Rodent Models Of Breast Cancer Research Articles

Tantalum oxide nanoparticles as versatile and high-resolution X-ray contrast agent for intraductal image-guided ablative procedure in rodent models of breast cancer

AbstractThere are limited options for primary prevention of breast cancer (BC). Experimental procedures to locally prevent BC have shown therapeutic efficacy in animal models. To determine the suitability of FDA-approved iodine-containing and various metal-containing (bismuth, gold, iodine, or tantalum) preclinical nanoparticle-based contrast agents for image-guided intraductal (ID) ablative treatment of BC in rodent models, we performed a prospective longitudinal study to determine the imaging performance, local retention and systemic clearance, safety profile, and compatibility with ablative solution of each contrast agent. At least six abdominal mammary glands (>3 female FVB/JN mice and/or Sprague-Dawley rats, 10–11 weeks of age) were intraductally injected with commercially available contrast agents (Omnipaque® 300, Fenestra® VC, MVivoTM Au, MVivoTM BIS) or in-house synthesized tantalum oxide (TaOx) nanoparticles. Contrast agents were administered at stock concentration or diluted in 70% ethanol (EtOH) and up to 1% ethyl cellulose (EC) as gelling agent to assess their compatibility with our image-guided ablative procedure. Mammary glands were serially imaged by microCT for up to 60 days after ID delivery. Imaging data were analyzed by radiologists and deep learning to measure in vivo signal disappearance of contrast agents. Mammary glands and major organs were ultimately collected for histopathological examination. TaOx-containing solutions provided best imaging performance for nitid visualization of ductal tree immediately after infusion, low outward diffusion (<1 day) and high homogeneity. Of all nanoparticles, TaOx had the highest local clearance rate (46% signal decay as stock and 36% as ablative solution 3 days after ID injection) and exhibited low toxicity. TaOx-containing ablative solution with 1% EC caused same percentage of epithelial cell death (88.62% ± 7.70% vs. 76.38% ± 9.99%, p value = 0.089) with similar minimal collateral damage (21.56 ± 5.28% vs. 21.50% ± 7.14%, p value = 0.98) in mouse and rat mammary glands, respectively. In conclusion, TaOx-nanoparticles are a suitable and versatile contrast agent for intraductal imaging and image-guided ablative procedures in rodent models of BC with translational potential to humans.

Advances in Immunocompetent Mouse and Rat Models.

Rodent models of breast cancer have played critical roles in our understanding of breast cancer development and progression as well as preclinical testing of cancer prevention and therapeutics. In this article, we first review the values and challenges of conventional genetically engineered mouse (GEM) models and newer iterations of these models, especially those with inducible or conditional regulation of oncogenes and tumor suppressors. Then, we discuss nongermline (somatic) GEM models of breast cancer with temporospatial control, made possible by intraductal injection of viral vectors to deliver oncogenes or to manipulate the genome of mammary epithelial cells. Next, we introduce the latest development in precision editing of endogenous genes using in vivo CRISPR-Cas9 technology. We conclude with the recent development in generating somatic rat models for modeling estrogen receptor-positive breast cancer, something that has been difficult to accomplish in mice.

X-Ray Visualization of Intraductal Ethanol-Based Ablative Treatment for Prevention of Breast Cancer in Rat Models

There are still a limited number of primary interventions for prevention of breast cancer. For women at a high risk of developing breast cancer, the most effective intervention is prophylactic mastectomy. This is a drastic surgical procedure in which the mammary epithelial cells that can give rise to breast cancer are completely removed along with the surrounding tissue. The goal of this protocol is to demonstrate the feasibility of a minimally invasive intraductal procedure that could become a new primary intervention for breast cancer prevention. This local procedure would preferentially ablate mammary epithelial cells before they can become malignant. Intraductal methods to deliver solutions directly to these epithelial cells in rodent models of breast cancer have been developed at Michigan State University and elsewhere. The rat mammary gland consists of a single ductal tree that has a simpler and more linear architecture compared to the human breast. However, chemically induced rat models of breast cancer offer valuable tools for proof-of-concept studies of new preventive interventions and scalability from mouse models to humans. Here, a procedure for intraductal delivery of an ethanol-based ablative solution containing tantalum oxide nanoparticles as X-ray contrast agent and ethyl cellulose as gelling agent into the rat mammary ductal tree is described. Delivery of aqueous reagents (e.g., cytotoxic compounds, siRNAs, AdCre) by intraductal injection has been described previously in mouse and rat models. This protocol description emphasizes methodological changes and steps that pertain uniquely to delivering an ablative solution, formulation consideration to minimize local and systemic side effects of the ablative solution, and X-ray imaging for in vivo assessment of ductal tree filling. Fluoroscopy and micro-CT techniques enable to determine the success of ablative solution delivery and the extent of ductal tree filling thanks to compatibility with the tantalum-containing contrast agent.

