Sarah P. Sherlock

Drug delivery with carbon nanotubes for in vivo cancer treatment.

Chemically functionalized single-walled carbon nanotubes (SWNT) have shown promise in tumor-targeted accumulation in mice and exhibit biocompatibility, excretion, and little toxicity. Here, we show in vivo SWNT drug delivery for tumor suppression in mice. We conjugate paclitaxel (PTX), a widely used cancer chemotherapy drug, to branched polyethylene glycol chains on SWNTs via a cleavable ester bond to obtain a water-soluble SWNT-PTX conjugate. SWNT-PTX affords higher efficacy in suppressing tumor growth than clinical Taxol in a murine 4T1 breast cancer model, owing to prolonged blood circulation and 10-fold higher tumor PTX uptake by SWNT delivery likely through enhanced permeability and retention. Drug molecules carried into the reticuloendothelial system are released from SWNTs and excreted via biliary pathway without causing obvious toxic effects to normal organs. Thus, nanotube drug delivery is promising for high treatment efficacy and minimum side effects for future cancer therapy with low drug doses.

Deep-tissue anatomical imaging of mice using carbon nanotube fluorophores in the second near-infrared window

Fluorescent imaging in the second near-infrared window (NIR II, 1–1.4 μm) holds much promise due to minimal autofluorescence and tissue scattering. Here, using well-functionalized biocompatible single-walled carbon nanotubes (SWNTs) as NIR II fluorescent imaging agents, we performed high-frame-rate video imaging of mice during intravenous injection of SWNTs and investigated the path of SWNTs through the mouse anatomy. We observed in real-time SWNT circulation through the lungs and kidneys several seconds postinjection, and spleen and liver at slightly later time points. Dynamic contrast-enhanced imaging through principal component analysis (PCA) was performed and found to greatly increase the anatomical resolution of organs as a function of time postinjection. Importantly, PCA was able to discriminate organs such as the pancreas, which could not be resolved from real-time raw images. Tissue phantom studies were performed to compare imaging in the NIR II region to the traditional NIR I biological transparency window (700–900 nm). Examination of the feature sizes of a common NIR I dye (indocyanine green) showed a more rapid loss of feature contrast and integrity with increasing feature depth as compared to SWNTs in the NIR II region. The effects of increased scattering in the NIR I versus NIR II region were confirmed by Monte Carlo simulation. In vivo fluorescence imaging in the NIR II region combined with PCA analysis may represent a powerful approach to high-resolution optical imaging through deep tissues, useful for a wide range of applications from biomedical research to disease diagnostics.

Supramolecular Stacking of Doxorubicin on Carbon Nanotubes for In Vivo Cancer Therapy

Doxorubicin (DOX) is a member of the anthracycline class of chemotherapeutic agents that are used for the treatment of many common human cancers, including aggressive non-Hodgkin’s lymphoma.[1,2] However, DOX is highly toxic in humans and can result in severe suppression of hematopoiesis, gastrointestinal toxicity,[3] and cardiac toxicity.[4] To date, several approaches, including delivery using liposomes (DOXIL),[5] have been developed to reduce the toxicity and enhance the clinical utility of this highly active antineoplastic agent.

High performance in vivo near-IR (>1 μm) imaging and photothermal cancer therapy with carbon nanotubes

Short single-walled carbon nanotubes (SWNTs) functionalized by PEGylated phospholipids are biologically non-toxic and long-circulating nanomaterials with intrinsic near infrared photoluminescence (NIR PL), characteristic Raman spectra, and strong optical absorbance in the near infrared (NIR). This work demonstrates the first dual application of intravenously injected SWNTs as photoluminescent agents for in vivo tumor imaging in the 1.0–1.4 μm emission region and as NIR absorbers and heaters at 808 nm for photothermal tumor elimination at the lowest injected dose (70 μg of SWNT/mouse, equivalent to 3.6 mg/kg) and laser irradiation power (0.6 W/cm2) reported to date. Ex vivo resonance Raman imaging revealed the SWNT distribution within tumors at a high spatial resolution. Complete tumor elimination was achieved for large numbers of photothermally treated mice without any toxic side effects after more than six months post-treatment. Further, side-by-side experiments were carried out to compare the performance of SWNTs and gold nanorods (AuNRs) at an injected dose of 700 μg of AuNR/mouse (equivalent to 35 mg/kg) in NIR photothermal ablation of tumors in vivo. Highly effective tumor elimination with SWNTs was achieved at 10 times lower injected doses and lower irradiation powers than for AuNRs. These results suggest there are significant benefits of utilizing the intrinsic properties of biocompatible SWNTs for combined cancer imaging and therapy.

Photothermally enhanced drug delivery by ultrasmall multifunctional FeCo/graphitic shell nanocrystals.

