Em formação

12.6: Receptor de tirosina quinases - Biologia

12.6: Receptor de tirosina quinases - Biologia


We are searching data for your request:

Forums and discussions:
Manuals and reference books:
Data from registers:
Wait the end of the search in all databases.
Upon completion, a link will appear to access the found materials.

Receptor de tirosina quinases (RTK)

Cascata de eventos: Um receptor transmembrana COM ATIVIDADE ENZIMÁTICA DEPENDENTE DE HORMÔNIO (tirosina quinase) se liga a um sinal químico extracelular, causando uma mudança conformacional no receptor que se propaga através da membrana. Exemplos são o receptor de insulina e o receptor do fator de crescimento epidérmico.

O receptor Tyr quinases autofosforila-se, em um processo necessário para sua atividade. Quando o receptor é autofosforilado, outras proteínas podem se ligar ao domínio citoplasmático do receptor Tyr quinase, onde são fosforiladas. Os substratos alvo fosforilados pelo receptor Tyr quinase são proteínas com um domínio comum de 100 aminoácidos denominado SH para homologia src, com base na homologia estrutural com outra proteína citoplasmática, Src. Src é uma Tyr quinase intracelular ativada quando se liga através de 2 domínios SH ao receptor autofosforilado Tyr quinase. Especificamente, foi demonstrado que o domínio SH2 se liga a peptídeos fosforilados. Estes domínios direcionam proteínas para o receptor autofosforilado Tyr quinase.

Figura: Proteína Quinases Dependentes de Receptor / Ligante

Muitas proteínas envolvidas na transdução de sinal têm domínios SH2. Algumas dessas proteínas também possuem domínios catalíticos com atividade quinase. Outros têm fosfatase, fator de transcrição. ou domínios de andaimes.

Src Kinase Domínio SH2 Domínio SH2


As tirosina quinases da família Src são necessárias para a transdução de sinal mediada por fator de crescimento derivado de plaquetas em células NIH 3T3.

Três membros da família Src de proteínas tirosina quinases Src, Fyn e Yes se associam ao receptor do fator de crescimento derivado de plaquetas ativado (PDGF) in vivo. Esta interação requer o domínio de homologia 2 (SH2) Src do membro da família Src e causa a ativação da atividade intrínseca das quinases da família Src. Nós microinjetamos células com DNA codificando formas cataliticamente inativas das proteínas Src e Fyn e examinamos seus efeitos na sinalização mediada por PDGF in vivo. Src e Fyn inativas por cinase inibiram a entrada de células estimulada por PDGF na fase S, enquanto as formas ativas por cinase das proteínas não tiveram efeitos inibitórios. Um domínio SH2 intacto foi necessário para a inibição. Além disso, quando a Fyn inativa por quinase foi injetada em quadrinhos com um plasmídeo que expressa Ras ativada, as células puderam entrar na fase S, indicando que a expressão de Fyn inativa por quinase não danificou a viabilidade celular. A injeção de um anticorpo específico para Src, Fyn e Yes também reduziu a transdução de sinal através do receptor de PDGF, mas apenas quando injetado dentro de 8 horas de estimulação de PDGF. Juntos, esses resultados indicam que os membros da família Src ubiquamente expressos são necessários para a sinalização mitogênica induzida por PDGF.


Siegel, R. L., Miller, K. D., & amp Jemal, A. (2016). Estatísticas do câncer, 2016. CA: a Cancer Journal for Clinicians, 66(1), 7–30.

Perou, C. M., Sorlie, T., Eisen, M. B., van de Rijn, M., Jeffrey, S. S., Rees, C. A., et al. (2000). Retratos moleculares de tumores de mama humanos. Nature, 406(6797), 747–752.

Sorlie, T., Perou, C.M., Tibshirani, R., Aas, T., Geisler, S., Johnsen, H., et al. (2001). Os padrões de expressão gênica de carcinomas de mama distinguem subclasses de tumor com implicações clínicas. Anais da Academia Nacional de Ciências dos Estados Unidos da América, 98(19), 10869–10874.

Sorlie, T., Tibshirani, R., Parker, J., Hastie, T., Marron, J. S., Nobel, A., et al. (2003). Observação repetida de subtipos de tumor de mama em conjuntos de dados de expressão gênica independentes. Anais da Academia Nacional de Ciências dos Estados Unidos da América, 100(14), 8418–8423.

Voduc, K. D., Cheang, M. C., Tyldesley, S., Gelmon, K., Nielsen, T. O., & amp Kennecke, H. (2010). Subtipos de câncer de mama e o risco de recidiva local e regional. Journal of Clinical Oncology, 28(10), 1684–1691.

Cancer Genome Atlas N (2012). Retratos moleculares abrangentes de tumores de mama humanos. Natureza, 490(7418), 61–70.

