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Experimental and clinical results with Avemar (a dried extract from fermented weath germ) in animal cancermodels and in cancer patients MICHELE NICHELATTI, M.D., MÁTÉ HIDVÉGI, M.D.
Avemar Research Group, Limbiate (MI), Italy and Biromedicina Co., Budapest INTRODUCTION Relationships between food and health today and some nonnutrient substances like echinacea, saw palmetto, must be studied by taking into account the expanding role of rosemary, cat’s claw, mistletoe, kombucha, shiitake mush- dietary supplements, specialised medical foods and functional rooms, and shark cartilage. Most of these compounds are being foods, collectively named as nutraceuticals. Nonnutrient bio- studied with some relation to cancer; however, the reality is logically active components of foods are receiving increasing that not long after any information is publicized, patients start scientific attention. Health professionals, consumers and self-experimenting with these remedies. They self-dose at a industry are incorporating this new knowledge into their own range of levels, both high and low; with consequences that may practice, behavior and strategies. In most cases, nutraceuticals prove effective, useless, or harmful results. can challenge the standard distinctions existing between foods In the late 1990’s, reports were published about a biotech process by which a fermented wheat germ extract could been The family of dietary supplements includes not only essential produced. The product, called Avemar, available as a water sol- nutrients but also botanical and herbal products, which offer a uble granulate for oral consumption, has gained much attention particular challenge in evaluation of biological activity, active from cancer researchers of several countries, like Israel, constituents, and interactions with conventional medicines.
Hungary, the United States, England and Russia. The reason Medical foods include a somewhat limited category of foods why this extract has got so much dedication from researchers targeted to existing health problems. Functional foods repre- was possibly the fact that it has been produced from one of the sent an emerging category of food products with claims to most common food sources of mankind, and it has shown a offer health benefits. There is a great need for ongoing research good synergism with some anticancer drugs used in standard and documentation regarding the efficacy, safety, and regula- tion of both dietary supplements and the other specialized food products. Health professionals need to actively follow these Wheat kernel contains 2-4% germ (also called embryo), which scientific advances to be credible sources of information for is separated from the endosperm by milling operations like rolling, sieving, etc. In the wheat grain, most nutrients with the exception of starch, are concentrated in the germ. Even though Beyond allowed cancer-related health claims, patients are it is nutritious, wheat germ is mainly used as an animal feed.
today invested by popular press and advertising with a confus- Besides its proteins of high biological value and its oil (show- ing array of remedies found in dietary supplements and bioac- ing a good fatty acid pattern), wheat germ is the richest known tive substances found in foods. Included are specific foods natural source of tocopherols and also abundant in B-group vit- (tomatoes, broccoli, sprouts, chili peppers, yogurt, soybeans), amins. The most significant antinutrients of wheat germ are the drinks (green tea, grapefruit and orange juice), vitamins (C, D, lectin WGA (wheat germ agglutinin) and a group of trypsin E, folic acid and beta-carotene), minerals (selenium, calcium) inhibitors, which can be destroyed by heat treatment. Other remarkable nonnutrients of wheat germ are the bioquinones which are present as glycosides of the corresponding methoxy- hydroquinones (3), whose potential anticancer effects have been firstly investigated in experiemental systems by Nobel 1137 Budapest, Szent István park 18.