Intraductal Delivery and X-ray Visualization of Ethanol-Based Ablative Solution for Prevention and Local Treatment of Breast Cancer in Mouse Models

Breast cancer is the most prevalent cancer and the second-leading cause of cancer-related death for women in the USA. For high-risk women, prophylactic mastectomy is the most effective primary prevention strategy. Prophylactic mastectomy is an aggressive surgical procedure that completely removes the mammary epithelial cells from which breast cancer arises along with the surrounding tissue. We seek to develop a minimally invasive intraductal procedure as an alternative to prophylactic mastectomy to locally ablate the mammary epithelial cells before they can become malignant. We and others have developed an intraductal delivery procedure to reach and treat these epithelial cells in rodent models of breast cancer. While the mouse mammary gland with a single non-anastomosed ductal tree opening at the nipple has a much less complex and tortuous architecture than the human breast, chemically induced and genetically engineered mouse models of breast cancer are valuable to produce proof-of-concept studies of new preventative strategies. Here, we describe a procedure for intraductal delivery of an ethanol-based ablative solution containing micro-CT/X-ray tantalum-based contrast agent within the mouse mammary ductal tree for the therapeutic purpose of primary prevention of breast cancer. Intraductal delivery of aqueous reagents (e.g., cytotoxic compounds, siRNAs, AdCre) has been previously described in mouse models. Thus, we focus our protocol description on methodological modifications and unique experimental considerations for optimizing delivery of ethanol, for minimizing local and systemic side effects of ethanol administration, and for in vivo visualization of ductal tree filling via micro-CT/fluoroscopy imaging. Visualization of the ductal tree immediately after injection of a contrast-containing solution allows for confirmation of complete filling or unsuccessful outcomes such as underfilling or overfilling. This procedure can be applied for delivery and imaging of other ablative compounds aimed at either preventing tumor formation or locally treating early-stage tumors accessible via the ductal tree.

Intraductal Delivery and X-ray Visualization of Ethanol-Based Ablative Solution for Prevention and Local Treatment of Breast Cancer in Mouse Models.

Breast cancer is the most prevalent cancer and the second-leading cause of cancer-related death for women in the USA. For high-risk women, prophylactic mastectomy is the most effective primary prevention strategy. Prophylactic mastectomy is an aggressive surgical procedure that completely removes the mammary epithelial cells from which breast cancer arises along with the surrounding tissue. We seek to develop a minimally invasive intraductal procedure as an alternative to prophylactic mastectomy to locally ablate the mammary epithelial cells before they can become malignant. We and others have developed an intraductal delivery procedure to reach and treat these epithelial cells in rodent models of breast cancer. While the mouse mammary gland with a single non-anastomosed ductal tree opening at the nipple has a much less complex and tortuous architecture than the human breast, chemically induced and genetically engineered mouse models of breast cancer are valuable to produce proof-of-concept studies of new preventative strategies. Here, we describe a procedure for intraductal delivery of an ethanol-based ablative solution containing micro-CT/X-ray tantalum-based contrast agent within the mouse mammary ductal tree for the therapeutic purpose of primary prevention of breast cancer. Intraductal delivery of aqueous reagents (e.g., cytotoxic compounds, siRNAs, AdCre) has been previously described in mouse models. Thus, we focus our protocol description on methodological modifications and unique experimental considerations for optimizing delivery of ethanol, for minimizing local and systemic side effects of ethanol administration, and for in vivo visualization of ductal tree filling via micro-CT/fluoroscopy imaging. Visualization of the ductal tree immediately after injection of a contrast-containing solution allows for confirmation of complete filling or unsuccessful outcomes such as underfilling or overfilling. This procedure can be applied for delivery and imaging of other ablative compounds aimed at either preventing tumor formation or locally treating early-stage tumors accessible via the ductal tree.

