Ординатура / Офтальмология / Английские материалы / Retinal and Choroidal Angiogenesis_Penn_2008

control by angiogenic factors, proenzyme activation, and inhibition of activated MMPs by exogenous or endogenous inhibitors, which include the tissue inhibitors of metalloproteinase (TIMPs)290 and the membraneanchored glycoprotein RECK.291 Any imbalance between MMPs and TIMPs could theoretically lead to a pro-angiogenic environment. Recently, accumulating evidence indicates that the role of each MMP and TIMP during angiogenesis depends on the particular stage of the process, as well as local microenvironmental factors.292 The potential for MMPs to play both positive and negative roles in angiogenesis highlights the need to develop selective synthetic MMP inhibitors. This would enable specific targeting of those MMPs involved mainly in promoting angiogenesis (i.e., MMP-2, MMP-9 and the membrane-type (MT)-MMPs) while sparing those involved in the generation of angiostatic proteins (such as MMP-7 and MMP-12).

Prinomastat (AG3340; Agouron Pharmaceuticals, Inc.) is a relatively selective MMP inhibitor. It potently inhibits MMP-2, -3, -9, -13, and MT- MMP-1 (MMP-14), but weakly inhibits MMP-1 and MMP-7.293 Preclinical evidence supports the ability of prinomastat administered by intraperitoneal injections to inhibit preretinal NV in a mouse OIR model.294 Intravitreal injection can prevent CNV in the laser-induced CNV model.295 However, preliminary data from phase II clinical trials of oral prinomastat in humans show the compound to be tolerated, but of no visual benefit to patients with CNV.294 Similar safe-but-no-efficacy data were collected during its oncology

trials. Further human studies with prinomastat were halted in both oncology and AMD trials in 2000.296

3.4Integrin Antagonists

Integrins are a widely expressed family of cell adhesion receptors via which cells attach to the ECM, to each other’s surfaces, or to different cell types. All integrins are composed of 2 heterodimeric units and expressed on a wide variety of cells, and most cells express several different integrins.297 In relation to angiogenesis, the integrins α2β1, αVβ3, αVβ5, and α5β1 are of

interest since the expression of these is upregulated in activated endothelial

cells.298-306

α5β1 integrin plays a key role in anchoring the abluminal surface of endothelial cells to their basement membrane, is overexpressed on tumor vessels, and is rapidly accessible from the bloodstream.307 Antibodies against α5β1 integrin inhibit angiogenesis induced by a variety of angiogenic stimuli (FGF2, TNFalpha, and IL8), but not VEGF, suggesting a potential divergence of roles for α5β1 versus αVβ3 integrins in angiogenesis.308 Similarly, these integrins may play different roles in lymphogenesis.309 Friedlander and co-workers reported the presence of αVβ3 in CNV

membranes from patients with exudative AMD or presumed ocular histoplasmosis.299,300 In addition, pharmacological antagonists of αVβ3 are efficacious inhibitors of angiogenesis. An anti-αVβ3 function-blocking

antibody310 or cyclic peptides311 suppress corneal NV,299 hypoxia-induced preretinal NV (OIR model),312 and murine tumor angiogenesis.298,313-315 A

small molecule antagonist (EMD121974 or Cilengitide) and a humanized blocking antibody (LM609 or Vitaxin®) were tested in clinical trials for their efficacy in cancer and have been found to be relatively safe and well tolerated.316

Protein Design Labs (www.pdl.com) is developing a chimeric monoclonal antibody (M200) and its Fab fragment (F200) that inhibit the functionality of α5β1 integrin; i.e., they impair binding to fibronectin. M200 is currently in a phase I trial for solid tumor treatment, while preclinical safety data are being gathered related to an exudative AMD indication. Intravenous injection of M200 or F200 has provided preliminary efficacy against laser-induced CNV in monkeys.317 Jerini Pharmaceuticals is also developing an α5β1 integrin antagonist, JSM6427, and has shown that their compound can both suppress and regress laser-induced CNV in the mouse following continuous infusion through a subcutaneous osmotic minipump.318

3.5Inhibitors with Polypharmacological Activity

3.5.1Glucocorticoids

Glucocorticoids or corticosteroids have been used therapeutically in posterior segment disease for decades. Machemer et al. first described the utility of local glucocorticoids for intraocular proliferative disease.319 Their potent and efficacious anti-inflammatory properties are most commonly used to treat conjunctivitis, keratitis, and anterior and/or posterior uveitis, where posterior uveitis typically requires local subconjunctival, subtenon, or intravitreal injection, and/or systemic administration. Unfortunately, concomitant with their anti-inflammatory activity, serious ocular adverse events have been associated with corticosteroid use, including ocular hypertension, cataract formation, and retinal toxicity.320 Typical corticosteroid drugs selected for ocular use are dexamethasone, hydrocortisone, prednisolone, and triamcinolone. Currently, several local delivery systems that provide sustained release of a variety of steroid compounds are under evaluation in clinical trials for posterior segment inflammation, edema, and pathological angiogenesis.

