Property- and Structure-Guided Discovery of a Tetrahydroindazole Series of Interleukin‑2 Inducible T‑Cell Kinase Inhibitors

Interleukin-2 inducible T-cell kinase (ITK), a member of the Tec family of tyrosine kinases, plays a major role in T-cell signaling downstream of the T-cell receptor (TCR), and considerable efforts have been directed toward discovery of ITK-selective inhibitors as potential treatments of inflammatory disorders such as asthma. Using a previously disclosed indazole series of inhibitors as a starting point, and using X-ray crystallography and solubility forecast index (SFI) as guides, we evolved a series of tetrahydroindazole inhibitors with improved potency, selectivity, and pharmaceutical properties. Highlights include identification of a selectivity pocket above the ligand plane, and identification of appropriate lipophilic substituents to occupy this space. This effort culminated in identification of a potent and selective ITK inhibitor (GNE-9822) with good ADME properties in preclinical species

Battling Btk Mutants With Noncovalent Inhibitors That Overcome Cys481 and Thr474 Mutations

The Bruton’s tyrosine kinase (Btk) inhibitor ibrutinib has shown impressive clinical efficacy in a range of B-cell malignancies. However, acquired resistance has emerged, and second generation therapies are now being sought. Ibrutinib is a covalent, irreversible inhibitor that modifies Cys481 in the ATP binding site of Btk and renders the enzyme inactive, thereby blocking B-cell receptor signal transduction. Not surprisingly, Cys481 is the most commonly mutated Btk residue in cases of acquired resistance to ibrutinib. Mutations at other sites, including Thr474, a gatekeeper residue, have also been detected. Herein, we describe noncovalent Btk inhibitors that differ from covalent inhibitors like ibrutinib in that they do not interact with Cys481, they potently inhibit the ibrutinib-resistant Btk C481S mutant <i>in vitro</i> and in cells, and they are exquisitely selective for Btk. Noncovalent inhibitors such as GNE-431 also show excellent potency against the C481R, T474I, and T474M mutants. X-ray crystallographic analysis of Btk provides insight into the unique mode of binding of these inhibitors that explains their high selectivity for Btk and their retained activity against mutant forms of Btk. This class of noncovalent Btk inhibitors may provide a treatment option to patients, especially those who have acquired resistance to ibrutinib by mutation of Cys481 or Thr474

Pyridones as Highly Selective, Noncovalent Inhibitors of T790M Double Mutants of EGFR

The rapid advancement of a series of noncovalent inhibitors of T790M mutants of EGFR is discussed. The optimization of pyridone <b>1</b>, a nonselective high-throughput screening hit, to potent molecules with high levels of selectivity over wtEGFR and the broader kinome is described herein

Tetrahydroindazoles as Interleukin‑2 Inducible T‑Cell Kinase Inhibitors. Part II. Second-Generation Analogues with Enhanced Potency, Selectivity, and Pharmacodynamic Modulation in Vivo

The medicinal chemistry community has directed considerable efforts toward the discovery of selective inhibitors of interleukin-2 inducible T-cell kinase (ITK), given its role in T-cell signaling downstream of the T-cell receptor (TCR) and the implications of this target for inflammatory disorders such as asthma. We have previously disclosed a structure- and property-guided lead optimization effort which resulted in the discovery of a new series of tetrahydroindazole-containing selective ITK inhibitors. Herein we disclose further optimization of this series that resulted in further potency improvements, reduced off-target receptor binding liabilities, and reduced cytotoxicity. Specifically, we have identified a correlation between the basicity of solubilizing elements in the ITK inhibitors and off-target antiproliferative effects, which was exploited to reduce cytotoxicity while maintaining kinase selectivity. Optimized analogues were shown to reduce IL-2 and IL-13 production in vivo following oral or intraperitoneal dosing in mice

Discovery of Potent and Selective Pyrazolopyrimidine Janus Kinase 2 Inhibitors

The discovery of somatic Jak2 mutations in patients with chronic myeloproliferative neoplasms has led to significant interest in discovering selective Jak2 inhibitors for use in treating these disorders. A high-throughput screening effort identified the pyrazolo­[1,5-<i>a</i>]­pyrimidine scaffold as a potent inhibitor of Jak2. Optimization of lead compounds <b>7a</b>–<b>b</b> and <b>8</b> in this chemical series for activity against Jak2, selectivity against other Jak family kinases, and good in vivo pharmacokinetic properties led to the discovery of <b>7j</b>. In a SET2 xenograft model that is dependent on Jak2 for growth, <b>7j</b> demonstrated a time-dependent knock-down of pSTAT5, a downstream target of Jak2

Tetrahydroindazoles as Interleukin‑2 Inducible T‑Cell Kinase Inhibitors. Part II. Second-Generation Analogues with Enhanced Potency, Selectivity, and Pharmacodynamic Modulation in Vivo

