Publications by authors named "Forrest M Kievit"

51 Publications

Antioxidant thioether core-crosslinked nanoparticles prevent the bilateral spread of secondary injury to protect spatial learning and memory in a controlled cortical impact mouse model of traumatic brain injury.

Biomaterials 2021 05 22;272:120766. Epub 2021 Mar 22.

Department of Biological Systems Engineering, University of Nebraska - Lincoln, 200LW Chase Hall, Lincoln, NE, 68583, USA. Electronic address:

The secondary phase of traumatic brain injury (TBI) is partly caused by the release of excess reactive oxygen species (ROS) from the primary injury. However, there are currently no therapies that have been shown to reduce the secondary spread of injury beyond the primary insult. Nanoparticles offer the ability to rapidly accumulate and be retained in injured brain for improved target engagement. Here, we utilized systemically administered antioxidant thioether core-cross-linked nanoparticles (NP1) that scavenge and inactivate ROS to reduce this secondary spread of injury in a mild controlled cortical impact (CCI) mouse model of TBI. We found that NP1 treatment protected CCI mice from injury induced learning and memory deficits observed in the Morris water maze (MWM) test at 1-month post-CCI. This protection was likely a result of NP1-mediated reduction in oxidative stress in the ipsilateral hemisphere as determined by immunofluorescence imaging of markers of oxidative stress and the spread of neuroinflammation into the contralateral hippocampus as determined by immunofluorescence imaging of activated microglia and neuron-astrocyte-microglia triad formation. These data suggest NP1-mediated reduction in post-traumatic oxidative stress correlates with the reduction in the spread of injury to the contralateral hippocampus to protect spatial memory and learning in CCI mice. Therefore, these materials may offer an improved treatment strategy to reduce the secondary spread of TBI.
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http://dx.doi.org/10.1016/j.biomaterials.2021.120766DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8068673PMC
May 2021

siRNA nanoparticle suppresses drug-resistant gene and prolongs survival in an orthotopic glioblastoma xenograft mouse model.

Adv Funct Mater 2021 Feb 6;31(6). Epub 2020 Nov 6.

Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195, United States; Department of Neurological Surgery, University of Washington, Seattle, WA 98195, United States.

Temozolomide (TMZ) is the standard of care chemotherapy drug for treating glioblastomas (GBMs), the most aggressive cancer that affects people of all ages. However, its therapeutic efficacy is limited by the drug resistance mediated by a DNA repair protein, O-methylguanine-DNA methyltransferase (MGMT), which eliminates the TMZ-induced DNA lesions. Here we report the development of an iron oxide nanoparticle (NP) system for targeted delivery of siRNAs to suppress the TMZ-resistance gene (MGMT). We show that our NP is able to overcome biological barriers, bind specifically to tumor cells, and reduce MGMT expression in tumors of mice bearing orthotopic GBM serially-passaged patient-derived xenografts. The treatment with sequential administration of this NP and TMZ resulted in increased apoptosis of GBM stem-like cells, reduced tumor growth, and significantly-prolonged survival as compared to mice treated with TMZ alone. This study introduces an approach that holds great promise to improve the outcomes of GBM patients.
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http://dx.doi.org/10.1002/adfm.202007166DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7942690PMC
February 2021

Ultrasmall Mixed Eu-Gd Oxide Nanoparticles for Multimodal Fluorescence and Magnetic Resonance Imaging of Passive Accumulation and Retention in TBI.

ACS Omega 2020 Jul 23;5(26):16220-16227. Epub 2020 Jun 23.

Department of Biological Systems Engineering, University of Nebraska-Lincoln, 3605 Fair Street, Lincoln, Nebraska 68583-0726, United States.

Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. TBI can have a long-term impact on the quality of life for survivors of all ages. However, there remains no approved treatment that improves outcomes following TBI, which is partially due to poor delivery of therapies into the brain. Therefore, there is a significant unmet need to develop more effective delivery strategies that increase the accumulation and retention of potentially efficacious treatments in the injured brain. Recent work has revealed that nanoparticles (NPs) may offer a promising approach for site-specific delivery; however, a detailed understanding of the specific NP properties that promote brain accumulation and retention are still being developed. Multimodal imaging plays a vital role in the understanding of physicochemical properties that initiate the uptake and accumulation of NPs in the brain at both high spatial (e.g., fluorescence imaging) and temporal (e.g., magnetic resonance imaging, MRI) frequency. However, many NP systems that are currently used in TBI only provide contrast in a single imaging modality limiting the imaging data that can be obtained, and those that offer multimodal imaging capabilities have complicated multistep synthesis methods. Therefore, the goal of this work was to develop an ultrasmall NP with simple fabrication capable of multimodal imaging. Here, we describe the development, characterization, accumulation, and retention of poly(ethylene glycol) (PEG)-coated europium-gadolinium (Eu-Gd) mixed magnetic NPs (MNPs) in a controlled cortical impact mouse model of TBI. We find that these NPs having an ultrasmall core size of 2 nm and a small hydrodynamic size of 13.5 nm can be detected in both fluorescence and MR imaging modalities and rapidly accumulate and are retained in injured brain parenchyma. These NPs should allow for further testing of NP physicochemical properties that promote accumulation and retention in TBI and other disease models.
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http://dx.doi.org/10.1021/acsomega.0c01890DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7346268PMC
July 2020

Evaluating differential nanoparticle accumulation and retention kinetics in a mouse model of traumatic brain injury via K mapping with MRI.

Sci Rep 2019 11 6;9(1):16099. Epub 2019 Nov 6.

Department of Biological Systems Engineering, University of Nebraska, 200 LW Chase Hall, Lincoln, NE, 68583, USA.

