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Dr. Ursula Jacob Medical director USA 2012 Sonntag, 7. Mai 2017 • The clinic has 17 rooms • Over 7,000 patients treated, from Europe, Australia and North America. • Staff of 4 doctors, 12 nurses, nutritionist and psychologist Indications • • • • • • • preventive diagnostics and treatments regenerative treatments hematological diseases metabolic disorders chronic neurological diseases chronic viral diseases oncological diseases Diagnostic spectrum: • electrocardiogram at rest • sonography • bone marrow cytology with possibility of fast diagnosis • biopsies under appropriate control Diagnostic spectrum in cooperation: • • • • • • • • • • • • • • CST chemo sensitivity test (testing of chemotherapeutics, natural substances, antibodies and tumor genes) CTC test (checking for circulating tumor cells) CFS test (checking for autoimmune-/viral disease) X-ray, CT and MRT pulmonary function test exercise electrocardiogram bronchoscopy gastroscopy coloscopy rectoscopy PET (positron-emission-tomography) scintigraphy laboratory diagnostics pathology Therapeutic spectrum: • • • • • • • hyperthermia immunological treatment naturopathic treatments psycho-oncology nutritional therapies individually tested chemotherapy / immune therapy individual preventive therapy (based on gene analysis) • vaccination Therapeutic spectrum in cooperation: • • • • • • • surgical interventions radiation therapy venous port implantations embolisation perfusion therapy laser therapy cyberknife Micro tumor testing (CTC)chemosensitivity Assays and R&D CFS -Testings AUTHOR-PRESENTER: DR. Ursula Jacob R.G.C.C s Services • Clinical Services 1. 2. Chemosensitivity testing Detection , quantification and immunophenotyping CTCs (MRD test) • Research activities 1. 2. 3. Evaluation of substances candidates for drugs “whanabe”. Detection of new targets for new therapeutic approaches Basic research in molecular Oncology Cancer Hallmarks CARCINOGENESIS STEPS INITIATION •Viral interference •Chemical interference •Radiation influence PROMOTION •Cell cycle instability •DNA repair aberration •Apoptosis instabiliity PROGRESS •Invasion •Neo-angiogenesis •EMT and MET VOGELSTEIN MODEL OF DEVELOPING COLON CANCER Cancer Hallmarks Novel techniques • FEATURES OF NOVEL ASSAYS-METHODS 1. 2. 3. 4. 5. 6. 7. 8. TECHNIQUES Accuracy •Flow Cytometry Sensitivity Specificity •Quantitative PCR Cellular profile •Micro-arrays Cellular abilities and potentials •Protein arrays Detection of heterogeneity Detection of subpopulation Detection and isolation of MRD cells Flow cytometry 1. 2. 3. 4. 5. METHODOLOGY Single cell analysis. Information about size and granularity for each cell. Information through fluorescence about any antigen we wish to analyze (in theory). Information about DNA quantity and quality. Viability and apoptosis detection. THE MORE PARAMETERS YOU CAN ANALYZE THE MORE INFORMATION YOU RECEIVE AND THE DISCRIMINATION ABILITY RISES. Quantitative PCR • THE METHODOLODY OF qPCR 1. 2. 3. ISOLATION OF mRNA FROM ALL CELLS PRODUCTION OF cDNA FROM mRNA USING RT-PCR. USING SPECIFIC PROBE FOR EACH GENE THAT WE ARE INTERESTED IN (WITH FLUOROPHORE AND QUENCER) IN EACH CYCLE THE PROBE IS RELEASED AND THE QUENCER IS SEPARATED FROM THE FLUOROPHORE AND ITS BLEACHES. VISUALISATION AND QUANTIFICATION OF FLUORESCENCE 4. 5. • DATA ANALYSIS 1. THE CYCLE THRESHOLD REFLECT THE PRIMARY ORIGINAL AMOUNT OF TEMPLATE mRNA . THE LEVEL OF FLUORESCENCE REFLECT THE NUMBER OF AMPLIFICATIONS THAT cDNA HAS RECEIVED IT IS POSSIBLE EITHER TO PERFORM RELATIVE QUANTIFICATION OF A GENE IN COMPARE WITH AN ENDOGENOUS ONE (HOUSEKEEPING OR ABSOLUT QUNATIFICATION BASED ON 2. 3. Micro-Arrays analysis • THE METHODOLODY OF MICRO-ARRAY 1. 2. 