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Purpose: In this study, we investigate the precision and accuracy of mid‐to‐high frequency ultrasound imaging to assess non‐invasively cell death for incorporation of this method in pre‐clinical and clinical practice to characterize tumor response to radiotherapy.Method and Materials:Tumor xenografts (n=8) of head and neck cancer were exposed to radiationdoses of 2, 4 and 8Gy. Ultrasoundimages were collected with an ultrasoundscanner using frequencies of 15–35MHz before and 24 hours after exposure to radiation. Irradiated tumors exhibited large hyperechoic regions in ultrasoundsimages 24 hours after exposure to radiation that corresponded to areas of cell death in histology. The ultrasoundimages were registered with the histological images of the tumor slices taken at regular intervals. The tumor was contoured on histological slices and ultrasoundimages, the regions of cell death were contoured on histological slices and the hyperechoic regions were contoured on ultrasoundimages. Each set of contours was converted to a surface mesh. The volume and center of mass were calculated for each representation determined by a surface mesh. Results: The average difference between the relative (to histology) volume representations in histology and ultrasound were 10.7%±8.9% for tumor and 21.7%±12.2% for cell death. The average differences between the relative (to the maximum dimension of the tumor) center of mass of volume representations in histology and ultrasoundimages were 2.7%±2.0% for tumor and 15.5%±8.9 % for cell death. Conclusion: The method provides the correspondence between the volumes of cell death assessed from histology and from ultrasound imaging and can be used to assess early tumor response to radiotherapy. Part of the differences associated with cell death representation in histological and ultrasoundimages (21.7%) was caused by the differences in tumor representation (10.7%) in these images. The effect of these uncertainties is the subject of ongoing investigation.

High intensity focused ultrasound has considerable potential for the noninvasive treatment of localised disease. A detailed understanding of the kinetics of tissue coagulation is required to optimise ultrasonic parameters. In this presentation a theoretical model was used to examine the effects of temperature dependent ultrasonic attenuation and absorption on the transient tissue temperature distributions and lesion dimensions. A finite difference algorithm was used to solve numerically the nonlinear form of the bioheat transfer equation in cylindrical coordinates. The lesion dimensions were calculated based on the time-temperature distributions in tissue by using a thermal dose threshold to define the lesion boundaries. The results were compared to published experimental data in which the the location of maximal energy deposition during short duration high intensity focused ultrasound irradiation of in vitro tissue was examined. It was found that the theoretical model did not predict the size and shape of the experimental lesions. To correctly predict lesion size and shape much higher values of attenuation and absorption were required than can be accounted for by thermal coagulation of the tissue alone. The values used suggest that for intensities greater than 3030 W/cm2 the effective local attenuation/absorption in the focal region increased by a factor of 10–20. It is finally shown that temperature dependent tissue changes should be incorporated in thermal models to avoid underestimation of the induced temperature distributions during high intensity focused ultrasound therapy.

Temperature distributions measured during thermal therapy are a major prognostic factor of the efficacy and success of the procedure. Thermal models are used to predict the temperature elevation of tissues during heating. Theoretical work has shown that blood flow through large blood vessels plays an important role in determining temperature profiles of heated tissues. In this paper, an experimental investigation of the effects of large vessels on the temperature distribution of heated tissue is performed. The blood flow dependence of steady state and transient temperature profiles created by a cylindrical conductive heat source and an ultrasound transducer were examined using a fixed porcine kidney as a flow model. In the transient experiments, a 20 s pulse of hot water, 30 degrees C above ambient, heated the tissues. Temperatures were measured at selected locations in steps of 0.1 mm. It was observed that vessels could either heat or cool tissues depending on the orientation of the vascular geometry with respect to the heat source and that these effects are a function of flow rate through the vessels. Temperature gradients of 6 degrees C mm(-1) close to large vessels were routinely measured. Furthermore, it was observed that the temperature gradients caused by large vessels depended on whether the heating source was highly localized (i.e. a hot needle) or more distributed (i.e. external ultrasound). The gradients measured near large vessels during localized heating were between two and three times greater than the gradients measured during ultrasound heating at the same location, for comparable flows. Moreover, these gradients were more sensitive to flow variations for the localized needle heating. X-ray computed tomography data of the kidney vasculature were in good spatial agreement with the locations of all of the temperature variations measured. The three dimensional vessel path observed could account for the complex features of the temperature profiles. The flow dependences of the transient temperature profiles near large vessels during the pulsed experiments were consistent with the temperature distributions measured in the steady state experiments and provided unique insights into the process of convective heat transfer in tissues. Finally, it was shown that even for very short treatment times (3-20 s), large vessels had significant effects on the tissue temperature distributions.

