RESUMO
DNA methyltransferase inhibitor (DNMTi) efficacy in solid tumors is limited. Colon cancer cells exposed to DNMTi accumulate lysine-27 trimethylation on histone H3 (H3K27me3). We propose this Enhancer of Zeste Homolog 2 (EZH2)-dependent repressive modification limits DNMTi efficacy. Here, we show that low-dose DNMTi treatment sensitizes colon cancer cells to select EZH2 inhibitors (EZH2is). Integrative epigenomic analysis reveals that DNMTi-induced H3K27me3 accumulates at genomic regions poised with EZH2. Notably, combined EZH2i and DNMTi alters the epigenomic landscape to transcriptionally up-regulate the calcium-induced nuclear factor of activated T cells (NFAT):activating protein 1 (AP-1) signaling pathway. Blocking this pathway limits transcriptional activating effects of these drugs, including transposable element and innate immune response gene expression involved in viral defense. Analysis of primary human colon cancer specimens reveals positive correlations between DNMTi-, innate immune response-, and calcium signaling-associated transcription profiles. Collectively, we show that compensatory EZH2 activity limits DNMTi efficacy in colon cancer and link NFAT:AP-1 signaling to epigenetic therapy-induced viral mimicry.
Assuntos
Neoplasias do Colo , Proteína Potenciadora do Homólogo 2 de Zeste , Histonas , Humanos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Histonas/metabolismo , Metilação , Transdução de Sinais , Fator de Transcrição AP-1/metabolismoRESUMO
Three undescribed sesquiterpenes, designed as pichinenoid A-C (1-3), along with nine known ones (4-12) were isolated from the stems and leaves of Picrasma chinensis. The new isolates including their absolute configurations were elucidated based on extensive spectroscopic methods, single crystal X-ray diffraction, and electronic circular dichroism (ECD) experiments, as well as comparison with literature data. Structurally, compounds 1 and 2 are descending sesquiterpenes, while pichinenoid C (3) is a rare sesquiterpene bearing a 2-methylenebut-3-enoic acid moiety at the C-6 side chain. All the isolated compounds were tested for their neuroprotective effects against the H2O2-induced damage on human neuroblastoma SH-SY5Y cells, and most of them showed moderate neuroprotective activity. Especially, compounds 1, 3-5, and 7 showed a potent neuroprotective effect at 25 or 50 µM. Moreover, the neuroprotective effects of compounds 1 and 4 were tested on a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) mouse model. Results of western blot and immunofluorescence indicated that compound 4 significantly counteract the toxicity of MPTP, and reversed the expression of tyrosine hydroxylase (TH) in substantia nigra (SN) and striatum (ST) of the mouse brain. Interestingly, western blot data suggested compound 4 also enhanced B-cell lymphoma-2 (Bcl-2) and heme oxygenase 1 (HO-1) expressions in the brain tissues from MPTP damaged mouse.
Assuntos
Fármacos Neuroprotetores , Picrasma , Folhas de Planta , Caules de Planta , Sesquiterpenos , Animais , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/isolamento & purificação , Sesquiterpenos/farmacologia , Sesquiterpenos/isolamento & purificação , Camundongos , Humanos , Linhagem Celular Tumoral , Estrutura Molecular , Picrasma/química , Caules de Planta/química , Folhas de Planta/química , Masculino , Heme Oxigenase-1/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , China , Compostos Fitoquímicos/farmacologia , Compostos Fitoquímicos/isolamento & purificação , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , Camundongos Endogâmicos C57BLRESUMO
Nonreceptor tyrosine kinase c-Src plays a crucial role in cell signaling and contributes to tumor progression. However, the development of selective c-Src inhibitors turns out to be challenging. In our previous study, we performed posttranslational modification-inspired drug design (PTMI-DD) to provide a plausible way for designing selective kinase inhibitors. In this study, after identifying a unique pocket comprising a less conserved cysteine and an autophosphorylation site in c-Src as well as a promiscuous covalent inhibitor, chemical optimization was performed to obtain (R)-LW-Srci-8 with nearly 75-fold improved potency (IC50 = 35.83 ± 7.21 nM). Crystallographic studies revealed the critical C-F···CâO interactions that may contribute to tight binding. The kinact and Ki values validated the improved binding affinity and decreased warhead reactivity of (R)-LW-Srci-8 for c-Src. Notably, in vitro tyrosine kinase profiling and cellular activity-based protein profiling (ABPP) cooperatively indicated a specific inhibition of c-Src by (R)-LW-Srci-8. Intriguingly, (R)-LW-Srci-8 preferentially binds to inactive c-Src with unphosphorylated Y419 both in vitro and in cells, subsequently disrupting the autophosphorylation. Collectively, our study demonstrated the feasibility of developing selective kinase inhibitors by cotargeting a nucleophilic residue and a posttranslational modification site and providing a chemical probe for c-Src functional studies.