5. ISOLATION OF mRNA FROM CANCER CELLS PRODUCTION OF cDNA FROM mRNA USING cDNA SYNTHESIS (first and second strand). MODIFICATION OF cDNA WITH BIOTIN-STREPTAVIDIN OR FLUORESCENT DYES. HYBRIDIZATION OF cDNA WITH SPECIFIC PROBES FOR GENES VISUALISATION OF HYBRIDIZED PROBES • GENES THAT ARE ANALYSED 1. GENES THAT BECOME TARGETS FOR CYTOSTATICS (DHFR,TS, TOPOI, TOPOII etc) GENES THAT PRODUCE GROWTH FACTORS OR GROWTH FACTOR RECEPTORS (EGF, IGF , TGF etc) GENES THAT ARE INVOLVED TO METASTASIS (VEGF, EGF, PDGF ETC) GENES THAT REGULATE CELL CYCLE (p53, p27, p16 etc) GENES THAT ARE INVOLVED TO RESISTANCE PHENOTYPE (MDR1, DNMT1,BCL, NP, NFK, HTERT etc) 3. 4. 2. 3. 4. 5. The role of SNPs in drug metabolism • The majority of drugs need to transformed into active formulations by several enzymatic transformations • Most of the active substances needed to deactivated in order to be removed from the organism. • SNPs can cause either accumulate toxicities due slow removal of an active drug (accumulators) or • By specific SNPs drugs will never sustain their active forms in order to act (rapid metabolizers) Chemosensitivity molecular assays may proved powerful tool of prediction of adverse reaction and drug side effects Chemosensitivity and targeted therapy • • • • • • Recently monoclonal antobodies or biomolecules used as inhibitors or modifiers of a target antigen (CD20, Her/neu 2, CD33, CD52, EGF-r, VEGFRII, Bcr-Abl etc). Only molecular techniques can detect and quantify such molecules (flow cytometry, IHC, qPCR, hybridization assay). Mutation of such target molecules may induce resistant phenotype of cancer cells. Down regulation of those antigens may also induce resistance Antigen depletion may stress and promote more resistant subclones if there is no control. Flip-flop of membrane antigen may hide targets for biological drugs Molecular techniques may control and personalize the therapy Chemo-sensitivity / resistance Testing and targeted treatments in cancer Our clinical experience ONCOTRACE ONCOTRAIL ONCOCOUNT TU profile + What is Artemisinin? • Artemisinin is a sesquiterpene lactone isolated from the plant Artemesia annua L. (has been used for the treatment of malaria). • Dr. Zhenxing Wei was first to isolate artemisinin in 1970. • The artemisinin molecule contains an endoperoxide bridge that reacts with a ferrous iron to form free radicals • There are several analogs of artemisinin including artesunate and artemether. How does Artemisinin work? • Artemisinin causes the cancer cell to commit suicide. • The artemisinin molecule contains an endoperoxide bridge that reacts with a ferrous iron atom to form free radicals • Generation of free radicals leads to macromolecular damages and cell death. • Cancer cells have a very high iron uptake and thus they are more susceptible. Research on Artemisinin: MOLT-4 (Leukemia Cell Line) Studies • First study on artemisinin (1995) was done in cell culture (MOLT-4 lymphoblastoid leukemia cell line). • Results show all MOLT-4 cells were killed in 8 hours by 200 micromolar of dihydroartemisinin. • The drug is 100 times less toxic to human lymphocytes in culture. Research on Artemisinin: Leukemia Cell Line APOPTOSIS IN MOLT-4 LEUKEMIA CELLS (dihydro) Artemisinin (8 hours only) 0% Control 100 % 200 M Hyperthermia (24 hr incubation) 3.26 % Control 5.01 % 44C for 1 hr Hydrogen Peroxide (24 hr incubation) 3.52 % Control 40.09 % 176 M Mitoxantrone (24 hr incubation) 3.51 % Control 55.02 % 0.5 M Novobiocin (24 hr incubation) 3.75 % Control 22.68 % 800 M Sodium Ascorbate (24 hr incubation) 3.47 % Control 62.59 % 2000 M X-ray (24 hr incubation) 3.2 % Control 9.5 % 100 rads Research on Artemisinin: Trials in Dogs • Dog trials were begun soon after encouraging results in MOLT-4 experiments (1994-1995). • Dogs of different breeds (male and female) having various types of cancers (lymphosarcoma, breast adenocarcinoma, osteosarcoma, ETC) were treated. • Results: Specific results varied with dogs, but generally positive. Tumor sizes were drastically reduced. No reoccurrence of cancer in 5 dogs operated and given artemisinin. Research on Artemisinin: Human Breast Cancer Cells in vitro • Most recent research was published (2001) on a breast cancer cell line (HB 27) in vitro. • Breast cancer cells treated with dihydroartemisinin and holotransferrin were almost completely eliminated (after 16 hrs of treatment cell count was only 2% of that at time zero). Research on Artemisinin: Human Breast Cancer Cells in vitro • A morphological examination of breast cancer cells treated with dihydroartemisinin and holotransferrin showed that they were undergoing apoptosis and necrosis. • Drug had no effect on normal breast cells. FIGURE 1 Breast Cancer Cells in vitro undergo rapid and almost complete cell death (98%) after treatment with dihydro-artemisinin and holotransferrin. 