Nuvisan (Germany)

Additional therapeutic options are needed for patients with lung cancer with HER2 activating mutations, including exon 20 insertions. Sevabertinib shows activity against ERBB2-encoded HER2 exon 20 insertions in preclinical models of lung cancer, corroborated by early data from a phase 1/2 clinical trial.

In this manuscript, we summarize the discussions and key messages from developed in the e-environment team of the European Bioanalysis forum, which were the basis of a subsequent workshop on cloud applications in a regulated bioanalysis lab environment, hosted by the European Bioanalysis Forum e-environment team at their 16th Open Symposium in Barcelona, Spain in November 2023. The purpose of our discussions is to provide further insight and understanding on the status of having cloud applications implemented in a regulated bioanalysis laboratory and the challenges experienced. The discussions highlight the importance of cross functional collaboration during the entire process of cloud implementation and some of the uncertainties in the different functions' roles and responsibilities.

While grouping/read-across is widely used to fill data gaps, chemical registration dossiers are often rejected due to weak category justifications based on structural similarity only. Metabolomics provides a route to robust chemical categories via evidence of shared molecular effects across source and target substances. To gain international acceptance, this approach must demonstrate high reliability, and best-practice guidance is required. The MetAbolomics ring Trial for CHemical groupING (MATCHING), comprising six industrial, government and academic ring-trial partners, evaluated inter-laboratory reproducibility and worked towards best-practice. An independent team selected eight substances (WY-14643, 4-chloro-3-nitroaniline, 17α-methyl-testosterone, trenbolone, aniline, dichlorprop-p, 2-chloroaniline, fenofibrate); ring-trial partners were blinded to their identities and modes-of-action. Plasma samples were derived from 28-day rat tests (two doses per substance), aliquoted, and distributed to partners. Each partner applied their preferred liquid chromatography–mass spectrometry (LC–MS) metabolomics workflows to acquire, process, quality assess, statistically analyze and report their grouping results to the European Chemicals Agency, to ensure the blinding conditions of the ring trial. Five of six partners, whose metabolomics datasets passed quality control, correctly identified the grouping of eight test substances into three categories, for both male and female rats. Strikingly, this was achieved even though a range of metabolomics approaches were used. Through assessing intrastudy quality-control samples, the sixth partner observed high technical variation and was unable to group the substances. By comparing workflows, we conclude that some heterogeneity in metabolomics methods is not detrimental to consistent grouping, and that assessing data quality prior to grouping is essential. We recommend development of international guidance for quality-control acceptance criteria. This study demonstrates the reliability of metabolomics for chemical grouping and works towards best-practice.

The ICH M10 guideline on bioanalytical method validation and sample analysis is being adopted since 2023. However, and inevitably, some paragraphs or requirements remain ambiguous and are open for different interpretations. In support of a harmonized interpretation by the industry and health authorities, the European Bioanalysis Forum organized a workshop on 14 November 2023 in Barcelona, Spain, to discuss unclear and/or ambiguous paragraphs which were identified by the European Bioanalysis Forum community and delegates of the workshop prior to the workshop. This manuscript reports back from the workshop with recommendations and aims at continuing an open scientific discussion within the industry and with regulators in support of a science-driven guideline for the bioanalytical community and in line with the ICH mission - that is, achieve greater harmonization worldwide to ensure that safe, effective and high-quality medicines are developed and registered in the most resource-efficient manner.