Távbeszélô: (36 1) 266 1023 Távmásoló: (36 1) 266 1026 A group of chemists has produced a per os applicable stan- dardized complex of multiple, biologically active molecules Nõgyógyászati Onkológia 2002; 7:40–41 Experimental and clinical results with Avemar (a dried extract from fermented weath germ) in animal cancer models and in cancer patients obtained from the aqueous extract of fermented wheat germ The B16 melanoma was used as muscle-lung metastasis (5). The standardized extract – named as Avemar – has been model, while the C38 mouse colorectal carcinoma cell line was registered and is now marketed as an over-the-counter dietary applied for serving as spleen-liver metastasis model. Mice supplement in various countries, like Hungary, Israel, Italy, bearing the C38 colorectal carcinoma implanted into the spleen Austria, Slovakia, Czech, Cyprus and Switzerland. It is there- were treated with 5-FU administered via intraperitoneal injec- fore to point out that Avemar is not a drug, nor an alternative tion 3 times a week in a dosage of 1 mg/kg, while the mice to standard anticancer drugs or standard therapies: Avemar is a inoculated with the B16 melanoma received DTIC treatment dietary supplement to be given to cancer patients to help drugs daily (60 mg/kg i.p.) Synchronously, the animals treated with antineoplastic agents also received Avemar daily (3 g/kg). In the case of combined (Avemar + DTIC) treatment the number As any natural product, Avemar exerts several biological of lung metastases of B16 melanoma practically decreased to effects, which can be theoretically explained, according to zero, and this effect was significant. The results show that in some possible metabolic modeling (6-7). In general, the bio- therapeutic composition, Avemar – having metastasis inhibito- logical activity of Avemar can be divided in effects useful for ry effect also alone – exerted a more than additive effect, that the treatment of neoplastic diseases (8-16) and effects which is, it synergically enhanced the metastasis inhibitory effect of can be well used in the treatment of certain immune distur- DTIC used in clinical practice to decrease metastasis in proto- bances (17-19). Avemar even improves the patients’ quality of cols for treatment of patients with melanoma. Treatment of life which latter effects can be independent from the previous C38 colorectal carcinoma with the therapeutic composition of Avemar and 5-FU decreased the number of liver metastases synergically. This effect was also significant. The mass of the ANIMAL EXPERIMENTS: SINGLE USE OF AVEMAR In all experi- diseased spleen also displayed a marked decrease as a conse- ments, 8-10 week-old inbred mice of 20-22 g body weight were used. The following transplantable tumor lines, grown on mice or rats, were used in the experiments: a highly metastatic Although the therapeutic effects at both of the combination variant of Lewis lung carcinoma (3LL-HH), B16 mouse experiments were considerable, the usual toxic side effects of melanoma, C38 mouse colorectal tumor and HCR-25, a human cytostatics, e.g. decrease of body mass were not observed. It colon carcinoma xenograft (17). In all experiments, Avemar can be concluded that Avemar treatment does not reduce the treatment was started 24 hours after tumor implantation.
antitumoral effects of chemotherapeutic drugs upon the prima- Avemar was dissolved in water and administered by means of ry tumors but, dramatically enhances their antimetastatic a gastric tube. The daily dose was 3 g/kg body weight per os effects. Using several other cytostatics (data not shown) it was administered in 0.1 ml of water. Control animals received tap also proved that Avemar did not reduce their cytostatic effects water daily (0.1 mL), also via gastric tube.
Avemar treatment resulted in a statistically significant 71% CHEMOPREVENTIVE EFFECTS OF AVEMAR It has been demonstrat- decrease in the number of liver metastases of the 3LL-HH ed that Avemar treatment prevents colon cancer in laboratory tumor inoculated into the spleen (17). In case of the HCR-25 animals; in this case, four weeks old inbred male F-344 rats human colon carcinoma, the 50 days of Avemar treatment were used (9). Colon carcinogenesis has been induced by decreased the amount of liver metastases, in addition to reduc- injections of azoxymethane (AOM), a well-known carcino- ing the weight of the tumorous spleen. The number of metas- genic chemical. Ten rats served as untreated controls (group 1).
