Automated, Accelerated Nanoscale Synthesis of Iminopyrrolidines

Miniaturization and acceleration of synthetic chemistry is an emerging area in pharmaceutical, agrochemical, and materials research and development. Herein, we describe the synthesis of iminopyrrolidine-2-carboxylic acid derivatives using chiral glutamine, oxo components, and isocyanide building blocks in an unprecedented Ugi-3-component reaction. We used I-DOT, a positive-pressure-based low-volume and non-contact dispensing technology to prepare more than 1000 different derivatives in a fully automated fashion. In general, the reaction is stereoselective, proceeds in good yields, and tolerates a wide variety of functional groups. We exemplify a pipeline of fast and efficient nanomole-scale scouting to millimole-scale synthesis for the discovery of a useful novel reaction with great scope

Sustainability by design:Automated nanoscale 2,3,4-trisubstituted quinazoline diversity

Small molecule synthesis is equally important for materials and pharmaceuticals. However, the traditional approach performed in pharmaceutical R&D including maintenance of million-sized libraries and optimization synthesis of hundreds or even thousands of chemicals on a mmol or larger scale lacks sustainability. Here, we exemplify the synthetic execution of a newly designed quinoxaline reaction towards a transformative sustainability in chemistry. This includes nanoscale synthesis, deep chemical space exploration, scalability over 6 orders of magnitude from milligram up to 10-gram resynthesis of quinazolines enabled by the simultaneous variation of four classes of building blocks. Benefits of our approach include a simple to perform, one-step procedure, mild reaction conditions and access to a very large chemical space through accessing many available building blocks. More than thousand derivatives were produced in an automated fashion on a nanoscale using positive pressure facilitated dispensing. Along with these advantages, there is a considerable reduction in synthetic effort, reagents, solvent, glass and plastic consumables and power consumption to decrease the footprint of synthetic chemistry

Multicomponent reaction-derived covalent inhibitor space

The area of covalent inhibitors is gaining momentum due to recently introduced clinical drugs, but libraries of these compounds are scarce. Multicomponent reaction (MCR) chemistry is well known for its easy access to a very large and diverse chemical space. Here, we show that MCRs are highly suitable to generate libraries of electrophiles based on different scaffolds and three-dimensional shapes and highly compatible with multiple functional groups. According to the building block principle of MCR, acrylamide, acrylic acid ester, sulfurylfluoride, chloroacetic acid amide, nitrile, and alpha,beta-unsaturated sulfonamide warheads can be easily incorporated into many different scaffolds. We show examples of each electrophile on 10 different scaffolds on a preparative scale as well as in a high-throughput synthesis mode on a nanoscale to produce libraries of potential covalent binders in a resource-and time-saving manner. Our operational procedure is simple, mild, and step economical to facilitate future covalent library synthesis

PROTACs- a game-changing technology

Introduction: Proteolysis - targeting chimeras (PROTACs) have emerged as a new modality with the potential to revolutionize drug discovery. PROTACs are heterobifunctional molecules comprising of a ligand targeting a protein of interest, a ligand targeting an E3 ligase and a connecting linker. The aim is instead of inhibiting the target to induce its proteasomal degradation. Areas covered: PROTACs, due to their bifunctional design, possess properties that differentiate them from classical inhibitors. A structural analysis, based on published crystal aspects, kinetic features and aspects of selectivity are discussed. Specific types such as homoPROTACs, PROTACs targeting Tau protein and the first PROTACs recently entering clinical trials are examined. Expert opinion: PROTACs have shown remarkable biological responses in challenging targets, including an unprecedented selectivity over protein family members and even efficacy starting from weak or unspecific binders. Moreover, PROTACs are standing out from classical pharmacology by inducing the degradation of the target protein and not merely its inhibition. However, there are also challenges in the field, such as the rational structure optimization, the evolution of computational tools, limited structural data and the greatly anticipated clinical data. Despite the remaining hurdles, PROTACs are expected to soon become a new therapeutic category of drugs

Direct-to-biology, automated, nano-scale synthesis, and phenotypic screening-enabled E3 ligase modulator discovery

Thalidomide and its analogs are molecular glues (MGs) that lead to targeted ubiquitination and degradation of key cancer proteins via the cereblon (CRBN) E3 ligase. Here, we develop a direct-to-biology (D2B) approach for accelerated discovery of MGs. In this platform, automated, high throughput, and nano scale synthesis of hundreds of pomalidomide-based MGs was combined with rapid phenotypic screening, enabling an unprecedented fast identification of potent CRBN-acting MGs. The small molecules were further validated by degradation profiling and anti-cancer activity. This revealed E14 as a potent MG degrader targeting IKZF1/3, GSPT1 and 2 with profound effects on a panel of cancer cells. In a more generalized view, integration of automated, nanoscale synthesis with phenotypic assays has the potential to accelerate MGs discovery.</p

