A novel system consisting of Rh-DuPHOS and ionic liquid for asymmetric hydrogenations.

The ionic liquid [bmim][PF6] was found to provide extra stability to the air-sensitive chiral catalyst Rh-MeDuPHOS in asymmetric hydrogenation of enamides.

Treatment of immune cell-mediated liver damage by nonpeptidic mimetics of the extracellular matrix-associated Arg-Gly-Asp epitope.

The etiology of T-cell-mediated liver injury involves the migration of immune cells, notably CD4+ T lymphocytes, into liver tissues. This process is mediated primarily by integrin-recognition of the sub-endothelium basement membranes and the extracellular matrix. The Arg-Gly-Asp-containing peptide, a major cell-adhesive ligand of extracellular matrix, is present in various plasma- and matrix-glycoproteins, such as fibronectin. Recently, we have described the design and usage of nonpeptide mimetics of Arg-Gly-Asp which bind specifically to integrins, and thereby, inhibit T cell immunity in vivo. We examined the efficacy of Arg-Gly-Asp-mimetics as potential therapeutic compounds for the treatment of experimental T-cell-mediated liver injury induced in mice by injection of Concanavalin-A. We now report that the Arg-Gly-Asp-mimetics specifically inhibited the binding of murine T cells to fibronectin, but did not affect the proliferative response of these cells in vitro. Intraperitoneal or oral administration of the Arg-Gly-Asp-mimetics but not the Arg-Gly-Asp-containing peptide, inhibited liver damage in mice if given before their inoculation with Con-A, as manifested by a lesser elevation in their serum levels of hepatic enzymes. The inhibitory effect of the Arg-Gly-Asp-mimetics was dose-dependent, the ED50 of the tested molecules being in the range of 100 micrograms per mouse and reaching maximal effect, e.g. approximately 95% inhibition, at 500 micrograms per mice. Thus, the Arg-Gly-Asp-mimetics described here may be used therapeutically to prevent immune-cell-mediated acute or chronic pathological reactions in the liver.

Novel psi-S-CH2 peptide-bond replacement and its utilization in the synthesis of nonpeptidic surrogates of the Leu-Asp-Val sequence that exhibit specific inhibitory activities on CD4+ T cell binding to fibronectin.

The Leu-Asp-Val-(LDV)-containing amino acid sequence, derived from the alternatively spliced first connecting segment region of fibronectin (FN), was shown to be recognized primarily by the alpha 4 beta 1-integrin receptor expressed on the surface of various cell types. This adhesion epitope may therefore inhibit integrin-mediated cell interactions with the extracellular matrix glycoprotein, including adhesion, migration, activation and differentiation. To probe the structural requirements for LDV recognition by integrins and examine the feasibility of inhibition of LDV-dependent cell-FN interactions, we have designed and constructed a novel psi-S-CH2 peptide bond surrogate that was employed in the formation of LDV surrogates. The synthesis of the psi-S-CH2 surrogates reported herein is based on Michael addition of 4-methylpentane thiol to an itaconic acid diester to form an S-CH2 bond. We have found that the LDV surrogates comprises of 4-methylpentanoate-Asp-i-butyl amide and 8-methyl-3-(2-methylpropylaminocarbonyl)-5-thianonanoic acid interfered with CD4+ human T-cell adhesion to FN in vitro, with an ED50 of 280 micrograms/mL. A control structural mimetic of the Leu-Glu-Val (LEV) peptide did not interfere with the T-cell-FN interaction. The specificity of the reaction was substantiated by the finding that the LDV mimetics did not interfere with T-cell adhesion to laminin, another major cell-adhesive glycoprotein of the extracellular matrix.(ABSTRACT TRUNCATED AT 250 WORDS)

Nonpeptidic analogues of the Arg-Gly-Asp (RGD) sequence specifically inhibit the adhesion of human tenon's capsule fibroblasts to fibronectin.