tases in the Avemar-treated animals as compared to the control For the treatment of the animals in group 2, AOM was dis- group was around 50% (17). In case of the B16 melanoma solved in physiologic saline and the animals were given 3 sub- inoculated into the muscle, also a significant decrease of 85% cutaneous injections 1 week apart, 15 mg/kg body weight was observed in the number of metastases as compared to the (BW) each. In two additional groups the basal diet was sup- plemented with Avemar. The extract was dissolved in water and was given at a dose of 3 g/kg BW once a day. In group 3, ANIMAL EXPERIMENTS: COMBINED USE OF AVEMAR AND CYTO- animals started to receive Avemar two weeks prior to the first STATICS In these experiments the B16 mouse melanoma and injection of AOM daily and continuously thereafter until sacri- C38 mouse colorectal tumor strains were used. The aim ficed 32 weeks later. In group 4 the basal diet was supple- of these experiments was to find out how the daily treat- mented by Avemar administration only. At the end of the ment with Avemar (3 g/kg body weight) would influence the experiment all the rats were sacrificed by exsanguination, the tumor growth and metastasis inhibiting effect of treatment abdominal large vessels were cut under a light ether anaesthe- with some of the well known antineoplastic agents 5-Fluo- sia and a complete autopsy was performed. The percentage of rouracil (5-FU) and Dacarbazine (DTIC), which are widely animals developing colon tumors and the number of tumors used in clinical oncology in the frame of various treatment per animals were: 0 and 0 (group 1); 83.0 and 2.3 (group 2); 44.8 (p < 0.001) and 1.3 (p < 0.004) (group 3); 0 and 0 (group Nõgyógyászati Onkológia 2002; 7:40–41 4); all the tumors resulted of neoplastic nature also at histolog- the probability that survival time Tis greater than a given time ical inspection. Thus, the overall chemopreventive effect of Avemar (see also Table 1) was neraly 70%, as one obtains from where f(u) and respectively are the probability density and the cumulative probability of T; obviously, the survivor function is The numbers of the aberrant crypt foci (ACF) per area (mea- very sensitive to the shape of the probability density. In this sured in cm2) was 4.85 in group 2, while in group 3 the num- model, the survivor function must be estimated, assuming that ber of ACF numbers was 2.03 only (p <0.0001). its value is constant between two consecutive events, thus the plot of S(t) versus time is represented by a stepwise decreasing Table 1. Macroscopic findings in the large intestine of F-344 rats treated with Avemar or with Avemar + AOM; statistical significance was: p <0.001 (*); p<0.004 (**) If all the observed individuals are followed up until the event occurs to each of them, the Sˆ (t) value, estimating the true S(t), may be evolved from the ratio S(t) = N(T ³ t) / N(0) , where N(T ³ t) is the number of subjects surviving at time T ³ t , and N(0) is the number of subject originally enrolled in the trial. In the case of censored data (like in this second trial), however, this simple calculation cannot be done, and the esti- mated Sˆ (t) value must be evaluated by some other methods.
One of the most used is the Kaplan-Meier product limit esti- mator, which is obtained with the formula where rk is the number of subjects at risk (including censored CLINICAL STUDIES: NEW METASTASES AND PROGRESSION-FREE subjects) at time immediately preceding tk, and dk is the num- SURVIVAL IN CANCER PATIENTS An early open-label phase II ber of subjects experiencing the event at time tk.
clinical trial with Avemar was conducted in colorectal cancer patients, involving 30 consecutive subjects undergoing cura- Survival analysis allows the assessment of the periods where a tive surgery, accrued since 1998 up to June 1999 (20). Patients given clinical event of interest (death, or any disease progres- were divided into control cohort (n = 18, 11 men and 7 women sion event like a new metastasis, a relapse, or the death itself) with mean age of 70 years) and Avemar cohort (n = 12, 6 men has the highest and the lowest chance. For this purpose, it is and 6 women with mean age of 64 years) according to their used the hazard function h(t) defined by the relationship own preference. Patients of the control group received adju- vant chemotherapy alone (if necessary), whereas patients of the Avemar group received adjuvant chemotherapy (if neces- sary) plus 9 g of Avemar once or twice daily, depending on from which we easily obtain the survivor function in terms of their body weight. The median follow-up of all patients was 9 At the end of the study, no patients treated with Avemar did show new metastases, neither hepatic, nor in other organs, The survival analysis uses its own regression models. In gen- while 4 patients (22%) did develop new metastases in the con- eral, its multiplicative factor must be assumed costant, so that the hazards in the studied cohorts must be proportional. In this case, one is dealing with proportional hazards regression, with This first clinical result was so encouraging that it was decid- hazard ratio constant over time, and different individuals have ed to evaluate the impact of Avemar in a second trial involving proportional hazards, so that, if the covariate row vector of more patients, and comparing the disease progression-free sur- subject A is, say, xA = (xA1, xA2, K, xAn), and the covariate row vival as well as the overall survival in two groups of colorec- vector of subject B is xB = (xB1, xB2, K, xBn), then the ratio tal patients differing just for the Avemar intake. In the survival h(t | NA) / h(t | NB) must not change with time along all the analysis trial done with Avemar, as well as in all similar trials, study period. Under this assumption (to be verified at time of the absolute survival and the disease progression-free survival data analysis), the hazard function could be written as are normally assessed by a survivor function S(t), defined as Nõgyógyászati Onkológia 2002; 7:40–41 Experimental and clinical results with Avemar (a dried extract from fermented weath germ) in animal cancer models and in cancer patients is a relative risk function of the vector of covariates alone.