Relatively independent of the experimental paradigm, corticosteroids demonstrated the ability to inhibit vascular permeability and NV in a variety

of ocular tissues. Numerous publications demonstrate the anti-angiogenic efficacy of glucocorticoids, such as dexamethasone and triamcinolone, in rodent models of corneal angiogenesis,321 OIR, branch retinal vein occlusion (preretinal NV), and laser-induced CNV.322-328 In addition, intravitreal triamcinolone has been shown to reduce VEGF-induced retinal vascular permeability in adult rats [Bingaman et al., unpublished data, 2005] and rabbits.329

Although they appear to provide robust efficacy, the exact mechanism by which glucocorticoids inhibit ocular angiogenesis and vascular permeability/ edema has not been fully elucidated. Most commonly, glucocorticoids are known to inhibit the pro-inflammatory activities of leukocytes and other immune cells, and in turn reduce the angiogenic/edematous stimuli. However,

glucocorticoids can act directly upon vascular endothelial cells and inhibit growth factor-induced proliferation, migration, and tube formation.328,330 In

cell culture, glucocorticoids also have been to shown to inhibit VEGF expression and/or production in a various cell types, including human vascular smooth muscle cells, ARPE-19 cells, and human Muller cells.331-335 The mechanism by which glucocorticoids can inhibit VEGF expression appears to be a post-transcriptional destabilization of VEGF mRNA.332,335 Although triamcinolone may not alter basal VEGF expression,336 recent evidence in animal models328 and patients with DME337 supports the ability of triamcinolone acetonide to downregulate VEGF induction.

Triamcinolone Acetonide

Over the last several years, the off-label use of triamcinolone acetonide, TAA (Kenalog® 40 mg/mL, Shering-Plough), administered via intravitreal injection has become standard-of-care during the treatment of exudative AMD and DME. Although patients with exudative AMD have been reported to benefit for months following local treatment with TAA (4–25 mg),338-344 results at 1 year following a single intravitreal injection in a prospective randomized trial did not support the use of intravitreal TAA as a

monotherapy.345 Subsequently, concurrent use of intravitreal TAA with PDT has become common practice346,43 and has a two fold scientific rationale: (1)

TAA has the ability to acutely reduce subretinal edema and (2) it has the potential to reduce the burst of VEGF produced immediately post-PDT. Numerous small, physician-sponsored clinical studies suggest that local administration of TAA provides efficacy in patients with DME, as evidenced by reduced retinal thickness via noninvasive imaging (OCT) and stabilized or improved visual acuity measures.344,347-352 Not surprisingly, intravitreal

injection of TAA or crystalline cortisone also has been shown to provide empirical efficacy in patients with PDR.347,348

Although intravitreal injection of 50–100 μL Kenalog® is the most commonly reported route of administration and dose volume, posterior subconjunctival and subtenon injections also have provided clinical efficacy. Intravitreal injection was shown to be superior to posterior subtenon infusion in a recent study involving 28 patients with DME.353 Because of the excipients found in Kenalog® and their propensity to induce a limited frequency of sterile endophthalmitis,354 some ophthalmologists have begun to compound their own steroid formulation by centrifuging the Kenalog® product and then resuspending the insoluble drug particles with excipients known to be compatible with intraocular use.355 Regardless of the formulation used, cataract formation and ocular hypertension in predisposed patients are still major risk factors with triamcinolone use, especially following repeated administration. Because of the ocular liabilities associated with intraocular injection of an off-label formulation designed for intra-articular injection, the National Eye Institute (NEI, NIH, Bethesda, MD) has partnered with Allergan (Irvine, CA) and initiated clinical studies using a TAA formulation that is compatible with intraocular use.

