The purpose of this study was to determine the relative importance of blood vessels (hemangiogenesis) versus lymphatic vessels (lymphangiogenesis) in mediating immunological responses after transplantation. Using the murine model of corneal transplantation, graft survival was compared in differentially prevascularized and avascular recipient beds. Donor corneas (C57BL/6) were transplanted into uninflamed or inflamed avascular, prehemvascularized only or prehemvascularized and prelymphvascularized recipient murine eyes (BALB/C). Selective inhibition of lymphangiogenesis was achieved using antivascular endothelial growth factor receptor 3 Abs and anti-integrin α5 small molecules. Grafts placed into only prehemvascularized recipient beds had a similarly good graft survival compared with grafts placed into completely avascular, normal recipients, whereas the pre-existence of lymphatic vessels significantly deteriorated corneal graft survival (p < 0.05). Lymphatic vessels seem to contribute significantly to graft rejection after (corneal) transplantation. That may allow for selective, temporary, perioperative antilymphangiogenic treatment to promote graft survival without affecting blood vessels, even after solid organ transplantation.

Topical or systemic application of bevacizumab inhibits both inflammation-induced angiogenesis and lymphangiogenesis in the cornea. This finding suggests an important role of VEGF-A in corneal lymphangiogenesis. Bevacizumab may be useful in preventing immune rejections after penetrating keratoplasty or tumor metastasis via lymphatic vessels.

Rational drug design relies on an iterative procedure of initial protein-structure determination, followed by the design, chemical synthesis, and subsequent biological evaluations of specific compounds. However, there is still a large gap between known protein sequences and 3D structures. To date, the most successful theoretical approach to bridge this gap is homology modeling. It is possible to construct an approximate 3D model of the structural unknown protein if the sequence homology to the known 3D structure of the reference protein is higher than 40%. Such a homologymodeled structure is suitable for rational drug design.[1]Herein we describe the successful use of our recently published homology model of the integrin a5b1[2] to design potent (with activities up to the subnanomolar range) and selective ligands for the two highly similar integrin receptors a5b1 and avb3. Structural considerations were used to trigger potency and selectivity in both directions. These ligands could allow functional studies in vivo of the role of these two integrin subtypes and might be used as lead structures for antiangiogenic cancer therapy.Integrins constitute an important class of heterodimeric cell-adhesion receptors that are involved in many severe pathological processes, such as tumor metastasis, thrombosis, inflammation, and osteoporosis.[3] Therefore, they have been attractive therapeutic targets for several years.[4] Since Brooks et al. reported that various low-molecular-weight ligands (for example, our synthesized cyclopentapeptide cyclo(-Arg-GlyAsp-D-Phe-Val-) = c(-RGDfV-) [5a] ), which are recognized by the avb3 and avb5 integrins, block angiogenesis in response to growth factors in tumors,[5b] many selective avb3-ligands have been developed and some compounds have reached clinical trials.[7] As a result of our research, the cyclic Nmethylated pentapeptide c(-RGDf[NMe]V-), [5b] known as cilengitide, has entered phase II trials for patients with glioblastoma.Recent knock-out experiments showed, however, that genetically altered mice (lacking the av integrin) show extensive angiogenesis in some cases, whereas other mice (lacking the b3 or b5 integrins) show no significant effects, and as such, the idea that these two integrins are proangiogenic was seriously questioned. [8,9] On the other hand, the proangiogenic function of the a5b1 receptor has been clearly demonstrated [10,11] so that the a5b1 integrin moved into the focus of research. Although crystal structures of the extracellular domains of the avb3 and aIIbb3 integrins have been solved and provided a deep insight into the ligand binding, [12,13] very little detailed structural information about the a5b1 receptor itself or about ligand-receptor interactions have been obtained until now. [14] Furthermore, there are only a few small-molecule ligands known to bind a5b1, [15,16] which prompted us to focus our research on this integrin subtype. A first hint for the design of new a5b1 ligands came from our homology model of the a5b1 integrin in complex with a rece...

An unselective cyclic peptide integrin ligand was sequentially N-methylated by a designed approach, where only the externally oriented (solvent exposed) amide bonds were N-methylated. The N-methylation resulted in tremendous enhancement in selectivity among the different integrin receptor subtypes (alpha5beta1, alphavbeta3, and alphaIIbbeta3). Conformational and docking studies were performed, which suggested that the receptor selectivity is principally caused by reduced backbone flexibility due to N-methylation.