Traumatic brain injury (TBI) is a leading cause of injury-related death worldwide, yet there are no approved neuroprotective therapies that improve neurological outcome post-injury. Transient opening of the blood-brain barrier following injury provides an opportunity for passive accumulation of intravenously administered nanoparticles through an enhanced permeation and retention-like effect. However, a thorough understanding of physicochemical properties that promote optimal uptake and retention kinetics in TBI is still needed. In this study, we present a robust method for magnetic resonance imaging of nanoparticle uptake and retention kinetics following intravenous injection in a controlled cortical impact mouse model of TBI. Three contrast-enhancing nanoparticles with different hydrodynamic sizes and relaxivity properties were compared. Accumulation and retention were monitored by modelling the permeability coefficient, K, for each nanoparticle within the reproducible mouse model. Quantification of K for different nanoparticles allowed for non-invasive, multi-time point assessment of both accumulation and retention kinetics in the injured tissue. Using this method, we found that 80 nm poly(lactic-co-glycolic acid) nanoparticles had maximal K in a TBI when injected 3 hours post-injury, showing significantly higher accumulation kinetics than the small molecule, Gd-DTPA. This robust method will enable optimization of administration time and nanoparticle physicochemical properties to achieve maximum delivery.
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http://dx.doi.org/10.1038/s41598-019-52622-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6834577PMC
November 2019

A Role for Nanoparticles in Treating Traumatic Brain Injury.

Pharmaceutics 2019 Sep 13;11(9). Epub 2019 Sep 13.

Department of Biological Systems Engineering, University of Nebraska-Lincoln, Lincoln, NE 68583-0726, USA.

Traumatic brain injury (TBI) is one of the main causes of disability in children and young adults, as well as a significant concern for elderly individuals. Depending on the severity, TBI can have a long-term impact on the quality of life for survivors of all ages. The primary brain injury can result in severe disability or fatality, and secondary brain damage can increase the complexities in cellular, inflammatory, neurochemical, and metabolic changes in the brain, which can last decades post-injury. Thus, survival from a TBI is often accompanied by lifelong disabilities. Despite the significant morbidity, mortality, and economic loss, there are still no effective treatment options demonstrating an improved outcome in a large multi-center Phase III trial, which can be partially attributed to poor target engagement of delivered therapeutics. Thus, there is a significant unmet need to develop more effective delivery strategies to overcome the biological barriers that would otherwise inhibit transport of materials into the brain to prevent the secondary long-term damage associated with TBI. The complex pathology of TBI involving the blood-brain barrier (BBB) has limited the development of effective therapeutics and diagnostics. Therefore, it is of great importance to develop novel strategies to target the BBB. The leaky BBB caused by a TBI may provide opportunities for therapeutic delivery via nanoparticles (NP). The focus of this review is to provide a survey of NP-based strategies employed in preclinical models of TBI and to provide insights for improved NP based diagnostic or treatment approaches. Both passive and active delivery of various NPs for TBI are discussed. Finally, potential therapeutic targets where improved NP-mediated delivery could increase target engagement are identified with the overall goal of providing insight into open opportunities for NP researchers to begin research in TBI.
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http://dx.doi.org/10.3390/pharmaceutics11090473DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6781280PMC
September 2019

Nanoparticle Biokinetics in Mice and Nonhuman Primates.

ACS Nano 2017 09 18;11(9):9514-9524. Epub 2017 Sep 18.

Clinical Research Division, Fred Hutchinson Cancer Research Center , Seattle, Washington 98109, United States.

Despite the preponderance of iron oxide nanoparticles (NPs) designed for theranostic applications, widespread clinical translation of these NPs lags behind. A better understanding of how NP pharmacokinetics vary between small and large animal models is needed to rapidly customize NPs for optimal performance in humans. Here we use noninvasive magnetic resonance imaging (MRI) to track iron oxide NPs through a large number of organ systems in vivo to investigate NP biokinetics in both mice and nonhuman primates. We demonstrate that pharmacokinetics are similar between mice and macaques in the blood, liver, spleen, and muscle, but differ in the kidneys, brain, and bone marrow. Our study also demonstrates that full-body MRI is practical, rapid, and cost-effective for tracking NPs noninvasively with high spatiotemporal resolution. Our techniques using a nonhuman primate model may provide a platform for testing a range of NP formulations.
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http://dx.doi.org/10.1021/acsnano.7b05377DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6002853PMC
September 2017

Core-Cross-Linked Nanoparticles Reduce Neuroinflammation and Improve Outcome in a Mouse Model of Traumatic Brain Injury.

ACS Nano 2017 09 16;11(9):8600-8611. Epub 2017 Aug 16.

Department of BioEngineering, Molecular Engineering and Sciences Institute , Box 355061, Seattle, Washington 98195, United States.

Traumatic brain injury (TBI) is the leading cause of death and disability in children and young adults, yet there are currently no treatments available that prevent the secondary spread of damage beyond the initial insult. The chronic progression of this secondary injury is in part caused by the release of reactive oxygen species (ROS) into surrounding normal brain. Thus, treatments that can enter the brain and reduce the spread of ROS should improve outcome from TBI. Here a highly versatile, reproducible, and scalable method to synthesize core-cross-linked nanoparticles (NPs) from polysorbate 80 (PS80) using a combination of thiol-ene and thiol-Michael chemistry is described. The resultant NPs consist of a ROS-reactive thioether cross-linked core stabilized in aqueous solution by hydroxy-functional oligoethylene oxide segments. These NPs show narrow molecular weight distributions and have a high proportion of thioether units that reduce local levels of ROS. In a controlled cortical impact mouse model of TBI, the NPs are able to rapidly accumulate and be retained in damaged brain as visualized through fluorescence imaging, reduce neuroinflammation and the secondary spread of injury as determined through magnetic resonance imaging and histopathology, and improve functional outcome as determined through behavioral analyses. Our findings provide strong evidence that these NPs may, upon further development and testing, provide a useful strategy to help improve the outcome of patients following a TBI.
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http://dx.doi.org/10.1021/acsnano.7b03426DOI Listing
September 2017

Nanoparticle-mediated knockdown of DNA repair sensitizes cells to radiotherapy and extends survival in a genetic mouse model of glioblastoma.

Nanomedicine 2017 Oct 11;13(7):2131-2139. Epub 2017 Jun 11.

