Zhen Gu, UNC at Chapel Hill | NC State
Samir Mitragotri, University of California, Santa Barbara
Chenjie Xu, Nanyang Technological University
Symposium Support Aldrich Materials Science
KK2: Nanomaterials for Tissue Engineering
Tuesday PM, April 07, 2015
Park Central Hotel, 2nd Floor, Metropolitan I
2:30 AM - *KK2.01
Next-Gen Bio-Inspired Tissue Adhesive Technologies
Jeffrey Michael Karp 1
1Brigham and Women's Hospital/Harvard Medical School Cambridge United StatesShow Abstract
Unfortunately there has not been significant innovation to provide effective technologies for sealing tissues, especially internal tissues that are wet. While sutures and staples have been used for decades, they exhibit considerable limitations (they can induce significant tissue damage and their placement is often slow and challenging during minimally invasive procedures). This talk will explore multiple Next-Gen bio-inspired adhesive technologies to address several unmet medical needs. Through inspiration from the gecko, nano- and micro-topography has been harnessed to achieve biocompatible and controlled tissue adhesion. Towards development of a tissue adhesive glue to seal septal defects in children, we have harnessed inspiration from insect footpads, slugs and sandcastle worm viscous secretions that can create stable underwater adhesive bonds. We have been inspired by porcupine quills to develop mechanically interlocking adhesives that could replace staples with strong adhesion yet less tissue damage. We have also been inspired by the spiny-headed worm to develop an array of microneedles with biomimetic swellable tips that can achieve strong bonds to tissue through a mechanically interlocking mechanism and can be used to securely affix skin grafts to prevent seromas. This design provides universal soft tissue adhesion with minimal damage and reduced risk of infection compared to staples, and provides opportunity for delivery of bioactive therapeutics.
3:00 AM - *KK2.02
Cryo-Nanoprocessing of Bioresorbable Composites for Bone Tissue Engineering
SweeHin Teoh 1
1Nanyang Technological University Nanyang SingaporeShow Abstract
Bone is a composite structure that is made up of collagen (organic), calcium phosphates and trace elements (inorganic). Together with strict mechanical requirements, many scaffold-based tissue engineering strategies are now focused on developing composite systems that provide sufficient mechanical stability and biological stimulation to dictate osteogenic events in vivo. On this note, processing techniques for bioresorbable composites for bone tissue engineering have traditionally involved solvents and heat, which may not be ideal due to toxicity and material degradation, respectively. As such, alternative processing methods are actively pursued and evaluated. To this end, cryo-nanoprocessing has recently surfaced as a potential alternative processing technique for composites. By subjecting composite materials to temperatures sufficiently below glass transition, effective attrition into fine powders (sub-micron and nano) may be achieved. In addition, this attrition process facilitates distribution of the filler, resulting in a homogeneous composite powder. This homogeneous distribution may be retained in further post-processing techniques. A mechanistic explanation for this phenomenon may be that of solid-state diffusion, where effective interfacial interactions are created, resulting in the retention of their homogeneous distributions. The implications of this phenomenon such as mechanical and biological responses are important for bone tissue engineering strategies centered on scaffold technology.
3:30 AM - KK2.03
Nitinol-Based Nanotubular Coatings for the Modulation of Human Vascular Cell Function
Phin Peng Lee 2 1 Alec Cerchiari 1 2 Tejal A Desai 2 1
1UCB UCSF San Francisco United States2University of California, San Francisco San Francisco United StatesShow Abstract
We present the synthesis of an upright nanotubular coating with discrete, exposed nanotubes on top of superelastic Nitinol via anodization and the characterization of the surface elemental composition and nickel release rates of these substrates. We demonstrate, for the first time, that this coating could improve re-endothelialization by increasing the cell spreading and migration of primary human aortic endothelial cells on Nitinol. The coating also has the potential for reducing neointimal hyperplasia by decreasing the proliferation and expression of collagen I and MMP-2 in primary human aortic smooth muscle cells (HASMC). Furthermore, we did not observe the nanotubular surface to induce inflammation through ICAM-1 expression in HASMC as compared to the flat control. This coating could be used to improve Nitinol stents by reducing restenosis rates and, given the extensive use of Nitinol in other implantable devices, act as a generalized coating strategy for other medical devices.
4:15 AM - *KK2.04
Electrospun Nanofibers for Translational Applications
Younan Xia 1
1Georgia Institute of Technology Atlanta United StatesShow Abstract
Electrospun nanofibers can be routinely prepared from a wide variety of polymers, incuding those biocompatible and biodegradable polymers that have been approved by the FDA for clinical applications. Owning to their small diameters, controllable structures and alignments, electrospun nanofibers represent an ideal class of substitutes for the extracellular matrix and have found widespread use in regenerative medicine. In this talk, I will focus on their use in the repair or regeneration of tendons, tendon-to-bone insertions, dural tissues, and peripheral nerves. I will specifically discuss the issues that still need to be addressed before this class of nanomaterials can eventually find use in clinical applications.
4:45 AM - KK2.05
Versatile Click Alginate Hydrogels
Rajiv Desai 1 2 Sandeep T. Koshy 1 2 3 David J. Mooney 1 2 Neel S. Joshi 1 2
1Harvard University Cambridge United States2Harvard University Boston United States3Harvard-MIT Division of Health Sciences and Technology Cambridge United StatesShow Abstract
Alginate hydrogels are well-characterized, biologically inert materials that are attractive for many biomedical applications including the encapsulation of small molecules, proteins, and cells. Unfortunately, canonical covalent alginate hydrogels are limited in their usage for cell or protein encapsulation and subsequent delivery, due to crosslinking and functionalization strategies that can be biologically harmful. In this work we modified alginate polymers with tetrazine or norbornene groups and subsequently formed click alginate hydrogels with a wide range of mechanical properties. The highly specific, bio-orthogonal, and rapid click reaction is irreversible and allows for easy incorporation of cells with high postencapsulation viability. Additionally, preformed click alginate hydrogels can be easily modified with cell adhesive ligands to create a cytocompatible substrate with tunable ligand densities for 2D cell culture. Furthermore, click alginate hydrogels are minimally inflammatory, maintain structural integrity over several months, and reject cell infiltration when injected subcutaneously in mice. Click alginate hydrogels combine the numerous benefits of alginate hydrogels with powerful bio-orthogonal click chemistry to create new materials for a variety of tissue engineering applications.
Funding Sources: Wyss Institute, NIH 2 R01 HL069957, ARO W911NF-13-1-0242
5:00 AM - KK2.06
Anomalous Diffusion of Nanoparticles in Spheroid Three-Dimensional Tissue Model
Yichun Wang 1
1University of Michigan Ann Arbor United StatesShow Abstract
Understanding transport of nanoscale carriers within organs/tissues is crucial for their adequate use in biomedical imaging and drug delivery.Animal studies of such processes encounter formidable challenges associated with spatial resolution of in-vivo real-time imaging, inherent variability of animals, and high costs.Conversely, Petri dish cultures of immortalized cell are too simplistic to account for cellular sophistication of organs.Three-dimensional (3D) cell cultures approach the complexity of the actual organs and may enable high temporal and spatial resolution of permeation dynamics. Here, we investigated transport behavior of CNTs with and without targeting ligand -- transforming growth factor β1 (TGFβ1) -- in highly uniform spheroids of hepatocellular carcinoma (HCC) obtained in inverted colloidal crystals (ICC) scaffold that closely replicate human liver tumor tissue. Surprisingly, apparent diffusion rate of CNTs in tissue replica is higher than can be predicted theoretically and comparable with diffusion rates of similarly charged small molecule. Moreover, efficiency of CNT transport in tissues is enhanced after functionalization by TGFβ1 despite the considerable increase of volume and molecular mass. Such unexpected diffusion is attributed to surface diffusion of nanotubes on cellular membrane. These findings demonstrate that tissue spheroids allow for accurate optical tracking and theoretical description of CNT transport. They also indicate that nanotubes and similar nanostructures can serve as efficient drug delivery vehicles deep into the tissue that was often difficult to realize with other drug delivery carriers.
5:15 AM - KK2.07
Peripheral Nerve Regeneration by Synthetic Laminin and Glycosaminoglycans Mimetic Peptide Nanofibers
Mevhibe Gecer 1 Busra Mammadov 1 Melike Sever 1 Mustafa O. Guler 1 Ayse Tekinay 2
1UNAM, Bilkent University Ankara Turkey2Unam Ankara TurkeyShow Abstract
Peripheral nerve injury incidences are quite high mainly due to the unprotected nature of peripheral nervous system when compared to the central nervous system. Although regenerative capacity of peripheral nerves is much higher than those of the central nervous system, therapy is not much successful when the gap between proximal and distal stumps is long and because of poor functional outcomes. Autografts are applied in such cases although this method holds donor site morbidity (sense loss) and neuroma formation risks leading to intense pain experience by patients. Moreover unsatisfactory operational recovery after nerve injury is still a significant clinical challenge. Polymeric nerve guides are developed to be used in such cases to provide peripheral nerve regeneration by isolating nerve stumps in a hollow tube. However, although these nerve guides improve regeneration up to some level, clinical success ratio is not much high. There are some reports indicating patients undergone recession surgery due to intense pain at the injury site and neuroma formation and without regeneration after nerve guide transplantation. Considering the high cost of these nerve guides along with being not as effective as desired, it is clear that further modifications are required to improve their therapeutic effect. Commercially available polymeric nerve guides are inert materials not possessing any regenerative signal; they are just hollow tubes in which nerve stumps are being isolated from the surrounding tissue. In this study, it is aimed to improve effectiveness of nerve guides used in peripheral nerve injury therapy. For this purpose, peptide nanofiber gels are made up of neuroinductive peptide molecules one of which is derived from laminin (IKVAV-PA) while the other mimics growth factor binding glycosaminoglycans (GAG-PA). It is recently published that neural cells cultured on PA gels formed from these two molecules can extend quite long neurites. Nerve guides were filled with various form of peptide gels or sucrose that were applied to 10 mm gaps formed by full transaction in rat sciatic nerve. Functional recovery of sciatic nerve in nerve guide/PA and autograft was remarkably faster in comparison of other groups. Behavioral tests, electrophysiological measurements and histological analysis revealed that these laminin and glycosaminoglycan mimetic peptide nanofibers induce neuritogenesis with a great efficacy compared with autologous nerve grafts. Our results demonstrated that these two bioactive peptide amphiphile are fundumental for the optimal regeneration in peripheral nerve injuries and also they will be investigated further for their clinical potentials.
