Justin J. Law

Stable confinement of positron emission tomography and magnetic resonance agents within carbon nanotubes for bimodal imaging

AIMSnSimultaneous positron emission tomography/MRI has recently been introduced to the clinic and dual positron emission tomography/MRI probes are rare and of growing interest. We have developed a strategy for producing multimodal probes based on a carbon nanotube platform without the use of chelating ligands.nnnMATERIALS & METHODSnGd(3+) and (64)Cu(2+) ions were loaded into ultra-short single-walled carbon nanotubes by sonication. Normal, tumor-free athymic nude mice were injected intravenously with the probe and imaged over 48 h.nnnRESULTS & CONCLUSIONnThe probe was stable for up to 24 h when challenged with phosphate-buffered saline and mouse serum. Positron emission tomography imaging also confirmed the stability of the probe in vivo for up to 48 h. The probe was quickly cleared from circulation, with enhanced accumulation in the lungs. Stable encapsulation of contrast agents within ultra-short single-walled carbon nanotubes represents a new strategy for the design of advanced imaging probes with variable multimodal imaging capabilities.

Subcellular Partitioning and Analysis of Gd3+-Loaded Ultrashort Single-Walled Carbon Nanotubes.

Magnetic resonance imaging (MRI) is of vast clinical utility, with tens of millions of scans performed annually. Chemical contrast agents (CAs) can greatly enhance the diagnostic potential of MRI, and ∼50% of MRI scans use CAs. However, CAs have significant limitations such as low contrast enhancement, lack of specificity, and potential toxicity. Recently developed, Gd3+-loaded ultrashort single-walled carbon nanotubes, also referred to as gadonanotubes or GNTs, exhibit ∼40 times the relaxivities of clinical CAs, representing a potential major advance in clinically relevant MRI CA materials. Although initial cytotoxicity and MRI studies have suggested great promise for GNTs, relatively little is known regarding their subcellular interactions, which are crucial for further, safe development of GNTs as CAs. In this work, we administered GNTs to a well-established human cell line (HeLa) and to murine macrophage-like cells (J774A.1). GNTs were not acutely cytotoxic and did not reduce proliferation, except for the highest exposure concentration of 27 μg/mL for J774A.1 macrophages, yet bulk uptake of GNTs occurred in minutes at picogram quantities, or millions of GNTs per cell. J774A.1 macrophages internalized substantially more GNTs than HeLa cells in a dose-dependent manner, and Raman imaging of the subcellular distribution of GNTs revealed perinuclear localization. Fluorescence intensity and lifetime imaging demonstrated that GNTs did not grossly alter subcellular compartments, including filamentous-actin structures. Together, these results provide subcellular evidence necessary to establish GNTs as a new MRI CA material.

Relaxivity enhancement of aquated Tris(β-diketonate)gadolinium(III) chelates by confinement within ultrashort single-walled carbon nanotubes.

Ultrashort single-walled carbon nanotubes loaded with gadolinium ions (gadonanotubes) have been previously shown to exhibit extremely high T1 -weighted relaxivities (>100 mm(-1) s(-1) ). To further examine the effect of nanoconfinement on the relaxivity of gadolinium-based contrast agents for magnetic resonance imaging, a series of ultrashort single-walled carbon nanotube (US-tube) materials internally loaded with gadolinium chelates have been prepared and studied. US-tubes were loaded with Gd(acac)3 u2009·u20092H2 O, Gd(hfac)3 u2009·u20092H2 O, and Gd(thd)3 (acacu2009=u2009acetylacetone, hfacu2009=u2009hexafluoroacetylacetone, thdu2009=u2009tetramethylheptanedione). The longitudinal relaxivities of the prepared materials determined at 25°C in a 1.5 T field were 103 mm(-1) s(-1) for Gd(acac)3 u2009·u20092H2 O@US-tubes, 105 mm(-1) s(-1) for Gd(hfac)3 u2009·u20092H2 O@US-tubes and 26 mm(-1) s(-1) for Gd(thd)3 @US-tubes. Compared with the relaxivities obtained for the unloaded chelates (<10 mm(-1) s(-1) ) as well as accounting for the T1 reduction observed for the empty US-tubes, the boost in relaxivity for chelate-loaded US-tubes is attributed to confinement within the nanotube and depends on the number of coordinated water molecules.