Prostate cancer is a frequent malignancy in older men and has a very high 5‐year survival rate if diagnosed early. The prognosis is much less promising if the tumor has already spread outside the prostate gland. Targeted treatments mainly aim at blocking androgen receptor (AR) signaling and initially show good efficacy. However, tumor progression due to AR‐dependent and AR‐independent mechanisms is often observed after some time, and novel treatment strategies are urgently needed. Dysregulation of the PI3K/AKT/mTOR pathway in advanced prostate cancer and its implication in treatment resistance has been reported. We compared the impact of PI3K/AKT/mTOR pathway inhibitors with different selectivity profiles on in vitro cell proliferation and on caspase 3/7 activation as a marker for apoptosis induction, and observed the strongest effects in the androgen‐sensitive prostate cancer cell lines VCaP and LNCaP. Combination treatment with the AR inhibitor darolutamide led to enhanced apoptosis in these cell lines, the effects being most pronounced upon cotreatment with the pan‐PI3K inhibitor copanlisib. A subsequent transcriptomic analysis performed in VCaP cells revealed that combining darolutamide with copanlisib impacted gene expression much more than individual treatment. A comprehensive reversal of the androgen response and the mTORC1 transcriptional programs as well as a marked induction of DNA damage was observed. Next, an in vivo efficacy study was performed using the androgen‐sensitive patient‐derived prostate cancer (PDX) model LuCaP 35 and a superior efficacy was observed after the combined treatment with copanlisib and darolutamide. Importantly, immunohistochemistry analysis of these treated tumors showed increased apoptosis, as revealed by elevated levels of cleaved caspase 3 and Bcl‐2‐binding component 3 (BBC3). In conclusion, these data demonstrate that concurrent blockade of the PI3K/AKT/mTOR and AR pathways has superior antitumor efficacy and induces apoptosis in androgen‐sensitive prostate cancer cell lines and PDX models.

As a step towards implementing non-aversive handling techniques at a big mouse breeding facility in Germany, tunnel handling was introduced in a breeding unit comprising three inbred mouse strains. To assess whether tunnel handling would be feasible for the animal technicians in their everyday work and beneficial for the mice when being handled during weekly cage change only, the behaviour of tunnel- and tail-handled animals of both sexes was examined before, during and after the handling events over a period of nine weeks. Moreover, the time expenditure was compared between both handling techniques. It was possible to use the tunnel in all three mouse strains. However, the impact of the handling techniques on the behavioural parameters investigated in the present study were strain-specific. All behavioural parameters suggested that NZW mice benefited the most from tunnel handling. The results obtained from Hello Kitty and WNK mice were ambiguous, which may suggest that a brief handling session during the cage clean may have not been sufficient to habituate them to the process of handling. It took the animal technicians approximately 3 seconds longer per mouse when using a tunnel. The strain-specific results should encourage researchers to share their experiences with non-aversive handling techniques in different mouse strains, for example, along with their research articles.

BackgroundThe second messenger diacylglycerol plays a key role in T-cell receptor (TCR) downstream signaling and T-cell activation. Diacylglycerol kinase zeta (DGKζ) is a lipid kinase that regulates T-cell activation by phosphorylating diacylglycerol to produce phosphatidic acid, thereby acting as a ligand-independent, intracellular immune checkpoint. The inhibition of DGKζ offers the potential to enhance T-cell priming against suboptimal tumor antigens and to overcome multiple immunosuppressive mechanisms in the tumor microenvironment in a TCR engagement-dependent manner. We evaluated the specificity, efficacy, and safety of the DGKζ inhibitor BAY 2965501 in various preclinical in vitro and in vivo studies.Methods and ResultsBAY 2965501 is a highly selective, potent human/mouse cross-reactive DGKζ inhibitor. In vitro, BAY 2965501 increased natural killer cell- and T-cell-mediated tumor cell killing and enhanced interleukin 2-induced natural killer cell activation. Importantly, the inhibition of DGKζ by BAY 2965501 was able to overcome inhibitory signals conferred by transforming growth factor beta, prostaglandin E2, and adenosine signaling in T cells. However, BAY 2965501 showed no direct anti-proliferative effects on human tumor cell lines in vitro. Single-cell sequencing of human tumor-infiltrating lymphocytes isolated from primary human tumors revealed high expression of DGKζ specifically in exhausted CD8+ TCR clonotypes, suggesting a potential immunosuppressive role in this T-cell subpopulation. Furthermore, BAY 2965501 effectively enhanced the in vitro anti-tumor reactivity of human T cells expressing tumor-reactive TCRs. In vivo, BAY 2965501 reduced T-cell exhaustion markers, such as programmed cell death receptor 1 and T-cell immunoglobulin mucin 3, and enhanced antiviral T-cell responses in mice chronically infected with lymphocytic choriomeningitis virus. In MB49, F9, and Hepa129 syngeneic mouse tumor models, BAY 2965501 as monotherapy reduced tumor growth when compared with vehicle treatment. Combining BAY 2965501 with an anti-programmed cell death ligand-1 (anti-PDL-1) antibody reduced tumor growth vs anti-PDL-1 monotherapy. Preclinical toxicology studies showed only low-grade gastrointestinal effects, suggesting a tolerable clinical profile.ConclusionsIn summary, BAY 2965501 is a highly potent and selective, orally available DGKζ inhibitor. A first-in-human clinical trial of BAY 2965501 in solid tumors is currently enrolling patients (NCT05614102). This study will evaluate the safety, tolerability, maximum tolerated or administered dose, pharmacokinetics, pharmacodynamics, and tumor response profile of BAY 2965501.AcknowledgementsEditing support was provided by Rachel Fairbanks, BA (Hons), Complete HealthVizion, IPG Health Medical Communications, and was funded by Bayer AG.Ethics ApprovalAll animal experiments were conducted in accordance with the German Animal Welfare Law and approved by Berlin authorities (Landesamt für Arbeitsschutz, Gesundheitsschutz und technische Sicherheit Berlin, LAGetSi; code number A0378/12)