PURPOSE: Scar formation from the process of wound healing is mediated primarily by fibroblasts and is a major problem in ophthalmology in general and in glaucoma therapy in particular. Interactions between fibroblasts and glycoprotein components of the extracellular matrix (ECM) are among the major causes of scar formation. Recognition of ECM glycoproteins occurs via cell surface integrins that are specific for adhesion epitopes, such as the Arg-Gly-Asp (RGD-) containing amino acid sequence. The RGD sequence, which is present in several matrix and plasma proteins, including fibronectin (FN) and vitronectin, is involved in cell-ECM interactions that occur during inflammatory and homeostatic reactions. The present study was designed to examine the possibility of using RGD-mimetics as inhibitors of fibroblasts-FN interactions. METHODS: Two nonpeptidic mimetics of RGD were designed and constructed, and their inhibitory effect on the adhesion of human tenon's capsule fibroblasts to FN was examined. SF-6,5 consisting of the carboxylate group of Asp and the guanidinium group of Arg divided by a linear atom spacer and a newly designed mimetic, designated NS-11, were examined herein. RESULTS: The RGD mimetics, SF-6,5 and NS-11, but not an Arg-Gly-Glu (RGE) mimetic inhibited the adhesion of ocular fibroblasts to immobilized FN. SF-6,5 and NS-11 did not inhibit adhesion of the fibroblasts to laminin or spontaneous proliferation of fibroblasts. Trypsin pretreatment of the nonpeptidic RGD mimetics did not affect their ability to inhibit adhesion of the ocular fibroblasts to FN, whereas that of the RGD-containing peptide was abolished by the pretreatment. CONCLUSION: Nonpeptidic mimetics of RGD are attractive candidates for therapeutic agents to prevent the formation of scar tissue and related RGD-dependent pathologic events.

Inhibition of CD4+ T lymphocyte binding to fibronectin and immune-cell accumulation in inflammatory sites by non-peptidic mimetics of Arg-Gly-Asp.

The Arg-Gly-Asp (RGD) cell adhesion motif has been demonstrated in various studies to play a pivotal role in leucocyte and platelet interactions with plasma and extracellular matrix (ECM) glycoproteins. The recognition of the RGD sequence is mediated by heterodimeric receptors designated integrins of the beta 1 subfamily, expressed on distinct cell types, including T lymphocytes. We have recently shown that flexible non-peptidic mimetics of RGD, in which the two ionic side groups were separated by a linear spacer of 11 atoms, bound specifically to the platelet integrin alpha 11b beta 3, and inhibited T cell-mediated immune responses. The present study was designed to (i) further characterize the structural requirements for RGD interactions with CD4+ T cells, and (ii) examine the mechanisms by which the RGD mimetics interfere with immune cell reactivity in vivo. We now report that freezing the conformational degrees of freedom in the spacer chain, which fixes the relative orientation of the guanidinium and carboxylate side groups in a favourable manner, results in a higher level of inhibition of T cell binding to immobilized fibronectin, an RGD-containing ECM glycoprotein. In vivo, treatment of mice with relatively low doses of the RGD mimetics, but not the RGD peptide, inhibited the elicitation of an adoptively transferred DTH reaction. This inhibition was achieved by direct impairment of the ability of antigen-primed lymph node cells to migrate and accumulate in inflammatory sites. Hence, we suggest that the design and production of non-peptidic mimetics of RGD offers a novel approach to study defined parameters related to the structure-function requirements of small adhesion epitopes. Furthermore, this approach could be used therapeutically to inhibit pathological processes which depend on RGD recognition.

Inhibition of integrin-mediated platelet aggregation, fibrinogen-binding, and interactions with extracellular matrix by nonpeptidic mimetics of Arg-Gly-Asp.

The interaction of the activated platelet integrin, glycoprotein IIb-IIIa (GPIIb-IIIa) with fibrinogen and von-Willebrand factor (vWF) is essential for platelet aggregation. The minimal structure required for this integrin's binding to fibrinogen is the Arg-Gly-Asp (RGD) sequence. Inasmuch as normal level of GPIIb-IIIa-RGD interactions are required for maintaining hemostasis, elevated platelet aggregation can cause adverse pathological effects. We have previously reported that nonpeptidic mimetics of RGD, consisting of carboxylate and guanidinium groups of Asp and Arg divided by a linear 11-atom spacer, acquired a significant affinity for the GPIIb-IIIa integrin and inhibited platelet aggregation. The structural requirements for the interactions of the RGD sequence with GPIIb-IIIa and the inhibitory potential of a newly designed series of mimetics on platelet aggregation and interactions with extracellular matrix (ECM) were assayed herein. Adenosine-diphosphate (ADP)-induced platelet aggregation was inhibited in a dose-dependent manner by various RGD mimetics, with a maximal inhibition of 80-100% with an IC50 of 3 microM for the most potent inhibitor, NS-11, in which a six-membered ring was introduced into the spacer chain, which exceeded the IC50 attained with the original RGDS peptide. The inhibitory effect of the RGD mimetics was attributed to their specific interaction with the GPIIb-IIIa integrin, since these mimetics inhibited the binding of the PAC-1 mAb to GPIIb-IIIA. Furthermore, the binding of 125I-labeled fibrinogen to platelets was inhibited by the RGD surrogates in a dose-dependent and saturable manner. The RGD-mimetics also inhibited up to 70% the adhesion, aggregation, and deposition of platelets onto ECM.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of metastatic cell colonization in murine lungs and tumor-induced morbidity by non-peptidic Arg-Gly-Asp mimetics.

