Acoustic Angiography: A New Imaging Platform for High Resolution Mapping of Microvasculature and Tumor Assessment

Statistically, one in four Americans will die from cancer. Many new tumor detection and therapeutic approaches have improved patient outcomes, but cancer continues to run rampant in our country; it claimed the lives of 1.6 million Americans in 2012. To put this number of annual deaths in perspective, it is over 500 times the number of people who died in the horrific attacks on September 11, 2001. This dissertation does not offer either an antidote to the disease, nor a detection mechanism appropriate for all tumor types. It does, however, present the description and characterization of a novel dual-frequency ultrasound imaging transducer, capable of operating in a new imaging mode we call `acoustic angiography.' These images offer high resolution and high contrast 3D depictions of the microvasculature; herein we demonstrate its cancer assessment utility by way of multiple imaging studies. Throughout this dissertation, image data from both healthy and diseased tissues are presented. Additionally, acoustic assessments of vasculature within an ex vivo biomatrix scaffold model (a platform for creating of artificial organs) are presented. A vessel mapping algorithm, originally developed for human magnetic resonance angiography images, has been implemented in both in vivo and ex vivo tissue volumes. A novel microvessel phantom generation technique is presented, which allows ground-truth coordinates for vascular networks to be defined and imaged. Finally, the ultrasound pulsing technique, radiation force, was used as a method to improve the diagnostic sensitivity of ultrasound to malignant tumors. Together, the results of these studies suggest that the imaging approach, acoustic angiography, enabled by our new dual-frequency ultrasound transducer, could eventually be used to detect and monitor tumors in a clinical imaging context. This dissertation supports the following three hypotheses: 1) A prototype dual-frequency ultrasound transducer can be used to depict in vivo microvasculature, 2) These microvascular images can be quantitatively assessed as a means to characterize the presence of a tumor, and evaluate tumor response to therapy, and 3) Radiation force can be used as a method to improve ultrasonic diagnostic sensitivity to the presence of a tumor.Doctor of Philosoph

Advances in Molecular Imaging with Ultrasound

Ultrasound imaging has long demonstrated utility in the study and measurement of anatomic features and noninvasive observation of blood flow. Within the last decade, advances in molecular biology and contrast agents have allowed researchers to use ultrasound to detect changes in the expression of molecular markers on the vascular endothelium and other intravascular targets. This new technology, referred to as ultrasonic molecular imaging, is still in its infancy. However, in preclinical studies, ultrasonic molecular imaging has shown promise in assessing angiogenesis, inflammation, and thrombus. In this review, we discuss recent advances in microbubble-type contrast agent development, ultrasound technology, and signal processing strategies that have the potential to substantially improve the capabilities and utility of ultrasonic molecular imaging

Early Assessment of Tumor Response to Radiation Therapy using High-Resolution Quantitative Microvascular Ultrasound Imaging

Measuring changes in tumor volume using anatomical imaging weeks to months post radiation therapy (RT) is currently the clinical standard for indicating treatment response to RT. For patients whose tumors do not respond successfully to treatment, this approach is suboptimal as timely modification of the treatment approach may lead to better clinical outcomes. We propose to use tumor microvasculature as a biomarker for early assessment of tumor response to RT. Acoustic angiography is a novel contrast ultrasound imaging technique that enables high-resolution microvascular imaging and has been shown to detect changes in microvascular structure due to cancer growth. Data suggest that acoustic angiography can detect longitudinal changes in the tumor microvascular environment that correlate with RT response

Mapping Microvasculature with Acoustic Angiography Yields Quantifiable Differences between Healthy and Tumor-bearing Tissue Volumes in a Rodent Model

To determine if the morphologies of microvessels could be extracted from contrast material–enhanced acoustic angiographic ultrasonographic (US) images and used as a quantitative basis for distinguishing healthy from diseased tissue

Quantitative Volumetric Perfusion Mapping of the Microvasculature Using Contrast Ultrasound