The human pituitary adenylate cyclase-activating polypeptide receptor (hPAC1-R), a class B G-protein-coupled receptor (GPCR) identified almost 30 years ago, represents an important pharmacological target in the areas of neuroscience, oncology, and immunology. Despite interest in this target, only a very limited number of small molecule modulators have been reported for this receptor. We herein describe the results of a drug discovery program aiming for the identification of a potent and selective hPAC1-R antagonist. An initial high-throughput screening (HTS) screen of 3.05 million compounds originating from the Bayer screening library failed to identify any tractable hits. A second, completely revised screen using native human embryonic kidney (HEK)293 cells yielded a small number of hits exhibiting antagonistic properties (4.2 million compounds screened). BAY 2686013 (1) emerged as a promising compound showing selective antagonistic activity in the submicromolar potency range. In-depth characterization supported the hypothesis that BAY 2686013 blocks receptor activity in a noncompetitive manner. Preclinical, pharmacokinetic profiling indicates that BAY 2686013 is a valuable tool compound for better understanding the signaling and function of hPAC1-R. SIGNIFICANCE STATEMENT: Although the human pituitary adenylate cyclase-activating polypeptide receptor (hPAC1-R) is of major significance as a therapeutic target with a well documented role in pain signaling, only a very limited number of small-molecule (SMOL) compounds are known to modulate its activity. We identified and thoroughly characterized a novel, potent, and selective SMOL antagonist of hPAC1-R (acting in an allosteric manner). These characteristics make BAY 2686013 an ideal tool for further studies.

Dear Editor, Metabolically induced cancer heterogeneity creates a large source of novel potential targets towards an enhanced therapeutic window alone and in combination with classic chemo-and radiotherapy [1, 2] . This is of particular interest for non-small cell lung cancer (NSCLC), which accounts for more than 80% of all lung tumor types characterized by limited responses to current treatment options [3] [4] [5] . Genetically-defined KRAS/LKB1-mutant NSCLC tumors exploit the proximal urea cycle enzyme carbamoyl phosphate synthetase 1 (CPS1) as an intermediate step for pyrimidine biosynthesis, thereby contrasting its crucial hepatic role in ammonia detoxification [6] [7] [8] . Thus, CPS1 could serve as a novel target for NSCLC susceptibility (Figure 1A ). In this study, we investigated the metabolic changes observed in both NSCLC and healthy hepatic systems following the identification, characterization and application of a small molecule inhibitor of ectopic CPS1 functionality. To search for CPS1 ligands, we performed a highthroughput screening approach as part of a thermal stability assay with recombinant human CPS1 and identified 5 out of more than 11,000 compounds as potential CPS1 inhibitory ligands (Figure 1B , Supplementary

