Evaluating the oral delivery of GalNAc-conjugated siRNAs in rodents and non-human primates.

Oral delivery is the most widely used and convenient route of administration of medicine. However, oral administration of hydrophilic macromolecules is commonly limited by low intestinal permeability and pre-systemic degradation in the gastrointestinal (GI) tract. Overcoming some of these challenges allowed emergence of oral dosage forms of peptide-based drugs in clinical settings. Antisense oligonucleotides (ASOs) have also been investigated for oral administration but despite the recent progress, the bioavailability remains low. Given the advancement with highly potent and durable trivalent N-acetylgalactosamine (GalNAc)-conjugated small interfering RNAs (siRNAs) via subcutaneous (s.c.) injection, we explored their activities after oral administration. We report robust RNA interference (RNAi) activity of orally administrated GalNAc-siRNAs co-formulated with permeation enhancers (PEs) in rodents and non-human primates (NHPs). The relative bioavailability calculated from NHP liver exposure was <2.0% despite minimal enzymatic degradation in the GI. To investigate the impact of oligonucleotide size on oral delivery, highly specific GalNAc-conjugated single-stranded oligonucleotides known as REVERSIRs with different lengths were employed and their activities for reversal of RNAi effect were monitored. Our data suggests that intestinal permeability is highly influenced by the size of oligonucleotides. Further improvements in the potency of siRNA and PE could make oral delivery of GalNAc-siRNAs as a practical solution.

Design of an injectable synthetic and biodegradable surgical biomaterial.

We report the design of an injectable synthetic and biodegradable polymeric biomaterial comprised of polyethylene glycol and a polycarbonate of dihydroxyacetone (MPEG-pDHA). MPEG-pDHA is a thixotropic physically cross-linked hydrogel, displays rapid chain relaxation, is easily extruded through narrow-gauge needles, biodegrades into inert products, and is well tolerated by soft tissues. We demonstrate the clinical utility of MPEG-pDHA in the prevention of seroma, a common postoperative complication following ablative and reconstructive surgeries, in an animal model of radical breast mastectomy. This polymer holds significant promise for clinical applicability in a host of surgical procedures ranging from cosmetic surgery to cancer resection.

Poly(carbonate-ester)s of dihydroxyacetone and lactic acid as potential biomaterials.

The synthesis of new polymeric biomaterials using biocompatible building blocks is important for the advancement of the biomedical field. We report the synthesis of statistically random poly(carbonate-ester)s derived from lactic acid and dihydroxyacetone by ring-opening polymerization. The monomer mole feed ratio and initiator concentration were adjusted to create various copolymer ratios and molecular weights. A dimethoxy acetal protecting group was used to stabilize the dihydroxyacetone and was removed using elemental iodine and acetone at reflux to produce the final poly(lactide-co-dihydroxyacetone) copolymers. The characteristics of the copolymers in their protected and deprotected forms were characterized by (1)H NMR, (13)C NMR, GPC, TGA, and DSC. Hydrolytic degradation of the deprotected copolymers was tracked over an 8-week time frame. The results show that faster degradation occurred with increased carbonate content in the copolymer backbone. The degradation pattern of the copolymers was visualized using SEM and revealed a trend toward surface erosion as the primary mode of degradation.

Prediction of in vivo performance of oral extended release formulations prior to clinical evaluation: A case study for enteric coated polymeric beads formulation.

To reduce cost and time for product development, an ideal strategy for the development of oral extended release (ER) product is to identify the desired formulation with minimum needsfor clinical evaluation. The aim of this work was to demonstrate the feasibility of adopting a "prediction-then-validation" strategy for the development of oral ER formulations. Instead of the traditional approach using multiple ER formulations for IVIVC development, an enteric-coated fast release formulation was successfully utilized for the development of a biopredictive tool to estimate the drug release from enteric coated polymeric ER formulations in the intestine. A TS1 (time scale factor between Tvitro and Tvivo equals to 1) system was designed and developed, based on which the in vivo pharmacokinetic (PK) performance of ER formulations in dog and in human were well predicted prior to in vivo evaluations. The model further passed a posteriori validation using the criteria for level A IVIVC and, as designed, provided a Tscale value of 1 for the IVIVC model.

Engineered particles demonstrate improved flow properties at elevated drug loadings for direct compression manufacturing.

Optimizing powder flow and compaction properties are critical for ensuring a robust tablet manufacturing process. The impact of flow and compaction properties of the active pharmaceutical ingredient (API) becomes progressively significant for higher drug load formulations, and for scaling up manufacturing processes. This study demonstrated that flow properties of a powder blend can be improved through API particle engineering, without critically impacting blend tabletability at elevated drug loadings. In studying a jet milled API (D50=24µm) and particle engineered wet milled API (D50=70µm and 90µm), flow functions of all API lots were similarly poor despite the vast difference in average particle size (ffc<4). This finding strays from the common notion that powder flow properties are directly correlated to particle size distribution. Upon adding excipients, however, clear trends in flow functions based on API particle size were observed. Wet milled API blends had a much improved flow function (ffc>10) compared with the jet milled API blends. Investigation of the compaction properties of both wet and jet milled powder blends also revealed that both jet and wet milled material produced robust tablets at the drug loadings used. The ability to practically demonstrate this uncommon observation that similarly poor flowing APIs can lead to a marked difference upon blending is important for pharmaceutical development. It is especially important in early phase development during API selection, and is advantageous particularly when material-sparing techniques are utilized.

Characterizing the structure/function parameter space of hydrocarbon-conjugated branched polyethylenimine for DNA delivery in vitro.

