Jessica Webber

Photothermal nanodrugs: potential of TNF-gold nanospheres for cancer theranostics

Nanotechnology has been extensively explored for drug delivery. Here, we introduce the concept of a nanodrug based on synergy of photothermally-activated physical and biological effects in nanoparticle-drug conjugates. To prove this concept, we utilized tumor necrosis factor-alpha coated gold nanospheres (Au-TNF) heated by laser pulses. To enhance photothermal efficiency in near-infrared window of tissue transparency we explored slightly ellipsoidal nanoparticles, its clustering, and laser-induced nonlinear dynamic phenomena leading to amplification and spectral sharpening of photothermal and photoacoustic resonances red-shifted relatively to linear plasmonic resonances. Using a murine carcinoma model, we demonstrated higher therapy efficacy of Au-TNF conjugates compared to laser and Au-TNF alone or laser with TNF-free gold nanospheres. The photothermal activation of low toxicity Au-TNF conjugates, which are in phase II trials in humans, with a laser approved for medical applications opens new avenues in the development of clinically relevant nanodrugs with synergistic antitumor theranostic action.

Indocyanine green enhanced near-infrared laser treatment of murine mammary carcinoma.

It is well accepted that near‐infrared (NIR) lasers are appropriate to ablate benign lesions and induce irreversible thermal injury in deeply seated blood vessels. At this wavelength, the laser light penetrates deep (3–5 mm) into the skin. However, many researchers have reported noticeable pain, extending from mild to severe, during and immediately after NIR laser treatment. Intravenous administration of an exogenous chromophore [indocyanine green (ICG), dye] can effectively convert NIR laser light into heat. In this approach, the presence of ICG has shown to enhance thermal injury of blood vessels in the treatment of healthy tissues. However, the effectiveness of thermal injury on the regression of cutaneous carcinomas during ICG/NIR laser therapy has not been assessed. The purpose of our study was to evaluate the potential benefit of using ICG/NIR laser therapy to regress superficial carcinoma with thermal injury. Two groups of A/J mice with subcutaneous mammary adenocarcinoma tumors (7–9 mm) were irradiated with a 808‐nm NIR laser preceded by tail vein injection of ICG dye or sterile saline. Histological evaluation of the subcutaneous tissue revealed minor thermal damage and necrosis in the laser/saline group and substantial damage (up to 100% necrosis) in the laser/ICG group. The laser/ICG‐treated group showed a steady reduction in tumor volume compared to the laser/saline group: 48% by day 5 (p = 0.045) and 69–70% by days 8, 9 and 10 (p values 0.0005 or less). The vascular‐targeted ICG–NIR laser therapy appears to have potential for treating superficial tumors.

Hyperthermia-enhanced indocyanine green delivery for laser-induced thermal ablation of carcinomas

Abstract Purpose: Intravenous administration of indocyanine green (ICG) dye can effectively convert near-infrared (NIR) laser light into heat and enhance thermal injury of blood vessels; however, there is no selective uptake of ICG by the tumour compared to the other tissues, which impacts the therapeutic ratio of this strategy unless uptake can be selectively increased in tumour tissue. Here we investigated the use of local hyperthermia prior to intravenous ICG administration to enhance ICG uptake in tumour tissue, thereby enhancing laser thermal ablation of solid tumours. Methods: Murine SCK breast or SCCVII head and neck tumours were treated with a 755-nm laser light either alone or with prior intravenous administration of 4 mg/kg ICG and/or local tumour hyperthermia at 42.5 °C for 60 min. Retention of ICG was quantified using a NIR animal imaging system. Treatment effects were assessed by growth delay and histology. Results: ICG accumulation in the heated tumours was 1.23-fold greater on average compared to non-heated tumours, in both models. In SCK tumours, animals receiving either laser irradiation alone or in conjunction with ICG had a 1.86- or 3.91-fold increase in tumour growth delay, respectively. The addition of local hyperthermia before ICG injection resulted in complete regression of SCK tumours. Uptake of ICG increased in SCCVII tumours; however, little change in tumour growth delay was observed. Conclusion: Using local hyperthermia may improve the delivery of ICG to the tumour and thereby increase the extent of laser thermal ablation of smaller superficial malignancies that can be effectively exposed to laser therapy.