Electrical impedance spectroscopy (EIS) and high-frequency ultrasound (HFU) have been evaluated in various in vivo and in vitro models as potential methods to monitor biological changes induced by photodynamic therapy (PDT). EIS was assessed in tumor-bearing rat leg in vivo, in multicell tumor spheroids in vitro, and in normal rat liver tissue in vivo. HFU measurements, both imaging and backscatter frequency scanning, were tested in normal rat brain and skin treated in vivo. Marked changes in the EIS spectra were seen in all 3 models following PDT, depending on the photosensitizer and treatment parameters. With HFU, significant increases in echogenicity of the PDT-treated tissues were observed, with evidence of dose dependency and correlation with apoptotic cell death in vivo. While the results are encouraging for both modalities, a number of technical problems remain, particularly in the case of EIS, before these methods can be used reliably in mechanistic and clinical PDT applications.

Changes in magnetic resonance (MR) signals during interstitial microwave heating are reported, and correlated with simultaneously acquired temperature readings from three fiber-optic probes implanted in a polyacrylamide gel phantom. The heating by a MR-compatible microwave antenna did not interfere with simultaneous MR image data acquisition. MR phase-difference images were obtained using a fast two-dimensional-gradient echo sequence. From these images the temperature-sensitive resonant frequency of the 1H nuclei was found to decrease approximately by 0.008 ppm/ degree C. The method and results presented here demonstrate that noninvasive MR-temperature imaging can be performed simultaneously with interstitial microwave thermal treatment.

Purpose: Currently, no clinical imaging modality is used routinely to assess tumor response to cancer therapies within hours to days of the delivery of treatment. Here, the authors demonstrate the efficacy of ultrasound at a clinically relevant frequency to quantitatively detect changes in tumors in response to cancer therapies using preclinical mouse models. Methods: Conventional low-frequency and corresponding high-frequency ultrasound (ranging from 4 to 28 MHz) were used along with quantitative spectroscopic and signal envelope statistical analyses on data obtained from xenograft tumors treated with chemotherapy, x-ray radiation, as well as a novel vascular targeting microbubble therapy. Results: Ultrasound-based spectroscopic biomarkers indicated significant changes in cell-death associated parameters in responsive tumors. Specifically changes in the midband fit, spectral slope, and 0-MHz intercept biomarkers were investigated for different types of treatment and demonstrated cell-death related changes. The midband fit and 0-MHz intercept biomarker derived from low-frequency data demonstrated increases ranging approximately from 0 to 6 dBr and 0 to 8 dBr, respectively, depending on treatments administrated. These data paralleled results observed for high-frequency ultrasound data. Statistical analysis of ultrasound signal envelope was performed as an alternative method to obtain histogram-based biomarkers and provided confirmatory results. Histological analysis of tumor specimens indicated up to 61% cell death present in the tumors depending on treatments administered, consistent with quantitative ultrasound findings indicating cell death. Ultrasound-based spectroscopic biomarkers demonstrated a good correlation with histological morphological findings indicative of cell death (r2 = 0.71, 0.82; p < 0.001). Conclusions: In summary, the results provide preclinical evidence, for the first time, that quantitative ultrasound used at a clinically relevant frequency, in addition to high-frequency ultrasound, can detect tissue changes associated with cell deathin vivo in response to cancer treatments.

We have previously measured the speed of sound in individual cells and cell pellets. Spheroids can be used to more accurately model non-vascularized tumors due to their three-dimensional growth pattern, cell-cell interaction, disorganized growth, and development of a necrotic core when grown to sufficiently large sizes. To examine cell death in spheroids due to necrosis or chemotherapy, quantitative ultrasound methods were used on the ultrasound backscatter power spectrum throughout the spheroid. MCF7 spheroids were probed at 55 MHz using a VEVO770 high frequency ultrasound machine, and at 80 and a scanning acoustic microscope. Changes in spheroid structure as the necrotic core develops were recorded and analysed. This work furthers our understanding of non-invasively identifying the viability of cancerous tumours.