American Cancer Society (2016). Fatos e números sobre o câncer. Atlanta: American Cancer Society., 2016.

Rivera, E., & amp Gomez, H. (2010). Resistência à quimioterapia no câncer de mama metastático: a evolução do papel da ixabepilona. Breast Cancer Research, 12(Suplemento 2), S2.

Yarden, Y. & amp Shilo, B. Z. (2007). SnapShot: via de sinalização de EGFR. Cell, 131(5), 1018.

Lemmon, M. A., & amp Schlessinger, J. (2010). Sinalização celular por tirosina quinases receptoras. Cell, 141(7), 1117–1134.

Chen, M. K., & amp Hung, M. C. (2015). Clivagem proteolítica, tráfego e funções das tirosina quinases receptoras nucleares. The FEBS Journal, 282(19), 3693–3721.

Schlessinger, J. (1988). Transdução de sinal por oligomerização de receptor alostérico. Tendências em Ciências Bioquímicas, 13(11), 443–447.

Gordus, A., Krall, J.A., Beyer, E.M., Kaushansky, A., Wolf-Yadlin, A., Sevecka, M., et al. (2009). Combinações lineares de afinidades de acoplamento explicam as diferenças quantitativas na sinalização RTK. Biologia de Sistemas Moleculares, 5, 235.

Casaletto, J. B., & amp McClatchey, A. I. (2012). Regulação espacial de tirosina quinases receptoras em desenvolvimento e câncer. Nature Reviews. Câncer, 12(6), 387–400.

Schlessinger, J. 2014. Receptor tirosina quinases: legado das primeiras duas décadas. Perspectivas de Cold Spring Harbor em biologia, 6(3).

Waterman, H., & amp Yarden, Y. (2001). Mecanismos moleculares subjacentes à endocitose e seleção de tirosina quinases do receptor ErbB. FEBS Letters, 490(3), 142–152.

von Zastrow, M., & amp Sorkin, A. (2007). Sinalização na via endocítica. Opinião Atual em Biologia Celular, 19(4), 436–445.

Wang, S. C., & amp Hung, M. C. (2009). Translocação nuclear dos receptores de tirosina quinase de membrana da família do fator de crescimento epidérmico. Pesquisa Clínica do Câncer, 15(21), 6484–6489.

Lee, H. H., Wang, Y. N., & amp Hung, M. C. (2015). Modo de sinalização não canônico da família de receptores do fator de crescimento epidérmico. American Journal of Cancer Research, 5(10), 2944–2958.

The, Y. Y. (2001). Família EGFR e seus ligantes no câncer humano. Mecanismos de sinalização e oportunidades terapêuticas. European Journal of Cancer, 37(Suplemento 4), S3 – S8.

Hynes, N. E., & amp MacDonald, G. (2009). Receptores ErbB e vias de sinalização no câncer. Opinião Atual em Biologia Celular, 21(2), 177–184.

Neal, J. W., & amp Sledge, G. W. (2014). Década na terapia direcionada à revisão: sucessos, toxicidades e desafios em tumores sólidos. Nature reviews Clinical oncology., 11(11), 627–628.

Remon, J., Moran, T., Majem, M., Reguart, N., Dalmau, E., Marquez-Medina, D., et al. (2014). Resistência adquirida aos inibidores do receptor tirosina quinase do fator de crescimento epidérmico no câncer de pulmão de células não pequenas mutante de EGFR: uma nova era começa. Revisões do tratamento do câncer., 40(1), 93–101.

Yarden, Y. & amp Sliwkowski, M. X. (2001). Desembaraçando a rede de sinalização ErbB. Nature Reviews. Biologia Molecular Celular, 2(2), 127–137.

Prenzel, N., Fischer, O. M., Streit, S., Hart, S., & amp Ullrich, A. (2001). Família de receptores do fator de crescimento epidérmico como elemento central para a transdução e diversificação do sinal celular. Câncer Relacionado ao Endócrino, 8(1), 11–31.

Citri, A., & amp Yarden, Y. (2006). Sinalização EGF-ERBB: em direção ao nível do sistema. Nature Reviews. Biologia Molecular Celular, 7(7), 505–516.

Schneider, M. R. & amp Wolf, E. (2009). Os ligantes do receptor do fator de crescimento epidérmico em um relance. Journal of Cellular Physiology, 218(3), 460–466.

Chaffer, C. L., & amp Weinberg, R. A. (2011). Uma perspectiva sobre a metástase de células cancerosas. Ciência, 331(6024), 1559–1564.

Avraham, R., & amp Yarden, Y. (2011). Regulação por feedback da sinalização EGFR: tomada de decisão por loops antecipados e atrasados. Nature Reviews. Biologia Molecular Celular, 12(2), 104–117.