Contrast-enhanced ultrasound imaging has demonstrated significant potential as a noninvasive technology for monitoring blood flow in the microvasculature. With the application of nondestructive contrast imaging pulse sequences combined with a clearance-refill approach, it is possible to create quantitative time-to-refill maps of tissue correlating to blood perfusion rate. One limitation to standard two-dimensional (2D) perfusion imaging is that the narrow elevational beamwidth of 1- or 1.5-D ultrasound transducers provides information in only a single slice of tissue, and thus it is difficult to image exactly the same plane from study to study. We hypothesize that inhomogeneity in vascularization, such as that common in many types of tumors, makes serial perfusion estimates inconsistent unless the same region can be imaged repeatedly. Our objective was to evaluate error in 2D quantitative perfusion estimation in an in vivo sample volume because of differences in transducer positioning. To mitigate observed errors due to imaging plane misalignment, we propose and demonstrate the application of quantitative 3-dimensional (3D) perfusion imaging. We also evaluate the effect of contrast agent concentration and infusion rate on perfusion estimates. Contrast-enhanced destruction-reperfusion imaging was performed using parametric mapping of refill times and custom software for image alignment to compensate for tissue motion. Imaging was performed in rats using a Siemens Sequoia 512 imaging system with a 15L8 transducer. A custom 3D perfusion mapping system was designed by incorporating a computer-controlled positioning system to move the transducer in the elevational direction, and the Sequoia was interfaced to the motion system for timing of the destruction-reperfusion sequence and data acquisition. Perfusion estimates were acquired from rat kidneys as a function of imaging plane and in response to the vasoactive drug dopamine. Our results indicate that perfusion estimates generated by 2D imaging in the rat kidney have mean standard deviations on the order of 10%, and as high as 22%, because of differences in initial transducer position. This difference was larger than changes in kidney perfusion induced by dopamine. With application of 3D perfusion mapping, repeatability in perfusion estimated in the kidney is reduced to a standard deviation of less than 3%, despite random initial transducer positioning. Varying contrast agent administration rate was also observed to bias measured perfusion time, especially at low concentrations; however, we observed that contrast administration rates between 2.7 × 108 and 3.9 × 108 bubbles/min provided results that were consistent within 3% for the contrast agent type evaluated. Three-dimensional perfusion imaging allows a significant reduction in the error caused by transducer positioning, and significantly improves the reliability of quantitative perfusion time estimates in a rat kidney model. When performing perfusion imaging, it is important to use appropriate and consistent contrast agent infusion rates to avoid bias

A Pilot Study to Assess Markers of Renal Damage in the Rodent Kidney After Exposure to 7 MHz Ultrasound Pulse Sequences Designed to Cause Microbubble Translation and Disruption

Acoustic radiation force has been proposed as a mechanism to enhance microbubble concentration for therapeutic and molecular imaging applications. It is hypothesized that once microbubbles are localized, bursting them with acoustic pressure could result in local drug delivery. It is known that low-frequency, high-amplitude acoustic energy combined with cavitation nuclei can result in bioeffects. However, little is known about the bioeffects potential of acoustic parameters involved in radiation-force and microbubble destruction pulse sequences applied at higher frequencies. In this pilot study, rat kidneys are exposed to high-duty cycle, low-amplitude pulse sequences known to cause substantial bubble translation due to radiation force, as well as high-amplitude short pulse sequences known to cause microbubble destruction. Both studies are performed at 7 MHz on a clinical ultrasound system, and implemented in 3-D for entire kidney exposure. Analysis of biomarkers of renal injury and renal histopathology indicate that there was no significant renal damage due to these ultrasound parameters in conjunction with microbubbles within the study group

Improving Sensitivity in Ultrasound Molecular Imaging by Tailoring Contrast Agent Size Distribution: In Vivo Studies