Department of Neurological Surgery, University of Washington, Seattle, WA, United States; Department of Materials Science and Engineering, University of Washington, Seattle, WA, United States; Department of Radiology, University of Washington, Seattle, WA, United States. Electronic address:

Glioblastoma (GBM) remains incurable, and recurrent tumors rarely respond to standard-of-care radiation and chemo-therapies. Therefore, strategies that enhance the effects of these therapies should provide significant benefits to GBM patients. We have developed a nanoparticle delivery vehicle that can stably bind and protect nucleic acids for specific delivery into brain tumor cells. These nanoparticles can deliver therapeutic siRNAs to sensitize GBM cells to radiotherapy and improve GBM treatment via systemic administration. We show that nanoparticle-mediated knockdown of the DNA repair protein apurinic endonuclease 1 (Ape1) sensitizes GBM cells to radiotherapy and extend survival in a genetic mouse model of GBM. Specific knockdown of Ape1 activity by 30% in brain tumor tissue doubled the extended survival achieved with radiotherapy alone. Ape1 is a promising target for increasing the effectiveness of radiotherapy, and nanoparticle-mediated delivery of siRNA is a promising strategy for tumor specific knockdown of Ape1.
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http://dx.doi.org/10.1016/j.nano.2017.06.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6002851PMC
October 2017

3D Porous Chitosan-Alginate Scaffolds Promote Proliferation and Enrichment of Cancer Stem-Like Cells.

J Mater Chem B 2016 Oct 31;4(38):6326-6334. Epub 2016 Aug 31.

Department of Materials Science & Engineering, University of Washington, Seattle, WA 98195.

Cancer stem cells are increasingly becoming a primary target for new cancer treatment development. The ability to study their transient behavior in vitro will provide the opportunity for high-throughput testing of more effective therapies. We have previously demonstrated the use of 3D porous chitosan-alginate (CA) scaffolds to promote cancer stem-like cell (CSC) proliferation and enrichment in glioblastoma. Here we use 3D porous CA scaffolds to promote cancer stem-like cell enrichment in cell lines from prostate, liver, and breast cancers, and investigate the proliferation, morphology, and gene expressions of cells cultured in CA scaffolds as compared to 2D controls. The 3D CA scaffold cultures for all three cancer types showed reduced proliferation, formation of tumor spheroids, and increased expression of CSC associated mark genes (CD133 and NANOG), as opposed to monolayers. Additionally, we present a putative mechanism for the cancer stem-like cell enrichment on CA scaffolds. This study demonstrates that the cancer stem-like cell enrichment in CA scaffolds is a robust process that is not restricted to particular cancer types.
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http://dx.doi.org/10.1039/C6TB01713DDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5260821PMC
October 2016

Nanoparticles for cancer gene therapy: Recent advances, challenges, and strategies.

Pharmacol Res 2016 12 19;114:56-66. Epub 2016 Oct 19.

Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195, USA; Department of Neurological Surgery, University of Washington, Seattle, WA 98195, USA. Electronic address:

Compared to conventional treatments, gene therapy offers a variety of advantages for cancer treatment including high potency and specificity, low off-target toxicity, and delivery of multiple genes that concurrently target cancer tumorigenesis, recurrence, and drug resistance. In the past decades, gene therapy has undergone remarkable progress, and is now poised to become a first line therapy for cancer. Among various gene delivery systems, nanoparticles have attracted much attention because of their desirable characteristics including low toxicity profiles, well-controlled and high gene delivery efficiency, and multi-functionalities. This review provides an overview on gene therapeutics and gene delivery technologies, and highlight recent advances, challenges and insights into the design and the utility of nanoparticles in gene therapy for cancer treatment.
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http://dx.doi.org/10.1016/j.phrs.2016.10.016DOI Listing
December 2016

Culture on 3D Chitosan-Hyaluronic Acid Scaffolds Enhances Stem Cell Marker Expression and Drug Resistance in Human Glioblastoma Cancer Stem Cells.

Adv Healthc Mater 2016 12 2;5(24):3173-3181. Epub 2016 Nov 2.

Department of Materials Science and Engineering, University of Washington, Seattle, WA, 98195, USA.

The lack of in vitro models that support the growth of glioblastoma (GBM) stem cells (GSCs) that underlie clinical aggressiveness hinders developing new, effective therapies for GBM. While orthotopic patient-derived xenograft models of GBM best reflect in vivo tumor behavior, establishing xenografts is a time consuming, costly, and frequently unsuccessful endeavor. To address these limitations, a 3D porous scaffold composed of chitosan and hyaluronic acid (CHA) is synthesized. Growth and expression of the cancer stem cell (CSC) phenotype of the GSC GBM6 taken directly from fresh xenogratfs grown on scaffolds or as adherent monolayers is compared. While 2D adherent cultures grow as monolayers of flat epitheliod cells, GBM6 cells proliferate within pores of CHA scaffolds as clusters of self-adherent ovoid cells. Growth on scaffolds is accompanied by greater expression of genes that mediate epithelial-mesenchymal transition and maintain a primitive, undifferentiated phenotype, hallmarks of CSCs. Scaffold-grown cells also display higher expression of genes that promote resistance to hypoxia-induced oxidative stress. In accord, scaffold-grown cells show markedly greater resistance to clinically utilized alkylating agents compared to adherent cells. These findings suggest that our CHA scaffolds better mimic in vivo biological and clinical behavior and provide insights for developing novel individualized treatments.
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http://dx.doi.org/10.1002/adhm.201600684DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5253135PMC
December 2016

Modeling the tumor microenvironment using chitosan-alginate scaffolds to control the stem-like state of glioblastoma cells.

Biomater Sci 2016 Apr 21;4(4):610-3. Epub 2015 Dec 21.

Department of Neurological Surgery, University of Washington, Seattle, WA 98195, USA.

Better prediction of in vivo drug efficacy using in vitro models should greatly improve in vivo success. Here we utilize 3D highly porous chitosan-alginate complex scaffolds to probe how various components of the glioblastoma microenvironment including extracellular matrix and stromal cells affect tumor cell stem-like state.
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http://dx.doi.org/10.1039/c5bm00514kDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4803622PMC
April 2016

Iron-Oxide-Based Nanovector for Tumor Targeted siRNA Delivery in an Orthotopic Hepatocellular Carcinoma Xenograft Mouse Model.

Small 2016 Jan 7;12(4):477-87. Epub 2015 Dec 7.

Department of Materials Science and Engineering, University of Washington, Seattle, WA, 98195, USA.