The spreading and colonization of tumor cells require their migration to metastatic sites via blood vessels. To penetrate blood-vessel walls, cells, including malignant ones, must recognize and associate with the sub-endothelium extracellular matrix (ECM) and its glycoproteins. Recognition of ECM-glycoproteins, such as fibronectin (FN) and vitronectin (VN), is mediated by integrin receptors expressed on various cell types, including platelets, leukocytes and tumor cells. The Arg-Gly-Asp (RGD)-containing peptide, a major adhesive ligand of ECM, is present in various plasma and matrix glycoproteins, such as FN and VN. Non-peptidic mimetics of RGD, consisting of carboxylate and guanidinium groups of Asp and Arg divided by a linear atom spacer, express a high affinity for the alpha IIb-beta 3 integrin and inhibit platelet aggregation. Herein, the ability of RGD mimetics to inhibit adhesive interactions between tumor cells and RGD, and tumor progression in vivo, was examined. RGD-containing peptides and the RGD mimetic, compound SF-6,5, but not the Arg-Gly-Glu (RGE) peptide or the corresponding mimetic, specifically inhibited B16-F10 melanoma cell adhesion to immobilized VN and FN. Daily administration in vivo of SF-6,5 to mice inhibited the formation of B16-F10 colonies in experimental and spontaneous models of metastases. Moreover, SF-6,5 could prevent mouse death caused by massive colonization of tumor cells in the lungs. The therapeutic effect of RGD-containing peptides on tumor metastasis formation was marginal, probably due to the small amounts used, and its susceptibility to proteolysis in situ. Thus, non-peptidic mimetics of small adhesive epitopes may provide a novel therapeutic tool to prevent an adverse pathological event involving integrin-dependent cell-cell and cell-ECM interactions.

Structural analysis of integrin recognition and the inhibition of integrin-mediated cell functions by novel nonpeptidic surrogates of the Arg-Gly-Asp sequence.

The pivotal role of the Arg-Gly-Asp (RGD) peptide motif in integrin-mediated cell adhesive interactions with extracellular matrix and plasma proteins stimulated the present design of nonpeptidic mimetics of this sequence. To probe the structural requirements for RGD recognition by integrins, we designed various structural mimetics of the tripeptide sequence, which consist of differentially spaced guanidinium and carboxylic groups. We now report that structures which contain guanidinium and carboxylic groups separated by an 11-carbon atom backbone mimic the distal configuration of functional RGD sequence. These compounds acquire a considerable affinity for the RGD-dependent platelet alpha IIb beta 3 integrin. As a result, these mimetics specifically inhibited platelet aggregation with an IC50 at the submillimolar range and interfered with RGD-dependent adhesion of CD4+ T-lymphocytes and metastatic tumor cells to immobilized fibronectin and vitronectin. A structural mimetic of the Arg-Gly-Glu (RGE) sequence, and structures with incorrect spacing between the functional groups, failed to inhibit these adhesive interactions. Furthermore, substitution of the guanidinium group by a primary amine abrogated the RGD-mediated biological effects. In vivo, an RGD surrogate effectively inhibited the elicitation of a delayed-type hypersensitivity reaction mediated by CD4+ T-cells, while the RGE mimetic did not. This interference suggests for a central role for RGD recognition in the regulation of immune responses. These proteolytically stable RGD mimetics may thus serve as useful therapeutic agents in versatile pathologic processes which depend on RGD recognition.