MedChemCASES | Gesellschaft Deutscher Chemiker e.V. (2023)

Dr. Benjamin Horning
April 26, 2023
5PM (Berlin time, CET)

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Industrialized chemoproteomics enables the discovery of allosteric modulators of Janus Kinase 1 and Cereblon targeting novel sites

Interest in covalent inhibitors has seen a resurgence in recent years, owing in large part to proteomics technology to assess selectivity and de-risk off-target concerns. We have utilized our proteomics platform to guide the development of a screening library containing small molecules armed with covalent reactive groups. From these efforts we have discovered inhibitors of Janus kinase 1 (JAK1) that potently and selectively inhibit JAK1 mediated cytokine signaling through a novel allosteric pocket on the pseudokinase domain, distinct from the pocket bound by recent clinical stage inhibitors of the related kinase Tyk2. We demonstrate that our inhibitors function via inhibiting trans-phosphorylation of JAK1 via its heterodimeric JAK family member, a key step in the signaling cascade. These covalent, allosteric JAK1 inhibitors avoid the selectivity issues common to traditional kinase inhibitors, which bind to highly conserved kinase active sites. Moreover, since the target cysteine in JAK1 is not highly conserved, they have the potential to avoid potential clinical safety issues associated with inhibiting off targets such as JAK2. In addition, we have discovered a novel pocket on the E3 ligase Cereblon that can be targeted by covalent molecules. Binding to this pocket allosterically prevents imide binding, and the allosteric relationship between these two sites can be utilized to monitor imide binding to Cereblon in bifunctional molecules.

Velcrin compounds are small molecules which act as molecular glues to form a PDE3A-SLFN12 complex. This complex formation induces apoptosis in cancer cells expressing elevated levels of PDE3A and SLFN12 (de Waal et al., Nat Chem Biol 2016; Wu et al., J Bio Chem 2020). Unlike traditional targeted therapies that leverage dependencies created by genomic alterations in cancer cells, velcrins instead kill cancer cells by a gain-of-function mechanism dependent on stimulation of SLFN12 RNase activity by complex formation with PDE3A (Garvie et al., Nat Commun 2021).

Based on in-vivo tool compounds which were discovered by hit-to-lead optimization from a phenotypic screen (Lewis et al., ACS Med Chem Lett 2019), the team set out to optimize the properties of the tool compounds. While already showing excellent potency and in-vivo efficacy in various tumor models, the compounds were only approximately tenfold selective against PDE3A when compared to their cell killing activity. PDE3A/B activity most notably influences cardiohemodynamics and inhibition of PDE3A/B was to be reduced to generate a larger safety window. Co-crystallization of analogs with PDE3A showed the binding mode of the velcrin class and changes in the solvent exposed region of the PDE3A binders led to steep activity changes in complex formation and cell killing. Careful optimization of the ratio of cell killing to PDE3A activity while optimizing DMPK and safety pharmacology led to the discovery of BAY 2666605.

BAY 2666605 has recently entered a First-in-Human study (NCT04809805) in patients with advanced solid tumors that co-express PDE3A and SLFN12, including melanoma, ovarian cancer, and sarcoma. We will discuss the discovery and structure activity relationships of BAY 2666605.

Prof. Dr. Anna K.H. Hirsch (Helmholtz Institut, Saarland)
December 14, 2022
5:00 PM (Berlin time)

Addressing unusual anti-infective targets

The challenges associated with anti-infective drug-discovery programmes can be tackled by combining several established and unprecedented hit-identification strategies with phenotypic antibacterial screening. I will illustrate this approach with a selection of un(der)explored targets. The first is a vitamin transporter from the energy-coupling factor (ECF) class, which is unique to Gram-positive bacteria. Here, we report on the structure-based virtual screening (SBVS), design, synthesis and structure–activity relationships of the first classes of selective, antibacterial inhibitors of the energycoupling factor (ECF) transporters with good in vitro and whole-cell activity and a good in vitro ADMET and in vivo PK profiles. A newly established cell-based uptake assay in Lactobacillus casei greatly facilitated our screening and hit-to-lead optimisation campaign. The second is the β-subunit of the bacterial DNA polymerase III (sliding clamp, DnaN), an attractive antibacterial target. We pursued several hit-identification strategies, including a SBVS campaign, affording novel chemotypes with micromolar affinity and promising antibacterial activity. Mode-ofaction studies confirmed DnaN as the molecular target. The new compound displays broad-spectrum antibacterial activity against mycobacteria, Gram-positive and Gram-negative pathogens also against multidrug-resistant bacteria with no cytotoxicity and good in vivo PK profiles. Finally, we succeeded in fragment merging and linking, affording highly selective and potent inhibitors of the extracellular metalloprotease and virulence factor of Pseudomonas aeruginosa, the elastase LasB. Multiparameter optimisation is currently ongoing based on extensive in vitro and ex vivo profiling, including the establishment of complex biological assays. Our approach promises to deliver the urgently needed anti-infective agents featuring both new chemical scaffolds and unprecedented modes of action. Multiparameter optimisation is currently ongoing based on extensive in vitro, wholecell, ex vivo and in vivo profiling, including the establishment of complex biological assays. A particular emphasis will be placed on the lead optimisation of frontrunners for permeation and achieving good lung exposure.

