Sustained release systems for the perivascular administration of atorvastatin to prevent vein graft failure

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Thesis

Reference

Sustained release systems for the perivascular administration of atorvastatin to prevent vein graft failure

MYLONAKI, Ioanna

Abstract

The aim of the present thesis work was to explore sustained and local statin delivery approaches for the prevention of vein graft failure. These systems are meant to be applied perivascularly (around the vessel) during the open surgical intervention. We developed an atorvastatin delivery system containing cross-linked hyaluronic acid hydrogel and poly(lactic-co-glycolic) acid microparticles. This system allowed for a fast (over three days) and sustained (over four weeks) release of atorvastatin. The formulation was applied in vivo on the carotid artery of a rodent model of vein graft failure. Intimal hyperplasia development was inhibited by 61 % compared to control. On a porcine model of vein graft failure, a dose of 10 mg of atorvastatin applied perivascularly was not sufficient to inhibit intimal hyperplasia. In a parallel project we combined the fast and sustained release potential with mechanical support, a polyethylene terephthalate mesh coated with a layer of poly(lactic-co-glycolic) acid containing ATV was developed.

MYLONAKI, Ioanna. Sustained release systems for the perivascular administration of atorvastatin to prevent vein graft failure. Thèse de doctorat : Univ. Genève, 2017, no. Sc.

5057

URN : urn:nbn:ch:unige-929625

DOI : 10.13097/archive-ouverte/unige:92962

Available at:

http://archive-ouverte.unige.ch/unige:92962

Disclaimer: layout of this document may differ from the published version.

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UNIVERSITÉ DE GENÈVE Section de Sciences Pharmaceutiques Laboratoire de Technologie Pharmaceutique

FACULTÉ DES SCIENCES Dr. Florence Delie

Dr. Olivier Jordan

Sustained release systems for the perivascular administration of atorvastatin to prevent

vein graft failure

THÈSE

présentée à la Faculté des sciences de l’Université de Genève pour obtenir le grade de Docteur ès sciences, mention sciences pharmaceutiques

par

Ioanna Mylonaki

de

Réthymnon (GRÈCE)

These N°: 5057 Genève

Atelier de reproduction Repromail 2017

Publications issues de cette thèse J. Control. Release, 2016; 232:93-102

Biomaterials, 2017; 128:56-68

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To the wonderful feeling of discovery

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Acknowledgements

First and foremost, I would like to thank my dear supervisors Dr. Florence Delie and Dr. Olivier Jordan. Thank you, for having taken me aboard this amazing journey. It has been five long years that allowed me to discover the world of science, my child’s dream. Thank you for your patience and availability, for giving me the freedom to explore my own ideas, for allowing me to learn through trial and error. Special thanks to Florence for teaching me the importance of details and for having showed me the way to efficiently work in a collaborative group, for her positivism and her encouragement. Also to Olivier, that has always been a mine of new ideas, for propagating his excitement for science, always smiling and in a good mood. I always tend to mention Florence first but don’t take me wrong Olivier, it is just a habit! Also thank you for having shared with me moments of relaxation in the snowy Alps and the sunny Cretan beaches.

I would also like to thank Prof. Eric Allémann for integrating me in his group and for having kept me under his watchful eye. For his constructive and to the point remarks on the course of my project.

But also for his very human profile when it comes to practical problems as to renting an apartment in Geneva (!). I also have to point out how immensely I appreciated that you have tolerated my - often -caustic sense of humour. Eric, I hope you will feel younger once I will leave the lab.

A big thank you to all the wonderful collaborations I participated. The team from the university hospital of Lausanne: Dr. F. Saucy for his passionate collaboration in the project, Prof. J.-A.

Haefliger for his accurate comments, Dr. C. Dubuis, Dr. E. Allain and Dr. F. Strano for their involvement in the in vivo experiments. Also to Dr. A.-L. Rougemont from the university hospital of Geneva, for her patience when dealing with my countless histological slides. I would also like to thank Dr. A. Tzika from the university of Geneva, for having generously shared the equipment of her lab. To F. Guth and O. Deloche from the technology transfer department of the university, for taking care of the patent, the business discussions and assuring the cash to advance our projects. Also a deep appreciation goes to the team from the Centre d'Investigation Clinique - Innovation Technologique, Biomatériaux of Bordeaux, Prof. L. Bordenave, Dr. M. Durand and Dr. A. Purnama.

Their warm welcome, constant support and fruitful discussions made my stay in Bordeaux an opening towards new horizons. Also Dr. X. Berard and Dr. Barandon from the university hospital of Bordeaux, for taking the time to provide me with the necessary human material, despite their busy schedules.

My gratitude goes also to the team of FATEC and FABIO. My wonderful office mates Nathalie Boulens, Karolina Janikowska and Cedric Thauvin. I acknowledge you for having tolerated my crazy behaviour: I know that asking you to be quiet and five minutes later hiding in the recycling bin, was

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sometimes hard to deal with. My colleagues Jordan Bouilloux, Brigitte Delavy, Stella Ehrenberger, Tiziana di Francesco, Katrin Fuchs, Floriane Groell, Viktorija Herceg, Pierre Maudens, Marco Perdigao and Carlota Salgado. You guys constituted the milieu in which I want to be, thank you for teaching me how hard and also constructive it is to work with people from different cultures. For the fun we had (and hopefully we will continue having) in and outside the lab. Also my master students Naser Rehalia and Orio Trosi, for having sustained my pressure and delivered their master theses.

To my greek friends Kostas, Anna, and little Yorgos that supported me from far and assured my divertissement when I was away from Geneva. But also to the wonderful friends I made in Geneva, to Ðorđe, to Estel, to Evi, to Thanasis, to Bruno, to Davide and to Paolo, for the parties and for the good time we had together in and outside the Geneva. To Vittorio, to Vito, to Yorgos. Thank you for having undertaken the tedious challenge of sharing your lives with a stubborn PhD student. For showing me the other side of life and revitalizing me with new ideas, attitudes, and philosophies that allowed me to advance in my life.

Thank you parents, Pantelis and Désirée for having brought me up to be diligent, to have a restless character, persistent and durable to hard work. For having inspired me with the attitude that life is only a set of open doors, that nothing is impossible. Additionally, by the nature of the family you two created, you made my integration to other cultures easy and somehow natural. Finally, words might not be enough to address my thanks to my dearest and beloved friend Maria. For showing me the big picture, the other side of life, for showing me how to enjoy and do what I do with passion.

For showing me the way to be what I want to be.

Greek version: Ευχαριστώ τους γονείς μου, Παντελή και Désirée. που μου ενέπνευσαν τις αρετές της επιμονής στους στόχους μου, της συνέπειας στις υποχρεώσεις μου, της αντοχής στη σκληρή δουλειά. Που μου εμφυσήσαν μια θετική στάση απέναντι στη ζωή, βοηθώντας με να καταλάβω πως τίποτα δεν είναι αδύνατο αρκεί να έχει πίστη και επιμονή στο στόχο. Τους ευχαριστώ τέλος γιατί ο χαρακτήρας της οικογένειας που δημιούργησαν, κατέστησε εύκολη και σχεδόν φυσική την προσαρμογή και ενταξή μου σε αλλους πολιτισμούς. Τέλος, τα λόγια ίσως να μην είναι αρκετά για να εκφράσω την ευγνομωσύνη μου στην αγαπημένη φίλη μου Μαρία. Που μου έδειξε μια πιο συνολική θεώρηση των πραγμάτων, την άλλη πλευρά της ζωής, το δρόμο της ευχαρίστης και της παθιασμένης προσέγγισης. Που μου δείχνει την οδό για να είμαι αυτό που θέλω πραγματικά.

