Natalie Mendez

Encapsulation of adenovirus serotype 5 in anionic lecithin liposomes using a bead-based immunoprecipitation technique enhances transfection efficiency.

Oncolytic viruses (OVs) constitute a promising class of cancer therapeutics which exploit validated genetic pathways known to be deregulated in many cancers. To overcome an immune response and to enhance its potential use to treat primary and metastatic tumors, a method for liposomal encapsulation of adenovirus has been developed. The encapsulation of adenovirus in non-toxic anionic lecithin-cholesterol-PEG liposomes ranging from 140 to 180 nm in diameter have been prepared by self-assembly around the viral capsid. The encapsulated viruses retain their ability to infect cancer cells. Furthermore, an immunoprecipitation (IP) technique has shown to be a fast and effective method to extract non-encapsulated viruses and homogenize the liposomes remaining in solution. 78% of adenovirus plaque forming units were encapsulated and retained infectivity after IP processing. Additionally, encapsulated viruses have shown enhanced transfection efficiency up to 4 × higher compared to non-encapsulated Ads. Extracting non-encapsulated viruses from solution may prevent an adverse in vivo immune response and may enhance treatment for multiple administrations.

Assessment of in vivo systemic toxicity and biodistribution of iron-doped silica nanoshells

Silica nanoparticles are an emerging class of biomaterials which may be used as diagnostic and therapeutic tools for biomedical applications. In particular, hollow silica nanoshells are attractive due to their hollow core. Approximately 70% of a 500 nm nanoshell is hollow, therefore more particles can be administered on a mg/kg basis compared to solid nanoparticles. Additionally, their nanoporous shell permits influx/efflux of gases and small molecules. Since the size, shape, and composition of a nanoparticle can dramatically alter its toxicity and biodistribution, the toxicology of these nanomaterials was assessed. A single dose toxicity study was performed in vivo to assess the toxicity of 500 nm iron-doped silica nanoshells at clinically relevant doses of 10-20 mg/kg. This study showed that only a trace amount of silica was detected in the body 10 weeks post-administration. The hematology, biochemistry and pathological results show that the nanoshells exhibit no acute or chronic toxicity in mice.

Ultrasound Responsive Macrophase-Segregated Microcomposite Films for in Vivo Biosensing

Ultrasound imaging is a safe, low-cost, and in situ method for detecting in vivo medical devices. A poly(methyl-2-cyanoacrylate) film containing 2 μm boron-doped, calcined, porous silica microshells was developed as an ultrasound imaging marker for multiple medical devices. A macrophase separation drove the gas-filled porous silica microshells to the top surface of the polymer film by controlled curing of the cyanoacrylate glue and the amount of microshell loading. A thin film of polymer blocked the wall pores of the microshells to seal air in their hollow core, which served as an ultrasound contrast agent. The ultrasound activity disappeared when curing conditions were modified to prevent the macrophase segregation. Phase segregated films were attached to multiple surgical tools and needles and gave strong color Doppler signals in vitro and in vivo with the use of a clinical ultrasound imaging instrument. Postprocessing of the simultaneous color Doppler and B-mode images can be used for autonomous identification of implanted surgical items by correlating the two images. The thin films were also hydrophobic, thereby extending the lifetime of ultrasound signals to hours of imaging in tissues by preventing liquid penetration. This technology can be used as a coating to guide the placement of implantable medical devices or used to image and help remove retained surgical items.

Quantification of endocytosis using a folate functionalized silica hollow nanoshell platform

Abstract. A quantification method to measure endocytosis was designed to assess cellular uptake and specificity of a targeting nanoparticle platform. A simple N-hydroxysuccinimide ester conjugation technique to functionalize 100-nm hollow silica nanoshell particles with fluorescent reporter fluorescein isothiocyanate and folate or polyethylene glycol (PEG) was developed. Functionalized nanoshells were characterized using scanning electron microscopy and transmission electron microscopy and the maximum amount of folate functionalized on nanoshell surfaces was quantified with UV-Vis spectroscopy. The extent of endocytosis by HeLa cervical cancer cells and human foreskin fibroblast (HFF-1) cells was investigated in vitro using fluorescence and confocal microscopy. A simple fluorescence ratio analysis was developed to quantify endocytosis versus surface adhesion. Nanoshells functionalized with folate showed enhanced endocytosis by cancer cells when compared to PEG functionalized nanoshells. Fluorescence ratio analyses showed that 95% of folate functionalized silica nanoshells which adhered to cancer cells were endocytosed, while only 27% of PEG functionalized nanoshells adhered to the cell surface and underwent endocytosis when functionalized with 200 and 900  μg, respectively. Additionally, the endocytosis of folate functionalized nanoshells proved to be cancer cell selective while sparing normal cells. The developed fluorescence ratio analysis is a simple and rapid verification/validation method to quantify cellular uptake between datasets by using an internal control for normalization.

Targeting Tumors Using Nanoparticle Platforms: A Phase I Study of a Systemically Administered Gene Therapy System

The potential for nonviral gene therapy to treat cancer has been limited by the difficulty of targeting therapeutic genes to tumors. In this issue of Molecular Therapy, Senzer et al.1 show evidence of specific tumor targeting as well as clinical efficacy in a phase I trial of a nonviral vector. This is the first phase I clinical trial to demonstrate tumor specificity and uptake by metastatic tumors of a systemically delivered liposomal nanoparticle. Importantly, the therapeutic nanoparticle elicited minimal side effects and the majority of patients demonstrated stable disease. This phase I trial has shown this delivery system to be a safe, stable, efficient, and tumor-specific drug delivery platform for systemic administration.

Identifying lost surgical needles with visible and near infrared fluorescent light emitting microscale coating

Background. Retained foreign bodies (RFOs) have substantial clinical and financial consequences. In laparoscopic surgery, RFOs can be a cause of needing to convert a minimally invasive surgery (MIS) procedure to an open operation. A coating for surgical models was developed to augment localization of needles using fluorescence appropriate for open and minimally invasive surgeries procedures. Methods. An epoxy matrix containing both dansyl chloride and indocyanine green was coated as visible and near infrared labels, respectively. With ultraviolet excitation, dansyl chloride emits green fluorescence and with NIR excitation, the ICG dye emits radiation observable with specialized near infrared capable laparoscopes. To evaluate the coatings, open and laproscopic surgeries were simulated in rabbits. Surgeons blinded to the type of needles (coated or non‐coated) were timed while finding needles in standard conditions and with the use of the adjunct coatings. Control needles not located within 300 seconds were researched with the corresponding near infrared or ultraviolet light. Localization time was evaluated for statistical significance, P < .05. Results. All dual dye coated needles searched utilizing the near infrared camera (n = 26) or ultraviolet light (n = 26) were located within 300 seconds. Conversely, 9 needles in both control settings (no dye usage) were not located within 300 seconds. Mean time to locate control needles in open surgery and laparoscopic surgery was statistically 2–3× greater than time to localization with the use of dye as an adjunct (P = .0027 open, P < .001 laparoscopic). Conclusion. Incorporation of a dual‐dye fluorescent coating on surgical needles improved the efficiency of locating needles, may minimize the need to convert minimally invasive surgeries procedures to open, and may decrease the consequences of a missed RFO.