Therapeutic Regulation of Plaque Angiogenesis in Atherosclerosis: Pros and Cons

Transcription

1 Therapeutic Regulation of Plaque Angiogenesis in Atherosclerosis: Pros and Cons Karen Moulton, M.D. Dept. of Medicine, Cardiology University of Colorado Anschutz Medical Campus Learning Objectives: 1. Functions of neovascularization in atherosclerotic plaques: evidence and hypotheses. 2. Endogenous regulators of neovascularization in atherosclerotic plaques 3. Rationale for selective regulation plaque neovascularization. Disclosure: Sponsored research grant to test effects of VEGF164 aptamer on atherosclerosis in mice.

2 Functions of Plaque Neovascularization Promote atheroma growth: Werber, Atherosclerosis (1987). Moulton K, Circulation (1999). Khurana R, Circulation (2004). Vessel wall perfusion: Geiringer, J Path Bact. (1951). Heistad DD, Circulation Res. (1981). Williams JK, Circulation Res. (1988) Conduits for inflammatory cell entry: O Brien K, Circulation (1996). Moulton K, PNAS (2003). Plaque hemorrhage and vulnerability for rupture: Patterson, Arch. Pathology (1938). RBCs-atherogenic stimulus Kolodgie, NEJM (2003). Neovascularization-rupture Virmani, ATVB (2005); Moreno, Circulation (2004).

3 Neovascularization in advanced atheromas of atherosclerosis-prone mice CD31 vwf In Situ CD31 MicroCT WT Ae -/- Moulton et al (1999) Circ 99: ApoE;ApoB100: Celleti, Nat Med (2001). ApoE: Heeschen, Nat Med (2001). LDLR-/-: Petrovan, ATVB (2007) ApoE;LDLR KO, 80 wks Langhenrich. ATVB (2006) LDLR-ApoB100 Drinane. Circ Res. (2009)

4 Angiogenesis and Plaque Growth Angiogenesis inhibitors delay growth of advanced plaques Endostatin EC Moulton, Circulation (1999) TNP-470 fumagillin EC Moulton, Circulation (1999) VEGFR1-Ab EC, immune Luttun, Nat Med (2002). Angiostatin EC Moulton, PNAS (2003) sol. Flt1 (rabbit, mice) EC, immune Khurana, Circ (2004); Zhou, ATVB 2004 FGFR-dn EC, SMC Khurana, Circulation (2004) αvβ3-tnp nanoparticle EC Winter, ATVB (2006) VEGFR2-vaccine EC Petrovan, ATVB (2007) rpai-1(23) EC, SMC Drinane, Circ. Res. (2009) Angiogenesis stimulators promote plaque growth VEGF (iv) EC, immune Lazarous Circulation (1996) VEGF (iv) EC, immune Celletti, Nat Med (2001) Nicotine EC, Heeschen, Nat Med (2001) PlGF EC, immune Khurana, Circ (2005) VEGF-A,B,C,D (adeno) EC, immune Leppanen, Circ (2005) ns - Angiogenesis independent and dependent phases of plaque growth. - Characterize effects of angiogenesis regulators in atherosclerosis.

5 Vasa Vasorum: wall and plaque perfusion Anatomy: correlations with plaque size, or lamellar units. Geiringer, J Path Bact. (1951). Wolinsky, Circ Res (1967) Vessel wall perfusion: microsphere measures of flow, vasoactive changes in flow, compressive forces of artery wall.: Heistad DD, Circulation Res. (1981). Williams JK, Circulation Res. (1988); Ritman and Lerman, Cardiovasc. Res (2007) End-vessels: implications for drainage and vascular targeting. Gossl et al, Am J Phys Heart (2003). Hypoxia and oxidative stress markers in vessel wall: Bjornheden (Wiglund) ATVB (1999): Hypoxic areas in vessel wall in vivo. Heterogeneous distributions of vasa vasorum and hypoxia. Levin ATVB (2003). Mapping glucose, ATP, glycogen and lactate in vessel wall

6 Angiogenesis and Chronic Inflammation are Linked Mononuclear cells in plaque neovascularization. mice (Moulton, Circ (1999) Human (Daemen JACC (2009). Permeability: Vessels have limited pericytes. Some have abnormal junctions basement membranes and membrane blebs. (Sluimer et al JACC (2009). ApoE EC and various leukocytes share receptors for various angiogenesis factors

7 Vulnerable Plaque: Intraplaque hemorrhage is not the only potential mechanism ESC position paper on stabilisation of atherosclerotic plaques. Thromb Haemo (2011). Diverse mechanisms: cap rupture, erosions, bleeding. Diverse mechanisms of stabilization - cholesterol reduction (AngT, neovascularization, inflammatory pathways?) Role of intraplaque hemorrhage in evolution of some vulnerable plaque morphologies.

8 Title change: The question is not whether plaque neovascularization should be modified, but how it should be modified. Anti-angiogenesis agents are in widespread clinical use as standard treatments for cancer and macular degeneration. Patients with existing atherosclerosis (known and unknown) ARE receiving them. Neovascularization remodels autonomously during physiologic growth, wound healing and inflammation. Intervene on major active molecular pathways. Different goals for treatment: permeability, normalization, means for targeting specific vascular beds. Modification Regression!