Advanced Small Animal Conformal Radiation Therapy Device

We have developed a small animal conformal radiation therapy device that provides a degree of geometrical/anatomical targeting comparable to what is achievable in a commercial animal irradiator. small animal conformal radiation therapy device is capable of producing precise and accurate conformal delivery of radiation to target as well as for imaging small animals. The small animal conformal radiation therapy device uses an X-ray tube, a robotic animal position system, and a digital imager. The system is in a steel enclosure with adequate lead shielding following National Council on Radiation Protection and Measurements 49 guidelines and verified with Geiger-Mueller survey meter. The X-ray source is calibrated following AAPM TG-61 specifications and mounted at 101.6 cm from the floor, which is a primary barrier. The X-ray tube is mounted on a custom-made “gantry” and has a special collimating assembly system that allows field size between 0.5 mm and 20 cm at isocenter. Three-dimensional imaging can be performed to aid target localization using the same X-ray source at custom settings and an in-house reconstruction software. The small animal conformal radiation therapy device thus provides an excellent integrated system to promote translational research in radiation oncology in an academic laboratory. The purpose of this article is to review shielding and dosimetric measurement and highlight a few successful studies that have been performed to date with our system. In addition, an example of new data from an in vivo rat model of breast cancer is presented in which spatially fractionated radiation alone and in combination with thermal ablation was applied and the therapeutic benefit examined.

An alternating focused ultrasound system for thermal therapy studies in small animals

PURPOSE To develop an alternating focused ultrasound system (AFUS) for preclinical studies of thermal and acoustic responses of tumors in small animal models. This work was motivated by the need of noninvasively creating relatively small spheroidal thermal lesions in small targets (e.g., a murine tumor) without damaging the surrounding tissues. METHODS The AFUS consists of two lead zirconate titanate (PZT-4) spherically curved ultrasound transducers with focal zones crossing each other at a 90 degrees angle. The transducers were independently powered following a programed alternating firing scheme. Before the device design and construction, an acoustic and biothermal model was developed to simulate the ultrasound pressure field and the resulting temperature and thermal dose distributions. A shape factor, sphericity, to quantify the roundness of the lesions was calculated based on the 240 equivalent minutes at 43 degrees C thermal dose contours. A prototype of the AFUS was constructed with two identical transducers of an operating frequency of 2.25 MHz, 38 mm in diameter, and F-number equal to 1.33. To evaluate the performance of the AFUS experimentally, a series of heating in polyacrylamide phantoms, ex vivo porcine liver tissues, and in implanted mouse tumors fibrosarcoma (FSaII) in vivo was conducted. In these experimental cases, the sphericity was calculated and compared based on the visible lesion (a marked change in coloration). RESULTS As shown in the simulations, the lesions induced in polyacrylamide phantoms, ex vivo porcine liver tissues, and in vivo mouse tumors, the sphericities of the lesions yielded by AFUS heating were approximately 50% higher than those of single focused ultrasound heating as long as moderate intensities were used and the duty cycle pulses were distributed equally among the transducers. CONCLUSIONS The AFUS is a device capable of noninvasively creating spheroidal thermal lesions in small targets such as murine tumors.

Conductive thermal ablation of 4T1 murine breast carcinoma reduces severe hypoxia in surviving tumour

Purpose: The purpose of this study was to quantify hypoxia changes in viable tumour volumes after thermal ablation of a murine breast carcinoma. Methods: Murine breast 4T1 tumours were grown in the rear leg of BALB/c mice to an average diameter of 10–12 mm. Tumours were treated with conductive interstitial thermal therapy (CITT) at a peak temperature of 80–90°C for 10 min. The animals were euthanised 72 h later, and the tumours were removed for immunohistochemical staining with pimonidazole – a marker of partial pressure of oxygen. The levels of pimonidazole staining intensity were used to quantify changes in hypoxia gradients in terms of strong, medium and weak positive pixel fractions. Results: The pimonidazole staining ratio of viable control tumour tissue to viable tissue in tumours that were ablated was 0.7 for weak staining, 2.7 for medium staining and 8.0 (p < 0.03) for strong pimonidazole staining. Conclusion: This shift of pimonidazole staining toward lower intensity pixels in the remaining tumour indicates that tumour ablation with CITT may increase radiosensitivity of the remaining tumour tissue and presents a rationale for combination therapy.