A clinical study was undertaken investigating the efficacy of ultrasound to quantify cell death in tumor responses with cancer treatment. Patients (n = 25) with locally advanced breast cancer received anthracyline and taxane-based chemotherapy treatments over four to six months. The majority of patients went on to have a modified radical mastectomy and correlative whole mount histopathology. Data collection was carried out using an Ultrasonix-RP and an L15-5 6 cm transducer pulsed at 10 MHz with RF data collected five times during neoadjuvant chemotherapy. Data indicated increases of approximately 9 dBr (+/−1.67) maximally in ultrasound backscatter in patients who clinically responded to treatment. Patients assessed as responding poorly demonstrated significantly lower increases (2.3 +/− 1.7 dBr). Increases in 0-MHz intercept followed similar trends while increases in spectral slope were observed locally from tumor regions demonstrating increases in tissue echogenicity. This study demonstrates the potential of ultrasound to quantify changes in tumors in response to cancer treatment administration in a clinical setting. The results indicate that such responses can be detected early during a course of chemotherapy and should permit ineffective treatments to be changed to more efficacious ones potentially leading to improved treatment outcomes.

Large blood vessels can produce steep temperature gradients in heated tissues leading to inadequate tissue temperatures during hyperthermia. This paper utilizes a finite difference scheme to solve the basic equations of heat transfer and fluid flow to model blood vessel cooling. Unlike previous formulations, heat transfer coefficients were not used to calculate heat transfer to large blood vessels. Instead, the conservation form of the finite difference equations implicitly modelled this process. Temperature profiles of heated tissues near thermally significant vessels were calculated. Microvascular heat transfer was modelled either as an effective conductivity or a heat sink. An increase in perfusion in both microvascular models results in a reduction of the cooling effects of large vessels. For equivalent perfusion values, the effective conductivity model predicted more effective heating of the blood and adjacent tissue. Furthermore, it was found that optimal vessel heating strategies depend on the microvascular heat transfer model adopted; localized deposition of heat near vessels could produce higher temperature profiles when microvascular heat transfer was modelled according to the bioheat transfer equation (BHTE) but not the effective thermal conductivity equation (ETCE). Reduction of the blood flow through thermally significant vessels was found to be the most effective way of reducing localized cooling.

Red blood cell (RBC) aggregation and oxygenation are important markers for a variety of blood disorders. No current technique is capable of simultaneously measuring aggregation/oxygenation levels noninvasively. We propose using photoacoustic ultrasound spectroscopy (PAUS) for assessing both phenomena. This technique relies on frequency-domain analysis of the PA signals by extracting parameters such as the ultrasound spectral slope and the midband fit. To investigate the effect of hematocrit, aggregation, and oxygenation levels on PAUS parameters, a Monte Carlo-based theoretical model and an experimental protocol using porcine RBCs were developed. The samples were illuminated at 750 and 1064 nm and changes in the PAUS parameters were compared to the oxygen-dependent optical absorption coefficients to assess the oxygenation level. Good agreement between the theoretical and experimental spectral parameters was obtained for the spectral slope of the nonaggregated spectra ( ~ 0.3 dB / MHz). The experimental midband fit increased by ~ 5 dB for the largest aggregate size. Based on the analysis of the PA signals, the oxygen saturation level of the most aggregated sample was > 20% greater than the nonaggregated sample. The results provide a framework for using PA signals' spectroscopic parameters for monitoring the aggregation and oxygenation levels of RBCs.

A theoretical model investigating the dependence of optoacoustic (OA) signal on blood oxygen saturation (SO2) is discussed. The derivations for the nonbandlimited and bandlimited OA signals from many red blood cells (RBCs) are presented. The OA field generated by many RBCs was obtained by summing the OA field emitted by each RBC approximated as a fluid sphere. A Monte Carlo technique was employed generating the spatial organizations of RBCs in two-dimensional. The RBCs were assumed to have the same SO2 level in a simulated configuration. The fractional number of oxyhemoglobin molecules, confined in a cell, determined the cellular SO2 and also defined the blood SO2. For the nonbandlimited case, the OA signal amplitude decreased and increased linearly with blood SO2 when illuminated by 700 and 1000 nm radiations, respectively. The power spectra exhibited similar trends over the entire frequency range (MHz to GHz). For the bandlimited case, three acoustic receivers with 2, 10, and 50 MHz as the center frequencies were considered. The linear variations of the OA amplitude with blood SO2 were also observed for each receiver at those laser sources. The good agreement between simulated and published experimental results validates the model qualitatively. © 2012 Society of Photo-Optical