Hepatocellular carcinoma (HCC) is one of the deadliest cancers worldwide. Small interfering RNA (siRNA) holds promise as a new class of therapeutics for HCC, as it can achieve sequence-specific gene knockdown with low cytotoxicity. However, the main challenge in the clinical application of siRNA lies in the lack of effective delivery approaches that need to be highly specific and thus incur low or no systemic toxicity. Here, a nonviral nanoparticle-based gene carrier is presented that can specifically deliver siRNA to HCC. The nanovector (NP-siRNA-GPC3 Ab) is made of an iron oxide core coated with chitosan-polyethylene glycol (PEG) grafted polyethyleneimine copolymer, which is further functionalized with siRNA and conjugated with a monoclonal antibody (Ab) against human glypican-3 (GPC3) receptor highly expressed in HCC. A rat RH7777 HCC cell line that coexpresses human GPC3 and firefly luciferase (Luc) is established to evaluate the nanovector. The nanoparticle-mediated delivery of siRNA against Luc effectively suppresses Luc expression in vitro without notable cytotoxicity. Significantly, NP-siLuc-GPC3 Ab administered intravenously in an orthotopic model of HCC is able to specifically bound to tumor and induce remarkable inhibition of Luc expression. The findings demonstrate the potential of using this nanovector for targeted delivery of therapeutic siRNA to HCC.
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http://dx.doi.org/10.1002/smll.201501985DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4829640PMC
January 2016

Nanoparticle-Mediated Target Delivery of TRAIL as Gene Therapy for Glioblastoma.

Adv Healthc Mater 2015 Dec 26;4(17):2719-26. Epub 2015 Oct 26.

Department of Materials Science and Engineering, University of Washington, Seattle, WA, 98195, USA.

Human tumor necrosis factor α-related apoptosis-inducing ligand (TRAIL) is an attractive cancer therapeutic because of its ability to induce apoptosis in tumor cells while having a negligible effect on normal cells. However, the short serum half-life of TRAIL and lack of efficient in vivo administration approaches have largely hindered its clinical use. Using nanoparticles (NPs) as carriers in gene therapy is considered as an alternative approach to increase TRAIL delivery to tumors as transfected cells would be induced to secrete TRAIL into the tumor microenvironment. To enable effective delivery of plasmid DNA encoding TRAIL into glioblastoma (GBM), we developed a targeted iron oxide NP coated with chitosan-polyethylene glycol-polyethyleneimine copolymer and chlorotoxin (CTX) and evaluated its effect in delivering TRAIL in vitro and in vivo. NP-TRAIL successfully delivers TRAIL into human T98G GBM cells and induces secretion of 40 pg mL(-1) of TRAIL in vitro. Transfected cells show threefold increased apoptosis as compared to the control DNA bound NPs. Systemic administration of NP-TRAIL-CTX to mice bearing T98G-derived flank xenografts results in near-zero tumor growth and induces apoptosis in tumor tissue. Our results suggest that NP-TRAIL-CTX can potentially serve as a targeted anticancer therapeutic for more efficient TRAIL delivery to GBM.
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http://dx.doi.org/10.1002/adhm.201500563DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4715716PMC
December 2015

Anti-HER2/neu peptide-conjugated iron oxide nanoparticles for targeted delivery of paclitaxel to breast cancer cells.

Nanoscale 2015 Nov;7(43):18010-4

Departments of Materials Science and Engineering, University of Washington, Seattle, Washington 98195, USA.

Nanoparticles (NPs) for targeted therapy are required to have appropriate size, stability, drug loading and release profiles, and efficient targeting ligands. However, many of the existing NPs such as albumin, liposomes, polymers, gold NPs, etc. encounter size limit, toxicity and stability issues when loaded with drugs, fluorophores, and targeting ligands. Furthermore, antibodies are bulky and this can greatly affect the physicochemical properties of the NPs, whereas many small molecule-based targeting ligands lack specificity. Here, we report the utilization of biocompatible, biodegradable, small (∼30 nm) and stable iron oxide NPs (IONPs) for targeted delivery of paclitaxel (PTX) to HER2/neu positive breast cancer cells using an anti-HER2/neu peptide (AHNP) targeting ligand. We demonstrate the uniform size and high stability of these NPs in biological medium, their effective tumour targeting in live mice, as well as their efficient cellular targeting and selective killing in human HER2/neu-positive breast cancer cells.
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http://dx.doi.org/10.1039/c5nr04867bDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4683026PMC
November 2015

3D Porous Chitosan-Alginate Scaffolds as an In Vitro Model for Evaluating Nanoparticle-Mediated Tumor Targeting and Gene Delivery to Prostate Cancer.

Biomacromolecules 2015 Oct 16;16(10):3362-72. Epub 2015 Sep 16.

Department of Materials Science and Engineering and ‡Department of Neurological Surgery, University of Washington , Seattle, Washington 98195, United States.

Cationic nanoparticles (NPs) for targeted gene delivery are conventionally evaluated using 2D in vitro cultures. However, this does not translate well to corresponding in vivo studies because of the marked difference in NP behavior in the presence of the tumor microenvironment. In this study, we investigated whether prostate cancer (PCa) cells cultured in three-dimensional (3D) chitosan-alginate (CA) porous scaffolds could model cationic NP-mediated gene targeted delivery to tumors in vitro. We assessed in vitro tumor cell proliferation, formation of tumor spheroids, and expression of marker genes that promote tumor malignancy in CA scaffolds. The efficacy of NP-targeted gene delivery was evaluated in PCa cells in 2D cultures, PCa tumor spheroids grown in CA scaffolds, and PCa tumors in a mouse TRAMP-C2 flank tumor model. PCa cells cultured in CA scaffolds grew into tumor spheroids and displayed characteristics of higher malignancy as compared to those in 2D cultures. Significantly, targeted gene delivery was only observed in cells cultured in CA scaffolds, whereas cells cultured on 2D plates showed no difference in gene delivery between targeted and nontarget control NPs. In vivo NP evaluation confirmed targeted gene delivery, indicating that only CA scaffolds correctly modeled NP-mediated targeted delivery in vivo. These findings suggest that CA scaffolds serve as a better in vitro platform than 2D cultures for evaluation of NP-mediated targeted gene delivery to PCa.
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http://dx.doi.org/10.1021/acs.biomac.5b01032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4831622PMC
October 2015

Temozolomide nanoparticles for targeted glioblastoma therapy.

ACS Appl Mater Interfaces 2015 Apr 18;7(12):6674-82. Epub 2015 Mar 18.

‡Department of Materials Science and Engineering, University of Washington, Seattle, Washington 98195, United States.

