Phenex Pharmaceuticals (Germany)

A key metabolic alteration in tumor cells is an elevated glycolysis rate, resulting in the production of increased amounts of protons and lactate leading to lactic acidosis within the tumor microenvironment. The tumor-promoting effects of lactic acidosis are versatile, ranging from stimulation of proliferation, angiogenesis and metastasis to the reshaping of the tumor immune environment to a tumor-permissive phenotype. In tumors lactate is transported across the plasma membrane by the monocarboxylate transporters 1 and 4 (MCT 1/4) in symport with a proton. While MCT1 is expressed almost ubiquitously at low levels, MCT4 expression is regulated by hypoxia-inducible factor 1α (Hif1α), the central player in the cellular response to hypoxia. Thus, MCT4 is present in hypoxic, highly glycolytic cells. Given their central role in prevention of intracellular acidification and lactate transport, MCTs appear to be indispensable for glycolytic tumor growth. Since MCT4 has the strongest implication on the development and aggressiveness of diverse cancer types it has emerged as a new target for the treatment of solid tumors.We have identified MCT4 specific inhibitors structurally based on an internal compound library using cell-based lactate export assays. A subsequent extensive medicinal chemistry program was able to significantly improve activity and selectivity of the primary structures. Cellular target engagement was verified by cell-based assays like cellular thermal shift assay (CETSA). These efforts resulted in the identification of highly potent, orally bioavailable and selective MCT4 inhibitors (MCT4 IC50 < 20nM; MCT1 IC50 > 100µM) with good physicochemical properties and beneficial pharmacokinetics in mouse.In vivo efficacy and tumor lactate modulation were evaluated in NCI-H358 xenograft and MC38 syngeneic mouse models. Thereby we explore the potential of our MCT4 inhibitor to act as a single anti-cancer agent or to synergize with other established therapies like angiogenesis inhibitors or cancer immunotherapeutics.Citation Format: Nina Hambruch, Barbara Herkert, Christian Gege, Aurelie Mallinger, Johannes Fabian, Olaf Kinzel, Yansong Wang, Gero Fink, Michael Albers, Anette Funk, Annika Bittman-Waetzig, Tilly Fleckenstein, Floriane Braun, Claus Kremoser. Identification of a potent, orally bioavailable and selective MCT4 Inhibitor for the treatment of solid Warburg tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3588.

The aryl hydrocarbon receptor (AhR) is a ligand-controlled transcription factor that is primarily known to be a sensor of xenobiotics and tumor-promoting activities of halogenated hydrocarbons and polycyclic aromatic hydrocarbons. Several endogenous ligands such as metabolites produced by commensal microorganisms on skin and in gut and metabolites of L-Tryptophan - produced under control of the Tryptophan dioxygenases IDO1 and TDO2 such as L-Kynurenine and Kynurenic acid are known to modulate the transcriptional activity of AhR. The activity of AhR is predominant in many different immune and epithelial cells, thereby balancing immune responses towards various signals.Activation of the IDO1-Kyn-AhR pathway and accumulation of nuclear AhR protein is frequently seen in different tumor types and possibly linked to the observed diminished anti-tumor immune response. AhR agonizing ligands produced by cancer cells and/or lymphocytes recruited to the TME reduce the anti-tumor immune response through increasing the numbers and function of regulatory T cells while reducing that of cytotoxic CD8+ T cells.In order to reduce AhR-mediated immune-suppression in cancer patients, Phenex Pharmaceuticals initiated a program to identify small molecule AhR antagonists to block activated downstream signaling of AhR. To this end, we have identified novel AhR antagonists, which showed strong antagonistic activity against agonist-activated human AhR in a cell based CYP1A1 promoter-driven luciferase reporter assay in HepG2 cells. The compound series display good oral bioavailability and low clearance in mice. Our AhR antagonist molecules show single agent anti-tumor activity and increase the efficacy of Gemcitabine and anti-PD-L1 checkpoint inhibition in different syngeneic mouse tumor models. Overall, AhR antagonist treatment increased CD8+ T cell and M1 macrophage numbers, and decreased IL22 levels, which might be of functional relevance in the tumor context. Future research is directed to determine PK-PD markers suitable for clinical development and identification of human malignancies likely of responding to AhR antagonist treatment.Citation Format: Sheena Pinto, Christoph Steeneck, Michael Albers, Simon Anderhub, Manfred Birkel, Larisa Buselic-Wölfel, Gisela Eisenhardt, Claus Kremoser, Thomas Hoffmann, Ulrich Deuschle. Targeting the IDO1-Kynurenine-AhR pathway for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1210.