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Acronyms

ANOVA: analysis of variance AVF: arteriovenous fistula

CABG: coronary artery bypass graft CAL: carotid artery ligature

clHA: cross-linked hyaluronic acid DAPI: 4',6-diamidino-2-phenylindole fluorescent stain

DDS: drug-delivery systems FFD: farnesyl pyrophosphate EVAc: polyethylene vinyl acetate E.U.: European Union

G: gel

Gatv: atorvastatin loaded gel

GGPP: geranylgeranyl pyrophosphate HA: hyaluronic acid

HE: hematoxylin and eosin staining hGSV: human great saphenous veins HMG-CoA: 3-hydroxy-3-methyl-glutaryl- coenzyme A reductase

hVSMC: human vascular smooth muscle cells

IH: intimal hyperplasia M: microparticles

Matv: atorvastatin loaded microparticles MDs: medical devices

NO: nitric oxide

PBS: phosphate buffer saline

PABG: peripheral artery bypass graft PCL: polycaprolactone

PEAD-SA: poly (erucic acid dimer-sebacic anhydride)

PECC: poly(ethylene carbonate-ε- caprolactone)

PEG: polyethylene glycol PET: polyethylene terephthalate PMS: post-marketing surveillance PLA: polylactic acid

PLCL: poly(L-lactide-co-caprolactone) PLGA: poly(lactic-co-glycolic) acid PTFE: polytetrafluoroethylene PPO: polypropylene oxide PVAL: polyvinyl alcohol SD: standard deviation SDS: sodium dodecyl sulfate

SEM: scanning electron microscopy SEoM: standard error of the mean SVG: saphenous vein graft U.S.: United States

VGEL: Van Gieson elastin VGF: vein graft failure

α-SMA: alpha smooth muscle actin

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2

Foreword

Open surgical revascularization consists in the implantation of a vascular graft that reroutes blood flow to circumvent an area of arterial obstruction. It is used to treat peripheral and coronary artery diseases resulting from atherosclerosis. More specifically, coronary artery bypass graft (CABG) surgery is performed in patients with left main coronary artery disease and three-vessel coronary disease, whereas peripheral artery bypass graft (PABG) surgery is used to treat patients with late- stage peripheral artery occlusive disease [1]. The first CABG procedures were performed in the mid- 60’s [2]. Despite the advancements of balloon angioplasty with or without stent placement, the recent SYNTAX clinical trial showed that CABG should remain the standard of care for patients with complex lesions [3]. Notwithstanding, the differences between the two revascularization strategies should be recognized. In CABG, bypass grafts are placed on the mid-coronary vessel, providing extra sources of blood-flow to the myocardium and offering protection against the consequences of further proximal obstructive disease. In contrast, angioplasty aims at restoring normal blood- flow of the native coronary vasculature by local treatment of obstructive lesions without offering protection against new disease proximal to the stent [4]. In the United States (U.S.), 1.5 million CABG were performed in 2000 [5]. PABG referring to iliac, femoro-femoral and femoral-popliteal accounted for 450.000 annually in the U.S [6]. In the European Union (E.U.) an average of 220.000 CABG and 70.000 femoral-popliteal bypass were reported in 2014 [7]. Roughly, it is estimated that more than 2.000.000 open surgical revascularizations are performed annually in the U.S. and the E.U.

Table 1.: Clinical trials studying the patency of saphenous vein grafts post-CABG.

Saphenous vein grafts (SVG) are widely used for CABG interventions, despite the higher patency of arterial grafts [13]. This is due to their ease of harvesting, expendability, adequate length and additionaly they are employed when for more than one grafts are necessary. For PABG, SVG is the

Study Patients enrolled

Follow-up Patency rate Ref

NCT00054847 367 1 year 89 % [8]

#207/297/364 617 10 years 61 % [9]

NCT00042081 1828 12-18 months 57 % [10,11]

- 1388

1 year 88 %

[12]

5 years 75 %

> 15 years 50 %

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FOREWORD

3 conduit of choice. However, patency rates of SVG are low especially in the long term as shown in Table 1. The PREVENT IV multicenter, randomized, double-blind, clinical trial conducted in 107 U.S. sites from 2002 to 2003, is the most recent clinical trial, giving statistical evidence for vein graft failure (VGF). Angiograms were assessed to 1828 patients up to 18 months after surgery. VGF, considered as > 75 % stenosis or occlusion, was observed in 43 % of the patients, while 25 % of the grafts actually failed [11, 12].

The pathophysiology of VGF is presented in Fig. 1. It is triggered by lesions of the endothelial layer caused by the surgical process. During the first hours following the intervention, the luminal surface is covered by fibrin rich layers. Circulating white blood cells including neutrophils, monocytes and lymphocytes would attach and infiltrate the fibrin-rich layer and the intima. The migration of smooth muscle cells in the media and fibroblasts in the adventitia of the injured vessel is initiated during the first weeks. Intimal hyperplasia (IH) is at that point initiated. Growth factors and cytokines released by cells in the vessel wall, such as inflammatory cells, enhance proliferation of smooth muscle cells and induce extracellular matrix deposition, resulting in further growth of the IH. Under atherogenic conditions, macrophages in the vessel wall can take lipids up to become foam cells. A necrotic core is formed through the years, by dying foam cells, and cholesterol depositions. Ultimately, the vessel lumen occlusion leads to VGF.

Figure 1. Time frame of the development of VGF. Reproduced with permission [1]. Upper part:

Atherosclerotic process. Lower part: Intimal hyperplasia formation.

For the treatment of VGF, therapeutic approaches consist in thrombectomy, redo bypass grafting or balloon angioplasty with or without stenting [4]. Repeated bypass graft revascularization is associated with a fourfold higher mortality than primary CABG surgery [14]. Additionally, saphenous veins might no longer be available and other graft materials need to be used. Let alone

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FOREWORD

4 the burden added to the health of the patient, such treatments are associated with an explosion of health costs. These elements justify the urgent need for developing prevention strategies for VGF.

Box 1. Pleiotropic effects of statins. Adapted from [17]

Improved endothelial function/stabilizes the atherosclerotic plaque

 Inhibition of isoprenoids synthesis via inhibition of HMG-CoA reductase

 Up-regulation of the endothelial nitric oxide synthase and NAD(P)H oxidase

 Regulation of the vascular smooth muscle cell migration and proliferation

 Regulation of the cell cycle

 Regulation of tissue plasminogen activator (tPA)/tPA inhibitor expression

 Inhibition of the expression of endothelin-1 and its vasoconstriction/mitogenic

 Increasing the collagen content/reducing metalloproteinase activity, decreasing lipid content, inflammation and cell death in atherosclerotic plaques

Anti-inflammatory and immunomodulatory effects

 Reducing plasma concentrations of pro-inflammatory cytokines

 Decreasing the proliferative response of mononuclear cells

 Inhibition of leukocyte–endothelial cell adhesion by inhibition of geranylgeranylation of GTP-binding protein Rho

 Inhibition of interferon-γ-induced expression of major histocompatibility complex II molecules on human endothelial cells and macrophages

 Inhibition of T cells' activation Effects on bone metabolism

 Stimulation of the production of bone morphogenetic protein-2 (BMP-2)