Abstract CSNK1A1 is a serine/threonine kinase involved in multiple cellular processes, including cell division, beta catenin signaling, and TP53 activation. Inhibition of CSNK1A1 has previously been validated as a therapeutic strategy in hematologic malignancy, and degradation of CSNK1A1 protein is the downstream mechanism of action for lenalidomide in 5q- myelodysplasia (Krönke, et al. Nature. 2015.). However, lenalidomide is inactive in most solid tumor models, thus limiting the study of CSNK1A1 inhibition in other contexts. Analysis of genetic loss-of-function data from the Cancer Dependency Map reveals multiple sensitive models, including lineage-specific enrichment in colorectal and gastric cancer. In an academic-industry collaboration, we a) developed first-in-class potent and selective ATP-competitive CSNK1A1 small molecule inhibitors with preclinical anti-cancer efficacy in vivo, and b) identified FAM83 expression as a key determinant of inhibitor sensitivity. We identified a tetrahydro-pyrrolopyridinone scaffold that was subsequently optimized to yield BAY-888 (CSNK1A1 IC50 4 nM @ 10 μM ATP; 63 nM @ 1 mM ATP) and BAY-204 (CSNK1A1 IC50 2 nM @ 10 μM ATP; 12 nM @ 1 mM ATP). The crystal structure of CSNK1A1 in complex with BAY-888 confirmed compound binding in the ATP binding pocket. Across the PRISM barcoded cell line panel of more than 500 solid tumor cell lines, inhibitors phenocopy the CSNK1A1 shRNA knockdown profile. To determine downstream mediators of CSNK1A1 inhibitor sensitivity, we performed co-IP mass spectrometry following CSNK1A1 pulldown and global phosphoproteomic assays following inhibitor treatment. We identified multiple interacting proteins that are also phosphorylation targets, including FAM83 family members. FAM83 was recently reported to mediate the subcellular localization of CSNK1A1 (Fulcher, et al. Sci Signal. 2018.). Excitingly, the baseline expression of FAM83B and FAM83H correlates with inhibitor and shRNA cell line sensitivity. Modulation of FAM83H expression altered CSNK1A1 localization and sensitivity to CSNK1A1 inhibition. BAY-888 and BAY-204 are orally bioavailable and were evaluated in multiple murine cell line xenograft models. We observed promising efficacy in DLBCL (TMD8) in vivo as well as in multiple FAM83-high solid tumor models, including colorectal (HCT116 and HT29), gastric (IM95), and urothelial cancer (KU19-19). We identified RPS6 phosphorylation as one of the PD biomarkers correlating with efficacy in vivo. In summary, CSNK1A1 is a promising target with anti-tumor efficacy and achievable therapeutic index in preclinical models of FAM83-high solid tumors. Citation Format: Steven M. Corsello, Huajia Zhang, Rajesha Rupaimoole, Volker K. Schulze, Clara Lemos, Kasia B. Handing, Douglas L. Orsi, Mrinal Shekhar, Ulrike Sack, Sven Christian, Wilhelm Bone, Ranad Humeidi, William Colgan, Stephanie Hoyt, Andrew Cherniack, Jens Schroder, Stefan Kaulfuss, Krzysztof Brzezinka, Oliver von Ahsen, Anne Mengel, Roman C. Hillig, Detlev Suelzle, Jeremie Mortier, Caitlin Harrington, Rohith Nagari, Justyna Wierzbinska, Derek Chiang, Georg Beckmann, Meagan Olive, Namrata Udeshi, Annie Apffel, Steven Carr, Philip Lienau, Christian Lechner, Ulf Boemer, Alisha Caliman, David McKinney, Florence Wagner, Dominik Mumberg, Marcus Bauser, Andrea Haegebarth, Knut Eis, Ashley Eheim, Todd R. Golub. Discovery of potent and selective CSNK1A1 inhibitors for solid tumor 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 3588.