[i] ANTIMETASTATIC ACTIVITY OF PROPOLIS, CAFFEIC ACID PHENETHYL ESTER AND CAFFEIC ACID AGAINST MAMMARY CARCINOMA OF CBA MICE

312        I. BASIC, N. ORSOLIC, A. BRBOT-SARANOVIC,   Z. TADIC, 
                                              D. SULIMANOVIC

                         (CROATIA)

We have studied the antimetastatic efficacy of caffeic acid phenethyl ester (CAPE), a propolis-derived compound and compared it with the antimetastatic effect of either water-soluble derivatives of propolis (WSDP) or caffeic acid (CA).  The tumor was a transplantable mammary carcinoma (MCA) of spontaneous origin, weakly immunogenic, to a syngeneic CBA mouse.  Metastases in the lung were generated by injecting 2 x 10⁵ viable tumor cells intravenously.  Tested compounds were given per os before or after tumor cell inoculation, the dose comprises 12.5 mg/mouse of either CAPE or CA, and 50 mg/mouse of WSDP, respectively.  Both the preventive and the curative therapy significantly reduced the number of tumor nodules in the lung of mice.  The antimetastatic effectiveness of both WSDP and CAPE was equally high and also higher than that of CA.  Two approaches for testing the possible mode of antimetastatic activity in the lungs were proposed: a modulation of immune reaction of recipients or an induction of apoptosis during the formation of metastatic nodules.  Changes in several immunological parameters, such as the response of lymphocytes to polyclonal mitogens in vitro, the production of tumor necrosis factor (a (TNF a) by lymphoid cells in vitro and the rosette formation of lymphoid cells with SRBC, correlated well with the antimetastatic properties of the tested compounds.  A direct relationship was also found between the antimetastatic activity of the compounds studied and the induction of apoptosis.

                                                                                                p.119

[ii]

Kittharz – die antibiotische  Alternative

von Walter Binder (Naturheilpraxis 5/79)

Das Kittharz der Bienen hat nach Auffassung zahlreicher Forscher in aller Welt, insbesondere in Rumänien, der UdSSR, den USA und in Dänemark eine hervorragende antibiotische Wirkung.  Eine stattliche Kasuistik untermauert diese Meinung Zunächst muß festgestellt werden, daß die Lebensform Biene, so wie sie heute in Erscheinung tritt, seit 42 Millionen Jahren existiert und eine abgrundtiefe Zahl anderer Insektenformen überlebt hat.  Das hat sicher seinen Grund.  Die ihr zugedachte Rolle, einmal die Bestäubung der Blüten, zum anderen die Verbreitung verschiedenster Pflanzensamen, hat ja auf die Pflanzenwelt eine mutagen-dynamisierende Wirkung und hilft ihren Bestand sichern.

Wenn eine bestimmte Tierart in unserer Biosphäre überlebt, dann muß ihr Organismus eine Reihe von wirksamen Programmen haben, die im Notfall rechtzeitig einsetzen.  Unter den entscheidenden Eigenschaften ist wohl die wichtigste jene, wirksame Abwehrmechanismen gegen Mikro- wie auch Makro-aggressoren zu besitzen.  Gerade diese Eigenschaften, eine universelle bakterizide und eine bakteriostatische Sekretproduktion, besitzen die Bienen.  Daß z. B. in einem Bienenkorb von, Kubikmeter Volumen, ohne endemische Einbrüche 50.000 bis 60.000 Insekten auf engsten Raum und unter feuchttropischen Klima existieren, erscheint wie ein Wunder.  Deshalb ist es auch einleuchtend, daß alle Produkte, die mit dem Speichel der Biene verarbeitet werden, von Bakterien, Viren und Pilzen nicht angegriffen werden können.

Den-Kittharz (auch Propolis genannt) besorgen die Kittharzbienen im Spätsommer und Herbst (August bis Oktober), wenn durch starke Sonneneinstrahlung.die Bäume harzen und das Harz flüssig genug ist für den Transport.  Die Harzpartikel werden mit dem enzymreichen Speichel löslich gemacht und in den Kitthöschen deponiert. Im Stock übernehmen andere Arbeitsbienen den Kittharz, speicheiln ihn erneut ein und verschließen damit alle Ritzen und Spalten in der Stockwandung.  Auch das Flugloch wird mit Kittharz eingefaßt.  Es kommt den Bienen darauf an, sich für die kommenden Wintermonate vor Feuchtigkeit, Kälte, Zugluft, und, was das allerwichtigste ist, vor Mikro- und Makroorganismen zu schützen.