 Stimulation of osteoblast differentiation and activity

 Inhibition of osteoclast development Anti-proliferative effects

 Inhibition of HMG-CoA reductase, with consecutive reduction of cellular pool of non- sterol isoprenoids FPP and GGPP

 Inhibition of the isoprenylation of GTP-binding proteins Rho, Rac, Rab, Rap, Ras, responsible for important cell signaling in cell proliferation and migration

 Inhibition of tumor growth and differentiation Effects on risk of dementia

 Reducing β-amyloid formation and deposition

 Anti-inflammatory effects

 Regulation of microglial activation

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FOREWORD

5 For the prevention of VGF, the ‘no-touch’ surgical technique, as well as anti-platelet and lipid- lowering therapies are recommended by the European and American guidelines [4, 15]. Anti-platelet therapy is currently employed to prevent thrombotic incidents [1]. Lipid-lowering hydroxymethylglutaryl-CoA reductase inhibitors (statins), which are traditionally used to treat dyslipidemia, are known to have pleiotropic effects [16]. Indeed statins have beneficial effects on endothelial function, act as anti-inflammatories, impact cell proliferation and migration (Box 1.), which are of primary importance for IH [17]. The pleiotropic effects of statins are mediated through the inhibition of cholesterol biosynthesis intermediates: the isoprenoids farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), which are ligands of lipid molecules involved in cell signaling processes [16]. The GTPase proteins Ras and Rho need to be isoprenylated by these ligands to participate in numerous intracellular signaling pathways. Therefore, statins can modulate cellular processes, such as apoptosis, proliferation, and migration, by indirectly inhibiting isoprenylation of the GTPases.

Despite the fact that the vast majority of patients undergoing GABG or PABG are under anti-platelet and lipid-lowering treatment, the vein grafts patency rates remain remains low (Table 1). Overall, no efficient preventive treatment for VGF is available up to date.

The aim of the present thesis work was to explore novel local statin delivery system approaches for the prevention of vein graft failure. These systems are designed to be applied perivascularly (around the vessel) during the open surgical intervention.

No perivascular drug delivery system (DDS) or medical device (MD) for the prevention of VGF exists on the market due to lack of efficacy. CHAPTER I presents the state of the art of perivascular DDS and MD. It reviews the existing literature and investigates the outcomes of clinical trials. The different perivascular approaches were thoroughly evaluated under the scope of facile administration, system localization, vessel constriction, drug release profile and biodistribution.

Initialy, we conceived perivascular DDS consisting of a cross-linked hyaluronic acid (clHA) gel loaded with atorvastatin (ATV). The cross-linked gel would allow for the administration and the remanence of the drug in the perivascular area. In preliminary studies, we demonstrated that ATV released from this system successfully inhibited hVSMC migration and proliferation in vitro, and that ATV was released from the gel in the cell culture medium within 10 h (ANNEX I). To achieve a longer drug release duration fitting the time-development of the pathology (Fig. 1), ATV was loaded in poly-lactic-co-glycolic acid (PLGA) microparticles. Aspirin and paclitaxel were also successfully loaded in microparticles (ANNEX II). CHAPTER II presents an in-depth evaluation of the parameters affecting the drug release kinetics of ATV from PLGA microparticles of different molecular weights. A technique consisting in sectioning microparticles during the course of drug release, allowed to observe pore formation in the matrix of the particles.

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FOREWORD

6 ATV loaded microparticles releasing ATV over a duration of 37 days and/or non-encapsulated ATV, were embedded in the clHA gel. Accordingly, in CHAPTER III, we evaluated the impact of drug release profile on the inhibition of IH development in a rodent model. This project featured in the cover story of the Journal of Controlled Release, volume 232 (ANNEX III). The encouraging results generated, led us to undertake experiments on porcine animal models that are physiologically and anatomically closer to humans. The preliminary results of these experiments are presented CHAPTER IV.

Finally, the early development of a DDS combining the sustained release of ATV with a mechanical support for the vessel is shown in CHAPTER V. The development of a PET macroporous mesh coated with PLGA containing ATV is described. The aim was to achieve a sustained release of ATV from the mesh, while preserving the mechanical properties of the system.

Refferences

[1] M.R. de Vries, K.H. Simons, J.W. Jukema, J. Braun, P.H.A. Quax, Vein graft failure: from pathophysiology to clinical outcomes, Nat. Rev. Cardiol. 13(8) (2016) 451-470.

[2] S.J. Head, T.M. Kieser, V. Falk, H.A. Huysmans, A.P. Kappetein, Coronary artery bypass grafting: Part 1—the evolution over the first 50 years, Eur. Heart J. 34(37) (2013) 2862-2872.

[3] F.W. Mohr, M.-C. Morice, A.P. Kappetein, T.E. Feldman, E. Ståhle, A. Colombo, M.J. Mack, D.R.

Holmes Jr, M.-A. Morel, N.V. Dyck, V.M. Houle, K.D. Dawkins, P.W. Serruys, Coronary artery bypass graft surgery versus percutaneous coronary intervention in patients with three-vessel disease and left main coronary disease: 5-year follow-up of the randomised, clinical SYNTAX trial, The Lancet 381(9867) (2013) 629-638.

[4] W. Stephan, P. Kolh, F. Alfonso, J.-P. Collet, J. Cremer, V. Falk, G. Filippatos, C. Hamm, S.J. Head, P.

Juni, A.P. Kappetein, A. Kastrati, J. Knuuti, U. Landmesser, G. Laufer, F.-J. Neumann, D.J. Richter, P.

Schauerte, M.S. Uva, G.G. Stefanini, D.P. Taggart, L. Torracca, M. Valgimigli, W. Wijns, A. Witkowski, 2014 ESC/EACTS guidelines on myocardial revascularization, Eur. Heart J. 68(2) (2015) 144.

[5] A.B. Bernstein, E. Hing, A. Moss, K. Allen, A. Siller, R. Tiggle, Health care in America: Trends in utilization., Hyattsville, Maryland: National Center for Health Statistics, 2003.

[6] P.P. Goodney, A.W. Beck, J. Nagle, H.G. Welch, R.M. Zwolak, National trends in lower extremity bypass surgery, endovascular interventions, and major amputations, J. Vasc. Surg. 50(1) (2009) 54-60.

[7] Eurostat, Cardiovascular diseases statistics, European Comission, 2016.

[8] S. Goldman, G.K. Sethi, W. Holman, et al., Radial artery grafts vs saphenous vein grafts in coronary artery bypass surgery: A randomized trial, JAMA 305(2) (2011) 167-174.

[9] S. Goldman, K. Zadina, T. Moritz, T. Ovitt, G. Sethi, J.G. Copeland, L. Thottapurathu, B. Krasnicka, N.

Ellis, R.J. Anderson, W. Henderson, Long-term patency of saphenous vein and left internal mammary artery grafts after coronary artery bypass surgery: Results from a Department of Veterans Affairs Cooperative Study, J. Am. Coll. Cardiol. 44(11) (2004) 2149-2156.

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FOREWORD

7 [10] C.N. Hess, R.D. Lopes, C.M. Gibson, R. Hager, D.M. Wojdyla, B.R. Englum, M. Mack, R. Califf, N.T.

Kouchoukos, E.D. Peterson, J.H. Alexander, Saphenous Vein Graft Failure after Coronary Artery Bypass Surgery: Insights from PREVENT IV, Circulation (2014).

[11] R.D. Lopes, R.H. Mehta, G.E. Hafley, J.B. Williams, M.J. Mack, E.D. Peterson, K.B. Allen, R.A.

