Preclinical characterization of CPL304110 as a potent and selective inhibitor of fibroblast growth factor receptors 1, 2, and 3 for gastric, bladder, and squamous cell lung cancer.

Fibroblast Growth Factor Receptors (FGFRs) are a family of receptor tyrosine kinases expressed on a plethora of cell membranes. They play crucial roles in both embryonic development and adult tissue functions. There is an increasing amount of evidence that FGFR-mediated oncogenesis is mainly related to gene amplification, activating mutations, or translocation in tumors of various histological types. Dysregulation of FGFRs has been implicated in a wide variety of neoplasms, such as bladder, gastric, and lung cancers. Given their functional significance, FGFRs emerge as promising targets for cancer therapy. Here, we introduce CPL304100, an innovative and highly potent FGFR1-3 kinase inhibitor demonstrating excellent in vitro biological activity. Comprehensive analyses encompassed kinase assays, cell line evaluations, PK/PD studies surface plasmon resonance studies, molecular docking, and in vivo testing in mouse xenografts. CPL304110 exhibited a distinctive binding profile to FGFR1/2/3 kinase domains, accompanied by a good safety profile and favorable ADMET parameters. Selective inhibition of tumor cell lines featuring active FGFR signaling was observed, distinguishing it from cell lines lacking FGFR aberrations (FGFR1, 2, and 3). CPL304110 demonstrated efficacy in both FGFR-dependent cell lines and patient-derived tumor xenograft (PDTX) in vivo models. Comparative analyses with FDA-approved FGFR inhibitors, erdafitinib and pemigatinib, revealed certain advantages of CPL304110 in both in vitro and in vivo assessments. Encouraging preclinical results led the way for the initiation of a Phase I clinical trial (01FGFR2018; NCT04149691) to further evaluate CPL304110 as a novel anticancer therapy.

Arginase 1/2 Inhibitor OATD-02: From Discovery to First-in-man Setup in Cancer Immunotherapy.

Pharmacologic inhibition of the controlling immunity pathway enzymes arginases 1 and 2 (ARG1 and ARG2) is a promising strategy for cancer immunotherapy. Here, we report the discovery and development of OATD-02, an orally bioavailable, potent arginases inhibitor. The unique pharmacologic properties of OATD-02 are evidenced by targeting intracellular ARG1 and ARG2, as well as long drug-target residence time, moderate to high volume of distribution, and low clearance, which may jointly provide a weapon against arginase-related tumor immunosuppression and ARG2-dependent tumor cell growth. OATD-02 monotherapy had an antitumor effect in multiple tumor models and enhanced an efficacy of the other immunomodulators. Completed nonclinical studies and human pharmacokinetic predictions indicate a feasible therapeutic window and allow for proposing a dose range for the first-in-human clinical study in patients with cancer. SIGNIFICANCE: We have developed an orally available, small-molecule intracellular arginase 1 and 2 inhibitor as a potential enhancer in cancer immunotherapy. Because of its favorable pharmacologic properties shown in nonclinical studies, OATD-02 abolishes tumor immunosuppression induced by both arginases, making it a promising drug candidate entering clinical trials.

OATD-02 Validates the Benefits of Pharmacological Inhibition of Arginase 1 and 2 in Cancer.

