Imidazolone as an Amide Bioisostere in the Development of β-1,3--Acetylglucosaminyltransferase 2 (B3GNT2) Inhibitors.

B3GNT2 is responsible for elongation of cell surface long-chain polylactosamine, which influences the regulation of the immune response, making it an attractive target for immunomodulation. In the development of amide containing B3GNT2 inhibitors guided by structure-based drug design, imidazolones were found to successfully serve as amide bioisosteres. This novel imidazolone isosteric strategy alleviated torsional strain of the amide bond on binding to B3GNT2 and improved potency, isoform selectivity, as well as certain physicochemical and pharmacokinetic properties.

Systematic Study of the Glutathione Reactivity of -Phenylacrylamides: 2. Effects of Acrylamide Substitution.

A comprehensive understanding of structure-reactivity relationships is critical to the design and optimization of cysteine-targeted covalent inhibitors. Herein, we report glutathione (GSH) reaction rates for -phenyl acrylamides with varied substitutions at the α- and β-positions of the acrylamide moiety. We find that the GSH reaction rates can generally be understood in terms of the electron donating or withdrawing ability of the substituent.

Discovery of a Covalent Inhibitor of KRAS (AMG 510) for the Treatment of Solid Tumors.

KRAS has emerged as a promising target in the treatment of solid tumors. Covalent inhibitors targeting the mutant cysteine-12 residue have been shown to disrupt signaling by this long-"undruggable" target; however clinically viable inhibitors have yet to be identified. Here, we report efforts to exploit a cryptic pocket (H95/Y96/Q99) we identified in KRAS to identify inhibitors suitable for clinical development.

The clinical KRAS(G12C) inhibitor AMG 510 drives anti-tumour immunity.

KRAS is the most frequently mutated oncogene in cancer and encodes a key signalling protein in tumours. The KRAS(G12C) mutant has a cysteine residue that has been exploited to design covalent inhibitors that have promising preclinical activity. Here we optimized a series of inhibitors, using novel binding interactions to markedly enhance their potency and selectivity.

Discovery of -(1-Acryloylazetidin-3-yl)-2-(1-indol-1-yl)acetamides as Covalent Inhibitors of KRAS.

KRAS regulates many cellular processes including proliferation, survival, and differentiation. Point mutants of KRAS have long been known to be molecular drivers of cancer. , which occurs in approximately 14% of lung adenocarcinomas, 3-5% of colorectal cancers, and low levels in other solid tumors, represents an attractive therapeutic target for covalent inhibitors.

Discovery of ( R)-8-(6-Methyl-4-oxo-1,4,5,6-tetrahydropyrrolo[3,4- b]pyrrol-2-yl)-3-(1-methylcyclopropyl)-2-((1-methylcyclopropyl)amino)quinazolin-4(3 H)-one, a Potent and Selective Pim-1/2 Kinase Inhibitor for Hematological Malignancies.

Pim kinases are a family of constitutively active serine/threonine kinases that are partially redundant and regulate multiple pathways important for cell growth and survival. In human disease, high expression of the three Pim isoforms has been implicated in the progression of hematopoietic and solid tumor cancers, which suggests that Pim kinase inhibitors could provide patients with therapeutic benefit. Herein, we describe the structure-guided optimization of a series of quinazolinone-pyrrolodihydropyrrolone analogs leading to the identification of potent pan-Pim inhibitor 28 with improved potency, solubility, and drug-like properties.

Targeted Degradation of MDM2 as a New Approach to Improve the Efficacy of MDM2-p53 Inhibitors.

MDM2 is a key oncogenic protein that serves as a negative regulator of the tumor suppressor p53. While a number of inhibitors of the MDM2-p53 interaction have progressed to clinical testing as treatments for a variety of hematologic and solid tumor cancers, the results thus far have been mixed, with perhaps the strongest responses observed in relapsed/refractory acute myeloid leukemia (AML). In an effort to improve the efficacy for this class of compounds, researchers have turned to targeted degradation of MDM2.