Dr. Kai Schiemann (Merck KGaA)
October 26, 2022
5:00 PM (Berlin time)

Discovery of M1069, a highly selective dual inhibitor of adenosine A2a/A2b receptors

While the A2A adenosine receptor has been considered a major contributor to adenosine mediated tumor immunosuppression, the A2B receptor has recently emerged as another potential therapeutic target. Considering the importance of both targets, we developed M1069 as a selective dual antagonist of the A2A and A2B adenosine receptors to counteract the immunosuppressive effects of adenosine signaling and stimulate anti-tumor immune responses in patients with advanced malignancies.

M1069 was discovered through the medicinal chemistry optimization of a thiazolo[4,5-c]pyridin-2-amine scaffold. The binding mode was confirmed by X-ray structure analysis of close derivatives of M1069 in a complex with the A2A receptor domain. Initial cellular optimization resulted in a very high A2AR potency and a low potency for A2BR. Homology models of the A2BR, along with docking into the published A1R, supported further optimization. Increasing the potency on A2BR was a major challenge, which required some fine-tuning of the substituents, while keeping a high split ratio to the A1R, which was defined as an exclusion criterion to avoid unwanted side-effects. Finally, potent dual and selective A2A/A2B antagonists were further profiled. Surprisingly, some of these showed species’ differences in human and murine T cells, as well as myeloid cells at high (10 µM) NECA (stable analog of adenosine) concentrations and needed to be excluded from the evaluation of in vivo pharmacology using murine tumor models. In contrast, M1069 demonstrated dual A2A and A2B antagonist activity in assays with both human and murine immune cells, favorable physchem/ADME properties, good PK in various species and predicted minimal brain penetration in humans.

Preclinically, M1069 has demonstrated superior activity in comparison to A2A-selective reference antagonists in high (10uM) NECA/adenosine settings in vitro, which translated into superior anti-tumor activity for M1069 in vivo in the 4T1 tumor model. In addition, M1069 showed combination activity with the chemotherapeutic agents’ cisplatin and paclitaxel in the 4T1 tumor model. These preclinical data are consistent with a primary mechanism of action of M1069 that involves the reversal of immune suppression via dual antagonism of pro-tumorigenic A2A and A2B adenosine receptors in an adenosine rich TME, where A2B could act complementary, or compensatory to A2A. Together, this data suggests higher potential for dual A2A/A2B antagonism of M1069 compared to A2A antagonism to block the immune-suppressive/pro-tumorigenic impact of adenosine.

M1069 is currently being evaluated as a monotherapy in a Phase I, first-in-human clinical trial in patients with advanced solid tumors (NCT05198349).

Dr. Nis Halland (Sanofi)
September 22, 2022
5:00 PM (Berlin time)

Rational design and optimization of SGK1 inhibitors for the treatment osteoarthritis
The serine/threonine kinase SGK1 is an activator of the β-catenin pathway and a powerful stimulator of cartilage degradation that is found to be upregulated under genomic control in diseased osteoarthritic cartilage. Today no oral disease-modifying treatments are available and chronic treatment in this indication sets high requirements for the drug selectivity, pharmacokinetic and safety profile. We describe the preparation of a highly selective druglike 1H-pyrazolo[3,4-d]pyrimidine SGK1 inhibitor that matches both safety and pharmacokinetic requirements for oral dosing. Rational compound design was facilitated by a novel hSGK1 cocrystal structure and classic medicinal chemistry was applied to guide the chemical optimization of ADMET and selectivity profiles.

