Lecture VII Inhibitors of angiogenesis 22.05.2006
mały nieunaczyniony guz rosnący unaczyniony guz VEGF (i inne...) rosnące kapilary
Wzrost nowotworu zależy od miejsca jego lokalizacji, a dokładnie od dostępu do naczyń krwionośnych
Metastasis National Cancer Institute
Angiogenic mediators in tumor growth Ferrara and Kerbel, Nature 2005
Unaczynienie guza model unaczynienia rzeczywiste naczynia
Obszary niedotlenione w guzie (a) (b) (c) Kolor niebieski (barwnik: Hoechst) przebieg naczyń krwionośnych w guzie. Kolor czerwony (barwnik: nitroimidazol EF5) obszar niedotleniony w guzie. Obrazy nalożone na siebie obszary niedotlenione znajdują się tuż przy naczyniu, co sugeruje, że naczynie nie działa prawidłowo. Giordano and Johnson 2001.
Naczynia w nowotworze wyglądają inaczej niż w zdrowej tkance: widok z zewnątrz Naczynie ze zdrowej tkanki Naczynie z nowotworu
Naczynia w nowotworze wyglądają inaczej niż w zdrowej tkance: widok z wewnątrz Naczynie ze zdrowej tkanki Naczynie z nowotworu
Wydłużenie kapilary o 1 mm pozwala na wzrost około 10 000 komórek nowotworowych
Terapia anty-angiogenna angiogenna pośrednia bezpośrednia hamuje syntezę czynników angiogennych przez komórki nowotworowe hamuje odpowiedź śródbłonka na czynniki angiogenne produkowane przez komórki nowotworowe Rosnący nowotwór Komórki śródbłonka
Wpływ hamowania angiogenezy na rozwój guza Guz nieleczony Guz leczony terapią antyangiogenną Guz wciąż rośnie, a komórki nowotworowe rozsiewają się Guz zmniejsza się
Prof. Judah Folkman ze współpracownikami
Angiostatyna hamuje wzrost nowotworów u myszy
Endostatyna hamuje wzrost nowotworów u myszy
VEGF-induced signaling in endothelial cells Zachary, Cardiovasc Res 2001
Urbich et al., FASEB 2002
Different approaches for inhibition of tumor angiogenesis
Direct and indirect inhibitors of angiogenesis
Various strategies to inhibit VEGF signaling Ferrara and Kerbel, Nature 2005
Angiogenesis inhibitors in clinical trials
Antiangiogenic effects of chemotherapeutic agents Ferrara and Kerbel, Nature 2005
Effect of Avastin and chemotherapy on lung cancer Ferrara and Kerbel, Nature 2005
RNA interference for inhibition of angiogenesis 4 th May 2005 Sirna Therapeutics Announces Results From Interim Analysis of Phase 1 Single Dose Trial With Sirna-027 in Patients With Age-Related Macular Degeneration Sirna-027 is the first chemically optimized short interfering RNA (sirna) to be tested in a human clinical trial. The compound targets Vascular Endothelial Growth Factor Receptor-1 (VEGFR-1), a key component of the clinically validated vascular endothelial growth factor (VEGF) pathway. VEGFR-1 is found primarily on vascular endothelial cells and is stimulated by both VEGF and placental growth factor (PlGF) resulting in the growth of new blood vessels. By targeting VEGFR-1, Sirna-027 is designed to reduce pathologic angiogenesis mediated by both VEGF and PlGF. To date, fourteen patients have each received a single intravitreal dose of Sirna-027 ranging from 100 to 800 micrograms. So far, all patients have experienced visual acuity stabilization during their trial participation -- the longest assessment being 84 days.
Anti-angiogenic gene therapy
Anti-angiogenic therapy for other diseases...
