A Novel Combination of Piant Extracts withPromising Anti-Prostate Cancer Activity

by J. L. Hall, PhD, N. Springer, MSc, and D. L. Bemis, PhD

IntroductionProstate cancer is the most

common cancer in men. The causesof this disease are not wellunderstood, but prostate cancer isclearly associated with age, making Itseem like an almost inevitableaffliction for older men.' The diagnosisof prostate cancer is based onanalysis of a tissue biopsy -microscopic examination of a tinysample of prostate tissue. A positivebiopsy means that cancer cells arepresent and the conventionaltreatments include hormonal therapy,radiation therapy, and surgery with theactual regimen keyed to the stage ofcancer detected in the prostate andwhether there is evidence that it hasspread.

The majority of biopsies arenegative. The good news for thesemen - that they don't have cancer -Is tempered by the fact that theyremain at high risk for developingprostate cancer, especially those witha family history of the disease.Moreover, they must cope with thesymptoms that indicated prostateproblems in the first place, includingelevated PSA {Prostate SpecificAntigen, a marker for prostate cancer)and BPH (Benign ProstaticHyperplasia or swollen prostate, oftenassociated with urinary and sexualproblems).

There is no widely acceptedconventional treatment for theprevention of prostate cancer in high-risk men or for the alleviation of thesymptoms associated with enlargedprostate. The significant side effectsfrequently associated with hormonesand surgery has generatedconsiderable interest in the potentialof alternative treatments includingherbal remedies and nutritionalsupplements for alleviating symptomsand promoting prostate health. One ofthe challenges for anyone surveying

the growing list of vitamins, herbs, andminerals claimed to keep the prostatehealthy is to sort out whether thevarious promises are evidence-based.In other words: is there any sciencethat supports the use of the product,what is the quality of the research, andwhat do the data tell us? This reportsummarizes the development of anovel combination of herbalsupplements with anti-prostate canceractivity and describes the scientificmethods behind the product.

The Scientific Procedures forTesting and Validating PotentialRemedies

While the ultimate goal ofidentifying and researching naturalcompounds is to justify their use forpromoting or restoring the health ofhuman beings, there are a series oflaboratory experiments that areessential before any of thesecompounds can be evaluated forclinical use. These experiments,broadly referred to as pre-clinical,have evolved into a relativelystandardized progression starting within vitro or test tube procedures,followed by cell-based assays, andculminating with in vivo or animal-based studies, and finally, whenjustified, clinical trials in humanpopulations. This series ofexperiments is focused both onassuring that the compound may besafely tolerated by humans as well ason providing a scientific rationale forits application, hopefully to includesome understanding of how thecompound exerts its effect.

For natural compounds proposedfor use in humans, the priority is todetermine whether they exhibit toxicityand if so, to what extent. Considerableinformation can be gained from toxicitystudies in which various cellsmaintained in tissue culture areexposed to a range of concentrations

of the compound. The lower theconcentration at which cells are killed,the higher the toxicity and the lesslikely the compound is to be toleratedby animals. Compounds with limitedtoxicity may be further tested in a vastarray of in vitro studies. These mightinclude assays for their effect on cellproliferation (important for cancer), forexamining whether their effect istissue specific {important if the goal isto target a specific cancer), and forlocalizing their site of action in the celland for determining their biochemicalimpact {important if there is anindication of a specific anti-canceractivity). Many of these biochemicalexperiments, including enzymeassays, can be performed in cell-freesystems that take place in a miniaturetest tube containing test compoundsand their molecular targets.

What emerges from these in vitrostudies is a series of observationsconstituting a profile of thecompound's toxicity, physiologicaleffect, and mechanism of action. Thisprofile will dictate whether further testsand animal studies are worthwhile.The decision to pursue animal studiesis critical because these studies serveas a 'proof of principle' enablingexamination of a compound's effect inan organism, often a mammal, whosebiology closely resembles that of ahuman being. These tests can revealboth the true potential of acompound's benefits as well asunanticipated side effects that are onlyevoked in a complex organism. Theoptions for the design of animalstudies are too numerous and diverseto summarize here so a single relevantexample of an animal model systemthat has proven useful for cancerresearch will be reviewed.