alone, and therefore, to obtain information on the feasibility of Thus, since the hazard ratio between individuals A and B must long term administration of Avemar as well as to estimate the expected difference of treatment outcome between cohorts of colorectal cancer patients receiving standard treatment and standard treatment plus Avemar supplementation. For the tri- als, the chosen values for sample size calculus were b = 0.05 In the case of exponential relative risk, the effect on a log-lin- (i.e., 5%) and 1–b = 0.9 (i.e., the power was 90%), so that a ear scale is additive, and the baseline hazard function is multi- minimal sample size of 50 patients was needed for each cohort.
plied by the covariate vector: for this reason, each individual accrued in the trial shows an hazard function of the form: Beyond the standard oncological treatment used in both groups, the patients assigned to the Avemar cohort did take 9 gramms of Avemar per os once or twice daily, along all the If one models parametrically only the relative r isk, as pro- study period, for which the minimal follow-up was at least 6 posed by Cox (21-22), then the shape of baseline hazard may months. The treatment time period was measured as the inter- be left unspecified, and a semiparametric model can be con- val between the time 0 (baseline) and the last completed visit.
structed, allowing to estimate b from a partial likelihood func- Patients of the control cohort received the standard oncological tion which takes into account ties among survival times and treatment alone, consisting of 5-fluorouracil (5-FU) based chemotherapy and/or radiation therapy, following surgery. All patients were evaluated at baseline, after one month, and then every 12 weeks. Evaluation included imaging quantification of all measurable lesions (by usual radiographic, ultrasonic, or magnetic resonance techniques), laboratory tests (hematology, chemistry, and urinalysis), physical examination, as well as where s is the vector sum of the covariates of the m individu- data regarding treatment compliance and toxicity. Tumor pro- gression was defined as an increase of at least 25 percent in the overall tumor size or the appearance of any new lesions; deaths To analyse the influence of Avemar (added to surgery and stan- were also recorded. All the time-related events were measured dard radiotherapy and/or chemotherapy) on the disease pro- gression-free survival (disease progression events were defined as deaths, relapses and new metastases occurring in The primary end-point of this study was to compare progres- both cohorts) and on overal survival (deaths only) in colorec- sion-free survivals of the two cohorts. For this purpose, it was tal cancer patients, an open-label comparative cohort trial has used the two-tailed, unstratified log-rank tests (Kaplan-Meier method), while for other comparisons, z-test, Mann-Whitney’s test, Fisher’s exact test and Student’s t-test were applied as For the analysis of the effects exerted by different variables (disease staging, Avemar administration, age, sex, chemother- apy, radiotherapy) on survival, the Cox regression (proportion- The multicenter trial started in November 1998 and patient al hazards model) was used, after verifying that this method recruitment lasted up to March 2001, so that 170 consecutive was suitable according to the study data, by means of the colorectal cancer subjects entered the study (24), to be includ- Schoenfeld residuals (23) of the general form: ed in the Avemar or in the control cohorts according to the patient’s own decison. The patients had either new diagnosis of their cancer or arrived for routine check-up of their previously The age of the patients of the control cohort was significantly for each covariate x , such that r is the existing difference higher than that of the Avemar one (mean was 66.1 years in the between the covariate value for any failed j-th observation and controls versus 61.7 years in the Avemar cohort; p <0.01). In the average value of the