9 Endogenous Regulators of Plaque Angiogenesis I. Angiogenesis stimulators in atheromas. VEGF-A (VEGF164) is major angiogenesis activity in atheromas VEGF164 and its receptors. (ATVB 2009) II. III. IV. Endogenous inhibitors in normal arteries that are depleted by atherosclerosis and vascular inflammation. Collagen XVIII (and proteolytic endostatin domain). Moulton et al. Circ (2004) and Zeng et al. JLR (2005). Other candidates: Collagen IV, fibulin-5, elastin and perlecan have cryptic anti-angiogenesis molecules that are depleted by atherosclerosis. Replacement therapy to restore a normal artery wall component. Diabetes. Moulton (unpublished 2010) Plaque angiogenesis is induced by T1D and accelerates atherosclerosis. Hypoxia-related transcription factors or metabolic stress. Sluimer & Daemen J Path (2009)

10 Sprouts Bioassay for Angiogenesis Activities in Atheromas Aorta: + Plaque nl Moulton, PNAS (2003) Fc Flt-Fc VEGF physiologic effects: EC survival, vasoreactivity, anti-thrombotic VEGF pathologic effects: VEGFR1+ monocytes, neuropilin 1+ T cells, EC and hematopoietic progenitors in angiogenesis and inflammation.

11 Differential effects of VEGF isoforms in atherosclerosis Ae-/-VEGF-wt Ae-/-VEGF 164/164 Ae-/- VEGF 188/188 VEGF isoform mice: Same total VEGF transcripts, Isoform pattern is restricted 0.15% chol. diet. Early stage,10 wks * 24 wks angiogenic stage * * *

12 Selective inhibition of VEGF164 isoform Pegylated aptamer, short half life in plasma 1st VEGF inhibitor approved for macular degeneration. Tested on tumor angiogenesis and retinopathy models in animals.

13 VEGF164 aptamer inhibited early and late-stage atherosclerosis. LDLR -/- ;ApoB100 tm/tm mice on 0.15% chol. Diet for 12 and 24 wks. Macugen: Macular Degeneration NEJM (2004)

14 Effects of VEGF164 Aptamer Treatment: Control Tx mice Inhibited plaque angiogenesis (indirect effect). VEGF164 antagonist reduced macrophages in atheromas. Matrigel plugs containing hvegf165, bfgf or no growth factors were implanted 2 weeks before the end of treatment. V164 aptamer mice developed 88% and 52% reduced vessel areas compared to controls.

15 Endogenous Regulators of Plaque Angiogenesis I. Angiogenesis stimulators in atheromas. VEGF-A (VEGF164) is major angiogenesis activity in atheromas VEGF164 and its receptors. (ATVB 2009) II. III. IV. Endogenous inhibitors in normal arteries that are depleted by atherosclerosis and vascular inflammation. Collagen XVIII (and proteolytic endostatin domain). Moulton et al. Circ (2004) and Zeng et al. JLR (2005). Other candidates: Collagen IV, fibulin-5, elastin and perlecan have cryptic anti-angiogenesis molecules that may be depleted by atherosclerosis. Replacement therapy to restore a normal artery wall component. Diabetes. Moulton (unpublished 2010) Plaque angiogenesis is induced by T1D and accelerates atherosclerosis. Hypoxia-related transcription factors or metabolic stress. Sluimer & Daemen J Path (2009)

16 Plaque Area (%) Collagen XVIII and ES product is abundant in aorta. WT HET KO Measurement WT HET KO P W v H P W v K Sudan IV+ area (%) Arch thickness (m) Cholesterol (mg/dl) Weight (g) Moulton et al. Circ. (2004)

17 Mouse Strain with Highly Angiogenic Plaques WT KO Measurement Fold-WT P VV count VV density Int.Neov. (%) % of KO arches had neovascularization compared to 13% in WT % incidence in human atheromas. (O Brien, AJP 1994).

18 Clinical Effects of high dose VEGF Inhibitors: not selective for pathologic functions of VEGF Agent Mice Rabbit Pig Human Sol Flt-1, + + BP, proteinuria, preeclampsia VEGFRinhibitors BP, proteinuria Pro-thrombotic Vegf-ab + + BP, proteinuria Pro-thrombotic Target microvascular plaque network and spare normal arteries.

19 Neuropilins: VEGFisoform selective receptors

20 ApoE -/- versus ApoE -/- ;NRP2/LacZ Nrp2 promoter LacZ activity (blue)

21 Future Goals of Neovascular Therapy I. Cancer and Macular Degeneration: chronic treatments Targets beyond growth factors and receptors. Combined agents for synergy: tumor vessels normalize and facilitate efficacy of XRT and chemo. II. Plaque Angiogenesis: Target beyond growth factor and receptors Treatment duration? Microvascular versus large vessel endothelium CV goals might reduce permeability, leukocyte adhesion and recruitment Therapy may require selective targeting or lower doses.

22 Future Exploits of Plaque Neovascularization Monitor plaque angiogenesis-related biomarkers. Neovascular imaging to assess risk and treatment response. Specific delivery mechanisms (improve efficacy and safety). Integrin-targeted fumagillin or contrast-loaded nanoparticles to plaque neovascularization Winter et, Circulation 2003.

23 Acknowledgements Moulton Lab Ian McKittrick Laura Allmon Weston Truman Ashley Johns Dmitry Belchencko Joslin Diabetes Center Giulio R. Romeo Vascular Biology (Children s Boston) Marsha Moses Diane Bielenberg Rick Rogers Elke Pravda UC Denver (Diabetes Biomarker) Kristen Nadeau (TCH, Denver) Janet Snell-Bergeon (Barbara Davis) Moshe Levi (Renal Division) Yolanda Bogaert (Renal) Animal Imaging Core Schepens Eye Institute (VEGF) Patricia D Amore UCSD (C18 LpL project) Jeff Esko Rusty Bishop Jon Gonzales