Microbubble contrast agents used in conjunction with ultrasound are currently approved by the FDA and other global agencies as vascular agents for enhancing echocardiographic and hepatic imaging. They are also used in the research arena as molecular imaging agents for detection of angiogenesis, inflammation, and vulnerable plaque to name but a few. In addition, they have been translated to the clinic as ultrasound-mediated thrombolytic agents as well (i.e. Sonothrombolysis). Yet despite their utility, they are also restricted primarily to the vasculature due to their relatively large size (1 - 5 microns) and inability to extravasate into the tissue space. Recently, our laboratory has developed new phase-change contrast agents (PCCAs) that are comprised of a 100 nm, ultrasound-activatable, lipid-coated, perfluorocarbon-filled droplet designed for extravascular distribution followed by conversion to microbbubbles for ultrasound tissue imaging. These PCCAs consist of very low boiling liquid perfluorocarbons (decafluorobutane, bp = -1.1?C, octafluoropropane, bp = -36.7?C), stable at physiologic temperatures (37?C) and activatable into imagable microbubbles with low ultrasound energy outputs (FDA approved mechanical indices). Furthermore, conversion to microbubbles is acheivable using standard clinical ultrasound machines. Studies are ongoing with respect to their ability to extravasate into tissue and are the focus of a previou grant. Currently, we are measuring levels of PCCA extravasation as an indication of enhanced permeability and retention effect (EPR). In the focus of this new proposal, we wish to now study methodology for intracellular delivery of these PCCAs for activation inside the cellular milieu of breast cancer cells in vitro and in vivo for the purposes of selective breast tumor cell imaging. I addition, we wish to embark on in vitro studies to determine if intracellular cavitation of the resultant microbubbles can promote tumor cell destruction. As a medium for targeting, we will use the human epidermal growth factor receptor (hEGFR) as a biomarker for more precise delivery and intracellular uptake. Thus, the goal of this proposal is to formulate EGFR-targeted PCCAs for;1) highly selective intracellular imaging and;2) initiate monitoring for selective breast tumor cell destruction in vitro via ultrasound-mediated intracellular cavitation of the resultant microbubble. Successful accomplishment of these goals will poise our laboratory for designing the first ultrasound-mediated diagnostic and therapeutic agent for extravascular applications.

Public Health Relevance

Breast cancer in women is the second largest cause of cancer-related deaths. Despite alarming numbers, one key to reducing mortality is the early detection of disease before it becomes symptomatic. Recently, we have developed an ultrasound contrast agent that is comprised of a 100 nm, stabilized, liquid phase-change contrast agent (PCCA) that is small enough to diffuse into the tissue, target cancer cells and be activated into microbubbles for imaging with clinical ultrasound machines. Preliminary data in our laboratory now has shown that these PCCAs may also have the ability to be taken up by breast cancer cells and activated to microbubbles inside for more precise tumor cell imaging. We now wish to explore this concept of intracellular delivery and imaging for the early detection of breast cancer. Furthermore, it is the objective of this feasibility study to test our PCCAs as theranostic agents by imaging the resultant microbubbles and then bursting them to see if one can selectively damage the breast cancer cells from the inside.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (ZRG1-CMIP-W (09))
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Nordstrom, Robert J
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University of Arizona
Schools of Medicine
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Hadinger, Kyle P; Marshalek, Joseph P; Sheeran, Paul S et al. (2018) Optimization of Phase-Change Contrast Agents for Targeting MDA-MB-231 Breast Cancer Cells. Ultrasound Med Biol 44:2728-2738
Lin, Shengtao; Shah, Anant; Hernández-Gil, Javier et al. (2017) Optically and acoustically triggerable sub-micron phase-change contrast agents for enhanced photoacoustic and ultrasound imaging. Photoacoustics 6:26-36
Sheeran, Paul S; Matsuura, Naomi; Borden, Mark A et al. (2017) Methods of Generating Submicrometer Phase-Shift Perfluorocarbon Droplets for Applications in Medical Ultrasonography. IEEE Trans Ultrason Ferroelectr Freq Control 64:252-263
Marshalek, Joseph P; Sheeran, Paul S; Ingram, Pier et al. (2016) Intracellular delivery and ultrasonic activation of folate receptor-targeted phase-change contrast agents in breast cancer cells in vitro. J Control Release 243:69-77
Li, Sinan; Lin, Shengtao; Cheng, Yi et al. (2015) Quantifying activation of perfluorocarbon-based phase-change contrast agents using simultaneous acoustic and optical observation. Ultrasound Med Biol 41:1422-31