Baselga, J., & amp Swain, S. M. (2009). Novos alvos anticâncer: revisitando o ERBB2 e descobrindo o ERBB3. Nature Reviews. Câncer, 9(7), 463–475.

Tebbutt, N., Pedersen, M. W., & amp Johns, T. G. (2013). Visando a família ERBB no câncer: terapia de casais. Nature Reviews. Câncer, 13(9), 663–673.

Cantley, L. C. (2002). A via da fosfoinositida 3-quinase. Ciência, 296(5573), 1655–1657.

Scaltriti, M., & amp Baselga, J. (2006). A via do receptor do fator de crescimento epidérmico: um modelo para terapia direcionada. Pesquisa Clínica do Câncer, 12(18), 5268–5272.

Quesnelle, K. M., Boehm, A. L., & amp Grandis, J. R. (2007). Sinalização de EGFR mediada por STAT no câncer. Journal of Cellular Biochemistry, 102(2), 311–319.

Lurje, G., & amp Lenz, H. J. (2009). Sinalização de EGFR e descoberta de drogas. Oncologia, 77(6), 400–410.

Fischer, O. M., Hart, S., Gschwind, A., & amp Ullrich, A. (2003). Transativação do sinal EGFR em células cancerosas. Biochemical Society Transactions, 31(Pt 6), 1203-1208.

Liu, D., Aguirre Ghiso, J., Estrada, Y., & amp Ossowski, L. (2002). EGFR é um transdutor do sinal iniciado pelo receptor de uroquinase que é necessário para na Vivo crescimento de um carcinoma humano. Célula cancerosa, 1(5), 445–457.

Sainsbury, J. R., Farndon, J. R., Needham, G. K., Malcolm, A. J., & amp Harris, A. L. (1987). Status do receptor do fator de crescimento epidérmico como preditor de recorrência precoce e morte por câncer de mama. Lanceta, 1(8547), 1398–1402.

Tsutsui, S., Ohno, S., Murakami, S., Hachitanda, Y., & amp Oda, S. (2002). Valor prognóstico do receptor do fator de crescimento epidérmico (EGFR) e sua relação com o status do receptor de estrogênio em 1.029 pacientes com câncer de mama. Pesquisa e tratamento do câncer de mama, 71(1), 67–75.

Witton, C. J., Reeves, J. R., Going, J. J., Cooke, T. G., & amp Bartlett, J. M. (2003). Expressão da família HER1-4 de receptor de tirosina quinases no câncer de mama. The Journal of Pathology, 200(3), 290–297.

Rakha, E. A., El-Sayed, M. E., Green, A. R., Lee, A. H., Robertson, J. F., & amp Ellis, I. O. (2007). Marcadores prognósticos no câncer de mama triplo-negativo. Câncer, 109(1), 25–32.

Lehmann, B. D., Bauer, J. A., Chen, X., Sanders, M. E., Chakravarthy, A. B., Shyr, Y., et al. (2011). Identificação de subtipos de câncer de mama triplo-negativo humano e modelos pré-clínicos para seleção de terapias direcionadas. The Journal of Clinical Investigation, 121(7), 2750–2767.

Masuda, H., Zhang, D., Bartholomeusz, C., Doihara, H., Hortobagyi, G. N., & amp Ueno, N. T. (2012). Papel do receptor do fator de crescimento epidérmico no câncer de mama. Pesquisa e tratamento do câncer de mama, 136(2), 331–345.

Frederick, L., Wang, X. Y., Eley, G., & amp James, C. D. (2000). Diversidade e frequência de mutações no receptor do fator de crescimento epidérmico em glioblastomas humanos. Cancer Research, 60(5), 1383–1387.

Ooi, A., Takehana, T., Li, X., Suzuki, S., Kunitomo, K., Iino, H., et al. (2004). Superexpressão de proteínas e amplificação de genes de HER-2 e EGFR em cânceres colorretais: uma análise imunohistoquímica e fluorescente no local estudo de hibridização. Patologia Moderna, 17(8), 895–904.

Bhargava, R., Gerald, W. L., Li, A. R., Pan, Q., Lal, P., Ladanyi, M., et al. (2005). Amplificação do gene EGFR no câncer de mama: correlação com o mRNA do receptor do fator de crescimento epidérmico e a expressão da proteína e o status de HER-2 e ausência de mutações ativadoras de EGFR. Patologia Moderna, 18(8), 1027–1033.

Hanawa, M., Suzuki, S., Dobashi, Y., Yamane, T., Kono, K., Enomoto, N., et al. (2006). Superexpressão da proteína EGFR e amplificação gênica em carcinomas de células escamosas do esôfago. International Journal of Cancer, 118(5), 1173–1180.

Hirsch, F. R., Varella-Garcia, M., & amp Cappuzzo, F. (2009). Valor preditivo da superexpressão de EGFR e HER2 no câncer avançado de pulmão de células não pequenas. Oncogene, 28(Suplemento 1), S32 – S37.

