In the complex network of cellular physiology, the maintenance of cellular proteostasis emerges as a critical factor for cell survival, particularly under stress conditions. This homeostasis is largely governed by a sophisticated network of molecular chaperones and co-chaperones, among which Bcl-2-associated athanogene 3 (BAG3), able to interact with the ATPase domain of Heat Shock Protein 70 (HSP70), plays a pivotal role. The BAG3-HSP70 functional module is not only essential for cellular homeostasis but is also involved in the pathogenesis of various diseases, including cancer, neurodegenerative disorders, and cardiac dysfunction, making it an attractive target for therapeutic intervention. Inspired by our continuous interest in the development of new chemical platforms able to interfere with BAG3 protein, herein we report the discovery of compound 16, the first-in-class BAG3/HSP70 dual modulator, obtained by combining the multicomponent Ugi reaction with the alkyne-azide Huisgen procedure in a sequential tandem reaction approach. Through a combination of biophysical analysis, biochemical assays, and cell-based studies, we elucidated the mechanism of action of this inhibitor and assessed its potential as a therapeutic agent. Hence, this study can open new avenues for the development of novel anticancer strategies that leverage the simultaneous disruption of multiple chaperone pathways.

The antiapoptotic BAG3 protein plays a crucial role in cellular proteostasis and it is involved in several signalling pathways governing cell proliferation and survival. Owing to its multimodular structure, it possesses an extensive interactome including the molecular chaperone HSP70 and other specific cellular partners, which make it an eminent factor in several pathologies, particularly in cancer. Despite its potential as a therapeutic target, very few BAG3 modulators have been disclosed so far. Here we describe the identification of a promising BAG3 modulator able to bind the BAG domain of the protein featuring an imidazopyridine scaffold and obtained through the application of the Groebke–Blackburn–Bienaymé chemical synthesis procedure. The disclosed compound 10 showed a relevant cytotoxic activity, and in line with the biological profile of BAG3 disruption, it induced the activation of caspase 3 and 9.

Screening libraries of low molecular weight (LMW) compounds are foundational in early-stage drug discovery. However, the parallel synthesis of milligram-scale large libraries – especially for covalent inhibitors – remains constrained by resource-intensive purification protocols, high solvent consumption, and limited scalability. Electrophilic fragments such as acrylamides are widely used to target nucleophilic cysteines in disease-relevant proteins, but practical workflows to synthesize diverse covalent libraries in sufficient quantities and quality are lacking. Here, we report a high-throughput, chromatography-free workflow to synthesize and analyze 1235 acrylamides using a modified Ugi four-component reaction (U-4CR) featuring ammonia and acrylic acid. This strategy exploits direct product precipitation to replace chromatographic purification steps with simple filtration, enabling synthesis at scale with common parallel lab instrumentation. Our protocol provides a sustainable and modular solution to library generation, especially suitable for covalent drug discovery. Using complementary analytical methods – ¹H NMR, UPLC-MS-UV, and AEMS – we systematically evaluated compound purity, identity, and long-term stability. The large dataset was systematically processed using PeakSel® (live dashboard at https://peaksel.elsci.io/a/domling_group/post/8gY7CqUMVtb), providing complementary information on compound quality and long-term stability. This platform delivers the largest fully profiled covalent electrophile library to date and offers a generalizable framework for sustainable library production and high-content analytical workflows. It complements emerging analytics-only approaches by offering a complete synthesis-to-analysis pipeline accessible with standard laboratory instrumentation.

Miniaturized chemical synthesis through acoustic droplet ejection (ADE) is transforming reaction discovery by reducing material consumption and enabling highly parallelized, autonomous workflows. However, its application in organic synthesis has remained largely confined to DMSO and aqueous media, restricting solvent exploration and limiting broader process innovation. Often attributed to the intrinsic physics of acoustic transfer, these constraints may instead reflect historical calibration practices. Here, we establish a data-driven framework for expanding ADE-compatible solvent space toward a more applicable nanoscale reaction chemistry. Through systematic gravimetric analysis of 19 high-boiling organic solvents, including water and DMSO as reference fluids, we identify a broad operating window that supports accurate (deviation < 10%) and reproducible (RSD < 8%) dispensing using existing instrumentation. We further validate this framework via automated assembly of a 384-member Suzuki–Miyaura reaction library, where testing of N,N-dimethylformamide and a bio-based alternative solvent dimethyl isosorbide resulted in broad product formation across diverse substrates. This work decouples solvent choice from historical conventions and opens the route to a reaction-agnostic, solvent-flexible, and sustainable approach for high-throughput reaction discovery, enabling ADE workflows to transition from condition-constrained screening tools to general engines for sustainable, data-driven chemical exploration.