BACKGROUND: Arginases play essential roles in metabolic pathways, determining the fitness of both immune and tumour cells. Along with the previously validated role of ARG1 in cancer, the particular significance of ARG2 as a therapeutic target has emerged as its levels correlate with malignant phenotype and poor prognosis. These observations unveil arginases, and specifically ARG2, as well-validated and promising therapeutic targets. OATD-02, a new boronic acid derivative, is the only dual inhibitor, which can address the benefits of pharmacological inhibition of arginase 1 and 2 in cancer. METHODS: The inhibitory activity of OATD-02 was determined using recombinant ARG1 and ARG2, as well as in a cellular system using primary hepatocytes and macrophages. In vivo antitumor activity was determined in syngeneic models of colorectal and kidney carcinomas (CT26 and Renca, respectively), as well as in an ARG2-dependent xenograft model of leukaemia (K562). RESULTS: OATD-02 was shown to be a potent dual (ARG1/ARG2) arginase inhibitor with a cellular activity necessary for targeting ARG2. Compared to a reference inhibitor with predominant extracellular activity towards ARG1, we have shown improved and statistically significant antitumor efficacy in the CT26 model and an immunomodulatory effect reflected by Treg inhibition in the Renca model. Importantly, OATD-02 had a superior activity when combined with other immunotherapeutics. Finally, OATD-02 effectively inhibited the proliferation of human K562 leukemic cells both in vitro and in vivo. CONCLUSIONS: OATD-02 is a potent small-molecule arginase inhibitor with optimal drug-like properties, including PK/PD profile. Excellent activity against intracellular ARG2 significantly distinguishes OATD-02 from other arginase inhibitors. OATD-02 represents a very promising drug candidate for the combined treatment of tumours, and is the only pharmacological tool that can effectively address the benefits of ARG1/ARG2 inhibition. OATD-02 will enter clinical trials in cancer patients in 2022.

Design, Synthesis, and Development of pyrazolo[1,5-a]pyrimidine Derivatives as a Novel Series of Selective PI3Kδ Inhibitors: Part I-Indole Derivatives.

Phosphoinositide 3-kinase δ (PI3Kδ), a member of the class I PI3K family, is an essential signaling biomolecule that regulates the differentiation, proliferation, migration, and survival of immune cells. The overactivity of this protein causes cellular dysfunctions in many human disorders, for example, inflammatory and autoimmune diseases, including asthma or chronic obstructive pulmonary disease (COPD). In this work, we designed and synthesized a new library of small-molecule inhibitors based on indol-4-yl-pyrazolo[1,5-a]pyrimidine with IC50 values in the low nanomolar range and high selectivity against the PI3Kδ isoform. CPL302253 (54), the most potent compound of all the structures obtained, with IC50 = 2.8 nM, is a potential future candidate for clinical development as an inhaled drug to prevent asthma.

Inhibition of arginase modulates T-cell response in the tumor microenvironment of lung carcinoma.

Immunotherapy has demonstrated significant activity in a broad range of cancer types, but still the majority of patients receiving it do not maintain durable therapeutic responses. Amino acid metabolism has been proposed to be involved in the regulation of immune response. Here, we investigated in detail the role of arginase 1 (Arg1) in the modulation of antitumor immune response against poorly immunogenic Lewis lung carcinoma. We observed that tumor progression is associated with an incremental increase in the number of Arg1+ myeloid cells that accumulate in the tumor microenvironment and cause systemic depletion of ʟ-arginine. In advanced tumors, the systemic concentrations of ʟ-arginine are decreased to levels that impair the proliferation of antigen-specific T-cells. Systemic or myeloid-specific Arg1 deletion improves antigen-induced proliferation of adoptively transferred T-cells and leads to inhibition of tumor growth. Arginase inhibitor was demonstrated to modestly inhibit tumor growth when used alone, and to potentiate antitumor effects of anti-PD-1 monoclonal antibodies and STING agonist. The effectiveness of the combination immunotherapy was insufficient to induce complete antitumor responses, but was significantly better than treatment with the checkpoint inhibitor alone. Together, these results indicate that arginase inhibition alone is of modest therapeutic benefit in poorly immunogenic tumors; however, in combination with other treatment strategies it may significantly improve survival outcomes.

A Novel Oral Arginase 1/2 Inhibitor Enhances the Antitumor Effect of PD-1 Inhibition in Murine Experimental Gliomas by Altering the Immunosuppressive Environment.

