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Overview of the probe and enzyme.
A reporter enzyme, engineered by directed evolution, does not bind to healthy tissue, only targeted cancers with particular antigens. A probe is activated by the reporter enzyme which glows under excitation light.
Image Credit: ©2026 Kojima et al. American Chemical Society
Scientific Frontline: "At a Glance" Summary: Precision Tumor Imaging
- Main Discovery: Researchers developed a bioorthogonal fluorescence probe and a matching engineered reporter enzyme that selectively activate at targeted tumor sites, enabling high-contrast tumor visualization with minimal background noise.
- Methodology: The research team used directed evolution to train a reporter enzyme through repeated mutation and selection. In tests utilizing a mouse model with peritoneal cancer, the engineered enzyme was delivered specifically to tumor sites, followed by the introduction of the bioorthogonal dye probe. The probe was designed to remain completely inactive until encountering its matching engineered enzyme.
- Key Data: The targeted bioorthogonal system successfully highlighted millimeter-sized tumor lesions in vivo, demonstrating exceptionally low background fluorescence from surrounding healthy tissues.
- Significance: Conventional fluorescent dyes frequently illuminate healthy tissue via endogenous enzyme activation, complicating surgical tumor excision. This highly selective enzyme-probe pairing effectively eliminates background noise, significantly enhancing surgical precision and minimizing the risk of leaving undetected malignant cells behind.
- Future Application: The system serves as a powerful near-term research tool with significant long-term clinical potential for surgical oncology. Furthermore, by substituting the antigen-targeting component, the same enzyme-probe pairing principles can be adapted to other cancer types for highly targeted drug delivery, ensuring therapeutics exclusively reach malignant sites.
- Branch of Science: Chemical Biology, Molecular Imaging, and Oncology.
- Additional Detail: Before human clinical trials can proceed, researchers must address the significant challenge of ensuring that the engineered reporter enzyme does not provoke an adverse immune response in patients.
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| Glowing tumors. The red arrows point to green glowing tumors. Image Credit: ©2026 Kojima et al. American Chemical Society |
Successful cancer surgery depends on a surgeon’s ability to remove tumors, while minimizing harm to healthy tissues. Surgeons currently use glowing dyes which mark cancer cells to help differentiate from healthy cells, but these dyes aren’t perfect and will light up some healthy tissues too. For the first time, researchers including those from the University of Tokyo developed what they call a bioorthogonal fluorescence probe and a matching reporter enzyme that can activate the probe selectively at targeted tumor sites. This enables high-contrast tumor visualization with very low background. This study was performed in mice.
Cancer is a universal issue which affects uncountably many people around the world. Many will turn to surgery in the hope a surgeon will be able to completely remove a tumor leaving healthy tissues unaffected. Various tools and techniques have been developed over the years to improve the way these surgeries are performed, and visual imaging methods such as glowing dyes have proven to be very useful. But one drawback is that some probes can also be activated in healthy tissues by endogenous enzymes, creating background fluorescence and making it harder to judge what should be removed. The opposite is also possible, where cancer cells are left unmarked and are missed during surgery, increasing the chance of recurrence.
“Our group acknowledged this current shortcoming and improved upon this way to make cancer cells light up inside the body. In tests on mice, we delivered a special enzyme to tumors and used a fluorescence probe that only turns on when that enzyme is present,” said Associate Professor Ryosuke Kojima from the Laboratory of Chemical Biology and Molecular Imaging at the University of Tokyo. “Older probes often light up healthy tissue by mistake, creating background noise, but our highly selective, or bioorthogonal, dye probe is designed to stay completely off unless it meets its matching engineered enzyme. We essentially trained the enzyme through repeated mutation and selection, a form of directed evolution, so it could activate the probe strongly enough to work inside living animals.”
Kojima, with Professor Yasuteru Urano and their team, created a special fluorescent probe that is not easily activated by natural enzymes in the body, which helps prevent unwanted background glow. This probe was paired with a matching reporter enzyme specially tailored to switch it on, so fluorescence appears mainly where the enzyme is delivered. When tested in mice bearing peritoneal cancer, the engineered enzyme reached the tumors in the abdominal wall lining and was followed up by the probe which lit up as expected.
“This allowed us to see tiny, millimeter-sized tumor lesions with extremely low background noise, a level of contrast that could be very useful during surgery,” said Kojima. “In the near term, this system could become a powerful research tool, and in the longer term, it may help surgeons remove tumors more completely by clearly highlighting cancer cells. A major hurdle for clinical use will be ensuring that the engineered enzyme does not trigger an unwanted immune response in patients.”
The system could be adapted to other kinds of cancers too, beyond the peritoneal cancer used in these trials, as many cancers present corresponding antigens, telltale markers of tumor tissue. By swapping the tumor-targeting component (for example, an antibody or nanobody against a chosen antigen), the same enzyme–probe pair could in principle be redirected to other cancer types. Looking even further ahead, this research could even be helpful in highly targeted drug delivery, where instead of glowing dyes, cancer-fighting drugs would be sent to the sites they’re needed, and nowhere else. But as Kojima stresses, it's still early days, the trials have only been done in mice, and much work is needed before it’s deemed safe enough for human trials.
Published in journal: Journal of the American Chemical Society
Authors: Ziyi Wang, Ryosuke Kojima, Rikuki Kiji, Kyohhei Fujita, Ryo Tachibana, Reiko Tsuchiya, Taku Uchiyama, Yoshihiro Minagawa, Tadahaya Mizuno, Kiyohiko Igarashi, Hiroyuki Noji, Mako Kamiya, and Yasuteru Urano
Source/Credit: University of Tokyo
Reference Number: ongy030326_01