SU‐FF‐J‐160: Spatially Fractionated Radiation Therapy (GRID) On Implanted Tumors Using a Small Animal Conformal Radiation Therapy System

Purpose: To deliver spatially fractionated (GRID) radiation fields to murine tumors using a high precision small animal conformal radiation therapy device (SACRTD). Method and Materials: The SACRTD is a “gantry” based image‐guide system with a special collimating assembly that allows field sizes down to 0.5 mm in diameter at isocenter. The precise positioning of the target/beam is achieved by a six‐degree of freedom robotic system (Adept Technology, Pleasanton, CA) which has positional repeatability of ±0.020mm in XYZ directions. The system can be used to deliver small GRID fields with varying open‐to‐close area ratios. Immunohistochemistry for DNA damage after 10 Gy to a 1cm depth GRID therapy was performed in B16 murine melanoma tumors. The GRID pattern consisted of an array of 7 beams of 1 mm in diameter at isocenter with center‐to‐center separation of 2 mm. Tumorgrowth rates and cell survival were assessed. Results : The exit dose recorded by Gafchromic film confirmed that the GRID radiation pattern was delivered with high precision. Histology showed the radiation induced DNA strand breaks in the open regions of the GRID field. Approximately 55% of clonogenic cells in the B16 tumors were killed by the single GRID dose. Interestingly, mice receiving GRID alone did not have significant tumorgrowth delay compared to untreated tumors. However, in the group that received GRID therapy followed by Anginex anti‐angiogenic peptide for 5 days there was a noticeable delay in tumorgrowth compared to tumors receiving 10 Gy GRID alone. Conclusion: This preliminary biological study shows the usefulness and flexibility of the SACRTD to deliver precise radiation patterns to implanted tumors in small animals. The system facilitates in vivo radiobiological studies and promotes translational research in radiationoncology.

Sclerostin Antibody Treatment Stimulates Bone Formation to Normalize Bone Mass in Male Down Syndrome Mice: SclAb INCREASES BONE FORMATION IN DS MICE

Down syndrome (DS), characterized by trisomy of human chromosome 21, is associated with a variety of endocrine disorders as well as profound skeletal abnormalities. The low bone mass phenotype in DS is defined by low bone turnover due to decreased osteoclast and osteoblast activity, decreasing the utility of antiresorptive agents in people with DS. Sclerostin antibody (SclAb) is a therapeutic candidate currently being evaluated as a bone anabolic agent. Scl, the product of the sclerostin gene (SOST), inhibits bone formation through its inhibition of Wnt signaling. SclAb increases bone mass by suppressing the action of the endogenous inhibitor of bone formation, Scl. To examine the effects of SclAb on the DS bone phenotype, 8‐week‐old male wild‐type (WT) andTs65Dn DS mice were treated with 4 weekly iv injections of 100 mg/kg SclAb. Dual‐energy X‐ray absorptiometry (DXA), microCT, and dynamic histomorphometry analyses revealed that SclAb had a significant anabolic effect on both age‐matched WT littermate controls and Ts65Dn DS mice that was osteoblast mediated, without significant changes in osteoclast parameters. SclAb treatment significantly increased both cortical and trabecular bone mass at multiple sites; SclAb treatment resulted in the normalization of Ts65Dn bone mineral density (BMD) to WT levels in the proximal tibia, distal femur, and whole body. Ex vivo bone marrow cultures demonstrated that SclAb increased the recruitment of the mesenchymal progenitors into the osteoblast lineage, as indicated by increased alkaline phosphatase–positive colonies, with no effect on osteoclast differentiation. Together, in the setting of a murine model of DS and decreased bone turnover, SclAb had a potent anabolic effect. SclAb stimulated bone formation and increased osteoblastogenesis without affecting osteoclastogenesis or bone resorption. These data suggest that SclAb is a promising new therapy to improve bone mass and reduce fracture risk in the face of the low bone mass and turnover prevalent in the DS population.