Unter anderem verwenden die Bienen den Kittharz auch zur Mumifizierung größerer Tiere, die nicht mehr aus dem Stock entfernt werden können, z.-B. kleine Eidechsen, größere Käfer usw.  Der Kittharz hat eine hervorragende konservierende Wirkung, und verhindert die bakterielle Zersetzung.  Befallen verschiedene Mikroorganismen z. B. der Pilz Aspergillus flavus die Bienenbrut, so werden die Pilzverseuchten Strecklarven sofort mit Kittharz bedeckt und damit auch die resistenten Sporen unschädlich gemacht.

Ersatz für Penicillin

Nun zum medizinischen Teil : Mikrobiologische Untersuchungen mit Tuberkelbazillen wurden bereits in Dänemark (1948 ) gemacht.  Dabei hat sich ein wäßriger Extrakt aus Propolis gegen die an sich resistenten Bakterien als wirksam erwiesen.  Später wurden erstmals an der TU in Kopenhagen Propolisextrakte an 30 verschiedenen Mikroorganismen angewandt und eine wachstumshemmende Wirkung beobachtet. 1968 untersuchten die dänischen Forscher Jens Hoiriis Nielsen und Hans Erik Christensen die Wirksamkeit von äthanolischen Propolisauszug gegen Schmarotzerpilze.  Das Ergebnis verlief für alle Pilze tödlich.  In einem Parallelversuch wurden 15 Propolisproben gegen 39 Bakterienstämme und 20 Schmarotzerpilzen angewandt.  Davon wurden 24 Bakterienstämme in ihrem Wachstum gehemmt und alle 20 Pilzarten getötet.  Die rumänischen Wissenschaftler.  A. Derevici und A. Popesku fanden heraus, daß-Propolisextrakte auf Enterococcen eine absolut tödliche Wirkung haben, jedoch nicht auf Staphylococcen.  Am staatlichen Forschungsinstitut für Medizin in Kazan (UdSSR) (1963) wurden  50 Tuberkulosepatienten mit einer propolishaltigen Ernährung kurmäßig versorgt.  Das Resultat war außerordentlich zufriedenstellend.  Todorov, ein bekannter bulgarischer Wissenschaftler, entdeckte die günstige Wirkung von Propolisextrakt auf das vegetative Nervensystem und beobachtete eine Gefäßerweiterung im mittleren Arterienbereich.  Das wissenschaftliche Institut für Röntgenologie, Radiologie und Onkologie in Kiew (UdSSR) hat nach Strahlenverbrennungen die Haut mit Propolissalben erfolgreich behandeln können.  Am staatlichen Laboratorium in Baschkirien (UdSSR) veröffentlichte der Wissenschaftler Tjeannitsjev einen profunden Bericht über die spezifischen Eigenschaften des Propolis gegenüber der Abwehr und wies auch eine hämatogene Wirkung nach.  Seine Forschungsergebnisse belegte er mit 2000 Versuchen an Patienten.

Nach seiner statistischen Studien der Kasuistik sind die größten Erfolge bei Infektionen des Nasen-Rachenraumes zu verzeichnen.  Es folgen Rhinitis, Sinusitis, Otitis media, Conjunktivitis, verschiedene Influenzaformen, unklare grippöse Infekte mit bronchitischer, Tendenz, Pyelonephritis, Cystitis, Prostatitis, Colitis, alle Enteritiden, Gastritis, Ulcus Duodenum sowie Magengeschwüre.

Extern:  Ekzeme vor allem Mykosen, Ulcus cruris, nekrotische Gewebsprozesse, Brand, Karbunkel, und auch Phlegmonöse Entartungen.

Es läßt sich ohne Übertreibung sagen, daß Propolis die allmählich stumpf werdende Penicillinwaffe ersetzen könnte.  Vor allem die einmaligen Ergebnisse von Z. G. Tjeannitsjev geben zu denken.  Er hebt im besonderen hervor, daß die Phagozytenzahl merklich ansteigt, die Emigrationsgeschwindigkeit der Leukozyten beträchtlich zunimmt und im serologischen Test die Antikörper erhöht waren.  Ihre Reaktionsgeschwindigkeit mit dem Antigen scheint vergrößert.  Außerdem sein die fibrinolytischen Faktoren vermehrt und die roten Blutkörperchen zahlreicher. Er konnte auch an Hand seiner Kasuistik nachweisen, daß die Zahl der Koagulopathien spürbar sank.