Reis-Filho, J. S., Pinheiro, C., Lambros, M. B., Milanezi, F., Carvalho, S., Savage, K., et al. (2006). Amplificação de EGFR e falta de mutações ativadoras em carcinomas de mama metaplásicos. The Journal of Pathology, 209(4), 445–453.

Burga, L. N., Hu, H., Juvekar, A., Tung, N. M., Troyan, S. L., Hofstatter, E. W., et al. (2011). A perda de BRCA1 leva a um aumento na expressão do receptor do fator de crescimento epidérmico em células epiteliais mamárias e a inibição do receptor do fator de crescimento epidérmico previne cânceres negativos para o receptor de estrogênio em camundongos mutantes BRCA1. Pesquisa de câncer de mama: BCR., 13(2), R30.

Zhang, J., Antonyak, M. A., Singh, G., & amp Cerione, R. A. (2013). Um mecanismo para a regulação positiva dos níveis do receptor de EGF em glioblastomas. Relatórios de células, 3(6), 2008–2020.

Verma, A. & amp Mehta, K. (2007). Quimiorresistência tecidual mediada por transglutaminase em células cancerosas. Atualizações de resistência a drogas, 10(4–5), 144–151.

Huang, L., Xu, A. M., & amp Liu, W. (2015). Transglutaminase 2 no câncer. American Journal of Cancer Research, 5(9), 2756–2776.

Lynch, T. J., Bell, D. W., Sordella, R., Gurubhagavatula, S., Okimoto, R. A., Brannigan, B. W., et al. (2004). Ativação de mutações no receptor do fator de crescimento epidérmico subjacente à responsividade do câncer de pulmão de células não pequenas ao gefitinibe. The New England Journal of Medicine, 350(21), 2129–2139.

Paez, J. G., Janne, P. A., Lee, J. C., Tracy, S., Greulich, H., Gabriel, S., et al. (2004). Mutações de EGFR em câncer de pulmão: correlação com a resposta clínica à terapia com gefitinibe. Ciência, 304(5676), 1497–1500.

Pao, W., Miller, V., Zakowski, M., Doherty, J., Politi, K., Sarkaria, I., et al. (2004). As mutações no gene do receptor de EGF são comuns em cânceres de pulmão de “nunca fumantes” e estão associadas à sensibilidade de tumores a gefitinibe e erlotinibe. Anais da Academia Nacional de Ciências dos Estados Unidos da América, 101(36), 13306–13311.

Teng, Y. H., Tan, W. J., Thike, A. A., Cheok, P. Y., Tse, G. M., Wong, N. S., et al. (2011). Mutações no gene do receptor do fator de crescimento epidérmico (EGFR) no câncer de mama triplo negativo: possíveis implicações para a terapia direcionada. Breast Cancer Research, 13(2), R35.

Nakai, K., Hung, M.C., Yamaguchi, H. 2016. Uma perspectiva sobre terapias anti-EGFR direcionadas ao câncer de mama triplo-negativo. American Journal of Cancer Research, na imprensa.

Pedersen, M. W., Meltorn, M., Damstrup, L., & amp Poulsen, H. S. (2001). A mutação do receptor do fator de crescimento epidérmico do tipo III. Significado biológico e alvo potencial para terapia anticâncer. Annals of Oncology, 12(6), 745–760.

Gan, H. K., Cvrljevic, A. N., & amp Johns, T. G. (2013). A variante III do receptor do fator de crescimento epidérmico (EGFRvIII): onde as coisas selvagens são alteradas. The FEBS Journal, 280(21), 5350–5370.

Del Vecchio, C. A., Jensen, K. C., Nitta, R. T., Shain, A. H., Giacomini, C. P., & amp Wong, A. J. (2012). A variante III do receptor do fator de crescimento epidérmico contribui para os fenótipos de células-tronco cancerígenas no carcinoma de mama invasivo. Cancer Research, 72(10), 2657–2671.

Reya, T., & amp Clevers, H. (2005). Sinalização Wnt em células-tronco e câncer. Nature, 434(7035), 843–850.

Burgess, A. W. (2008). Família EGFR: estrutura de sinalização fisiológica e alvos terapêuticos. Fatores de crescimento, 26(5), 263–274.

Marti, U., Burwen, S.J., Wells, A., Barker, M.E., Huling, S., Feren, A.M., et al. (1991). Localização do receptor do fator de crescimento epidérmico nos núcleos dos hepatócitos. Hepatologia, 13(1), 15–20.

Han, W., & amp Lo, H. W. (2012). Paisagem da rede de sinalização de EGFR em cânceres humanos: biologia e resposta terapêutica em relação às localizações subcelulares do receptor. Cartas de Câncer, 318(2), 124–134.

