what could form the basis of future studies to further validate the presence and place of ACTV in TCM-derived cancer therapies.
Keyword Prostate cancer
Traditional Chinese medicine
Quality control
Australian cane toad venom
HPLC
LC-MS
MTT assay
biotransformation

 
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Created: Tue, 11 Mar 2014, 13:48:32 EST by Ms Jing Jing on behalf of Scholarly Communication and Digitisation Service