Michel Wathier

Synthesis and properties of supramolecular ionic networks.

The synthesis of a supramolecular ionic network, its physical properties, and the use of this network property to form macroscopic porphyrin fibers are described. These ionic networks are compared to ionic liquids. Current ionic liquid compositions have a charge and molar ratio of 1:1 where an anionic species is matched with a cationic species; however, alteration of this molar ratio while maintaining the charge ratio of 1:1 by using multivalent cationic/anionic molecular pairs affords new ionically cross-linked networks with interesting properties.

Synthesis and characterization of dendron cross-linked PEG hydrogels as corneal adhesives.

In pursuit of a wound-specific corneal adhesive, hydrogels formed by the reaction of propionaldehyde, butyraldehyde, or 2-oxoethyl succinate-functionalized poly(ethylene glycol) (PEG) with a peptide-based dendritic cross-linker (Lys(3)Cys(4)) were characterized. These macromers react within minutes of mixing to form transparent and elastic hydrogels with in vitro degradation times that range from hours to months based on the type of bonds formed during the cross-linking reaction, either thiazolidine or pseudoproline. The mechanical properties of these materials, determined via parallel plate rheology, were dependent on the polymer concentration, as was the hydrogel adhesive strength, which was determined by lap shear adhesive testing. In addition, these hydrogels were efficacious in closing ex vivo 4.1 mm central corneal lacerations: wounds closed with these hydrogel adhesives were able to withstand intraocular pressure values equivalent to, or in excess of, those obtained by closing the wounds with suturing.

A Large-Molecular-Weight Polyanion, Synthesized via Ring-Opening Metathesis Polymerization, as a Lubricant for Human Articular Cartilage

A large-molecular-weight polyanion is found to possess lubricating properties for cartilage. The polyanion, sodium poly(7-oxanorbornene-2-carboxylate), is synthesized by ring-opening metathesis polymerization of methyl 5-oxanorbornene-2-carboxylate. When dissolved in aqueous solution and applied to the surface of human cartilage it reduces the friction at the interface and acts as a lubricant. Its performance is similar to that of synovial fluid and superior to those of saline and Synvisc in an ex vivo human cartilage plug-on-plug model. The polymer is also not readily degraded by hyaluronidase or cytotoxic to human chondrocytes in vitro. As such, this polymer is a new type of viscosupplement, and the results provide insight into the design requirements for synthesizing highly efficacious synthetic biolubricants.

In Situ Polymerized Hydrogels for Repairing Scleral Incisions Used in Pars Plana Vitrectomy Procedures