covariate, which is weighted on the contrary, the Avemar patients had significantly more advanced basis of estimated hazards from Cox model. The residual disease stages (Mann-Whitney probe: z = 4.618; p <0.001), analysis for this trial has shown no evidence of violations of since 27.3 per cent of the Avemar patients were at UICC stage the assumptions at the basis of Cox proportional hazards IV (metastatic), while this value for the control patients was 3.8 % only (p <0.001); moreover, the average time from diag- nosis to the study enrolment was significantly longer for the The goal was to determine if the use of Avemar adds any ther- Avemar cohort (11.2 months and 1.1 months, respectively, p apeutic benefit compared to standard therapeutical protocols <0.001). There were no significant differences between the Nõgyógyászati Onkológia 2002; 7:40–41 average length of time from diagnosis to the last visit (29.6 dictors. Interestingly, similarly to the previously observed months and 34.0 months, respectively; Student’s t = 1.494; p = nearly 70% preventing effect exerted by Avemar in rat colon 0.137), nor significant difference between the number of carcinogenesis model (9), in this last clinical trial Avemar patients receiving chemotherapy (z = 1.819; p = 0.069), while increased the probability of survival still by nearly seventy the controls did receive significantly more radiotherapy (z = percent (see the exp(b) value in Table 3), since one obtains 1 3.406; p <0.001). Generally, the prognoses of the Avemar patients at baseline were poorer than those of the control Table 2. Occurrence of progression-related events 1. Ashley JM, Harrison G. Dietary supplements, medical foods, and func- tional foods. In: Nutritional oncology. Heber D, Blackburn GL, Go VLW, eds. Academic Press, New York, 1999: 371-378. 2. Bonney RC. Nutraceuticals and functional foods – A new market for the pharmaceutical industry. Scrip Reports BS 1017, PJB Publications Ltd, 3. Cosgrove DJ, Daniels DGH, Greer EN, Hutchinson JB, Moran T, Whitehead JK. Isolation of methoxy- and 2:6-dimethoxy-p-benzoquinone from fermented wheat germ. Nature 1952; 169:966-967.
At end-point analysis, observed progression-related events 4. Szent-Györgyi A. Metabolism and cancer. Int J Quant Chem Quant Biol (relapsed tumors, new metastatic lesions, deaths) were signifi- cantly more abundant in the control cohort (Table 2). The log- 5. Tömösközi-Farkas R, Rásó E, Lapis K, Szende B, Paku S, Hidvégi M.
rank test showed significant differences in favor of the Avemar Avemar: Fermentation product of wheat germ with antimetastatic effect.
patients, in both the cumulative probabilities of disease pro- In: Functional Foods – A new challenge for the food chemists. Vol. 2.
Lásztity R, Pfannhauser W, Simon-Sarkadi L, Tömösközi S, eds.
gression-free survival (primary endpoint) and overall sur- Federation of European Chemical Societies, Food Chemistry Division, vivals. Among all analysed covariates (age, sex, UICC staging, Avemar treatment, radiotherapy and chemotherapy), the only 6. Hidvégi M. Results of the research on the mode of action of Avemar. (In strong predictors of survival in the Cox proportional hazards Hungarian). Nõgyógy Onkol 1998; 3:241-243.
model were UICC stage and Avemar treatment (Table 3). 7. Cascante M, Boros LG, Comin-Anduix B, Atauri P, Centelles JJ, Lee W- NP. Metabolic control analysis in drug discovery and disease. Nat Table 3. Multivariate analysis of survival of colorectal patients (Cox regres- sion), proportional hazards model: c2 = 22.756; p = 0.0009. Among all the 8. Hidvégi M, Rásó E, Tömösközi-Farkas R, Paku S, Lapis K, Szende B.
six tested variables, only the strong predictors (with significant p value) are Effect of Avemar and Avemar + vitamin C on tumor growth and metasta- sis in experimental animals. Anticancer Res 1998; 18:2353-2358.
9. Zalatnai A, Lapis K, Szende B, Rásó E, Telekes A, Resetár Á, Hidvégi M. Wheat germ extract inhibits experimental colon carcinogenesis in F- 344 rats. Carcinogenesis 2001; 22:1649-1652.