Changes in cell structure as tumours respond to cancer therapies can be detected using high frequency ultrasound (20-60 MHz). However, it is unknown how single cell variations influence the ultrasound signal as these ultrasound frequencies cannot resolve individual cells. Acoustic microscopy uses ultra-high frequency ultrasound (over 100 MHz) to quantify the structural and mechanical properties of single cells. The backscattered echoes from the cell membrane and substrate were used to determine the thickness, sound speed, density, bulk modulus and attenuation of breast cancer cells undergoing various biological processes, such as mitosis (cell division) and apoptosis (programmed death). Moreover, measurements were made to examine differences between malignant and benign breast tumour cells. Statistically significant changes in the thickness, sound speed and bulk modulus (2%), and attenuation (60%) between malignant and benign cells, and thickness (70%) and attenuation (21%) between cells undergoing mitosi...

Increasing the nonlinear response of ultrasound contrast agents (UCAs) in response to two consecutive pulses in amplitude modulation (AM) techniques can significantly increase the contrast to tissue ratio (CTR). It has been recently shown that one way to increase the nonlinearity is to take advantage of the buckling behavior of lipid shell bubbles. However, this method is limited to frequencies below the resonance of the bubble. To achieve enhanced nonlinearity at high frequencies, the oscillations of the UCAs should be optimized. In this work, the Hoff model was solved for a wide range of the parameters of the UCAs. Results showed that when the bubbles are sonicated with a frequency slightly less than the integer multiples of its resonance frequency (e.g., 2.7 and 3.6), and above a pressure threshold (PT), there is a significant increase in the UCA stable radial oscillations, which corresponds to a significant backscattered signal enhancement. If the sonication frequency of the AM technique is optimized according to the mentioned frequencies, and the amplitudes of the AM first and second pulses are smaller and greater than this PT, the CTR of the AM technique may be enhanced significantly.

Conventional methods of subharmonic (SH) imaging of ultrasound contrast agents (UCAs) employ a frequency which is approximately twice the resonance frequency of the UCA (the SH resonance frequency). However, this approach may be limited by a low signal to noise ratio. To address this problem, the Hoff model was solved for several UCAs and the SH resonance frequencies of the bubble were calculated for different applied pressures (PAs). By increasing the PA above the pressure threshold for SH generation, the SH resonance frequency shifts toward lower values. Calculation of the scattered pressure in the regime of stable oscillations show that, if the bubble is sonicated with its pressure dependent SH resonance frequencies, the backscattered signal can be significantly stronger than when sonicating with the conventional SH resonance frequency (although the initiation of the SH occurs at a higher PA). The enhancement of the SH and ultraharmonic amplitudes can be up to 7 and 14 dB, respectively. This enhancemen...

The aim of many cancer therapies is to induce cell death within a target tumor. A substantial body of research using in vitro and in vivo models has demonstrated that cell death can be detected via quantitative ultrasound techniques. This study investigates for the first time the potential to quantify tumor responses to therapy in patients, using spectral and signal envelope statistics analysis of ultrasound data. A clinical study was undertaken investigating the efficacy of ultrasound to quantify cell death in tumor responses with cancer treatment. Patients (n=20) with locally advanced breast cancer received anthracyline and taxane‐based chemotherapy treatments. Data collection consisted of acquiring tumor images and radiofrequency data prior to treatment onset and at 4 times during treatment (weeks 1, 4, and 8, and preoperatively). Data indicate increases of approximately 9 dBr (±1.67) in ultrasound backscatter in patients who responded to treatment. Patients assessed as responding poorly demonstrated s...

In this study, we investigate for the first time the potential to quantify tumor responses to therapy in vivo, using spectral and signal envelope statistics analysis of conventional frequency ultrasound data. Tumors were grown in SCID mice using a human prostate cell line (PC‐3) and treated in three groups consisting of combinations of ultrasonically stimulated micro‐bubble anti‐angiogenic treatment and 8 Gy doses of single fraction 100 kVp X‐ray radiation. Data collection consisted of tumor images in addition to radiofrequency data prior to treatment administration and at 24 h following. Analysis of the normalized power spectra of 10‐MHz data yielded an increase in mid‐band fit of approximately 3.8±0.7, 4.6±0.8, and 5.8±0.7 dBr compared to data acquired before treatment. Spectral slopes were relatively invariant. However, the 0‐MHz intercept followed the trend of the mid‐band fit. Signal envelope changes were compatible with observations made from spectral data and linked to correlative increases in cell...

