Advances In Chemical Biology Research Articles

A Rapid and Sensitive MicroPlate Assay (MPSA) Using an Alkyne-Modified CMP-Sialic Acid Donor to Evaluate Human Sialyltransferase Specificity.

Human sialyltransferases primarily utilize CMP-Sias, especially transferring Neu5Ac from CMP-Neu5Ac to various acceptors. Advances in chemical biology have led to the synthesis of novel CMP-Sia donors suitable for bioorthogonal reactions in cell-based assays. However, the compatibility of these donors with all human enzymes remains uncertain. We synthesized a non-natural CMP-Sia donor with an alkyne modification on the N-acyl group of Neu5Ac, which was effectively used by human ST6Gal I and ST3Gal I. A sensitive MicroPlate Sialyltransferase Assay (MPSA) was developed and expanded to a panel of 13 human STs acting on glycoproteins. All assayed enzymes tolerated CMP-SiaNAl, allowing for the determination of kinetic parameters and turnover numbers. This study enhances the biochemical characterization of human sialyltransferases and opens new avenues for developing sialyltransferase inhibitors.

Small chemical molecules regulating the phytohormone signalling alter the plant's physiological processes to improve stress adaptation, growth and productivity.

Small chemical molecules are attractive agents for improving the plant processes associated with plant growth and stress tolerance. Recent advances in chemical biology and structure-assisted drug discovery approaches have opened up new avenues in plant biology to discover new drug-like molecules to improve plant processes for sustained food production. Several compounds targeting phytohormone biosynthesis or signalling cascades were designed to alter plant physiological mechanisms. Altering Abscisic acid synthesis and its signalling process can improve drought tolerance, and the processes targeted are reversible. Molecules targeting cytokinin, Auxin, and gibberellic acid regulate plant physiological processes and can potentially improve plant growth, biomass and productivity. The potential of molecules may be exploited as agrochemicals to enhance agricultural productivity. The discovery of small molecules provides new avenues to improve crop production in changing climatic conditions and the nutritional quality of foods. We present the rational combinations of small molecules with inhibitory and co-stimulatory effects and discuss future opportunities in this field.

Abstract A026: Senescence driven by pharmacological RAS inhibition potentiates anti-tumor activity of combination therapies in pancreatic cancer

Abstract Senescence is a form of stable proliferative arrest that occurs in response to various forms of cellular stress. In established cancer, these include unresolved DNA damage triggered by chemotherapeutic agents or irradiation, re-establishment of tumor suppressive programs, and by drugs that target cell cycle progression. Senescent cells take on a “Senescence Associated Secretory Phenotype” or SASP, characterized by increased expression of proinflammatory cytokines and other factors that modulate the tissue environment. The SASP triggers immune cell infiltration and, in the context of cancer, potent antitumor immunity. This implicates the use of senescence-inducing agents clinically and suggests that senescence biology may be harnessed to increase the fraction of patients that respond to immunotherapy. RAS is mutated in over 90% of pancreatic cancers, but until recently, has been considered to be “undruggable” due to its small size and lack of drug binding pockets. Recent advances in chemical biology have led to the development of several mutant isoform-specific and broad-spectrum RAS-GTP inhibitors, which hold great clinical promise. We have found that, in mouse models of pancreatic cancer, the non-covalent broad-spectrum inhibitor RM-042 and the covalent KRASG12D specific inhibitor RM-044 trigger a senescence-like phenotype characterized by cell cycle arrest and increased SASP expression. RAS inhibitor-mediated senescence enhances the immune surveillance of tumor cells, and this effect is potentiated by certain combination agents. Our results suggest that senescence contributes to RAS inhibitor efficacy in PDAC and reveal strategies to harness the immune system to improve therapy outcomes and circumvent resistance. Citation Format: Caroline Broderick, Riccardo Mezzadra, Alejando Jiminez-Sanchez, Almudena Chaves-Perez, Janelle Simon, Yu-Jui Ho, Exequiel Sisso, Sha Tian, Amanda Kulick, Elisa DeStanchina, Scott W. Lowe. Senescence driven by pharmacological RAS inhibition potentiates anti-tumor activity of combination therapies in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr A026.

Proteomics Update and Perspectives from the Proteomics in Cell Biology and Disease Mechanisms Conference.

Proteomics, or the large-scale study of proteomes, has benefitted from many recent advances in chemical biology, mass spectrometry, and machine learning. The Proteomics in Cell Biology and Disease Mechanisms conference showcased the synergy between these elements and the vast range of biological questions that proteomics can now help us to answer.

Chemical biology to dissect molecular mechanisms underlying plant circadian clocks.