Discovery and Optimization of Quinazolinone-pyrrolopyrrolones as Potent and Orally Bioavailable Pan-Pim Kinase Inhibitors.

The high expression of proviral insertion site of Moloney murine leukemia virus kinases (Pim-1, -2, and -3) in cancers, particularly the hematopoietic malignancies, is believed to play a role in promoting cell survival and proliferation while suppressing apoptosis. The three isoforms of Pim protein appear largely redundant in their oncogenic functions. Thus, a pan-Pim kinase inhibitor is highly desirable.

Discovery and Optimization of Macrocyclic Quinoxaline-pyrrolo-dihydropiperidinones as Potent Pim-1/2 Kinase Inhibitors.

The identification of Pim-1/2 kinase overexpression in B-cell malignancies suggests that Pim kinase inhibitors will have utility in the treatment of lymphoma, leukemia, and multiple myeloma. Starting from a moderately potent quinoxaline-dihydropyrrolopiperidinone lead, we recognized the potential for macrocyclization and developed a series of 13-membered macrocycles. The structure-activity relationships of the macrocyclic linker were systematically explored, leading to the identification of 9c as a potent, subnanomolar inhibitor of Pim-1 and -2.

High-Throughput Mass Spectrometric Analysis of Covalent Protein-Inhibitor Adducts for the Discovery of Irreversible Inhibitors: A Complete Workflow.

We have implemented a solid-phase extraction based time-of-flight mass spectrometer system in combination with novel informatics to rapidly screen and characterize the covalent binding of different irreversible inhibitors to intact proteins. This high-throughput screening platform can be used to accurately detect and quantitate the extent of formation of different covalent protein-inhibitor adducts between electrophilic inhibitors and nucleophilic residues such as cysteine or lysine. For a representative 19.

Systematic Study of the Glutathione (GSH) Reactivity of N-Arylacrylamides: 1. Effects of Aryl Substitution.

Success in the design of targeted covalent inhibitors depends in part on a knowledge of the factors influencing electrophile reactivity. In an effort to further develop an understanding of structure-reactivity relationships among N-arylacrylamides, we determined glutathione (GSH) reaction rates for a family of N-arylacrylamides independently substituted at ortho-, meta-, and para-positions with 11 different groups common to inhibitor design. We find that substituent effects on reaction rates show a linear Hammett correlation for ortho-, meta-, and para-substitution.

Discovery of 5-(1H-indol-5-yl)-1,3,4-thiadiazol-2-amines as potent PIM inhibitors.

PIM kinases are a family of Ser/Thr kinases that are implicated in tumorigenesis. The discovery of a new class of PIM inhibitors, 5-(1H-indol-5-yl)-1,3,4-thiadiazol-2-amines, is discussed with optimized compounds showing excellent potency against all three PIM isoforms.

The discovery of novel 3-(pyrazin-2-yl)-1H-indazoles as potent pan-Pim kinase inhibitors.

The three Pim kinases are a small family of serine/threonine kinases regulating several signaling pathways that are fundamental to tumorigenesis. As such, the Pim kinases are a very attractive target for pharmacological inhibition in cancer therapy. Herein, we describe our efforts toward the development of a potent, pan-Pim inhibitor.

Phosphoinositide-3-kinase inhibitors: evaluation of substituted alcohols as replacements for the piperazine sulfonamide portion of AMG 511.

Replacement of the piperazine sulfonamide portion of the PI3Kα inhibitor AMG 511 (1) with a range of aliphatic alcohols led to the identification of a truncated gem-dimethylbenzylic alcohol analog, 2-(5-(4-amino-6-methyl-1,3,5-triazin-2-yl)-6-((5-fluoro-6-methoxypyridin-3-yl)amino)pyridin-3-yl)propan-2-ol (7). This compound possessed good in vitro efficacy and pharmacokinetic parameters and demonstrated an EC50 of 239 ng/mL in a mouse liver pharmacodynamic model measuring the inhibition of hepatocyte growth factor (HGF)-induced Akt Ser473 phosphorylation in CD1 nude mice 6 h post-oral dosing.

