Synthesis of Water-Soluble, Nonimmunogenic Polyamide Cross-Linking Agents

Abstract

Novel polyamides were developed that can be used as cross-linking agents for proteins such as hemoglobin. Water-soluble, nonimmunogenic polyamides containing oxygen and sulfur atoms in the backbone were prepared by the polycondensation of the diacids bis(carboxymethyloxyacetyl)-1,4-diaminobutane (1a) or 3, 3'-thiodipropionic acid (1b) with diethylene glycol bis(3-aminopropyl) ether (2). The resulting alpha,omega-diacids were converted to the corresponding activated esters using any of a variety of carboxylic acid activating reagents including the novel reagent diphenyl(1-methylimidazol-2-thiyl)phosphonate (9). The resulting polyamides could be activated with a broad spectrum of groups that allow for the cross-linking and surface modification of proteins.

Stepwise Solid-Phase Synthesis of Polyamides as Linkers

“…Previous syntheses of polyamides of repeat unit NHYNH−COXCO, for example, Nylon 66 and more recent solution polymerization of diacids with diamines, yield products with a wide range of chain lengths . Acute toxicity screening in rodents suggests that such polyamides are neither toxic nor immunogenic . We have shown that it is possible, by selecting an appropriate diacid and diamine for each step, to modulate factors such as hydrophobicity along the length of the chain (e.g., diamines such as 1,6-diaminohexane), to add reactive groups to the chain termini (e.g., oxime-forming groups), and to extend the chain by standard techniques of peptide synthesis.…”

“…Results of biological assays of the several PEG-polyamide linked EPO-mimetic dimers and the many multimeric constructs we have made will be reported elsewhere. What we describe here is a facile stepwise method for making a new class of polymers: biocompatible polyamides with a defined number of repeat units, which uses commercially available diacids and diamines (for an extensive list see ref ) and involves solid-phase chemistry which can be performed by machine. Previous syntheses of polyamides of repeat unit NHYNH−COXCO, for example, Nylon 66 and more recent solution polymerization of diacids with diamines, yield products with a wide range of chain lengths .…”

“…What we describe here is a facile stepwise method for making a new class of polymers: biocompatible polyamides with a defined number of repeat units, which uses commercially available diacids and diamines (for an extensive list see ref ) and involves solid-phase chemistry which can be performed by machine. Previous syntheses of polyamides of repeat unit NHYNH−COXCO, for example, Nylon 66 and more recent solution polymerization of diacids with diamines, yield products with a wide range of chain lengths . Acute toxicity screening in rodents suggests that such polyamides are neither toxic nor immunogenic .…”

“…In biochemistry and medicine, water-soluble polymer chains are useful as linkers to connect multiple copies of a structural motif − as well as for shielding proteins from the immune system and increasing biological half-life. − Ideally, these chains should be of defined structure rather than a mixture of chains of different lengths. Polypeptide chains offer one possibility 4 but are in principle susceptible to proteolytic cleavage and might prove to be immunogenic.…”

Polymerization of modified diaspirin cross-linked hemoglobin (DCLHb) with 1,6-bismaleimic-hexane

“…In the present study, DCLHb reacted first with bromoacetic acid to block the residual thiol groups of protein chains, and then an amino group on the surface of one DCLHb tetramer was converted into thiol group with 2-iminothiolane hydrochloride (2-IT); therefore, a thiol group was located on the DCLHb, which was called ''DCLHb-SH'' and could react with cross-linking reagent (Figure 1). Next, the ''DCHLb-SH'' reacted with 1,6bismaleimic-hexane (1,6-BMH, Figure 2) so that a specific intermolecular cross-linking could be formed between two converted thiol groups (Hai et al 1998), aiming at an appropriate increase of molecular mass of hemoglobin ( Figure 2). The properties of the cross-linked DCLHb were characterized, and P 50 (the O 2 pressure at which Hb is half saturated) of the product was also evaluated.…”