Molecular Ultrasound Imaging

Abstract

In the last decade, molecular ultrasound imaging has been rapidly progressing. It has proven promising to diagnose angiogenesis, inflammation, and thrombosis, and many intravascular targets, such as VEGFR2, integrins, and selectins, have been successfully visualized in vivo. Furthermore, pre-clinical studies demonstrated that molecular ultrasound increased sensitivity and specificity in disease detection, classification, and therapy response monitoring compared to current clinically applied ultrasound technologies. Several techniques were developed to detect target-bound microbubbles comprising sensitive particle acoustic quantification (SPAQ), destruction-replenishment analysis, and dwelling time assessment. Moreover, some groups tried to assess microbubble binding by a change in their echogenicity after target binding. These techniques can be complemented by radiation force ultrasound improving target binding by pushing microbubbles to vessel walls. Two targeted microbubble formulations are already in clinical trials for tumor detection and liver lesion characterization, and further clinical scale targeted microbubbles are prepared for clinical translation. The recent enormous progress in the field of molecular ultrasound imaging is summarized in this review article by introducing the most relevant detection technologies, concepts for targeted nano- and micro-bubbles, as well as their applications to characterize various diseases. Finally, progress in clinical translation is highlighted, and roadblocks are discussed that currently slow the clinical translation.

Keywords: active targeting; angiogenesis; clinical translation; inflammation; molecular imaging; molecular ultrasound; nanobubbles; targeted microbubbles; thrombosis.

Figure 1

Non-linear response from UCA to US. The US transducer is transmitting ultrasound waves to tissue and UCA. For tissue (A) the sent and received signals are similar (linear response), while for UCA (B) the received signal has a different frequency than the initial frequency (non-linear response).

Figure 2

Theoretical background of imaging techniques based on the non-linear response of UCA. During pulse inversion two pulses are transmitted. The second pulse is shifted by 180 degrees. After receiving the echoes, the responses are summed up. In tissue the transmitted and received signals are the same and thus cancel out. For UCA the summed signal does not cancel out due to the non-linear response. During power modulation two pulses are transmitted. The second pulse has a two-fold difference in amplitude. For the summation of the responses the second pulse is multiplied by two. For tissue the response is near zero, for UCA the summed response is not zero due to the irregularity in UCA echoes. Contrast pulse sequencing is a combination of both mentioned methods. Two pulses are used where the second one is shifted by 180 degrees and has an amplitude twice the magnitude as the first one. Reproduced with permission from [11]. Copyright Elsevier, 2011.

Figure 3

Schematic representation of microbubbles, nanobubbles, and nanodroplets.

Figure 4

Schematic representation of different functionalization methods of UCA.

Figure 5

CEUS images of treated (top) and untreated tumors (bottom). Left side: early vascular phase with VEGFR2-targeted MB as a functional imaging biomarker; Right side: late phase of VEGFR2-specific binding with the targeted MB as a molecular imaging biomarker 8 min after contrast injection. Reproduced with permission from [51]. Copyright Eschbach et al., 2017.

Figure 6

B-mode US images of the left ventricle 4 h after conorary occlusion overlayed with the signal of E-selectin targeted MB (A), non-targeted MB (B) and non-specific IgG targeted MB (C). Enhanced contrast signal is seen using E-selectin targeted MB. Reproduced with permission from [88]. Copyright SAGE Publications, 2014.

Figure 7

Molecular US imaging of thrombosis. Spatial maps of the molecular US signal color-coded and overlaid on B-mode images acquired at baseline after infusion of MB and washing with saline using targeted MB (left), non-targeted MB (middle), and targeted MB co-injected with hirudin (right). Targeted MB give specific signal only in the presence of thrombin. Reproduced with permission from [97]. Copyright American Chemical Society, 2017.

Figure 8

In vitro microscopy images of control (MB_C), single (MB_S), double (MB_D), and triple (MB_T) targeted MB adhered on cells. Triple targeted MB adhere more on the cells than single or double targeted ones. Reproduced with permission from [139]. Copyright John Wiley and Sons, 2011.

Figure 9

Contrast harmonic images of NB and commercially available Definity^® MB (DEF) at different concentrations and frequencies. Reproduced with permission from [161]. Copyright Elsevier, 2013.