Assuntos
Proteína Tirosina Quinase CSK , Inibidores de Proteínas Quinases , Humanos , Proteína Tirosina Quinase CSK/antagonistas & inibidores , Proteína Tirosina Quinase CSK/metabolismo , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Transdução de Sinais , Quinases da Família srcRESUMO
Picrachinentins A-F (1-6, respectively), six novel cyclopeptide alkaloid-type burpitides (CPABs), were isolated and fully elucidated from the EtOH extract of the stems and leaves of Picrasma chinensis. Structurally, compounds 1-6 have a 14-membered paracyclophane ring system that was closed through an ether bond between the ß-hydroxy amino acid and tyrosine and modified with a 4,5-methylenedioxybenzoyloxy (MDBz, 3 and 5) or hexanoyl (Hexa, 1, 2, 4, and 6) group at the N-terminus. Interestingly, this is the first report on the isolation and characterization of CPABs from plants of the Simaroubaceae family. In addition, all compounds showed a neuroprotective effect against H2O2-damaged SH-SY5Y cells. Compound 1 was further investigated for its neuroprotective activities using a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease animal model, and it dramatically improved MPTP-impaired motor behavioral performance. Biochemical analysis revealed compound 1 restored the tyrosine hydroxylase expression in the striatum of the MPTP-damaged mouse brain, which demonstrates its protective effect on dopaminergic neurons.
Assuntos
Alcaloides , Fármacos Neuroprotetores , Peptídeos Cíclicos , Picrasma , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/isolamento & purificação , Animais , Peptídeos Cíclicos/química , Peptídeos Cíclicos/farmacologia , Peptídeos Cíclicos/isolamento & purificação , Camundongos , Picrasma/química , Alcaloides/farmacologia , Alcaloides/química , Alcaloides/isolamento & purificação , Estrutura Molecular , Humanos , Peróxido de Hidrogênio/farmacologia , Peróxido de Hidrogênio/antagonistas & inibidores , Folhas de Planta/química , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/farmacologiaRESUMO
The RING E3 ubiquitin ligase UHRF1 is an established cofactor for DNA methylation inheritance. Nucleosomal engagement through histone and DNA interactions directs UHRF1 ubiquitin ligase activity toward lysines on histone H3 tails, creating binding sites for DNMT1 through ubiquitin interacting motifs (UIM1 and UIM2). Here, we profile contributions of UHRF1 and DNMT1 to genome-wide DNA methylation inheritance and dissect specific roles for ubiquitin signaling in this process. We reveal DNA methylation maintenance at low-density CpGs is vulnerable to disruption of UHRF1 ubiquitin ligase activity and DNMT1 ubiquitin reading activity through UIM1. Hypomethylation of low-density CpGs in this manner induces formation of partially methylated domains (PMD), a methylation signature observed across human cancers. Furthermore, disrupting DNMT1 UIM2 function abolishes DNA methylation maintenance. Collectively, we show DNMT1-dependent DNA methylation inheritance is a ubiquitin-regulated process and suggest a disrupted UHRF1-DNMT1 ubiquitin signaling axis contributes to the development of PMDs in human cancers.