Paclitaxel Chemotherapy Elicits Widespread Brain Anisotropy Changes in a Comprehensive Mouse Model of Breast Cancer Survivorship: Evidence From In Vivo Diffusion Weighted Imaging.

Breast cancer is one of the most common diseases in the United States with 1 in 8 women developing the disease in her lifetime. Women who develop breast cancer are often post-menopausal and undergo a complex sequence of treatments including surgery, chemotherapy, and aromatase inhibitor therapy. Both independently and through potential interactions, these factors and treatments are associated with behavioral comorbidities reported in patients (e.g., fatigue), although the underlying neurobiological mechanisms are poorly understood. Currently, brain imaging is the most feasible way to assess neurobiology in patients. Indeed, breast cancer patients display alterations in white matter connections and chemotherapy is associated with decreased white and gray matter in the corpus callosum and cortex as well as decreased hippocampal volume. However, imaging in breast cancer rodent models is lacking, impeding translation of the mechanistic neurobiological findings made possible through modeling. Furthermore, current rodent models of breast cancer often lack the complexity of typical multimodal breast cancer treatments, thereby limiting translational value. The present study aimed to develop a comprehensive model of post-menopausal breast cancer survival using immunocompetent ovariectomized mice, including an orthotopic syngeneic tumor, surgical tumor removal, chemotherapy, and aromatase inhibitor therapy. Using this model, we systematically investigated the cumulative effects of chemotherapy and hormone replacement therapy on neurostructure and behavior using diffusion weighted imaging, open field test, and spontaneous alternation test. Our previous findings, in a simplified chemotherapy-only model, indicate that this regimen of chemotherapy causes circulating and central inflammation concurrent with reduced locomotor activity. The current study, in the more comprehensive model, has recapitulated the peripheral inflammation coincident with reduced locomotor activity as well as demonstrated that chemotherapy also drives widespread changes in brain anisotropy. Validating the clinical relevance of this comprehensive rodent breast cancer model will allow for additional neurobiological investigations of the interactions among various cancer components associated with behavioral comorbidities, as well as the relationship between these mechanisms and neurostructural imaging changes that can be measured in cancer patients.

Large Animal Models of Breast Cancer.

In this mini review the status, advantages, and disadvantages of large animal modeling of breast cancer (BC) will be discussed. While most older studies of large animal BC models utilized canine and feline subjects, more recently there has been interest in development of porcine BC models, with some early promising results for modeling human disease. Widely used rodent models of BC were briefly reviewed to give context to the work on the large animal BC models. Availability of large animal BC models could provide additional tools for BC research, including availability of human-sized subjects and BC models with greater biologic relevance.

Advances in Rodent Models for Breast Cancer Formation, Progression, and Therapeutic Testing.

Breast cancer is a highly complicated disease. Advancement in the treatment and prevention of breast cancer lies in elucidation of the mechanism of carcinogenesis and progression. Rodent models of breast cancer have developed into premier tools for investigating the mechanisms and genetic pathways in breast cancer progression and metastasis and for developing and evaluating clinical therapeutics. Every rodent model has advantages and disadvantages, and the selection of appropriate rodent models with which to investigate breast cancer is a key decision in research. Design of a suitable rodent model for a specific research purpose is based on the integration of the advantages and disadvantages of different models. Our purpose in writing this review is to elaborate on various rodent models for breast cancer formation, progression, and therapeutic testing.

Effect of Zinc Supplementation on the Serum Metabolites Profile at the Early Stage of Breast Cancer in Rats.

The cytotoxic properties of zinc nanoparticles have been evaluated in vitro against several types of cancer. However, there is a lack of significant evidence of their activity in vivo, and a potential therapeutic application remains limited. Herein we report the effective inhibition of tumor growth by zinc nanoparticles in vivo, as the effect of the dietary intervention, after the chemical induction in a rodent model of breast cancer. Biopsy images indicated grade 1 tumors with multiple inflammatory infiltrates in the group treated with zinc nanoparticles, whereas, in the other groups, a moderately differentiated grade 2 adenocarcinoma was identified. Moreover, after the supplementation with zinc nanoparticles, the levels of several metabolites associated with cancer metabolism, important to its survival, were found to have been altered. We also revealed that the biological activity of zinc in vivo depends on the size of applied particles, as the treatment with zinc microparticles has not had much effect on cancer progression.