Circadian clocks regulate the diel rhythmic physiological activities of plants, enabling them to anticipate and adapt to day-night and seasonal changes. Genetic and biochemical approaches have suggested that transcription-translation feedback loops (TTFL) are crucial for Arabidopsis clock function. Recently, the study of chemical chronobiology has emerged as a discipline within the circadian clock field, with important and complementary discoveries from both plant and animal research. In this review, we introduce recent advances in chemical biology using small molecules to perturb plant circadian clock function through TTFL components. Studies using small molecule clock modulators have been instrumental for revealing the role of post-translational modification in the clock, or the metabolite-dependent clock input pathway, as well as for controlling clock-dependent flowering time.

Abstract 3727: Chemical and genetic targeting of epigenetic proteins for cancer therapy

Abstract Recent evidence suggests that some cancers turn on or off some genes through an entirely a different mechanism, called epigenetics. The robust epigenetic landscape plays a vital role in gene expression via its ability to promote or repress transcription. Reprogramming of these processes can lead to genomic instability, which may propagate a cancerous state. Thus, there are proteins that turn on and off genes can control cell growth and cancer behaviors. Our research focuses to combat this reprogramming utilizing novel small molecules; targeting epigenetic proteins involved in these gene regulatory pathways. Among all the cancer type, the pediatric cancers posted significant challenge for effective therapy development. Despite intense study and cooperative group trials, patients diagnosed with aggressive, high-risk pediatric tumors such as sarcomas, neuroblastoma, high grade gliomas and embryonal brain tumors have poor overall survival outcomes. In addition to the limited efficacy of conventional chemotherapy and radiation in such patients, these modalities are fraught with toxicities, leaving survivors with long-term deficits including infertility, heart failure, endocrine disruption and a risk of secondary malignancies. Thus, one of the most pressing clinical needs in pediatric oncology remains the identification of agents with potent anti-tumor activity that have limited toxicities. Recently, we have utilized both inhibitors and degraders to study the EP300 function in neuroblastoma, an aggressive, hard to treat pediatric cancer. Advances in chemical biology have recently identified a novel group of molecules, called proteolysis-targeted chimaeras (PROTACs) or degrader. These agents may be generated toward any target that has a small molecule binder. To fully assess the therapeutic potential of this novel strategy for pediatric cancers, and potentially for other cancers as well, we set out to target the histone acetyltransferase EP300 that regulates growth and survival in nearly 65% of cancer cell lines, especially in high-risk neuroblastoma. Here, we generated potent and specific reagents, including a lead molecule called JQAD1, that are capable of driving specific degradation of EP300 with limited off-target effects. These compounds induce on-target apoptosis and growth delay in xenografted neuroblastoma tumors, and display excellent pharmacokinetics in vivo without detectable hematologic, hepatic, renal, neurologic or other toxicities. This study showcased our approach to advance cancer research by combining the power of chemistry, chemical biology, structural biology, computational biology and cellular biology. We cover the full spectrum of drug discovery and mechanistic studies for cancer, and also make meaningful and strong collaborations within the cancer research community. Citation Format: Jun Qi. Chemical and genetic targeting of epigenetic proteins for cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3727.

Membrane Mimetic Chemistry in Artificial Cells.

Lipid membranes in cells are fluid structures that undergo constant synthesis, remodeling, fission, and fusion. The dynamic nature of lipid membranes enables their use as adaptive compartments, making them indispensable for all life on Earth. Efforts to create life-like artificial cells will likely involve mimicking the structure and function of lipid membranes to recapitulate fundamental cellular processes such as growth and division. As such, there is considerable interest in chemistry that mimics the functional properties of membranes, with the express intent of recapitulating biological phenomena. We suggest expanding the definition of membrane mimetic chemistry to capture these efforts. In this Perspective, we discuss how membrane mimetic chemistry serves the development of artificial cells. By leveraging recent advances in chemical biology and systems chemistry, we have an opportunity to use simplified chemical and biochemical systems to mimic the remarkable properties of living membranes.

Recent Advances in Chemical Biology of Mitochondria Targeting.

Mitochondria are vital subcellular organelles that generate most cellular chemical energy, regulate cell metabolism and maintain cell function. Mitochondrial dysfunction is directly linked to numerous diseases including neurodegenerative disorders, diabetes, thyroid squamous disease, cancer and septicemia. Thus, the design of specific mitochondria-targeting molecules and the realization of real-time acquisition of mitochondrial activity are powerful tools in the study and treatment of mitochondria dysfunction in related diseases. Recent advances in mitochondria-targeting agents have led to several important mitochondria chemical probes that offer the opportunity for selective targeting molecules, novel biological applications and therapeutic strategies. This review details the structural and physiological functional characteristics of mitochondria, and comprehensively summarizes and classifies mitochondria-targeting agents. In addition, their pros and cons and their related chemical biological applications are discussed. Finally, the potential biomedical applications of these agents are briefly prospected.