Polyethylenimine is a popular DNA transfection reagent, and many approaches have been explored to further enhance its transfection efficiency. Substitution of branched polyethylenimine's primary amine groups is an attractive approach because it is amenable to a variety of chemistries and is also implicated as a primary factor in its cytotoxicity. The purpose of this work was to serially substitute saturated hydrocarbons to branched polyethylenimine and determine what structure/function relationships exist between the hydrocarbon length and its degree of substitution, relative to transfection efficiency in multiple cell lines. Specifically, acetate, butanoate and hexanoate were conjugated to branched polyethylenimine (M(w) = 25,000) using an aqueous condensation protocol. Transfections were performed in culture using HeLa, NIH/3T3 and Clone 9 cell lines. Biophysical characteristics of the polyelectrolyte complexes were also measured (hydrodynamic diameter, relative binding affinity) and correlated to transfection efficiency. The results show that substitution of the primary amines generally increases transfection efficiency relative to unconjugated polyethylenimine, but increasing the degree of substitution beyond approximately 25 mol% generally decreases transfection efficiency from the optimum. Additionally, increasing hydrocarbon length generally decreased transfection efficiency. There was little correlation between particle size and binding efficiency to transfection efficiency.

A rapidly resorbable hemostatic biomaterial based on dihydroxyacetone.

We have developed a rapid acting, rapidly resorbable, non-toxic, topical hemostatic agent comprised of a PEGylated, polymerized sequence of dihydroxyacetone (MPEG-pDHA) that is highly effective in vivo. Twenty-eight Sprague-Dawley rats underwent left lateral hepatectomy. To the cut edge of the liver, rats received MPEG-pDHA (50 mg), normal saline (0.5 mL), or Instat (50 mg), a commercially available hemostatic compound. Bleeding time and total blood loss were quantified. Coagulation studies and scanning electron microscopy were performed on phlebotomized blood combined with MPEG-pDHA. Rats treated with MPEG-pDHA had significantly decreased bleeding time (97 s) and total blood loss (1.35 g) compared to normal saline (464 s and 3.83 g, p < 0.05 for each), and a significantly shorter bleeding time compared to Instat (165 s, p < 0.05). Histology confirmed that all MPEG-pDHA was metabolized within 3 weeks. The addition of MPEG-pDHA to whole blood did not significantly affect prothrombin time (12.0 s vs. 13.2 s, p = 0.130), partial thromboplastin time (27.0 s vs. 21.8 s, p = 0.118), or thrombin clotting time. MPEG-pDHA is an effective and rapidly resorbable hemostatic agent that may find broad hemostatic application in a wide range of surgical procedures.

Materials in surgery: a review of biomaterials in postsurgical tissue adhesion and seroma prevention.

Postoperative tissue adhesion is a complex inflammatory disorder in which tissues that normally remain separated in the body grow into each other. Seroma is a common postoperative complication that arises when serous fluid collects in the space generated following surgeries that require extensive dissection and that create large empty spaces. Postsurgical tissue adhesion and seroma formation are two serious surgical complications that have received more attention recently from the biomaterials community. This paper provides a review of the pathogenesis and treatment of these surgical complications, with a thorough overview of biomaterial-based treatment and prevention methods.

A functionalizable biomaterial based on dihydroxyacetone, an intermediate of glucose metabolism.

A biomaterial and its potential degradation products should be biocompatible, nontoxic, and removed by the body upon expiration of its functional lifetime. One historically successful approach is to create new materials from biomolecules that naturally occur in the human body. Herein, we report the synthesis and characterization of a polycarbonate based on dihydroxyacetone, a 3-carbon ketose, and an intermediate in the glucose metabolic pathway. The polymer was synthesized in a range of molecular weights ( approximately 8000 to approximately 37,500) by ring-opening polymerization. The C2 carbonyl of dihydroxyacetone is reactive to amines, and this reactivity was used to functionalize the polymer's surface in a one-step reaction by reductive amination. Additionally, contact angle measurements show the surface of poly(2-oxypropylene carbonate) is hydrophilic even though it is insoluble in water. Mechanical analysis of the polymer revealed it is exceptionally strong for an aliphatic polycarbonate. Specifically, poly(2-oxypropylene carbonate), M(w) 37 500, yielded a Young's modulus of 0.5 GPa and a compressive yield stress of 50 MPa. These values equal or exceed those of cancellous bone with similar dimensions.

Diblock copolymers based on dihydroxyacetone and ethylene glycol: synthesis, characterization, and nanoparticle formulation.

Polymeric biomaterials have played an integral role in tissue engineering, biomedical devices, and targeted drug delivery. Block copolymers are especially important because their physical and chemical properties can be controlled by adjusting the ratio, size, and type of constituting blocks. Herein, the synthesis and characterization of diblock copolymers composed of poly(ethylene glycol) and a polycarbonate based on the metabolic intermediate, dihydroxyacetone, are reported. The length of the dihydroxyacetone-based block was controlled by adjusting the reactant feed ratios and initiator injection conditions. Intermediates and final products were characterized via (1)H NMR, GPC, DSC, TGA, and diffusion-ordered NMR spectroscopy. The dihydroxyacetone-based hompolymer is insoluble in water and most organic solvents, but is hydrophilic in nature. This, coupled with poly(ethylene glycol)'s solubility characteristics, allows the block copolymer to form nanoparticles in aqueous and organic anti-solvents. Dynamic light scattering and TEM results indicated the formation of spherical nanoparticles.