FeCo/graphitic carbon shell (FeCo/GC) nanocrystals (∼4-5 nm in diameter) with ultrahigh magnetization are synthesized, functionalized, and developed into multifunctional biocompatible materials. We demonstrate the ability of this material to serve as an integrated system for combined drug delivery, near-infrared (NIR) photothermal therapy, and magnetic resonance imaging (MRI) in vitro. We show highly efficient loading of doxorubicin (DOX) by π-stacking on the graphitic shell to afford FeCo/GC-DOX complexes and pH sensitive DOX release from the particles. We observe enhanced intracellular drug delivery by FeCo/GC-DOX under 20 min of NIR laser (808 nm) induced hyperthermia to 43 °C, resulting in a significant increase of FeCo/GC-DOX toxicity toward breast cancer cells. The synergistic cancer cell killing by FeCo/GC-DOX drug delivery under photothermal heating is due to a ∼two-fold enhancement of cancer cell uptake of FeCo/GC-DOX complex and the increased DOX toxicity under the 43 °C hyperthermic condition. The combination of synergistic NIR photothermally enhanced drug delivery and MRI with the FeCo/GC nanocrystals could lead to a powerful multimodal system for biomedical detection and therapy.

Separation of nanoparticles in a density gradient: FeCo@C and gold nanocrystals.

Size and geometric control of nanomaterials are important to the discovery of intrinsic size/shape dependent properties and bottom up approaches for the fabrication of functional nanodevices. Two general strategies have been employed to create size-uniform nanocrystals. One method is direct particle size control during synthesis by adjusting growth parameters; 7–9] the other is post-synthesis separation. Much capacity exists to improve size separation efficacy in the latter case. Differential centrifugation can remove large and unstable particles from colloidal systems, but lacks precise control over particle size. Addition of adjustable amounts of “anti-solvent” (including CO2) [12] into colloidal systems may make precipitation processes more controllable. Other methods include filtration (including diafiltration), electrophoresis, and chromatographic methods that can produce particle fractions with narrow shape and size distributions. To maintain or improve the quality of nanoparticle (NP) separation, whilst addressing the issues of adhesion and clogging in liquid–solid phase separation processes, a completely liquid phase separation method is highly appealing. Isopycnic centrifugation, which is often used for biomacromolecule separation, relies upon a density gradient and ultracentrifugation to separate components according to subtle density differences, and has been applied for diameter and electronic-dependent separation of single-walled carbon nanotubes (SWNT). However, the isopycnic densitygradient centrifugation method reaches a limitation when it is extended to the separation of metal nanoparticles. Such a method requires that the components for separation have densities within a gradient range. Aqueous density gradient media usually have densities less than 1.4 gcm , which is much less than the density of metal nanoparticles. Size or shape separation of such heavy nanocrystals remains an issue, both in their preparation and utility for various applications. In contrast to isopycnic separation, ultracentrifugal rate separation can utilize density gradients to separate nanocrystals with higher densities than the gradient media itself. We have previously applied such a method to achieve length separation of suspended SWNTs and pegylated graphene oxide. In this report, the method was extended to metallic NP size separation. Nanoparticles of various size, suspension chemistry, and composition, including FeCo@C and gold nanoparticles (Au NPs), were separated using the method. FeCo nanocrystals coated in graphitic shells have superior magnetic properties, and have shown promise for applications in biolabeling and magnetic resonance imaging (MRI). However, the chemical deposition method used for their preparation produces nanocrystals with wide size distribution. They are thus ideal candidates for post-synthesis separation. FeCo@C NPs with average diameters of about 4 nm were separated first by our density gradient rate (DGR) separation method by using a 10+ 20+ 30+ 40% gradient and centrifugation for 3.5 h. TEM results of typical fractions (Figure 1A) indicate that fraction 8 (labeled as “f8” in Figure 1B) contained circa 1.5 nm NPs. The average particle diameter of subsequent fractions (f11, 15, 19, 24, and 27) gradually increased from 2.5 to 5.6 nm. By varying the step gradient densities and centrifuge exposure time, this method could be used for separation of nanoparticles of a larger size range, which was demonstrated by (on average) 7 nm FeCo@C NP separation (see TEM images of initial 7 nm FeCo@C NPs in the Supporting Information). A gradient of higher density steps (20+ 30+ 40+ 60%) was used. The use of higher density gradient media helps to control the sedimentation rate by reducing the density difference between the NPs and the environmental medium, and increasing the medium viscosity. After centrifugation for 2.5 h, several bands formed in the centrifuge vessel (Figure 2A), just as in the 4 nm NP case. Sampling fractions along the centrifuge vessel yielded nanoparticles of increasing size (with increasing density), as revealed by TEM (Figure 2B). From f5 to f16, the average particle size increased from 2 to 6.5 nm. It is noteworthy that the Fe/Co atomic ratios of f8 and f16 were measured by energy dispersive spectra (EDS) and found to be different (Figure 2B, bottom right). For f8, the ratio Fe/Co= 48:52; for f16, Fe/Co= 40:60. Previously such stoichiometry was analyzed by calcination/burning of the graphitic shells at 500 8C, dissolving the metal species in an HCl solution, and measuring the iron and cobalt concentrations on the basis of the ultraviolet[*] Prof. Dr. X. M. Sun, L. Bai State Key Laboratory of Chemical Resource Engineering Beijing University of Chemical Technology Beijing 100029 (China)