Retisert® & MedidurTM

pSivida, Inc. (www.pSivida.com), formerly Control Delivery Systems (CDS; Watertown, MA) has been a pioneering company in the area of local delivery to the eye. Retisert® (Bausch & Lomb (www.bausch.com), NJ/pSivida, Inc., Watertown, MA), an intravitreal, nonerodible implant based on CDS’s Envision TD technology and containing fluocinolone acetonide, was approved in early 2005 for the treatment of noninfectious, posterior uveitis.356 Phase III DME studies also were being conducted. In the Retisert® device, fluocinolone is released in a zero-order fashion through dissolution of the drug from microscopic pores in the device. Releasekinetics studies estimate sustained drug delivery for up to 36 months.357 Results from two 34-week phase III randomized, multicenter, dose-masked trials in patients with noninfectious posterior uveitis demonstrated that eyes receiving the 0.5 or 2 mg Retisert® implant had a decreased recurrence rate

and improvement in visual acuity and that 20–25% had vision improvement of ≤ 3 lines.358,359 The nonerodible device requires a short surgical procedure

for implantation. One of the major issues affecting the approval of the Retisert® implant in DME has been a relatively high number of patients who experience steroid-related ocular side effects. At 12 months following implantation in a phase III DME trial, 42% of implanted eyes exhibited serious ocular adverse events, such as cataract and elevated intraocular pressure, as compared to 13% of the control (standard of care) eyes.359

MedidurTM (Alimera Sciences (www.alimerasciences.com), Atlanta, GA/Psivida, Watertown, MA) is an erodible intravitreal implant that

contains fluocinolone acetonide and is administered in the physician’s office. This delivery platform has recently been moved into a 36-month phase III study in 900 patients with DME.

Posurdex®

Posurdex® (Allergan (www.allergan.com), Irvine, CA) is a bioerodible pellet containing dexamethasone that is currently being investigated in phase III trials involving patients with persistent macular edema, where the pellet is administered intravitreally via a single-use, disposable injector containing a 23 gauge needle.360-362 The eroding implant reportedly releases dexamethasone in a zero-order fashion into the vitreous for approximately 28 days. Likely because of the shorter duration of action as compared to an intravitreal depot of TAA, Posurdex® has been associated with a reduced frequency of steroid-induced side-effects. Moreover, although the drug is estimated to be depleted from the vitreous around 1 month following implantation, patient benefit appears to persist. If this paradigm is shown to be effective through the phase III trials, it will substantiate the concept that pulse therapy may be a viable alternative to continuous therapy during the treatment of macular edema.

3.5.2RETAANE® 15mg (anecortave acetate suspension): an Angiostatic Cortisene

Anecortave acetate (Alcon® Laboratories (www.alcon.com), Ft. Worth, TX) is an angiostatic derivative of cortisol, first identified by Clark et al. at Alcon®,363 that inhibits multiple steps within the angiogenic cascade, both upstream and downstream of angiogenic growth factor ligand-receptor interaction.364-366 Three modifications were made to cortisol to generate anecortave acetate: the 11-hydroxyl, essential for glucocorticoid activity, was removed; a double bond was added between C-9 and C-11 to prevent in vivo enzymatic rehydroxylation at C-11; and an acyl group was added to the hydroxyl group at C-21 to enhance ocular penetration (Figure 3). The addition at C-21 also enhances the drug’s physicochemical properties as a slow-release depot. These modifications have resulted in the creation of a unique angiostatic molecule with no evidence of glucocorticoid receptormediated side effects.367-369

O O

Figure 23-3. Chemical design of anecortave acetate, a novel angiostatic cortisene. Panel A is the parent structure, cortisol, and Panel B is the cortisene derivative, anecortave acetate.

Anecortave acetate inhibits pathological ocular angiogenesis independent of the underlying etiological process by acting at several steps within the neovascular pathway. In early work with the compound, anecortave acetate was found to inhibit angiogenic proteolysis of the basement membrane and ECM by downregulating the expression of urokinase plasminogen activator

(uPA) and MMPs, as well as upregulating expression of plasminogen activator inhibitor-1 (PAI-1), a uPA inhibitor.364,370,371 These actions block

the proteolytic cascade and hinder migration of proliferating vascular endothelial cells through the surrounding interstitial tissues. More recently, preclinical studies demonstrate that anecortave acetate can significantly

downregulate the expression and production of both VEGF and IGF-1, as well as block VEGF-induced angiogenesis in vitro.372,365,366 Anecortave

acetate and/or its deacetylated active metabolite, anecortave desacetate, have demonstrated significant angiostatic activity in 7 different species and 12 distinct preclinical models of NV, including a surrogate model of exudative

AMD where pathological CNV is induced by laser rupture of Bruch’s membrane in mice.369,373