5:30 AM - KK2.08
Bone Engineering via Provisional Cartilage Matrix: Can Bone-Like Nano Hydroxyapatite Play a Role?
Melika Sarem 1 2 3 Andrea Barbero 4 Pavel Salavei 3 Ivan Martin 4 V.Prasad Shastri 1 2 3
1Institute for Macromolecular Chemistry, University of Freiburg Freiburg Germany2Helmholtz Virtual Institute on Multifunctional Biomaterials for Medicine Freiburg Germany3Bioss, Centre for Biological Signalling Studies, University of Freiburg Freiburg Germany4Department of Biomedicine, University Hospital Basel Basel SwitzerlandShow Abstract
Despite the tremendous progress in the past decade in translating nanomaterial&’s into cell contacting applications, there is still a huge knowledge gap to be filled with regards to deciphering the interplay between soluble signals and nanoscale physical cues present in the cellular microenvironment, on cell fate decisions. De novo bone regeneration is still challenging due to the complexity of the bone environment. Bone formation during development from cartilaginous tissue via endochondral ossification (EO) provides a unique framework to implement a developmental engineering paradigm in restoration of large volumes of bone. Recently, human nasal septum chondrocytes (hNCs) have been successfully used in nasal and knee cartilage reconstruction. However, the main challenge in using chondrocytes (Ch) in bone repair lies in invoking hypertrophy in phenotypically stable Ch.
We hypothesized that the stiff mineral phase juxtaposed with the softer collagen matrix, provides a biophysically heterogeneous environment that could play an important role in driving hypertrophy in Ch. With the aim of unraveling the role of physical cues offered by the bone environment, we have developed a platform that brings together concepts of self-assembly with control over protein conformation, to realize bone-like biomimetic hydroxyapatite (BBHAp) on synthetic substrates. Using polyelectrolyte-mediated induction of conformation changes in proteins, a nanoscale coating of biomineralization promoting proteins derived from avian and mammalian sources was realized on a fibrous polymer mesh, which when exposed to source of calcium and phosphate yielded BBHAp. That BBHAp coating in addition to being uniform, had similarity to the HAp phase found in bone in both the orientation of crystals along c-axis (002) and crystal dimensions (average BBHAp dimensions L: 58 nm; W: 30 nm; T: 3.90 nm versus HAp phase in bone: L: ~50 nm; W: 25 nm; T: 4 nm).
The ability of the BBHAp environment to upregulate EO related events was studied using hNCs and human articular chondrocytes (hACs)in vitroand in vivo in nude mouse model. In vitro, RT-PCR analysis of hNCs and hACs cultured on BBHAp revealed 12-fold down-regulation in COL II, and respectively 4- and 8-fold up-regulation in COL X in comparison to non-HAp controls. This drastic change in COL X/Col II ratio is one of the hallmarks for initiation of hypertrophy in Ch, which was also confirmed by immunohistochemistry (IHC). On going in vivo studies indicate the onset of EO-like events within the provisional cartilaginous matrix in an ectopic environment, including the ingress of blood vessels into the remodelling matrix, which was confirmed by the presence of CD31+ cells and concomitant positive staining for bone sialoprotein. These finings in sum implicate an important role for BBHAp in triggering EO related cellular events.
KK3: Poster Session I
Tuesday PM, April 07, 2015
Marriott Marquis, Yerba Buena Level, Salon 7/8/9
9:00 AM - KK3.01
Highly Moldable 3D Electrospun Scaffolds for Cell and Gene Delivery with Enhanced Mechanical Properties
Slgirim Lee 1 Sung Hwan Cho 2 Wuyong Choi 1 Unyong Jeong 2 Jae-Hyung Jang 1
1Yonsei University Seoul Korea (the Republic of)2Yonsei University Seoul Korea (the Republic of)Show Abstract
In the current presentation, development of three-dimensional and macroporous electrospun biodegradable scaffolds with the moldability and increased mechanical properties is discussed. The presented electrospun scaffolds were developed to overcome the limitations of the conventional electrospinning technique including i) restricted shapes (two-dimensional and sheet-like shapes) and ii) limited cellular infiltration into the scaffolds. In our previous study, by the core/sheath electrospinning technique with the polystyrene (PS) and the poly (ε-caprolactone) (PCL) followed by selective leaching procedure, three-dimensional, macroporous and highly moldable nanofibrous structure was fabricated. The scaffolds could be shaped into a variety of shapes with various shaping methods including molding, rolling and manual shaping without compromise of nanofibrous structures on the outer surface and inside of the scaffolds. The scaffolds, however, have relatively low mechanical strength which possibly limits the applications. To resolve the issue, various additives were added to PCL. With additives such as carbon nanotubes (CNTs) and polydopamines, the ultimate tensile strength and Young&’s modulus of the scaffolds were increased to 2- or 3-fold without compromising their flexibility, moldability and macroporosity. In in vitro characterization with the cells, the cells in the scaffolds showed the homogeneous cellular infiltration into the scaffolds and increased cell viability. Furthermore, the characterization with the viral vectors adsorbed on the surface of the scaffolds showed significantly enhanced the intensity of transgene expression. In vivo experiments showed that cells from host tissues infiltrated into the moldable scaffolds, whilst the cells were located only on the surface of the conventional electrospun scaffolds. In conclusion, nanofibrous and biodegradable scaffolds with macroporosity with high moldability were fabricated and with the inorganic and organic additives, the mechanical strength could be significantly enhanced. The presented system is a facile and versatile technology which has substantial potential to contribute to tissue engineering and regenerative medicine.
9:00 AM - KK3.02
Osteoblastic Cell Response on Calcium Modified Alumina Prepared by Hydrothermal Treatment with Calcium Chloride
Akari Takeuchi 1 Abdullah Al-Mahmood Tarique 2 Kanji Tsuru 2 Kunio Ishikawa 2
1Shinshu University Matsumoto Japan2Kyushu University Fukuoka JapanShow Abstract
Alumina ceramics is one of the most important materials for bone repairing. Although alumina has been used as prostheses for many years due to its chemical stability, mechanical strength and fracture toughness, it has no direct bonding ability to bone. To obtain long-term fixation of the materials to bone, osteoconductive materials have been paid much attention because it can bond to bone through a bone-like apatite layer. Thus, surface modification technique to change alumina from bioinert to octeoconductive has been awaited for a long time. We previously reported that calcium-modified alumina prepared by hydrothermal treatment with CaCl2 aqueous solution formed bone-like apatite on its surface after soaking in a simulate body fluid. This result suggested that the calcium-modified alumina might be an osteoconductive material. In this study, we performed in vitro biological analyses using rat bone marrow stroma cells to evaluate the osteoconductivity of the calcium-modified alumina.
Commercial alumina disks were immersed in distilled water or 50 mmol/L of CaCl2 (aq) in Teflon vessel with stainless steel jacket. Then, it was kept 125°C for 7 days for hydrothermal treatment. After rinsed with distilled water, one of the disks was analyzed by X-ray photoelectron spectroscopy (XPS) to confirm the Ca binding onto the surface. Bone marrow stroma cells harvested from rat tibias were cultured in various periods on the surface of the sterilized alumina disks, and then analyzed in terms of initial cell attachment, proliferation, and bone-like nodule formation. The alumina disks without hydrothermal treatment were denoted as NT, and the alumina disks after hydrothermal treatment in distilled water or 50 mmol/L of CaCl2 (aq) were denoted as HT0 or HT50.
From the XPS results, Ca binding was detected only on the surface of HT50 whereas no Ca binding was detected on the other alumina disks. The initial cell attachment rate after 7 hours of culture on the surface of hydrothermally treated alumina disks was the same as sintered hydroxyapatite. The cell number on the surface of Ca0 and Ca50 was significantly higher than that on NT. In the scanning electron microscopic observation after 24 hours, well spreading cells on the surface of Ca50 whereas cells were spherical shape on the surface of NT. Cells on all alumina disks proliferated greatly over 7 days of culture. After 3 days, the cell number on HT50 was significantly higher than other samples. Bone-like nodule formation was observed only on HT50 surface after 9 days culture. In contrast, there were no differences regardless of the type of the disks after 15 days of culture. These results showed that biological properties of alumina to bone marrow stroma cells could be significantly improved by the surface modification with Ca. Therefore, it is concluded that calcium-modified alumina prepared by the hydrothermal treatment with calcium chloride solution has high possibility to be osteoconductive material.
