Accelerating PROTACs Discovery Through a Direct-to-Biology Platform Enabled by Modular Photoclick Chemistry.

Proteolysis targeting chimeras (PROTACs) have emerged as a promising strategy for drug discovery and exploring protein functions, offering a revolutionary therapeutic modality. Currently, the predominant approach to PROTACs discovery mainly relies on an empirical design-synthesis-evaluation process involving numerous cycles of labor-intensive synthesis-purification and bioassay data collection. Therefore, the development of innovative methods to expedite PROTAC synthesis and exploration of chemical space remains highly desired. Here, a direct-to-biology strategy is reported to streamline the synthesis of PROTAC libraries on plates, enabling the seamless transfer of reaction products to cell-based bioassays without the need for additional purification. By integrating amide coupling and light-induced primary amines and o-nitrobenzyl alcohols cyclization (PANAC) photoclick chemistry into a plate-based synthetic process, this strategy produces PROTAC libraries with high efficiency and structural diversity. Moreover, by employing this platform for PROTACs screening, we smoothly found potent PROTACs effectively inhibit triple-negative breast cancer (TNBC) cell growth and induce rapid, selective targeted degradation of cyclin-dependent kinase 9 (CDK9). The study introduces a versatile platform for assembling PROTACs on plates, followed by direct biological evaluation. This approach provides a promising opportunity for high-throughput synthesis of PROTAC libraries, thereby enhancing the efficiency of exploring chemical space and accelerating the discovery of PROTACs.

Spatiotemporal and global profiling of DNA-protein interactions enables discovery of low-affinity transcription factors.

Precise dissection of DNA-protein interactions is essential for elucidating the recognition basis, dynamics and gene regulation mechanism. However, global profiling of weak and dynamic DNA-protein interactions remains a long-standing challenge. Here, we establish the light-induced lysine (K) enabled crosslinking (LIKE-XL) strategy for spatiotemporal and global profiling of DNA-protein interactions. Harnessing unique abilities to capture weak and transient DNA-protein interactions, we demonstrate that LIKE-XL enables the discovery of low-affinity transcription-factor/DNA interactions via sequence-specific DNA baits, determining the binding sites for transcription factors that have been previously unknown. More importantly, we successfully decipher the dynamics of the transcription factor subproteome in response to drug treatment in a time-resolved manner, and find downstream target transcription factors from drug perturbations, providing insight into their dynamic transcriptional networks. The LIKE-XL strategy offers a complementary method to expand the DNA-protein profiling toolbox and map accurate DNA-protein interactomes that were previously inaccessible via non-covalent strategies, for better understanding of protein function in health and disease.

Light-induced primary amines and o-nitrobenzyl alcohols cyclization as a versatile photoclick reaction for modular conjugation.

The advent of click chemistry has had a profound impact on many fields and fueled a need for reliable reactions to expand the click chemistry toolkit. However, developing new systems to fulfill the click chemistry criteria remains highly desirable yet challenging. Here, we report the development of light-induced primary amines and o-nitrobenzyl alcohols cyclization (PANAC) as a photoclick reaction via primary amines as direct click handle, to rapid and modular functionalization of diverse small molecules and native biomolecules. With intrinsic advantages of temporal control, good biocompatibility, reliable chemoselectivity, excellent efficiency, readily accessible reactants, operational simplicity and mild conditions, the PANAC photoclick is robust for direct diversification of pharmaceuticals and biorelevant molecules, lysine-specific modifications of unprotected peptides and native proteins in vitro, temporal profiling of endogenous kinases and organelle-targeted labeling in living systems. This strategy provides a versatile platform for organic synthesis, bioconjugation, medicinal chemistry, chemical biology and materials science.