Abstract 5243: Averting tumor growth in rodent breast cancer models with a liquid ablation approach

Abstract In low-resource settings, surgery or radiation are inaccessible to 90% of cancer patients due to inadequate infrastructure and lack of trained personnel. Resource appropriate technologies must be developed to make cancer treatments accessible. Ablative techniques, such as radiofrequency ablation are often used as a minimally invasive alternative to surgery. However, energy-based ablation techniques require expensive machinery and a steady supply of electricity and expensive consumables. To address these shortcomings, we have developed a low-cost ablation technique that stabilizes tumor growth with a simple injection of ethanol in combination with a polymer, ethylcellulose (EC). The EC injected with ethanol creates a slow-release depot of ethanol at the injection site by undergoing a solvent induced phase-transition initiated by water in tissues. EC-ethanol improves upon traditional ethanol ablation by providing increased precision while remaining low-cost and simple to administer. We evaluated the precision of EC-ethanol as well as its vascular sclerosing capabilities as a strategy to avert tumor growth in rodent models of breast cancer. EC-ethanol injected at 1 mL/hr with 6% w/w EC was used as this demonstrated to be the optimal formulation and rate for causing tumor necrosis (Morhard et al., Scientific Reports, 2017) and enhanced the anti-tumor immunostimulatory response (Nief et al., 2019, Under Review). We demonstrate the precision delivery of EC-ethanol in orthotopic 67NR mammary tumor as with small-animal MRI. Further, ethanol devascularization is monitored with photoacoustic imaging 24 hours post ablation. Local tumor control was demonstrated both in 67NR mammary tumor and in 7,12-Dimethylbenz[a]anthracene (DMBA) induced rat mammary tumor models. Repeated ablations on significantly decrease primary tumor growth compared to untreated tumors in both the mouse and rat models (P<0.05). In rats weekly 200 µL EC-ethanol injections began when tumors reached 2 mL in volume (n=5), and in 30 days tumors decreased 20% ± 3% untreated controls grew an average of 721%. For mice (n=5), 50 µL was given daily for 6 days on day 3 after tumor implantation to account for small body weight and rapidly growing tumors. Mice treated with EC-ethanol lived significantly longer (P<0.05 log-rank test) than untreated mice and mice injected with the same volume of pure ethanol. EC-ethanol is a low-cost, non-invasive tumor destruction method that shows a clear control of tumor growth when applied as a fractionated dose. We utilized a mouse and rat tumor model to ensure that the EC-ethanol's success is not dependent on the model. Ongoing studies will examine the impact of varying injected volumes at well-defined fractionation schedules on ethanol distribution, necrosis, tumor growth, and survival. Citation Format: Corrine A. Nief, Erika Chelales, Robert Morhard, Maomao Chem, Jeffrey Everitt, Jenna Mueller, Junjie Yao, Mark W. Dewhirst, Nirmala Ramanujam. Averting tumor growth in rodent breast cancer models with a liquid ablation approach [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5243.