9:00 AM - KK3.03
Translational Medicine Study on Peritendinous Anti-Adhesion Barriers Made of Nanofibrous Membranes
Jyh-Ping Chen 1 Shih-Hsien Chen 1 Chih-Hao Chen 2 1
1Chang Gung University Taoyuan Taiwan2Chang Gung Memorial Hospital Taoyuan TaiwanShow Abstract
To improve current problems facing anti-adhesion products, we propose to use elctrospinning to fabricate porous nanofibrous membranes as anti-adhesion barriers. Nanofibrous membranes have microporous pores to allow diffusion of nutrients, while block penetration of cells at surgical sites to prevent adhesion formation. The membranes also have good mechanical properties and flexibility for application at surgical sites by surgeons after surgery. Anti-adhesive molecules, platelet-rich plasma (PRP) or silver nanoparticles could be incorporated into nanofibers to develop a multi-functional anti-bacterial membrane to prevent peritendinous adhesion and to promote tendon healing. We propose to combine FDA-approved biomaterials with anti-adhesion properties, polycaprolactone (PCL), chitosan (CS), polyethylene glycol (PEG), hyaluronic acid (HA), silver nanoparticles (Ag) and platelet-rich-plasma (PRP), to produce anti-adhesion barrier nanofibrous membranes for translational medicine study. The electrospun membranes contain single, mixed polymers or core-shell nanofibers. Five kinds of membranes be studied including (1) CS-grafted PCL nanofibrous membranes, (2) HA-grafted PCL nanofibrous membranes, (3) PCL+PEG blended nanofibrous membranes, (4) HA/PCL+Ag core-shell nanofibrous membranes, and (5) HA+PRP/PCL core-shell nanofibrous membranes. Grafted CS or HA on nanofibers can effectively reduce cell adhesion, thereby reducing adhesion formation. Blended PEG and PCL nanofibrous membrane can increase the efficiency of penetration of nutrients, and it is also can reduce the formation of adhesions. The HA releasing core-shell nanofibrous membranes contain a HA core region and a PCL shell layer for extended release of HA to reduce peritendinous adhesions. With the release of growth factors from incorporated PRP to promote tendon healing and the anti-bacterial activity from incorporated silver nanoparticles, multi-functional anti-adhesion barrier membranes could be obtained. Silver nanoparticles were added to the outer PCL shell layer during electrospinning to endow the nanofibrous membrane with anti-bacterial properties. The long-term growth factors releasing core-shell nanofibrous membranes contain a PRP/HA core region and a PCL shell layer for extended release of growth factors in PRP to promote long-term tendon healing. We characterize the physico-chemical properties of the fabricated membranes, and carry out in vitro and in vivo tests of the safety and efficacy of those membranes.
9:00 AM - KK3.04
Hypothesis: Bones Toughness Arises from the Suppression of Elastic Waves
Benjamin Rhys Davies 1 Hala Zreiqat 1 Andrew Minett 2
1University of Sydney Sydney Australia2Univ of Sydney Sydney AustraliaShow Abstract
Bone and other natural material exhibit a combination of strength and toughness that far exceeds that of synthetic structural materials. Bone&’s toughness is a result of numerous extrinsic and intrinsic toughening mechanisms that operates synergistically at multiple length scales to produce a tough material. At the system level however no explanation or organizational principle exists to explain how so many individual toughening mechanisms can work together. In this paper we utilize the concept of phonon localization to explain, at the system level, the role of hierarchy, material heterogeneity, and the nanoscale dimensions of biological materials in producing tough composites. We show that phonon localization and attenuation, using a simple energy balance, dynamically arrests crack growth, prevents the co-operative growth of cracks, and allow for multiple toughening mechanism to work simultaneously in heterogeneous materials. In turn the heterogeneous, hierarchal and multi-scale structure of bone (which is generic to biological materials such as bone and nacre) can be rationalized because of the unique ability of such a structure to localize phonons of all wavelengths.
9:00 AM - KK3.05
Nanofibrillar Scaffolds in Translational Medicine
Virginia M. Ayres 1 Volkan Mudat Tiryaki 1 David I Shreiber 2 Ijaz Ahmed 2
1Michigan State University East Lansing United States2Rutgers, The State University of New Jersey Piscataway United StatesShow Abstract
There is immense potential for biomaterials in translational medicine, when quantitative measures of biomaterial properties coupled with quantitative measures of biological responses lead to predictive, mechanistic models. We will present our recent studies [1,2] of an implantable electrospun polyamide nanofibrillar scaffold that has demonstrated promising wound healing properties for the central nervous system including in vivo mitigation of astrocytic glial scarring. The biomaterial properties of the nanofibrillar scaffolds and three controls were characterized by novel AFM and contact angle measurements. When four properties: nanoscopic elasticity, work of adhesion and surface roughness, and surface polarity were compared statistically, it became clear that the four environments presented significantly different sets of properties to astrocytes (ANOVA followed by post hoc comparisons with Tukey&’s test, P<0.05). Properties of the nanofibrillar scaffolds uniquely regulated multiple astrocyte features that were consistent with reduced reactivity. Reactive dBcAMP-treated and untreated astrocytes cultured on nanofiber scaffolds demonstrated minimal morphologic features changes, including process length, cell spreading and cell shape index. The relative levels of RhoGTPase expressions did not change between treated and untreated astrocytes cultured on nanofibrillar scaffolds, unlike all other cultures surfaces. Pairwise comparisons of untreated and treated astrocytes on the nanofibrillar scaffolds also indicated a statistically significant decrease in expression of inhibitory proteoglycans (CSPGs) with no accompanying decrease in cell density. A separate study demonstrated FGF-2 growth factor up-regulation by astrocytes on the nanofibrillar scaffolds with increased neurite outgrowth by co-cultured neurons. In sharp contrast, the responses to the properties of one control exhibited features reminiscent of glial scar formation. Confocal, super-resolution and AFM imaging revealed details of chain-like clustering with interwoven processes.
The emerging disciplinary field of translational medicine has identified “prevention of basic research findings being tested in a clinical setting” as a major roadblock. Often the block is at the first translation, when the correspondence between in vitro laboratory findings and in vivo animal model results is weak. Recent work, including ours, indicates that biomaterial properties can be tailored to reproduce in vivo responses in the laboratory with far greater accuracy and further suggests that precise control holds potential for inducing preferential cell differentiation in both in vitro and in vivo environments.
 Tiryaki, VM, Ayres, VM, Ahmed, I, Shreiber, DI. Nanomedicine, Epub ahead of print 2014 (DOI: 10.2217/NNM.14.33)
 Tiryaki, VM, Ayres, VM, Khan, AA, Ahmed, I, Shreiber, DI, Meiners, S. Int. J. Nanomedicine. 2012 7:3891-3905 (DOI: 10.2147/IJN.S32681)
9:00 AM - KK3.06
Towards a Scalable Biomimetic Antibacterial Coating
Mary Nora Dickson 1 Elena Liang 1 Patrick Lo 1 Chuandi Zhang 1 Albert F Yee 1
1University of California, Irvine Irvine United StatesShow Abstract
It has been found that the nanopillars on cicada wings are inherently antibacterial, irrespective of surface chemistry (Ivanova et al., Small, 2012). Thus, fabrication of devices presenting such nanostructures would obviate the requirement for any special surface chemical modification. Nano- and microstructured antibacterial surfaces have been previously proposed, including the Sharklet microstructured film (Chung et al., 2007), black silicon (Ivanova et al., 2013) and multi-scale wrinkled polymer films (Freschauf et al., 2012); none of these approaches can be used on ordinary polymer surfaces or easily scaled up. Thus, we applied industrial nanostructring techniques to generate biomimetic antibacterial nanostructures at the surfaces of the commonly used polymer poly(methylmethacrylate) (PMMA). Compared to flat films, replicated cicada wings 1) exhibited reduced surface adhesion of live E. coli determined by a standard fluorescence based viability assay, and 2) killed these bacteria, as evidenced by a decrease in colony forming units in suspension over time (up to 24 hours). To make the nanopatterning technique more industrially viable and generate a larger patterned area, we next employed nanoimprint lithography. We utilized several nano-lined and nano-holed molds (Lightsmyth) as well as a commercially available antireflective stamp (Holotools, Germany) with a nanopillared pattern very similar to that of the cicada&’s wing, to imprint large, flat, nanostructured polymer thin films. We determined that, compared to flat films, the pillars generated by the antireflective stamp were more antibacterial, as assessed by live/dead staining. Recent efforts have focused on using polymer coatings with self-assembled nanostructures to decease the feature size of the nanopatterns, and to allow for the application of nanopillars to curved surfaces. Our surfaces could be used for a wide variety of environmental and medical applications, including surgical trays / instruments and door handles (which function in air), and for implantable medical devices or catheter tubes (which function in aqueous environments).