Glioblastoma (GBM) is a deadly and debilitating brain tumor with an abysmal prognosis. The standard therapy for GBM is surgery followed by radiation and chemotherapy with Temozolomide (TMZ). Treatment of GBMs remains a challenge, largely because of the fast degradation of TMZ, the inability to deliver an effective dose of TMZ to tumors, and a lack of target specificity that may cause systemic toxicity. Here, we present a simple method for synthesizing a nanoparticle-based carrier that can protect TMZ from rapid degradation in physiological solutions and can specifically deliver them to GBM cells through the mediation of a tumor-targeting peptide chlorotoxin (CTX). Our nanoparticle, namely NP-TMZ-CTX, had a hydrodynamic size of <100 nm, exhibited sustained stability in cell culture media for up to 2 weeks, and could accommodate stable drug loading. TMZ bound to nanoparticles showed a much higher stability at physiological pH, with a half-life 7-fold greater than that of free TMZ. NP-TMZ-CTX was able to target GBM cells and achieved 2-6-fold higher uptake and a 50-90% reduction of IC50 72 h post-treatment as compared to nontargeted NP-TMZ. NP-TMZ-CTX showed great promise in its ability to deliver a large therapeutic dose of TMZ to GBM cells and could serve as a template for targeted delivery of other therapeutics.
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http://dx.doi.org/10.1021/am5092165DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4637162PMC
April 2015

Bionanotechnology and the future of glioma.

Surg Neurol Int 2015 13;6(Suppl 1):S45-58. Epub 2015 Feb 13.

Department of Neurological Surgery, University of Washington, Seattle, Washington 98195, USA.

Designer nanoscaled materials have the potential to revolutionize diagnosis and treatment for glioma. This review summarizes current progress in nanoparticle-based therapies for glioma treatment including targeting, drug delivery, gene delivery, and direct tumor ablation. Preclinical and current human clinical trials are discussed. Although progress in the field has been significant over the past decade, many successful strategies demonstrated in the laboratory have yet to be implemented in human clinical trials. Looking forward, we provide examples of combined treatment strategies, which harness the potential for nanoparticles to interact with their biochemical environment, and simultaneously with externally applied photons or magnetic fields. We present our notion of the "ideal" nanoparticle for glioma, a concept that may soon be realized.
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http://dx.doi.org/10.4103/2152-7806.151334DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4338483PMC
February 2015

Nanoparticle mediated silencing of DNA repair sensitizes pediatric brain tumor cells to γ-irradiation.

Mol Oncol 2015 Jun 29;9(6):1071-80. Epub 2015 Jan 29.

Department of Neurological Surgery, University of Washington, Seattle, WA 98195, USA; Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195, USA; Department of Radiology, University of Washington, Seattle, WA 98195, USA. Electronic address:

Medulloblastoma (MB) and ependymoma (EP) are the most common pediatric brain tumors, afflicting 3000 children annually. Radiotherapy (RT) is an integral component in the treatment of these tumors; however, the improvement in survival is often accompanied by radiation-induced adverse developmental and psychosocial sequelae. Therefore, there is an urgent need to develop strategies that can increase the sensitivity of brain tumors cells to RT while sparing adjacent healthy brain tissue. Apurinic endonuclease 1 (Ape1), an enzyme in the base excision repair pathway, has been implicated in radiation resistance in cancer. Pharmacological and specificity limitations inherent to small molecule inhibitors of Ape1 have hindered their clinical development. Here we report on a nanoparticle (NP) based siRNA delivery vehicle for knocking down Ape1 expression and sensitizing pediatric brain tumor cells to RT. The NP comprises a superparamagnetic iron oxide core coated with a biocompatible, biodegradable coating of chitosan, polyethylene glycol (PEG), and polyethyleneimine (PEI) that is able to bind and protect siRNA from degradation and to deliver siRNA to the perinuclear region of target cells. NPs loaded with siRNA against Ape1 (NP:siApe1) knocked down Ape1 expression over 75% in MB and EP cells, and reduced Ape1 activity by 80%. This reduction in Ape1 activity correlated with increased DNA damage post-irradiation, which resulted in decreased cell survival in clonogenic assays. The sensitization was specific to therapies generating abasic lesions as evidenced by NP:siRNA not increasing sensitivity to paclitaxel, a microtubule disrupting agent. Our results indicate NP-mediated delivery of siApe1 is a promising strategy for circumventing pediatric brain tumor resistance to RT.
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http://dx.doi.org/10.1016/j.molonc.2015.01.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4439369PMC
June 2015

Glypican-3-targeting F(ab')2 for 89Zr PET of hepatocellular carcinoma.

J Nucl Med 2014 Dec 30;55(12):2032-7. Epub 2014 Oct 30.

Department of Surgery, University of Washington, Seattle, Washington

Unlabelled: Hepatocellular carcinoma (HCC) is an increasingly lethal malignancy for which management is critically dependent on accurate imaging. Glypican-3 (GPC3) is a cell surface receptor overexpressed in most HCCs and provides a unique target for molecular diagnostics. The use of monoclonal antibodies (mAbs) that target GPC3 (αGPC3) in PET imaging has shown promise but comes with inherent limitations associated with mAbs such as long circulation times. This study used (89)Zr-conjugated F(ab')2 fragments directed against GPC3 ((89)Zr-αGPC3-F(ab')2) to evaluate the feasibility of the fragments as a diagnostic immuno-PET imaging probe.

Methods: Immobilized ficin was used to digest αGPC3, creating αGPC3-F(ab')2 fragments subsequently conjugated to (89)Zr. In vivo biodistribution and PET studies were performed on GPC3-expressing HepG2 and GPC3-nonexpressing RH7777 orthotopic xenografts.

Results: Reliable αGPC3-F(ab')2 production via immobilized ficin digestion was verified by high-performance liquid chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. (89)Zr-αGPC3-F(ab')2 demonstrated F(ab')2-dependent, antigen-specific cell binding. HepG2 tumor uptake was higher than any other tissue, peaking at 100 ± 21 percentage injected dose per gram (%ID/g) 24 h after injection, a value 33- to 38-fold higher than GPC3-nonexpressing RH7777 tumors. The blood half-life of the (89)Zr-αGPC3-F(ab')2 conjugate was approximately 11 h, compared with approximately 115 h for historic mAb controls. This shorter half-life enabled clear tumor visualization on PET 4 h after administration, with a resultant peak tumor-to-liver contrast ratio of 23.3. Blocking antigen-expressing tumors with an excess of nonradiolabeled αGPC3 resulted in decreased tumor uptake similar to native liver. The kidneys exhibited high tissue uptake, peaking at 24 h with 83 ± 12 %ID/g. HepG2 tumors ranging from 1.5 to 7 mm were clearly visible on PET, whereas larger RH7777 tumors displayed signal lower than background liver tissue.