Molecular imaging with ultrasound relies on microbubble contrast agents (MCAs) selectively adhering to a ligand-specific target. Prior studies have shown that only small quantities of microbubbles are retained at their target sites, therefore, enhancing contrast sensitivity to low concentrations of microbubbles is essential to improve molecular imaging techniques. In order to assess the effect of MCA diameter on imaging sensitivity, perfusion and molecular imaging studies were performed with microbubbles of varying size distributions. To assess signal improvement and MCA circulation time as a function of size and concentration, blood perfusion was imaged in rat kidneys using nontargeted size-sorted MCAs with a Siemens Sequoia ultrasound system (Siemans, Mountain View, CA) in cadence pulse sequencing (CPS) mode. Molecular imaging sensitivity improvements were studied with size-sorted αvβ3-targeted bubbles in both fibrosarcoma and R3230 rat tumor models. In perfusion imaging studies, video intensity and contrast persistence was ≈8 times and ≈3 times greater respectively, for “sorted 3-micron” MCAs (diameter, 3.3 ± 1.95 μm) when compared to “unsorted” MCAs (diameter, 0.9 ± 0.45 μm) at low concentrations. In targeted experiments, application of sorted 3-micron MCAs resulted in a ≈20 times video intensity increase over unsorted populations. Tailoring size-distributions results in substantial imaging sensitivity improvement over unsorted populations, which is essential in maximizing sensitivity to small numbers of MCAs for molecular imaging

3-D Microvessel-Mimicking Ultrasound Phantoms Produced With a Scanning Motion System

Ultrasound techniques are currently being developed which can assess the vascularization of tissue as a marker for therapeutic response. Some of these ultrasound imaging techniques seek to extract quantitative features about vessel networks, while high-frequency imaging also allows individual vessels to be resolved. The development of these new techniques, and subsequent imaging analysis strategies, necessitates an understanding of their sensitivities to vessel and vessel network structural abnormalities. Constructing in-vitro flow phantoms for this purpose can be prohibitively challenging, as simulating precise flow environments with non-trivial structures is often impossible using conventional methods of construction for flow phantoms. Presented in this manuscript is a method to create predefined structures with < 10 μm precision using a three-axis motion system. The application of this technique is demonstrated for the creation of individual vessel and vessel networks, which can easily be made to simulate the development of structural abnormalities typical of diseased vasculature in-vivo. Additionally, beyond facilitating the creation of phantoms which would be otherwise very challenging to construct, the method presented herein enables one to precisely simulate very slow blood flow, respiration artifacts, and to measure imaging resolution

Acoustic Angiography: A New Imaging Modality for Assessing Microvasculature Architecture

The purpose of this paper is to provide the biomedical imaging community with details of a new high resolution contrast imaging approach referred to as “acoustic angiography.” Through the use of dual-frequency ultrasound transducer technology, images acquired with this approach possess both high resolution and a high contrast-to-tissue ratio, which enables the visualization of microvascular architecture without significant contribution from background tissues. Additionally, volumetric vessel-tissue integration can be visualized by using b-mode overlays acquired with the same probe. We present a brief technical overview of how the images are acquired, followed by several examples of images of both healthy and diseased tissue volumes. 3D images from alternate modalities often used in preclinical imaging, contrast-enhanced micro-CT and photoacoustics, are also included to provide a perspective on how acoustic angiography has qualitatively similar capabilities to these other techniques. These preliminary images provide visually compelling evidence to suggest that acoustic angiography may serve as a powerful new tool in preclinical and future clinical imaging

An In Vivo Validation of the Application of Acoustic Radiation Force to Enhance the Diagnostic Utility of Molecular Imaging Using 3-D Ultrasound

For over a decade, the application of acoustic radiation force (ARF) has been proposed as a mechanism to increase ultrasonic molecular imaging (MI) sensitivity in vivo. Presented herein is the first noninvasive in vivo validation of ARF-enhanced MI with an unmodified clinical system. First, an in vitro optical-acoustical setup was used to optimize system parameters and ensure sufficient microbubble translation when exposed to ARF. 3D ARF-enhanced MI was then performed on 7 rat fibrosarcoma tumors using microbubbles targeted to αvβ3 and non-targeted microbubbles. Low-amplitude (< 25 kPa) 3D ARF pulse sequences were tested and compared to passive targeting studies in the same animal. Our results demonstrate that a 78% increase in image intensity from targeted microbubbles can be achieved when using ARF relative to the passive targeting studies. Furthermore, ARF did not significantly increase image contrast when applied to non-targeted agents, suggesting that ARF did not increase non-specific adhesion