Allan S. Hoffman

Design and development of polymers for gene delivery

The lack of safe and efficient gene-delivery methods is a limiting obstacle to human gene therapy. Synthetic gene-delivery agents, although safer than recombinant viruses, generally do not possess the required efficacy. In recent years, a variety of effective polymers have been designed specifically for gene delivery, and much has been learned about their structure–function relationships. With the growing understanding of polymer gene-delivery mechanisms and continued efforts of creative polymer chemists, it is likely that polymer-based gene-delivery systems will become an important tool for human gene therapy.

Applications of thermally reversible polymers and hydrogels in therapeutics and diagnostics

Abstract Polymers and hydrogels have been synthesized which phase separate from aqueous solutions when the temperature is raised through a critical temperature region (LCST). These polymeric systems may be used for the delivery or removal of drugs or other selected molecules and biomolecules. Enzymes immobilized on LCST polymers or within LCST hydrogels may be used in diagnostic assays, biochemical processes or in therapeutics for physiological conditions associated with temperature changes. Antibodies and other binding ligands may also be immobilized on LCST polymers or within LCST hydrogels for therapeutic, diagnostic or separation purposes. This paper describes our ongoing work on these interesting polymers and hydrogels.

A novel approach for preparation of pH-sensitive hydrogels for enteric drug delivery

Abstract A new rationale of preparing pH-sensitive hydrogels based on thermally reversible polymers is proposed, according to which a new class of hydrogels has been synthesized. These special gels are sensitive to both temperature and pH, and respond to the pH change to a much greater extent than hydrogels based on poly(hydroxyethyl methacrylate-coacrylic acid) or poly(acrylamide-co-acrylic acid) hydrogels. At 37°C, they are above their lower critical solution temperature when uncharged. Therefore, at gastric pH (1.4), these gels contain only a moderate amount of water, all of it as bound water. But at enteric pH (6.8–7.4), they swell extensively, permitting sustained enteric delivery of drugs such as indomethacin, which would otherwise cause stomach irritation. An in vitro study has shown that only a negligible amount of indomethacin is released at pH 1.4 in 24 hours, while at pH 7.4 more than 90% of the total drug in the gels is gradually released over ca. 5 hours. The release rate at pH 7.4 increases with the fraction of non-freezing, free water in the gels, which is in parallel with the gel acrylic acid content. An anomalous release profile was observed for all the pH-sensitive heterogeneous hydrogels at this pH, suggesting a swelling-controlled mechanism.

The design and synthesis of polymers for eukaryotic membrane disruption

The intracellular trafficking of drugs is critical to the efficacy of drugs that are susceptible to attack by lysosomal enzymes. It is therefore an important goal to design and synthesize molecules which can enhance the transport of endocytosed drugs from the endosomal compartments to the cytoplasm. The pH of an endosome is lower than that of the cytosol by one to two pH units, depending on the stage of endosomal development. This pH gradient is a key factor in the design of membrane-disruptive polymers which could enhance the endosomal release of drugs. Such polymers should disrupt lipid bilayer membranes at pH 6.5 and below, but should be non-lytic at pH 7.4. We have designed and synthesized pH-sensitive synthetic polymers which efficiently disrupt red blood cells within a sharply defined pH range. One of these polymers, poly(ethyl acrylic acid) (PEAAc) has been previously shown to disrupt synthetic vesicles in a pH-dependent fashion [6]. PEAAc hemolyzes red blood cells with an activity of 10(7) molecules per red blood cell, which is as efficient on a molar basis as the peptide melittin. The mechanism of RBC hemolysis by PEAAc is consistent with the colloid osmotic mechanism. PEAAcs hemolytic activity rises rapidly as the pH decreases from 6.3 to 5.0, and there is no hemolytic activity at pH 7.4. A related polymer, poly(propyl acrylic acid) (PPAAc), was synthesized to test whether making the pendant alkyl group more hydrophobic by adding one methylene group would increase the hemolytic activity. PPAAc was found to disrupt red blood cells 15 times more efficiently than PEAAc at pH 6.1. PPAAc was also not active at pH 7.4 and displayed a pH-dependent hemolysis that was shifted toward higher pHs. Random 1:1 copolymers of ethyl acrylate (EA) and acrylic acid (AAc) (which contain random -COOH and -C(2)H(5) groups that are present and regularly repeat in PEAAc) also displayed significant hemolytic activity, with an efficiency close to PEAAc. These results demonstrate that pH-sensitive synthetic polymers can be molecularly engineered to efficiently disrupt eukaryotic membranes within defined and narrow pH ranges. Thus, these polymers might serve as endosomal disruptive agents with specificities for early or late endosomes.

