Research
(1) New cysteine-reactive probes to expand the pool of ligandable cysteines for biological studies and drug development
Understanding functions and reactivity of cysteines on proteins have arouse lots of interest. This is not only for getting better ideas on biological processes initiated/regulated by proteins containing these functional cysteines, but also for the huge potentials to develop drug compounds targeting these cysteines for therapy.
Activity-based protein profiling (ABPP) is one of the most widely used platforms for proteome-wide cysteine profiling to identify functional cysteines. Cysteine-reactive probe is the key component in ABPP platform and can define the pool of ligandable cysteines and hence the population of proteins that can be targeted by covalent ligands. The conventional cysteine-reactive probe, iodoacetamide-alkyne (IAA), shows only fair reaction kinetics and selectivity with cysteine. Together with its high cellular toxicity and low biostability, this limits the full potential of ABPP platform for biological studies and drug research.
Against this backdrop, our lab is developing novel cysteine-reactive probes which show better cysteine reaction kinetics and selectivity. Together with their high biostability and low cytotoxicity, they have been found to capture more cysteines than IAA in cell lysates and live cells, in both gel-based and fluorescence imaging experiments. More interestingly, in MS experiments, our probes capture a larger and a significantly different population of cysteines than IAA. This should expand the pool of ligandable hotspots in whole-proteome cysteine profiling experiments, and facilitates further research and study on the development of new covalent ligands and potential lead compounds for targeting these new proteins for therapy.
Representative work:
Koo, T.-Y.,† Lai, H.,† Nomura, D. K., & Chung, C. Y.-S.* (2023). N-Acryloylindole-alkyne (NAIA) enables imaging and profiling new ligandable cysteines and oxidized thiols by chemoproteomics. Nature Communications, 14(1), 3564. (†These authors contributed equally) https://doi.org/10.1038/s41467-023-39268-w
(2) New chemical tools and technologies to study cellular redox biology
Cellular redox biology is governed by interesting classes of redox-active molecules, which can mediate reduction/oxidation of biomolecules. Notably examples of redox-active molecules include reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) and superoxide (O2–), and hydrogen sulfide (H2S) which belongs to reactive sulfur species (RSS) family. These redox-active molecules were once classified as detrimental compounds in biology and known to contribute to serious disease development such as cancer and neurodegenerative disorders. Yet, recently these redox-active molecules are found to be essential to life and can serve as important signaling molecules. This is mainly through their Redox Biology, which induces reduction/oxidation of biomolecules and subsequently changes their functions, activities and downstream signals.
Despites nowadays there are better ideas on redox signaling and its biological functions, the molecular mechanism on initiation, transduction and regulation of redox signaling, as well as the identity of proteins involved in the signaling process, remain insufficiently understood. This is because redox modifications of proteins are reversible, dynamic and unstable. In contrast to protein phosphorylation/dephosphorylation which are more stable modifications, there are no good conventional biochemical and biological experiments to study protein redox modifications and hence Cellular Redox Biology.
To overcome this challenge, our lab is developing new chemical probes that can: (1) induce “permanent covalent tag” onto proteins associated with Redox Biology. This allows us to identify the proteins by proteomics and mass spectrometry (MS)-based experiments. (2) We can also real-time visualize redox signaling events by turning the permanent covalent tag into a fluorescent one. This enables fluorescence imaging with superior spatial resolution.
Representative work:
Lai, H., & Chung, C. Y.-S.*(2024). Superoxide-responsive quinone methide precursors (QMP-SOs) to studyuperoxide biology by proximity labeling and chemoproteomics. RSC ChemicalBiology. https://doi.org/10.1039/D4CB00111G
**In the themed collection: 2024 RSC Chemical Biology Emerging Investigators
(3) Therapeutic covalent ligands for cancer therapy
As compared to chemotherapy, targeted therapy is generally more specific and shows less toxic side effects, and is considered to be a better treatment option to cancer patients. Nonetheless, only a small population of cancer-related proteins have been drugged by targeted therapeutic agents, thus not much alternative treatment options are available when drug resistance develops or poor prognosis is found. This limitation can be explained by the fact that close to 90% of human proteins are lack of nice and deep pockets for drug compound binding, and they were once considered as undruggable.
Recently, covalent drugs have been successfully developed to target G12C mutation in KRAS, one of the most famous undruggable proteins. This motivates us to explore covalent ligands to target new cancer driver proteins and investigate their potential applications for targeted cancer therapy. Through chemoproteomics-enabled screening platform, we have identified novel covalent ligands targeting different cancer driver proteins which have not been drugged so far.By integrative chemical biology and MS-based ABPP experiments, we have confirmed the specific binding of the covalent ligands on the cancer driver proteins in cancer cells. Some of the covalent ligands have been found to show promising in vivo antitumor effects, demonstrating the good potential of using these compounds for targeted cancer therapy.
(4) Nanovaccines for cancer therapy
Over the past few decades, there have been significant advancements in applying nanotechnology in drug delivery system, particularly for cancer therapy. One of the emerging fields is to utilize nanosystems to deliver cancer vaccines specifically to immune cells for triggering immunity to eradicate tumor cells. Unlike checkpoint inhibitor-based immunotherapy, cancer vaccines contain tumor antigens and function by targeting antigen-presenting cells (APCs), so that the APCs presenting these antigens can prime and activate T cells for killing cancer cells. Notably, if the cancer antigens are somatic mutation products, these antigens, which are called neoantigens, should be found in cancer cells specifically but not normal cells and tissues, and also specific in patients with these cancer mutations. As a result, in principle cancer vaccines should allow the development of personalized cancer therapy for preventing cancers in high-risk individuals or for treating existing cancers in patients. Yet, challenges in clinical translation of cancer vaccines have been found due to tumor immunosuppressive microenvironment, insufficient antigen delivery to APCs and ineffective stimulation of T cells. To address these problems, preparation of new generation of cancer vaccines using nanotechnology, i.e. cancer nanovaccines, can be a good solution.
Our lab is developing novel polymer-peptide hybrid nanosystems for their uses in cancer immunotherapy. They show triggered release of the cargo in APCs, thus allowing cross-presentation of the neoantigens to CD8+ T cells and subsequent cytotoxic killing of the cancer cells.