ABSTRACTRetinoic acid receptor-related orphan nuclear receptor gamma t (RORγt or RORC2) is a key transcription factor for the differentiation of naïve proinflammatory CD4+ T cells and the production of T helper-17 (TH17) cells. Inhibiting RORγt activity is thought to be beneficial in targeting a variety of inflammatory and autoimmune disorders, however current candidates remain to be validated in the clinic. Recently Vitae Pharmaceuticals successfully finished its Phase 1 single ascending dose clinical study with their proprietary RORγt inverse agonist VTP-43742. On the basis of the reported promising results, Vitae Pharmaceuticals could currently be considered as having the leading clinical candidate in the RORγt inverse agonist category. This prompts the interest on the exact chemical structure of their clinical candidate. The first relevant patent application (WO2014179564) from Vitae Pharmaceuticals describes RORγt inverse agonists with a 5,6-dihydro-4H-pyrrolo[3,4-d]thiazole core, while in the second and latest patent application (WO2015116904) this element has changed towards a 6,7-dihydro-5H-pyrrolo[3,4-b]pyridine core. By combining information from Vitae’s patent applications and trustworthy online information, the potential elucidation of the chemical structure of clinical candidate VTP-43742 is described.

Chemical random mutagenesis techniques with the germ line supermutagen N-ethyl-N-nitrosourea (ENU) have been established to provide comprehensive collections of mouse models, which were then mined and analyzed in phenotype-driven studies. Here, we applied ENU mutagenesis in a high-throughput fashion for a gene-driven identification of new mutations. Selected members of the large superfamily of G protein-coupled receptors (GPCR), melanocortin type 3 (Mc3r) and type 4 (Mc4r) receptors, and the orphan chemoattractant receptor GPR33, were used as model targets to prove the feasibility of this approach. Parallel archives of DNA and sperm from mice mutagenized with ENU were screened for mutations in these GPCR, and in vitro assays served as a preselection step before in vitro fertilization was performed to generate the appropriate mouse model. For example, mouse models for inherited obesity were established by selecting fully or partially inactivating mutations in Mc4r. Our technology described herein has the potential to provide mouse models for a GPCR dysfunction of choice within <4 mo and can be extended to other gene classes of interest.

Partial, selective activation of nuclear receptors is a central issue in molecular endocrinology but only partly understood. Using LXRs as an example, we show here that purely agonistic ligands can be clearly and quantitatively differentiated from partial agonists by the cofactor interactions they induce. Although a pure agonist induces a conformation that is incompatible with the binding of repressors, partial agonists such as GW3965 induce a state where the interaction not only with coactivators, but also corepressors is clearly enhanced over the unliganded state. The activities of the natural ligand 22(R)-hydroxycholesterol and of a novel quinazolinone ligand, LN6500 can be further differentiated from GW3965 and T0901317 by their weaker induction of coactivator binding. Using biochemical and cell-based assays, we show that the natural ligand of LXR is a comparably weak partial agonist. As predicted, we find that a change in the coactivator to corepressor ratio in the cell will affect NCoR recruiting compounds more dramatically than NCoR-dissociating compounds. Our data show how competitive binding of coactivators and corepressors can explain the tissue-specific behavior of partial agonists and open up new routes to a rational design of partial agonists for LXRs.