Discovery of an L-alanine ester prodrug of the Hsp90 inhibitor, MPC-3100.

Various types of Hsp90 inhibitors have been and continue to undergo clinical investigation. One development candidate is the purine-based, synthetic Hsp90 inhibitor 1 (MPC-3100), which successfully completed a phase I clinical study. However, further clinical development of 1 was hindered by poor solubility and consequent formulation issues and promoted development of a more water soluble prodrug.

Discovery of a new HIV-1 inhibitor scaffold and synthesis of potential prodrugs of indazoles.

A new oxazole scaffold showing great promise in HIV-1 inhibition has been discovered by cell-based screening of an in-house library and scaffold modification. Follow-up SAR study focusing on the 5-aryl substituent of the oxazole core has identified 4k (EC50=0.42μM, TI=50) as a potent inhibitor.

Discovery of (2S)-1-[4-(2-{6-amino-8-[(6-bromo-1,3-benzodioxol-5-yl)sulfanyl]-9H-purin-9-yl}ethyl)piperidin-1-yl]-2-hydroxypropan-1-one (MPC-3100), a purine-based Hsp90 inhibitor.

Modulation of Hsp90 (heat shock protein 90) function has been recognized as an attractive approach for cancer treatment, since many cancer cells depend on Hsp90 to maintain cellular homeostasis. This has spurred the search for small-molecule Hsp90 inhibitors. Here we describe our lead optimization studies centered on the purine-based Hsp90 inhibitor 28a containing a piperidine moiety at the purine N9 position.

Characterization of the cellular and antitumor effects of MPI-0479605, a small-molecule inhibitor of the mitotic kinase Mps1.

Mps1 is a dual specificity protein kinase that is essential for the bipolar attachment of chromosomes to the mitotic spindle and for maintaining the spindle assembly checkpoint until all chromosomes are properly attached. Mps1 is expressed at high levels during mitosis and is abundantly expressed in cancer cells. Disruption of Mps1 function induces aneuploidy and cell death.

A novel series of IKKβ inhibitors part I: Initial SAR studies of a HTS hit.

A novel series of (E)-1-((2-(1-methyl-1H-imidazol-5-yl) quinolin-4-yl) methylene) thiosemicarbazides was discovered as potent inhibitors of IKKβ. In this Letter we document our early efforts at optimization of the quinoline core, the imidazole and the semithiocarbazone moiety. Most potency gains came from substitution around the 6- and 7-positions of the quinoline ring.

A novel series of IKKβ inhibitors part II: description of a potent and pharmacologically active series of analogs.

A novel series of (E)-1-((2-(1-methyl-1H-imidazol-5-yl) quinolin-4-yl) methylene) thiosemicarbazides was discovered as potent inhibitors of IKKβ. In this Letter we document our efforts at further optimization of this series, culminating in 2 with submicromolar potency in a HWB assay and efficacy in a CIA mouse model.

Inhibition of eEF2-K by thieno[2,3-b]pyridine analogues.

Several series of thieno[2-3-b]pyridine analogues were synthesized and screened for inhibitory activity against eukaryotic elongation factor-2 kinase (eEF2-K). Modifications around several regions of the lead molecules were made, with a ring fusion adjacent to the nitrogen on the thienopyridine core being critical for activity. The most active compound 34 shows an IC(50) of 170 nM against eEF2-K in vitro.

Triterpene based compounds with potent anti-maturation activity against HIV-1.

Efforts towards developing orally bioavailable HIV-1 maturation inhibitors starting from betulinic acid 1 are described. SAR resulted in improved potency, physicochemical properties, and enhanced oral absorption in rats.

Discovery of 2-chloro-N-(4-methoxyphenyl)-N-methylquinazolin-4-amine (EP128265, MPI-0441138) as a potent inducer of apoptosis with high in vivo activity.

