The present invention provides compositions comprising an anti-angiogenic factor, and a polymeric carrier. Representative examples of anti-angiogenic factors include Anti-Invasive Factor, Retinoic acids and derivatives thereof, and paclitaxel. Also provided are methods for embolizing blood vessels, and eliminating biliary, urethral, esophageal, and tracheal/bronchial obstructions.

Anti-angiogenic compositions and methods of use

Plasminogen activation and regulation of pericellular proteolysis.

Activation of the alternative pathway of complement commences with the formation of an initial fluid-phase C3 convertase. Treatment of C3 with the nucleophilic reagent methylamine has previously been shown to result in the cleavage of an intramolecular thioester bond and to induce C3b-like properties, including the ability to form a fluid-phase C3 convertase. This report examines the hypothesis that spontaneous hydrolysis of the thioester generates a derivative of C3 that is responsible for the formation of the initial C3 convertase of the alternative pathway. The rate of spontaneous decay of C3 hemolytic activity in buffer was found to be between 0.2 and 0.4%/h. In the presence of other alternative pathway proteins, the rate of inactivation was 1%/h. The rate of spontaneous inactivation was greatly accelerated by low concentrations of chaotrophic agents such as KSCN or guanidine. Liberation of a sulfhydryl group, not present in native C3, correlated with loss of hemolytic activity, indicating that exposure to chaotropic agents resulted in thioester hydrolysis. Unlike native C3, C3 bearing a single reactive sulfhydryl group was capable of generating fluid-phase C3 convertase with Factors B, D, and P and was cleaved by Factor I (C3b inactivator) in the presence of Factor H (beta 1H). The fragmentation patterns indicated that the C3a domain was covalently associated with the functionally C3b-like C3. Organomercurial agarose was employed for the rapid removal of sulfhydryl-bearing, hemolytically inactive forms of C3 and C3b from native hemolytically active C3.

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Formation of the initial C3 convertase of the alternative complement pathway. Acquisition of C3b-like activities by spontaneous hydrolysis of the putative thioester in native C3.

Clearance and binding of two electrophoretic “fast” forms of human alpha 2-macroglobulin.

Publisher Summary This chapter presents the proteins of the alternative pathway and their special properties as enzymes, enzyme substrates, and regulators. The participation with the classical pathway components, C5–C9, in membrane attack is considered. The surprising discovery of the activation of the alternative pathway by many systems other than antibody is emphasized along with the key role played by C3 and its component parts. The alternative pathway of complement activation is closely linked to natural resistance to infections. The chapter discusses biological manifestations of alternative pathway activation. This area of research has been facilitated by isolated cytolytic alternative pathway that includes the five proteins of the membrane attack pathway. Thus, antibody-independent killing by the cytolytic pathway of bacterial and animal cells, including virus-infected human cells, is illuminated. The biomedical relevance of the alternative pathway as part of the complement system is unquestionable. Once activated, it can mark particles for phagocytosis, induce killing of nucleated cells or of bacteria, and cause lysis of enveloped viruses or virus-infected cells. It generates biologically active protein fragments, such as Bb, C3a, C3b, and C5a that influence function and behavior of inflammatory cells and cells of the immune system.

Molecular biology and chemistry of the alternative pathway of complement.

Methylamine reacts with the plasma protease inhibitor, alpha 2-macroglobulin, to form an irreversible, covalent modification. Quantitation of the reaction indicates 3.9 +/- (SD) 0.4 reactive sites per native tetrameric protein (Mr = 725,000) or one site per subunit. The reaction is selective and specific in that only 1 or 2 labeled peptides are observed on radioautography of peptide maps derived from [14C]methylamine-treated alpha 2-macroglobulin. A single chymotryptic peptide was isolated in 56% overall yield from the labeled protein. The peptide sequence by Edman degradation was found to be Gly-Cys-Gly-Glu-X-Asn-Met-(Val, Leu), in which X was the only radiolabeled phenylthiohydantoin derivative. Amino acid analysis and mass spectral analysis of the derivative suggests that X is gamma-glutamylmethylamide. Because glutamic acid and glutamine residues do not normally react with alkylamines, this work presents presumptive evidence for an alternative activated center in selected proteins.

Characterization of alkylamine-sensitive site in alpha 2-macroglobulin.

The inactivation of alpha 2M by the nucleophiles, NH4+, hydrazine, and methylamine, follows pseudo-first-order (second order, overall) rates. The rate of incorporation of nucleophiles however, is biphasic with the faster rate consistent with inactivation. Protease cleavage of alpha 2M is prevented by complete inactivation of alpha 2M by methylamine. Loss of protease cleavage is slower than methylamine incorporation but parallels inactivation. Our results are consistent with a complex model of sequential conformation changes leading to binding and inactivation of proteases by alpha 2M. Our results suggest that cross-linking of the protease to alpha 2M by aminolysis of the thiolester is not required for inactivation of the protease.

Richard P. Swenson

Papers

Characterization of alkylamine-sensitive site in alpha 2-macroglobulin.

THE METHYLAMINE REACTIVE SITE AND PROTEASE INHIBITION IN α2‐MACROGLOBULINa