DETERMINING THE EFFECTS OF PRO‐ANGIOGENIC ELP‐VEGF THERAPY ON TUMOR GROWTH AND PROGRESSION

The members of the Vascular Endothelial Growth Factor (VEGF) family are endogenous growth factors with a central role in angiogenesis and endothelial cell health. We recently showed the therapeutic potential of renal VEGF delivery using a fusion protein of VEGF bound to the carrier protein Elastin‐like Polypeptide (ELP‐VEGF) designed in our laboratory, to promote angiogenesis in chronic kidney diseases. Indeed, we showed that ELP‐VEGF therapy improves renal vascular density, decreased renal fibrosis, and improved glomerular filtration rate (GFR) in swine models of chronic renovascular disease and chronic kidney disease at therapeutic doses of 0.1 – 1.0 mg/kg.A potential limitation of the use of ELP‐VEGF as a therapeutic is the possibility of promoting uninhibited off‐target angiogenic growth which may be undesirable in certain disease processes such as cancer. In order to determine whether ELP‐VEGF therapy may be harmful for a patient with an undiagnosed tumor, the effects of a single administration of escalating doses (0.1 m/kg, 1 mg/kg, 10 mg/kg, 10 times the highest therapeutic dose) of ELP‐VEGF on tumor size and vascular density were determined in a mouse xenograft model of breast cancer.Human triple‐negative breast cancer cells (MDA‐MB‐231) that stably express a luciferase reporter were implanted into the mammary fat pad of athymic nude mice. Tumors were monitored thrice weekly for 7 weeks with external caliper measurements (blind) and bioluminescence imaging to assess tumor growth. Body weights of the mice were also recorded 3 times per week. At the end of the 7‐week period, mice were fully perfused with heparinized saline and microfil. Tumors were removed and scanned using micro‐computed tomography (u‐CT), and 3D reconstructions of the tumor vascular network were used to blindly quantify tumor vascular density.Caliper measurements and bioluminescence imaging showed no significant difference in tumor size or growth rate amongst the groups. There were also no significance differences in mouse body weights throughout the study or final weights of the tumors (0.1–10mg/kg 1.12±0.39 g, 1.233±0.46 g, 1.247±0.52 g, respectively; p=0.8695). However, analysis of the renal vasculature by u‐CT revealed a significant increase in total microvascular density after a single injection of ELP‐VEGF at 1mg/kg (78.88 ± 34.34 vessels/mm2; p = 0.0489 vs. controls) and 10mg/kg (97.17 ± 48.65 vessels/mm2; p = 0.0012 vs. controls). In conclusion, we determined that one‐time injection of ELP‐VEGF had no toxic effects on body weights and did not influence tumor growth rate, but it did significantly and dose‐dependently increase tumor vasculature density in a breast cancer xenograft model.Support or Funding InformationR01HL095638 and IPA34170267Figure 2. Analysis and results from ELP‐VEGF‐A administration to a rodent model of breast cancer. Results include body weight (a), tumor caliper measurements (b) tumor bioluminescence (c), final tumor mass(d).Figure 1Figure 3. Vasculature of each tumor was visualized using uCT scanner after microfi fixation. Blood vessels were divided and quantified by diameter between different ranges (a). Kidneys from these mice were also perfused with microfil, scanned through uCT scanner, and quantified by vessel diameter range (b).Figure 2

Withaferin A-mediated apoptosis in breast cancer cells is associated with alterations in mitochondrial dynamics

Withaferin A (WA), a steroidal lactone derived from a medicinal plant (Withania somnifera), inhibits cancer development in transgenic and chemically-induced rodent models of breast cancer but the underlying mechanism is not fully grasped. We have shown previously that WA treatment causes apoptotic cell death in human breast cancer cells that is preceded by inhibition of complex III of the mitochondrial electron transport chain. This study extends these observations to now demonstrate alterations in mitochondrial dynamics in WA-induced apoptosis. Assembly of complex III was decreased in MCF-7 and SUM159 cells but not in MDA-MB-231 as determined by native blue gel electrophoresis. Because WA is a Michael acceptor (electrophile), we explored the possibility of whether it covalently modifies cysteine residue(s) in ubiquinol-cytochrome c reductase, Rieske iron-sulfur polypeptide 1 (UQCRFS1). Covalent modification of cysteine in UQCRFS1 was not observed after WA treatment. Instead, WA treatment inhibited chemically-induced mitochondrial fusion and decreased the mitochondrial volume, and this effect was accompanied by a decrease in the expression of proteins involved in fusion process, including mitofusin1, mitofusin2, and full-length optic atrophy protein 1 (OPA1). A loss of volume in fragmented mitochondria also occurred in WA-exposed cells when compared to vehicle-treated control. WA treatment also caused a decrease in protein level of mitochondrial fission-regulating protein dynamin-related protein 1 (DRP1). Functional studies revealed that DRP1 deficiency and OPA1 knockdown attenuated apoptotic potential of WA. Taken together, these results indicate that WA not only alters Complex III assembly but also inhibits mitochondrial dynamics in breast cancer cells.

Ultrasound-mediated delivery of siESE complexed with microbubbles attenuates HER2+/- cell line proliferation and tumor growth in rodent models of breast cancer.

