Jerald E. Dumas

Synthesis, mechanical properties, biocompatibility, and biodegradation of polyurethane networks from lysine polyisocyanates

Bone defects, such as compressive fractures in the vertebral bodies, are frequently treated with acrylic bone cements (e.g., PMMA). Although these biomaterials have sufficient mechanical properties for fixing the fracture, they are non-degradable and do not remodel or integrate with host tissue. In an alternative approach, biodegradable polyurethane (PUR) networks have been synthesized that are designed to integrate with host tissue and degrade to non-cytotoxic decomposition products. PUR networks have been prepared by two-component reactive liquid molding of low-viscosity quasi-prepolymers derived from lysine polyisocyanates and poly(epsilon-caprolactone-co-DL-lactide-co-glycolide) triols. The composition, thermal transitions, and mechanical properties of the biomaterials were measured. The values of Youngs modulus ranged from 1.20-1.43 GPa, and the compressive yield strength varied from 82 to 111 MPa, which is comparable to the strength of PMMA bone cements. In vitro, the materials underwent controlled biodegradation to non-cytotoxic decomposition products, and supported the attachment and proliferation of MC3T3 cells. When cultured in osteogenic medium on the PUR networks, MC3T3 cells deposited mineralized extracellular matrix, as evidenced by von Kossa staining and tetracycline labeling. Considering the favorable mechanical and biological properties, as well as the low-viscosity of the reactive intermediates used to prepare the PUR networks, these biomaterials are potentially useful as injectable, biodegradable bone cements for fracture healing.

Synthesis, characterization, and remodeling of weight-bearing allograft bone/polyurethane composites in the rabbit

The process of bone healing requires the restoration of both anatomy and physiology, and there is a recognized need for innovative biomaterials that facilitate remodeling throughout this complex process. While porous scaffolds with a high degree of interconnectivity are known to accelerate cellular infiltration and new bone formation, the presence of pores significantly diminishes the initial mechanical properties of the materials, rendering them largely unsuitable for load-bearing applications. In this study, a family of non-porous composites has been fabricated by reactive compression molding of mineralized allograft bone particles (MBPs) with a biodegradable polyurethane (PUR) binder, which is synthesized from a polyester polyol and a lysine-derived polyisocyanate. At volume fractions exceeding the random close-packing limit, the particulated allograft component presented a nearly continuous osteoconductive pathway for cells into the interior of the implant. By varying the molecular weight of the polyol and manipulating the surface chemistry of the MBP via surface demineralization, compressive modulus and strength values of 3-6 GPa and 107-172 MPa were achieved, respectively. When implanted in bilateral femoral condyle plug defects in New Zealand White rabbits, MBP/PUR composites exhibited resorption of the allograft and polymer components, extensive cellular infiltration deep into the interior of the implant, and new bone formation at 6 weeks. While later in vivo timepoints are necessary to determine the ultimate fate of the MBP/PUR composites, these observations suggest that allograft bone/polymer composites have potential for future development as weight-bearing devices for orthopedic applications.

Injectable reactive biocomposites for bone healing in critical-size rabbit calvarial defects

Craniofacial injuries can result from trauma, tumor ablation, or infection and may require multiple surgical revisions. To address the challenges associated with treating craniofacial bone defects, an ideal material should have the ability to fit complex defects (i.e. be conformable), provide temporary protection to the brain until the bone heals, and enhance tissue regeneration with the delivery of biologics. In this study, we evaluated the ability of injectable lysine-derived polyurethane (PUR)/allograft biocomposites to promote bone healing in critical-size rabbit calvarial defects. The biocomposites exhibited favorable injectability, characterized by a low yield stress to initiate flow of the material and a high initial viscosity to minimize the adverse phenomena of extravasation and filter pressing. After injection, the materials cured within 10-12 min to form a tough, elastomeric solid that maintained mechanical integrity during the healing process. When injected into a critical-size calvarial defect in rabbits, the biocomposites supported ingrowth of new bone. The addition of 80 µg mL(-1) recombinant human bone morphogenetic protein-2 (rhBMP-2) enhanced new bone formation in the interior of the defect, as well as bridging of the defect with new bone. These observations suggest that injectable reactive PUR/allograft biocomposites are a promising approach for healing calvarial defects by providing both mechanical stability as well as local delivery of rhBMP-2.

Biocompatibility and chemical reaction kinetics of injectable, settable polyurethane/allograft bone biocomposites

