Jung Woo Choe

Ultrasound-guided delivery of microRNA loaded nanoparticles into cancer

Ultrasound induced microbubble cavitation can cause enhanced permeability across natural barriers of tumors such as vessel walls or cellular membranes, allowing for enhanced therapeutic delivery into the target tissues. While enhanced delivery of small (<1nm) molecules has been shown at acoustic pressures below 1MPa both in vitro and in vivo, the delivery efficiency of larger (>100nm) therapeutic carriers into cancer remains unclear and may require a higher pressure for sufficient delivery. Enhanced delivery of larger therapeutic carriers such as FDA approved pegylated poly(lactic-co-glycolic acid) nanoparticles (PLGA-PEG-NP) has significant clinical value because these nanoparticles have been shown to protect encapsulated drugs from degradation in the blood circulation and allow for slow and prolonged release of encapsulated drugs at the target location. In this study, various acoustic parameters were investigated to facilitate the successful delivery of two nanocarriers, a fluorescent semiconducting polymer model drug nanoparticle as well as PLGA-PEG-NP into human colon cancer xenografts in mice. We first measured the cavitation dose produced by various acoustic parameters (pressure, pulse length, and pulse repetition frequency) and microbubble concentration in a tissue mimicking phantom. Next, in vivo studies were performed to evaluate the penetration depth of nanocarriers using various acoustic pressures, ranging between 1.7 and 6.9MPa. Finally, a therapeutic microRNA, miR-122, was loaded into PLGA-PEG-NP and the amount of delivered miR-122 was assessed using quantitative RT-PCR. Our results show that acoustic pressures had the strongest effect on cavitation. An increase of the pressure from 0.8 to 6.9MPa resulted in a nearly 50-fold increase in cavitation in phantom experiments. In vivo, as the pressures increased from 1.7 to 6.9MPa, the amount of nanoparticles deposited in cancer xenografts was increased from 4- to 14-fold, and the median penetration depth of extravasated nanoparticles was increased from 1.3-fold to 3-fold, compared to control conditions without ultrasound, as examined on 3D confocal microscopy. When delivering miR-122 loaded PLGA-PEG-NP using optimal acoustic settings with minimum tissue damage, miR-122 delivery into tumors with ultrasound and microbubbles was 7.9-fold higher compared to treatment without ultrasound. This study demonstrates that ultrasound induced microbubble cavitation can be a useful tool for delivery of therapeutic miR loaded nanocarriers into cancer in vivo.

Integrated Circuits for Volumetric Ultrasound Imaging With 2-D CMUT Arrays

Real-time volumetric ultrasound imaging systems require transmit and receive circuitry to generate ultrasound beams and process received echo signals. The complexity of building such a system is high due to requirement of the front-end electronics needing to be very close to the transducer. A large number of elements also need to be interfaced to the back-end system and image processing of a large dataset could affect the imaging volume rate. In this work, we present a 3-D imaging system using capacitive micromachined ultrasonic transducer (CMUT) technology that addresses many of the challenges in building such a system. We demonstrate two approaches in integrating the transducer and the front-end electronics. The transducer is a 5-MHz CMUT array with an 8 mm × 8 mm aperture size. The aperture consists of 1024 elements (32 × 32) with an element pitch of 250 μm. An integrated circuit (IC) consists of a transmit beamformer and receive circuitry to improve the noise performance of the overall system. The assembly was interfaced with an FPGA and a back-end system (comprising of a data acquisition system and PC). The FPGA provided the digital I/O signals for the IC and the back-end system was used to process the received RF echo data (from the IC) and reconstruct the volume image using a phased array imaging approach. Imaging experiments were performed using wire and spring targets, a ventricle model and a human prostrate. Real-time volumetric images were captured at 5 volumes per second and are presented in this paper.

Forward-looking volumetric intracardiac imaging using a fully integrated CMUT ring array

Atrial fibrillation is the most common type of cardiac arrhythmia that now affects over 2.2 million adults in the United States alone. Currently fluoroscopy is the most common method for guiding interventional electrophysiological procedures. We are developing a 9-F forward-looking intracardiac ultrasound catheter for real-time volumetric imaging. We designed and fabricated a 64-element 10-MHz CMUT ring array with through-wafer via interconnects. We also designed custom front-end electronics to be closely integrated with the CMUT array at the tip of the catheter for improved SNR. This integrated circuit (IC) is composed of preamplifiers and protection circuitry, and can directly interface a standard imaging system. This multi-channel IC is capable of passing up to ±50-V bipolar pulses. An 8-channel front-end IC was fabricated based on this circuit topology. Additionally, a flexible PCB was designed for the integration of ring array with front-end electronics. We have acquired a PC-based real-time imaging platform and demonstrated real-time imaging with the ring array. We have also shown volume images using off-line full synthetic aperture image reconstruction method. The presented experimental results demonstrate the performance of our forward-looking volumetric intracardiac imaging approach. We are currently working on the final catheter integration and further development of our real-time imaging methods.