9:00 AM - KK3.07
Mesoporous Silica Nanoparticles to Study Molecular Mechanism of HIV Neurotoxicity
Francesca Taraballi 1 Valeria Avdoshina 2 Italo Mocchetti 2 Ennio Tasciotti 1
1Houston Methodist Research Institute Houston United States2Georgetown University Medical CenterNeuroscience Washington United StatesShow Abstract
Nanocarriers have been studied to prolong the circulation time, bioavailability or stability of a certain biomolecules and to direct its delivery to a specific tissues or cells. In the whole scenario, mesoporous silica nanoparticles (MSNs) have several advantages; they are a very tunable platform for size and shape. Due to the flexibility of this platform and the vast possibilities for its further functionalizations MSNs represent a suitable targeted delivery system. Further, surface tailoring allows circumventing unwanted biological interactions, facilitating bioavailability and cellular uptake. In the present study we exploit this feature in order to study a molecular mechanism that leads to Human immunodeficiency virus-1 (HIV) neurotoxicity. HIV promotes neuronal injury. This process contributes to HIV-associated neurocognitive disorders, affecting more than 50% of HIV-infected individuals, even in the presence of combined antiretroviral therapy. However, the mechanisms of HIV-associated neurotoxicity remain unclear, thus precluding effective treatment of the neurological complications. Here, we report that the HIV envelop protein gp120 binds with high affinity to neuronal specific b-tubulin isotype III (TUBB3), by a twenty amino acid domain of the α-helix region that we synthetized (Helix-A), but not to tubulin isoforms expressed by other cell types. Using in vitro studies we have found that gp120 binds to recombinant TUBB3, tubulin dimers, and assembled MTs. The aim of the present study was to evaluate the neuronal protective effect of Helix-A from gp120 toxicity. Unfortunately, Helix-A peptide did not penetrate inside cells cytoplasm. Thus, we bound Helix-A to MSNs (Helix-A nano) to allow for its penetration into primary rat cortical neurons. We characterized the Helix-A nano delivery system for its stability and biological activity. Helix-A nano prevented gp120-mediated decrease in mitochondrial function (MTT) as well as alteration in mitochondrial morphology. In addition, Helix-A nano prevented gp120-mediated neuronal loss (Hoechst/PI). Since Helix-A did not bind to gp120 the proposed mechanism of neuroprotection of Helix-A involves its competitive binding to TUBB3 and MTs. Thus, neuroprotection conferred by Helix-A strongly suggests that the direct interaction of gp120 with tubulin is one of the main central mechanisms by which gp120 promotes axonal degeneration. Hence, our study identifies a novel mechanism of HIV neurotoxicity and provides a potential novel therapeutic approach to reduce HIV-mediated neurodegeneration.
9:00 AM - KK3.08
Cyclodextrin Strengthened Polyvinylpyrrolidone (PVP) for Transdermal Drug Delivery
Wei Chen 1
1City University of Hong Kong Hong Kong Hong KongShow Abstract
The fast dissolving property of PVP makes it very suitable for transdermal delivery. But high water absorption and low mechanical strength limits its microneedles to penetrate into skin. In this work, we increase the mechanical strength of PVP by simply compositing cyclodextrin (CD) derivatives. PVP/CD film and microneedles are prepared by mixing and casting composite solution to PDMS mould, respectively. The water absorption, mechanical and thermal properties are tested by saturated salt method, microindentation and DSC, respectively. Comparing to pure PVP, The water absorption of PVP was reduced by 36%-40% at different RHs as the PVP/CD inclusion complexes formed, even after 10 or 20 days. The Young&’s modulus and hardness of the PVP/CD could be greatly improved, especially for low molecular weight PVP. Besides these, the thermal properties, the glass transition temperature (Tg) of the composite increased by up to 39 0C. With these improved properties, the composite microneedles can easily penetrate into pig skin with fewer cracks than pure PVP microneedles. This kind of microneedles is more suitable in transdermal delivery than pure PVP.
9:00 AM - KK3.09
Silk Fibroin/Hydroxyapatite Composite Scaffolds for Bone Tissue Engineering
Min Hee Kim 1 Dong Su Im 1 Won Ho Park 1
1Chungnam National University Daejeon Korea (the Republic of)Show Abstract
Regenerated Bombyx mori silk fibroin (SF) has excellent biological and mechanical properties, including biocompatibility, programmable biodegradability, and remarkable strength and toughness. Diverse and adaptable properties of Sf are possible by varying the structural form using different processing conditions. One of the important physical forms for biomaterials is the formation of hydrogels, which has been extensively studied for a variety of polymers. The sol-gel transition depended on the concentration of the protein, temperature, and pH. In the SF hydrogel, random coil to β-sheet(physical cross-linking) structural transitions were noted during the process of hydrogelation. Due to the β-sheet formation, SF exhibits relatively slow degradation in vitro and in vivo, compared to collagen and many other biopolymers.
Tissue engineering has potential to address this need through the combination of biomaterials, growth factors, and cells. Highly porous scaffolds are generally used as the substrate for anchorage dependent cells and to facilitate nutrient and metabolite distribution to guide cell growth leading to new bone tissue formation. For bone tissue engineering, biodegradable synthetic polymers such as poly(glycolic acid) (PGA), poly(lactic acid) (PLA), and copolymers of poly(DL-lactic-glycolic acid) (PLGA), and biodegradable naturally derived polymers such collagen and fibrin.
Hydroxyapatite (HAP) has been investigated for bone replacement since this material mimics natural bone mineral features. HAP has been studied extensively in cell culture and possesses osteoconductivity.
In this study, the synthesis and characterization of bone-like mineral HAP into highly porous biodegradable silk fibroin scaffold with via chemical cross-linking reaction of SF by gamma-ray (γ-ray) were investigated. These 3-D SF scaffolds had different secondary structures, elasticity and nanostructure compared with β-sheet induced hydrogels.The effect of degradation rate, elasticity and mineralization on osteogenic responses of osteoblast was assessed with respect to bone tissue engineering
9:00 AM - KK3.10
ATP-Triggered Anticancer Drug Delivery
Ran Mo 1 2 Tianyue Jiang 1 2 Wujin Sun 1 2 Zhen Gu 1 2
1University of North Carolina at Chapel Hill and North Carolina State University Raleigh United States2University of North Carolina at Chapel Hill Chapel Hill United StatesShow Abstract
Stimuli-responsive drug-delivery systems (DDSs) are playing an increasingly crucial role in a variety of applications for cancer treatment. We herein report an ATP-responsive nanogel consisting of three distinct functional components: an ATP-responsive DNA scaffold with doxorubicin (Dox), protamine and a hyaluronic acid (HA)-crosslinked shell. The DNA scaffold was composed of the ATP aptamer and its complementary single-stranded DNA (cDNA),which also provided faithful loading sites for Dox. When exposed to ATP, the dissociation of the DNA duplex resulting from the formation of the ATP/ATP aptamer complex caused the intercalated Dox to be released from the duplex. Cationic protamine was utilized to complex the DNA motif, which had enhanced cell penetration, endosomal escape and nuclear targeting effects. Anionic HA was coated on the complex core followed by UV irradiation-mediated photocrosslinking to form a protective crosslinked gel shell, and also serves as ligands to bind the CD44 receptors that are overexpressed on the cell surface of many cancers. This ATP-responsive nanocarrier showed an enhancement in the chemotherapeutic inhibition of tumor growth using xenograft breast tumor-bearing mice. In addition, a liposome-based co-delivery system, composed of a fusogenic liposome encapsulating the Dox-loaded ATP-responsive DNA scaffold and an ATP-loaded liposome, was further developed for ATP-mediated drug release triggered by liposomal fusion. Directly delivery of extrinsic liposomal ATP facilitated the Dox release from the fusogenic liposome in the acidic intracellular compartments by a pH-sensitive membrane fusion and therapeutic efficacy was enhanced both in vitro and in vivo.
9:00 AM - KK3.11
Actin-Polymerization Driven Phenomena: Experiments and Simulation
Qian Zhu 1
1The University of Hong Kong Hong Kong ChinaShow Abstract
Actin exists in almost all of the eukaryotic cells (except nematode sperm) and participates in many important cellular processes, including cell motility, morphogenesis and endocytosis. Specifically, the force generated by polymerization and depolymerization of actin filaments is commonly believed to play a key role in how these processes take place. Substantial progress has been achieved in the past decade in understanding how actin networks assemble themselves and evolve. In particular, essential proteins crucial for actin based motility have been identified allowing researchers to reconstruct actin-propelled motions of artificial cargos (microspheres or lipid vesicles coated with proper proteins) in vitro. Based on this approach, fundamental issues like the temporal and spatial distribution of polymerization-induced stress and the interplay between load surface geometry, as well as elasticity, and actin assembly can all be probed.
My present study is focusing on:
(1) Investigating the rotational movements of artificial cargos (microspheres) propelled by actin polymerization.
Many experiments have proved that Listeria monocytogenes will rotate along its long axis and form right-handed helical trajectories during actin-based motility. But whether artificial cargos propelled by actin polymerization have rotational movements has not been verified. To look into this subject, polystyrene beads bound with small fluorescent beads and coated by VCA were used to generate motility in assays reconstituted from purified proteins. The spatial positions of polystyrene beads and fluorescent beads were recorded by confocal microscope during the motion process. Whether rotational movements exist can then be confirmed by tracking their relative positions.
(2) Modeling actin polymerization induced shape changes of artificial lipid vesicles based on energetic considerations.
Lipid vesicles have been applied to investigate actin-propelled motions in vitro. It&’s been discovered that lipid vesicles can induce the structural deformation of actin filaments from branched networks to bundles. Bundled actin filaments exert forces on the surfaces of vesicles which will then generate filopodium-like protrusions. Built on the experimental results and the principle of minimum energy, we come up with a model attempting to explain the shape changes of artificial lipid vesicles induced by actin polymerization.
Experimental findings, theoretical frameworks and some preliminary results will be presented.
