nan Hiromi

Total synthesis and biological evaluation of (-)-apicularen A and its analogues.

The total synthesis of (-)-apicularen A (1), a highly cytostatic 12-membered macrolide, and its analogues is described. The convergent and distinct approach not only provides 1, but also opens the opportunity to synthesize C10-C11 functional analogues of 1. The key steps of the total synthesis include assembling of iodoalkene 12 and aldehyde 13by Nozaki-Hiyama-Kishi (NHK) coupling, stereospecific construction of 2,6-trans-disubstituted dihydropyran by Pd(II)-catalyzed 1,3-chirality transfer reaction, and Yamaguchi macrolactonization. The (17E,20Z,22Z)-heptadienoylenamine moiety in the side chain is installed by an efficient Cu(I)-mediated coupling to complete the synthesis. Analogues of C11-epi-, C11-deoxy-C10-α-hydroxy-, and C10-C11 dehydrated apicularen A 3-5 were also prepared. Cytostatic activities of (-)-apicularen A and the three analogues for three different cancer cell lines are described.

Gamma-Glutamylcyclotransferase: A Novel Target Molecule for Cancer Diagnosis and Treatment

Gamma-glutamylcyclotransferase (GGCT) is one of the major enzymes involved in glutathione metabolism. However, its gene locus was unknown for many years. Recently, the gene for GGCT was found to be identical to C7orf24, which is registered as a hypothetical protein. Orthologs have been found in bacteria, plants, and nematodes as well as higher organisms, and the GGCT gene is highly preserved among a wide range of species. GGCT (C7orf24) was also reported as an upregulated protein in various cancers. Although the function of GGCT in cancer cells has not been determined, the following important activities have been reported: (1) high expression in various cancer tissues and cancer cell lines, (2) low expression in normal tissues, (3) inhibition of cancer cell proliferation via anti-GGCT RNAi, (4) inhibition of cancer cell invasion and migration via anti-GGCT RNAi, (5) an epigenetic transcriptional regulation in cancer cells, and (6) an antitumor effect in cancer-bearing xenograft mice. Therefore, GGCT is promising as a diagnostic marker and a therapeutic target for various cancers. This review summarizes these interesting findings.

A fluorogenic probe for γ-glutamyl cyclotransferase: application of an enzyme-triggered O-to-N acyl migration-type reaction.

Light it up: human chromosome 7 ORF 24, a tumor-related protein, has been identified as a γ-glutamyl cyclotransferase (GGCT) in the glutathione homeostasis cycle. The singular substrate preference of the enzyme has hampered chemical probe development, and no fluorogenic probe has been reported. Here we report the first fluorogenic dipeptide probe, LISA-4, which should contribute toward further understanding of GGCT.

Synthesis and GGCT Inhibitory Activity of N-Glutaryl-L-alanine Analogues.

γ-Glutamylcyclotransferase (GGCT) is an important enzyme that cleaves γ-glutamyl-amino acid in the γ-glutamyl cycle to release 5-oxoproline and amino acid. Eighteen N-acyl-L-alanine analogues including eleven new compounds have been synthesized and examined for their inhibitory activity against recombinant human GGCT protein. Simple N-glutaryl-L-alanine was found to be the most potent inhibitor for GGCT. Other N-glutaryl-L-alanine analogues having methyl and dimethyl substituents at the 2-position were moderately effective, while N-(3R-aminoglutary)-L-alanine, the substrate having an (R)-amino group at the 3-position or N-(N-methyl-3-azaglutaryl)-L-alanine, the substrate having an N-methyl substituent on the 3-azaglutaryl carbon, in constract, exhibited excellent inhibition properties.

Depletion of γ-glutamylcyclotransferase inhibits breast cancer cell growth via cellular senescence induction mediated by CDK inhibitor upregulation

BackgroundChromosome 7 open reading frame 24 (C7orf24) was originally identified as a highly expressed protein in various types of cancer, and later shown to be a γ-glutamylcyclotransferase (GGCT). GGCT depletion in cancer cells has anti-proliferative effects in vitro and in vivo, and it is therefore considered a promising candidate as a therapeutic target. However, the cellular events induced by GGCT depletion remain unclear.MethodsGGCT was depleted by siRNA in MCF7, MDA-MB-231, PC3, A172, Hela, and LNCaP cells. Induction of cellular senescence was evaluated with senescence-associated β-galactosidase (SA-β-Gal) staining. Expression levels of p21WAF1/CIP1 and p16INK4A were assessed by qRT-PCR and Western blotting. Effects of simultaneous double knockdown of p21WAF1/CIP1 and p16INK4A together with GGCT on cell cycle regulation and cell growth was measured by flow cytometry, and trypan blue dye exclusion test.ResultsWe found that GGCT knockdown induces significant cellular senescence in various cancer cells. Cyclin dependent kinase inhibitor p21WAF1/CIP1 and/or p16INK4A were upregulated in all cell lines tested. Simultaneous knockdown of p21WAF1/CIP1 recovered the cell cycle arrest, attenuated cellular senescence induction, and rescued the subsequent growth inhibition in GGCT-silenced MCF7 breast cancer cells. In contrast, in GGCT silenced MDA-MB-231 breast cancer cells, GGCT depletion upregulated p16INK4A, which played a regulatory role in senescence induction, instead of p21WAF1/CIP1.ConclusionsOur findings demonstrate that induction of cellular senescence mediated by the upregulation of cyclin-dependent kinase inhibitors is a major event underlying the anti-proliferative effect of GGCT depletion in breast cancer cells, highlighting the potential of GGCT blockade as a therapeutic strategy to induce cellular senescence.