Harrington, C.M. Gibson, R.M. Califf, N.T. Kouchoukos, T.B. Ferguson, J.H. Alexander, Relationship Between Vein Graft Failure and Subsequent Clinical Outcomes After Coronary Artery Bypass Surgery, Circulation 125(6) (2012) 749-756.

[12] G.M. Fitzgibbon, H.P. Kafka, A.J. Leach, W.J. Keon, G.D. Hooper, J.R. Burton, Coronary bypass graft fate and patient outcome: Angiographic follow-up of 5,065 grafts related to survival and reoperation in 1,388 patients during 25 years, J. Am. Coll. Cardiol. 28(3) (1996) 616-626.

[13] J.F. Sabik Iii, B.W. Lytle, E.H. Blackstone, P.L. Houghtaling, D.M. Cosgrove, Comparison of Saphenous Vein and Internal Thoracic Artery Graft Patency by Coronary System, Ann. Thor. Surg. 79(2) (2005) 544-551.

[14] C.-H. Yap, L. Sposato, E. Akowuah, S. Theodore, D.T. Dinh, G.C. Shardey, P.D. Skillington, J.

Tatoulis, M. Yii, J.A. Smith, M. Mohajeri, A. Pick, S. Seevanayagam, C.M. Reid, Contemporary Results Show Repeat Coronary Artery Bypass Grafting Remains a Risk Factor for Operative Mortality, Ann. Thor.

Surg. 87(5) (2009) 1386-1391.

[15] L.D. Hillis, P.K. Smith, J.L. Anderson, J.A. Bittl, C.R. Bridges, J.G. Byrne, J.E. Cigarroa, V.J. DiSesa, L.F. Hiratzka, A.M. Hutter, M.E. Jessen, E.C. Keeley, S.J. Lahey, R.A. Lange, M.J. London, M.J. Mack, M.R. Patel, J.D. Puskas, J.F. Sabik, O. Selnes, D.M. Shahian, J.C. Trost, M.D. Winniford, 2011 ACCF/AHA Guideline for Coronary Artery Bypass Graft Surgery: Executive Summary, A Report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines 124(23) (2011) 2610-2642.

[16] J.K. Liao, U. Laufs, Pleiotropic effects of statins, Annu. Rev. Pharmacol. Toxicol. 45 (2005) 89-118.

[17] I. Buhaescu, H. Izzedine, Mevalonate pathway: A review of clinical and therapeutical implications, Clin. Biochem. 40(9–10) (2007) 575-584.

[18] J.K. Liao, U. Laufs, PLEIOTROPIC EFFECTS OF STATINS, Annual review of pharmacology and toxicology 45 (2005) 89-118.

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CHAPTER I

Perivascular medical devices and

drug delivery systems: Making the

right choices-delivery

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11

CHAPTER I

Perivascular medical devices and drug delivery systems:

Making the right choices

Ioanna Mylonaki¹, Eric Allémann¹, François Saucy², Jacques-Antoine Haefliger², Florence Delie¹, Olivier Jordan¹

1 School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, rue Michel Servet 1, CH-1211 Geneva 4, Switzerland

2 Department of Vascular Surgery, Lausanne University Hospital, rue du Bugnon 46, CH-1011 Lausanne, Switzerland

Biomaterials, 2017; 128:56-68 doi: 10.1016/j.biomaterials.2017.02.028

Abstract

Perivascular medical devices and drug delivery systems were conceived for local application around a blood vessel during open vascular surgery. These systems provide mechanical support and/or pharmacological activity for the prevention of intimal hyperplasia following vessel injury. Despite abundant reports in the literature and numerous clinical trials, no efficient perivascular treatment is available. In this review, the existing perivascular medical devices and drug delivery systems, such as polymeric gels, meshes, sheaths, wraps, matrices, and metal meshes, are jointly evaluated. The key criteria for the design of an ideal perivascular system are identified. Perivascular treatments should have mechanical specifications that ensure system localization, prolonged retention and adequate vascular constriction. From the data gathered, it appears that a drug is necessary to increase the efficacy of these systems. As such, the release kinetics of pharmacological agents should match the development of the pathology. A successful perivascular system must combine these optimized pharmacological and mechanical properties to be efficient.

Keywords: Perivascular administration, periadventitial administration, intimal hyperplasia, wrap, mesh, cuff

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CHAPTER I - Perivascular medical devices and drug eluting systems: Making the right choices

12

Introduction

Perivascular systems, also referred to as periadventitial (Greek περί; perí, “around” the vessel or the adventitia) systems, are intended for the local treatment of focal vascular pathologies. In contrast to stents placed endovasculary when minimally invasive percutaneous interventions are performed, perivascular systems are conceived to improve the outcome of coronary or peripheric bypass interventions, arteriovenous fistulas placements and carotid surgery. According to recent clinical trials and despite the advancements of percutaneous coronary interventions (e.g. stent and/or balloon placement), the coronary artery bypass graft remains the standard of care for patients with complex lesions [1, 2]. For lower extremity vascular surgery, guidelines recommend the use of bypasses for long and diffuse arterial disease [3]. Overall, more than 2 million coronary and peripheral bypasses are performed annually in the United States and European Union [4-6]. Nevertheless, open surgery induces trauma to the vasculature associated with inflammation, thrombosis or cell proliferation, which may lead to intimal hyperplasia (IH) and consequently partial or total lumen occlusion.

Systemic administration of compounds is recommended to improve long-term surgery outcomes.

However, these treatments offer poor local efficacy and may present adverse effects [7]. As a result, up to 40 % of the vein grafts implanted occlude within five years following the intervention [8]. To address this problem, the first local perivascular formulation was described in the late 1980s [9, 10].

This was followed by the release of numerous reports on the perivascular approach. The superiority of local drug delivery system (DDSs) was demonstrated in a comparison of the anti-restenotic effect between local and systemic administration [11]. Perivascular systems range from drug-free medical

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13 devices (MDs), which exert their action through mechanical support of the vasculature, to DDSs, which offer a complementary pharmacological effect. Although numerous pre-clinical results are encouraging, the overall outcome of the clinical trials is poor. An efficient system is not yet available for perivascular application in the clinical routine [12]. It has been suggested that these poor outcomes might be partly related to inappropriate experimental models due to physiological, anatomical or interventional differences from clinical conditions [8, 13, 14].

To improve efficacy, the selection of effective compound(s) and the design of the system have to be considered together. Compound selection for the local treatment of focal vascular pathologies has been extensively reviewed elsewhere; anti-proliferative drugs, statins, enzyme inhibitors, receptor antagonists, matrix metalloproteinases, nitric oxide donors and gene therapy approaches were suggested for reducing cell proliferation [15-17]. Importantly, it was emphasized that, due to the multifactorial nature of the restenosis etiology, a single pharmacological agent may not suffice to inhibit lumen occlusion [18]. This can also be interpreted as the need to reinforce the pharmacological action with a mechanical action. Formulation design has been previously reviewed;

in the 1990s, mainly heparin perivascular DDSs were investigated [19-21]. Later, MDs with mechanical properties for vascular support were compared [22, 23]. The possible mechanisms involved in the external support strategy for preventing vein graft failure were also reviewed [24].

Furthermore, some DDSs for perivascular delivery were recently discussed [25].

The aim of the present review is to discuss the efficacy of both perivascular MDs and DDSs in order to highlight the importance of system design. Endovascular systems and compounds applied directly on the perivascular area by ultrasound-guided injection or micro-infusion catheters are beyond the scope of this review. After a brief overview of the pathology, the perivascular systems involved in the clinical trials will be discussed. The different approaches will be thoroughly evaluated under the scope of facile administration, system localization, vessel constriction, drug release profile and biodistribution. The appropriate physicochemical characteristics of the systems needed to fit the vascular pathology will be elaborated.