100 50 0 0 4 8 12 16 12 16 Time (hr) Percent of cell count from time zero Research on Artemisinin: Percent of cell count from time zero Breast cancer cells (HTB 27) 150 Normal breast cells (HTB125) 150 100 50 0 0 4 8 Time (hr) Control holotrans ferrin dihydroartemisinin dihydroartemisinin + holotrans ferrin FIGURE 2 Research on Artemisinin: 150 24 hrs after replating 8 hrs and replating 200 time zero Perecnt of cell count from time zero Breast cancer cells were completely non-viable after 8 hours of treatment with holotransferrin and dihydroartemisinin, as proved by replating. BREAST CANCER CELLS (HTB 27) holotransferrin 100 dihydroartemisinin + holotransferrin 50 0 24 hrs after replating 8 hrs and replating 150 time zero Normal breast cell counts slightly decreased with the same treatment, suggesting some damage to cells. Percent of cell count from time zero NORMAL BREAST CELLS (HTB 125) 100 holotransferrin 50 0 dihydroartemisinin + holotransferrin Principles of Artemisinin Therapy: How to kill cancer cells • Starvation by depletion of nutrients • Exercise by generating H2O2 • Drugs including vitamin C, vitamin D and artemisinin and analogs • Alkaline pH in body Benefits of Exercise For killing of Cancer Cells: • • • • Generates Hydrogen Peroxide. Results in high concentrations of oxygen in the body. With the help of vitamin D, puts calcium in bones. Increases circulation allowing immune cells to reach cancer General Benefits: • Feeling of well-being (increased appetite) • Increased excretory processes • Raises Pain Threshold Conclusion • Artemisinin can be used to treat various types of cancer. • Side effects are minimal and it can be taken orally. Catumaxomab Mode of Action Trifunctional antibodies: Proof of principle 1. Activation of T cells • Methods: – – • Co-culture of tumour cell lines (EpCAM-positive and EpCAM-negative) with peripheral blood mononuclear cells (PBMCs) Incubated with trifunctional antibody (trAb; catumaxomab) for up to 10 days prior to immunohistochemical analysis Results: – – – – – – – After addition of trAb, EpCAM-positive cells had altered morphology and were surrounded by PBMCs After 72 hours no EpCAM-positive tumour cells could be detected Clustering of PBMCs occurred in all cultures containing trAb In cultures without trAb, no clusters of PBMCs were seen Clusters consisted predominantly of CD3+ T cells EpCAM-negative tumour cells were unaffected, however clustering of PBMCs was seen No difference in cell behaviour between PBMCs from healthy donors or those with prostate cancer EpCAM=Epithelial Adhesion Molecule; PBMC=peripheral blood mononuclear cell; trAb=trifunctional antibody. Riesenberg, et al. J Histochem Cytochem 2001; 49:911. Trifunctional antibodies: Proof of principle 1. Activation of T cells • Activation of immune effector cells (CD3+ T cells) • Results: – Increased proliferation of lymphocytes – Cluster consisted predominantly of CD3+ T cells Trifunctional antibodies: Proof of principle 1. Activation of T cells • Expression of lytic proteins specific for cytotoxic T lymphocytes (CTLs) – trAb-mediated release of perforine • Results: – Lymphocytes released perforine • Cytotoxic T cell – Evidence that tumour cells are killed by cytotoxic T cells via necrotic mode (via osmotic lysis) rather than apoptosis Trifunctional antibodies: Proof of principle 1. Activation of T cells (cont’d) • Activation of T cells and expression of lytic proteins ELISPOT (IFNg) • Methods: – Coculture of the tumour cell line BHY with CD8+ cytotoxic T lymphocytes (CTLs) – Incubated with or without trAb BHY CD8+ – ELISPOT analysis