Resümee:              Die Wirkung.von Kittharz und seine komplexe Einflußnahme auf den Organismus ist um so rätselhafter, wenn man die chemische Inhaltsanalyse dazu vergleicht: 55 % Harz und Balsam, 30% Wachs, 10% flüchtige Öle, 5% Pollen, und als mineralische Verbrennungsrückstände: Calcium, Aluminium, Vanadium, Mangan und Silicium.

Einerseits scheint es auf das erythropoetische System eine gewisse Anstoßwirkung zu haben, das gesamte RES zu aktivieren, die fibrinolytische Entstehung in der Leber und seine hämatogene Entfaltung zu begünstigen, auf die Bakterienbesiedelung des enteralen Abschnittes selektpositiv einzuwirken, andererseits den adrenerg bedingten Hochdruck im  mittleren Arterienbereich und Arteriolensystem durch sympathatikolytische Einflüsse zu senken.  Fraglich ist auch eine anabol-hormonale Wirkung.  Extern fördert Propolis die Granulierung und Epithelisierung von Wunden, Ekzemen und gutartigen Hautentartungen.

[iii] [26]    El-Ghazaly, M. A., & Khayyal, M. T. (1995). The use of aqueous

propolis extract against radiation-induced damage. Drugs Exp Clin Res,

21(6), 229-36, Whole body exposure to gamma radiation has been

experimentally shown to exaggerate inflammatory responses and to enhance

the release of mediators. A thirteen per cent aqueous extract of propolis

(bee glue) was previously shown to have potent antiinflammatory activity.

The present study was carried out to show whether the extract could

influence the exaggerated inflammatory response in irradiated animals. Rats

were exposed to acute (2 and 6 Gy) & fractionated (1 Gy/week) doses of

gamma ionizing radiation. Treatment with the aqueous extract orally (5

ml/kg) before and after radiation exposure markedly reduced the exaggerated

paw oedema response to carrageenan. In the acute phase of adjuvant-induced

arthritis, exposure to ionizing radiation caused an increase in serum acid

phosphatase level. Malondialdehyde concentration in plasma and superoxide

dismutase activity in blood significantly increased. Treatment with aqueous

propolis extract prior to irradiation reduced malondialdehyde concentration

in plasma and normalized the serum acid phosphatase level. The extract

stimulated the release of superoxide dismutase enzyme. Aqueous propolis

extract could possibly be of therapeutic value in protecting against

inflammatory responses induced by gamma radiation.

[iv] HASHIMOTO, Takaharu; AGA, Hajime;
TABUCHI, Akihiko; SHIBUYA, Takashi; CHAEN,
Hiroto; FUKUDA, Shigeharu; KURIMOTO, Masashi (1998) (Japan)  –  Anti-Helicobacter
pylori compounds in Brazilian propolis. Natural Medicins 2(),  pp.518-520.

An ethanol extract from Brazilian propolis has antibacterial activity on
Helicobacter pylori, from which two active fractions were obtained by silica
gel column chromatography. From one fraction, an anti-H. pylori compound was
isolated which was identified as 4-hydroxycinnamic acid (p-coumaric acid:1),
and from the other fraction, 3-prenyl-4-hydroxy-cinnamic acid,
3-prenyl-4-dihydrocinnamoloxycinnamic acid, and
3,5-diprenyl-4-hydroxycinnamic acid were isolated, whose anti-H. pylori
activities expressed as the minimal inhibitory concentration (MIC) in mg/ml,
were 15.6-31.3, 62.5, 125, and 250, respectively. 4-Hydroxycinnamic acid
derivatives are likely to be major anti-H. pylori compounds in Brazilian
propolis.

[v] Jaiswal, A. K.; Venugopal, R.; Mucha, J.; Carothers, A. M.;  Grunberger, D. (1997)  –  Caffeic acid phenethyl ester stimulates human antioxidant response element-mediated expression of the NAD(P)H:quinone oxidoreductase (NQO1) gene,
            in Cancer Res, 57(3),  pp.440-46 (***-abstract).