Injectable and settable bone grafts offer significant advantages over pre-formed implants due to their ability to be administered using minimally invasive techniques and to conform to the shape of the defect. However, injectable biomaterials present biocompatibility challenges due to the potential toxicity and ultimate fate of reactive components that are not incorporated in the final cured product. In this study the effects of stoichiometry and triethylenediamine (TEDA) catalyst concentration on the reactivity, injectability, and biocompatibility of two component lysine-derived polyurethane (PUR) biocomposites were investigated. Rate constants were measured for the reactions of water (a blowing agent resulting in the generation of pores), polyester triol, dipropylene glycol (DPG), and allograft bone particles with the isocyanate-terminated prepolymer using an in situ attenuated total reflection Fourier transform infrared spectroscopy technique. Based on the measured rate constants, a kinetic model predicting the conversion of each component with time was developed. Despite the fact that TEDA is a well-known urethane gelling catalyst, it was found to preferentially catalyze the blowing reaction with water relative to the gelling reactions by a ratio >17:1. Thus the kinetic model predicted that the prepolymer and water proceeded to full conversion, while the conversions of polyester triol and DPG were <70% after 24h, which was consistent with leaching experiments showing that only non-cytotoxic polyester triol and DPG were released from the reactive PUR at early time points. The PUR biocomposite supported cellular infiltration and remodeling in femoral condyle defects in rabbits at 8weeks, and there was no evidence of an adverse inflammatory response induced by unreacted components from the biocomposite or degradation products from the cured polymer. Taken together, these data underscore the utility of the kinetic model in predicting the biocompatibility of reactive biomaterials.

Synthesis, characterization of calcium phosphates/polyurethane composites for weight-bearing implants†

Calcium phosphate (CaP)/polymer composites have been studied as an alternative graft material for the treatment of bone defects. In this study, lysine-triisocyanate-based polyurethane (PUR) composites were synthesized from both hydroxyapatite (HA) and β-tricalcium phosphate (TCP) to reduce the brittleness of CaP and increase the bioactivity of the polymer. The mechanical properties and in vitro cellular response were investigated for both HA/PUR and TCP/PUR composites. The composites were implanted in femoral defects in rats, and in vivo bioactivity was evaluated by X-rays, micro-computed tomography (μCT), and histological sections. In biomechanical testing, PUR improved the mechanical properties of the CaP, thus rendering it potentially suitable for weight-bearing applications. In vitro cell culture studies showed that CaP/PUR composites are biocompatible, with β-TCP enhancing the cell viability and proliferation relative to HA. CaP/PUR composites also supported the differentiation of osteoblastic cells on the materials. When implanted in rat femoral defects, the CaP/PUR composites were biocompatible and osteoconductive with no adverse inflammatory response, as evidenced by X-rays, μCT images, and histological sections. Additionally, a histological examination showed evidence of cellular infiltration and appositional remodeling. These results suggest that CaP/PUR composites could be potentially useful biomaterials for weight-bearing orthopaedic implants.

Abstract C60: Snail transcription factor contributes to bone metastasis in prostate and breast cancer cells

Prostate cancer that is hormone refractory and has metastasized preferentially to bone is the main cause of prostate cancer death, especially in African American men. Receptor activator of nuclear factor kappa B ligand (RANKL) and its receptor (RANK) contributes to bone metastatic lesions and bisphosphonates such as Zometa and Fosamax have been used as antagonists of RANKL for the treatment of breast and prostate cancer metastasis. African American men have the highest bone mineral density compared to any other race, and the role this may play in prostate cancer metastasis to bone is not clear. A better understanding of bone metastasis may lead to alternative treatment options for metastatic prostate cancer. Snail transcription factor is important early in development and in cancer cells and promotes cancer cell migration and progression by inducing epithelial mesenchymal transition (EMT). We have observed increased expression of Snail in prostate cancer bone metastatic human patient samples. We hypothesized that Snail can mediate EMT-mediated prostate cancer migration towards bone of high bone mineral density and mediate the vicious cycle of tumor-tumor microenvironment reciprocal interactions through calcium and RANKL signaling. We generated an EMT model for prostate and breast cancer utilizing the ARCaP human prostate and MCF-7 breast cancer cells overexpressing Snail and identified increased RANKL expression that was associated with increased osteoclastogenesis both in vitro and in vivo, as well as decreased bone volume and density. We utilized pre-molded bone discs which are allograft bone/polyurethane (PUR) composite bone void fillers with tunable properties that have advantage over existing bone implant models in that it contains bigger pore sizes that support rapid cellular infiltration and remodeling. We treated the bone discs with hydrochloric acid which decreased the bone density to a ratio of 1:1.18 for low (HCL-treated): high (untreated) bone density, which is quite close to the 1: 1.2 ratio seen in Caucasian vs African American men. Incubation of these bone discs with prostate or breast cancer cells overexpressing Snail led to increased calcium release from bone of high density as compared to low density. We are currently testing whether this increased calcium release in response to Snail may promote paracrine cell proliferation. Since Snail is not required by adult cells except during injury, targeting Snail that is mainly expressed by cancer cells may antagonize metastatic lesions in bone without affecting normal bone in other areas of the body. Citation Format: Basil A. Smith, Veronica Henderson, Christopher Coke, Jerald Dumas, Cimona Hinton, Manu Platt, Leland K. Chung, Majd Zayzafoon, Valerie A. Odero-Marah. Snail transcription factor contributes to bone metastasis in prostate and breast cancer cells. [abstract]. In: Proceedings of the Sixth AACR Conference: The Science of Cancer Health Disparities; Dec 6–9, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2014;23(11 Suppl):Abstract nr C60. doi:10.1158/1538-7755.DISP13-C60