Multiplexed five-color molecular imaging of cancer cells and tumor tissues with carbon nanotube Raman tags in the near-infrared

AbstractSingle-walled carbon nanotubes (SWNTs) with five different C13/C12 isotope compositions and well-separated Raman peaks have been synthesized and conjugated to five targeting ligands in order to impart molecular specificity. Multiplexed Raman imaging of live cells has been carried out by highly specific staining of cells with a five-color mixture of SWNTs. Ex vivo multiplexed Raman imaging of tumor samples uncovers a surprising up-regulation of epidermal growth factor receptor (EGFR) on LS174T colon cancer cells from cell culture to in vivo tumor growth. This is the first time five-color multiplexed molecular imaging has been performed in the near-infrared (NIR) region under a single laser excitation. Near zero interfering background of imaging is achieved due to the sharp Raman peaks unique to nanotubes over the low, smooth autofluorescence background of biological species.

Phospholipid-Dextran with a Single Coupling Point: a Useful Amphiphile for Functionalization of Nanomaterials

Nanomaterials hold much promise for biological applications, but they require appropriate functionalization to provide biocompatibility in biological environments. For noncovalent functionalization with biocompatible polymers, the polymer must also remain attached to the nanomaterial after removal of its excess to mimic the high-dilution conditions of administration in vivo. Reported here are the synthesis and utilization of singly substituted conjugates of dextran and a phospholipid (dextran-DSPE) as stable coatings for nanomaterials. Suspensions of single-walled carbon nanotubes were found not only to be stable to phosphate buffered saline (PBS), serum, and a variety of pHs after excess polymer removal, but also to provide brighter photoluminescence than carbon nanotubes suspended by poly(ethylene glycol)-DSPE. In addition, both gold nanoparticles (AuNPs) and gold nanorods (AuNRs) were found to maintain their dispersion and characteristic optical absorbance after transfer into dextran-DSPE and were obtained in much better yield than similar suspensions with PEG-phospholipid and commonly used thiol-PEG. These suspensions were also stable to PBS, serum, and a variety of pHs after removal of excess polymer. dextran-DSPE thus shows great promise as a general surfactant material for the functionalization of a variety of nanomaterials, which could facilitate future biological applications.

Near Infrared Imaging and Photothermal Ablation of Vascular Inflammation Using Single-Walled Carbon Nanotubes

Background Macrophages are critical contributors to atherosclerosis. Single-walled carbon nanotubes (SWNTs) show promising properties for cellular imaging and thermal therapy, which may have application to vascular macrophages. Methods and Results In vitro uptake and photothermal destruction of mouse macrophage cells (RAW264.7) were performed with SWNTs (14.7 nmol/L) exposed to an 808-nm light source. SWNTs were taken up by 94±6% of macrophages, and light exposure induced 93±3% cell death. In vivo vascular macrophage uptake and ablation were then investigated in carotid-ligated FVB mice (n=33) after induction of hyperlipidemia and diabetes. Two weeks postligation, near-infrared fluorescence (NIRF) carotid imaging (n=12) was performed with SWNT-Cy5.5 (8 nmol of Cy5.5) given via the tail vein. Photothermal heating and macrophage apoptosis were evaluated on freshly excised carotid arteries (n=21). NIRF of SWNTs showed higher signal intensity in ligated carotids compared with sham, confirmed by both in situ and ex vivo NIRF imaging (P<0.05, ligation versus sham). Immunofluorescence staining showed colocalization of SWNT-Cy5.5 and macrophages in atherosclerotic lesions. Light (808 nm) exposure of freshly excised carotids showed heating and induction of macrophage apoptosis in ligated left carotid arteries with SWNTs, but not in control groups without SWNTs or without light exposure. Conclusions Carbon nanotubes accumulate in atherosclerotic macrophages in vivo and provide a multifunctional platform for imaging and photothermal therapy of vascular inflammation.

Graphite-Coated Magnetic Nanoparticle Microarray for Few-Cells Enrichment and Detection

Graphite-coated, highly magnetic FeCo core-shell nanoparticles were synthesized by a chemical vapor deposition method and solubilized in aqueous solution through a unique polymer mixture modification, which significantly improved the biocompatibility and stability of the magnetic nanoparticles (MNPs). Such functionalized MNPs were proven to be very stable in different conditions which would be significant for biological applications. Cell staining, manipulation, enrichment, and detection were developed with these MNPs. Under external magnetic manipulation, the MNP-stained cells exhibited directed motions. Moreover, MNPs were printed on substrates to modulate the magnetic field distribution on the surface. Capture and detection of sparse populations of cancer cells spiked into whole blood has been explored in a microarray fashion. Cancer cells from hundreds down to only two were able to be simply and efficiently detected from 1 mL of whole blood on the MNP microarray chips. Interestingly, the cells captured through the MNP microarray still showed viability and adhered to the MNP spots after incubation, which could be utilized for cancer cell detection, localized growth, and proliferation.