Design of fluorogenic probes and fluorescent-tagged inhibitors for γ-glutamyl cyclotransferase

A tumor‐related protein, human chromosome 7 ORF 24 (C7orf24), is involved in regulation of the glutathione homeostasis cycle as a γ‐glutamyl cyclotransferase (GGCT). The singular substrate preference of this enzyme had long hampered its chemical probe development. That is, substrate of GGCT is definitely ‘γ‐Glu‐Xaa’, where Xaa is an L‐α‐amino acid. Based on the structure of substrates and GGCT fluorogenic probes, LISA‐4/101, we succesfully developed a fluorescent‐tagged inhibitor, gKFA. These chemical tools will assist cancer‐related researches in the future. Copyright

A Novel Prodrug of a γ‐Glutamylcyclotransferase Inhibitor Suppresses Cancer Cell Proliferation in vitro and Inhibits Tumor Growth in a Xenograft Mouse Model of Prostate Cancer

γ‐Glutamylcyclotransferase (GGCT) depletion inhibits cancer cell proliferation. However, whether the enzymatic activity of GGCT is critical for the regulation of cancer cell growth remains unclear. In this study, a novel diester‐type cell‐permeable prodrug, pro‐GA, was developed based on the structure of N‐glutaryl‐l‐alanine (GA), by structure optimization using temporary fluorophore‐tagged prodrug candidates. The antiproliferative activity of pro‐GA was demonstrated using GGCT‐overexpressing NIH‐3T3 cells and human cancer cells including MCF7, HL‐60, and PC3 cells. By contrast, normal cells were not significantly affected by pro‐GA treatment. Moreover, pro‐GA administration exhibited anticancer effects in a xenograft model using immunocompromised mice inoculated with PC3 cells. These results indicate that the enzymatic activity of GGCT accelerates tumor growth and that GGCT inhibition is a promising therapeutic strategy for the treatment of GGCT‐overexpressing tumors.

Prohibitin-2 is a novel regulator of p21 WAF1/CIP1 induced by depletion of γ-glutamylcyclotransferase

Previous studies show that gamma-glutamylcyclotransferase (GGCT) is expressed at high levels in various cancer tissues and that its knockdown inhibits MCF7 cancer cell growth via upregulation of p21WAF1/CIP1 (p21). However, the detailed underlying mechanism is unclear. Here, we used yeast two-hybrid screening and co-immunoprecipitation to identify Prohibitin-2 (PHB2) as a novel protein that interacts with GGCT. We also show that nuclear expression of PHB2 in MCF7 cells falls upon GGCT knockdown, and that overexpression of PHB2 inhibits p21 upregulation. A chromatin immunoprecipitation assay revealed that nuclear PHB2 proteins bind to the p21 promoter, and that this interaction is abrogated by GGCT knockdown. Moreover, knockdown of PHB2 alone led to significant upregulation of p21 and mimicked the cellular events induced by GGCT depletion, including G0/G1 arrest, cellular senescence, and growth inhibition, in a p21 induction-dependent manner. Taken together, the results indicate that PHB2 plays a central role in p21 upregulation following GGCT knockdown and as such may promote deregulated proliferation of cancer cells by suppressing p21.

Mechanisms of Tumor Growth Inhibition by Depletion of γ-Glutamylcyclotransferase (GGCT): A Novel Molecular Target for Anticancer Therapy

γ-Glutamylcyclotransferase (GGCT), which is one of the major enzymes involved in glutathione metabolism, is upregulated in a wide range of cancers—glioma, breast, lung, esophageal, gastric, colorectal, urinary bladder, prostate, cervical, ovarian cancers and osteosarcoma—and promotes cancer progression; its depletion leads to the suppression of proliferation, invasion, and migration of cancer cells. It has been demonstrated that the suppression or inhibition of GGCT has an antitumor effect in cancer-bearing xenograft mice. Based on these observations, GGCT is now recognized as a promising therapeutic target in various cancers. This review summarizes recent advances on the mechanisms of the antitumor activity of GGCT inhibition.