Geneticists have developed strainsof mice that have proven to bevaluable in vivo models for studyingthe biological response to treatments

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Plant Extracts with Anti-Prostate Cancer Activity

for human cancers. Some of thesespecialized mice lack a thymus andhence cannot mount an immuneresponse.^ When human cancer cellsare transplanted into these mice, aprocess called a graft, the cells willgrow into a tumor without interferencefrom the host. Since the firstsuccessful transplantation of humantumors into athymic mice, manydifferent human tumor cell lines (e.g.,mammary, lung, genitourinary, gastro-intestinal, head and neck,glioblastoma. bone, and malignantmelanomas) have been transplantedand successfully grown in theseanimals. The mice are then treatedwith compounds to assess their affecton the transplanted tumor cells.Positive results in these experiments,meaning that the compound reducesor abolishes the tumor with little or noharm to the mice, provides anextremely powerful argument fortesting the compounds in humanclinical trials.

The experimental proceduresbriefly summarized above representthe pathway for establishing ascientific basis for the safety andactivity of any extract or supplementintended for improving human health.Products that have been subjected tothese procedures stand apart fromthose that have not been rigorouslytested. It is precisely this scientificlineage that was followed for thedevelopment of the novel combinationof plant extracts described below.

Beljanski's Experiments withPao Pereira and RauwoifiaVomitoria; Anti-Cancer Activity

Both Pao Pereira, a tree that growsin the Amazon rain forest andRauwoifia Vomitoria, a plant thatgrows in Africa, have a history of useas herbal remedies for a variety ofailments. The fact that extracts fromthese plants contain compounds withanti-cancer activity was firstdemonstrated by the Frenchbiochemist, Mirko Beljanskt, workingat the Pasteur Institute.

Beljanski developed a cell-free invitro assay called the Oncotest whichhe used to test compounds for theirpropensity to act as carcinogens."This test, at once simple and powerful,was based on his observation that theDNA isolated from cancer cells isdifferent from the DNA from healthycells in two respects: cancer cell DNAabsorbs more UV light and is moreactive as a template for in vitro DNAsynthesis. These observations areconsistent with the cancer DNA doublehelix being relatively more open orrelaxed in structure resulting in morefrequent loops of single stranded DNA.Beljanski saw this destabilization ofthe DNA duplex as an inherentproperty of cancer DNA and reasonedthat it accounts for the characteristicenhancement of UV absorption andtemplate activity. When Beljanskiadded known carcinogens to thecancer DNA he found that both thehyperchromicity (elevated UVabsorption) and the rate of DNAsynthesis were further increasedwhereas the results with normal DNAwere almost unchanged. Compoundswithout cancer causing effects wereneutral in the Oncotest, having nosignificant effect on chromicity ortemplate activity of either cancer orhealthy DNA,

Having established the validity ofthe Oncotest to identify carcinogens,Beljanski went on to use the test to tryand identify compounds that inhibit orreduce the UV absorption andtemplate activity of cancer DNAs whileat the same time have no effect onnormal DNAs. In a sense, thisinnovation amounts to using theOncotest in reverse and enablesscreening for compounds thatcounteract the effects of carcinogens.Any compounds found in thesescreens could be considered potentialanti-cancer agents. In the course ofthese experiments, Beljanski foundthat extracts from Pao Pereira andRauwoifia Vomitoria reduced UVabsorption and template activity ofcancer DNA in the Oncotest. Thepotential of these compounds was first

observed in these in vitroexperiments.^

Beljanski proceeded to test the Paoand Rauwoifia extracts in a fairlyextensive series of cell-basedassays.*"^° The extracts showedseveral consistent and noteworthyproperties. First, at optimalconcentrations, they stopped theproliferation of cancer cell linesmaintained in the laboratory, whilesparing healthy cells. This selectivitywas maintained across a broadspectrum of cell types including brain,colon, liver, kidney, and skin. In thecourse of these experiments Beljanskishowed that the extracts were activeagainst the prostate cancer cell line.

Finally, Beljanski performed in vivotests to explore the effects of the activecompound in the Rauwoifia extract oncancer cells transplanted into mice.'^In these experiments mouse cancercell lines from a lymphoma weretransplanted into mice of the samestrain in which the cancer originated.This ensured that the mice did notdevelop an immune response to thetransplanted cells, thus enabling theeffects of the Rauwoifia compound tobe clearly discerned. At the highestdoses, up to 80% of the mice treatedwith the Rauwoifia compound survivedfor ninety days whereas all of theuntreated mice were dead by day forty.As noted in the report of this study:

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Plant Extracts with Anti-Prostate Cancer Activity

"All mice that did not survive haddeveloped tumors (average weight 8.6+/- 2.1 grams). Cured mice survivedin excellent condition," These dataclearly demonstrated the anti-canceractivity of the Rauwoifia compound inan animal model system. Perhaps,most striking was the absence ofdeleterious side effects.