Isolation and molecular characterization of novel glucarpidases:Enzymes to improve the antibody directed enzyme pro-drug therapy for cancer treatment

<div><p>Repeated cycles of antibody-directed enzyme pro-drug therapy (ADEPT) and the use of glucarpidase in the detoxification of cytotoxic methotrexate (MTX) are highly desirable during cancer therapy but are hampered by the induced human antibody response to glucarpidase. Novel variants of glucarpidase (formal name: carboxypeptidase G2, CPG2) with epitopes not recognized by the immune system are likely to allow repeated cycles of ADEPT for effective cancer therapy. Towards this aim, over two thousand soil samples were collected and screened for folate hydrolyzing bacteria using folate as the sole carbon source. The work led to the isolation and the characterization of three new glucarpidase producing strains, which were designated as: <i>Pseudomonas lubricans</i> strain SF168, <i>Stenotrophomonas</i> sp SA and <i>Xenophilus azovorans</i> SN213. The <i>CPG2</i> genes of <i>Xenophilus azovorans</i> SN213 (named <i>Xen CPG2</i>) and <i>Stenotrophomonas sp</i> SA (named <i>Sten CPG2</i>) were cloned and molecularly characterized. Both Xen CPG2 and Sten CPG2 share very close amino acid sequences (99%); we therefore, focused on the study of Xen CPG2. Finally, we demonstrated that a polyclonal antibody raised against our new CPG2, Xen CPG2, does not react with the CPG2 from <i>Pseudomonas sp</i>. strain RS-16 (Ps CPG2) that are currently in clinical use. The two enzymes, therefore could potentially be used consecutively in the ADEPT protocol to minimize the effect of the human antibody response that hampers current treatment with Ps CPG2. The identified novel CPG2 in this study will, therefore, pave the way for safer antibody directed enzyme pro-drug therapy for cancer treatment.</p></div

Direct-to-biology, automated, nano-scale synthesis, and phenotypic screening-enabled E3 ligase modulator discovery

Thalidomide and its analogs are molecular glues (MGs) that lead to targeted ubiquitination and degradation of key cancer proteins via the cereblon (CRBN) E3 ligase. Here, we develop a direct-to-biology (D2B) approach for accelerated discovery of MGs. In this platform, automated, high throughput, and nano scale synthesis of hundreds of pomalidomide-based MGs was combined with rapid phenotypic screening, enabling an unprecedented fast identification of potent CRBN-acting MGs. The small molecules were further validated by degradation profiling and anti-cancer activity. This revealed E14 as a potent MG degrader targeting IKZF1/3, GSPT1 and 2 with profound effects on a panel of cancer cells. In a more generalized view, integration of automated, nanoscale synthesis with phenotypic assays has the potential to accelerate MGs discovery.</p

Direct-to-biology, automated, nano-scale synthesis, and phenotypic screening-enabled E3 ligase modulator discovery

Thalidomide and its analogs are molecular glues (MGs) that lead to targeted ubiquitination and degradation of key cancer proteins via the cereblon (CRBN) E3 ligase. Here, we develop a direct-to-biology (D2B) approach for accelerated discovery of MGs. In this platform, automated, high throughput, and nano scale synthesis of hundreds of pomalidomide-based MGs was combined with rapid phenotypic screening, enabling an unprecedented fast identification of potent CRBN-acting MGs. The small molecules were further validated by degradation profiling and anti-cancer activity. This revealed E14 as a potent MG degrader targeting IKZF1/3, GSPT1 and 2 with profound effects on a panel of cancer cells. In a more generalized view, integration of automated, nanoscale synthesis with phenotypic assays has the potential to accelerate MGs discovery.</p

Nanoscale, automated, high throughput synthesis and screening for the accelerated discovery of protein modifiers

Hit finding in early drug discovery is often based on high throughput screening (HTS) of existing and historical compound libraries, which can limit chemical diversity, is time-consuming, very costly, and environmentally not sustainable. On-the-fly compound synthesis and in situ screening in a highly miniaturized and automated format has the potential to greatly reduce the medicinal chemistry environmental footprint. Here, we used acoustic dispensing technology to synthesize a library in a 1536 well format based on the Groebcke–Blackburn–Bienaymé reaction (GBB-3CR) on a nanomole scale. The unpurified library was screened by differential scanning fluorimetry (DSF) and cross-validated using microscale thermophoresis (MST) against the oncogenic protein–protein interaction menin–MLL. Several GBB reaction products were found as μM menin binder, and the structural basis of the interactions with menin was elucidated by co-crystal structure analysis. Miniaturization and automation of the organic synthesis and screening process can lead to an acceleration in the early drug discovery process, which is an alternative to classical HTS and a step towards the paradigm of continuous manufacturing