Perivascular physiology and vascular pathophysiology

Blood vessels are composed of three layers, the tunica intima, the tunica media and the tunica adventitia. Intimal hyperplasia develops on medium-sized vein grafts (Fig. 1A, 1B) and to a lesser extent on coronary and carotid arteries. The tunica intima, forming the endothelium, consists of a monolayer of endothelial cells. They line the luminal surface of the vascular system, providing a structural and metabolic barrier between the blood and the underlying tissues. The tunica intima is separated from the tunica media by an internal elastic lamina. The tunica media has concentrically arranged contractile or secretory smooth muscle cells. The perivascular area is delimited by the

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14 adventitia, the outer tunica of the vessel. The adventitia is an active layer of the vasculature, and it is now established that this vessel layer is the most complex and dynamic compartment of the vessel wall, containing fibroblasts, vasa vasorum, nerves and immune cells [26]. Indeed, the tunica adventitia of the vein plays an active role during vascular injury induced by surgical manipulations.

The vasa vasorum of the adventitia is a network of small blood vessels that supplies the adventitial vascular wall and drives inflammatory cells to the vessel wall in case of injury. Maiellaro et al.

present an ‘outside-in’ hypothesis in which the vascular inflammation response, prior to IH development, is initiated in the adventitia and progresses inward towards the intima [27]. In the same review, drug uptake from the adventitia to the intima was shown ex vivo, demonstrating that the diffusion of heparin from the perivasculature to the lumen area was possible (perivascular delivery) [28]. Additionally, it was calculated that this inward diffusion was associated with an outweighed convection of heparin from the lumen to the vascular tissue (endovascular delivery). Moreover, the concentration of growth factors in the rat artery tissue was markedly increased (40-fold) when administered perivascularly, compared to an intravenous administration [29]. These data document that the accumulation of a compound directly in contact with the selected tissue could be efficient when delivered perivascularly. Perivascular drug delivery actually presents distinct advantages over systemic administration, which is associated systemically with blood flow, large circulating volumes and a first-pass effect on the liver, which decrease the likelihood of the drug permeating the vascular tissue at the desired site of action.

The pathology targeted by the perivascular administration is mainly IH, triggering the atherosclerotic process and promoting lumen occlusion [30]. Following vascular intervention, a vessel injury is generated. After bypass surgery or arteriovenous fistula confection, the grafted vein needs to adapt to a new hemodynamic environment; mechanical forces, and particularly low shear stress and high wall tension, contribute to the development of IH, which is the physiological response of the vessel to various hemodynamic forces. The development of IH can be divided into three different stages (Fig. 1C) [8, 18, 31-34]. IH is initiated by endothelium disruption and extensive smooth muscle cell death in the tunica media of the vessel. Within the first 24 h, inflammatory cells, platelets and fibrin adhere and infiltrate the lesion. Fibroblasts are phenotypically modulated into myofibroblasts, while contractile smooth muscle cells change into a secretory phenotype. These cells then proliferate and migrate to the intima, contributing to the intimal thickening. Extracellular matrix is also secreted during this ‘first wave’ of the response. The second stage consists of the migration of secretory smooth muscle cells towards the intima, creating a predominant thickening after a period of four weeks. In the third stage, smooth muscle cells divide at a decreasing rate during the following weeks or months. It has been shown that approximately 50 % of the neointimal cells derive from the adventitia [35-37]. Adventitial non-migratory fibroblasts regulate collagen matrix organization,

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15 which significantly impacts constrictive remodeling [27]. It is henceforth widely accepted that the role of the adventitia is fundamental to the development of IH [38, 39].

IH causes restenosis in approximately 30–50 % of superficial coronary and femoral angioplasties, 10–30 % of coronary vein graft bypasses and 42–50 % of the arteriovenous fistulas for hemodialysis [18, 40, 41]. When autologous grafts are not available, materials such as polytetrafluoroethylene (Teflon®, PTFE) may be used instead, but they are also associated with IH development [42].

Overall, lumen occlusion following vascular surgery affects millions of patients around the world.

Figure 1. (A) Schematic diagram of a medium-sized vein. (B) Hematoxylin (dark blue nuclei)-Eosin (cytoplasm pink)-Saffron (yellow collagen) staining of a human saphenous vein. (C) Schematic diagram depicting the three stages of intimal hyperplasia (IH) development. Stage 1: Fibroblast differentiation to myofibroblasts, endothelial disruption attracting macrophages and platelets. Stage 2: Smooth muscle cell migration in the intima, extra cellular matrix production. Stage 3: Lumen narrowing due to established intimal hyperplasia.

Classification of perivascular systems

Perivascular application can either refer to MDs or DDSs, which vary in architecture and materials.

In the literature, the nomenclature of perivascular systems is somewhat confusing. Terms such as mesh, wrap, film, sheet, cuff and sheath are frequently encountered and might have overlapping

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16 meanings. In this review, we use a nomenclature based on the design of the different systems. A decision tree illustrates the different terms we suggest for the purposes of this review (Fig. 2).

Photographs of these systems applied in vivo are presented in section 5.1 (Fig. 3).

Two main categories are defined depending on the texture: semi-solids and solids. Semi-solids can be applied using a syringe and due to their plasticity may even envelop the vessel. In the category of solids, meshes are tubular structures made of a net of metallic wires or polymeric threads forming a macroporous system. Sheaths, cuffs, wraps (also called films or sheets) and matrices are formed by amorphous polymers. They can have a tubular, curved or planar geometry. The tubular systems (sheaths) can be applied like a sock on the periphery of the vein. Planar systems can be wrapped around (wrap) the vein or sutured (cuff), while matrices are placed near the vessel wall. Overall, these systems can be either MDs if they exert their effect through their mechanical properties or DDSs if they are drug carriers or are a combination of both.

Figure 2. Nomenclature of perivascularly applied systems.

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17

Clinical trials

A review of past clinical trials provides a wide range of data that may serve as a basis to optimize the development of new systems. A list of these trials using perivascular systems is presented in Table 1. Since for some trials, especially those completed before 2005, no results were published, this list is not exhaustive.

During the first decade of the 21st century, trials mainly focused on MDs such as metal and polymeric meshes. The use of steel alloy meshes (Biocompound graft®) did not improve vein graft patency (native vein 68.7 % vs. Biocompound graft 68.3 %, p ≤ 0.05), but their use could be beneficial for varicose veins [43]. In fact, a polyethylene terephthalate (Dacron®, PET) mesh (Provena®, B. Braun Inc.) is used to support varicose veins during peripheral bypass [44]. A non- constrictive Dacron® sheath reinforced with PTFE ribs failed to improve patency due to frequent thrombosis, likely because of stent rigidity or oversizing, resulting in kinking [45]. To date, the most successful effort for improving vein graft patency is eSVS® (Kips Bay Medical Inc.), consisting of a flexible kink-resistant nitinol mesh that received marketing authorization for patients undergoing coronary artery bypass grafting, which was consistently reported as safe [46]. However, enthusiasm was moderated as the outcomes of studies on graft patency following coronary bypass were contradictory. While some showed a patency up to 100 % [47, 48], others demonstrated that the patency rate of mesh-supported grafts ranged from 28 to 49 % after 9-12 months [14, 46, 49]. It was suggested that trials had confusing confounding variables other than the evaluation of the mesh that could impact results. In fact, the failures were attributed to inappropriate surgical manipulations and modifications of the application technique were suggested [50]. Cell-containing MDs have also been developed. Vascugel® is a spongy gelatin matrix containing endothelial cells that gave encouraging preclinical results [51]. Despite the safety demonstrated in clinical trials, Vascugel® did not enter later phase clinical trials due to low efficacy [52].

