Antitumor Agents XLVIII: Structure-Activity Relationships of Quassinoids as In Vitro Protein Synthesis Inhibitors of P-388 Lymphocytic Leukemia Tumor Cell Metabolism

Liou, Y.F.; Hall, Iris H.; Okano, Masayoshi; Lee, K.H.; Chaney, Stephen G.

doi:10.1002/jps.2600710414

Cited by 22 publications

(7 citation statements)

References 16 publications

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“…A pulse analysis using a methionine analogue to monitor newly synthesized proteins confirmed that nascent protein synthesis is essentially abolished following brusatol treatment. A previous study suggested that brusatol inhibited formation of the first peptide bond between puromycin and [3H]methionyl-transfer RNA bound to the initiation complex (40). Why this mechanism would cause increased levels of ribosomal components is not clear.…”

Section: Resultsmentioning

confidence: 98%

Application of Mass Spectrometry Profiling to Establish Brusatol as an Inhibitor of Global Protein Synthesis

Vartanian¹,

Taylur

Lee³

et al. 2016

Molecular & Cellular Proteomics

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The KEAP1/Nrf2 pathway senses and responds to changes in intracellular oxidative stress. Mutations that result in constitutive activation of Nrf2 are present in several human tumors, especially non-small cell lung cancer. Therefore, compounds that inhibit Nrf2 activity might be beneficial in treating patients whose tumors show activation of this pathway. Recent reports suggest that the natural product brusatol can potently and selectively inhibit Nrf2 activity, resulting in cell cytotoxicity, and can be effectively combined with chemotherapeutic agents. Here, we analyzed the effects of brusatol on the cellular proteome in the KEAP1 mutant non-small cell lung cancer cell line A549. Brusatol was found to rapidly and potently decrease the expression of the majority of detected proteins, including Nrf2. The most dramatically decreased proteins are those that display a short half-life, like Nrf2. This effect was confirmed by restricting the analysis to newly synthesized proteins using a labeled methionine analogue. Moreover, brusatol increased the expression of multiple components of the ribosome, suggesting that it regulates the function of this macromolecular complex. Finally, we show that brusatol induces its potent cellular cytotoxicity effects on multiple cancer cell lines in a manner independent of KEAP1/Nrf2 activity and with a profile similar to the protein translation inhibitor silvestrol. In conclusion, our data show that the activity of brusatol is not restricted to Nrf2 but,

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Section: Resultsmentioning

confidence: 98%

Application of Mass Spectrometry Profiling to Establish Brusatol as an Inhibitor of Global Protein Synthesis

Vartanian¹,

Taylur

Lee³

et al. 2016

Molecular & Cellular Proteomics

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“…Inhibition of overall protein synthesis was observed in rabbit reticulocytes, as well as in cell lines derived from lymphocytic leukemia, Ehrlich carcinoma, hepatoma, and lymphoid leukemia (41)(42)(43)(44). It should be noted that nanomolar concentrations of brusatol were used in our study, whereas micromolar concentrations were used in the in vitro studies conducted by the aforementioned groups (42,43). In our study, brusatol (20-80 nM) specifically inhibited the protein level of Nrf2 in a dose-dependent manner (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…This group reported that bruceantin, brucein D, brucein E, bruceoside A, and brusatol inhibited DNA, RNA, and protein synthesis as well as oxidative phosphorylation in p-388 lymphocytic leukemia cells (39,40). Inhibition of overall protein synthesis was observed in rabbit reticulocytes, as well as in cell lines derived from lymphocytic leukemia, Ehrlich carcinoma, hepatoma, and lymphoid leukemia (41)(42)(43)(44). It should be noted that nanomolar concentrations of brusatol were used in our study, whereas micromolar concentrations were used in the in vitro studies conducted by the aforementioned groups (42,43).…”

Section: Discussionmentioning

confidence: 99%

Brusatol enhances the efficacy of chemotherapy by inhibiting the Nrf2-mediated defense mechanism

Ren

Villeneuve

Jiang

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

557

506

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The major obstacle in cancer treatment is the resistance of cancer cells to therapies. Nrf2 is a transcription factor that regulates a cellular defense response and is ubiquitously expressed at low basal levels in normal tissues due to Keap1-dependent ubiquitination and proteasomal degradation. Recently, Nrf2 has emerged as an important contributor to chemoresistance. High constitutive expression of Nrf2 was found in many types of cancers, creating an environment conducive for cancer cell survival. Here, we report the identification of brusatol as a unique inhibitor of the Nrf2 pathway that sensitizes a broad spectrum of cancer cells and A549 xenografts to cisplatin and other chemotherapeutic drugs. Mechanistically, brusatol selectively reduces the protein level of Nrf2 through enhanced ubiquitination and degradation of Nrf2. Consequently, expression of Nrf2-downstream genes is reduced and the Nrf2-dependent protective response is suppressed. In A549 xenografts, brusatol and cisplatin cotreatment induced apoptosis, reduced cell proliferation, and inhibited tumor growth more substantially when compared with cisplatin treatment alone. Additionally, A549-K xenografts, in which Nrf2 is expressed at very low levels due to ectopic expression of Keap1, do not respond to brusatol treatment, demonstrating that brusatol-mediated sensitization to cisplatin is Nrf2 dependent. Moreover, a decrease in drug detoxification and impairment in drug removal may be the primary mechanisms by which brusatol enhances the efficacy of chemotherapeutic drugs. Taken together, these results clearly demonstrate the effectiveness of using brusatol to combat chemoresistance and suggest that brusatol can be developed into an adjuvant chemotherapeutic drug.