Target specific and long-acting delivery of protein, peptide, and nucleotide therapeutics using hyaluronic acid derivatives

Hyaluronic acid (HA) is a biodegradable, biocompatible, non-toxic, non-immunogenic and non-inflammatory linear polysaccharide, which has been used for various medical applications such as arthritis treatment, ocular surgery, tissue augmentation, and so on. In this review, the effect of chemical modification of HA on its distribution throughout the body was reported for target specific and long-acting delivery applications of protein, peptide, and nucleotide therapeutics. According to the real-time bio-imaging of HA derivatives using quantum dots (QDot), HA-QDot conjugates with 35mol% HA modification maintaining enough binding sites for HA receptors were mainly accumulated in the liver, while those with 68mol% HA modification losing much of HA characteristics were evenly distributed to the tissues in the body. The results are well matched with the fact that HA receptors are abundantly present in the liver with a high specificity to HA molecules. Accordingly, slightly modified HA derivatives were used for target specific intracellular delivery of nucleotide therapeutics and highly modified HA derivatives were used for long-acting conjugation of peptide and protein therapeutics. HA has been also used as a novel depot system in the forms of physically and chemically crosslinked hydrogels for various protein drug delivery. This review will give you a peer overview on novel HA derivatives and the latest advances in HA-based drug delivery systems of various biopharmaceuticals for further clinical development.

Hydrogels for biomedical applications

Abstract: This paper reviews the composition and synthesis of hydrogels, the character of their absorbed water, and permeation of solutes within their swollen matrices. The most important properties of hydrogels relevant to their biomedical applications are also identified, in particular for use of hydrogels as drug and cell carriers, and as tissue engineering matrices.

Development of a novel endosomolytic diblock copolymer for siRNA delivery

The gene knockdown activity of small interfering RNA (siRNA) has led to their use as target validation tools and as potential therapeutics for a variety of diseases. The delivery of these double-stranded RNA macromolecules has proven to be challenging, however, and in many cases, is a barrier to their deployment. Here we report the development of a new diblock copolymer family that was designed to enhance the systemic and intracellular delivery of siRNA. These diblock copolymers were synthesized using the controlled reversible addition fragmentation chain transfer polymerization (RAFT) method and are composed of a positively-charged block of dimethylaminoethyl methacrylate (DMAEMA) to mediate siRNA condensation, and a second endosomal-releasing block composed of DMAEMA and propylacrylic acid (PAA) in roughly equimolar ratios, together with butyl methacylate (BMA). A related series of diblock compositions were characterized, with the cationic block kept constant, and with the ratio of DMAEMA and PAA to BMA varied. These carriers became sharply hemolytic at endosomal pH regimes, with increasing hemolytic activity seen as the percentage of BMA in the second block was systematically increased. The diblock copolymers condensed siRNA into 80-250 nm particles with slightly positive Zeta potentials. SiRNA-mediated knockdown of a model protein, namely glyceraldehyde 3-phosphate dehydrogenase (GAPDH), in HeLa cells generally followed the hemolytic activity trends, with the most hydrophobic second block (highest BMA content) exhibiting the best knockdown. This pH-responsive carrier designed to mediate endosomal release shows significant promise for the intracellular delivery of siRNA.

An AB block copolymer of oligo(methyl methacrylate) and poly(acrylic acid) for micellar delivery of hydrophobic drugs

An AB block copolymer of oligo(methyl methacrylate) (oMMA) and poly(acrylic acid) (PAAc) has been synthesized. The block copolymer forms micelles in an aqueous medium, as confirmed by a fluorescence probe technique using pyrene. Doxorubicin hydrochloride was incorporated into the micelle and the release profile of doxorubicin hydrochloride was investigated. Slow and prolonged release of doxorubicin hydrochloride from the micelle was observed. The AB block copolymer micelle can be useful for prolonged mucosal drug delivery of hydrophobic drugs.

Really smart bioconjugates of smart polymers and receptor proteins

Over the past 18 years we have been deeply involved with the synthesis and applications of stimuli-responsive polymer systems, especially polymer-biomolecule conjugates. This article summarizes our work with one of these conjugate systems, specifically polymer-protein conjugates. We include conjugates prepared by random polymer conjugation to lysine amino groups, and also those prepared by site-specific conjugation of the polymer to specific amino acid sites that are genetically engineered into the known amino acid sequence of the protein. We describe the preparation and properties of thermally sensitive random conjugates to enzymes and several affinity recognition proteins. We have also prepared site-specific conjugates to streptavidin with temperature-sensitive polymers, pH-sensitive polymers, and light-sensitive polymers. The preparation of these conjugates and their many fascinating applications are reviewed in this article.

Thermally reversible hydrogels. III: Immobilization of enzymes for feedback reaction control

Hydrogels have been synthesized which exhibit a lower critical solution temperature (LCST). The gels shrink and deswell in aqueous solutions when the temperature is raised through their LCST. This phenomenon is reversible on cooling below the LCST where the gels reswell and expand. An enzyme has been immobilized within a thermally reversible hydrogel for the first time. We have shown that the enzyme activity is “shut off” when the gel is raised above its LCST. This phenomenon is reversible and the enzyme-gel regains activity below its LCST. Such catalytic hydrogels may be used to control reaction rates and temperatures by a thermal feedback mechanism.