Proliferative diabetic retinopathy Normal macula Hemorrhages in non-proliferative diabetic retinopathy New blood vessels around optic nerve in proliferative diabetic retinopathy Hemorrhage from new blood vessel growth in proliferative diabetic retinopathy
RNA interference Dicer dsrna sirna (small interfering RNA) RISC (RNA-induced silencing complex) 5 3 mrna degradacja mrna
Clinical trials of sirna therapy of macular degeneration
DMB Macular degeneration
sirna against mrna VEGF Acuity Pharmaceuticals Normal Macula Dry AMD:Drusen formation under the Macula Wet AMD:Macula with abnormal blood vessels + sirna Carla McCullough, a 79-year-old woman in Cleveland, Ohio - 9 November 2004 first injection of sirna
sirna-027 inhibits pathological angiogenesis Trial Sites: Kontrola Wilmer Ophthalmologic Institute, Johns Hopkins Hospital School of Medicine Joint Clinical Research Center, Harvard Medical School, Massachusetts Eye and Ear Infirmary sirna-027 Cole Eye Institute, Cleveland Clinic Foundation Jules Stein Eye Institute, University of California Los Angeles
Potential application of sirna - therapy of viral infections (HCV, HIV) - inhibition of angiogenesis AMD, psoriasis, rheumatoid arthritis, cancer -Cancer inhibition of the expression of oncogenes - neurodegenerative diseases Huntington s diseases - asthma inhibition of expression of inflammatory cytokines
Angiogenesis and atherosclerosis JA Ware, Nature Med 2001
VEGF in human atherosclerotic lesions Chen et al., ATVB, 1999
Inhibitory angiogenezy zmniejszają rozmiary blaszki miażdżycowej
Looking after your heart... www.healthyheart.nbs.uk
Effects of inhibition of HMG-CoA reductase by statins STATINS
Lipid-independent statin effects Endothelial function Plaque stability Anti-thrombotic Angiogenesis STATINS Anti-inflammatory Vascular cytoprotection Anti-oxidant Immunomodulatory
Statins decrease VEGF synthesis by NIH 3T3 Ras Cells Feleszko et al., Int J Cancer. 1999 May 17;81(4):560-7.
However, the pharmacological concentration of statins in patients receiving those drugs for treatment of hypercholesterolemia are in the nanomolar range also in patients receiving experimentally very high doses of statins for treatment of cancer the highest determined plasma concentration was below 5 µm
Pro-angiogenic effect of statins... in vitro 0.1 µm simvastatin/pravastatin in vivo Kureishi et al. Nature Med., 2000 0.1 mg/kg body weight/i.p.
Biphasic effects of statins on angiogenesis stimulation of angiogenesis at low, nanomolar concentrations, inhibition at micromolar Weiss et al. Circulation 2002:105:739-745; Urbich et al. Circ Res 2002:90: 737-744 Atorvastatin (µm) Effect of atorvastatin on tube formation
Statins are pro-angiogenic at nanomolar concentrations Frick, Dulak et al., Atherosclerosis 2003 No of tubes as % of control 250 200 150 100 50 # # 0 Co VEGF 20 ng Simv 0.1 µm Simv 1 µm Simv 10 µm Mev Acid 100 µm Simv 0.1+ Mev Acid Non-treated VEGF 20ng Simv 0.1 µm Simv 1 µm
Biphasic effects of statins on angiogenesis Increased length of sprouts Disruption of sprouts control Atv 0.01 µm Atv 0.1 µm Atv 10 µm 250 * cumulative length [arbitrary units] 200 150 100 50 * 0 control 0.01 0.1 1.0 10 Atorvastatin [µm] Dulak et al., Endothelium 2005
Effect of atorvastatin on basal and hypoxia- induced VEGF synthesis in HMEC-1 NORMOXIA HYPOXIA VEGF EF2 RT-PCR qualitative and real-time control Atv 100 nm control Atv 100 nm VEGF [rel.expression] 3 2 1 NORMOXIA VEGF [rel.expression] 3 2 1 HYPOXIA 0 control Atv 100 nm 0 control Atv 100 nm N = 2 independent experiments (real-time)
Effect of atorvastatin on IL-8 synthesis in HMEC-1 RT-PCR Real-time PCR Normoxia NORMOXIA control Atv 100 nm IL-8 EF2 HYPOXIA control Atv 100 nm IL-8 [rel.expression] 1,2 0,8 0,4 0 control Hypoxia Atv 100nM % of control 100 80 60 40 20 # * * 0 control Atv 10 nm Atv 100 nm Atv 1 µm
Pharmacology & angiogenesis
Thalidomide www.celgene.com (N-phthalimido)glutarimide C 13 H 10 N 2 O 4 Derivative of glutamic acid phtalimide ring glutarimide ring Biały, krystaliczny, niemal bezwonny proszek, rozpuszczalny w DMSO.
Deformacje spowodowane talidomidem
Thalidomide inhibits angiogenesis D Amato et al., PNAS 1994 Inhibition of corneal-induced neovascularisation
Unexpected findings
Cancer Res 2004: 64: 7822-7835 VEGF 165 b 1. Is expressed in normal cells and tissues 2. Is downregulated in prostate cancer 3. binds VEGFR-2 but does not activate it
VEGF 165 b does not phosphorylate VEGF receptor
VEGF 165 b inhibits VEGF 165 -mediated angiogenesis in rabbit cornea
VEGF 165b inhibits VEG 165 mediated angiogenesis
VEGF 165b inhibits tumor growth
Always have your eyes open to unexpected findings!
Thank you for your interest in vascular biology Good luck at the exam!