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“…Baseado nos nossos resultados, em que a IsoB reduz a produção/liberação de citocinas pró-inflamatórias, mas não inibe a transcrição do mesmo gene (TNF RNAm), e em outros estudos que mostram potencial efeito dos quassinóides na inibição da síntese proteica eucariótica (FUKAMIYA et al, 2005;LIOU et al, 1982), foi testada a hipótese de que a IsoB atua por este mecanismo. Células HEK transfectadas com uma plasmídeo para expressão constitutiva de luciferase, por conter uma região promotora PKG que recruta inespecificamente fatores de transcrição, incubadas com…”

Section: Discussionunclassified

“…Muitos quassinóides foram estudados, principalmente, na década de 70 e 80, por demonstrarem atividade terapêutica no tratamento de leucemia, e que resultou na aprovação do uso clínico da bruceatina para quimioterapia deste câncer (EIGEBALY et al, 1979;POLONSKY;MERIENNE;SEVENET, 1981;LIOU et al, 1982;ALMEIDA, 2007…”

Section: Figura 4 -Estrutura Química Da Isobruceina Bunclassified

Mecanismos envolvidos nos efeitos anti-inflamatório e anti-hipernociceptivo da Isobruceina B

Silva¹

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Objetivo: Avaliar o efeito da Isobruceina B como anti-hipernociceptivo e antiinflamatório, e identificar o mecanismo molecular envolvido. Métodos: Inicialmente, o efeito da IsoB sobre a hipernocicepção inflamatória do tecido plantar foi avaliada por von Frey eletrônico. Os tecidos plantares foram removidos para quantificação indireta da infiltração de neutrófilos por mieloperoxidase (MPO). Seu efeito nocicepção térmica foi avaliado em camundongos pelo teste da placa quente a 55º C. Camundongos pré-tratados com IsoB (s.c.) e tratados com carragenina (i.pl), tiveram seus tecidos plantares removidos para quantificação da liberação de citocinas TNF, IL-1β e KC/CXCL1. Para os estudos in vitro, foram utilizados macrófagos extraídos da cavidade peritoneal de camundongos naïve, pré-tratados com 3% de tioglicolato (i.p.) ou de linhagem RAW 264.7. Macrófagos peritoneais foram incubadas com lipopolissacarídeo (LPS) na presença ou ausência de diferentes concentrações de IsoB, para a quantificação da liberação das citocinas TNF, IL-1β, KC/CXCL1 e IL-10 no sobrenadante pelo método de ELISA. A produção in vitro de óxido nítrico (NO) foi indiretamente determinada pelo método de Griess. Para verificar o efeito da IsoB sobre a via do NF-kB, foram utilizados macrófagos RAW 264.7 estavelmente transfectados com um gene para expressão de luciferase dependente de NF-kB (RAW-Luc), incubados com diferentes estímulos (LPS, peptideoglicano, TNF e acetato de forbol miristato [PMA]) na presença ou ausência de IsoB. A atividade da luciferase foi detectada por meio do Luminômetro. A citotoxicidade da IsoB foi avaliada por MTT, lactato desidrogenase e ensaios de citotoxicidade pelo azul de tripan em macrófagos, in vitro. O efeito da IsoB em macrófagos, sobre a degradação o IkB-α e translocação do NF-kB (pela marcação da subunidade p65) para o núcleo e foram examinados por Western Blot de extratos nucleares e citoplasmáticos, e por Microscopia Confocal. O efeito da IsoB sobre a transcrição do RNAm do gene TNF induzido por LPS, foi avaliada por RT-PCR. Células HEK 293 transfectadas foram utilizadas para verificar o efeito da IsoB sobre a síntese de proteínas. Resultados: A IsoB inibe a hipernocicepção inflamatória, mas não tem efeito sobre a nocicepção térmica. IsoB inibiu a produção/libertação das citocinas IL-1β e KC/CXCL1 in vivo, e de TNF, IL-1β, KC/CXCL1, IL-10 e NO in vitro de forma concentração-dependente. O composto analisado também inibiu a atividade da luciferase, NF-kB dependente, por todos os estímulos testado, de forma concentração-dependente. Não foi detectada atividade citotóxica em nenhuma das concentrações testadas. A IsoB não inibe a degradação do IkB-α ou a translocação de NF-kB para o núcleo. Inesperadamente, o composto natural testado também não reduziu a transcrição de RNAm do gene TNF, porém inibiu a síntese de luciferase inespecificamente induzida. Conclusão: A redução da liberação de citocinas próinflamatórias, tais como TNF, IL-1β e KC/CXCL1 e da migração de neutrófilos são mecanismos envolvidos nos efeitos anti-inflamat...

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Antitumor Agents XLVIII: Structure-Activity Relationships of Quassinoids as In Vitro Protein Synthesis Inhibitors of P-388 Lymphocytic Leukemia Tumor Cell Metabolism

Cited by 22 publications

References 16 publications

Application of Mass Spectrometry Profiling to Establish Brusatol as an Inhibitor of Global Protein Synthesis

Application of Mass Spectrometry Profiling to Establish Brusatol as an Inhibitor of Global Protein Synthesis

Brusatol enhances the efficacy of chemotherapy by inhibiting the Nrf2-mediated defense mechanism

Mecanismos envolvidos nos efeitos anti-inflamatório e anti-hipernociceptivo da Isobruceina B

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