KK1: Nanomaterials for Biomacromolecule Delivery
Tuesday AM, April 07, 2015
Park Central Hotel, 2nd Floor, Metropolitan I
9:30 AM - KK1.01
Development of New mRNA Delivery Materials
Yizhou Dong 1
1The Ohio State University Columbus United StatesShow Abstract
Biomaterials and biotechnology have made significant advances in mRNA therapeutics for a variety of biomedical applications in the past decade. One major challenge to the broad application of nucleic acid therapeutics in humans is their safe, selective and efficient delivery to target tissues. We will describe the development of new nanomaterials for delivery of messenger RNA for transient protein expression. These results offer promise towards the future treatment of genetic disorders that require complex gene regulation.
9:45 AM - KK1.02
Cationic Block Copolymers for the Co-Delivery of siRNA and Chemotherapeutics
David Spencer 1 Bryan Luu 1 Nicholas A. Peppas 1
1The University of Texas at Austin Austin United StatesShow Abstract
Cationic polymers capable of encapsulating chemotherapeutic drugs and electrostatically binding small interfering RNA (siRNA) are a promising platform for the treatment of drug resistant cancer. Specifically, pH responsive cationic polymers that are in a collapsed state in the blood and swollen at the slightly acidic pH of the endosomes allow for enhanced therapeutic effect.
In this work, block copolymers of dimethylamino(ethyl methacrylate), diethylamino(ethyl methacrylate), or diisopropylamino(ethyl methacrylate) and poly(ethylene) glycol were synthesized by activators regenerated by electron transfer atom transfer radical polymerization (ARGET ATRP). The hydrophobic pH responsive monomer as well as the ratio of hydrophobic to hydrophilic block lengths were varied to modulate pH responsive behavior. NMR was used to verify incorporation of monomers and critical micelle concentration was determined by pyrene fluorescence. Particles were typically 70-100nm at pH 7 as determined by dynamic light scattering and carried a positive zeta potential. The ratio of amine containing methacrylate monomers to poly(ethylene) glycol allowed for the tunable modulation of the pKa 5.5 to 7.0. Cytotoxicity of the cationic polymers was assessed in human adenocarcinoma cells, and nanoparticle uptake in murine macrophages was observed using confocal microscopy. Chemotherapeutic delivery potential was studied by imbibition of doxorubicin at low pH followed by release in vitro at pH 7.4 for 2 hours and then pH 5.2 for 6 hours.
Block copolymers synthesized by ARGET ATRP demonstrated tunable pH responsive properties appropriate for co-delivery via endosomes. These block copolymer were cyto-compatible with model cells lines and demonstrated the ability to effectively load and release a model chemotherapeutic.
This material is based upon work supported by the National Science Foundation Graduate Research Fellowship Program (DGE-1110007).
10:00 AM - *KK1.03
Nucleic Acid Delivery Systems for RNA Therapy and Gene Editing
Daniel G. Anderson 1
1Massachusetts Institute of Technology Cambridge United StatesShow Abstract
High throughput, combinatorial approaches have revolutionized small molecule drug discovery. Here we describe our work on high throughput methods for developing and characterizing RNA delivery and gene editing systems. Libraries of degradable polymers and lipid-like materials have been synthesized, formulated and screened for their ability to delivery RNA, both in vitro and in vivo. A number of delivery formulations have been developed with in vivo efficacy, and show potential therapeutic application for the treatment of genetic disease, viral infection, and cancer.
10:30 AM - KK1.04
Non-Viral Delivery of microRNA from 3D Silk Fibroin Scaffolds to Promote Osteogenesis of Mesenchymal Stem Cells
Chuanxu Yang 1 Chi-Chih Chang 1 Jorgen Kjems 1
1Aarhus University Aarhus DenmarkShow Abstract
Stem cell-based tissue engineering holds great potential for the regeneration and/or replacement of damaged tissue due to their capability of differentiation to several lineages. Controlling Mesenchymal stem cells (MSCs) differentiation into osteogenic lineage provides an important strategy for the restoration the function of bones after osteoporosis, osteoarthritis or orthopedic surgeries. To control the osteogenic differentiation, microRNAs (miRNAs) are found to be highly potent tools due to the delicate control of gene expression through RNA interference (RNAi) mechanism. Several promising miRNAs have been identified as key regulators of multiple growth factors simultaneously. Despite their great potential, miRNAs are negatively charged macromolecules with limited stability, thus hampering the biological functions. Therefore, a key challenge for controlling MSCs differentiation by miRNAs is the development of an effective delivery system. In addition, an ideal platform for tissue regeneration usually involves a three-dimensional (3D) macroporous scaffold to provide mechanical support for cell seeding and organization.
To address the delivery issue, we synthesized a lipid-like-material, so-called “lipidoid” by conjugating alkyl-epoxides to spermine. Thereafter, the lipid nanoparticles (LNP) self-assembled together with cholesterol and DSPE-PEG by the ethanol-injection method. We evaluated the delivery efficiency by silencing the expression of GFP in GFP expressing MSCs and the demonstrated a significant knockdown effect, higher than that of Lipofectamine 2000, with no toxicity detected.
To generate a proper mechanical support for MSCs, a 3D porous scaffold was fabricated by lyophilization of a silk fibroin (SF) solution. SF is a naturally occurring protein with great biocompatibility, high strength and elasticity. Our water insoluble 3D SF scaffold with macroporous structure was obtained after crystallization by ethanol treatment. The functionalization of scaffold with siRNA/miRNA containing LNP was achieved by a simple soaking and freeze-drying process. By tuning the feed concentration of particles, various loading amounts could be achieved. 3D culture of MSCs in siGFP functionalized scaffold resulting in gene silencing and the efficacy is proportional to the loading amount.
Furthermore, miRNAs including anti-miR-138 and anti-miR-34c could also be loaded into the 3D scaffold. Under osteogenic culture, significantly enhanced alkaline phosphatase (APL) activity was observed in MSCs after 14 days and increased calcium content was achieved after 28 days, indicating the promoting of osteogenesis.
In summary, we have engineered a novel platform based on 3D silk fibroin scaffold functionalized with miRNAs loaded LNP to promote osteogenesis of MSCs. The facile fabrication and the versatility of our delivery platform to load different miRNA payloads, favor its broad regenerative applications and targeted personalized therapy.
10:45 AM - KK1.05
Calcium Phosphate Nanoparticles as Versatile Carriers of Drugs and Biomolecules: Gene Therapy, Immunology and Drug Delivery
Matthias Epple 1
1University of Duisburg-Essen Essen GermanyShow Abstract
Calcium phosphate nanoparticles consist of the same material as human hard tissue (bone and teeth) and therefore have a high biocompatibility. They can be loaded with all kinds of (bio-)molecules which are protected from enzymatic degradation below a nanometer-thin shell of silica. On their surface, targeting molecules like antibodies or peptides can be covalently attached. The incorporation of fluorescing molecules permits to track their pathway into cells, in the case of near-infrared dyes (NIR) also in-vivo after intraveneous injection.
They are entering a cell by endocytosis or macropinocytosis, as shown by fluorescence microscopy in the presence of endocytosis inhibitors. Inside the cell, they first occur inside an endosome that later turns into a lysosome where the low pH leads to the dissolution of calcium phosphate, an increase of osmotic pressure and a rapid endosomal escape to release the cargo into the cytoplasm.
Different applications are shown, including in-vivo results: The delivery of DNA leads to an efficient transfection to turn on the production of proteins. The delivery of siRNA leads to an efficient gene silencing. The delivery of toll-like receptor ligands together with an antigen stimulates dendritic cells and specifically turns on the immune system against viruses. This is demonstrated both for prophylactic and for therapeutic vaccination. In additional experiments, it is shown that all kinds of molecules, both synthetic as well as biological (like proteins) can be carried across the cell membrane with the help of calcium phosphate nanoparticles.
11:30 AM - *KK1.06
Self-Assembled Lipid-Polymer Hybrid Nanoparticles for siRNA Delivery
Jinjun Shi 1
1Brigham and Women's Hospital, Harvard Medical School Boston United StatesShow Abstract
RNA interference (RNAi), which can selectively knockdown target genes, has shown great potential in the treatment of various diseases including cancer. Thus far, numerous nanoparticle (NP) platforms, such as lipoplexes and polyplexes, have been developed to facilitate the safe and effective delivery of small interfering RNA (siRNA), which represents a major hurdle for the clinical applications of RNAi. Nevertheless, these NP systems lack the sustained siRNA release property, and thus can only induce transient gene silencing due to the short lifetime of siRNA. Therefore, it is expected that the development of controlled-release siRNA NPs will lead to sustained gene silencing and more effective cancer treatment.
Herein, we present a robust NP platform for safe and effective siRNA delivery in a sustained manner, which can be developed through the self-assembly of biodegradable and biocompatible polymers and lipids. The lipid-polymer hybrid NPs show excellent knockdown efficacy at low doses of siRNA. More importantly, these NPs can control the temporal release of siRNA, with the half-release time of ~ 9 days, for sustained silencing of target gene expression. For example, results demonstrate that the expression of PHB can be effectively inhibited for over two weeks after short-term transfection with the NPs. The NP-mediated sustained silencing of PHB in turn generates more effective tumor cell growth inhibition in vitro and in vivo than the lipofectamine-siRNA complexes. Furthermore, this NP platform has been applied to deliver siRNAs against drug resistance and chemotherapeutic drugs for synergistic cancer treatment. We expect that the lipid-polymer hybrid NP platform with the property of sustained siRNA release could be of interest in both fundamental biological studies and clinical applications.