A shift of research focus towards DDSs has occurred over the last fifteen years. Vascular WrapTM (Angiotech Pharmaceuticals) is a paclitaxel-eluting poly(lactic-co-glycolic) acid (PLGA) wrap that was designed for the improvement of graft patency [53]. These wraps elute paclitaxel over 21 days and are fully hydrolyzed after 2-3 months. A statistically significant difference compared to the control was reported initially, and the results were encouraging for the prevention of peripheral PTFE graft failure. Nevertheless, later on, enrollment in this trial was withdrawn due to the occurrence of a high infection rate [54, 55]. A phase III clinical trial is currently ongoing for Coll-RTM, a sirolimus-eluting collagen wrap for use on patients on hemodialysis, receiving PTFE grafts. The first-in-human trial showed a 100 % surgical success rate and 76 % or 38 % unassisted graft patency at 12 or 24 months, respectively; however, the study lacks an appropriate control group [56, 57].

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18 Table 1. Perivascular systems tested in clinical trials. Clinical phases 1, 2: The primary endpoints are safety and feasibility, while the secondary endpoint is graft patency. Clinical phases 3, 4: The primary endpoint is graft patency. Patency refers to lack of lumen stenosis as assessed by angiography. PMS: post- marketing surveillance. The time frame of evaluation in months is mentioned in parentheses. CABG: coronary artery bypass graft, PABG: peripheral artery bypass graft, AVF: arteriovenous fistula, PTFE: Polytetrafluoroethylene graft.

Name System Active principle Trial Outcome Ref

Medical Devices

Biocompound Graft® Steel alloy mesh None Phase 4 68 % CABG patency (36 m) [40]

Provena® PET mesh None Phase 1 Safe, 82 % PABG patency (6 m) [41, 55]

Extent^TM PET sheath + PTFE ribs None Phase 1 Terminated - Thrombosis [42]

eSVS Mesh® Nitinol mesh None

Phase 1 Safe, 28 % CABG patency (9 m) [11, 56]

Phase 1 Safe, 49 % CABG patency (9 m) [43]

Phase 2 Unknown [57]

PMS 34 % CABG patency (12 m) [58]

PMS Terminated-Company closed [59, 60]

Vascugel® Gel matrix Endothel. cells Phase 1,2 Safe, 38 % AVF patency (6 m) [49]

Drug Delivery Systems

Heparin-alginate microcapsules Fibr. growth fact. Phase 1 Safe CABG [61]

Vascular Wrap^TM PLGA wrap Paclitaxel Phase 1,2 Terminated - Infection PTFE [50-52]

Coll-R^TM Collagen wrap Sirolimus Phase 1 Safe, 38 % PTFE patency (24 m) [53, 54]

Phase 3 Ongoing [62]

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19 Overall, most of the polymeric and metal perivascular systems have been proven safe and feasible to be applied. However, efficacy is still questionable. For perivascular DDSs, trials are still at an early stage. The efficacy evaluation is time consuming, as patency should be evaluated after several months or even years post-operation. The lack of evident positive results warrants further system development and relevant preclinical models.

Properties of perivascular systems

Once the physiochemical properties of the selected active compound(s) have been taken into account, the design of efficient systems for perivascular administration must consider the following criteria:

(1) administration mode, system localization and appropriate vascular constriction, which are related to the mechanical properties of the material (strength, flexibility, stability); and (2) drug release kinetics, which are related to the chemical properties of the polymer.

Administration – Localization

Different types of perivascular systems administered during open surgery are presented in Figure 3.

These systems should have adequate mechanical properties in order to be bent, sutured or injected and to remain on site for the desired period of time. Semi-solids, most frequently gels, present the advantages of easy handling and conformable covering of the graft [66]. Indeed, a gel should be easily manipulated by the surgeon to cover the anastomosis as well as the graft itself (Fig. 3 D and E). Additionally, appropriate gel viscosity would allow the gel to be administered or re-administered via ultrasound-guided injection or catheter micro-injection [67, 68]. On the other hand, gels may spread or move away from the site of application due to vascular tone, and in the case of coronary bypass, due to the contractile movements of the beating heart. However, anatomical constraints, e.g., muscle tissue/plane, might contribute to avoid gel migration as shown by magnetic resonance imaging [69]. Alternatively, to limit the spreading of a hydrophilic heparin-loaded polyvinyl alcohol (PVA) gel, Jones et al. enclosed it in a silicone shell to maintain the semi-liquid PVA-heparin at the perivascular site [9, 70]. Nevertheless, employing a silicone shell to limit the gel diffusion is not an appealing approach due to the non-degradable nature of the polymer.

Poloxamer (triblock copolymer: hydroxypoly(oxyethylene)/poly(oxypropylene)/ poly (oxyethylene) gels have been frequently tested in preclinical studies [79-86]. At concentrations of 20-30 %, these copolymers present the property of gelation above a critical temperature. They can be handled and injected easily at low temperatures as liquids, but they gel at body temperature [87]. Still, poloxamers are likely not appropriate for perivascular application due to their fast washout and clearance when in contact with body fluids. Indeed, in 1992, Simons et al. noticed that poloxamer F127 Pluronic®

disappeared after 1-2 h in all animals after application to the carotid artery [88]. Using magnetic

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20 resonance imaging, the signals of poloxamers, applied subcutaneously, disappeared after 45 h, confirming the short retention time of this polymer [89].

Figure 3. Images of different perivascular administration systems, adapted from the literature: A:

Braided nitinol mesh [68]; B: Knitted nitinol mesh [69]; C: Provena® PET mesh, courtesy of Dr. Fr.

Saucy; D: Dipyridamole microspheres in PLGA-PEG-PLGA Regel® gel [70]; E: Sirolimus in PLGA-PEG-PLGA Regel® gel [64]; F: Sirolimus on PET mesh [71]; G: Sirolimus in PCL wrap [72]; H: Mithramycin in EVAc (polyethylene vinyl acetate) + PEG cuff [73]; I: Paclitaxel-loaded PLGA microneedle cuff [74]; J: Sunitinib-loaded PLGA wrap [75]. Copyright owners granted their permission for re-use.

A concept combining thermogelation and biodegradability has been proposed for an injectable gel system with improved safety and an appropriate retention time [90]. Cheung’s and other groups suggested the use of PLGA-PEG-PLGA gels, (PEG: polyethylene glycol) (Regel®) [67, 69, 91-93],

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21 which are thermoresponsive and degrade within approximately 30 days [94]. This hydrophobic polymer solidifies at room temperature in contact with water within seconds upon body contact [93]

and thus requires delicate coordination in the operating room to ensure the application of its liquid form that would allow conformable covering of the graft. Another thermogelling and biodegradable gel is Atrigel®, consisting of a solution of PLGA in N-methyl-2-pyrrolidone (a class II solvent) [95].

Once injected in the body, water displaces the solvent, and the polymer precipitates instantly, forming a porous solid structure [96]. Tissue exposure to such solvents may cause local toxicity and teratogenicity [97-99]. Therefore, using a formulation with this solvent for prevention of IH is questionable.