The highly tunable, noninvasive and spatially targeted nature of microbubble-enhanced, ultrasound-guided (MB+US) drug delivery makes it desirable for a wide variety of therapies. In breast cancer, both HER2+ and HER2- type neoplasms pose significant challenges to conventional therapeutics in greater than 40% of breast cancer patients, even with the widespread application of biologics such as trastuzumab. To address this therapeutic challenge, we examined the novel combination of tumor-injected microbubble-bound siRNA complexes and monodisperse size-isolated microbubbles (4-µm diameter) to attenuate tumor growth in vivo, as well as MB+US-facilitated shRNA and siRNA knockdown of ESE-1, an effector linked to dysregulated HER2 expression in HER2+/- cell line propagation. We first screened six variants of siESE and shESE for efficient knockdown of ESE in breast cancer cell lines. We demonstrated efficient reduction of BT-474 (PR+, ER+, HER2+; luminal B) and MDA-MB-468 (PR-, ER-, HER2-; triple-negative) clonogenicity and non-adherent growth after knockdown of ESE-1. A significant reduction in proliferative potential was seen for both cell lines using MB+US to deliver shESE and siESE. We then demonstrated significant attenuation of BT-474 xenograft tumor growth in Nod/SCID female mice using direct injection of microbubble-adsorbed siESE to the tumor and subsequent sonication. Our results suggest a positive effect on drug delivery from MB+US, and highlights the feasibility of using RNAi and MB+US for breast cancer pathologies. RNAi coupled with MB+US may also be an effective theranostic approach to treat other acoustically accessible tumors, such as melanoma, thyroid, parotid and skin cancer.

Daily or weekly dosing with EGFR inhibitors, gefitinib and lapatinib, and AKt inhibitor MK2206 in mammary cancer models.

Daily vs. weekly dosing with EGFR inhibitors (gefitinib and lapatinib) and an AKT inhibitor (MK2206) were compared in two rodent breast cancer models. Female Sprague-Dawley rats were administered methylnitrosourea (MNU) at 50 days of age, and gefitinib (daily/weekly dosing at 10/70 mg/kg BW) or lapatinib (daily/weekly dosing at 75/525 mg/kg BW) were administered by gavage beginning 5 days after MNU. For the prevention studies, weekly or daily dosing with gefitinib or lapatinib reduced cancer multiplicity >75%, and all treatments reduced tumor weights by >90%. For the therapeutic studies, MNU-treated rats were followed until small palpable mammary cancers developed. The rats were then treated daily or weekly as above for 6 weeks. Either daily or weekly dosing with lapatinib or gefitinib caused regression in >50% of the tumors. Immunohistochemistry biomarker studies in palpable mammary cancers following a weekly dose of gefitinib showed that 1 day (but not 7 days) after treatment, the levels of phosphorylated EGFR1 were significantly decreased. In an ER-negative (ER−) Neu-overexpressing model employing MMTV-Neu/P53KO mice, daily (100 mg/kg BW/day, 5 days each week), or weekly dosing (500 or 250 mg/kg BW) with gefitinib reduced tumor multiplicity 65, 85 and 75%, respectively. In the MNU prevention model, daily dosing (100 mg/kg BW/day) with the allosteric AKT inhibitor MK2206 was ineffective, while weekly dosing (700 mg/kg BW) reduced the final tumor weight >70%. Combining weekly MK2206 with the aromatase inhibitor vorozole (0.12 mg/kg BW/day) showed that each compound alone reduced tumor multiplicity 40–50%. The combination reduced cancer multiplicity ~70%. These studies demonstrate the efficacy of weekly dosing with various protein kinase inhibitors; raising the possibility of employing these agents in a breast cancer preventive setting.

Disease modifying actions of interleukin-6 blockade in a rat model of bone cancer pain.

Metastasis of cancer to the skeleton represents a debilitating turning point in the lives of patients. Skeletal metastasis leads to moderate to severe ongoing pain along with bone remodeling that can result in fracture, events that dramatically diminish quality of life. Interleukin-6 (IL-6) levels are elevated in patients with metastatic breast cancer and are associated with a lower survival rate. We therefore determined the consequences of inhibition of IL-6 signaling using a novel small molecule antagonist, TB-2-081, on bone integrity, tumor progression, and pain in a rodent model of breast cancer. Rat MAT B III mammary adenocarcinoma cells were injected and sealed within the tibia of female Fischer rats. Growth of these cells within the rat tibia elicited increased IL-6 levels both within the bone exudate and in the plasma, produced ongoing pain and evoked hypersensitivity, and bone fracture that was observed by approximately day 12. Systemic TB-2-081 delivered by subcutaneous osmotic minipumps starting at tumor implantation prevented tumor-induced ongoing bone pain and evoked hypersensitivity without altering tumor growth. Remarkably, TB-2-081 infusion significantly reduced osteolytic and osteoblastic bone remodeling and time to fracture likely by decreasing osteoclastogenesis and associated increase in bone resorption. These findings indicate that blockade of IL-6 signaling may represent a viable, disease-modifying strategy to prevent tumor-induced bone remodeling allowing for stabilization of bone and decreased fractures as well as diminished ongoing pain that may improve quality of life of patients with skeletal metastases. Notably, anti-IL-6 antibodies are clinically available allowing for rapid testing of these possibilities in humans.