Proteases and Their Modulators in Cancer Therapy: Challenges and Opportunities.

Proteostasis is the process of regulating intracellular proteins to maintain the balance of the cell proteome, which is crucial for cancer cell survival. Several proteases located in the cytoplasm, mitochondria, lysosome, and extracellular environment have been identified as potential antitumor targets because of their involvement in proteostasis. Although the discovery of small-molecule inhibitors targeting proteases faces particular challenges, rapid advances in chemical biology and structural biology, and the new technology of drug discovery have facilitated the development of promising protease modulators. In this review, the protein structure and function of important tumor-related proteases and their inhibitors are presented. We also provide a prospective on advances and the outlook of new drug strategies that target these proteases.

Navigating the Biological-Material Interface with the Guide of Chemical Biology.

Research at the biological-material interface often has translation in mind, with applications in medical implants, drug delivery, and regenerative medicine. While the clinical impact of this research is undeniable, a clearer picture of the in vivo behavior of materials is needed to address longstanding limitations in performance and function. Advances in chemical biology and biotechnology have propelled our understanding of how small molecules and biologics behave in living systems. Adapting these techniques to the study of synthetic materials, enabled by modern polymer chemistry, will bring molecular resolution to biological-material interactions and guide the development of next-generation biomaterials for therapeutic and diagnostic applications.

Recent Advances in Chemical Biology Using Benzophenones and Diazirines as Radical Precursors.

The use of light-activated chemical probes to study biological interactions was first discovered in the 1960s, and has since found many applications in studying diseases and gaining deeper insight into various cellular mechanisms involving protein–protein, protein–nucleic acid, protein–ligand (drug, probe), and protein–co-factor interactions, among others. This technique, often referred to as photoaffinity labelling, uses radical precursors that react almost instantaneously to yield spatial and temporal information about the nature of the interaction and the interacting partner(s). This review focuses on the recent advances in chemical biology in the use of benzophenones and diazirines, two of the most commonly known light-activatable radical precursors, with a focus on the last three years, and is intended to provide a solid understanding of their chemical and biological principles and their applications.

Photo-controllable bioorthogonal chemistry for spatiotemporal control of bio-targets in living systems.

The establishment of bioorthogonal chemistry is one of the most significant advances in chemical biology using exogenous chemistry to perturb and study biological processes. Photo-modulation of biological systems has realized temporal and spatial control on biomacromolecules in living systems. The combination of photo-modulation and bioorthogonal chemistry is therefore emerging as a new direction to develop new chemical biological tools with spatiotemporal resolution. This minireview will focus on recent development of bioorthogonal chemistry subject to spatiotemporal control through photo-irradiation. Different strategies to realize photo-control on bioorthogonal bond-forming reactions and biological applications of photo-controllable bioorthogonal reactions will be summarized to give a perspective on how the innovations on photo-chemistry can contribute to the development of optochemical biology. Future trends to develop more optochemical tools based on novel photochemistry will also be discussed to envision the development of chemistry-oriented optochemical biology.

All about that fat: Lipid modification of proteins in Cryptococcus neoformans.

Lipid modification of proteins is a widespread, essential process whereby fatty acids, cholesterol, isoprenoids, phospholipids, or glycosylphospholipids are attached to polypeptides. These hydrophobic groups may affect protein structure, function, localization, and/or stability; as a consequence such modifications play critical regulatory roles in cellular systems. Recent advances in chemical biology and proteomics have allowed the profiling of modified proteins, enabling dissection of the functional consequences of lipid addition. The enzymes that mediate lipid modification are specific for both the lipid and protein substrates, and are conserved from fungi to humans. In this article we review these enzymes, their substrates, and the processes involved in eukaryotic lipid modification of proteins. We further focus on its occurrence in the fungal pathogen Cryptococcus neoformans, highlighting unique features that are both relevant for the biology of the organism and potentially important in the search for new therapies.

Isolation of a non-genomic origin fluoroquinolone responsive regulatory element using a combinatorial bioengineering approach.

