Giovanni Cervi

Identification of N,1,4,4-Tetramethyl-8-{[4-(4-Methylpiperazin-1-Yl)Phenyl]Amino}-4,5-Dihydro-1H-Pyrazolo[4,3-H]Quinazoline-3-Carboxamide (Pha-848125), a Potent, Orally Available Cyclin Dependent Kinase Inhibitor.

The discovery of a novel class of inhibitors of cyclin dependent kinases (CDKs) is described. Starting from compound 1, showing good potency as inhibitor of CDKs but being poorly selective against a panel of serine-threonine and tyrosine kinases, new analogues were synthesized. Enhancement in selectivity, antiproliferative activity against A2780 human ovarian carcinoma cells, and optimization of the physical properties and pharmacokinetic profile led to the identification of highly potent and orally available compounds. Compound 28 (PHA-848125), which in the preclinical xenograft A2780 human ovarian carcinoma model showed good efficacy and was well tolerated upon repeated daily treatments, was identified as a drug candidate for further development. Compound 28 is currently undergoing phase I and phase II clinical trials.

Thieno[3,2-C]Pyrazoles: A Novel Class of Aurora Inhibitors with Favorable Antitumor Activity.

A novel series of 3-amino-1H-thieno[3,2-c]pyrazole derivatives demonstrating high potency in inhibiting Aurora kinases was developed. Here we describe the synthesis and a preliminary structure-activity relationship, which led to the discovery of a representative compound (38), which showed low nanomolar inhibitory activity in the anti-proliferation assay and was able to block the cell cycle in HCT-116 cell line. This compound demonstrated favorable pharmacokinetic properties and good efficacy in the HL-60 xenograft tumor model.

Discovery and Optimization of Pyrrolo[1,2-A]Pyrazinones Leads to Novel and Selective Inhibitors of Pim Kinases.

A novel series of PIM inhibitors was derived from a combined effort in natural product-inspired library generation and screening. The novel pyrrolo[1,2-a]pyrazinones initial hits are inhibitors of PIM isoforms with IC50 values in the low micromolar range. The application of a rational optimization strategy, guided by the determination of the crystal structure of the complex in the kinase domain of PIM1 with compound 1, led to the discovery of compound 15a, which is a potent PIM kinases inhibitor exhibiting excellent selectivity against a large panel of kinases, representative of each family. The synthesis, structure-activity relationship studies, and pharmacokinetic data of compounds from this inhibitor class are presented herein. Furthermore, the cellular activities including inhibition of cell growth and modulation of downstream targets are also described.

Discovery of 2-(cyclohexylmethylamino)pyrimidines as a new class of reversible valosine containing protein inhibitors.

Valosine-containing protein (VCP), also known as p97 or cdc48 in yeast, is a highly abundant protein belonging to the AAA ATPase family involved in a number of essential cellular functions, including ubiquitin-proteasome mediated protein degradation, Golgi reassembly, transcription activation, and cell cycle control. Altered expression of VCP has been detected in many cancer types sometimes associated with poor prognosis. Furthermore, VCP mutations are causative of some neurodegenerative disorders. In this paper we report the discovery, synthesis, and structure-activity relationships of substituted 2-aminopyrimidines, representing a new class of reversible VCP inhibitors. This class of compounds, identified in a HTS campaign against recombinant VCP, has been progressively expanded and manipulated to increase biochemical potency and gain cellular activity.

Abstract 2940: Identification of potent VCP/p97/CDC48 inhibitors with distinct biochemical mechanisms including a reversible, allosteric inhibitor that activates the unfolded protein response, induce autophagy and cancer cell death

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Valosin Containing Protein (VCP), also called p97 in mammals and cdc48 in yeast, is an ubiquitously expressed and essential AAA ATPase important in specific cellular processes including Endoplasmic Reticulum Associated Degradation (ERAD), Golgi reformation, membrane fusion and autophagy. VCP is a hexameric complex formed by six identical protomers, each composed of three domains: an N-terminal domain responsible for the interaction with co-factors and adaptor proteins, and two AAA ATPase domains, D1 and D2. VCP acts as a protein-directed molecular machine that converts energy derived from ATP hydrolysis into mechanical force to cause disassembly of multiprotein complexes or extraction of molecules from the membrane to be delivered to the proteasome for degradation. VCP also affects autophagy and aggresome formation processes. Overall, VCP plays a key role in cellular homeostasis. The clinical success of proteasome inhibitors, and recent advances in the preclinical development of molecules that interfere with protein folding and degradation has highlighted that cancer cells can be extremely sensitive to perturbation of protein homeostasis. The availability of small molecules that specifically inhibit VCP function would help to clarify whether VCP is a valid target for cancer therapy. We found that VCP silencing by siRNA induces cancer cell death in a variety of tumor cell lines, and examined the main cellular pathways modulated upon VCP ablation, thus identifying biomarkers suitable for characterizing the cellular activity of VCP inhibitors. We then performed a High Throughput Screening campaign using recombinant VCP and identified multiple compound classes that inhibit VCP function with distinct biochemical mechanisms. Initial hits included a compound that covalently modifies VCP, representatives of two classes of ATP-sensitive inhibitors, and an inhibitor class characterized by a novel allosteric mechanism of action. Chemical expansion of initial hits resulted in improvement of biochemical potency, yielding highly active derivatives for each class, suggesting that VCP is a druggable target. Notably, the binding site of the allosteric class of inhibitors was identified by photo-affinity labeling in combination with available structural data. We will present data on a potent (30 nM) and specific compound emerging from this class that displays anti-proliferative activity related to the modulation of direct VCP biomarkers, activation of the Unfolded Protein Response (UPR) and perturbation of autophagy, which ultimately result in cancer cell death. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2940. doi:1538-7445.AM2012-2940