Lo, H. W., Xia, W., Wei, Y., Ali-Seyed, M., Huang, S. F., & amp Hung, M. C. (2005). Novo valor prognóstico do receptor do fator de crescimento epidérmico nuclear no câncer de mama. Cancer Research, 65(1), 338–348.

Psyrri, A., Yu, Z., Weinberger, P.M., Sasaki, C., Haffty, B., Camp, R., et al. (2005). Determinação quantitativa da expressão do receptor do fator de crescimento epidérmico nuclear e citoplasmático no câncer de células escamosas da orofaringe por meio de análise quantitativa automatizada. Pesquisa Clínica do Câncer, 11(16), 5856–5862.

Hoshino, M., Fukui, H., Ono, Y., Sekikawa, A., Ichikawa, K., Tomita, S., et al. (2007). A expressão nuclear de EGFR fosforilado está associada a um mau prognóstico de pacientes com carcinoma epidermóide de esôfago. Patobiologia, 74(1), 15–21.

Psyrri, A., Egleston, B., Weinberger, P., Yu, Z., Kowalski, D., Sasaki, C., et al. (2008). Correlaciona e determinantes do conteúdo do receptor do fator de crescimento epidérmico nuclear em um microarray de tecido de câncer orofaríngeo. Cancer Epidemiology, Biomarkers & amp Prevention, 17(6), 1486–1492.

Xia, W., Wei, Y., Du, Y., Liu, J., Chang, B., Yu, Y. L., et al. (2009). A expressão nuclear do receptor do fator de crescimento epidérmico é um novo valor prognóstico em pacientes com câncer de ovário. Carcinogênese molecular, 48(7), 610–617.

Hadzisejdic, I., Mustac, E., Jonjic, N., Petkovic, M., & amp Grahovac, B. (2010). EGFR nuclear em câncer de mama invasivo ductal: correlação com ciclina-D1 e prognóstico. Patologia Moderna, 23(3), 392–403.

Dittmann, K., Mayer, C., Fehrenbacher, B., Schaller, M., Kehlbach, R., & amp Rodemann, H. P. (2010). O transporte nuclear de EGFR induzido por radiação ionizante é regulado por fosforilação no resíduo Thr654. FEBS Letters, 584(18), 3878–3884.

Huo, L., Wang, Y. N., Xia, W., Hsu, S.C., Lai, C.C., Li, L.Y., et al. (2010). A RNA helicase a é um parceiro de ligação ao DNA para a ativação transcricional mediada por EGFR no núcleo. Anais da Academia Nacional de Ciências dos Estados Unidos da América, 107(37), 16125–16130.

Wheeler, D. L., Dunn, E. F., & amp Harari, P. M. (2010). Compreender a resistência aos inibidores de EGFR - impacto nas estratégias de tratamento futuras. Nature Reviews. Oncologia Clínica, 7(9), 493–507.

Chen, Y. J., Huang, W. C., Wei, Y. L., Hsu, S. C., Yuan, P., Lin, H. Y., et al. (2011). A expressão elevada de BCRP / ABCG2 confere resistência adquirida a gefitinib em células que expressam EGFR de tipo selvagem. PloS One, 6(6), e21428.

Huang, W. C., Chen, Y. J., Li, L. Y., Wei, Y. L., Hsu, S. C., Tsai, S. L., et al. (2011). A translocação nuclear do receptor do fator de crescimento epidérmico por fosforilação dependente de Akt aumenta a expressão da proteína resistente ao câncer de mama em células resistentes ao gefitinibe. The Journal of Biological Chemistry, 286(23), 20558–20568.

Wang, Y. N., & amp Hung, M. C. (2012). Funções nucleares e mecanismos de tráfego subcelular da família de receptores do fator de crescimento epidérmico. Cell & amp Bioscience, 2(1), 13.

Carpenter, G., & amp Liao, H. J. (2013). Receptor de tirosina quinases no núcleo. Cold Spring Harbor Perspectives in Biology, 5(10), a008979.

Wang, Y., Hsu, J. L., & amp Hung, M. C. (2013). Funções nucleares e tráfego de tirosina quinases receptoras. Em G. Yarden YaT (Ed.), Tráfico de vesículas em câncer (pp. 159-176). Nova York: Springer.

Lin, S. Y., Makino, K., Xia, W., Matin, A., Wen, Y., Kwong, K. Y., et al. (2001). Localização nuclear do receptor EGF e seu novo papel potencial como fator de transcrição. Nature Cell Biology, 3(9), 802–808.

Lo, H. W., Hsu, S. C., Ali-Seyed, M., Gunduz, M., Xia, W., Wei, Y., et al. (2005). Interação nuclear de EGFR e STAT3 na ativação da via iNOS / NO. Célula cancerosa, 7(6), 575–589.