Using a live cell, high-throughput caspase-3 activator assay, we have identified a novel series of 4-anilinoquinazolines as inducers of apoptosis. In this report, we discuss the discovery of 2-chloro-N-(4-methoxyphenyl)-N-methylquinazolin-4-amine, compound 2b (EP128265, MPI-0441138) as a highly active inducer of apoptosis (EC50 for caspase activation of 2 nM) and as a potent inhibitor of cell proliferation (GI50 of 2 nM) in T47D cells. Compound 2b inhibited tubulin polymerization, was effective in cells overexpressing ABC transporter Pgp-1, and was efficacious in the MX-1 human breast and PC-3 prostate cancer mouse models.

MPC-6827: a small-molecule inhibitor of microtubule formation that is not a substrate for multidrug resistance pumps.

A novel series of 4-arylaminoquinazolines were identified from a cell-based screening assay as potent apoptosis inducers. Through structure-activity relationship studies, MPC-6827 and its close structural analogue, MPI-0441138, were discovered as proapoptotic molecules and mitotic inhibitors with potencies at low nanomolar concentrations in multiple tumor cell lines. Photoaffinity and radiolabeled analogues of MPC-6827 were found to bind a 55-kDa protein, and this binding was competed by MPC-6827, paclitaxel, and colchicine, but not vinblastine.

Hydrophilic, pro-drug analogues of T138067 are efficacious in controlling tumor growth in vivo and show a decreased ability to cross the blood brain barrier.

The novel anticancer compound T138067 is an irreversible inhibitor of tubulin polymerization. Amides 3-6 were synthesized using standard methodologies and determined to be significantly less lipophilic than T138067 based on logP calculations. Tubulin polymerization and [(3)H]-T138067 competition assays revealed that these amides are pro-drugs for parent aniline 2.

Novel antineoplastic agents with efficacy against multidrug resistant tumor cells.

A novel series of pentafluorobenzenesulfonamides has been shown to inhibit the growth of a variety of human tumor cell lines. Among the cell types against which these agents were evaluated were the multidrug resistant (MDR) cell lines MCF-7/ADR and P388/ADR. The cytotoxic activity of members of this series of compounds was not affected by the multidrug resistant pump in MCF-7/ADR or P388/ADR cells.

Identification of ERF-1 as a member of the AP2 transcription factor family.

The ERF-1 transcription factor was previously shown to be involved in the regulation of estrogen receptor (ER) gene transcription in hormonally responsive breast and endometrial carcinomas. In this study we sought to identify the gene for ERF-1. ERF-1 activates ER gene transcription by binding to the imperfect palindrome CCCTGCGGGG within the promoter of the ER gene.

Activate NF-kappa B or die?

Activation of the transcription factor NF-kappaB has been linked to apoptosis, with the factor playing either an anti-apoptotic or a pro-apoptotic role, depending on the type of cell in which it is expressed.

Systematic mutational analysis of the death domain of the tumor necrosis factor receptor 1-associated protein TRADD.

Tumor necrosis factor receptor 1 (TNF-R1) mediates most of the biological properties of TNF including activation of the transcription factor NF-kappaB and programmed cell death. An approximately 80-amino acid region within the intracellular domain of the receptor, termed the death domain, is required for signaling NF-kappaB activation and cytotoxicity. A TNF-R1-associated protein TRADD has been discovered that interacts with the death domain of the receptor.

TNF-dependent recruitment of the protein kinase RIP to the TNF receptor-1 signaling complex.

The death domain of tumor necrosis factor (TNF) receptor-1 (TNFR1) triggers distinct signaling pathways leading to apoptosis and NF-kappa B activation through its interaction with the death domain protein TRADD. Here, we show that TRADD interacts strongly with RIP, another death domain protein that was shown previously to associate with Fas antigen. We also show that RIP is a serine-threonine kinase that is recruited by TRADD to TNFR1 in a TNF-dependent process.

Sp1 is a component of the cytokine-inducible enhancer in the promoter of vascular cell adhesion molecule-1.

Transcription of the vascular cell adhesion molecule-1 (VCAM-1) gene in endothelial cells is induced by the inflammatory cytokines interleukin-1 beta, tumor necrosis factor-alpha, and lipopolysaccharide. Previous studies demonstrated that the cytokine-response region in the VCAM1 promoter contains binding sites for the transcription factors nuclear factor-kappa B (NF-kappa B) and interferon regulatory factor-1. Using a saturation mutagenesis approach, we report that the cytokine-inducible enhancer consists of these previously characterized elements and a novel region located 3' of the NF-kappa B sites.