Conclusion: This study demonstrates the feasibility of using (89)Zr-αGPC3-F(ab')2 for intrahepatic tumor localization with small-animal PET. Faster blood clearance and lower background liver uptake enable excellent signal-to-noise ratios at early time points. Increased renal uptake is similar to that as has been seen with clinical radioactive peptide imaging. (89)Zr-αGPC3-F(ab')2 addresses some of the shortcomings of whole-antibody immuno-PET probes. Further optimization is warranted to maximize probe sensitivity and specificity in the process of clinical translation.
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http://dx.doi.org/10.2967/jnumed.114.145102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4259878PMC
December 2014

Redox-responsive magnetic nanoparticle for targeted convection-enhanced delivery of O6-benzylguanine to brain tumors.

ACS Nano 2014 Oct 29;8(10):10383-95. Epub 2014 Sep 29.

Department of Materials Science and Engineering, University of Washington , Seattle, Washington 98195, United States.

Resistance to temozolomide (TMZ) based chemotherapy in glioblastoma multiforme (GBM) has been attributed to the upregulation of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT). Inhibition of MGMT using O(6)-benzylguanine (BG) has shown promise in these patients, but its clinical use is hindered by poor pharmacokinetics that leads to unacceptable toxicity. To improve BG biodistribution and efficacy, we developed superparamagnetic iron oxide nanoparticles (NP) for targeted convection-enhanced delivery (CED) of BG to GBM. The nanoparticles (NPCP-BG-CTX) consist of a magnetic core coated with a redox-responsive, cross-linked, biocompatible chitosan-PEG copolymer surface coating (NPCP). NPCP was modified through covalent attachment of BG and tumor targeting peptide chlorotoxin (CTX). Controlled, localized BG release was achieved under reductive intracellular conditions and NPCP-BG-CTX demonstrated proper trafficking of BG in human GBM cells in vitro. NPCP-BG-CTX treated cells showed a significant reduction in MGMT activity and the potentiation of TMZ toxicity. In vivo, CED of NPCP-BG-CTX produced an excellent volume of distribution (Vd) within the brain of mice bearing orthotopic human primary GBM xenografts. Significantly, concurrent treatment with NPCP-BG-CTX and TMZ showed a 3-fold increase in median overall survival in comparison to NPCP-CTX/TMZ treated and untreated animals. Furthermore, NPCP-BG-CTX mitigated the myelosuppression observed with free BG in wild-type mice when administered concurrently with TMZ. The combination of favorable physicochemical properties, tumor cell specific BG delivery, controlled BG release, and improved in vivo efficacy demonstrates the great potential of these NPs as a treatment option that could lead to improved clinical outcomes.
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http://dx.doi.org/10.1021/nn503735wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4212796PMC
October 2014

Proliferation and enrichment of CD133(+) glioblastoma cancer stem cells on 3D chitosan-alginate scaffolds.

Biomaterials 2014 Nov 7;35(33):9137-43. Epub 2014 Aug 7.

Department of Neurological Surgery, University of Washington, Seattle, WA 98195, USA; Department of Materials Science & Engineering, University of Washington, Seattle, WA 98195, USA. Electronic address:

Emerging evidence implicates cancer stem cells (CSCs) as primary determinants of the clinical behavior of human cancers, representing an ideal target for next-generation anti-cancer therapies. However CSCs are difficult to propagate in vitro, severely limiting the study of CSC biology and drug development. Here we report that growing cells from glioblastoma (GBM) cell lines on three dimensional (3D) porous chitosan-alginate (CA) scaffolds dramatically promotes the proliferation and enrichment of cells possessing the hallmarks of CSCs. CA scaffold-grown cells were found more tumorigenic in nude mouse xenografts than cells grown from monolayers. Growing in CA scaffolds rapidly promoted expression of genes involved in the epithelial-to-mesenchymal transition that has been implicated in the genesis of CSCs. Our results indicate that CA scaffolds have utility as a simple and inexpensive means to cultivate CSCs in vitro in support of studies to understand CSC biology and develop more effective anti-cancer therapies.
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http://dx.doi.org/10.1016/j.biomaterials.2014.07.037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4167581PMC
November 2014

Thermoreversible poly(ethylene glycol)-g-chitosan hydrogel as a therapeutic T lymphocyte depot for localized glioblastoma immunotherapy.

Biomacromolecules 2014 Jul 25;15(7):2656-62. Epub 2014 Jun 25.

Department of Materials Science and Engineering, University of Washington , Seattle, Washington 98195, United States.

The outcome for glioblastoma patients remains dismal for its invariably recrudesces within 2 cm of the resection cavity. Local immunotherapy has the potential to eradicate the residual infiltrative component of these tumors. Here, we report the development of a biodegradable hydrogel containing therapeutic T lymphocytes for localized delivery to glioblastoma cells for brain tumor immunotherapy. Thermoreversible poly(ethylene glycol)-g-chitosan hydrogels (PCgels) were optimized for steady T lymphocyte release. Nuclear magnetic resonance spectroscopy confirmed the chemical structure of poly(ethylene glycol)-g-chitosan, and rheological studies revealed that the sol-to-gel transition of the PCgel occurred around ≥32 °C. T lymphocyte invasion through the PCgel and subsequent cytotoxicity to glioblastoma were assessed in vitro. The PCgel was shown to be cellular compatible with T lymphocytes, and the T lymphocytes retain their anti-glioblastoma activity after being encapsulated in the PCgel. T lymphocytes in the PCgel were shown to be more effective in killing glioblastoma than those in the Matrigel control. This may be attributed to the optimal pore size of the PCgel allowing better invasion of T lymphocytes. Our study suggests that this unique PCgel depot may offer a viable approach for localized immunotherapy for glioblastoma.
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http://dx.doi.org/10.1021/bm500502nDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4215871PMC
July 2014

Chitosan-based thermoreversible hydrogel as an in vitro tumor microenvironment for testing breast cancer therapies.

