Wenjun Du

Synthesis of Highly pH‐Responsive Glucose Poly(orthoester)

pH-Responsive polymers have great potential in biomedical applications, including the selective delivery of preloaded drugs to tissues with low pHu2005values. These polymers usually contain acid-labile linkages such as esters and acetals/ketals. However, these linkages are only mildly pH-responsive with relatively long half-lives (t1/2 ). Orthoester linkages are more acid-labile, but current methods suffer from synthetic challenges and are limited to the availability of monomers. To address these limitations, a sugar poly(orthoester) was synthesized as a highly pH-responsive polymer. The synthesis was achieved by using 2,3,4-tri-O-acetyl-α-D-glucopyranosyl bromide as a difunctional AB monomer and tetra-n-butylammonium iodide (TBAI) as an effective promoter. Under optimal conditions, polymers with molecular weights of 6.9u2005kDa were synthesized in a polycondensation manner. The synthesized glucose poly(orthoester), wherein all sugar units were connected through orthoester linkages, was highly pH-responsive with a half-life of 0.9, 0.6, and 0.2u2005hours at pHu20056, 5, and 4, respectively.

Conjugating an anticancer drug onto thiolated hyaluronic acid by acid liable hydrazone linkage for its gelation and dual stimuli-response release.

A prodrug gelation strategy was developed for the sustained and dual stimuli-response release of doxorubicin hydrochloride (DOX·HCl), a commonly used anticancer drug. For this purpose, the chemical conjugation of DOX·HCl onto thiolated hyaluronic acid (HA) was carried out by an acid liable hydrazone linkage and verified by (1)H NMR analyses. When exposed to the air, such a polysaccharide conjugate showed unique self-gelation ability in aqueous solution. The gelation time and extent depended mainly on the content of thiol groups on thiolated HA. The resultant hydrogel exhibited a dominant elastic response and a thixotropic property. In particular, it could release sustainably conjugated DOX·HCl in dual pH- and reduction-responsive modes. The cumulative drug release was found to be significantly accelerated under the conditions mimicking the intracellular environments of cancer cells. The in vitro cytotoxicity assays for the human nasopharyngeal carcinoma CNE2 cells treated with various release media confirmed the effectiveness of this conjugate hydrogel for cancer cell inhibition.

A general chemical synthesis platform for crosslinking multivalent single chain variable fragments

Multivalent single chain variable fragments (scFv) show increased affinity to tumor-associated antigens compared to monovalent scFv and intact monoclonal antibodies (mAb). Multivalent constructs can be derived from self-associating or covalent scFv with covalent constructs offering improved in vivo and in vitro stability. Covalent attachment of scFv can be achieved using genetically engineered expression vectors that afford scFv with site specific cysteine functionality. Expression vectors for di-scFv-C wherein the cysteine is located in the center of two scFv have also been developed for attaching chemically reactive linkers. In the example illustrated here, the di-scFv-C is derived from a mAb directed against the MUC1 epitope, which is presented on cancer cells. To achieve multivalency, a chemical crosslinking strategy utilizing various azide and multi-alkyne functionalized polyethylene glycol (PEG) linkers was implemented. Conjugation was achieved by attachment of these linkers to the scFv thiol functionality. Chemoselective ligation was employed to covalently link different protein conjugates via copper(I) catalyzed azide alkyne 1,3-dipolar cycloaddition reaction (CuAAC) chemistry. Ligations were achieved in >70% yield using a specific set of linkers as determined by SDS-PAGE and densitometry. ELISA showed increased tumor binding of a tetravalent scFv providing a versatile chemical crosslinking strategy for construction of multivalent and bi-specific immunoconjugates that retain biological activity and have potential application in pre-targeted radioimmunotherapy and imaging.

Water soluble cationic dextran derivatives containing poly(amidoamine) dendrons for efficient gene delivery.

To develop new dextran derivatives for efficient gene delivery, the conjugation of poly(amidoamine) dendrons onto biocompatible dextran was carried out by a Cu(I)-catalyzed azide-alkyne cycloaddition, as confirmed by FTIR and (1)H NMR analyses. For resultant dextran conjugates with various generations of poly(amidoamine) dendrons, their buffering capacity and in vitro cytotoxicity were evaluated by acid-base titration and MTT tests, respectively. In particular, their physicochemical characteristics for the complexation with plasmid DNA were investigated by the combined analyses from agarose gel electrophoresis, zeta potential, particle size, transmission electron microscopy and fluorescence emission spectra. Moreover, their complexes with plasmid DNA were studied with respect to their transfection efficiency in human embryonic kidney (HEK293) cell lines. In the case of a higher generation of poly(amidoamine) dendrons, such a dextran conjugate was found to have much lower cytotoxicity and better gene delivery capability when compared to branched polyethylenimine (bPEI, 25kDa), a commonly used gene vector.