The Best of Quantitative Ultrasound

The objective of these studies was to demonstrate that quantitative ultrasound (QUS) techniques could be used to improve diagnostic ultrasound imaging. Specifically, QUS techniques were applied to improving detection of breast and thyroid cancer, early detection of preterm birth, quantification of fatty liver and thermometry. QUS techniques were employed that utilized parameterization of the frequency-dependent backscatter coefficient, the attenuation coefficient and/or envelope statistics. The backscatter coefficient was parameterized through the effective scatterer diameter (ESD) and the effective acoustic concentration (EAC). Backscattered ultrasound data were acquired from tissue samples using clinical ultrasound scanners providing the RF signal and from laboratory systems utilizing single-element focused transducers. The data spanned frequency ranges from 2 to 50 MHz. Combinations of the ESD, EAC and envelope statistics were successful in differentiating malignant from benign tumors in rodent models of breast cancer. Similarly, the ESD combined with envelope statistics provided differentiation of malignant from benign nodules in mouse models of thyroid cancer. Cervical attenuation was an early predictor of preterm birth in rat models and in humans. Attenuation and the slope and magnitude of the backscatter coefficient were observed to predict the degree of fatty liver in mouse models and in humans. The EAC was observed to track temperature when using thermal therapies in different tissues. QUS techniques have continued to mature and have demonstrated new sources of contrast leading to improved diagnostic capabilities and the ability to monitor and assess therapy.

Ospemifene: A first-in-class, non-hormonal selective estrogen receptor modulator approved for the treatment of dyspareunia associated with vulvar and vaginal atrophy

Ospemifene is a selective estrogen receptor modulator (SERM) approved for the treatment of dyspareunia associated with vulvar and vaginal atrophy (VVA) due to menopause. As the first non-hormonal treatment for this indication, the approval of ospemifene represents a significant milestone in postmenopausal women’s health. Ospemifene is a triphenylethylene similar in chemical structure to tamoxifen and toremifene. Consistent with other SERMs such as tamoxifen, toremifene, and raloxifene, ospemifene possesses a distinctive mix of estrogenic and antiestrogenic tissue-specific effects in bone, breast tissue, serum lipids, and the vagina. Among the approved SERMs, ospemifene is the only agent with a nearly full estrogen agonist effect on the vaginal epithelium while having neutral to slight estrogenic effects in the endometrium, making ospemifene uniquely suited for the treatment of dyspareunia associated with VVA, also known as atrophic vaginitis, which affects up to 50% of postmenopausal women. This review begins with a brief history of the discovery of ospemifene, its mechanism of action, and its preclinical development, with an emphasis on its tissue-specific effects on bone, breast, uterus and endometrium, serum lipids and vagina. A brief discussion on the genotoxicity of ospemifene compared to tamoxifen and toremifene is included. The focus then shifts to the clinical development of ospemifene from Phase I through Phase III. We will close with the FDA approval of ospemifene and a justification of the future clinical evaluation of ospemifene as a potential breast cancer chemopreventive agent, where several preclinical studies in different rodent breast cancer models strongly suggest ospemifene is as effective as tamoxifen.

