More than just a traitorous lump of cells, tumors are clever instigators of rebellion in the body, encouraging the growth of a network of blood vessels around them to maintain a supply of essential nutrients and oxygen. By understanding of how blood vessels sprout around tumors—a process known as angiogenesis—scientists can develop drugs to cut off the tumor’s lifeline and improve patient outcomes.
To observe the architecture of blood vessels in tumors, researchers led by Lai Guan Ng, a Principal Investigator at A*STAR’s Singapore Immunology Network (SIgN) and Bin Liu, a Professor at the National University of Singapore, devised an imaging tool that not only allows for clear visualization of blood vessels in deep brain tissue, but also helps distinguish abnormal tumor blood vessels from normal ones. The research was carried out by Shaowei Wang, a Researcher from the National University of Singapore.
A major consideration for deep tissue imaging is the wavelength of light used—not all wavelengths penetrate deeply into biological tissue. “Traditional two-photon microscopy usually uses laser light in the near-infrared (NIR)-I region (700-950 nm) as the excitation light source, which limits the imaging depth to around 500 μm,” Ng explained. “Compared to NIR-I light, NIR-II light (1,000-1,700 nm) can penetrate much deeper to excite a fluorescent probe inside living tissues.”
Getting the right fluorescent probe was the second hurdle the team needed to overcome. Traditional NIR probes, which have poor solubility in water, suffer from aggregation-induced quenching—their fluorescence is largely reduced when they clump together in an aqueous biological environment. Hence, the researchers used a combination of nanotechnology and engineering to create fluorophores that emit a strong fluorescence when clumped together.
“We fabricated small nanoparticles comprising thousands of fluorophore molecules aggregated together for live tissue imaging. This is known as aggregation-induced emission,” said Wang.
Tested in mice, the researchers showed that their nanoparticles excited by NIR-II light could illuminate blood vessels in the brain with great clarity to a depth of around 1 mm—twice as deep as what has been possible by conventional imaging methods. The nanoparticles also differentiated normal brain vessels from tumor vessels; the intensity of the fluorescence was higher in tumor vessels.
“[The brighter fluorescence in tumors] is attributed to the unique leaky structure of tumor vasculature, which allows the fluorophores to stick to the vessel wall and form bright aggregates,” said Ng.
Compared to fluorophores presently approved by the FDA for clinical imaging, the nanoparticles in this study have higher stability in water, good biocompatibility and a longer circulating time in the blood. The high-resolution imaging afforded by the nanoparticles has the potential to improve the detection of residual tumor tissue after surgical excision of the bulk tumor.
The team is already thinking of ways to improve the performance and function of their fluorescent probe. “The imaging depth can be improved by using light with longer wavelengths. Additionally, antibody or peptides can be attached to the surface of our nanoparticles, giving them specific tumor-targeting capabilities,” said Wang.
The A*STAR-affiliated researchers contributing to this research are from the Singapore Immunology Network (SIgN).