12:00 PM - KK1.07
Biomimetic Vesicles for Glucose-Responsive Insulin Delivery
Wanyi Tai 1 2 Jin Di 1 2 Zhen Gu 1 2
1University of North Carolina at Chapel Hill and North Carolina State University Raleigh United States2University of North Carolina at Chapel Hill Chapel Hill United StatesShow Abstract
An artificial “closed-loop” system able to mimic pancreas activity and release insulin in response to glucose level changes has the potential to improve patient compliance and health. Herein, a new glucose-responsive formulation for self-regulated insulin delivery was constructed by packing insulin, glucose-specific enzymes into pH-sensitive polymersome-based nanovesicles assembled by a diblock copolymer. Glucose can passively transport across the bilayer membrane of the nanovesicle and be oxidized into gluconic acid by glucose oxidase, thereby causing a decrease in local pH. The acidic microenvironment causes the hydrolysis of the pH sensitive nanovesicle that in turn triggers the release of insulin in a glucose responsive fashion. In vitro studies validated that the release of insulin from nanovesicle was effectively correlated with the external glucose concentration. In vivo experiments, in which diabetic mice were subcutaneously administered with the nanovesicles, demonstrate that a single injection of the developed nanovesicle facilitated reduction of the blood glucose levels in for over one week.
12:15 PM -
12:30 PM - KK1.09
Bioorthogonal Hydrogel Chemistries for Enabling Therapeutic Protein Delivery Systems
Amin Famili 1 Jeffrey Aaronson 1 Karthikan Rajagopal 1
1Genentech South San Francisco United StatesShow Abstract
Bioorthogonal chemistries enable new avenues for materials design when encapsulation of therapeutic proteins without loss of function is required. Recently, inverse electron demand Diels-Alder (IEDDA) chemistries have garnered interest for hydrogel synthesis applications in which bioorthogonality is paramount. To date, a thorough evaluation of protein stability after exposure to IEDDA functionalities has been missing. In this work, we have characterized two therapeutic antibodies—a monoclonal antibody (mAb) and an antibody fragment (Fab)—by size exclusion chromatography (SEC), ion exchange chromatography (IEC), capillary electrophoresis (CE-SDS), antigen-binding surface plasmon resonance (SPR) and mass spectroscopy (MS) after exposure to several IEDDA functionalities. As an exemplary application, we produced hyaluronate hydrogels with in situ antibody encapsulation to demonstrate the applicability of this chemistry to the design of a protein delivery system. Tethering of the therapeutic antibody to the hydrogel matrix via a transient covalent or non-covalent linker enables the development of a system for the sustained delivery of proteins. This work sheds new light on the applicability of IEDDA chemistries to biomaterial design and synthesis and demonstrates their applicability in a translational protein delivery system.
12:45 PM - KK1.10
Disease-Directed Drug Delivery in Translational Medicine: A New Paradigm to Prevent Inflammatory Diseases
Praveen Kumar Vemula 1
1Institute for Stem Cell Biology and Regenerative Medicine (inStem) Bangalore IndiaShow Abstract
Inflammation or disease severity in inflammatory/autoimmune diseases is presented in highly fluctuated state. One such example is, immune response during post-transplantation of solids organs and vascularized composite allografts (VCA). Currently, systemic immunosuppression is used in vascularized composite allotransplantation (VCA). This treatment has considerable side effects and reduces the quality of life of VCA recipients. We have developed a self-assembled hydrogel in which immunosuppressive drug tacrolimus has been encapsulated. Hydrogel releases the drug in response to proteolytic enzymes that are overexpressed during inflammation (disease severity). This hydrogel showed robust hydrolytic stability to avoid burst release of drug, which is a hallmark of current delivery vehicles, and released the drug in response to the inflammation in a dose-dependent manner. We validated this hydrogel in limb transplantation model in rats. A one-time local injection of the tacrolimus-laden hydrogel significantly pro-longed graft survival in a Brown Norway-to-Lewis rat hindlimb transplantation model, leading to a median graft survival of >100 days compared to 33.5 days in tacrolimus only-treated recipients. Control groups with no treatment or hydrogel only showed a graft survival of 11 days. In conclusion, a single-dose local injection of an enzyme-responsive tacrolimus- hydrogel is capable of preventing VCA rejection for >100 days in a rat model and may offer a new approach for immunosuppression in VCA.
Zhen Gu, UNC at Chapel Hill | NC State
Samir Mitragotri, University of California, Santa Barbara
Chenjie Xu, Nanyang Technological University
Symposium Support Aldrich Materials Science
KK5: Smart Nanomaterials for Bioapplications
Wednesday PM, April 08, 2015
Park Central Hotel, 2nd Floor, Metropolitan I
2:30 AM - *KK5.01
Enhancing Nanoparticle Catalysis on Electrochemical Reduction of H2O2 for Sensitive Cancer Detection
Shouheng Sun 1
1Brown University Providence United StatesShow Abstract
Hydrogen peroxide (H2O2) is an important reactive oxygen species generated in cells via oxygen metabolism and is involved actively in cell signaling and cell growth. However its uncontrolled over-production can cause detrimental oxidation of biomolecules and lead to aging, cancer and other diseases. In this talk, I will present our new method of developing dumbbell (Au-Fe3O4 and PdPt-Fe3O4) and core/shell (Au/MnO) nanoparticles as sensitive probe for H2O2 detection. The dumbbell nanoaprticles were prepared by controlled nucleation and growth of Fe3O4 on the pre-synthesized noble nanoparticles, while the core/shell Au/MnO nanoparticles were made by controlled oxidation of AuMn alloy nanoparticles. Both dumbbell and core/shell nanoparticles are active for electrochemical reduction of H2O2 with the detection limit reaching as low as 5 nM. The highly sensitive electrochemical sensor has been used to monitor H2O2 concentration levels released from living cells, from which tumorigenic cells were detected to have higher level of H2O2 than the non-tumorigenic ones. The composite nanoparticle probe can be used for sensitive cancer detection and may also help to increase efficacy for cancer therapy.
3:00 AM - KK5.02
Self-Decomposable SiO2 Nanoparticles for Application in Sustained Drug Release
Quan Li 1
1The Chinese University of HongKong Hong Kong ChinaShow Abstract
One of the most important issues in nanoparticle-carrier drugs is the release of the drug molecules from the carrier. In particular, sustained release is a critical feature that is desired in modern drug design. It aims to prolong therapeutic effect and avoid systemic toxic effect caused by burst release. In the present work, we show that the sustained release can be achieved by employing a self-decomposable SiO2 nanoparticle system with specific loading schemes. We are able to manipulate the drug release profiles and achieved sustained drug release with long duration. In vivo results showed that under the same dose, the nanoparticle drug demonstrated prolonged and stable presence in plasma for 3 days, while the free drug itself was cleared out of the biological system in a couple of hours. In addition, the self-decomposable feature of the nanoparticles enables the carriers&’ easy renal clearance after the drug release, significantly reduced their undesired accumulation in livers and spleens. The present study shows a promising system with highly desired therapeutic features. It will have great impact on improving the management of chronic diseases which require long-term medication.
3:15 AM - KK5.03
Near-IR Responsive Drug Releasing System Based on Nano-Assembly of Gold Nanorods and Phase Change Material
Junseok Lee 1 Won Jong Kim 1
1POSTECH Pohang Korea (the Republic of)Show Abstract
Near-infrared (NIR)-responsive drug delivery system have been studied for a decade from inherent characteristics of NIR such as good biocompatibility or deep penetration depth against human tissue. Herein, we prepared a NIR-responsive drug delivery carrier by the facile fabrication methods including mixing and extraction procedure. Nano-assembly of mesoporous silica coated gold nanorods (AuNR@mSiO2) and a drug loaded phase change material (PCM) was fabricated and physicochemical properties were analyzed. The gold nanorods at the core of AuNR@mSiO2 showed outstanding NIR sensitivity and the mesoporous silica shell was utilized for reservoir of loaded drug and PCM. The PCM played a pivotal role for drug loading as both a thermosensitive gatekeeper and a medium for a hydrophobic anticancer drug. An enhanced cytotoxicity of this system was observed against various cell lines in comparison with that of free DOX. It can be explained as a result of enhanced intracellular drug releasing through the rapid melting process of PCM, which was easily monitored by cellular imaging. Since any complicated synthetic methods are required for fabrication and PCM is easily capable of medium for various hydrophobic drugs, this system has an enormous potential for broad application in the biomedical field as a delivery system for hydrophobic drugs with NIR-responsive behavior.
3:30 AM - KK5.04
Novel Light Triggered Therapeutics for Selective Cell Phenotypes
Samuel Martin Goodman 1 Colleen Courtney 1 Jessica McDaniel 1 Anushree Chatterjee 1 Prashant Nagpal 1
1University of Colorado Boulder United StatesShow Abstract
Current methodologies towards developing selective drugs and nanoparticle-based therapies rely on non-specific drugs or metal nanoparticle, and a selective drug-delivery system (colloids and vesicles coated with desired antibodies or proteins) to deliver therapeutics to desired cell sites. Typically these non-specific drugs, once spatially delivered, kill the targeted cells (e.g. tumor or pathogen) and even healthy cells in close proximity. Here we present a novel nanoparticle-based therapy where the nanoparticles selectively target specific cell phenotypes and biological processes, based on their biochemical potentials. Using a specific light induced targeting mechanism, these Light-Activated Reactive Species (LARS) have the capability to target and kill specific-pathogens upon illumination with visible or infrared light, without affecting the growth of other cells. Furthermore, the growth of selected cells and the targeted pathogens is unaffected in dark, and we can trigger these designed LARS species externally with light to demonstrate bactericidal properties. To demonstrate the efficacy of the specific design of these LARS species, we will also show both proliferation and cell death using two different LARS in the same targeted cell phenotypes. Using Escherichia coli cells and HEK293 mammalian cells as model systems, we will demonstrate selective LARS which either specifically kill or proliferate E. coli growth, while leaving HEK293 cells unaffected in individual cultures as well as a co-culture of E. coli and HEK293 cells. These unprecedented results demonstrate the ability to tune the biochemical potential using LARS nanoparticles, allow targeting selective cell phenotypes over a range of organisms and enable a new class of antimicrobials and targeted organism-specific therapeutics.