Crosslinked polymer networks possessing a high water content, otherwise known as hydrogels, are multipurpose materials for medical applications in areas such as drug delivery, tissue engineering, and wound healing. To form such hydrogels, dendritic or highly branched macromers are advantageous. These macromers provide opportunities to create hydrogels at low polymer concentration, to control swelling, and to vary the hydrogel mechanical properties. These favorACHTUNGTRENNUNGable attributes arise with dendritic polymers because of the well-defined composition, the large number of endgroups, the physiochemical properties (such as low viscosity), and the preparation methods that allow for precise structural control and optimization capabilities. We are synthesizing dendritic macromolecules composed of biocompatible building blocks and evaluating the corresponding hydrogels as ocular adhesives. Using a photocurable hydrogel system based on poly(glycerol-succinic acid)-polyethylene glycol hybrid dendritic-linear macromolecules, full-thickness 4.1 mm corneal lacerations in enucleated eyes and chicken eyes in vivo, as well as secured LASIK flaps in vitro, have been successfully repaired. In addition to light-activated hydrogel formation, other crosslinking strategies that quickly afford a hydrogel adhesive are also explored. Herein, we report the synthesis of lysineterm ACHTUNGTRENNUNGinated peptide dendrimers and dendrons, the formation of crosslinked hydrogels with a poly(ethylene glycol) di-activated ester, the analysis of hydrogel mechanical properties, and the closure of a sclerotomy incision—the wound created during a typical vitrectomy procedure. The dendrons ([G1]-Lys-NH2 and [G2]-Lys-NH2) and dendriACHTUNGTRENNUNGmers (([G1]-Lys-NH2)2-PEG and ([G2]-Lys-NH2)2-PEG) used for hydrogel formation were synthesized as shown in Scheme 1. Several amide-coupling approaches were explored (BOP, DCC, EDC, and oxalyl chloride), and the pentafluorophenol-ester strategy was found to yield the highest amide coupling reactions. Thus, we first prepared the pentafluorophenol-ester of ZLys(Z)OH and BocLysACHTUNGTRENNUNG(Boc)OH using N-N’-dicyclohexylcarbodiACHTUNGTRENNUNGimide (DCC) and 2,3,4,5,6-pentafluorophenol (PFP) in CH2Cl2. After crystallization in CH2Cl2/hexane, white crystalline products were obtained (98% and 92% yield, respectively). Next, ZLys(Z)OPFP was coupled to LysOMe·2HCl in the presence of diisopropylethylamine (DIEA) and 1-hydroxybenzotriazole (HOBT) to prevent racemization, to give 1. Compound 1 was purified by precipitation in ether and obtained in 98% yield. The Z-amino protecting groups were cleaved by hydrogenolysis (Pd/C, H2) in methanol (99% yield) and the amine functionality was subsequently acidified using 1m HCl (99% yield) to afford the [G1]-Lys-NH2 dendron, 2. The larger dendron [G2]-Lys-NH2, 3, was synthesized by reacting 2 with BocLysACHTUNGTRENNUNG(Boc)PFP in the presence of HOBT and DIEA, followed by treatment with TFA to remove the Boc protecting groups (70% and 99% yield, respectively). The protected intermediate was purified by silica gel chromatography (CH2Cl2/MeOH=95:5). The corresponding product obtained using ZLys(Z)OPFP was difficult to isolate and purify. The intermediates prepared in the dendron synthesis were also used to prepare the ([G1]-Lys-NH2)2-PEG, 4 and ([G2]-Lys-NH2)2-PEG 5 dendrimers. ZLys(Z)OPFP was coupled to diamino polyethylene glycol [MW=3400] in the presence of DIEA and HOBT in CH2Cl2, to afford ([G1]-Lys-NH2)2-PEG 4. The product was washed with water and purified by precipitation in ether (95% yield). The deprotection of the Z groups was achieved by hydrogenolysis (Pd/C, H2) in methanol followed by precipitation in ether to afford ([G1]-Lys-NH2)2-PEG 4 as a white powder (99% yield). The previously prepared, protected dendron 1 was used for the synthesis of the ([G2]-Lys-NH2)2-PEG dendrimer, 5. The methyl ester of 1 was hydrolyzed by saponACHTUNGTRENNUNGification with 1m NaOH in methanol followed by neutralization with 1m HCl (85% yield). The pentafluorophenol-ester of this acid was obtained by treatment with 2,3,4,5,6-pentafluorophenol and DCC in CH2Cl2 to afford a white powder after crystallization (95% yield). This activated ester dendron was then coupled to diamino polyethylene glycol [MW=3400] in CH2Cl2 to give, after precipitation in ether, a white powder in 96% yield. The Z groups were removed by hydrogenolysis and the product, ([G2]-Lys-NH2)2-PEG dendron 5, was isolated by precipitation in ether (99% yield). The structural identities of the various intermediates and dendritic macromolecules were determined by H, C NMR, and mass spectrometry (see Supporting Information). A hydrogel starts to form within one minute of mixing aqueous solutions of the dendrimer or dendron with a poly(ethylene glycol) disuccinimidyl proprionate, 6 ; PEG-NHS. The hydrogels are optically transparent and Figure 1 shows the 5 :6 hydrogel. All the hydrogels are formed using a 1:1 reactive group stoichiometry. The crosslinking reaction can be followed by infrared spectroscopy (IR), as shown in Figure 2a. For example, the IR stretch for the PEG-NHS at 1733 cm 1 disappears over time and the reaction between dendron 2 and 6 (1:1; 18 w/w%) is complete within 5000 seconds. As expected, the hydrogels swell when placed in an aqueous solution. Hydrogel swelling is linear with increasing concentrations of polymer [a] Dr. M. Wathier, Prof. M. W. Grinstaff Departments of Biomedical Engineering and Chemistry, Metcalf Center for Science and Engineering, Boston University, Boston, MA 02215 (USA) Fax: (+1)617-358-3186 E-mail : mgrin@bu.edu [b] Dr. M. S. Johnson, Dr. M. A. Carnahan, Dr. C. Baer, Dr. B. W. McCuen, Dr. T. Kim Department of Ophthalmology, Duke University Medical Center Durham NC 27710 (USA) [**] This work was supported by the NIH NEI. Supporting information for this article is available on the WWW under http://www.chemmedchem.org or from the author.

Acidic polysaccharide mimics via ring-opening metathesis polymerization.

An efficient and general synthetic strategy for the preparation of high-molecular-weight hydrophilic polymers bearing both carboxylic acid and hydroxyl pendant groups is described. Specifically, poly(5,6-dihydroxyoxanorbornane carboxylic acid) with molecular weight ranging from ∼100 000 to 5 000 000 g/mol was prepared by ring-opening metathesis polymerization of methyl 5-oxanorbornene-2-carboxylate in the presence of Grubbs catalyst II and subsequently modified to tune the hydrophobic/hydrophilic properties by the introduction of either hydroxyl or carboxylic acid functionalities. These polymers mimic the natural acidic polysaccharide alginate and form hydrogels with polylysine. These polymers belong to a class of carbohydrate-like polymers, which are of interest for investigating the relationships between chemical structure and rheological properties as well as for providing new synthetic polysaccharide substitutes for applications in the biotechnology and pharmaceutical industries.

