Development of 2-(4-pyridyl)-benzimidazoles as PKN2 chemical tools to probe cancer

Kinases are signalling proteins which have proven to be successful targets for the treatment of a variety of diseases, predominantly in cancers. However, only a small proportion of kinases (<20%) have been investigated for their therapeutic viability, likely due to the lack of available chemical tools across the kinome. In this work we describe initial efforts in the development of a selective chemical tool for protein kinase N2 (PKN2), a relatively unexplored kinase of interest in several types of cancer. The most successful compound, 5, has a measured IC50 of 0.064 μM against PKN2, with ca. 17-fold selectivity over close homologue, PKN1

Development of dihydropyrrolopyridinone-based PKN2/PRK2 chemical tools to enable drug discovery

The Protein Kinase N proteins (PKN1, PKN2 and PKN3) are Rho GTPase effectors. They are involved in several biological processes such as cytoskeleton organization, cell mobility, adhesion, and cell cycle. Recently PKNs have been reported as essential for survival in several tumor cell lines, including prostate and breast cancer. Here, we report the development of dihydropyrrolopyridinone-based inhibitors for PKN2 and its closest homologue, PKN1, and their associated structure–activity relationship (SAR). Our studies identified a range of molecules with high potency exemplified by compound 8 with Ki = 8 nM for PKN2 and 14x selectivity over PKN1. Membrane permeability and target engagement for PKN2 were assessed by a NanoBRET cellular assay. Importantly, good selectivity across the wider human kinome and other kinase family members was achieved. These compounds provide strong starting points for lead optimization to PKN1/2 development compounds

Unsaturated fatty acids as high-affinity ligands of the C-terminal Per-ARNT-Sim domain from the Hypoxia-inducible factor 3α\alpha

Hypoxia-inducible transcription factors (HIF) form heterodimeric complexes that mediate cell responses to hypoxia. The oxygen-dependent stability and activity of the HIF-α subunits is traditionally associated to post-translational modifications such as hydroxylation, acetylation, ubiquitination, and phosphorylation. Here we report novel evidence showing that unsaturated fatty acids are naturally occurring, non-covalent structural ligands of HIF-3$\alpha$, thus providing the initial framework for exploring its exceptional role as a lipid sensor under hypoxia

Binding and structural analyses of potent inhibitors of the human Ca2+/calmodulin dependent protein kinase kinase 2 (CAMKK2) identified from a collection of commercially-available kinase inhibitors

Calcium/Calmodulin-dependent Protein Kinase Kinase 2 (CAMKK2) acts as a signaling hub, receiving signals from various regulatory pathways and decoding them via phosphorylation of downstream protein kinases - such as AMPK (AMP-activated protein kinase) and CAMK types I and IV. CAMKK2 relevance is highlighted by its constitutive activity being implicated in several human pathologies. However, at present, there are no selective small-molecule inhibitors available for this protein kinase. Moreover, CAMKK2 and its closest human homolog, CAMKK1, are thought to have overlapping biological roles. Here we present six new co-structures of potent ligands bound to CAMKK2 identified from a library of commercially-available kinase inhibitors. Enzyme assays confirmed that most of these compounds are equipotent inhibitors of both human CAMKKs and isothermal titration calorimetry (ITC) revealed that binding to some of these molecules to CAMKK2 is enthalpy driven. We expect our results to advance current efforts to discover small molecule kinase inhibitors selective to each human CAMKK9CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP465651/2014-388887.136342/2017-00; 88887.136432/2017-00; 88887.136437/2017-00; 88887.146077/2017-002019/14275-8; 2016/09041-0; 2017/05697-0Brazilian agency FAPESP (Fundacao de Amparo a Pesquisa do Estado de Sao Paulo)Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP) [2013/50724-5, 2014/50897-0]; Brazilian agency CNPq (Conselho Nacional de Desenvolvimento Cientifico e Tecnologico)National Council for Scientific and Technological Development (CNPq) [465651/2014-3]; AbbVie; Bayer Pharma AG; Boehringer IngelheimBoehringer Ingelheim; Canada Foundation for InnovationCanada Foundation for Innovation; Eshelman Institute for Innovation; Genome CanadaGenome Canada; Innovative Medicines Initiative (EU/EFPIA) [ULTRA-DD] [115766]; JanssenJohnson & Johnson USAJanssen Biotech Inc; Merck KGaA Darmstadt Germany; MSD; Novartis Pharma AG; Ontario Ministry of Economic Development and Innovation; PfizerPfizer; TakedaTakeda Pharmaceutical Company Ltd; Wellcome [106169/ZZ14/Z]; National Cancer Institute of the National Institutes of HealthUnited States Department of Health & Human ServicesNational Institutes of Health (NIH) - USANIH National Cancer Institute (NCI) [R01CA218442]; CAPES (Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior)CAPES [88887.136342/2017-00, 88887.136432/2017-00, 88887.136437/2017-00, 88887.146077/2017-00]; FAPESPFundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP) [2019/14275-8, 2016/09041-0, 2017/05697-0]; National Institute of General Medical Sciences from the National Institutes of Health [GU51510, GU56413, P41 GM103403]; NIH-ORIP HEI grant [S10 RR029205]; DOE Office of Science by Argonne National LaboratoryUnited States Department of Energy (DOE) [DE-AC02-06CH11357