Contrast ultrasound-guided photothermal therapy using gold nanoshelled microcapsules in breast cancer

ObjectivesThe purpose of this study was to test whether dual functional gold nano-shelled microcapsules (GNS-MCs) can be used as an ultrasound imaging enhancer and as an optical absorber for photothermal therapy (PTT) in a rodent model of breast cancer. MethodsGNS-MCs were fabricated with an inner air and outer gold nanoshell spherical structure. Photothermal cytotoxicity of GNS-MCs was tested with BT474 cancer cells in vitro and non-obese diabetes-SCID (NOD/SCID) mice with breast cancer. GNS-MCs were injected into the tumor under ultrasound guidance and treated with near-infrared (NIR) laser irradiation. The photothermal ablative effectiveness of GNS-MCs was evaluated by measuring the surface and internal temperature of the tumor as well as the size of the tumor using histological confirmation. ResultsNIR laser irradiation resulted in significant tumor cell death in GNS-MCs-treated BT474 cells in vitro. GNS-MCs were able to serve as an ultrasound enhancer to guide the intratumoral injection of GNS-MCs and ensure their uniform distribution. In vivo studies revealed that NIR laser irradiation increased the intratumoral temperature to nearly 70°C for 8min in GNS-MCs-treated mice. Tumor volumes decreased gradually and tumors were completely ablated in 6 out of 7 mice treated with GNS-MCs and laser irradiation by 17 days after treatment. ConclusionThis study demonstrates that ultrasound-guided PTT with theranostic GNS-MCs is a promising technique for in situ treatment of breast cancer.

Quantitative receptor-based imaging of tumor proliferation with the sigma-2 ligand [(18)F]ISO-1.

The sigma-2 receptor is expressed in higher density in proliferating (P) tumor cells versus quiescent (Q) tumor cells, thus providing an attractive target for imaging the proliferative status (i.e., P:Q ratio) of solid tumors. Here we evaluate the utility of the sigma-2 receptor ligand 2-(2-[18F]fluoroethoxy)-N-(4-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)butyl)-5-methyl-benzamide, [18F]ISO-1, in two different rodent models of breast cancer. In the first study, small animal Positron Emission Tomography (PET) imaging studies were conducted with [18F]ISO-1 and 18FDG in xenografts of mouse mammary tumor 66 and tracer uptake was correlated with the in vivo P:Q ratio determined by flow cytometric measures of BrdU-labeled tumor cells. The second model utilized a chemically-induced (N-methyl-N-nitrosourea [MNU]) model of rat mammary carcinoma to correlate measures of [18F]ISO-1 and FDG uptake with MR-based volumetric measures of tumor growth. In addition, [18F]ISO-1 and FDG were used to assess the response of MNU-induced tumors to bexarotene and Vorozole therapy. In the mouse mammary 66 tumors, a strong linear correlation was observed between the [18F]ISO-1 tumor: background ratio and the proliferative status (P:Q ratio) of the tumor (R = 0.87). Similarly, measures of [18F]ISO-1 uptake in MNU-induced tumors significantly correlated (R = 0.68, P<0.003) with changes in tumor volume between consecutive MR imaging sessions. Our data suggest that PET studies of [18F]ISO-1 provide a measure of both the proliferative status and tumor growth rate, which would be valuable in designing an appropriate treatment strategy.

Chronic Social Isolation Is Associated with Metabolic Gene Expression Changes Specific to Mammary Adipose Tissue

Chronic social isolation is linked to increased mammary tumor growth in rodent models of breast cancer. In the C3(1)/SV40 T-antigen FVB/N (TAg) mouse model of "triple-negative" breast cancer, the heightened stress response elicited by social isolation has been associated with increased expression of metabolic genes in the mammary gland before invasive tumors develop (i.e., during the in situ carcinoma stage). To further understand the mechanisms underlying how accelerated mammary tumor growth is associated with social isolation, we separated the mammary gland adipose tissue from adjacent ductal epithelial cells and analyzed individual cell types for changes in metabolic gene expression. Specifically, increased expression of the key metabolic genes Acaca, Hk2, and Acly was found in the adipocyte, rather than the epithelial fraction. Surprisingly, metabolic gene expression was not significantly increased in visceral adipose depots of socially isolated female mice. As expected, increased metabolic gene expression in the mammary adipocytes of socially isolated mice coincided with increased glucose metabolism, lipid synthesis, and leptin secretion from this adipose depot. Furthermore, application of media that had been cultured with isolated mouse mammary adipose tissue (conditioned media) resulted in increased proliferation of mammary cancer cells relative to group-housed-conditioned media. These results suggest that exposure to a chronic stressor (social isolation) results in specific metabolic reprogramming in mammary gland adipocytes that in turn contributes to increased proliferation of adjacent preinvasive malignant epithelial cells. Metabolites and/or tumor growth-promoting proteins secreted from adipose tissue could identify biomarkers and/or targets for preventive intervention in breast cancer.