An Unbiased Integrated Chemical Biology Approach for Target Identification of Ruthenium-based Anticancer Drug Candidates on the Edge to Clinical Application

Breast cancer is one of the most common cancers worldwide. Triple-negative breast cancers (TNBCs), accounting for 15%–20% of breast cancer cases, are particularly challenging to treat owing to their aggressive nature, higher chance of spreading to other organs, and lack of effective targeted therapies. Although chemotherapy remains the standard treatment, it is often associated with severe side effects, and there is a need for new and safer treatment options. The proposed project will advance the development of a promising ruthenium-based anticancer compound, RUTEN3. Early studies have shown that RUTEN3 can effectively stop the growth of TNBC tumors and prevent their spread, and it has fewer side effects than traditional chemotherapy drugs. However, to advance RUTEN3 toward clinical use, it is essential to understand how it works at the molecular level. Specifically, we need to know the proteins it targets and how it interacts with them. The proposed project will explore three key questions:1. What are the molecular targets of RUTEN3? Using advanced techniques such as the thermal proteome profiling (TPP), we will identify which proteins RUTEN3 binds to within cancer cells. These interactions will be validated to ensure they are accurate and meaningful. 2. Is RUTEN3 stable in biological environments?We will study how RUTEN3 behaves in different biological fluids and whether it undergoes chemical changes, such as breakdown by enzymes in the liver. Stability is crucial for ensuring that the drug remains effective in the body. 3. Does RUTEN3 interact with its targets through reversible or permanent binding?We will investigate whether RUTEN3 forms reversible (non-covalent) or permanent (covalent) bonds with its target proteins. Reversible interactions can reduce the risk of long-term side effects, whereas permanent interactions may offer higher potency but require careful monitoring to avoid toxicity.Our interdisciplinary team, having expertise in bioinorganic chemistry, molecular biology and proteomics, will adopt state-of-the-art mass spectrometry to conduct these studies. The project will lay the groundwork for more effective, low-toxicity treatments for TNBCs. Within the three-year period covered by this proposal, this project will impact the medicinal chemistry community by clarifying the interactions between ruthenium drug candidates and their protein targets, providing valuable insights into their nature. In the long run, understanding the targets of RUTEN3 will help identify which patients can benefit from this drug candidate, predict any possible side effects it might have and guide future clinical trials.