Caffeic acid phenethyl ester (CAPE) is a phenolic antioxidant derived from the propolis of honeybee hives. CAPE was shown to inhibit the formation of intracellular hydrogen peroxide and oxidized bases in DNA of 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated HeLa cells and was also found to induce a redox change that correlated with differential growth effects in transformed cells but not the nontumorigenic parental ones. Mediated via the electrophile or human antioxidant response element (hARE), induction of the expression of NAD(P)H quinone oxidoreductase (NQO1) and glutathione S-transferase Ya subunit genes by certain phenolic antioxidants has been correlated with the chemopreventive properties of these agents. Here, we determined by Northern analysis that CAPE treatment of hepatoma cells stimulates NQO1 gene expression in cultured human hepatoma cells (HepG2), and we characterized the effects of CAPE treatment on the expression of a reporter gene either containing or lacking the hARE or carrying a mutant version of this element in rodent hepatoma (Hepa-1) transfectants. A dose-dependent transactivation of human hARE-mediated chloramphenicol acetyltransferase (cat) gene expression was observed upon treatments of the Hepa-1 transfectants with TPA, a known inducer, as well as with CAPE. The combined treatments resulted in an apparent additive stimulation of the reporter expression. To learn whether this activation of cat gene expression was effected by protein kinase C in CAPE-treated cells, a comparison was made of cat gene activity after addition of calphostin, a protein kinase C inhibitor. Calphostin reduced the cat gene induction by TPA but not by CAPE, suggesting that stimulation of gene expression in this system by these agents proceeds via distinct mechanisms. Band-shift experiments to examine binding of transactivator proteins from nuclear extracts of treated and untreated cells to a hARE DNA probe showed that TPA exposure increased the binding level. In contrast, binding of factors to this probe was inhibited after either in vivo treatment of cells with CAPE or in vitro addition of this compound to the nuclear extract. In view of the clear stimulation by CAPE of gene expression mediated by hARE, possible explanations of this result are discussed.

[vi] Lambert, Priscila (1999) (Brazil)  – Pesquisa mostra que resina produzida pelas abelhas impede o crescimento de células cancerígenas. Própolis pode proteger corpo de tumores,
          in Folha de São Paulo, 12 de Julho de 1999.

Investigación muestra que la resina producida por las abejas impide el crecimiento de células cancerígenas.

El Própoleo puede proteger al cuerpo de tumores.

PRISCILA LAMBERT

de un Reportage Local

El propoleo, aquella resina que las abejas producen para proteger la colmena de invasores, también puede tener propiedades que protegen al cuerpo humano de “invasores”.

            Un estudio realizado en Brasil recientemente demostró que determinados tipos de propoleos impedirían el crecimiento de células cancerígenas en estúdios de laboratorio. La investigación fué realizada por el bioquímico Yong Kun Park, profesor titular de la Facultad de Ingenieria de Alimentos de Unicamp (Universidade Estadual de Campinas).

            El ya había sido director en una tesis que concluyó que ciertos tipos de propoleos inhiben la acción de la bacteria Streptococcus mutans, causadora de carie dentaria (lea el texto en esta página).

            El profesor viene estudiando la actividad del propoleo desde 1990. Fueron analizadas 400 muestras recolectadas en todas las regiones del país.

Muestras

            La primer conclución, despues de un test de cromatografia (una especie de fotografía de los compuestos químicos), fue la de que, dentro de las 400 muestras, 12 eran diferentes entre sí. O sea, existen por lo menos 12 tipos de propoleos en Brasil, que varían de coloración y composición.

            Solamente en la región sur, Park descubrió siete tipos diferentes de resina. “El país tiene una biodiversidad muy rica, lo que explica las variaciones de propoleos. En paises europeos o en EUA, las resinas de las abejas son todas iguales”, dice el profesor. “Cada tipo de propoleo brasilero tiene una propiedad antimicrobiana o antibiótica diferente”.

            En su estúdio, los 12 tipos de propoleos fueron colocados en contacto con diferentes células cancerosas –del intestino, riñon, mamas, nariz y faringe.

Tiempo

            Despues de dos semanas –tiempo sufuciente para que las células se repordujeran y crecieran-, diez muestras habían presentado, en diferentes grados, no solo la inhibición de crecimiento, sino la destrucción parcial de las células.

            La muestra que presentó el mejor resultado fué un tipo de propoleo de Bahia (BA-8), que redujo las celulas de cancer nasofaríngeo en 97% y de mama en 96%, entre otros (vea quadro al lado).