The medicinal chemistry community has directed considerable efforts toward the discovery of selective inhibitors of interleukin-2 inducible T-cell kinase (ITK), given its role in T-cell signaling downstream of the T-cell receptor (TCR) and the implications of this target for inflammatory disorders such as asthma. We have previously disclosed a structure- and property-guided lead optimization effort which resulted in the discovery of a new series of tetrahydroindazole-containing selective ITK inhibitors. Herein we disclose further optimization of this series that resulted in further potency improvements, reduced off-target receptor binding liabilities, and reduced cytotoxicity. Specifically, we have identified a correlation between the basicity of solubilizing elements in the ITK inhibitors and off-target antiproliferative effects, which was exploited to reduce cytotoxicity while maintaining kinase selectivity. Optimized analogues were shown to reduce IL-2 and IL-13 production in vivo following oral or intraperitoneal dosing in mice

Discovery of GDC-0853: A Potent, Selective, and Noncovalent Bruton’s Tyrosine Kinase Inhibitor in Early Clinical Development

Bruton’s tyrosine kinase (Btk) is a nonreceptor cytoplasmic tyrosine kinase involved in B-cell and myeloid cell activation, downstream of B-cell and Fcγ receptors, respectively. Preclinical studies have indicated that inhibition of Btk activity might offer a potential therapy in autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus. Here we disclose the discovery and preclinical characterization of a potent, selective, and noncovalent Btk inhibitor currently in clinical development. GDC-0853 (<b>29</b>) suppresses B cell- and myeloid cell-mediated components of disease and demonstrates dose-dependent activity in an <i>in vivo</i> rat model of inflammatory arthritis. It demonstrates highly favorable safety, pharmacokinetic (PK), and pharmacodynamic (PD) profiles in preclinical and Phase 2 studies ongoing in patients with rheumatoid arthritis, lupus, and chronic spontaneous urticaria. On the basis of its potency, selectivity, long target residence time, and noncovalent mode of inhibition, <b>29</b> has the potential to be a best-in-class Btk inhibitor for a wide range of immunological indications

Discovery of GDC-0853: A Potent, Selective, and Noncovalent Bruton’s Tyrosine Kinase Inhibitor in Early Clinical Development

Bruton’s tyrosine kinase (Btk) is a nonreceptor cytoplasmic tyrosine kinase involved in B-cell and myeloid cell activation, downstream of B-cell and Fcγ receptors, respectively. Preclinical studies have indicated that inhibition of Btk activity might offer a potential therapy in autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus. Here we disclose the discovery and preclinical characterization of a potent, selective, and noncovalent Btk inhibitor currently in clinical development. GDC-0853 (<b>29</b>) suppresses B cell- and myeloid cell-mediated components of disease and demonstrates dose-dependent activity in an <i>in vivo</i> rat model of inflammatory arthritis. It demonstrates highly favorable safety, pharmacokinetic (PK), and pharmacodynamic (PD) profiles in preclinical and Phase 2 studies ongoing in patients with rheumatoid arthritis, lupus, and chronic spontaneous urticaria. On the basis of its potency, selectivity, long target residence time, and noncovalent mode of inhibition, <b>29</b> has the potential to be a best-in-class Btk inhibitor for a wide range of immunological indications

Cell Active Hydroxylactam Inhibitors of Human Lactate Dehydrogenase with Oral Bioavailability in Mice

A series of trisubstituted hydroxylactams was identified as potent enzymatic and cellular inhibitors of human lactate dehydrogenase A. Utilizing structure-based design and physical property optimization, multiple inhibitors were discovered with <10 μM lactate IC₅₀ in a MiaPaca2 cell line. Optimization of the series led to <b>29</b>, a potent cell active molecule (MiaPaca2 IC₅₀ = 0.67 μM) that also possessed good exposure when dosed orally to mice

Discovery of a Noncovalent, Mutant-Selective Epidermal Growth Factor Receptor Inhibitor

Inhibitors targeting the activating mutants of the epidermal growth factor receptor (EGFR) have found success in the treatment of EGFR mutant positive non-small-cell lung cancer. A secondary point mutation (T790M) in the inhibitor binding site has been linked to the acquired resistance against those first generation therapeutics. Herein, we describe the lead optimization of a series of reversible, pan-mutant (L858R, del_746–750, T790M/L858R, and T790M/del_746–750) EGFR inhibitors. By use of a noncovalent double mutant (T790M/L858R and T790M/del_746–750) selective EGFR inhibitor (<b>2</b>) as a starting point, activities against the single mutants (L858R and del_746–750) were introduced through a series of structure-guided modifications. The in vitro ADME-PK properties of the lead molecules were further optimized through a number of rational structural changes. The resulting inhibitor (<b>21</b>) exhibited excellent cellular activity against both the single and double mutants of EGFR, demonstrating target engagement in vivo and ADME-PK properties that are suitable for further evaluation. The reversible, noncovalent inhibitors described complement the covalent pan-mutant EGFR inhibitors that have shown encouraging results in recent clinical trials