Glioblastomas (GBM) are the common and aggressive primary brain tumors that are incurable by conventional therapies. Immunotherapy with immune checkpoint inhibitors is not effective in GBM patients due to the highly immunosuppressive tumor microenvironment (TME) restraining the infiltration and activation of cytotoxic T cells. Clinical and experimental studies showed the upregulation of expression of the arginase 1 and 2 (ARG1 and ARG2, respectively) in murine and human GBMs. The elevated arginase activity leads to the depletion of L-arginine, an amino-acid required for the proliferation of T lymphocytes and natural killer cells. Inhibition of ARG1/2 in the TME may unblock T cell proliferation and activate effective antitumor responses. To explore the antitumor potential of ARG1/2 inhibition, we analyzed bulk and single-cell RNA sequencing (scRNA-seq) data from human and murine gliomas. We found the upregulation of ARG1/2 expression in GBMs, both in tumor cells and in tumor infiltrating microglia and monocytes/macrophages. We employed selective arginase inhibitors to evaluate if ARG1/2 inhibition in vitro and in vivo exerts the antitumor effects. A novel, selective ARG1/2 inhibitor - OAT-1746 blocked microglia-dependent invasion of U87-MG and LN18 glioma cells in a Matrigel invasion assay better than reference compounds, without affecting the cell viability. OAT-1746 effectively crossed the blood brain barrier in mice and increased arginine levels in the brains of GL261 glioma bearing mice. We evaluated its antitumor efficacy against GL261 intracranial gliomas as a monotherapy and in combination with the PD-1 inhibition. The oral treatment with OAT-1746 did not affect the immune composition of TME, it induced profound transcriptomic changes in CD11b+ cells immunosorted from tumor-bearing brains as demonstrated by RNA sequencing analyses. Treatment with OAT-1746 modified the TME resulting in reduced glioma growth and increased antitumor effects of the anti-PD-1 antibody. Our findings provide the evidence that inhibition of ARG1/2 activity in tumor cells and myeloid cells in the TME unblocks antitumor responses in myeloid cells and NK cells, and improves the efficacy of the PD-1 inhibition.

Discovery and optimization of novel pyrazole-benzimidazole CPL304110, as a potent and selective inhibitor of fibroblast growth factor receptors FGFR (1-3).

The FGFR family is characterized by four receptors (FGFR 1-4), binding to 18 ligands called fibroblast growth factors (FGFs). Aberrant activation of FGFs and their FGFRs has been implicated in a broad spectrum of human tumors. We employed the scaffolds hybridization approach, scaffold-hopping concept to synthesize a series of novel pyrazole-benzimidazole derivatives 56 (a-x). Compound 56q (CPL304110) was identified as a selective and potent pan-FGFR inhibitor for FGFR1, -2, -3 with IC50s of 0.75 nM, 0.50 nM, 3.05 nM respectively, whereas IC50 of 87.90 nM for FGFR4. Due to its favorable pharmacokinetic profile, low toxicity and potent anti-tumor activity in vivo, compound 56q is currently under evaluation in phase I clinical trial for the treatment of bladder, gastric and squamous cell lung cancers (01FGFR2018; NCT04149691).

Discovery and Pharmacokinetics of Sulfamides and Guanidines as Potent Human Arginase 1 Inhibitors.

We designed and synthesized a series of arginase inhibitors as derivatives of the well-known 2-(S)-amino-6-boronohexanoic acid (ABH) with basic and neutral side chains in the α-position relative to the amino acid group. In an effort to improve the pharmacokinetic profile of literature examples and retain potent enzymatic activity, sulfamido moieties were introduced to generate hydrogen bond interaction with the aspartic acid residue in the arginase active site. The compounds with basic guanidine-containing side chains were even more potent arginase inhibitors. Both groups of compounds, as designed, demonstrated low clearance in their pharmacokinetic profile. The most active inhibitor 15aa showed high nanomolar potency with IC50 = 32 nM toward human arginase 1 and demonstrated low clearance (4.2 mL/min/kg), long t 1/2, and moderate volume of distribution in rat pharmacokinetic studies.