Novel tissue adhesives to secure laser in situ keratomileusis flaps

Purpose: To evaluate 2 novel biodendrimer tissue adhesives in sealing and securing laser in situ keratomileusis (LASIK) flaps. Setting: Duke University Eye Center, Durham, North Carolina, USA. Methods: Laser in situ keratomileusis flaps were created in 10 human eye‐bank eyes using the Hansatome microkeratome system (Bausch & Lomb). These eyes were divided into 2 groups. Flaps in the first group (n = 4) were secured with a laser‐activated biodendrimer adhesive along the flap edge. In the second group (n = 6), the flaps were secured with a self‐gelling dendritic adhesive. Dry Merocel sponges (Medtronic Solan) were used to test the strength of flap adherence in both groups. Further testing was performed in the second group. The hinges of these flaps were cut with a scalpel blade and fluorescein dye was injected under the flap to observe potential dye leakage along the flap edge. Results: Laser in situ keratomileusis flaps sealed with both adhesives were secure with no flap dislocation. There was no leakage of fluorescein dye observed in the second group. Both adhesives were easy to apply, clear when dry, and had a soft rubbery consistency. Conclusions: Two novel biodendrimer adhesives successfully sealed and secured LASIK flaps. These adhesives may prove to be an effective alternative for treating LASIK flap complications such as epithelial ingrowth or flap dislocation.

Real Time Imaging of Supramolecular Assembly Formation via Programmed Nucleolipid Recognition

Supramolecular assembly formation resulting from molecular recognition between complementary nucleolipids has been visualized in real time at the micrometer scale.

In vitro sealing of clear corneal cataract incisions with a novel biodendrimer adhesive.

OBJECTIVE To determine if a biodendrimer adhesive will seal a clear corneal cataract incision. DESIGN An experimental study in which 2.75-mm clear corneal cataract incisions were made in 8 human donor eyes. The corneas were mounted on an artificial anterior chamber. Leaking pressure was determined in 6 corneas. These corneas were then treated with adhesive and leaking pressure was again measured in 4 of them. India ink was then applied to the 2 remaining treated and the 2 untreated corneas. Chamber pressure was cycled between 100 and 0 mm Hg. Optical coherence tomography was used to visualize the wound dynamics of a ninth cornea treated with adhesive, mounted, and pressure cycled in a similar fashion. RESULTS The mean (SD) leaking pressure was 77 (14) mm Hg for the nonsealed wounds and 142 (22) mm Hg for the adhesive-sealed wounds. india ink entered the nonsealed wounds and anterior chamber when the intraocular pressure was cyclically raised and lowered, whereas no india ink entered the adhesive-sealed wounds. The optical coherence tomography-visualized corneal wound did gape under pressure cycling, but the adhesive remained intact and stretched to conform to the wound. CONCLUSIONS Biodendrimer adhesives may be used to seal cataract wounds to prevent leakage and influx of fluid.

Diphosphonium Ionic Liquids as Broad Spectrum Antimicrobial Agents

Purpose: One of the most disturbing trends in recent years is the growth of resistant strains of bacteria with the simultaneous dearth of new antimicrobial agents. Thus, new antimicrobial agents for the use on the ocular surface are needed. Methods: We synthesized a variety of ionic liquid compounds, which possess 2 positively charged phosphonium groups separated by 10 methylene units in a “bola”-type configuration. We tested these compounds for antimicrobial activity versus a variety of ocular pathogens, as well as their cytoxicity, in vitro in a corneal cell line and in vivo in mice. Results: The ionic liquid Di-Hex C10 demonstrated broad in vitro antimicrobial activity at low micromolar concentrations versus gram-negative and gram-positive organisms, including methicillin-resistant Staphylococcus aureus strains and ocular fungal pathogens. Treatment with Di-Hex C10 resulted in bacterial killing in as little as 15 minutes in vitro. Di-Hex C10 showed little cytotoxicity at 1 &mgr;M versus a corneal epithelial cell line or at 10 &mgr;M in a mouse corneal wound model. We also show that this bis-phosphonium ionic liquid structure is a key because a comparable monophosphonium ionic liquid is cytotoxic to both bacteria and corneal epithelial cells. Conclusions: Here, we report the first use of dicationic bis-phosphonium ionic liquids as antimicrobial agents. Our data suggest that diphosphonium ionic liquids may represent a new class of broad-spectrum antimicrobial agents for the use on the ocular surface.

Synthesis and Preclinical Characterization of a Cationic Iodinated Imaging Contrast Agent (CA4+) and Its Use for Quantitative Computed Tomography of Ex Vivo Human Hip Cartilage

Contrast agents that go beyond qualitative visualization and enable quantitative assessments of functional tissue performance represent the next generation of clinically useful imaging tools. An optimized and efficient large-scale synthesis of a cationic iodinated contrast agent (CA4+) is described for imaging articular cartilage. Contrast-enhanced CT (CECT) using CA4+ reveals significantly greater agent uptake of CA4+ in articular cartilage compared to that of similar anionic or nonionic agents, and CA4+ uptake follows Donnan equilibrium theory. The CA4+ CECT attenuation obtained from imaging ex vivo human hip cartilage correlates with the glycosaminoglycan content, equilibrium modulus, and coefficient of friction, which are key indicators of cartilage functional performance and osteoarthritis stage. Finally, preliminary toxicity studies in a rat model show no adverse events, and a pharmacokinetics study documents a peak plasma concentration 30 min after dosing, with the agent no longer present in vivo at 96 h via excretion in the urine.