            El método de cálculo de inhibición de tumores utilizado en el estudio tuvo como base de comparación los resultados obtenidos por la droga Etoposide, la más fuerte existente en el mercado para combatír el cancer. Ese método fué desarrollado por el Instituto Nacional del Cancer de los EUA.

            Los resultados presentados en el estúdio de Park fueron confirmados en un test realizado por investigadores norteamericanos de la Universidad de Carolina del Norte (EUA).

            La investigación rindió al profesor Park una invitación para pasar el próximo mes como profesor-visitante de la universidad norteamericana, donde comenzará nuevos estudios, ahora para descubrir cuales son los compuestos químicos presentes en el propoleo y responsables por la acción anticancerígena.

[vii] Lambert, Priscila (1999) (Brazil)  – Própolis puede proteger del sol,
                in Folha de São Paulo, 12 de Julho de 1999.

Outra propiedad atribuída al propoleo es la de filtrar los rayos ultravioletas tipo B.

            La farmacéutica Ana Luíza Mori, responsable por la farmacia universitaria de la Facultad de Ciencias Farmacéuticas de la USP, desarrollo una monografía de maestrado sobre esa acción del propoleo.

            Ella utilizó extractos de propoleos de diferentes regiones del país y del mundo para crear bases cosméticas, que fueron sometidas a un aparato que mide el grado de absorción de rayos ultravioletas.

            Como resultado, Ana Luíza obtuvo factores de protección de los rayos UVB que variaron de 1 (protección casi nula) hasta 8, en la muestra de propoleo de Pensilvania (EUA).

            “La utilización del propoleo como medicamento fitoterápico es complicada, porque la composición de la resina varía de acuerdo com el suelo, la vegetación y la época del año”, dice.

[viii] National Institute of Health, Tokyo, Japan.

[ix] A tumoricidal substance was isolated from Brazilian propolis as guided by cytotoxicity assay on HuH 13 (human hepatocellular carcinoma) cell and was characterized to be 3-[4-hydroxy-3,5-bis (3-methyl-2-butenyl)

phenyl]-2-propenoic acid (3,5-diprenyl-4-hydroxycinnamic acid (artepillin

C)). It exhibited preferential cytotoxicity to tumor cells cultured in

vitro. The cytotoxicity observed seemed to be partly attributable to

apoptosis-like DNA fragmentation. The compound showed anti-tumor activity more effective than that of 5-fluorouracil to transplantable human tumor cell lines when tested on histoculture drug response assay system.

[x] Universidade Estadual de Campinas – UNICAMP, Faculdade de Engenharia de Alimentos, Caixa Postal 6177, CEP 13081 – 970, Campinas – SP, Fone: 55-19-788-7055, Fax: 55-19-788-7890. Email: ykpark@fea.unicamp.br

[xi] We have collected 500 propolis samples which were collected by Africanized Apis mellifera in Southern, Southeastern, Central western, and Northeastern Brazil. The respective samples were extracted with ethanol. The ethanolic extracts of propolis (EEP) were analyzed by physicochemical methods such as appearance of EEP, measurement of absorption spectra by UV-spectrophotometry, reversed phase-thin layer chromatography (RP-HPTLC), reversed phase high performance liquid chromatography (RP-HPLC) and then the EEPs were evaluated their physiological activities such as antioxidant activity, antimicrobial activity, assay of cytotoxic activity to cancer cells, and HIV-growth inhibition assays. In accordance with results of EEP appearance, UV-absorption spectra, RP-HPTLC, and RP-HPLC, the propolis were classified as 12 groups. Among 12 groups of propolis, five groups of propolis were collected from Southern Brazil, whereas six groups of propolis were collected from Northeastern Brazil. Furthermore, one group of propolis was found in Southeastern Brazil. It was found that the variety of propolis is depending on plant ecology. Although rarely exceptional propolis was found in spite of geographic locations, these propolis were not included in this study. Physiological activities for 12 groups of propolis were also variables, depending on geographical location. This is due to fact that the compositions of propolis are depending on the compositions in plants.

[xii] Rao, C. V.; Desai, D.; Rivenson, A.; Simi, B.; Amin, S.; Reddy, B. S. (1995)  –  Chemoprevention of colon carcinogenesis by phenylethyl-3-methylcaffeate,
            in Cancer Res, 55(11),  pp.2310-15 (abstract).