Volumetric real-time imaging using a CMUT ring array

A ring array provides a very suitable geometry for forward-looking volumetric intracardiac and intravascular ultrasound imaging. We fabricated an annular 64-element capacitive micromachined ultrasonic transducer (CMUT) array featuring a 10-MHz operating frequency and a 1.27-mm outer radius. A custom software suite was developed to run on a PCbased imaging system for real-time imaging using this device. This paper presents simulated and experimental imaging results for the described CMUT ring array. Three different imaging methods-flash, classic phased array (CPA), and synthetic phased array (SPA)-were used in the study. For SPA imaging, two techniques to improve the image quality-Hadamard coding and aperture weighting-were also applied. The results show that SPA with Hadamard coding and aperture weighting is a good option for ring-array imaging. Compared with CPA, it achieves better image resolution and comparable signal-tonoise ratio at a much faster image acquisition rate. Using this method, a fast frame rate of up to 463 volumes per second is achievable if limited only by the ultrasound time of flight; with the described system we reconstructed three cross-sectional images in real-time at 10 frames per second, which was limited by the computation time in synthetic beamforming.

Forward-looking intracardiac imaging catheters using fully integrated CMUT arrays

Atrial fibrillation, the most common type of cardiac arrhythmia, now affects more than 2.2 million adults in the US alone. Currently, electrophysiological interventions are performed under fluoroscopy guidance, which besides its harmful ionizing radiation does not provide adequate soft-tissue resolution. Intracardiac echocardiography (ICE) provides realtime anatomical information that has proven valuable in reducing the fluoroscopy time and enhancing procedural success. We developed two types of forward-looking ICE catheters using capacitive micromachined ultrasonic transducer (CMUT) technology: MicroLinear (ML) and ring catheters. The ML catheter enables real-time forward-looking 2-D imaging using a 24-element 1-D CMUT phased-array that is designed for a center frequency of 10 MHz. The ring catheter uses a 64-element ring CMUT array that is also designed for a center frequency of 10 MHz. However, this ring-shaped 2-D array enables real-time forward-looking volumetric imaging. In addition, this catheter provides a continuous central lumen that enables convenient delivery of other devices such as RF ablation catheter, EP diagnostic catheter, biopsy devices, etc. Both catheters are equipped with custom front-end ICs that are integrated with the CMUT arrays at the tip of the catheters. The integration of the ICs with the CMUT arrays was accomplished using custom flexible PCBs. We also developed several image reconstruction schemes for the ring catheter on a PC-based imaging platform from VeraSonics. We performed a variety of bench-top characterizations to validate the functionality and performance of our fully integrated CMUT arrays. Using both catheters, we demonstrated in vivo images of the heart in a porcine animal model. We have successfully prototyped the first CMUT-based ICE catheters and proven the capabilities of the CMUT technology for implementing high-frequency miniature transducer arrays with integrated electronics.

3D volumetric ultrasound imaging with a 32×32 CMUT array integrated with front-end ICs using flip-chip bonding technology

3D ultrasound imaging is becoming increasingly prevalent in the medical field. Compared to conventional 2D imaging systems, 3D imaging can provide a detailed view of tissue structures that makes diagnosis easier for the physicians. In addition, 2D image slices can be formed at various orientations to the transducer, making the examination less dependent on the skill of the sonographer. However, various challenges exist in developing a 3D imaging system, such as integration of a large number of elements, as well as post-processing of datasets received from a large number of channels. 2D transducer arrays are typically integrated with custom ICs in the probe handle to perform some intermediate beamforming and to reduce the number of cable connections to the imaging system. Capacitive micromachined ultrasonic transducers (CMUTs) have emerged as an alternative to piezoelectric transducers. Being a MEMS device, they greatly benefit from flexibility and ease of fabrication, and can be seamlessly integrated with electronics. Previous work demonstrates 3D stacking of CMUTs and dummy ICs with an intermediate interposer layer. However, that represents more of a mechanical demonstration of 3D integration. In this paper, we present a fully functional 3D ultrasound imaging system comprising a 32×32 2D CMUT array, 3D-stacked with front-end ICs using flip-chip bonding technology. The imaging system is capable of capturing real-time volumetric ultrasound data, and displaying 2D and 3D ultrasound images.