Abstract 4644: A novel strategy for targeted drug delivery to the tumor vasculature by radiation-induced receptor expression on endothelial cells

One of the primary goals of a successful cancer treatment regimen is to deliver an effective combination of radiation and/or drugs to tumors while minimizing damage to normal tissues. Many anti-angiogenic agents, while not able to control tumor growth, possess the ability to selectively target the location and process of tumor blood vessel formation. Conversely many chemotherapy agents are highly cytotoxic and lack selective targeting ability thus decreasing the therapeutic ratio. We are investigating a new drug delivery strategy exploiting the tumor endothelium “stimulated” by ionizing radiation to preferentially target and deliver a nanoparticulate formulation of arsenic trioxide (ATO) encapsulated in liposomal vesicles or “nanobins” to the irradiated tumor tissue. The targeting is via the 33 amino acid anti-angiogenic peptide, anginex. The identification of galectin-1 as the receptor for anginex expressed on activated endothelial cells involved in tumor angiogenesis has revealed a solid basis for this therapeutic rationale. Unlike the antiangiogenic agent Avastin, which is an antibody that targets the vascular endothelial growth factor (VEGF) released by the tumor cells into the microenvironment, this peptide binds to galectin-1, a tumor endothelial cell specific antigen that is expressed in solid tumors. We have made the novel discovery that galectin-1 expression is further upregulated in the tumor after radiation exposure, particularly on the endothelial cell surface. Exposure of murine SCK breast tumors to a clinical radiation dose of 2Gy induced a substantial average increase of 141 +/− 49% in anginex uptake as assessed by [18]-F-labeled anginex biodistribution. These results suggest that anginex may be an effective targeting molecule for image and radiation-guided therapy of solid tumors. To assist in delineating the exact mechanisms for our radiation-guided drug delivery strategy, we have developed a system to grow 3D tumor cell-endothelial cell spheroids and have observed increased galectin-1 expression upon radiation exposure. We are using these spheroidal cultures to implant tumors in dorsal skin fold window chambers for intravital wavelength imaging of drug delivery to tumor tissue before and after radiation exposure. Understanding how anginex uptake selectively increases in endothelial cells after irradiation and its nexus to radiation-sensitivity and drug delivery is our current focus. Further studies are underway to characterize the in vivo targeting of arsenic trioxide chemotherapy via anginex-conjugated ‘nanobins’ in combination with radiation exposure and assess the therapeutic potential in our solid tumor model. Supported by NCI grant CA107160 and the Central Arkansas Radiation Therapy Institute. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4644. doi:1538-7445.AM2012-4644

Dual thermal ablation modality of solid tumors in a mouse model

Purpose: Develop a new combination therapy consisting of cryoablation and conductive high-temperature ablation for enhanced thermal ablation of solid tumors. Methods: We have constructed an invasive probe that can be used for consecutive cryoablation and hightemperature ablation (C/HTA), with a single insertion. The C/HTA probe was tested, in Balb/c mice bearing solid 4T1 tumors, in comparison to cryoablation and high temperature ablation, only. Three days after ablation, the diameter of the ablated zone was evaluated with pathological examination. Results: The C/HTA device can be used to induce larger ablation zones, in comparison to high temperature or cryoablation alone, and at lower thermal doses and temperatures than either modality alone. Conclusions: The relatively high thermal conductivity of ice, in comparison to water and native tissue, enables rapid heating of the ice-ball that result in improved conductive high temperature ablation. The new dual thermal modality improves ablation outcomes at lower thermal doses in comparison to a single ablation modality.