CD8+ trAb CD8+ BHY CD8+ BHY trAb ELISPOT (Granzyme B) • Results: – CTL activation indicated by IFNg – Cytotoxic activity mediated by granzyme B – Full induction of IFNg and granzyme B depends on the presence of EpCAM+ tumour cells IFNγ=interferon-γ. Schmitt M, et al. Int J Oncol 2004; 25:841–848. BHY CD8+ CD8+ trAb CD8+ BHY CD8+ BHY trAb Trifunctional antibodies: Proof of principle 2. Activation of dendritic cells • Tumour-cell killing mediated by monocytes/dendritic cells • Methods: – Coculture of PCI-1 (EpCAM+ squamous-cell carcinoma) cells with CD64+ monocytes and DCs, in the absence of T cells – Incubation with +/ - BiUII (anti-EpCAM x antiCD3)* – Lysis measured by cell viability assay (MTT) • Results – In co-culture with monocytes and DCs, tumour cell lysis depends on presence of trAb – Tumour cell lysis is most likely due to trAb-mediated ADCC *BiUII and catumaxomab originate from different parental anti-EpCAM antibodies, but recognize the same epitope region on human EpCAM. Zeidler R et al. J Immunol 1999;163:1246–1252. Trifunctional antibodies: Proof of principle 3. Lytic capacity • Tumour cell killing with BiUII compared with the parental antibodies (Abs) • Methods: – PBMCs were incubated with PCI-1 – Incubation with +/- BiUII (anti-EpCAM x anti-CD3) trAb or the monoclonal parental Abs (αCD3 and αEpCAM) – Lysis measured by cell viability assay (MTT) • Results – BiUII displays a much higher lytic activity compared with parental Abs Zeidler R et al. J Immunol 1999;163:1246–1252. Tumour cell lysis Trifunctional antibodies: Proof of principle 4. Activation of monocytes/macrophages • Tumour cell killing by monocytes/macrophages (phagocytosis) • Methods – Coculture of FITC-labelled PCI-1 cells with CD64+ monocytes – Incubation with +/- BiUII (anti-EpCAM x anti-CD3) trAb for 15h – Monocytes analysed by flow cytometry – Presence of FITC+ monocytes indicates phagocytosis of PCI cells • Results – Phagocytosis induced by BiUII – Activation of macrophages indicated by expression of neopterin and biopterin (not shown) – Activation of DCs indicated by upregulation of DC-CK1, CD83 and CD86 (co-stimulatory molecules) Zeidler R, et al. Br J Cancer 2000;83:261–266. Trifunctional antibodies: Proof of principle 5. Activation of natural killer cells • Binding and activation of NK cells resulting in tumour cell lysis by NK cells (ADCC) • Methods: – Highly purified CD56+/CD3– NK-cells were incubated with +/- BiUII (anti-EpCAM x anti-CD3) – Antibody binding assessed by flow cytometry • Results: Binding of trAb to highly purified NK cells Expression of CD95 (activation marker) after binding of trAb Tumor cell lysis by NKcells after binding of trAb Trifunctional antibodies: Proof of principle 6. Antitumour efficacy in vivo • Mouse model – Methods: • Antibody (BiLu = anti-human EpCAM x anti-mouse CD3) was administered on days 0, 2, 4 and 7 • Mice were injected intraperitoneally with a lethal dose of B16 (EpCAM+ melanoma) or A20 (EpCAM+ B-cell lymphoma) cells on day 2 Survival after challenge – Results: Survival after challenge with B16-EpCAM cells with A20-EpCAM cells 100 100 75 % survivors % survivors • Anti-mouse EpCAM x anti-CD3 monoclonal antibody kills tumour Bs(Fab)² = without cells in vivo Fc-part 50 75 Par Abs =parental antibodies 50 • No additional co-stimulation like IL-2 needed 25 25 0 0 0 24 48 72 96 120 days after tumor challenge bsAb bsAbW par.Abs Ruf and Lindhofer Blood 2001; 98(8):2526–34 contr. 0 30 60 90 120 150 days after tumor challenge bsAb par.Abs bs(Fab´)2 Trifunctional antibodies: Proof of principle 6. Antitumour efficacy in vivo (cont’d) • Long-lasting antitumour protection in vivo (mouse model) • Results – 14 of 18 BiLu treated mice survived the primary B16-EpCAM tumour challenge – Rechallenge with B16-EpCAM tumour cells without additional antibody treatment – Long-lasting protective immunity: tumorreactive antibodies are not restricted to EpCAM – Correlation between humoral (dominant IgG2a) immune response and survival of mice Ruf and Lindhofer. Blood 2001;98(8):2526–34. Survival after