Complete Depolymerization and Repolymerization of a Sugar Poly(orthoester)

The capability of a polymer to depolymerize, which regenerates its original monomer for further polymerization, is very attractive in terms of sustainability. Sugar poly(orthoester) was recently synthesized as a novel class of glycopolymer. The high sensitivity of the backbone orthoester linkage towards acidolysis provides a valuable model to study its depolymerization. Herein, we report that the sugar poly(orthoester) can be completely depolymerized at acidic conditions. However, in most cases, the depolymerization gave a stable cyclic product (1,6-anhydro glucopyranose), instead of its original monomer. We discovered that the formation of the cyclic structure was both kineticallyand thermodynamically-favored. However, this pathway could be shut down by chemically deactivating a key intermediate, shifting the reaction pathway that favors the formation of the original monomer. We further demonstrate that the regenerated monomer can be repolymerized efficiently. The fast depletion of fossil fuels, as well as the detrimental environmental impact by the non-degradable polymers urge the development of completely degradable polymers that can be obtained from renewable resources. Much effort has been invested, especially in the chemical syntheses of polymerizable monomers from naturally occurring building blocks, such as plant oil and carbohydrates, 11] as shown in Figure 1. Figure 1. Examples of polymerizable monomers synthesized from natually occuring building blocks; including those from plant oils (I-A and I-B); and from carbohydrates (II-A and II–B). Although the idea is attractive, the conversions of natural building blocks to polymerizable monomers are often challenging and tedious, requiring multiple synthetic steps, with each step in low yields. For example, Narine and co-workers reported that the synthesis of 1,7-heptamethylene diisocyanate monomer from oleic acid involved several steps including: 1) ozonolysis to cleave the double bonds; 2) oxidation; 3) conversion of the carboxylic acids to azides, followed by Curtius rearrangement to give the diisocyanate. The conversion was not only tedious but was also in low efficiency. Wooley and co-workers reported that the synthesis of a cyclic 4,6-carbonate glucose monomer (from glucose). The process involved a methylation; a selective protection-deprotection, as well as a ring-closing step. The process gave low efficiency, especially in the ring closing step. In another example, Du and co-workers reported the synthesis of a di-functional glucose monomer (2,3,4-tri-O-acetyl-alpha-Dglucosyl bromide), the process, starting from glucose, involved selective protection, acetylation, functional group transformation, and selective deprotection. Each of the steps were technically challenging. Other naturally occurring building blocks have been employed, but the conversions to polymerizable monomers also suffered from similar challenges. On the other hand, when polymers finish their duties, they are often dumped in landfills, where they are degraded into small molecules, usually, with the help of enzymes. Unfortunately, the processes are very slow, due to the lack of enzymes, and/or unfavorable conditions for the solid-state enzymatic catalysis. Furthermore, the degraded small molecules, which are often toxic, can leach into soil and underground water, causing secondary pollutions. Taking these challenges/problems into account, it would be highly desirable to develop fully sustainable polymers, which can be degraded directly into their original monomers, and can be repolymerized again. This concept represents a new class of polymers that can undergo repeated cycle of depolymerization and repolymerization, as illustrated as a cartoon in Figure 2. Figure 2. A schematic illustration of a complete cycle of depolymerization and repolymerization. [*] L. Li, S. Maiti, N. A. Thompson, I. J. Milligan, Prof. W. Du Department of Chemistry and Biochemistry Science of Advanced Materials Central Michigan University Mount Pleasant, MI 48858 (USA) E-mail: du1w@cmich.edu Supporting information for this article is given via a link at the end of the document. 10.1002/cssc.201701870 A cc ep te d M an us cr ip t ChemSusChem This article is protected by copyright. All rights reserved.The capability of a polymer to depolymerize, regenerating its original monomer for further polymerization, is very attractive in terms of sustainability. Recently discovered sugar poly(orthoesters) are an important class of glycopolymer. The high sensitivity of the backbone orthoester linkage toward acidolysis provides a valuable model to study the depolymerization. Herein, a sugar poly(orthoester) is shown to be completely depolymerized under acidic conditions. Interestingly, instead of the original monomer, the depolymerization gave a stable cyclic product (1,6-anhydro glucopyranose) in most cases, which was kinetically and thermodynamically favored. However, this pathway could be inhibited by chemically deactivating a key intermediate and thus favoring the formation of the original monomer. Efficient repolymerizaton of the regenerated monomer is also demonstrated.