**3. Potential clinical applications**

The main potential clinical application of labeled microbubbles includes early detection of tumors, monitoring tumor response, assessment of inflammation and/ or ischemia, early detection of transplant infection and potential for targeted drug delivery.

One of the challenges to use these microbubbles is their fabrication for clinical use. Specifically, the immunogenicity of the streptavidin used for attaching the ligand to the bubble requires that it be internalized within the microbubble. This is only available from a few manufacturers on an investigational basis. We reported our early experience developing a labeled microbubble [7].

Only a few studies using labeled microbubbles in humans have been reported [1]. The group headed by Willman showed that breast and ovarian malignant

### **Figure 3.**

*Time intensity curve in responsive (top) vs. non-responsive (bottom) hepatocellular cancer treated with anti-angiogenic medications.*

tumors had greater signal than benign ones. Specifically, in a study of 24 patients with ovarian cancers and 21 with breast lesions, 93% of malignant breast lesions and 77% of malignant ovarian cancers demonstrated increased signal [1].

Labeled microbubbles have also been used in detecting prostate cancer and sentinel lymph nodes [7, 8]. Molecular lymphosonography was used to detect metastatic involvement in a swine melanoma model. Metastatic involvement of the sentinel lymph node can distinguish local vs. metastatic disease in women with breast cancer. It has also been used in a variety of cancers such as melanoma, esophageal, head and neck, stomach and thyroid cancer. Molecular sonography using nanobubble targeted prostate specific membrane antigen has been investigated in an orthotopic murine model [8]. Increased extravasation and retention were observed in prostate cancer models as well as exhibiting different time/intensity curves (enhancement kinetics) in cancerous tissue.

Angiogenesis, the generation of new blood vessels, has also been extensively investigated with molecular sonography due to its role in the development and metastatic spread of numerous cancers. It is crucial in the growth of tumors and subsequent metastasis. VEGFR2 is one of the most studied markers of angiogenesis and is over expressed on tumor-associated endothelial cells [9]. In one study of a transgenic mouse model, microbubbles targeting VEGFR2 showed an increased contrast-enhanced ultrasound signal from hyperplasia to ductal carcinoma in situ and breast cancer compared to normal breast tissue [10]. In another study of laying hens, a microbubble targeting αvβ3 integrin sensitively detected ovarian cancer

### *Molecular Sonography: Current and Future Applications DOI: http://dx.doi.org/10.5772/intechopen.99903*

at an early stage [11]. These are just two studies that support the clinical utility of angiogenesis targeted microbubbles in early tumor detection.

Applying the principles of molecular sonography to atherosclerosis can help identify at-risk plaques before acute events occur, potentially enabling prevention of evolution into irreversible damage. Normally, patients do not exhibit symptoms until an episode of severe stenosis or rupture occurs; thus, there is valuable utility in monitoring the growth of atherosclerotic plaques. JAM-A is an example of a marker that has been linked to early plaque formation and vulnerability [12]. In a study performed on mice that were fed an atherogenic diet, molecular ultrasound of JAM-A showed early stages of atherosclerosis and detected acute blood flow variations [13].

Acute cardiac ischemia is another pathology that would benefit from molecular ultrasound imaging. Current methods of detection such as EKG and cardiac MRIs suffer from various shortcomings such as accuracy, convenience, or safety. Sonography with labeled microbubbles could resolve a number of these shortcomings when used to identify post-ischemic myocardium. E- and P- selectin are two potential markers that were used in murine models to identify ischemic tissue shortly after undergoing an ischemic event [14, 15]. While cardiac molecular ultrasound imaging has shown promising results regarding safety and efficacy, it is technically difficult and highly user-dependent. Further testing needs to be performed to assess its feasibility when compared with current gold standards.

Microbubbles also can be applied in the assessment of inflammatory bowel disease (IBD). Two methods of assessing IBD are endoscopy and ultrasound. The former requires a procedure under anesthesia, while the latter is restricted by its relatively limited sensitivity. Molecular ultrasound holds clinical potential to serve as a complementary tool with high sensitivity in the imaging of IBD. One study utilized porcine tissue with terminal ileitis imaged with dual E- and P-selectintargeted microbubbles. The study concluded that increased ultrasound signal correlated well with histologic grades of inflammation [16]. This study highlights selectin-targeted ultrasound's utility to serve as a high-sensitivity adjunctive tool for assessment of IBD in the clinical setting.