rechallenge with B16EpCAM cells without additional BiLu treatment T cells CD3 • CD3 is a pan T-cell marker expressed exclusively on all T cells • CD3 and T-cell receptor form the functional T-cell receptor complex • Binding of CD3 leads to an activation of T cells. MHC=major histocompatibility complex T cells CD3 • 2x CD3e • 1x CD3d • 1x CD3g TCR=T-cell receptor. Source: Holmes N. 2000. Available at: http://wwwimmuno.path.cam.ac.uk/~immuno/part1/lec14/lec15_00.html [Accessed 25 August 2008]. Case Report Patient with heavily pretreated pancreatic cancer Patient J14: Cytokine measurements (Luminex) I IL-2 50 Cytokines in pg/ml 45 40 35 30 25 20 15 10 5 0 18.9.2009 (5µg) 23.9.2009 (10µg) before infusion 29.9.2009 (10µg) end of infusion 13.10.2009 (10µg) 20.10.2009 (10µg) 24h after infusion IL-6 2000 Cytokines in pg/ml 1800 1600 1400 1200 1000 800 600 400 200 0 18.9.2009 (5µg) 23.9.2009 (10µg) before infusion 29.9.2009 (10µg) end of infusion 13.10.2009 (10µg) 24h after infusion 20.10.2009 (10µg) Patient J14: Human anti mouse antibody measurements (HAMA) Plasma; Mean value of six measurements, < 40 ng/ml = negative 18.9.2009 (5µg) 23.9.2009 (10µg) 29.9.2009 (10µg) 13.10.2009 (10µg) 20.10.2009 (10µg) Before therapy Negative (< 40 ng/ml) End of therapy Negative (< 40 ng/ml) 24h after therapy Not done Before therapy Negative (< 40 ng/ml) End of therapy Negative (< 40 ng/ml) 24h after therapy Negative (< 40 ng/ml) Before therapy Negative (< 40 ng/ml) End of therapy Negative (< 40 ng/ml) 24h after therapy Negative (< 40 ng/ml) Before therapy Negative (< 40 ng/ml) End of therapy Negative (< 40 ng/ml) 24h after therapy Negative (< 40 ng/ml) Before therapy Negative (< 40 ng/ml) End of therapy Negative (< 40 ng/ml) 24h after therapy Negative (< 40 ng/ml) Immun Plus +++++ a new form of HAELAN 951 immun plus Introduction Asian countries Western industrialized countries 20mg – 80mg Isoflavones amount per day Less hormone-dependent tumors ¹ ² ³ Less coronary disease Fewer menopausal symptoms 1 mg - 3 mg isoflavones amount per day Higher rate of hormonedependent tumors High rate of coronary disease Common menopausal symptoms Genetic differences have been excluded by studies ④. Through Westernization a sharp increase in breast cancer rate was observed 1.Lee HP, Gourley L, Duffy SW, Esteve J, Lee J und Day NE (1991). Dietary effects on breast-cancer risk in Singapore. Lancet 337(8751):1197-1200. 2. Wu AH, Ziegler RG, Horn-Ross PL, Nomura AM, West DW, Kolonel LN, Rosenthal JF, Hoover RN und Pike MC (1996). Tofu and risk of breast cancer in Asian-Americans. Cancer Epidemiol Biomarkers Prev 5(11):901-906. 3. Messina M und Wu AH (2009). Perspectives on the soy-breast cancer relation. Am J Clin Nutr 89(5):1673S-1679S. 4. Korde LA, Wu AH, Fears T, Nomura AM, West DW, Kolonel LN, Pike MC, Hoover RN und Ziegler RG (2009). Childhood Soy Intake and Breast Cancer Risk in Asian American Women. Cancer Epidemiol Biomarkers Prev 18(4):1050-1059. The History 1992 the National Cancer Institute (NCI) invests 23,7 Mio USD to research soy. The NCI Journal publishes the study and latest findings about the newly discovered active substances : The content of isoflavones in soyfoods was the main objective in studies on far more than 400 000 women. Soy is one of the best researched food plants, with more than 10,000 relevant publications and 700 to 800 new articles per year (Messina et al. 2009a). Messina M, Watanabe S und Setchell KD (2009a). Report on the 8th International Symposium on the Role of Soy in Health Promotion and Chronic Disease Prevention and Treatment. J Nutr 139(4):796S-802S. Bioavailability und Fermentation In the plant, isoflavones are bound to sugars as glycosides from Genistin Daidzin Glycitin In the fermentation (as in immune plus ™ ®) and also partly in the gastrointestinal tract, the glycosides are split by enzymes in the aglycones: Genistein Daidzein Glycitein also in other forms such as equol (from daidzein) for example converted z.B. Equol However, only about 30% of Europeans are capable to produce equol in their own