Structural insights into the high basal activity and inverse agonism of the orphan receptor GPR6 implicated in Parkinson's disease.

GPR6 is an orphan G protein-coupled receptor with high constitutive activity found in D2-type dopamine receptor-expressing medium spiny neurons of the striatopallidal pathway, which is aberrantly hyperactivated in Parkinson's disease. Here, we solved crystal structures of GPR6 without the addition of a ligand (a pseudo-apo state) and in complex with two inverse agonists, including CVN424, which improved motor symptoms in patients with Parkinson's disease in clinical trials. In addition, we obtained a cryo-electron microscopy structure of the signaling complex between GPR6 and its cognate G heterotrimer.

Discovery of TAK-041: a Potent and Selective GPR139 Agonist Explored for the Treatment of Negative Symptoms Associated with Schizophrenia.

The orphan G-protein-coupled receptor GPR139 is highly expressed in the habenula, a small brain nucleus that has been linked to depression, schizophrenia (SCZ), and substance-use disorder. High-throughput screening and a medicinal chemistry structure-activity relationship strategy identified a novel series of potent and selective benzotriazinone-based GPR139 agonists. Herein, we describe the chemistry optimization that led to the discovery and validation of multiple potent and selective in vivo GPR139 agonist tool compounds, including our clinical candidate TAK-041, also known as NBI-1065846 (compound ).

Inhibition of RNA polymerase I as a therapeutic strategy to promote cancer-specific activation of p53.

Increased transcription of ribosomal RNA genes (rDNA) by RNA Polymerase I is a common feature of human cancer, but whether it is required for the malignant phenotype remains unclear. We show that rDNA transcription can be therapeutically targeted with the small molecule CX-5461 to selectively kill B-lymphoma cells in vivo while maintaining a viable wild-type B cell population. The therapeutic effect is a consequence of nucleolar disruption and activation of p53-dependent apoptotic signaling.

Combined inhibition of EGFR and CK2 augments the attenuation of PI3K-Akt-mTOR signaling and the killing of cancer cells.

Ser/Thr protein kinase CK2 regulates multiple processes that play important roles in the sensitivity of cancer to epidermal growth factor receptor targeting therapeutics, including PI3K-Akt-mTOR signaling, Hsp90 activity, and inhibition of apoptosis. We hypothesized that top-down inhibition of EGFR, combined with lateral suppression of multiple oncogenic pathways by targeting CK2, would create a pharmacologic synthetic lethal event and result in an improved cancer therapy compared to EGFR inhibition alone. This hypothesis was tested by combining CX-4945, a first-in-class clinical stage inhibitor of CK2, with the EGFR tyrosine kinase inhibitor, erlotinib, in vitro and in vivo in models of non-small cell lung carcinoma, NCI-H2170, and squamous cell carcinoma, A431.

Discovery of CX-6258. A Potent, Selective, and Orally Efficacious pan-Pim Kinases Inhibitor.

Structure-activity relationship analysis in a series of 3-(5-((2-oxoindolin-3-ylidene)methyl)furan-2-yl)amides identified compound 13, a pan-Pim kinases inhibitor with excellent biochemical potency and kinase selectivity. Compound 13 exhibited in vitro synergy with chemotherapeutics and robust in vivo efficacy in two Pim kinases driven tumor models.

CK2 inhibitor CX-4945 suppresses DNA repair response triggered by DNA-targeted anticancer drugs and augments efficacy: mechanistic rationale for drug combination therapy.

Drug combination therapies are commonly used for the treatment of cancers to increase therapeutic efficacy, reduce toxicity, and decrease the incidence of drug resistance. Although drug combination therapies were originally devised primarily by empirical methods, the increased understanding of drug mechanisms and the pathways they modulate provides a unique opportunity to design combinations that are based on mechanistic rationale. We have identified protein kinase CK2 as a promising therapeutic target for combination therapy, because CK2 regulates not just one but many oncogenic pathways and processes that play important roles in drug resistance, including DNA repair, epidermal growth factor receptor signaling, PI3K/AKT/mTOR signaling, Hsp90 machinery activity, hypoxia, and interleukin-6 expression.

Protein kinase CK2 modulates IL-6 expression in inflammatory breast cancer.

Inflammatory breast cancer is driven by pro-angiogenic and pro-inflammatory cytokines. One of them Interleukin-6 (IL-6) is implicated in cancer cell proliferation and survival, and promotes angiogenesis, inflammation and metastasis. While IL-6 has been shown to be upregulated by several oncogenes, the mechanism behind this phenomenon is not well characterized.

Pre-clinical characterization of CX-4945, a potent and selective small molecule inhibitor of CK2 for the treatment of cancer.

In this article we describe the preclinical characterization of 5-(3-chlorophenylamino) benzo[c][2,6]naphthyridine-8-carboxylic acid (CX-4945), the first orally available small molecule inhibitor of protein CK2 in clinical trials for cancer. CX-4945 was optimized as an ATP-competitive inhibitor of the CK2 holoenzyme (Ki = 0.38 nM).

A method for detecting and preventing negative RNA interference in preparation of lentiviral vectors for siRNA delivery.

The lentiviral vector is a useful tool for delivery of hairpin siRNA (shRNA) into mammalian cells. However, the efficiency of this system for carrying double-stranded siRNA (dsRNA) has not been explored. In this study we cloned the two forms of siRNA-coding sequence, a palindromic DNA with a spacer loop for shRNA and a double-stranded DNA with opposing Pol III promoters for dsRNA, into lentiviral DNA vectors, and compared their viral vector production yields.

Structural and biochemical characterization of the therapeutic Anabaena variabilis phenylalanine ammonia lyase.

We have recently observed promising success in a mouse model for treating the metabolic disorder phenylketonuria with phenylalanine ammonia lyase (PAL) from Rhodosporidium toruloides and Anabaena variabilis. Both molecules, however, required further optimization in order to overcome problems with protease susceptibility, thermal stability, and aggregation. Previously, we optimized PAL from R.

A novel RNAi library based on partially randomized consensus sequences of nuclear receptors: identifying the receptors involved in amyloid beta degradation.

Combinatorial gene inactivation using an RNAi library is a powerful approach to discovering novel functional genes. However, generation of a comprehensive RNAi library remains technically challenging. In this report, we describe a simple and novel approach to designing gene-family-specific RNAi libraries by targeting conserved motifs using degenerate oligonucleotides.

A 96-well surrogate survival assay coupled with a special short interfering RNA vector for assessing cancer gene targets with enhanced signal/noise ratio and its utility in HTS for cancer therapeutic targets.

Transient transfection of short interfering RNAs to inactivate cancer therapeutic genes in cancer cells is an important method to induce therapeutic phenotypes (cell apoptosis, growth arrest, etc.) for cancer target validation. These phenotypes can be initially assessed by cell survival via colorimetric/fluorescence readings, e.