Development and application of bioluminescent probes to in vivo imaging

2023 Fiscal Year Final Research Report

• Project/Area Number: 22K20539
• Research Category: Grant-in-Aid for Research Activity Start-up
• Allocation Type: Multi-year Fund
• Review Section: 0501:Physical chemistry, functional solid state chemistry, organic chemistry, polymers, organic materials, biomolecular chemistry, and related fields
• Research Institution: University of Miyazaki
• Principal Investigator: Kiyama Masahiro 宮崎大学, 研究・産学地域連携推進機構, 研究員 (10968514)
• Project Period (FY): 2022-08-31 – 2024-03-31
• Keywords: 生物発光 / プローブ / イメージング / 近赤外発光 / プロテアーゼ

Outline of Final Research Achievements

We have developed a 3CL turn-on AkaLumine probe designed for detecting the SARS-CoV-2 main protease (3CL). This probe consists of a 3CL recognition peptide conjugated with AkaLumine, a luminescent substrate in the near-infrared bioluminescence system AkaBLI. In vitro analysis demonstrated a 20-fold increase in luminescence intensity when exposed to 3CL protease, confirming its high specificity and sensitivity. However, in vivo administration of the probe to AkaLuc-expressing mice in the absence of 3CL protease resulted in an unexpected 20% increase in luminescence compared to control conditions with AkaLumine alone. This unintended luminescence is hypothesized to result from the hydrolysis of the probe by serine proteases in the blood, leading to the premature release of AkaLumine.
Our current research is focused on enhancing the stability of the probe to prevent its hydrolysis by serine proteases in vivo.

Free Research Field

生物有機化学

Academic Significance and Societal Importance of the Research Achievements

AkaBLIはAkalucの局在、増殖過程、遺伝子発現動態を観察できる。さらに複雑な生命現象を観察するため、特定の生体分子に呼応して発光活性を変化させるプローブの開発が必要である。今回の研究では、AkaLumineに生体分子に呼応するペプチドを新しい修飾法で修飾したプローブを合成することができた。AkaLumine以外にも発光波長の異なる人工発光基質に適応可能であり、多色で観察できる技術となり得る。一方、ペプチドのアミド結合様の易生分解性構造を有していると、in vivoでは容易に分解されうるため、その回避方法の検討が必要であることも判明した。