In clinical trials, a specially designed blunt-tipped cannula (Figure 4) is used to deliver anecortave acetate as a periocular posterior juxtascleral depot (PJD) onto the outer surface of the sclera once every six months. Preclinical pharmacokinetic data demonstrate that optimal drug delivery is achieved when the drug is placed in direct contact with the posterior scleral surface.374

indications are being conducted with this pharmaceutical class (see

www.clinicaltrials.gov). NSAIDs inhibit the formation of prostaglandins, potent pro-angiogenic molecules,376,377 through inhibition of

cyclooxygenases, of which two isoforms have been described: COX-1 and COX-2. COX-1 is expressed constitutively and has been isolated in most cell lines and in almost all mammalian tissues. It is described as a housekeeping enzyme, responsible for cell-to-cell signaling, tissue homeostasis, and cytoprotection. Conversely, COX-2 is an inducible isoform influenced by a plethora of pro-inflammatory mediators.378 Levels of the COX-2 protein are negligible in most tissues without appropriate stimulation. However, in addition to being inducible, COX-2 is also expressed constitutively in some tissues, for example, brain,379 kidney,380 and iris.381 COX-2 expression is elevated in the retina, particularly in response to hypoxia or diabetes,382-384 where these and other data suggest that COX inhibition may be a rationale treatment modality for numerous retinal diseases. Consequently, a phase I/II safety/efficacy trial employing celecoxib (Celebrex®, Merck & Co.), a selective inhibitor of COX-2, has been completed (January 2005) to determine whether vision could be stabilized or improved in patients with exudative AMD undergoing PTD. At the time of publication, the results of this trial had not been released.

Successful results using COX-2 inhibitors, such as celecoxib, in experimental diabetic models highlights the therapeutic potential for these drugs in DR and possibly other ocular neovascular diseases.385-388 The positive results from these celecoxib studies complement the previous findings with aspirin, a nonspecific COX inhibitor, tested in a 5-year study in diabetic dogs.389 In comparison to oral administration, higher ocular concentrations were achieved with application of celecoxib through the conjunctiva.387 Ocular levels following subconjuctival injection of celecoxibcontaining microparticles were sufficient to reduce oxidative stress markers in the retina of diabetic rats.388 Another therapeutic NSAID that holds promise is the prodrug, Nepafenac, which is metabolized to amfenac, a potent COX-1/2 inhibitor.390 Topical 0.1% nepafenac (Nevanac®, Alcon® Laboratories, Ft. Worth, TX) has been approved for anterior segment inflammation following cataract surgery, and it has the unique ability to achieve bioactive tissue concentrations in the posterior segment following topical ocular delivery in numerous preclinical models.391 For example, topical ocular delivery of nepafenac inhibited laser-induced CNV and ischemia-induced preretinal NV in the mouse.392 Importantly, topical nepafenac now has been shown to prevent the development of various

manifestations of NPDR (e.g., acellular capillary formation, tunnel positive cells in the microvasculature, loss of pericytes, and oscillatory potential

changes in the electroretinogram) in the streptozotocin-induced diabetic rat.384

3.5.4Antimitotic Agents: CA4P

Combretastatin A4 phosphate (CA4P) is a water-soluble prodrug of combretastatin A4 (CA4), which is a tubulin-binding agent originally isolated from the African tree Combretum caffrum.393 CA4 is a structural

analogue of colchicine that binds tubulin at the same site, where the novel aspect of CA4 has been its selective toxicity toward tumor vasculature.394,395

CA4 has been shown to cause disruption of NV in non-neoplastic tissue as well.396 Although the mechanism responsible for its selectivity for neovascular tissues has not been completely elucidated, CA4 can dramatically change the three-dimensional shape of newly formed endothelial cells, with little or no effect on quiescent endothelial cells.397 One explanation may be that mature endothelial cells have a more highly developed actin cytoskeleton, which maintains the cell shape despite depolymerization of the tubulin cytoskeleton.398 In the mouse OIR model, daily intraperitoneal injection of CA4P starting before the onset of new vessel formation suppressed the development of preretinal NV. Histological and immunohistochemical analysis indicated that CA4P permitted the development of normal retinal vasculature while inhibiting aberrant neovascularization.399 Results in murine models of VEGF overexpression and laser-induced CNV demonstrated that CA4P suppressed the development of VEGF-induced subretinal NV as well as laser-induced CNV.400 More importantly, CA4P treatment also induced partial regression of the established CNV.

A phase I/II trial in patients with exudative AMD using intravenous CA4P administration once a week for 4 weeks with 6-month follow-up is currently ongoing. Preliminary results of 7 subjects found that CA4P was well tolerated in doses up to 36 mg/m2, with the most common side-effects including mildly increased pulse and blood pressure.401 A systemic dosing phase II study in patients with CNV associated with myopic macular degeneration (MMD) also was initiated in late 2004.402 Additionally, OXiGENE® is exploring ocular delivery methods for CA4P other than intravitreal injection.403