3:45 AM - KK5.05
The Effect of Polymer Structure on Drug-Polymer Interactions in Core/Shell Temperature Responsive Nanogels
Jonathan Thomas Peters 1 Isha Verma 2 Nicholas A. Peppas 1
1The University of Texas at Austin Austin United States2The University of Texas at Austin Austin United StatesShow Abstract
The issues encountered with current cancer therapies stem from a vast array of different diseases that fall under the heading of cancer. In order to address this issue in the past, treatments have been developed that leave much to be desired with respect to efficacy and deleterious side effects. This is especially apparent with the systemic delivery of chemotherapeutics. To this end, a wide range of nanocarriers have been developed over the years to take advantage of the enhanced permeability and retention effect that leads to the localization of large particulates in the cancerous tissue. One subset of nanocarriers that has been extensively studied is thermoresponsive hydrogels combined with an external stimuli responsive nanoparticle. The actuation of the temperature responsive hydrogel by the heating caused the external stimuli responsive nanoparticle leads to a release of a drug payload. In the past, these systems have been based around nanogels of N-isopropyl acrylamide and its copolymers. The issue that arises with these systems is that, due to the high surface area to volume ratio, and lack of consideration for drug polymer interactions, these systems result in inefficient loading and premature release of the drug payload.
To this end, a new core/shell system has been developed to enhance drug-polymer interactions while maintaining the responsive nature of N-alkyl substituted acrylamides. These systems utilize a core of temperature responsive N-isopropylmethacrylamide, to elicit a temperature response above 37°C, coated with a variety of drug compatible polymers. In order to identify the perameters that greatly improve drug-polymer interactions bulk hydrogels of the various polymer coatings are tested for partition coefficient and permeabilies of model chemotherapeutics. The results of these studies are then compared to the physical properties of the gels, including swelling ratio and relative hydrophobicity determined by pyrene fluorescence. The core shell systems are then synthesized and loading and release experiments are performed to determine the improvement of loading efficiency and the eventual release allowed by the different systems.
The bulk polymer films studied include tert-butyl methacrylate (TBMA), phenyl ehtyleneglycol ether acrylate (PEEA), and phenyl methacrylate (PMA). These systems are chosen due to hydrophobicity and structural properties that lend themselves to the interaction with model chemotherapeutics. The partition coefficients demonstrated that TBMA and PMA have a much higher drug affinity than PEEA, likely due to greater hydrophobicity. However, the overall release under gentle heating is inefficient, releasing a small percent of the loaded drug. This can be overcome via the extreme heating capable with the inclusion of magnetic nanoparticles, utilizing a burst-like release often attained by sharp temperature changes in N-alkyl substituted nanogels.
4:30 AM - KK5.06
Controlled Nanoparticle Release from Oscillating Magnetic Microbubbles
Chenjie Xu 1
1Nanyang Technological University Singapore SingaporeShow Abstract
Magnetic microbubbles, microbubbles coated with magnetic-nanoparticles, offer strong potential for targeted drug delivery due to their combined acoustic and magnetic properties. So far drug delivery based on magnetic microbubbles (MMBs) has been relying on their disruption/collapse upon applying strong ultrasonic forcing. During bubble collapse not only most of the carriers are released but also large shear stresses and microjets are generated. Yet the rapid collapse of bubbles may be harmful for the surrounding tissues and the synchronous burst release of drugs from many bubbles could exceed toxic limits. In summary, there is little control on the release of drugs using the bubble collapse.
In this talk, we will introduce on a new property of MMBs, in which MMBs continuously eject nanoparticles from the surface without collapsing under an acoustic filed. We drive the bubbles at their resonance frequency at a moderate amplitude inducing volume and surface modes. Particle release has been observed for both MMBs and functionalized MMBs (i.e. coated with a mixture of magnetic and drug containing particles) with medical relevant sized between 450 nm to 200 mu;m. We are able to predict the transport of the drug with a simple model force balance model. This technology posses interesting opportunities for targeted drug delivery through/into tissue barriers, which are hard to diffuse or penetrate without causing damage.
As a proof-of-concept, we deliver Doxorubicin-containing poly(lactic-co-glycolic acid) (PLGA) particles across the hydrogel barrier to target the cancer cells that are hard to reach for free Doxorubicin or Doxorubicin-PLGA particles.
4:45 AM - KK5.07
Targeted Delivery of Antibiotics Using Gold-Based Nanoparticle Platform for Antimicrobial Treatments
Jingyi Chen 1 Samir Jenkins 1 Daniel Meeker 2 Karen Beenken 2 Mark Smeltzer 2
1University of Arkansas Fayetteville United States2University of Arkansas for Medical Sciences Little Rock United StatesShow Abstract
Staphylococcus aureus (S. aureus) is a significant human pathogen responsible for a wide range of diseases including skin and wound infections, toxic shock syndrome, septic arthritis, endocarditis, and osteomyelitis. Increasingly, these infections are caused by bacterial strains resistant to standard antimicrobial treatments. We have demonstrated an effective alternative based on nanoparticle-mediated photothermal approach for elimination of S. aureus. In this work, we develop an integrated approach involving targeted delivery of antibiotics and photothermal effect to treat S. aureus. A smart polymer is developed to coat the surface of the nanoparticles for delivery of the antibiotics. The polymer responses to nanoparticle-mediated photothermal effect and expand its network to release controllable amount of antibiotics to eradicate the bacterial cells. Conjugation of a specific antibody to the nanoparticle platform increases the therapeutic effect to the S. aureus. This approach may lead to new treatment for implant-associated infections.
5:00 AM - KK5.08
Targeted Delivery of Antibiotics to Cells Infected with Francisella Tularensis Using Mesoporous Silica Nanoparticle-Supported Lipid Bilayers.
Carlee Ashley 1 Christopher Lino 1 Brandon Slaughter 1 Amber McBride 2 Marissa R. Anderson 1 Patrick Fleig 1 Andrew Gomez 1 Caroline Bouvie 2 C. Jeffrey Brinker 1 2 Eric C Carnes 1
1Sandia National Laboratories Albuquerque United States2University of New Mexico Albuquerque United StatesShow Abstract
Although nanotechnology promises to revolutionize the treatment of infectious disease, existing state-of-the-art nanoparticle delivery vehicles, including many liposomal and polymeric nanoparticle formulations, suffer from limited capacities, uncontrollable release profiles, and complex, specialized synthesis procedures that must be re-adapted for each new cargo molecule, leading to drug- and disease-specific ‘one-off&’ approaches. To address these limitations, we have developed mesoporous silica nanoparticle-supported lipid bilayers (‘protocells&’ - see Nature Materials (2011), 10: 389-397) for high capacity, cell-specific delivery of various therapeutic molecules, including antibiotics. Protocells are composed of a mesoporous silica nanoparticle (MSNP) core encased within a supported lipid bilayer (SLB). MSNPs have high surface areas and can, therefore, be loaded with 20-55 wt% of acidic, basic, and hydrophobic antibiotics, capacities that are 100 to 1000-fold higher than similarly-sized liposomes and polymeric nanoparticles. Furthermore, by controlling the degree of silica condensation in the MSNP core, release rates can be precisely tailored from 100% release within 12 hours to 2% release for nearly two months. Fusion of liposomes to antibiotic-loaded MSNPs creates a coherent SLB that enhances the colloidal stability of protocells in blood and helps retain encapsulated drugs within the MSNP core until protocells reach their target organ or cell. Furthermore, the SLB provides a biocompatible interface for display of targeting and endosomolytic moieties, which we have shown trigger efficient, cell-specific uptake of protocells, followed by cytosolic dispersion of encapsulated antibiotics. Specifically, protocells loaded with the antibiotic, levofloxacin, and targeted to cells infected by Francisella tularensis SCHU S4 are internalized by target cells 10,000-times more efficiently than by non-target cells and kill intracellular SCHU S4 more effectively than free levofloxacin and levofloxacin-loaded liposomes at just 2 wt% loading (one-twentieth the protocell&’s maximum loading capacity). In summary, protocells combine high cargo capacities, long-term stability in blood, high targeting specificity, and controllable release kinetics and are, therefore, a promising nanoparticle-based delivery platform. We are currently adapting protocells for inhalational administration and testing the pharmacokinetics, efficacy, and safety of levofloxacin-loaded protocells in rodents and non-human primates infected with aerosolized Francisella tularensis. Furthermore, to facilitate clinical translation, we are addressing reproducibility, purity, scalability, cost, and shelf-life in the absence of cold chain.