Hyaluronic acid (HA) gels have also been used for perivascular delivery [66, 78]. HA is a natural polymer, known for its low toxicity, as it is naturally distributed throughout the connective, epithelial, and neural tissues. In its crosslinked form, mucoadhesive HA gel can avoid the migration phenomena. Its cross-linked nature slows degradation, and consequently the gel remains in vivo for several months. Importantly, cross-linked HA will eventually be enzymatically degraded by hyaluronidases. It is used for rheumatology (Ostenil®), ophthalmology (Anikavisc™) and aesthetic surgery (Fortélis Extra®) applications. Furthermore, the role of HA on IH has been described in the literature. Most studies agree that HA is able to reduce IH formation [100, 101] or at least had no effect [102]. However, an increased cellular response has been reported in some cases [103]. Two recent reviews conclude that, at high concentrations, HA is a promising candidate for IH inhibition [104, 105].

Despite the ease of administration of semi-solid forms, solid forms are frequently selected for perivascular application. Their placement is only possible when the vasculature is readily exposed during surgery, as in the case of a bypass intervention or an arteriovenous fistula placement.

Compared to semi-solid systems, solid forms are less prone to migrate away from the desired location. Non-tubular devices (cuffs and wraps) can be adapted to the size of the vessel and to angular shapes such as anastomoses. They may be sutured in place or not. The non-sutured approach allows free movement of the vessel. The sutured systems allow the surgeons to decide the constriction to apply to the vein [106]. The application of tubular devices (meshes and sheaths) is more complex and requires special equipment.

To ensure the mechanical flexibility of solid perivascular systems, the optimal thickness of PCL, PLA and PLGA wraps was set to 50 µm [75]. Polymer selection is a crucial step. Due to their high elastic modulus, cuffs and wraps made of PLGA [75, 78, 107], PLA (polylactic acid) [75] or PCL [75, 108] are often hard and brittle to handle and to suture in place, and their stiffness further increases upon drug addition [78]. For PLGA, even when implantation was possible, the wrap turned brittle after a week in vivo [75, 78]. This could result in a non-conformable covering of the graft and

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22 could also trigger injury to the vasa vasorum, which may in turn, induce IH formation [109]. To improve elasticity, PEG or methylated PEG was added as a plasticizer to improve the elastic properties of cuffs and wraps [11, 76, 110, 111]. EVAc, usually used at 40 % vinyl acetate copolymer composition, can form flexible matrices with satisfactory elastic properties [11, 76, 106, 112-115]. Alternatively, highly elastic poly(L-lactide-co-caprolactone) (PLCL) was used [116, 117].

This polymer has additionally an excellent elongation at break performance, allowing shape adaptation (1,000 times better than PLGA).

Overall, gels developed for perivascular administration present the significant advantage of ease of administration. Thermoresponsive or cross-linked gels are the most appealing approaches to date, ensuring injectability, low toxicity and increased retention time. While solid forms require more effort for their placement, they are undoubtedly less prone to migrate compared to gels. However, they do not envelop anastomoses, which are the sites where IH is most abundantly developed. For solid formulations, PLCL has the most interesting mechanical properties for the appropriate placement of cuffs and wraps. However, as will be discussed in section 5.3, controlling the drug release is compromised with the use of this polymer.

Vessel constriction: Stress or support?

A venous graft is subjected to shear stress and expands to adapt to the new environment of arterial pressure [118]. Following implantation, the vein graft will arterialize, meaning that wall thickening will occur to adjust to the arterial pressure [23]. Mechanical support and even constriction of the vessel to increase shear stress has been suggested as beneficial for the vessel’s patency [119]. Gels or loose-fitting wraps offer no mechanical support to the vessel. Instead, the caliber of meshes or sheaths can be selected to constrict the graft (Fig. 3 A, B, C). Even though they are technically challenging and there is no possibility to quantitate the constriction force applied, cuffs could allow the surgeon to adjust the constriction while suturing.

It is very important to carefully select the diameter of the device. However, the influence of vessel constriction on IH reduction is still a matter of debate. This issue has been investigated and reviewed by different groups comparing the effect of tight or loose-fitting MDs [23, 24, 120]. More specifically, the term tight fitting is used for devices with a diameter equal or smaller than the dilated vessel, whereas loose fitting is characterized by a lack of intimate contact with the graft. Table 2 summarizes the results from studies on the effect of constriction on IH. It was suggested that non- constrictive MDs are preferred for leaving space for the development of neoadventitial vasa vasorum [120-125]. However, it was shown that a tight-fitting MD would prevent the vein from over- distension, thus favoring lumen size matching between the graft and the host artery. A tight fit would thus increase the shear stress and reduce IH [72, 126-129]. Importantly, constriction should be

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23 adequate to eliminate luminal irregularities in order to inhibit IH, but should not provoke vessel wall folding [71, 130].

Another important parameter regarding constricting systems is the duration of the constrictive effect.

It has been suggested that since IH is initiated within the first month following the intervention, the presence of a supportive material is not necessary after that period [120, 121, 125]. Bioresorbable systems, either semisolids or solids, would be preferred as the risk of infection linked to the application of a solid, permanent foreign body is high with potentially dramatic consequences for the patient. The nature of the biomaterial employed and particularly its biodegradability will determine: i) the duration of the constriction and ii) the retention time of the system.

When non-degradable nitinol metal meshes are used, they offer higher kink resistance and less fraying compared to polymeric meshes [72]. The choice of the filaments/wire arrangement (woven, knitted, braided) affects the radial pulse compliance, flexibility deployment and bending stress [131].

Knitted meshes have a higher pulse compliance, lower bending stiffness and an overall greater capacity to maintain a constant caliber diameter [72]. It was also shown that the type of weaving of the meshes impacts the in vivo effect. Indeed, the remaining medial muscle mass was superior in knitted compared to braided meshes [72].

In contrast, if biodegradable polymers are used, the constraint effect will evolve with time [22]. This parameter might be appealing, since it has been suggested that ideally the transfer of shear stress load to the vessel should occur gradually [132]. Indeed, biodegradable materials may cease to fulfill their supportive role within the first month following application due to alterations in the polymer crystallinity, making the device stiffer as biodegradation occurs. Researchers observed that PLGA wraps were stiff upon explantation after 14-28 d [75, 78]. If a stiff material remains perivascularly for such a long period of time, concerns are raised over its mechanical properties and over the impact on the prevention of IH. It has also been observed that polymer hardening generated thrombosis [75].

The data presented in Table 2 suggest that constriction is beneficial against IH. Several groups have shown that IH can be sufficiently reduced only with the use of devices offering mechanical support [71, 72, 117, 120-125, 128-130, 133-138]. It seems, however, that the biodegradable materials currently available will not fulfill the constriction demands of the perivascular systems. New materials need to be explored.

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24 Table 2. Effects of solid perivascular devices (mesh and sheath) on IH after an arteriovenous bypass intervention. The in vivo effect of intimal hyperplasia reduction is reported here as the percent reduction in intima thickness. P < 0.05 (*), P < 0.01 (**), P < 0.001 (***). Refer to the source for statistical analysis method. Qc = quotient of cross-sectional area of host artery to vein graft.