Induction of the gene encoding mucosal vascular addressin cell adhesion molecule 1 by tumor necrosis factor alpha is mediated by NF-kappa B proteins.

Mucosal vascular addressin cell adhesion molecule 1 (MAdCAM-1) is involved in trafficking of lymphocytes to mucosal endothelium. Expression of MAdCAM-1 is induced in the murine endothelial cell line bEnd.3 by tumor necrosis factor alpha (TNF-alpha), interleukin 1, and bacterial lipopolysaccharide.

Refolding and reconstitution of functionally active complexes of human leukocyte antigen DR2 and myelin basic protein peptide from recombinant alpha and beta polypeptide chains.

Major histocompatibility complex (MHC) class II molecules are cell surface heterodimeric glycoproteins consisting of one alpha and one beta polypeptide chain of similar size. These molecules play a critical role in immune recognition by displaying processed antigens to CD4-positive T helper cells. Several attempts to express the MHC class II molecules by recombinant methods in various systems resulted in either failure or poor recovery of the intact heterodimer.

Regulatory elements and transcription factors controlling basal and cytokine-induced expression of the gene encoding intercellular adhesion molecule 1.

The gene encoding intercellular adhesion molecule 1 (ICAM-1) is transcriptionally induced in response to inflammatory and immunomodulatory cytokines. To investigate the mechanisms controlling ICAM-1 gene expression, we have identified regulatory DNA sequences responsible for maintaining basal and mediating induced transcription in response to tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma). Regulatory elements centered 115, 60, and 40 bp upstream from the ICAM-1 transcription start site were implicated in cytokine-independent gene expression.

Three NF-kappa B binding sites in the human E-selectin gene required for maximal tumor necrosis factor alpha-induced expression.

Transcription of the gene encoding the endothelial cell-leukocyte adhesion molecule (ELAM-1; E-selectin) is induced in response to various cytokines, including tumor necrosis factor-alpha (TNF-alpha) and interleukin-1. A DNase I-hypersensitive site in the 5' proximal promoter region of the E-selectin gene is observed in human umbilical vein endothelial cells only following TNF-alpha treatment, suggesting the presence of a TNF-alpha-inducible element close to the transcriptional start site. Transient transfection studies in endothelial cells demonstrated that 170 bp of upstream sequences is sufficient to confer TNF-alpha inducibility.

Intramolecular charge heterogeneity in purified major histocompatibility class II alpha and beta polypeptide chains.

Major histocompatibility (MHC) class II antigens are heterodimeric cell surface glycoproteins consisting of an alpha and a beta chain. Although one-dimensional SDS-polyacrylamide gel electrophoresis analysis of purified MHC class II antigens shows a single diffuse band for each chain, multiple spots of identical molecular size were observed for each chain when analyzed by two-dimensional electrophoresis. The basis of this heterogeneity has not been clearly defined and has been predicted partially to be due to glycosylation and/or phosphorylation of the mature protein.

The ATP-binding component of a prokaryotic traffic ATPase is exposed to the periplasmic (external) surface.

The membrane-bound complex of bacterial periplasmic permeases consists of two hydrophobic integral membrane proteins and two copies of a hydrophilic ATP-binding protein. The ATP-binding proteins from all periplasmic permeases display a high level of sequence similarity and are referred to as "conserved components." The conserved component from the histidine permease, HisP, has been postulated on the basis of genetic evidence to be accessible at the exterior membrane surface, in contrast to the commonly postulated association with the interior membrane surface as peripheral membrane proteins.

Transcription factor based therapeutics: drugs of the future?

During the past decade there has been an explosion of information relating to our understanding of eukaryotic gene expression. The identification of transcription factors as the key regulatory molecules in this process, and the analysis of their structure and function have revealed that these proteins are potential targets for therapeutic intervention.