body. Aglycones are absorbed much better, so that the bioavailability is increased significantly . The amount of Aglykons represents 60% of the total weight of the glycoside: 100mg Isoflavonglykoside therefore correspond to about 60 mg of isoflavone aglycones Mechanisms of the Phytooestrogene from Soy • Anti-estrogen effect: Isoflavones have a weak estrogen effect (1: 1,000 to 1: 10,000). They bind to estrogen receptors (ER-alpha), without making an impact. They block the body's “strong” own estrogen. In this mechanism isoflavones act as anti-estrogens. • Stimulating the synthesis of SHBG (sex-hormone-binding globulin) in the liver, thus are more endogenous estrogens in bound (biologically inactive) form • Conversion of 17 beta-estradiol through 4- hydroxyestradiol into the „good“ 2hydroxyestradiol indole-3-carbino contained in soy derivative modulates the estrogen metabolism in such a manner that 4 - Hydroxöstradiol is turned into good 2 OH-estradiol. • Natural selective ER-beta agonist isoflavones dock selectively on to the ER-ß receptor and thus lead to an expression The Testosterone Metabolism ER-ß Anti-proliferation and anti - inflammatory Isoflavones are selective ER-ß agonists ER-α Proliferation and inflammatory 3ß-Adiol Estradiol 3ß-HSD* metabolised 3ß-Adiol 5 α-DHT Aromatase 5 α-Reductase Testosterone With increasing age less 3ß adiol is formed from testosterone, so an imbalance of ER-ß in favor of ER.α will be created . Age-related drop in estrogene and testosterone Inflammation + Inflammation plays an important role in tumor development 1)Huang, Y., Cao, S., Nagamani, M., Anderson, K. E., Grady, J. J., und Lu, L. J. (2005). Decreased circulating levels of tumor necrosis factor-alpha in postmenopausal women during consumption of soy-containing isoflavones. J. Clin Endocrinol. Metab 90 (7): 3956-3962. 2)Dijsselbloem, N., Vanden Berghe, W., De Naeyer, A., und Haegeman, G. (2004). Soy isoflavone phyto-pharmaceuticals in interleukin-6 affections. Multi-purpose nutraceuticals at the crossroad of hormone replacement, anti-cancer and anti-inflammatory therapy. Biochem. Pharmacol. 68 (6): 1171-1185. 3)Vanden Berghe, W., Dijsselbloem, N., Vermeulen, L., Ndlovu, N., Boone, E., und Haegeman, G. (2006). Attenuation of mitogen- and stress-activated protein kinase-1-driven nuclear factor-kappaB gene expression by soy isoflavones does not require estrogenic activity. Cancer Res. 66 (9): 4852-4862. 4)Vafeiadou, K., Hall, W. L., und Williams, C. M. (2006). Does genotype and equol-production status affect response to isoflavones? Data from a pan-European study on the effects of isoflavones on cardiovascular risk markers in post-menopausal women. Proc Nutr. Soc. 65 (1): 106-115. Two ways to inhibit the vascular endothelial growth factor VEGF (Vascular Endothelial Growth Factor) Genistein inhibits the transcription factor 1 alpha ① Genistein inhibits the transmission of signals from angiotensin II type 1 receptor for VEGF ② Inhibition of angiogenesis by genistein in human breast cancer cells and glioma cells was confirmed ③ Inhibition of VEGF effects → probably one of the main mechanisms of cancer protective effects, as observed in epidemiological and clinical studies on isoflavones. ①Guo, Y., Wang, S., Hoot, D. R., und Clinton, S. K. (2007). Suppression of VEGF-mediated autocrine and paracrine interactions between prostate cancer ells and vascular endothelial cells by soy isoflavones. J. Nutr. Biochem. 18 (6): 408-417. ②Anandanadesan, R., Gong, Q., Chipitsyna, G., Witkiewicz, A., Yeo, C. J., und Arafat, H. A. (2007). Angiotensin II Induces Vascular Endothelial Growth Factor in Pancreatic Cancer Cells Through an Angiotensin II Type 1 Receptor and ERK1/2 Signaling. J. Gastrointest. Surg. 12: 57-66. ③Schindler, R. und Mentlein, R. (2006). Flavonoids and vitamin E reduce the release of the angiogenic peptide vascular endothelial growth factor from human tumor cells. J. Nutr. 136 (6): 1477-1482. Immunological Studies of Supplement ZhenHua 851 Japan Health Management Center, Osaka University of Foreigner Studies