Optimization of an Enzymatic Antibody-Drug Conjugation Approach Based on Coenzyme A Analogs.

Phosphopantetheine transferases (PPTases) can be used to efficiently prepare site-specific antibody-drug conjugates (ADCs) by enzymatically coupling coenzyme A (CoA)-linker payloads to 11-12 amino acid peptide substrates inserted into antibodies. Here, a two-step strategy is established wherein in a first step, CoA analogs with various bioorthogonal reactivities are enzymatically installed on the antibody for chemical conjugation with a cytotoxic payload in a second step. Because of the high structural similarity of these CoA analogs to the natural PPTase substrate CoA-SH, the first step proceeds very efficiently and enables the use of peptide tags as short as 6 amino acids compared to the 11-12 amino acids required for efficient one-step coupling of the payload molecule.

Synthesis, structure-activity relationships, and in vivo efficacy of the novel potent and selective anaplastic lymphoma kinase (ALK) inhibitor 5-chloro-N2-(2-isopropoxy-5-methyl-4-(piperidin-4-yl)phenyl)-N4-(2-(isopropylsulfonyl)phenyl)pyrimidine-2,4-diamine (LDK378) currently in phase 1 and phase 2 clinical trials.

The synthesis, preclinical profile, and in vivo efficacy in rat xenograft models of the novel and selective anaplastic lymphoma kinase inhibitor 15b (LDK378) are described. In this initial report, preliminary structure-activity relationships (SARs) are described as well as the rational design strategy employed to overcome the development deficiencies of the first generation ALK inhibitor 4 (TAE684). Compound 15b is currently in phase 1 and phase 2 clinical trials with substantial antitumor activity being observed in ALK-positive cancer patients.

Design, synthesis, and structure-activity-relationship of phenyl imidazoles as potent Smoothened antagonists.

Through scaffold morphing of a known Smoothened antagonist Antag691, a series of novel phenyl imidazole derivatives were developed. Structure-activity-relationship studies and lead optimization led to the discovery of potent, selective and orally bioavailable Smoothened antagonist 19 that is suitable for in vivo studies.

Discovery of NVP-LDE225, a Potent and Selective Smoothened Antagonist.

The blockade of aberrant hedgehog (Hh) signaling has shown promise for therapeutic intervention in cancer. A cell-based phenotypic high-throughput screen was performed, and the lead structure (1) was identified as an inhibitor of the Hh pathway via antagonism of the Smoothened receptor (Smo). Structure-activity relationship studies led to the discovery of a potent and specific Smoothened antagonist N-(6-((2S,6R)-2,6-dimethylmorpholino)pyridin-3-yl)-2-methyl-4'-(trifluoromethoxy)biphenyl-3-carboxamide (5m, NVP-LDE225), which is currently in clinical development.

Discovery of selective aminothiazole aurora kinase inhibitors.

Aurora family kinases regulate important events during mitosis including centrosome maturation and separation, mitotic spindle assembly, and chromosome segregation. Misregulation of Aurora kinases due to genetic amplification and protein overexpression results in aneuploidy and may contribute to tumorigenesis. Here we report the discovery of new small molecule aminothiazole inhibitors of Aurora kinases with exceptional kinase selectivity and report a 1.

Identification of NVP-TAE684, a potent, selective, and efficacious inhibitor of NPM-ALK.

Constitutive overexpression and activation of NPM-ALK fusion protein [t(2:5)(p23;q35)] is a key oncogenic event that drives the survival and proliferation of anaplastic large-cell lymphomas (ALCLs). We have identified a highly potent and selective small-molecule ALK inhibitor, NVP-TAE684, which blocked the growth of ALCL-derived and ALK-dependent cell lines with IC(50) values between 2 and 10 nM. NVP-TAE684 treatment resulted in a rapid and sustained inhibition of phosphorylation of NPM-ALK and its downstream effectors and subsequent induction of apoptosis and cell cycle arrest.

Crystal structure of pyridoxal kinase in complex with roscovitine and derivatives.

Pyridoxal kinase (PDXK) catalyzes the phosphorylation of pyridoxal, pyridoxamine, and pyridoxine in the presence of ATP and Zn2+. This constitutes an essential step in the synthesis of pyridoxal 5'-phosphate (PLP), the active form of vitamin B6, a cofactor for over 140 enzymes. (R)-Roscovitine (CYC202, Seliciclib) is a relatively selective inhibitor of cyclin-dependent kinases (CDKs), currently evaluated for the treatment of cancers, neurodegenerative disorders, renal diseases, and several viral infections.

Roscovitine targets, protein kinases and pyridoxal kinase.

(R)-Roscovitine (CYC202) is often referred to as a "selective inhibitor of cyclin-dependent kinases." Besides its use as a biological tool in cell cycle, neuronal functions, and apoptosis studies, it is currently evaluated as a potential drug to treat cancers, neurodegenerative diseases, viral infections, and glomerulonephritis. We have investigated the selectivity of (R)-roscovitine using three different methods: 1) testing on a wide panel of purified kinases that, along with previously published data, now reaches 151 kinases; 2) identifying roscovitine-binding proteins from various tissue and cell types following their affinity chromatography purification on immobilized roscovitine; 3) investigating the effects of roscovitine on cells deprived of one of its targets, CDK2.

Synthesis and target identification of hymenialdisine analogs.

Hymenialdisine (HMD) is a sponge-derived natural product kinase inhibitor with nanomolar activity against CDKs, Mek1, GSK3beta, and CK1 and micromolar activity against Chk1. In order to explore the broader application of the pyrrolo[2,3-c]azepine skeleton of HMD as a general kinase inhibitory scaffold, we searched for additional protein targets using affinity chromatography in conjunction with the synthesis of diverse HMD analogs and profiled HMD against a panel of 60 recombinant enzymes. This effort has led to nanomolar to micromolar inhibitors of 11 new targets including p90RSK, KDR, c-Kit, Fes, MAPK1, PAK2, PDK1, PKCtheta, PKD2, Rsk1, and SGK.

Liposomes from novel photolabile phospholipids: light-induced unloading of small molecules as monitored by PFG NMR.

A molecular dithiane-based approach to synthesis of novel photolabile phospholipids is developed. These lipids are used in formulations with egg-POPC and cholesterol to prepare light-sensitive liposomes. Irradiation of such liposomes in PBS buffer (medium pressure mercury lamp, Pyrex filter, lambda > 300 nm) significantly increases the bilayer permeability and accelerates the release of entrapped small organic molecules by an order of magnitude.