Discovery of JDQ443, a structurally novel, potent and selective covalent oral inhibitor of KRAS^G12C
RAS is the most frequently mutated oncogene in cancer, with KRAS G12C mutations most commonly found in lung adenocarcinoma, colorectal cancer, and other solid tumor malignancies. Covalent inhibitors of KRASG12C have shown antitumor activity against advanced/metastatic KRAS G12C-mutated cancers. Here we report the identification of a new class of pyrazole based KRASG12C inhibitors discovered by structure based de-novo design. The compounds bind to the KRASG12C switch II pocket with a novel binding mode, exploiting unique interactions with the GDP-bound form of the KRASG12C protein. We describe the hit to lead and lead optimization by structure-based design of the novel chemical series leading to the discovery of NVP-JDQ443 (JDQ443), a novel, potent and selective, orally bioavailable KRASG12C covalent inhibitor. JDQ443 showed dose dependent antitumor efficacy in KRAS G12C-mutated cell-derived models and is currently in clinical development as monotherapy as well as in combination with TNO155, a SHP2 inhibitor (NCT04699188).

Dr. Tim Owens (Principia - Sanofi)
May 24, 2022
5:00 PM (Berlin time)

Development of Reversible Covalent Inhibitors as Chemical Probes and Clinical Candidates
In 2013,Serafimova assessed a series of electrophilic molecules that engaged non-catalytic cysteines in a covalent manner, but with variable target residence times, driven by the various stabilizing interactions of the molecule in the protein binding site. These reversible covalent inhibitors were shown to rapidly dissociate from common thiols while maintaining sustained inhibition of a target protein even after washout. This technology, therefore, has the potential to allow medicinal chemists to fashion inhibitors with the potential for selectivity gained by inhibiting only targets with a conserved cysteine, and minimal exposure requirements inherent with covalent inhibition. The reversible covalent approach furthermore mitigates risk of irreversible binding to non-target cysteines or endogenous thiols, but also allows the potential to improve selectivity among targets sharing a common cysteine. This technology has been used to identify selective inhibitors of kinases such as BTK and FGFR as well as LMP7 subunit of the immunoproteasome. Based on our interest in BTK inhibition for the treatment of immune mediated disorders, we embarked on a drug development effort to identify reversible covalent inhibitors culminating in the identification of PRN473 for topical administration and PRN1008 (rilzabrutinib) as an oral BTK inhibitor currently in multiple phase 3 studies.

High-throughput PROTAC synthesis for direct, biological assaying
Direct-to-biology (D2B) synthesis and assaying has the potential to cut drug-discovery cycle times by providing information rich experiments in a highly efficient manner. For PROTACs, this cycle time is often slower than conventional medicinal chemistry as there are a plethora of small molecule E3-ligase or protein-of-interest binders and vectors to investigate as well as a vast landscape of linker chemical space to explore; finding the optimum for these three individual components is time-consuming and resource intensive. An attractive alternative would be to conduct miniaturized reactions in 1536-well plates, where all three PROTAC components are simultaneously evaluated, and directly assessed as crude reactions in cell-based HiBiT assays. Judicious reagents choice is required to ensure high reaction conversion and, moreover, minimize any effect on cell-viability. This talk will showcase our Direct-to-biology protocol to assess crude PROTACs using two well established PROTAC target case-studies: HER2 and BRD4.

Dr. David Thaisrivongs (MSD)
February 23, 2022
5:00 PM (Berlin time)
Synthetic macrocyclic peptides that target protein-protein interactions: the discovery and early chemistry development of a PCSK9 inhibitor
Inhibition of Proprotein Convertase Subtilisin/Kexin type-9 (PCSK9) improves cardiovascular outcomes in patients requiring additional low-density lipoprotein cholesterol reduction on top of statins, but there has been limited uptake of the two commercialized PCSK9 antibody inhibitors due in part to cost and route of administration. The protein-protein interaction between PCSK9 and the low-density lipoprotein receptor which is targeted by these therapies is a large, flat surface, which has made the discovery of orally-bioavailable small molecule inhibitors highly challenging. This talk will introduce the use of macrocyclic peptides to interrupt such protein-protein interactions, and describe the discovery and early chemistry development of such a PCSK9 inhibitor.