GPU-Based Real-Time Volumetric Ultrasound Image Reconstruction for a Ring Array

Synthetic phased array (SPA) beamforming with Hadamard coding and aperture weighting is an optimal option for real-time volumetric imaging with a ring array, a particularly attractive geometry in intracardiac and intravascular applications. However, the imaging frame rate of this method is limited by the immense computational load required in synthetic beamforming. For fast imaging with a ring array, we developed graphics processing unit (GPU)-based, real-time image reconstruction software that exploits massive data-level parallelism in beamforming operations. The GPU-based software reconstructs and displays three cross-sectional images at 45 frames per second (fps). This frame rate is 4.5 times higher than that for our previously-developed multi-core CPU-based software. In an alternative imaging mode, it shows one B-mode image rotating about the axis and its maximum intensity projection, processed at a rate of 104 fps . This paper describes the image reconstruction procedure on the GPU platform and presents the experimental images obtained using this software.

Design optimization for a 2-D sparse transducer array for 3-D ultrasound imaging

In 3-D ultrasound imaging where 2-D transducer arrays with more than hundreds of elements are used, sparse arrays can be used to reduce the number of active ultrasound channels. Under a restriction of desired number of active channels, we can maximize the image quality by optimally choosing the positions of active elements. Here we use the method of simulated annealing to find the optimal configuration of a 2-D sparse array. This algorithm tries to minimize the value of an objective function defined as the energy ratio between the non-focal and focal regions in the point spread function (PSF). Optimal configurations were found for the cases of choosing 16, 20, 24, 28, and 32 transmit and receive elements from a 16×16-element rectangular transducer array. With only 32 transmit and 32 receive elements, we could achieve an energy ratio of 16%, compared to 6% of the full array, which is the gold standard utilizing all the 256 elements for both transmit and receive. Using Field II, we simulated imaging with the optimal sparse arrays, for off-axis targets as well as on-axis targets, and the resulting images were compared with those from some other configurations, such as full-transmit full-receive, full-transmit x-receive, x-transmit boundary-receive, and so on.

Real-time volumetric imaging system for CMUT arrays

We designed and implemented a flexible real-time volumetric ultrasound imaging system for capacitive micromachined ultrasonic transducer (CMUT) arrays, consisting of an ultrasound data acquisition system, an FPGA board, and a host PC. The system works with arbitrary-shaped CMUT arrays and non-standard beamforming methods, as well as with regular-shaped CMUT arrays and conventional beamforming methods. In this paper, we present the system design and real-time imaging results obtained using this system with a ring array, a rectangular array, and a linear array. In synthetic phased array (SPA) imaging with a 64-element ring array, we could display 3 image planes with a total of about 70,000 pixels in real time, at a frame rate of 9 frames per second (fps) which was limited by the computational load on the CPU required for synthetic beamforming. On the other hand, the frame rate in classic phased array (CPA) imaging is limited by the data transfer time. In CPA imaging with a 16×16-element rectangular array, a frame rate of 5.4 fps was achieved for 1,250 acquisitions per frame and a 2.5-cm imaging depth. The frame rate can be increased by reducing the number of pixels processed in SPA, or by reducing the number of beams received in CPA, at the expense of degraded image quality or reduced field of view.

Ultrasound-guided therapeutic modulation of hepatocellular carcinoma using complementary microRNAs.

Treatment options for patients with hepatocellular carcinoma (HCC) are limited, in particular in advanced and drug resistant HCC. MicroRNAs (miRNA) are non-coding small RNAs that are emerging as novel drugs for the treatment of cancer. The aim of this study was to assess treatment effects of two complementary miRNAs (sense miRNA-122, and antisense antimiR-21) encapsulated in biodegradable poly (lactic-co-glycolic acid) nanoparticles (PLGA-NP), administered by an ultrasound-guided and microbubble-enhanced delivery approach in doxorubicin-resistant and non-resistant human HCC xenografts. Proliferation and invasiveness of human HCC cells after miRNA-122/antimiR-21 and doxorubicin treatment were assessed in vitro. Confocal microscopy and qRT-PCR were used to visualize and quantitate successful intracellular miRNA-loaded PLGA-NP delivery. Up and down-regulation of miRNA downstream targets and multidrug resistance proteins and extent of apoptosis were assessed in vivo in treated human HCC xenografts in mice. Compared to single miRNA therapy, combination therapy with the two complementary miRNAs resulted in significantly (P<0.05) stronger decrease in cell proliferation, invasion, and migration of HCC cells as well as higher resensitization to doxorubicin. Ultrasound-guided delivery significantly increased in vivo miRNA-loaded PLGA-NP delivery in human HCC xenografts compared to control conditions by 5-9 fold (P<0.001). miRNA-loaded PLGA-NP were internalized in HCC cells and anti-apoptotic proteins were down regulated with apoptosis in ~27% of the tumor volume of doxorubicin-resistant human HCC after a single treatment with complementary miRNAs and doxorubicin. Thus, ultrasound-guided delivery of complementary miRNAs is highly efficient in the treatment of doxorubicin- resistant and non-resistant HCC. Further development of this new treatment approach could aid in better treatment of patients with HCC.