10. Fajka-Boja R, Hidvégi M, Ion G, Székely Szûcs K, Demidenko D, Monostori É (2000). Avemar triggers apoptosis and downregulation of major histocompatibility complex class 1 in leukocyte tumor cells. 1st The treatment with Avemar was generally safe (no serious Congress of the Hungarian Society of Clinical Oncology. Budapest, adverse events were recorded), and the compliance to protocol was good: practically, the only complaint reported by Avemar 11. Boros LG, Lee W-NP, Hidvégi M, Go VLW (2000). Metabolic effects of fermented wheat germ extract with anti-tumor properties in cultured MIA pancreatic adenocarcinoma cells. Combined Meeting of the International Association of Pancreatology and the American Pancreatic The results generally showed highly significant data in favor Association. Chicago, Illinois, USA, 1 – 5 November.
of Avemar treatment: that was somewhat surprising but not 12. Boros LG, Lapis K, Szende B, Tömösközi-Farkas R, Balogh Á, Boren entirely unexpected; rather, the main results confirmed what J, et al. Wheat germ extract decreases glucose uptake and RNA ribose for- previously seen in the phase II trial (20). It could be conclud- mation but increases fatty acid synthesis in MIA pancreatic adenocarcino- ma cells. Pancreas 2001; 23:141-147.
ed that the present study brought the first evidences that this wheat extract, in combination with surgery plus standard 13. Ribári O, Almay K, Hoffmann A, Hidvégi M. Early results on the sup- radio/chemotherapy, can significantly inhibit overall tumor portive treatment of head and neck cancer patients with Avemar. 1st Congress of the Hungarian Society of Clinical Oncology. Budapest, progression including the formation of new metastases, and could prolong the survival of colorectal cancer patients. The Cox analysis identified UICC staging and the Avemar treat- 14. Szende B, Rásó E, Hidvégi M, Tömösközi-Farkas R, Paku S, Prónai L, et al. A new, substituted benzoquinone-containing natural product with ment (for more than 6 months) as independent survival pre- antimetastatic effect. (In Hungarian). Orv Hetil 1998; 139:2893-2897.
Nõgyógyászati Onkológia 2002; 7:40–41 Experimental and clinical results with Avemar (a dried extract from fermented weath germ) in animal cancer models and in cancer patients 15. Fajka-Boja R, Hidvégi M, Shoenfeld Y, Ion G, Demydenko D, Th2 response in experimental SLE and promotes amelioration of the dis- Tömösközi-Farkas R, et al. Fermented wheat germ extract induces apop- tosis and downregulation of major histocompatibility complex class I pro- teins in tumor Tand B cell lines. Int J Oncol 2002; 20:563-570.
20. Jakab F, Mayer Á, Hoffmann A, Hidvégi M. First clinical data of a nat- ural immunomodulator in colorectal cancer. Hepatogastroenterology 2000; 16. Hidvégi M, Rásó E, Tömösközi-Farkas R, Szende B, Paku S, Prónai L, et al. MSC, a new benzoquinone-containing natural product with antimetastatic effect. Cancer Biother Radiopharm 1999; 14:277-289.
21. Cox DR. Partial likelyhood. Biometrika 1975; 62:269-276.
17. Hidvégi M, Rásó E, Tömösközi-Farkas R, Lapis K, Szende B. Effect of MSC on the immune response of mice. Immunopharmacology 1999; 22. Cox DR. Regression models and life tables. J Roy Statist Soc Ser B 18. Falkay Gy, Blazsó G. Antiinflammatory effect of Avemar. Scientific 23. Schoenfeld D. Partial residuals for the proportional hazards regression meeting of the Albert Szent-Györgyi Medical and Pharmaceutical Center of the University of Szeged. Szeged, Hungary, 14 November, 2000.
24. Jakab F, Shoenfeld Y, Balogh Á, Nichelatti M, Telekes A, Hoffmann A, 19. Ehrenfeld M, Blank M, Shoenfeld Y, Hidvégi M. Avemar (a new ben- et al. Anticancer activity of a wheat germ derived nutraceutical in colorec- zoquinone-containing natural product) administration interferes with the tal cancer patients. Am J Clin Oncol (submitted).
Nõgyógyászati Onkológia 2002; 7:40–41


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