Mol Pharm 2014 Jul 15;11(7):2134-42. Epub 2014 May 15.

Departments of †Materials Science and Engineering and ‡Neurological Surgery, University of Washington , Seattle, Washington 98195, United States.

Breast cancer is a major health problem for women worldwide. Although in vitro culture of established breast cancer cell lines is the most widely used model for preclinical assessment, it poorly represents the behavior of breast cancers in vivo. Acceleration of the development of effective therapeutic strategies requires a cost-efficient in vitro model that can more accurately resemble the in vivo tumor microenvironment. Here, we report the use of a thermoreversible poly(ethylene glycol)-g-chitosan hydrogel (PCgel) as an in vitro breast cancer model. We hypothesized that PCgel could provide a tumor microenvironment that promotes cultured cancer cells to a more malignant phenotype with drug and immune resistance. Traditional tissue culture plates and Matrigel were applied as controls in our studies. In vitro cellular proliferation and morphology, the secretion of angiogenesis-related growth factors and cytokines, and drug and immune resistance were assessed. Our results show that PCgel cultures promoted tumor aggregate formation, increased secretion of various angiogenesis- and metastasis-related growth factors and cytokines, and increased tumor cell resistance to chemotherapeutic drugs and immunotherapeutic T cells. This PCgel platform may offer a valuable strategy to bridge the gap between standard in vitro and costly animal studies for a wide variety of experimental designs.
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http://dx.doi.org/10.1021/mp5002119DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4096230PMC
July 2014

Glypican-3-targeted 89Zr PET imaging of hepatocellular carcinoma.

J Nucl Med 2014 May 13;55(5):799-804. Epub 2014 Mar 13.

Department of Surgery, University of Washington, Seattle, Washington.

Unlabelled: Hepatocellular carcinoma (HCC) is a devastating malignancy in which imperfect imaging plays a primary role in diagnosis. Glypican-3 (GPC3) is an HCC-specific cell surface proteoglycan overexpressed in most HCCs. This paper presents the use of (89)Zr-conjugated monoclonal antibody against GPC3 ((89)Zr-αGPC3) for intrahepatic tumor localization using PET.

Methods: Polymerase chain reaction confirmed relative GPC3 expression in cell lines. In vitro binding, in vivo biodistribution, and small-animal PET studies were performed on GPC3-expressing HepG2 and non-GPC3-expressing HLF and RH7777 cells and orthotopic xenografts.

Results: (89)Zr-αGPC3 demonstrated antibody-dependent, antigen-specific tumor binding. HepG2 liver tumors exhibited high peak uptake (836.6 ± 86.6 percentage injected dose [%ID]/g) compared with background liver (27.5 ± 1.6 %ID/g). Tumor-to-liver contrast ratio was high and peaked at 32.5. The smallest HepG2 tumor (<1 mm) showed lower peak uptake (42.5 ± 6.4 %ID/g) and tumor-to-liver contrast (1.57) but was still clearly visible on PET. Day 7 tissue activity was still substantial in HepG2 tumors (466.4 ± 87.6 %ID/g) compared with control RH7777 tumors (3.9 ± 1.3 %ID/g, P < 0.01), indicating antigen specificity by (89)Zr-αGPC3. HepG2 tumor treated with unlabeled αGPC3 or heat-denatured (89)Zr-αGPC3 demonstrated tumor activity (2.1 %ID/g) comparable to that of control xenografts, confirming antibody dependency.

Conclusion: This study demonstrated the feasibility of using (89)Zr-αGPC3 to image HCC in the liver, as well as the qualitative determination of GPC3 expression via small-animal PET. The ability to clarify the identity of small liver lesions with an HCC-specific PET probe would provide clinicians with vital information that could significantly alter patient management, warranting further investigation for clinical translation.
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http://dx.doi.org/10.2967/jnumed.113.132118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4116087PMC
May 2014

CCL21 and IFNγ recruit and activate tumor specific T cells in 3D scaffold model of breast cancer.

Anticancer Agents Med Chem 2014 Feb;14(2):204-10

Department of Materials Science and Engineering, University of Washington, 302L Roberts Hall, Box 352120, Seattle, WA, 98195, USA.

Effective elicitation of endogenous immunity is associated with improved prognosis for cancer patients. Clinical evidence in hematological and solid cancers shows that intratumoral injection of immunostimulatory genes primes and augments endogenous T cell responses. The ability of pro-inflammatory chemokines/cytokines to facilitate migration/activation of antigen-presenting cells (APC) and lymphocytes prompted our modeling of intratumoral delivery of a chemokine/cytokine combination for breast cancer treatment. Here, we demonstrate that expression of chemokine ligand 21 (CCL21) and interferon gamma (IFNγ) in tumors improves tumor specific T cell recruitment to tumor and activation in the tumor milieu. IFNγ and CCL21 were delivered into tumor cells via plasmids, and transfected cells were seeded to form spheroids on three-dimensional (3D) chitosan-alginate (CA) scaffolds. Co-expression of CCL21 and IFNγ, as evidenced by qRT-PCR and ELISA, induced increased recruitment, binding, and infiltration of anti-neu (p98) peptide specific T cells into the breast tumors as determined by SEM and immunofluorescence assays. The co-expression promoted recruitment of only p98 T cells, but not naïve T cells, demonstrating an antigen-restricted activation. Furthermore, the co-expression impacted T helper (Th) cell immunity, promoting an increase in secretion of pro-inflammatory Th-associated cytokine, tumor necrosis factor alpha (TNFα), and cytotoxic T lymphocyte (CTL)-associated protease, Granzyme B (GzB). Therefore, 3D CA scaffolds may be a useful breast cancer tumor microenvironment model to evaluate T cell function. Further characterization of CCL21-IFNγ mediated anti-tumor immunity will potentially benefit the development of chemokine/cytokine combination platforms as anti-cancer agents.
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http://dx.doi.org/10.2174/18715206113136660375DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4049463PMC
February 2014

In vivo safety evaluation of polyarginine coated magnetic nanovectors.

Mol Pharm 2013 Nov 21;10(11):4099-106. Epub 2013 Oct 21.