Previous studies from this laboratory have established that caffeic acid esters present in propolis, a natural resin produced by honey bees, are potent inhibitors of human colon adenocarcinoma cell growth, carcinogen-induced biochemical changes, and preneoplastic lesions in the rat colon. The present study was designed to investigate the chemopreventive action of dietary phenylethyl-3-methylcaffeate (PEMC) on azoxymethane-induced colon carcinogenesis and to examine the modulating effect of PEMC on phosphatidylinositol-specific phospholipase C (PI-PLC), phospholipase A2, lipoxygenase (LOX), and cyclooxygenase activities in the colonic mucosa and tumor tissues in male F344 rats. At 5 weeks of age, groups of rats were fed the control (modified AIN-76A) diet, or a diet containing 750 ppm of PEMC. At 7 weeks of age, all animals except those in the vehicle (normal saline)-treated groups were given 2 weekly s.c. injections of azoxymethane at a dose rate of 15 mg/kg body weight/week. All groups were maintained on their respective dietary regimen until the termination of the experiment 52 weeks after the carcinogen treatment. Colonic tumors were evaluated histopathologically. Both colonic mucosa and tumors were analyzed for PI-PLC, phospholipase A2, cyclooxygenase, and LOX activities. The results indicate that dietary administration of PEMC significantly inhibited the incidence and multiplicity of invasive, noninvasive, and total (invasive plus noninvasive) adenocarcinomas of the colon (P < 0.05-0.004). Dietary PEMC also suppressed the colon tumor volume by 43% compared to the control diet. Animals fed the PEMC diet showed significantly decreased activities of colonic mucosal and tumor PI-PLC (about 50%), but PEMC diet had no effect on phospholipase A2. The production of 5(S)-, 8(S)-, 12(S)-, and 15(S)- hydroxyeicosatetraenoic acids via the LOX pathway from arachidonic acid was reduced in colonic mucosa and tumors (30-60%) of animals fed the PEMC diet as compared to control diet. PEMC had no effect on the formation of colonic mucosal cyclooxygenase metabolites but inhibited the formation in colonic tumors by 15-30%. The precise mechanism by which PEMC inhibits colon tumorigenesis remains to be elucidated. It is likely that the chemopreventive action may be related, at least in part, to the modulation of PI-PLC-dependent signal transduction and LOX- mediated arachidonic acid metabolism.

[xiii] Scheller, Stanislaw; Krol, W.; Swiacik, J.; Owczarek, S.; Gabrys, J.; Shani, J. (1989, 1991)  –  Antitumoral property of ethanolic extract of propolis in mice-bearing Ehrlich carcinoma, as compared to bleomycin,
            in Z Naturforsch (C), 11, 44(11-12), 1989,  pp.1063-65 (abstract);
            in Kaal, 1991,  p.27-29 (***).

Antitumoral effect of ethanolic extract of propolis (EEP) was demonstrated in mature mice-bearing Ehrlich carcinoma. Survival rate after EEP treatment was compared to that of bleomycin, given alone or in combination every two days for 36 days and followed up for 14 additional days. The survival rate at 50 days was 55% after EEP and 40% after bleomycin, while all the mice-treated with EEP + bleomycin combination demonstrated shorter survival than the controls. It is concluded that while the in vivo activity of bleomycin is reduced in the presence of cytochrome-C-reductase inhibitors (like some of the EEP components are), the antitumoral property of EEP in the tumored animal model studied is significant and lasting.

[xiv] Su, Z. Z.; Lin, J.; Grunberger, D.; Fisher, P. B. (1994)  –  Growth suppression and toxicity induced by caffeic acid phenethyl ester (CAPE) in type 5 adenovirus-transformed rat embryo cells correlate directly with transformation progression,
            in Cancer Res, 54(7),  pp.1865-70 (abstract).