Hung, L. Y., Tseng, J. T., Lee, Y. C., Xia, W., Wang, Y. N., Wu, M. L., et al. (2008). O receptor do fator de crescimento epidérmico nuclear (EGFR) interage com o transdutor de sinal e ativador da transcrição 5 (STAT5) na ativação da expressão do gene da aurora-a. Pesquisa de ácidos nucléicos, 36(13), 4337–4351.

Dittmann, K., Mayer, C., Fehrenbacher, B., Schaller, M., Raju, U., Milas, L., et al. (2005). A importação nuclear do receptor do fator de crescimento epidérmico induzida pela radiação está ligada à ativação da proteína quinase dependente de DNA. The Journal of Biological Chemistry, 280(35), 31182–31189.

Wang, S.C., Nakajima, Y., Yu, Y.L., Xia, W., Chen, C.T., Yang, C.C., et al. (2006). A fosforilação da tirosina controla a função do PCNA através da estabilidade da proteína. Nature Cell Biology, 8(12), 1359–1368.

Harari, D. & amp Yarden, Y. (2000). Mecanismos moleculares subjacentes à ação do ErbB2 / HER2 no câncer de mama. Oncogene, 19(53), 6102–6114.

Lee-Hoeflich, S. T., Crocker, L., Yao, E., Pham, T., Munroe, X., Hoeflich, K. P., et al. (2008). Um papel central para HER3 no câncer de mama amplificado por HER2: implicações para a terapia direcionada. Cancer Research, 68(14), 5878–5887.

Garrett, J.T., Sutton, C.R., Kurupi, R., Bialucha, C.U., Ettenberg, S.A., Collins, S.D., et al. (2013). A combinação de anticorpo que inibe a dimerização de HER3 independente de ligante e um inibidor de p110alfa bloqueia potentemente a sinalização de PI3K e o crescimento de cânceres de mama HER2 +. Cancer Research, 73(19), 6013–6023.

Slamon, D. J., Clark, G. M., Wong, S. G., Levin, W. J., Ullrich, A., & amp McGuire, W. L. (1987). Câncer de mama humano: correlação de recidiva e sobrevida com a amplificação do oncogene HER-2 / neu. Ciência, 235(4785), 177–182.

Pegram, M., & amp Slamon, D. (2000). Justificativa biológica para HER2 / neu (c-erbB2) como um alvo para terapia com anticorpos monoclonais. Seminários em Oncologia, 27(5 Suplemento 9), 13-19.

Balko, J. M., Mayer, A. I., Levy, M., Arteaga, C. L. 2013. HER2 (ERBB2) in breast cancer, My Cancer Genome. https://www.mycancergenome.org/content/disease/breast-cancer/erbb2/ (atualizado em 10 de abril).

Bose, R., Kavuri, S.M., Searleman, A.C., Shen, W., Shen, D., Koboldt, D.C., et al. (2013). Ativando mutações HER2 no câncer de mama negativo para amplificação do gene HER2. Cancer Discovery, 3(2), 224–237.

Arteaga, C. L., & amp Engelman, J. A. (2014). Receptores ERBB: da descoberta do oncogene à ciência básica à terapêutica do câncer baseada em mecanismo. Célula cancerosa, 25(3), 282–303.

Carter, P., Presta, L., Gorman, C. M., Ridgway, J. B., Henner, D., Wong, W. L., et al. (1992). Humanização de um anticorpo anti-p185HER2 para terapia de câncer humano. Anais da Academia Nacional de Ciências dos Estados Unidos da América, 89(10), 4285–4289.

Agus, D. B., Akita, R. W., Fox, W. D., Lewis, G. D., Higgins, B., Pisacane, P. I., et al. (2002). O direcionamento da sinalização de ErbB2 ativada por ligante inibe o crescimento do tumor da mama e da próstata. Célula cancerosa, 2(2), 127–137.

Lewis Phillips, G. D., Li, G., Dugger, D. L., Crocker, L. M., Parsons, K. L., Mai, E., et al. (2008). Ter como alvo o câncer de mama HER2-positivo com trastuzumab-DM1, um conjugado anticorpo-droga citotóxica. Cancer Research, 68(22), 9280–9290.

Rabindran, S. K., Discafani, C. M., Rosfjord, E. C., Baxter, M., Floyd, M. B., Golas, J., et al. (2004). Atividade antitumoral de HKI-272, um inibidor irreversível e ativo por via oral da tirosina quinase HER-2. Cancer Research, 64(11), 3958–3965.

Konecny, G. E., Pegram, M. D., Venkatesan, N., Finn, R., Yang, G., Rahmeh, M., et al. (2006). Atividade do inibidor dual da quinase lapatinibe (GW572016) contra células de câncer de mama com superexpressão de HER-2 e tratadas com trastuzumabe. Cancer Research, 66(3), 1630–1639.