and Japan Institut of General Medical Sciences Design: Double-bind placebo Crossover (13 control group und 13 Placebogroup) Investigator:Weimu Xisuhen // Gaoqiao Li Control drug: 2400 mg Zhenhua 851 fermented Sojaextract Duration: 2 Weeks Date: March 2000 • Results: The substance Zhenhua 851 increases the activity of killer cells (NK *) cells after a few hours and increases the number of leukocytes (white blood cells, WBC) significantly. The cellular immune response was greatly increased and the sIL-2R (soluble interleukin 2 receptor) was improved. • Sojaextract in Immun plus® shows a positive Effect on NK cells and Interleukins Study of fermented Soy extract ZhenHua 851 Therapeutic effect of adjuvant treatment for cancer with ZhenHua 851 fermented soy Karnofsky performance scale 70 Karnofsky performance scale KPS Index Life quality index 68 66 67.31 65.31 65.64 Therapiebeginn 64.2 64 Therapieende nach 2 Monaten 62 60 Chemo 101 Pat. + ZhenHua Kontrollgruppe 103 Pat. nur Chemo Quality of life has improved significantly in the Zhenhua group while deteriorating the control group accordingly Sojaextract in Immun plus® has a positive effect on the quality of life Investigator:Prof. ZenHua Zang (Alle andern nicht aufgeführt) Studienort: Department of Radiotherapy and Department of Internal Medicine in Fujian Oncology Hospital, Fuzhou First Hospital, Division of Oncology Surgery, Department of Oncology, Fujian Provincial HospitalChemo Treatment 136//Control 131 Radiotherapy Treatment 34//Control 32 Period: April – Oktober 1992 Publikation:Oktober 1992 Chinese Medical Journal Study of fermented Soy extract ZhenHua 851 Therapeutic effect of adjuvant treatment for cancer with ZhenHua 851 fermented soy The serum protein, a protein for the authoritative malnutrition has improved significantly during the chemotherapy in the Zhenhua group while deteriorating the control group accordingly Sojaextract in immune plus ® has a positive effect on malnutrition during chemotherapy Studienleitung:Prof. ZenHua Zang (Alle andern nicht aufgeführt) Studienort: Department of Radiotherapy and Department of Internal Medicine in Fujian Oncology Hospital, Fuzhou First Hospital, Division of Oncology Surgery, Department of Oncology, Fujian Provincial Hospital Chemo Treatment 136//Control 131 Bestrahlung Treatment 34//Control 32 Zeitraum: April – Oktober 1992 Publikation:Oktober 1992 Chinese Medical Journal Study of fermented SojaextractZhenHua 851 Therapeutic effect of adjuvant treatment for cancer with ZhenHua 851 fermented soy The white blood cell count has improved significantly in the Zhenhua group while deteriorating the control group accordingly Soy extract in immune plus ® increases leukocytes Studienleitung:Prof. ZenHua Zang (Alle andern nicht aufgeführt) Studienort: Department of Radiotherapy and Department of Internal Medicine in Fujian Oncology Hospital, Fuzhou First Hospital, Division of Oncology Surgery, Department of Oncology, Fujian Provincial Hospital Chemo Treatment 136//Control 131 Bestrahlung Treatment 34//Control 32 Zeitraum: April – Oktober 1992 Publikation:Oktober 1992 Chinese Medical Journal Study update positive effects with soy „Shanghai Breast Cancer Survival Study“ Prof. Dr. Xiao Ou Shu - Vanderbilt University (Nashvill, USA) • 5043 women after breastcancer therapy • Ongoing study since 5 years • First preliminary results of this largest ongoing study of breast cancer patients show a more favorable course of the disease under the influence of soy isoflavones, with a relative risk of 0.67 (p = 0.01, 95% CI 0.50 to 0.88) for disease-specific mortality in the highest compared to the lowest quartile. These for isoflavones safety questions most relevant results were published recently (Shu et al. 2009) Prof. Shu has been able to use clinical data to show life-prolonging effects with isoflavones in breast cancer patients and the benefits of the combination of tamoxifen with isoflavones (Shu 2009) ① Shu XO, Zheng Y, Cai H, Gu K, Chen Z, Zheng W und Lu W (2009). Soy intake and breast cancer survival. JAMA 