5:15 AM - KK5.09
Dextran Coated Cerium Oxide Nanoparticles Act as Antioxidants
Ece Alpaslan 1 Merlyn Vargas 2 Amit K. Roy 1 Thomas J. Webster 1
1Northeastern University Boston United States2University of Antioquia UdeA Medelliacute;n ColombiaShow Abstract
The aim of this study was to evaluate the potential use of the antioxidant activity of surface -modified cerium oxide in rescuing human dermal fibroblast cells in the presence of hydrogen peroxide (H2O2) or hydroquinone (HYQ). It is widely known that various stressors like UV, heavy metals, drugs and other environmental agents constantly challenge cells. If unchecked, these stressors lead to diseases including inflammation, premature aging, neurodegeneration, various other disorders, and cancer. Oxidative stress is responsible for generating various types of reactive oxygen species (ROS), such as the superoxide anion (O2-), the hydroxyl radical (OH) and H2O2. ROS scavengers have become an important area to focus on nano-technology and apply to nano-medicine. With recent reports on cerium oxide nanoparticles being neuroprotective, radioprotective and anti-inflammatory, cerium oxide nanoparticles may be potential free radical scavengers which allow us to use it as a therapeutic agent to fight against cancer and other diseases due to a reduction in ROS. Considering these facts, cerium oxide nanoparticles may promote cell survival under oxidative stress. Ceria nanoparticles were synthesized from 1 mL aqueous solutions of 1 M cerium nitrate (Sigma Aldrich, St Louis, MO) and 2 mL of 0.1 M dextran T-10 (Pharmacosmos, Holback, Denmark) and these solutions were added drop wise to 6 mL of a 30% ammonium hydroxide (Sigma Aldrich, St. Louis, MO) solution while stirring for 24 hours at 25 omicron;C. Synthesized nanoparticles were characterized in terms of their size and chemistry via Transmission Electron Microscopy (TEM) and X-ray photoelectron spectroscopy (XPS). TEM results showed that the particles were around 3 nm. XPS data indicated the presence of cerium atoms in dried powders. Cytotoxicity (MTS) assays were also carried out with human dermal fibroblast (ATCC® PCS-201-012trade;) cells for 1 day in culture using DMEM (ATCC® 30-2003trade;), 10% FBS (ATCC® SCRR-30-2020trade;) and a 1% penicillin-streptomycin solution (ATCC® 30-2300trade;). Cells were seeded at a density 5,000 cells/well, allowed to adhere for 24 hours and the following day, the culture was treated with cytotoxic agents like H2O2 or HYQ with the concentration range from 100 to 800µM. After 24 hours of incubation, the MTS reagent was added and was determined. A dose of 500µM for H2O2 and 150µM HYQ cytotoxic agents was found effective in killing 50% of the cells. In order to determine the cyto-protective function of ceria nanoparticles, some of these cells were preincubated with ceria at a 500µg/mL concentration for 24 hours followed by the addition of cytotoxic agents. The culture was incubated for 24 hours before MTS assays. Results were compared with only 500µM of H2O2 and 150µM HYQ, but not ceria treated cells. The results showed that ceria treated cells were able to recover from the oxidative damage /cytotoxicity exerted by the drugs which suggests that ceria nanoparticles may act as antioxidants within the body.
5:30 AM - KK5.10
Novel Antioxidant Polymeric Prodrug Nanoparticles for the Treatment of Oxidative Stress-AssociatedDiseases
Dongwon Lee 1 Changsun Kang 1 Wooram Cho 1 Wooyoung You 1
1Chonbuk National University Jeonju Korea (the Republic of)Show Abstract
Hydrogen peroxide (H2O2) is one of reactive oxygen species (ROS) and plays a key role as a secondary messenger in normal cellular signaling. However, its overexpression leads to oxidative damages to tissues and organs and has been implicated in inflammatory responses and various life-threatening diseases. Therefore, H2O2 has great potential as a diagnostic and therapeutic biomarker of various inflammatory responses and there is increasing interest in the development of strategies to exploit ROS for diagnostics and therapy. We have developed copolyoxalate (PVAX) as an antioxidant polymeric prodrug that is able to rapidly scavenge H2O2 and exert antioxidant, anti-inflammatory and anti-apoptotic activities. PVAX was designed to incorporate H2O2-responsive peroxalate ester linkages and antioxidant vanillyl alcohol (VA) in its backbone. PVAX nanoparticles readily undergo H2O2-triggered hydrolytic degradation and release VA, which is able to reduce the generation of ROS and exert anti-inflammatory and anti-apoptotic activity. The main culprit in the pathogenesis of ischemia/reperfusion (I/R) injury is the overproduction of H2O2 which is one of the most abundant forms of ROS and causes inflammation, apoptosis and subsequent tissue damages. We therefore evaluated the therapeutic potential of PVAX nanoparticles using a mouse model of hepatic and myocardial I/R injuries. During the hepatic and myocardial I/R injuries, PVAX nanoparticles specifically reacted with overproduced H2O2 and exerted highly potent anti-inflammatory and anti-apoptotic activities, leading to the reduced cellular damages. We anticipate that PVAX nanoparticles have great potential as a novel therapeutic agent for the treatment of oxidative stress associated diseases.
5:45 AM - KK5.11
Exploiting Nanomaterials for Inhibition of UV-Induced Skin Damage
Krysta Biniek 1 Reinhold H. Dauskardt 1
1Stanford University Stanford United StatesShow Abstract
The outermost layer of skin, the stratum corneum (SC), protects the body from harmful environmental conditions such as ultraviolet (UV) exposure by serving as a selective barrier. We explore an emerging class of UV-blocking nanomaterials, inorganic zinc oxide (ZnO) and titanium dioxide (TiO2) nanoparticles (as opposed to conventional micron-sized particles). These are applied to the SC to protect against the harmful effects of solar UV exposure and induce regenerative repair after injury. They diffuse and partition into the SC intercellular boundaries, and the resulting construct is highly effective in preventing erythema from solar UV radiation. However, it remains unclear if these treatments can prevent degeneration in the biomechanical barrier of the SC. We have previously shown that solar UV radiation poses a double threat to the SC by increasing the driving force for cracking while simultaneously decreasing the SC&’s resistance to cracking by significantly reducing cellular cohesion, largely dominated by the intercellular lipids and corneodesmosomes, thereby impairing the critical barrier function of the skin.
We explored the interaction of zinc oxide and titanium dioxide nanoparticles with SC in the presence of UV radiation. We quantitatively determined the drying stresses, and hence the driving force for damage, that occur with UV exposure and the ability of inorganic nanoparticles to mitigate this damage. We also explored the ability of the nanoparticles to protect the innate SC resistance to corneocyte separation, which has been shown to significantly decrease under UV exposure. We found that the inorganic UV inhibitors protected the SC&’s mechanical properties remarkably well under relatively large doses of UV radiation. We also compared efficacy of the inorganic particles to chemical (UV absorbing) sunscreens. Clinical implications of this work include prevention and treatment of sunburn and long term skin damage such as photo-aging.
KK4: Rational Design of Nanomaterials for Bioapplications
Wednesday AM, April 08, 2015
Park Central Hotel, 2nd Floor, Metropolitan I
9:00 AM - *KK4.01
Liposomal Spherical Nucleic Acids: A New Approach to Immuno-Modulatory and Gene Regulation Therapies
Chad A. Mirkin 1
1Northwestern University Evanston United StatesShow Abstract
The natural defenses of biological systems for exogenous oligonucleotides, such as synthetic antisense DNA and siRNA, present many challenges for the delivery of nucleic acids in an efficient, non-toxic, and non-immunogenic fashion. Indeed, because nucleic acids are negatively charged and prone to enzymatic degradation, researchers have historically relied on transfection agents such as cationic polymers, liposomes, and modified viruses to facilitate cellular entry and protect delivered biomolecules from degradation. However, each of these platforms is subject to several drawbacks, which include toxicity at high concentrations, the requirement of specialty nucleic acids to enhance stability, and severe immunogenicity.
Spherical nucleic acid (SNA) nanoparticle conjugates pose one possible solution for circumventing these problems in the context of both immunomodulatory and RNAi pathways. SNAs are typically synthesized from inorganic nanoparticle templates that are functionalized with a spherical shell of densely organized, highly oriented nucleic acids. Remarkably, these highly negatively charged SNA structures do not require cationic transfection agents or additional particle surface modifications and naturally enter all cell lines tested to date (over 50, including primary cells). This talk describes novel liposomal SNAs representing a new class of single entity gene regulation agents made exclusively from components that are biocompatible and components of FDA-approved therapeutics. These novel therapeutics are constructed of 30 nm liposomal cores stabilized with a dense shell of oligonucleotide with a hydrophobic tail that can intercalate between the phospholipids that define the liposome structure. First, we have discovered that oligonucleotide shells can stabilize small (sub-50 nm) liposomal particles and in the process create a potentially very useful class of metal-free SNA. As with conventional SNAs based upon gold cores, these novel liposomal structures rapidly enter multiple cell lines without the need for ancillary transfection agents and can be used to effectively knockdown gene expression via antisense pathways. Moreover, they are indefinitely stable in serum containing media and can be easily made at scale from readily available starting materials. The fact that they do not require the polymeric, viral, or lipoplex co-carriers employed by most researchers is critical since such materials pose significant obstacles in terms of toxicity and immunogenicity.
KK6: Poster Session II
Wednesday PM, April 08, 2015
Marriott Marquis, Yerba Buena Level, Salon 7/8/9
9:00 AM - KK6.01
Self-Folded Redox/Acid Dual-Responsive Nanocarriers for Anticancer Drug Delivery
Yue Lu 1 2 Ran Mo 1 2 Wanyi Tai 1 <