Hudis, C. A. (2007). Trastuzumabe - mecanismo de ação e uso na prática clínica. The New England Journal of Medicine, 357(1), 39–51.

Vu, T., & amp Claret, F. X. (2012). Trastuzumab: mecanismos atualizados de ação e resistência no câncer de mama. Fronteiras em Oncologia, 2, 62.

Nagata, Y., Lan, K. H., Zhou, X., Tan, M., Esteva, F. J., Sahin, A. A., et al. (2004). A ativação do PTEN contribui para a inibição do tumor pelo trastuzumabe, e a perda do PTEN prediz a resistência ao trastuzumabe nos pacientes. Célula cancerosa, 6(2), 117–127.

Berns, K., Horlings, H. M., Hennessy, B. T., Madiredjo, M., Hijmans, E. M., Beelen, K., et al. (2007). Uma abordagem genética funcional identifica a via PI3K como o principal determinante da resistência ao trastuzumabe no câncer de mama. Célula cancerosa, 12(4), 395–402.

Zhang, S., Huang, W. C., Li, P., Guo, H., Poh, S. B., Brady, S. W., et al. (2011). Combater a resistência ao trastuzumabe visando SRC, um nó comum a jusante de múltiplas vias de resistência. Nature Medicine, 17(4), 461–469.

Nahta, R., Yuan, L. X., Zhang, B., Kobayashi, R., & amp Esteva, F. J. (2005). A heterodimerização do receptor do fator de crescimento semelhante à insulina / receptor 2 do fator de crescimento epidérmico humano contribui para a resistência ao trastuzumabe das células de câncer de mama. Cancer Research, 65(23), 11118–11128.

Ritter, C. A., Perez-Torres, M., Rinehart, C., Guix, M., Dugger, T., Engelman, J. A., et al. (2007). Células de câncer de mama humano selecionadas para resistência ao trastuzumabe na Vivo superexpressam o receptor do fator de crescimento epidérmico e ligantes ErbB e permanecem dependentes da rede de receptores ErbB. Pesquisa Clínica do Câncer, 13(16), 4909–4919.

Shattuck, D. L., Miller, J. K., Carraway 3rd, K. L., & amp Sweeney, C. (2008). O receptor Met contribui para a resistência ao trastuzumabe de células de câncer de mama com superexpressão de Her2. Cancer Research, 68(5), 1471–1477.

Zhuang, G., Brantley-Sieders, D. M., Vaught, D., Yu, J., Xie, L., Wells, S., et al. (2010). A elevação do receptor tirosina quinase EphA2 medeia a resistência à terapia com trastuzumabe. Cancer Research, 70(1), 299–308.

Scaltriti, M., Rojo, F., Ocana, A., Anido, J., Guzman, M., Cortes, J., et al. (2007). Expressão de p95HER2, uma forma truncada do receptor HER2 e resposta a terapias anti-HER2 no câncer de mama. Journal of the National Cancer Institute, 99(8), 628–638.

Price-Schiavi, S. A., Jepson, S., Li, P., Arango, M., Rudland, P. S., Yee, L., et al. (2002). O Muc4 de rato (complexo de sialomucina) reduz a ligação de anticorpos anti-ErbB2 às superfícies das células tumorais, um mecanismo potencial para a resistência à herceptina. International Journal of Cancer, 99(6), 783–791.

Thirumurthi, U., Shen, J., Xia, W., LaBaff, A. M., Wei, Y., Li, C. W., et al. (2014). A degradação mediada por MDM2 de SIRT6 fosforilada por AKT1 promove a tumorigênese e a resistência ao trastuzumabe no câncer de mama. Sinalização científica, 7(336), ra71.

Wang, S.C., Lien, H.C., Xia, W., Chen, I.F., Lo, H.W., Wang, Z., et al. (2004). Ligação e transativação do promotor COX-2 pelo receptor nuclear da tirosina quinase ErbB-2. Célula cancerosa, 6(3), 251–261.

Xie, Y., & amp Hung, M. C. (1994). Localização nuclear da tirosina quinase p185neu e sua associação com a transativação transcricional. Biochemical and Biophysical Research Communications, 203(3), 1589–1598.

Beguelin, W., Diaz Flaque, M.C., Proietti, C.J., Cayrol, F., Rivas, M.A., Tkach, M., et al. (2010). O receptor de progesterona induz a translocação nuclear ErbB-2 para promover o crescimento do câncer de mama por meio de um novo efeito de transcrição: a função ErbB-2 como um coativador de Stat3. Biologia Molecular e Celular, 30(23), 5456–5472.