Radiofrequency electric-field heating behaviors of highly enriched semiconducting and metallic single-walled carbon nanotubes

It is theorized that enhanced thermal heating may result from exposing single-walled carbon nanotubes (SWNTs) embedded in a conductive host to radiofrequency (RF) electric fields. We examine the RF-induced (13.56 MHz) heating behaviors of 95% metallic- and semiconducting-enriched SWNTs (m-/s-SWNTs) suspended in aqueous solutions with varying NaCl molarity (0.001 mM–1 M). The heating effects were only evident for host molarities below 1 mM (equivalent to 0.1 S/m) at which the s-SWNT heating rates dominated those of the m-SWNTs. The heating effects were localized to aligned and aggregated “SWNT ropes” ~1 cm in length that formed in suspension, parallel to the electric-field vector, during the RF exposure. For molarities above 1 mM, no enhancements were evident, owing to the large heating effects of the bulk ionic NaCl suspensions, which were observed in previous studies. Although larger enhancement effects proportional to the host conductivity have been theoretically predicted for m-/s-SWNT suspensions, this was not observed most likely because of the aggregation and screening effects, which diminished the scattered electric field near the m-/s-SWNTs. Our research may further the development of better nanoparticle heating agents for applications such as non-invasive RF-induced cancer hyperthermia.

Encapsulation of Alpha-particle Emitting Actinium-225 Ions within Carbon Nanotubes

225Ac3+ is a generator of α-particle–emitting radionuclides with 4 net α-particle decays that can be used therapeutically. Targeting 225Ac3+ by use of ligands conjugated to traditional bifunctional chelates limits the amount of 225Ac3+ that can be delivered. Ultrashort, single-walled carbon nanotubes (US-tubes), previously demonstrated as sequestering agents of trivalent lanthanide ions and small molecules, also successfully incorporate 225Ac3+. Methods: Aqueous loading of both 225Ac3+ ions and Gd3+ ions via bath sonication was used to construct 225Ac@gadonanotubes (225Ac@GNTs). The 225Ac@GNTs were subsequently challenged with heat, time, and human serum. Results: US-tubes internally loaded with both 225Ac3+ ions and Gd3+ ions show 2 distinct populations of 225Ac3+ ions: one rapidly lost in human serum and one that remains bound to the US-tubes despite additional challenge with heat, time, and serum. The presence of the latter population depended on cosequestration of Gd3+ and 225Ac3+ ions. Conclusion: US-tubes successfully sequester 225Ac3+ ions in the presence of Gd3+ ions and retain them after a human serum challenge, rendering 225Ac@GNTs candidates for radioimmunotherapy for delivery of 225Ac3+ ions at higher concentrations than is currently possible for traditional ligand carriers.

Encapsulation of α-Particle–Emitting 225Ac3+ Ions Within Carbon Nanotubes

225Ac3+ is a generator of α-particle–emitting radionuclides with 4 net α-particle decays that can be used therapeutically. Targeting 225Ac3+ by use of ligands conjugated to traditional bifunctional chelates limits the amount of 225Ac3+ that can be delivered. Ultrashort, single-walled carbon nanotubes (US-tubes), previously demonstrated as sequestering agents of trivalent lanthanide ions and small molecules, also successfully incorporate 225Ac3+. Methods: Aqueous loading of both 225Ac3+ ions and Gd3+ ions via bath sonication was used to construct 225Ac@gadonanotubes (225Ac@GNTs). The 225Ac@GNTs were subsequently challenged with heat, time, and human serum. Results: US-tubes internally loaded with both 225Ac3+ ions and Gd3+ ions show 2 distinct populations of 225Ac3+ ions: one rapidly lost in human serum and one that remains bound to the US-tubes despite additional challenge with heat, time, and serum. The presence of the latter population depended on cosequestration of Gd3+ and 225Ac3+ ions. Conclusion: US-tubes successfully sequester 225Ac3+ ions in the presence of Gd3+ ions and retain them after a human serum challenge, rendering 225Ac@GNTs candidates for radioimmunotherapy for delivery of 225Ac3+ ions at higher concentrations than is currently possible for traditional ligand carriers.