External support Animal model In vivo effect on IH Ref

Non-constrictive

PTFE sheath (vascular graft) Saph. vein interposition on carotid, pig -72 % (***) [119]

PET mesh VS PTFE sheaths Saph. vein interposition on carotid, pig IH reduction for PET [130]

PET mesh Saph. vein interposition on carotid, pig -72 % (**) 8 mm mesh [120]

PET mesh Saph. vein interposition on carotid, pig -97 % (**) [121]

PET mesh Saph. vein interposition on carotids, pig -42 % (*) [118]

PLGA sheath Saph. vein interposition on carotids, pig -73 % (*) [122]

Constrictive

Polypropylene mesh Jug. vein interposition on carotid, dog IH reduction [133]

PTFE sheath (vascular graft) Jug. vein interposition on carotid, rabbit IH reduction [132]

PTFE sheath (vascular graft) Saph. vein interposition on carotid, pig No IH reduction [133]

PLA sheath Saph. vein interposition on carotids, pig No IH reduction [134]

Polyurethane/PLA sheath Jug. vein interposition on carotid, rabbit IH reduction [135]

Collagen sheath Jug. vein interposition on carotid, rabbit -45 % (***) [125]

Knitted Nitinol mesh Femorofemoral interposition, baboon -88 % (***) [69]

Knitted Nitinol mesh Aortocoronary bypass, baboon -60 % (***) Qc=0.41 [136]

Braided alloy mesh Aortocoronary bypass, sheep -52 % (*) [137]

Braided Nitinol mesh Femorofemoral interposition, baboon -91 % (**) Qc=1.47 or 3.09 [68]

Comparison

PTFE sheath (vascular graft) Jug. vein interposition on carotid, rabbit IH reduction for tight fit [126]

Braided Nitinol mesh Femorofemoral interposition, baboon -98 % (**) Qc=0.45 or 1.16 [138]

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25 Drug release profile from the DDSs

The need for a controlled drug release

Although drug-free MDs such as meshes have proven their efficacy in controlling focal vascular pathologies, their efficiency is reinforced when active compounds are added to the device. Skalsky et al. compared the effect of a bare polyester mesh and a sirolimus-releasing mesh wrapped around an autologous vein grafted in a rabbit model. Although three weeks post-surgery, no difference was seen between both devices, after six weeks, only the sirolimus-eluting meshes ensured the desired effect [74]. A wide spectrum of pharmaceutical formulations has been designed for the perivascular application of bioactive compounds. The choice of the compound, which is of utmost importance, has been reviewed elsewhere [15-17]. In this section, we focus on the importance to deliver the compound at the right place with the right kinetics.

The use of biodegradable materials offers the possibility of controlling the drug release rate. Most of them were initially investigated for perivascular application to inhibit thrombosis following vascular surgery [70, 142]. Heparin was administered perivascularly, from fast-releasing hydrophobic polyanhydride wraps. More than a two-fold improvement in anastomotic patency was observed after 7 days, compared to the untreated control in a rat microvascular thrombosis model [142]. Rogers et al. reported that the sustained perivascular release of heparin from an EVAc matrix had not only an effect on thrombosis reduction but also reduced intimal thickness by 50 % in a rabbit’s iliac artery injury model [143, 144].

Apart from these early studies, perivascular formulations are mainly used to inhibit IH, which is described as a three-stage process (Fig. 1C). The most critical processes of the IH development occur in the first 4 weeks following the vascular injury. Ideally, a single application of a DDS should be sufficient to prevent IH. For this, the drug’s availability has to be appropriate to fit the time course of the evolution of the pathology [56, 145, 146]. In the literature describing perivascular DDS, the duration of the drug release ranges from a few hours to 3 months. Most of them focus on a 2- to 4- week duration. The importance of sustaining perivascular release was demonstrated in rats, as the fast uncontrolled release of high doses of heparin led to lethal bleeding as opposed to a controlled release formulation that allowed inhibition [147].

The selection of the drug release profile (e.g., diffusion, zero-order, biphasic) is equally as important as the release duration. A diffusion-like release should ensure that the burst is controlled, to avoid exposing the surrounding tissues to excessive or even toxic levels of drug. A release profile with a lag phase with no drug release might not be desirable for this pathology. We recently documented that the combination of a burst and a sustained release are necessary for atorvastatin to effectively reduce IH by 68 % [66]. By combining atorvastatin’s diffusion-driven release from a gel with PLGA

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26 microparticles release profile, the lag phase was avoided, and the treatment was efficient. Park has also noted the importance of an adequate drug release profile in perivascular delivery [148]. Table 3 summarizes the DDS found in the literature. Gels (with or without particulate systems) and solid formulations have been largely studied for drug release for perivascular application. This table investigates the relationship between the drug release duration in vitro and the in vivo efficacy of the drug for IH reduction.

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27 Table 3: Summary of DDS found in the literature for perivascular administration. In vitro release duration is indicated when available. The in vivo effect is presented as IH reduction as I/M: intima/media ratio or I: Intima reduction or L: Lumen increase compared to control. P < 0.05 (*), P < 0.01 (**), P <

0.001(***). Refer to the source for the statistical analysis method. PECC: poly(ethylene carbonate-ε-caprolactone), PEAD-SA: poly-(erucic acid dimer- sebacic anhydride), NA: data not available, PPO: Polypropylene Oxide.

Drug Delivery System Active compound In Vitro Release In Vivo Effect Ref Gels

Poloxamer Difluoromethylornithine NA L +49 % [77]

Poloxamer Suramin NA I -50 % (*) [80]

Poloxamer Cilostazol NA I/M -62 % (*) [81]

Poloxamer Prolif. cell nuclear antigen NA I/M -27 % (*) [82]

Poloxamer CDK2 oligonucleotides NA I/M -55 % (*) [146]

Poloxamer c-myb oligonucleotides NA I/M -63 % [85, 147]

Poloxamer c-myb oligonucleotides 3 d (diffusion) I/M -90 % [143]

Poloxamer c-myc oligonucleotides 3 d (diffusion) I/M +30 % [143]

Poloxamer Nitric Oxide donor 70-80 d I/M -46 % (*) [148, 149]

Poloxamer Sirolimus NA I/M -55 % (**) [78]

Poloxamer Sirolimus NA I -41 % (ns) [79]

Cross-linked HA Atorvastatin 3 d (diffusion) I/M -30 % (ns) [63]

Regel® Sirolimus 14 d (zero-order) L +52 %(*) [64, 66]

Regel® Paclitaxel NA I -47 % (*) [88]

Atrigel® Nitric oxide donor 14 d I/M -75 % (*) [150]

Atrigel® Nitric oxide donor t1⁄2: 39 min I -39 % (**) [93]

Fibrin glue Losartan NA I -82 % (*) [151]

PVAL in silastic shell Heparin NA I/M -51 % (**) [6]

PEG-Cys-NO photopolymerized Nitric Oxide donor NA I/M -77 % (*) [152, 153]

Peptide amphiphile nanofiber Nitric Oxide donor 5 d (diffusion) I/M -77 % (*) [154]

Sustained release systems for the perivascular administration of atorvastatin to prevent vein graft failure

Thesis

Reference

Sustained release systems for the perivascular administration of atorvastatin to prevent vein graft failure

Sustained release systems for the perivascular administration of atorvastatin to prevent

vein graft failure

THÈSE

par

Ioanna Mylonaki

Acknowledgements

Table of contents

Acronyms

Foreword

Refferences

CHAPTER I

Perivascular medical devices and

drug delivery systems: Making the

right choices-delivery

CHAPTER I

Perivascular medical devices and drug delivery systems:

Making the right choices

Abstract

Table of Contents

Introduction

Perivascular physiology and vascular pathophysiology

Classification of perivascular systems

Clinical trials

Properties of perivascular systems