302(22):2437-244 J Nutr. 2009 Apr;139(4):796S-802S. Epub 2009 Feb 18. Report on the 8th International Symposium on the Role of Soy in Health Promotion and Chronic Disease Prevention and Treatment. Messina M, Watanabe S, Setchell KD. Study update Randomized trials in women with breastcancer can be found in the published literature where 32- 132 mg isoflavones a day over a period after 3 years were despensed and had no adverse effects on cancer development Randomized trials in women with breast cancer show: 36-100 mg isoflavones per day over a period of up to three years, no adverse effects on cancer development was detected (Burke et al. 2003; Cheng et al. 2007; Hargreaves et al. 1999; Kaari et al. 2006; Marini et al. 2008; Maskarinec et al. 2009; Nahas et al. 2007; Powles et al. 2008; Qin et al. 2009; Verheus et al. 2008). A cohort study of 1954 breast cancer patients showed a lower relapse rate and no adverse interaction with tamoxifen. There have been explicitly stated positive effects against estrogen-receptor-positive breast cancer (Guha et al. 2009). Data points to protection against endometrial cancer with increased intake of isoflavones Goodman et al. 1997; Horn-Ross et al. 2003; Xu et al. 2004 Meta-analysis of 20 studies * with a duration of up to three years with a daily dose of 35-132 mg isoflavones. Not one case led to an influence on the structure of the endometrium. *Albertazzi et al. 2005; Baber et al. 1999; Balk et al. 2002; Burke et al. 2003; Caserta et al. 2005; Cheng et al. 2007; Crisafulli et al. 2004; D'Anna et al. 2007; Duncan et al. 1999b; Duncan et al. 1999a; Han et al. 2002; Kaari et al. 2006; Marini et al. 2008; Nahas et al. 2007; Nikander et al. 2005; Penotti et al. 2003; Petri Nahas et al. 2004; Powles et al. 2008; Sammartino et al. 2003; Scambia et al. 2000; Upmalis et al. 2000). Study update Phytoestrogenes wrongfully denunciated The risk of Breastcancer rises with syntetic gestagene (Conner et al. 2008)② WHI „Womens Health Initiative“ (Rossouw et al. 2002)① HET in der Menopause 16 000 postmenopausal women between the ages of 50 -79 • Duration 5.2 years • Terminated early because of 41% more strokes, 29% more heart attacks 50% thrombosis 26 %more breastcancer It seems that The development of complications (….)is not due by estrogene itself, but by the combination of synthetic gestagene and estrogenes. Increase in breast tissue density and cell division was only detected in women treated with high doses of estrogene and progestin combined (2 mg estradiol + 1 mg norethisterone acetate).(Söderqvist 2009) ③ ∙ Estrogen hypothesis = phytoestrogen = breast cancer → can no longer accurate ∙ Interplay of the estrogen receptor alpha / beta was not taken into account ∙ Phytoestrogens are ER-ß agonist ∙ Isoflavones no steroids but heterocyclic phenols ① Rossouw JE, Anderson GL, Prentice RL, LaCroix AZ, Kooperberg C, Stefanick ML, Jackson RD, Beresford SA, Howard BV, Johnson KC, Kotchen JM und Ockene J (2002). Risks and benefits of estrogen plus progestin in healthy postmenopausal women: principal results From the Women's Health Initiative randomized controlled trial. JAMA 288(3):321-333. ② Conner P, Lundström E und von Schoultz B (2008). Breast cancer and hormonal therapy. Clin Obstet Gynecol 51(3):592-606. ③ Söderqvist G (2009). Effects of conventional hormone therapy (HT) on breast tissue density and cell proliferation. Symposium on Evaluating the Efficacy and Safety of Isoflavones for Postmenopausal Women, 13-14 May. Milan (Italy): Council for Responsible Nutrition. I hope you didn’t fall asleep… Thank you very much for your kind attention! Thank you for your attention Privat Hospital Dr. Ursula Jacob Silberwaldstrasse 34 D 72280 Dornstetten – Hallwangen Germany Tel: +49 7443 964 240 Fax.: +49 7443 964 24 99 [email protected] www.ursula-jacob.de [email protected] www.rgcc-genlab.com privat clinic Dr. Ursula Jacob Silberwaldstraße 34 72280 Dornstetten-Hallwangen Tel.: +49 (0) 7443 – 964 24 – 0 Fax: +49 (0) 7443 – 964 24 – 99 [email protected] www.ursula-jacob.de Sonntag, 7. Mai 2017