Several studies have explored the potential of optoacoustic imaging for monitoring thermal therapies, yet the origin of the contrast in the images is not well understood. A technique is required to measure the changes in the optical and thermomechanical properties of tissues upon coagulation to better understand this contrast. An interferomet- ric method is presented for measuring simultaneously the optical and thermomechanical properties of native and coagulated ex-vivo bovine tissue samples based on analysis of the surface displacement of irra- diated samples. Surface displacements are measured after irradiation by short laser pulses at 750 nm. A 51% decrease in the optical at- tenuation depth is observed for coagulated liver samples compared to native samples. No significant differences in the Gr¨ uneisen coefficient are measured in the native and coagulated tissue samples. A mean value of 0.12 for the Gr¨ uneisen coefficient is measured for both native and coagulated liver tissues. The displacement profiles exhibit consistent differences between the two tissue types. To assess the changes in the sample mechanical properties, the experimental data also are compared to numerical solutions of the equation for thermoelastic deformation. The results demonstrate that differences in the tissue expansion dy- namics arise from higher values of elastic modulus for coagulated liver samples compared to native ones. C 2010 Society of Photo-Optical Instrumentation Engineers. (DOI: 10.1117/1.3517455)

We have discovered that high-frequency ultrasound imaging, or ultrasound biomicroscopy, can be used to detect apoptosis in a number of experimental systems. We have shown that such detection with 30-40 MHz ultrasound is possible using cells in an in vitro system (AML-3 leukemia cells) made to undergo apoptosis in response to treatment with a variety of cancer killing chemotherapeutic drugs. We have shown that ultrasound biomicroscopy can also detect programmed cell death in tissues made to undergo apoptosis in response to photodynamic therapy, currently an experimental cancer treating regimen. Lastly, we have shown that this ultrasound imaging approach works in vivo, using living animals where apoptosis has been induced similarly using photodynamic therapy. Specifically, apoptotic cells and regions of apoptosis in tissues exhibit up to a 36-fold increase in ultrasound backscatter intensity permitting this type of cell death to be readily discriminated from surrounding viable tissue.

High‐frequency ultrasound is a novel method to detect cell death based on changes in cell morphology that cause alterations in the acoustic properties of cell ensembles in tissues. In previous work, our group has suggested that for cells with a nucleus to cell volume ratio of 0.33, the backscatter response from single cells was best modeled as a fluid sphere. However, for cells with a larger nucleus, neither the fluid nor the elastic sphere models showed a good agreement. We hypothesize that cells with a large nucleus ratio (> 0.33) may be modeled as an elastic sphere surrounded by a fluid shell. In this work, the backscatter response from non‐nucleated cells (Strongylocentrotus purpuratus oocytes) was measured Experimental RF lines were acquired using the VisualSonics VS40B ultrasound biomicroscope using 20 and 40 MHz transducers. A finite‐element model of wave propagation was developed to solve high‐frequency ultrasound scattering from spheres. A very good agreement was found suggesting that the cytopla...

The goal of ultrasonic tissue characterization is to extract information over and above the information conventionally displayed on imaging instruments which is typically the backscatter strength (B‐mode image). The driving hypothesis of our work is that as the ultrasound wavelength approaches the size of a cell, the backscatter characteristics become more sensitive to the cells' physical attributes (size, structure, and composition) and changes in the cell structure due to therapeutic intervention. A better understanding of the interaction of acoustic waves with cells is required to interpret the backscatter data. An overview of our efforts to model scattering from individual cells, cell ensembles, and in‐vivo cancer will be presented. Theoretical results will be presented comparing scattering from individual cells with experimental data from a variety of cells in suspension, cell ensembles with pellets (compact aggregates of cells) and in‐vivo tumors. It will be shown that for the tumors we have studied...