Department of Materials Science and Engineering, University of Washington , Seattle, Washington 98195, United States.

Safety and efficacy are of critical importance to any nanomaterial-based diagnostic and therapy. The innocuity and functionality of a nanomaterial in vivo is largely dependent on the physicochemical properties of the material, particularly its surface coating. Here, we evaluated the influence of polycationic coating on the efficacy, clearance organ uptake, and safety of magnetic nanovectors designed for siRNA delivery. Polyethylene glycol (PEG) coated superparamagnetic iron oxide nanoparticles (NPs) of 12 nm in core diameter were modified with a polycationic coating of either poly-l-arginine (pArg) or polyethylenimine (PEI) and further covalently functionalized with siRNA oligonucleotides. The produced NP-pArg-siRNA and NP-PEI-siRNA nanovectors were similar in hydrodynamic size (21 and 22 nm, respectively) but significantly differed in zeta potentials (+2.1 mV and +29.8 mV, respectively). Fluorescence quantification assays revealed that the NP-pArg-siRNA nanovector was 3-fold more potent than NP-PEI-siRNA in delivering siRNA and 1.8-fold more effective in gene silencing when tested in rat C6 glioblastoma cells. In vivo, both nanovector formulations were similarly taken up by the spleen and liver as determined by histopathological and hemopathological assays. However, PEI coated nanovectors elicited severe hemoincompatibility and damage to the liver and spleen, while pArg coated nanovectors were found to be safe and tolerable. Combined, our findings suggest that polycationic coatings of pArg were more effective and safer than commonly used PEI coatings for preparation of nanovectors. The NP-pArg-siRNA nanovector formulation developed here shows great potential for in vivo based biomedical applications.
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http://dx.doi.org/10.1021/mp4005468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3946456PMC
November 2013

Porous chitosan-hyaluronic acid scaffolds as a mimic of glioblastoma microenvironment ECM.

Biomaterials 2013 Dec 26;34(38):10143-50. Epub 2013 Sep 26.

Department of Materials Science and Engineering, University of Washington, Seattle, WA, 98195, USA.

Cancer therapeutics are developed through extensive screening; however, many therapeutics evaluated with 2D in vitro cultures during pre-clinical trials suffer from lower efficacy in patients. Replicating the in vivo tumor microenvironment in vitro with three-dimensional (3D) porous scaffolds offers the possibility of generating more predictive pre-clinical models to enhance cancer treatment efficacy. We developed a chitosan and hyaluronic acid (HA) polyelectrolyte complex 3D porous scaffold and evaluated its physical properties. Chitosan-HA (C-HA) scaffolds had a highly porous network. C-HA scaffolds were compared to 2D surfaces for in vitro culture of U-118 MG human glioblastoma (GBM) cells. C-HA scaffold cultures promoted tumor spheroid formation and increased stem-like properties of GBM cells as evidenced by the upregulation of CD44, Nestin, Musashi-1, GFAP, and HIF-1α as compared with 2D cultures. Additionally, the invasiveness of GBM cells cultured in C-HA scaffolds was significantly enhanced compared to those grown in 2D cultures. C-HA scaffold cultures were also more resistant to chemotherapy drugs, which corresponded to the increased expression of ABCG2 drug efflux transporter. These findings suggest that C-HA scaffolds offer promise as an in vitro GBM platform for study and screening of novel cancer therapeutics.
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http://dx.doi.org/10.1016/j.biomaterials.2013.09.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3843957PMC
December 2013

Aligned chitosan-polycaprolactone polyblend nanofibers promote the migration of glioblastoma cells.

Adv Healthc Mater 2013 Dec 17;2(12):1651-9. Epub 2013 Jun 17.

Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195, USA; Department of Neurological Surgery, University of Washington, Seattle, WA 98195, USA.

In vitro models that accurately mimic the microenvironment of invading glioblastoma multiform (GBM) cells will provide a high-throughput system for testing potential anti-invasion therapies. Here, the ability of chitosan-polycaprolactone polyblend nanofibers to promote a migratory phenotype in human GBM cells by altering the nanotopography of the nanofiber membranes is investigated. Fibers are prepared with diameters of 200 nm, 400 nm, and 1.1 μm, and are either randomly oriented or aligned to produce six distinct nanotopographies. Human U-87 MG GBM cells, a model cell line commonly used for invasion assays, are cultured on the various nanofibrous substrates. Cells show elongation and alignment along the orientation of aligned fibers as early as 24 h and up to 120 h of culture. After 24 h of culture, human GBM cells cultured on aligned 200 nm and 400 nm fibers show marked upregulation of invasion-related genes including β-catenin, Snail, STAT3, TGF-β, and Twist, suggesting a mesenchymal change in these migrating cells. Additionally, cells cultured on 400 nm aligned fibers show similar migration profiles as those reported in vivo, and thus these nanofibers should provide a unique high-throughput in vitro culture substrate for developing anti-migration therapies for the treatment of GBM.
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http://dx.doi.org/10.1002/adhm.201300092DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3859701PMC
December 2013

Three-dimensional scaffolds to evaluate tumor associated fibroblast-mediated suppression of breast tumor specific T cells.

Biomacromolecules 2013 May 3;14(5):1330-7. Epub 2013 Apr 3.

Department of Medicine, University of Washington, Seattle, Washington 98109, United States.

In the tumor microenvironment, the signals from tumor-associated fibroblasts (TAF) that suppress antitumor immunity remain unclear. Here, we develop and investigate an in vitro three-dimensional (3D) scaffold model for the novel evaluation of TAF interaction with breast tumor cells and breast specific, neu antigen (p98) reactive T cells. Breast cancer cells seeded on 3D chitosan-alginate (CA) scaffolds showed productive growth and formed distinct tumor spheroids. Antigen specific p98 T cells, but not naïve T cells, bound significantly better to tumor cells on scaffolds. The p98 T cells induced potent tumor cell killing but T helper cell cytokine function was impaired in the presence of TAF coseeding on scaffolds. We found that the immunosuppression was mediated, in part, by transforming growth factor beta (TGF-b) and interleukin-10 (IL-10). Therefore, TAF appear capable of inducing potent T cell suppression. CA scaffolds can provide clinically relevant findings prior to preclinical testing of novel immunotherapies.
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http://dx.doi.org/10.1021/bm301928uDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3664178PMC
May 2013
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