The active component of the honeybee hive product propolis, caffeic acid phenethyl ester (CAPE), induces a selective growth suppressive and toxic effect toward cloned rat embryo fibroblast cells transformed by adenovirus type 5 (Ad5) or the Ad5 E1A transforming gene versus untransformed cloned rat embryo fibroblast cells (Z-z. Su et al., Mol. Carcinog., 4: 231-242, 1991). The present study was conducted to determine whether CAPE-induced growth suppression/toxicity was a direct result of expression of the Ad5 E1A and E1B transforming genes or a consequence of the action of these genes resulting in the transformed state. For this investigation we used somatic cell hybrids and 5- azacytidine-treated Ad5-transformed rat embryo cells that display different stages of expression of the transformed phenotype. This series of cell lines has permitted us to determine whether expression of the transformed state and the stage of transformation progression regulates CAPE sensitivity. Evidence is presented indicating that sensitivity to CAPE is directly determined by the state of expression of the transformed progression phenotype, as opposed to simply the expression of the Ad5 E1A and E1B transforming genes. These results provide further evidence that CAPE may represent a unique compound that can specifically target progressed transformed cells for growth suppression and toxicity. An understanding of the mechanism underlying this selective effect of CAPE could result in the identification of important biochemical pathways mediating cellular transformation and progression of the transformed state.

[xv] Su, Z. Z.; Lin, J.; Prewett, M.; Goldstein, N. I.; Fisher, P. B.  –  Apoptosis mediates the selective toxicity of caffeic acid phenethyl ester (CAPE) toward oncogene-transformed rat embryo fibroblast cells,
            in Anticancer Res, 15(5B),  pp.1841-8 (abstract).

The active component of the folk medicine propolis, caffeic acid phenethyl ester (CAPE), displays selective toxicity toward cloned rat embryo fibroblast (CREF) cells transformed by a spectrum of diverse acting oncogenes. Identification of the mode of action of CAPE should provide useful information for possible applications of this compound for cancer therapy. The present study uses a series of oncogene transformed, oncogene-reverted and CAPE-resistant oncogene transformed CREF cells to investigate the mechanism underlying the increased sensitivity of transformed cells to CAPE. A direct relationship exists between the cytotoxic effects of CAPE and the induction of DNA fragmentation and apoptosis. DNA degradation into nucleosomal fragments and apoptotic shifts in DNA cell cycle profiles occur in CAPE-treated CREF cells transformed by wild-type 5 adenovirus (Ad5), a mutant Ad5 (H5hr1), the wild-type Ad5 E1A transforming gene, v-src, Ha-ras and the human papilloma virus type 18 transforming genes (HPV-18). In contrast, untransformed CREF cells, human fibroblast expression library-induced morphological revertants of Ad5- and v-src-transformed CREF cells, and Krev-1 expressing revertant Ha-ras-transformed CREF cells are resistant to CAPE-induced toxicity and apoptosis. Similarly, mutant Ad5- transformed CREF cells selected by step-wise growth in increasing concentrations of CAPE are resistant to growth inhibition and apoptosis induced by CAPE. These findings indicate that expression of the transformed phenotype by rodent cells evokes sensitivity to CAPE induced toxicity through apoptosis. The acquisition of CAPE sensitivity in rodent cells is independent of the mode of action of the oncogenic agent. CAPE may prove useful as an antiproliferative agent in cancer cells transformed by mechanistically diverse acting oncogenes.

[xvi] Sud’ina, G. F.; Mirzoeva, O. K.; Pushkareva, M. A.; Korshunova, G. A.; Sumbatyan, N. V.; Varfolomeev, S. D. (1993)  –  Caffeic acid phenethyl ester as a lipoxygenase inhibitor with antioxidant properties,
            in FEBS Lett, 329(1-2),  pp.21-24 (abstract).

Caffeic acid phenethyl ester, an active component of propolis extract, inhibits 5-lipoxygenase in the micromolar concentration range. The inhibition is of an uncompetitive type, i.e. the inhibitor binds to the enzyme-substrate complex but not to the free enzyme. Caffeic acid phenethyl ester also exhibits antioxidant properties. At a concentration of 10 microM, it completely blocks production of reactive oxygen species in human neutrophils and the xanthine/xanthine oxidase system.

[xvii] Yamada, J.; Tomita, Y. (1996)  –  Antimutagenic activity of caffeic acid and related compounds,
            in Biosci. Biotechnol. Biochem., 60(2),  pp.328-29 (abstract).

Effects of caffeic acid and chlorogenic acid on mutagenicity were studied using the Salmonella typhimurium system. These compounds had inhibitory effects on the mutagenicity of Trp-P-1 and Glu-P-2. Caffeic acid completely eliminated the mutagenicity induced by activated Glu-P- 2. Some compounds analogous to caffeic acid, such as cinnamic acid, coumaric acid, and ferulic acid, also significantly decreased the mutagenicity of Glu-P-2.