Based on their track record in theseand other experiments, the Pao andRauwoifia extracts were successfullyused in numerous human case studiesand ultimately developed as dietarysupplements by Natural SourceInternational, Ltd. These products,called Pao V® and Rovol V®.respectively, have been sold in Europefor many years and have recentlybeen introduced in the United States.As described below, the anti-canceractivity of both of these extractsremains the subject of ongoingresearch and development.

New Research with Pao Pereiraand Rauwoifia Vomitoria: Anti-Prostate Cancer Activity

As part of an effort to developnatural products targeted towardprostate health. Natural Sourceestablished a collaboration with Drs.Katz and Bemis in the Department ofUrology at Columbia University. Thegoal of these studies was to examinethe potential activity of the Pao andRauwoifia extracts specifically withregard to prostate cancer. Thepreliminary results of theseexperiments were presented at theFirst International Conference of theSociety of Integrative Oncology andare published in an abstract of theproceedings. '̂ A more complete reportwill be published next year.

The experiments, carried out by Dr.Bemis, encompass in Wfro cell-basedassays that confirm the anti-canceractivity of the extracts in the humanprostate cancer cell line LNCaP.These results justified an animal studyinvolving transplantation of LNCaPcells Into athymic mice andsubsequent administration of the Paoextract to the mice. The results of

these experiments were positive. Dr.Bemis also performed a series of cellfree in vitro assays that shed new lighton the mechanism of action of the Paoand Rauwoifia extracts. Theseexperiments, the details of which canbe found in the abstract, suggest thatwhile the mechanisms of the anti-prostate cancer action of the twoextracts overlap, they arenevertheless distinct. This observationsupported the concept of combiningthe two extracts together in the formof a new product. This product, calledProstabel®, is designed to takeadvantage of the activity of both Paoand Rauwoifia as well as anysynergistic effect afforded by theirdifferent mechanisms of action, Aclinical trial to examine the effects ofProstabel® in men with elevated PSAis expected to begin next year.

ConclusionThe demonstration of the anti-

prostate cancer activity of the PaoPereira and Rauwoifia Vomitoriaextracts and the development of acombination of the two extracts in theform of Prostabel® represents aformidable example of the applicationof scientific methods to the analysisand validation of natural products forpromoting health.

Dr. Hail is a Technical Consultantfor Natural Source international, Ltd.in New York; Natacha Springer is aResearch Associate for NaturalSource Internationai, Ltd. New York:Dr Bemis is an Associate ResearchScientist in the Department of Urologyat Columbia University in New York.

For Further Information on NaturalSource Products and Dr. Beljanski'sscientific research please visit thefollowing websites:www,natu rai-source.comwww.beljanski.comwww.mbschachter.comhttp://www.findarticles.com/cf_dls/mOISW/244/111271889/p1/article.jhtml

References1, See Ihe following websites:

http://www.prostate-cancer,orghttp ://www, p rostate "Ca nee r, org/http://www,cancer.govhttp://www.cancer,gov/http://www.cumc.columbia.edii/dept/urology/

2, HA Azar, SB Fernandez. LM Bros, and JLSullivan. Human (umor xenografts inathymic (nude) mice: chemotherapy trialsin serially transplanted tumors. Annals ofClinical and Laboratory Science, Vol 12,Issue 1, 51-59 Copyright © 1982 byAssociation of Clinical Scientists

3, Beljanski, M.. Oncotest: a DNA assaysystem tor the screening ot carcinogenicsubstances. IRCS Medical Science. 1979.7: p. 476.

A. Beljanski. M,. Le Goff, L.. Beljanski. M.S.Ditterential susceptibility of cancer andnormal DNA templates allows the detectionof carcinogens and anticancer drugs. ThirdNCI-EORTC Symposium on new drugs incancer therapy. 1981. Institut Bordet.Bruxelles.

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8, Beljanski. M., Crochet, S., Beljanski, M.S.PB100: A Potent and Selective Inhibitor ofHuman BCNU Resistant Ghoblastoma CellMultiplication. Anticancer Research. 1993.13: pp, 2301-2308.

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11, Beljanski, M. "The anticancer agent PB-100, selectively active on malignant celllines, even multidrug resistant." Geneticsand Molecular Biology 23A (2000): 29-33.

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13, Abstracts of Ihe First InternationalConference of the Society ot IntegrativeOncology, November 17, 2004.

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