Advances in chemical biology have led to selection of synthetic functional nucleic acids for in vivo applications. Discovery of synthetic nucleic acid regulatory elements has been a long-standing goal of chemical biologists. Availability of vast genome level genetic resources has motivated efforts for discovery and understanding of inducible synthetic genetic regulatory elements. Such elements can lead to custom-design of switches and sensors, oscillators, digital logic evaluators and cell–cell communicators. Here, we describe a simple, robust and universally applicable module for discovery of inducible gene regulatory elements. The distinguishing feature is the use of a toxic peptide as a reporter to suppress the background of unwanted bacterial recombinants. Using this strategy, we show that it is possible to isolate genetic elements of non-genomic origin which specifically get activated in the presence of DNA gyrase A inhibitors belonging to fluoroquinolone (FQ) group of chemicals. Further, using a system level genetic resource, we prove that the genetic regulation is exerted through histone-like nucleoid structuring (H-NS) repressor protein. Till date, there are no reports of in vivo selection of non-genomic origin inducible regulatory promoter like elements. Our strategy opens an uncharted route to discover inducible synthetic regulatory elements from biologically-inspired nucleic acid sequences.

Chemical reporters for biological discovery

Functional tools are needed to understand complex biological systems. Here we review how chemical reporters in conjunction with bioorthogonal labeling methods can be used to image and retrieve nucleic acids, proteins, glycans, lipids and other metabolites in vitro, in cells as well as in whole organisms. By tagging these biomolecules, researchers can now monitor their dynamics in living systems and discover specific substrates of cellular pathways. These advances in chemical biology are thus providing important tools to characterize biological pathways and are poised to facilitate our understanding of human diseases.

Optogenetic pharmacology for control of native neuronal signaling proteins

The optical neuroscience revolution is transforming how we study neural circuits. By providing a precise way to manipulate endogenous neuronal signaling proteins, it also has the potential to transform our understanding of molecular neuroscience. Recent advances in chemical biology have produced light-sensitive compounds that photoregulate a wide variety of proteins underlying signaling between and within neurons. Chemical tools for optopharmacology include caged agonists and antagonists and reversibly photoswitchable ligands. These reagents act on voltage-gated ion channels and neurotransmitter receptors, enabling control of neuronal signaling with a high degree of spatial and temporal precision. By covalently attaching photoswitch molecules to genetically tagged proteins, the newly emerging methodology of optogenetic pharmacology allows biochemically precise control in targeted subsets of neurons. Now that the tools for manipulating endogenous neuronal signaling proteins are available, they can be implemented in vivo to enhance our understanding of the molecular bases of brain function and dysfunctions.

Chemical ‘omics’ approaches for understanding protein cysteine oxidation in biology

Oxidative cysteine modifications have emerged as a central mechanism for dynamic post-translational regulation of all major protein classes and correlate with many disease states. Elucidating the precise roles of cysteine oxidation in physiology and pathology presents a major challenge. This article reviews the current, targeted proteomic strategies that are available to detect and quantify cysteine oxidation. A number of indirect methods have been developed to monitor changes in the redox state of cysteines, with the majority relying on the loss of reactivity with thiol-modifying reagents or restoration of labeling by reducing agents. Recent advances in chemical biology allow for the direct detection of specific cysteine oxoforms based on their distinct chemical attributes. In addition, new chemical reporters of cysteine oxidation have enabled in situ detection of labile modifications and improved proteomic analysis of redox-regulated proteins. Progress in the field of redox proteomics should advance our knowledge of regulatory mechanisms that involve oxidation of cysteine residues and lead to a better understanding of oxidative biochemistry in health and disease.

Grand Challenge Commentary: Chemical transdifferentiation and regenerative medicine

The ability to alter cell identity with small molecules represents a powerful approach to restore biological function lost because of cellular deficiency. Developing this capability through advances in chemical biology could have an enormous impact on human health.

Fluorescent imaging of transition metal homeostasis using genetically encoded sensors

The ability to image the concentration of transition metals in living cells in real time is important for understanding transition metal (TM) homeostasis and its involvement in diseases. Genetically encoded fluorescent sensor proteins are attractive because they do not require cell-invasive procedures, can be targeted to different locations in the cell, and allow ratiometric detection. Important progress in the development of Zn(2+) sensors has allowed sensitive detection of the very low free concentrations of Zn(2+) in single cells, both in the cytosol and various organelles. Together with other recent advances in chemical biology, these tools seem particularly useful to interrogate the dynamics and compartmentation of TM homeostasis.

Chemical tools for understanding protein lipidation in eukaryotes

Lipidation of proteins is an important mechanism to regulate protein trafficking and activity in cell and tissues. The targeting of proteins to membranes by lipidation plays key roles in many physiological processes and when not regulated properly can lead to cancer and neurological disorders. Dissecting the precise roles of protein lipidation in physiology and disease is a major challenge. Recent advances in chemical biology have now enabled the semisynthesis of lipidated proteins for fundamental biochemical and cellular studies. In addition, new chemical reporters of protein lipidation have improved the detection and enabled the proteomic analysis of lipidated proteins. The expanding efforts in chemical biology are therefore providing new tools to dissect the mechanisms and functions of protein lipidation as well as develop therapeutics targeted at protein lipidation pathways in disease.