Effects of topical and subconjunctival bevacizumab in high-risk corneal transplant survival.

Allograft rejection is a leading cause of corneal graft failure and thus a leading indication for repeat penetrating keratoplasty.1 Indeed, repeat grafting as a result of previous failure has become the second leading indication for corneal transplantation, as reported.2 It has been known for many decades that the presence of preexisting blood vessels is a strong risk factor for corneal graft immune rejection.3–5 Grafting into vascularized corneal beds, or so-called high-risk corneal transplantations, leads to a rate of immune rejection of greater than 50%, even with a strict regimen of topical and systemic immunosuppressive drugs.6 In fact, stratification of risk factors for immunologic rejection in penetrating keratoplasty has identified recipient vascularization as a critical proximal cause for earlier and more fulminant rejection episodes.4,5,7,8 Targeting angiogenesis to modulate immune responses after corneal transplantation has been the core area of interest for many investigators.9–13 Why the relative immunologic quiescence of the eye, which is a central facet of its immunoprivileged state, is disturbed in patients with corneal neovascularization (NV) is not fully understood.14 However, experimental evidence strongly suggests that molecular factors such as the local immunosuppressive cytokine milieu (transforming growth factor-β, α-melanocyte-stimulating hormone) and functional attributes (anterior chamber-associated immune deviation), which play a critical role in maintaining the physiologic quiescence in the anterior segment, are subverted in the presence of corneal NV.14 In addition to blood vessels in vascularized high-risk corneas, lymphatic neovessels can ingrow in parallel with hemangiogenesis, facilitating effective access of donor and host antigen-presenting cells and antigenic material to regional lymph nodes, where accelerated sensitization to graft antigens occurs.15,16 Thus, treatment of corneal NV after corneal transplantation can potentially limit both the afferent (sensitization) and efferent (rejection) arms of alloimmunity and, hence, reduce the propensity for immunoinflammatory reactions that can jeopardize graft survival.14 Vascular endothelial growth factor (VEGF) is thought to be a key mechanistic mediator of NV.17 The prominent role of VEGF in the pathophysiology of corneal NV has been demonstrated in experimental models of corneal angiogenesis.18 It has been shown that VEGF is upregulated in inflamed and vascularized corneas in humans and in animal models.19 VEGF inhibitors, including pegaptanib sodium, ranibizumab, and bevacizumab, are used for the treatment of neovascular age-related macular degeneration.20 Recently, there has been growing interest in using topical and subconjunctival anti-VEGF for the treatment of corneal NV.21–26 Our data in a prospective clinical study have demonstrated a significant reduction in the severity of corneal NV in response to topical bevacizumab therapy in patients with stable corneal NV.27 In an animal model of high-risk corneal transplantation, it has also been shown that intraperitoneal (systemic) injection of a VEGF-neutralizing cytokine trap can improve corneal graft survival.12 These reports suggest that treatment with topical or locally injected anti-VEGF could offer an adjunctive measure to conventional therapies (e.g., corticosteroids) to curb the inciting factors of graft rejection in the setting of vascularized high-risk corneal transplantation. Therefore, we sought to evaluate whether corneal graft survival in vascularized high-risk corneal transplantation can be improved by initiating local (topical or subconjunctival) bevacizumab treatment in a murine model. To make a thorough and comprehensive assessment of corneal NV, a quantitative method was implemented to measure three primary metrics—neovascular area, vessel caliber, and neovessel invasion area. Our results indicate that while both topical and subconjunctival bevacizumab therapy inhibit corneal NV after high-risk transplantation, only the subconjunctival route is significantly effective in improving graft survival.