A finite‐element model of wave propagation using COMSOL Multiphysics (COMSOL Inc., Burlington, MA) was developed to solve the problem of high‐frequency ultrasound (10–70 MHz) scattering from spheres. This model is being used to predict ultrasound backscatter from cells for ultrasound tissue characterization. In this work, backscatter from an elastic (polystyrene) microsphere was used to validate the computational model against experimental results. Agreement between experimental results and finite‐element solutions were found in the frequency domain solutions. Moreover, conversion of the theoretical solutions to time‐domain backscatter data showed good agreement with experimental time domain measurements. To develop a better understanding of the interactions of the incident pressure field with the scattering object, oscillations of the elastic microsphere at various resonance frequencies were also investigated. A systematic relationship between the resonance frequency and its corresponding surface mode was found. An ultrasound scattering model by a single cell is also presented. The model treats the cell as an elastic sphere (nucleus) surrounded by a fluid shell (cytoplasm). Comparison of the theoretical backscatter predicted by the model and experimental measurements for acute Myeloid Leukemia (AML) cell is also shown. Finally, the implications of these results on ultrasound tissue characterization techniques are discussed.

High‐frequency ultrasound (HFU) imaging (in the range of 20–60 MHz) has been recently enabled by advances in transducer technology and electronics. These high frequencies afford greater image resolution, on the order of 40 microns, at the expense of penetration depth, limited to a few centimeters. The associated ultrasound wavelengths of HFU are 25–75 microns, on the order of the size of cells. Therefore, it is hypothesized that at these frequencies ultrasound tissue characterization will provide more meaningful information about the physical characteristics of cells and any changes that occur in their structure during cancer treatment. The late Dr. Frederic Lizzi pioneered ultrasound tissue characterization and the seminal papers he published provided the impetus for this work. Here, backscatter from individual cells and tightly packed cell aggregates of the same cells (emulating tumors) are examined and compared to theoretical models of ultrasound backscatter. Recent work with a series of cells lines with different sizes and physical characteristics will be presented. It will be shown that for in vitro and in vivo experiments, the backscatter signal characteristics best correlate with nuclear size.

A non-invasive method of monitoring apoptosis in cell culture, ex-vivo tissues and in-vivo tissues using high frequency ultrasound imaging is provided. The method includes the steps of: 1) imaging a selected site of the cell culture or tissues using high frequency (above 20 MHz) ultrasound imaging (before image); 2) exposing the selected site to an apoptosis-inducing stress; 3) imaging the selected site or a portion thereof, using ultrasound imaging at subsequent timed intervals (after image(s)); 4) measuring the signal amplitude of a region of interest of the selected site in the before and after images; 5) comparing the signal amplitude measurements for the regions of interest in the before and after images and determining whether the after image regions exhibit an increase in amplitude as compared to the before image regions which is an indication that apoptosis has begun; and 6) measuring the change in the frequency spectrum of the radiofrequency ultrasound backscatter signal in the region of interest in the before and after images and confirming that apoptosis has begun when the slope of the frequency spectrum has increased.

Three-dimensional quantitative ultrasound spectroscopic imaging of prostate was investigated clinically for the noninvasive detection and extent characterization of disease in cancer patients and compared to whole-mount, whole-gland histopathology of radical prostatectomy specimens. Fifteen patients with prostate cancer underwent a volumetric transrectal ultrasound scan before radical prostatectomy. Conventional-frequency (~ 5 MHz) ultrasound images and radiofrequency data were collected from patients. Normalized power spectra were used as the basis of quantitative ultrasound spectroscopy. Specifically, color-coded parametric maps of 0-MHz intercept, midband fit, and spectral slope were computed and used to characterize prostate tissue in ultrasound images. Areas of cancer were identified in whole-mount histopathology specimens, and disease extent was correlated to that estimated from quantitative ultrasound parametric images. Midband fit and 0-MHz intercept parameters were found to be best associated with the presence of disease as located on histopathology whole-mount sections. Obtained results indicated a correlation between disease extent estimated noninvasively based on midband fit parametric images and that identified histopathologically on prostatectomy specimens, with an r2 value of 0.71 (P < .0001). The 0-MHz intercept parameter demonstrated a lower level of correlation with histopathology. Spectral slope parametric maps offered no discrimination of disease. Multiple regression analysis produced a hybrid disease characterization model (r2 = 0.764, P < .05), implying that the midband fit biomarker had the greatest correlation with the histopathologic extent of disease. This work demonstrates that quantitative ultrasound spectroscopic imaging can be used for detecting prostate cancer and characterizing disease extent noninvasively, with corresponding gross three-dimensional histopathologic correlation.