Predictability of peripheral lymphocyte reduction of novel S1P1 agonists by in vitro GPCR signaling profile.

Surrogate readouts of G-protein-coupled receptor signaling pathways using highly engineered systems are often employed in the drug discovery process. However, accumulating data have demonstrated the importance of selecting relevant biological activity rather than technically facile assays to support high-throughout screening and subsequent structure-activity relationship studies. Here we report a case study using sphingosine-1-phosphate receptor 1 (S1P(1)) as the model system to compare compound activity in six different in vitro assays with their ability to predict in vivo efficacy.

Selective class I phosphoinositide 3-kinase inhibitors: optimization of a series of pyridyltriazines leading to the identification of a clinical candidate, AMG 511.

The phosphoinositide 3-kinase family catalyzes the phosphorylation of phosphatidylinositol-4,5-diphosphate to phosphatidylinositol-3,4,5-triphosphate, a secondary messenger which plays a critical role in important cellular functions such as metabolism, cell growth, and cell survival. Our efforts to identify potent, efficacious, and orally available phosphatidylinositol 3-kinase (PI3K) inhibitors as potential cancer therapeutics have resulted in the discovery of 4-(2-((6-methoxypyridin-3-yl)amino)-5-((4-(methylsulfonyl)piperazin-1-yl)methyl)pyridin-3-yl)-6-methyl-1,3,5-triazin-2-amine (1). In this paper, we describe the optimization of compound 1, which led to the design and synthesis of pyridyltriazine 31, a potent pan inhibitor of class I PI3Ks with a superior pharmacokinetic profile.

Structure-based design of a novel series of potent, selective inhibitors of the class I phosphatidylinositol 3-kinases.

A highly selective series of inhibitors of the class I phosphatidylinositol 3-kinases (PI3Ks) has been designed and synthesized. Starting from the dual PI3K/mTOR inhibitor 5, a structure-based approach was used to improve potency and selectivity, resulting in the identification of 54 as a potent inhibitor of the class I PI3Ks with excellent selectivity over mTOR, related phosphatidylinositol kinases, and a broad panel of protein kinases. Compound 54 demonstrated a robust PD-PK relationship inhibiting the PI3K/Akt pathway in vivo in a mouse model, and it potently inhibited tumor growth in a U-87 MG xenograft model with an activated PI3K/Akt pathway.

Optimization of a Potent, Orally Active S1P1 Agonist Containing a Quinolinone Core.

The optimization of a series of S1P1 agonists with limited activity against S1P3 is reported. A polar headgroup was used to improve the physicochemical and pharmacokinetic parameters of lead quinolinone 6. When dosed orally at 1 and 3 mg/kg, the azahydroxymethyl analogue 22 achieved statistically significant lowering of circulating blood lymphocytes 24 h postdose.

Quinolinone-based agonists of S1P₁: use of a N-scan SAR strategy to optimize in vitro and in vivo activity.

We reveal how a N-scan SAR strategy (systematic substitution of each CH group with a N atom) was employed for quinolinone-based S1P(1) agonist 5 to modulate physicochemical properties and optimize in vitro and in vivo activity. The diaza-analog 17 displays improved potency (hS1P(1) RI; 17: EC(50)=0.020 μM, 120% efficacy; 5: EC(50)=0.

Novel 5- and 6-subtituted benzothiazoles with improved physicochemical properties: potent S1P₁ agonists with in vivo lymphocyte-depleting activity.

An SAR campaign designed to increase polarity in the 'tail' region of benzothiazole 1 resulted in two series of structurally novel 5-and 6-substituted S1P(1) agonists. Structural optimization for potency ultimately delivered carboxamide (+)-11f, which in addition to possessing improved physicochemical properties relative to starting benzothiazole 1, also displayed good S1P(3) selectivity and acceptable in vivo lymphocyte-depleting activity.