Optimizing non-invasive radiofrequency hyperthermia treatment for improving drug delivery in 4T1 mouse breast cancer model

Interactions of high-frequency radio waves (RF) with biological tissues are currently being investigated as a therapeutic platform for non-invasive cancer hyperthermia therapy. RF delivers thermal energy into tissues, which increases intra-tumoral drug perfusion and blood-flow. Herein, we describe an optical-based method to optimize the short-term treatment schedules of drug and hyperthermia administration in a 4T1 breast cancer model via RF, with the aim of maximizing drug localization and homogenous distribution within the tumor microenvironment. This method, based on the analysis of fluorescent dyes localized into the tumor, is more time, cost and resource efficient, when compared to current analytical methods for tumor-targeting drug analysis such as HPLC and LC-MS. Alexa-Albumin 647u2009nm fluorphore was chosen as a surrogate for nab-paclitaxel based on its similar molecular weight and albumin driven pharmacokinetics. We found that RF hyperthermia induced a 30–40% increase in Alexa-Albumin into the tumor micro-environment 24u2009h after treatment when compared to non-heat treated mice. Additionally, we showed that the RF method of delivering hyperthermia to tumors was more localized and uniform across the tumor mass when compared to other methods of heating. Lastly, we provided insight into some of the factors that influence the delivery of RF hyperthermia to tumors.

Non-invasive radiofrequency field treatment of 4T1 breast tumors induces T-cell dependent inflammatory response

Previous work using non-invasive radiofrequency field treatment (RFT) in cancer has demonstrated its therapeutic potential as it can increase intratumoral blood perfusion, localization of intravenously delivered drugs, and promote a hyperthermic intratumoral state. Despite the well-known immunologic benefits that febrile hyperthermia can induce, an investigation of how RFT could modulate the intra-tumoral immune microenvironment had not beenxa0studied. Thus, using an established 4T1 breast cancer model in immune competent mice, we demonstrate that RFT induces a transient, localized, and T-cell dependent intratumoral inflammatory response. More specifically we show that multi- and singlet-dose RFT promote an increase in tumor volume in immune competent Balb/c mice, which does not occur in athymic nude models. Further leukocyte subset analysis at 24, 48, and 120u2009hours after a single RFT show a rapid increase in tumoral trafficking of CD4+ and CD8+ T-cells 24u2009hours post-treatment. Additional serum cytokine analysis reveals an increase in numerous pro-inflammatory cytokines and chemokines associated with enhanced T-cell trafficking. Overall, these data demonstrate that non-invasive RFT could be an effective immunomodulatory strategy in solid tumors, especially for enhancing the tumoral trafficking of lymphocytes, which is currently a major hindrance of numerous cancer immunotherapeutic strategies.

Improved, Shorter-Latency Carcinogen-Induced Hepatocellular Carcinoma Model in Pigs

Large animal models are important tools for hepatocellular carcinoma (HCC) research, especially in studies of hepatic vasculature, interventional techniques, and radiofrequency or microwave hyperthermia. Currently, diethylnitrosamine (DENA)-induced HCC in pigs is the only large animal model for in situ HCC with a tumor latency of 10–26 months. While phenobarbital (PB) is often used to accelerate DENA-induced HCC in rodents, it has not been previously studied in the porcine model. Therefore, we hypothesize that the addition of PB in the DENA-induced HCC porcine model will accelerate tumor latency compared to DENA alone. HCC and benign lesions were seen on serial MRI and confirmed on histopathology. Liver and tumors were further characterized by CT angiography, vascular corrosion casting, and permittivity measurements.