Name: Stephen Fesik (Vanderbilt University School of Medicine, Nashville, TN, USA)
Date: October 21, 2021, 4:00 pm
Chairs:Chistian Kuttruff,Julien Lefranc
Co-Chair:Franz von Nussbaum

Title:The Discovery of Potent WDR5 Inhibitors for the Treatment of Cancer
WDR5 is a component of multiple epigenetic regulatory complexes, including the mixed lineage leukemia (MLL)/SET complexes that deposit histone H3 lysine 4 methylation. Overexpression of WDR5 correlates with a poor clinical outcome in many human cancers, and WDR5 has emerged as an attractive target for therapy. We discovered potent WDR5 inhibitors using fragment-based methods and structure-based design. These inhibitors have been optimized for their drug-like properties and show potent oral activity in tumor xenograft mouse models. They act not by altering histone methylation but by displacing WDR5 from chromatin at protein synthesis genes, choking the translational capacity of these cells, and inducing death via a nucleolar stress response. Our inhibitors could thus have broad therapeutic utility as anti-cancer agents.

Name: Timo Heinrich (Merck KGaA, Darmstadt)
Date : September 23, 2021, 4:00 pm
Chair: Tatjana Ross, Franca Klingler
Co-Chair: Franz von Nussbaum

Title:Discovery and Optimization of Next Generation Reversible Methionine Aminopeptidase-2 Inhibitors – Identification of clinical compound M8891
Co- and post-translational protein processing is important to ensure the maturation tofunctional proteins. MetAP-2 proteolytically removes the amino-terminal methioninefrom nascent proteins, and inhibition of its activity has been shown to blockangiogenesis and tumor growth, suggesting that small-molecule inhibitors of MetAP-2may be promising options for the treatment of cancer. This talk describes how
persistence and serendipity helped to identify a previously unknown MetAP-2 inhibitoryscaffold. The biochemical activity of more conventional, purine base compounds didnot translate to cellular activity. Thorough analysis of HTS screening results led tothe identification of chiral tartronic diamide hits. Structure-based hit-to-lead optimizationis out-lined, including different synthetic strategies for most efficient access to improvedderivatives as well as asymmetric approaches. The initial lead compound suffered fromenterohepatic circulation, preventing further development. Binding kinetics wereinvestigated by fluorescence cross correlation spectroscopy and residence time on thetarget protein was one important characterization criterion. Adjustment of H-bonddonor/acceptor count and multiparameter analysis of the compound properties led tothe nomination of the clinical development compound M8891, which impedes thegrowth of primary endothelial cells, shows dose-dependent biomarker modulation andantitumoral activity in mouse xenograft models.

Stefan Knapp (Universität Frankfurt)
June 24, 2021
4:00 pm (Berlin time)
Chairs: Matthias Gehringer, Eleonora Diamanti
Co-Chair: Franz von Nussbaum

Title: Targeting protein scaffolding function in kinases
In living cells, protein kinases are organized in large signalling complexes comprising adapter proteins, diverse enzymes and regulatory proteins. In recent years it has become increasingly evident, that protein kinases act not only as independent enzymes but that they also function as protein interaction scaffolds organizing the assembly of signalling complexes in a conformation sensitive way. This complexity is also reflected by the response of kinases to inhibitors that may stabilize diverse conformations acting as inhibitors of enzymatic activity only, as modulators of kinase scaffolding roles or both. In this talk, I will exemplify the implications of altering protein interactions by allosteric small molecules as well as canonical ATP competitive inhibitors using a number of selective inhibitors that we developed recently. I will demonstrate how different binding modes that alter protein conformation and dynamics in a distinct way may result in diverse signalling outcomes and phenotypic responses. The important scaffolding roles of protein kinases will also enable targeting new and so far poorly explored members of the kinase family such as catalytically inactive pseudokinases, that represent a considerable number of largely unexplored kinase targets which have been linked to the development of many diseases.

Postponed dueto unforeseen events. The webinar will be rescheduled at a later date. We apologize for any inconvenience.

Cristina Nevado (University of Zurich, Switzerland)
Chairs: María Méndez Pérez, Franca Klingler
Co-Chair: Franz von Nussbaum

"Exploring the chemistry and biology of CREBBP and EP300 Bromodomains"
Expanding the chemical space and simultaneously ensuring synthetic accessibility is of upmost importance, not only for the discovery of effective binders for novel protein classes but, more importantly, for the development of compounds against hard-to-drug proteins. In this talk we will introduce AutoCouple, a de novo approach to computational ligand design focused on the diversity-oriented generation of chemical entities via virtual couplings. In a benchmark application, chemically diverse compounds with low-nanomolar potency for the CBP bromodomain and high selectivity against the BRD4(1) bromodomain were achieved by the synthesis of about 50 derivatives of the original fragment. The binding mode was confirmed by X-ray crystallography, target engagement in cells was demonstrated, and antiproliferative activity was showcased in three cancer cell lines.