Tan, M., Jing, T., Lan, K. H., Neal, C. L., Li, P., Lee, S., et al. (2002). A fosforilação na tirosina-15 de p34 (Cdc2) por ErbB2 inibe a ativação de p34 (Cdc2) e está envolvida na resistência à apoptose induzida por taxol. Molecular Cell, 9(5), 993–1004.

Schillaci, R., Guzman, P., Cayrol, F., Beguelin, W., Diaz Flaque, M.C., Proietti, C.J., et al. (2012). Relevância clínica da expressão nuclear de ErbB-2 / HER2 no câncer de mama. BMC Cancer, 12, 74.

Citri, A., Skaria, K. B., & amp Yarden, Y. (2003). Os surdos e mudos: a biologia de ErbB-2 e ErbB-3. Experimental Cell Research, 284(1), 54–65.

Carraway 3rd, K. L., Weber, J. L., Unger, M. J., Ledesma, J., Yu, N., Gassmann, M., et al. (1997). Neuregulina-2, um novo ligante das tirosina quinases receptoras ErbB3 / ErbB4. Nature, 387(6632), 512–516.

Shi, F., Telesco, S. E., Liu, Y., Radhakrishnan, R., & amp Lemmon, M. A. (2010). O domínio intracelular ErbB3 / HER3 é competente para ligar ATP e catalisar a autofosforilação. Anais da Academia Nacional de Ciências dos Estados Unidos da América, 107(17), 7692–7697.

Bieche, I., Onody, P., Tozlu, S., Driouch, K., Vidaud, M., & amp Lidereau, R. (2003). Valor prognóstico da expressão de mRNA da família ERBB em carcinomas de mama. International Journal of Cancer, 106(5), 758–765.

deFazio, A., Chiew, Y. E., Sini, R. L., Janes, P. W., & amp Sutherland, R. L. (2000). Expressão de receptores c-erbB, heregulina e receptor de estrogênio em linhagens de células mamárias humanas. International Journal of Cancer, 87(4), 487–498.

Sassen, A., Rochon, J., Wild, P., Hartmann, A., Hofstaedter, F., Schwarz, S., et al. (2008). Análise citogenética de HER1 / EGFR, HER2, HER3 e HER4 em 278 pacientes com câncer de mama. Breast Cancer Research, 10(1), R2.

Ocana, A., Vera-Badillo, F., Seruga, B., Templeton, A., Pandiella, A., & amp Amir, E. (2013). Superexpressão de HER3 e sobrevivência em tumores sólidos: uma meta-análise. Journal of the National Cancer Institute, 105(4), 266–273.

Chiu, C. G., Masoudi, H., Leung, S., Voduc, D. K., Gilks, B., Huntsman, D. G., et al. (2010). A superexpressão de HER-3 é um prognóstico de redução da sobrevida ao câncer de mama: um estudo com 4.046 pacientes. Annals of Surgery, 251(6), 1107–1116.

Morrison, M. M., Hutchinson, K., Williams, M. M., Stanford, J. C., Balko, J. M., Young, C., et al. (2013). A regulação negativa de ErbB3 aumenta a resposta do tumor luminal da mama aos antiestrogênios. The Journal of Clinical Investigation, 123(10), 4329–4343.

Jeong, E. G., Soung, Y. H., Lee, J. W., Lee, S. H., Nam, S. W., Lee, J. Y., et al. (2006). Mutações no domínio da quinase ERBB3 são raras em carcinomas de pulmão, mama e cólon. International Journal of Cancer, 119(12), 2986–2987.

Kan, Z., Jaiswal, B.S., Stinson, J., Janakiraman, V., Bhatt, D., Stern, H.M., et al. (2010). Diversos padrões de mutação somática e alterações de vias em cânceres humanos. Nature, 466(7308), 869–873.

Stephens, P. J., Tarpey, P. S., Davies, H., Van Loo, P., Greenman, C., Wedge, D. C., et al. (2012). O panorama dos genes do câncer e dos processos mutacionais no câncer de mama. Nature, 486(7403), 400–404.

Jaiswal, B. S., Kljavin, N. M., Stawiski, E. W., Chan, E., Parikh, C., Durinck, S., et al. (2013). Mutações oncogênicas ERBB3 em cânceres humanos. Célula cancerosa, 23(5), 603–617.

Zhang, N., Chang, Y., Rios, A., & amp An, Z. (2016). HER3 / ErbB3, um alvo terapêutico emergente do câncer. Acta Biochim Biophys Sin (Xangai)., 48(1), 39–48.

Offterdinger, M., Schofer, C., Weipoltshammer, K., & amp Grunt, T. W. (2002). C-erbB-3: uma proteína nuclear em células epiteliais mamárias. The Journal of Cell Biology, 157(6), 929–939.

Andrique, L., Fauvin, D., El Maassarani, M., Colasson, H., Vannier, B., & amp Seite, P. (2012). ErbB3 (80 kDa), uma variante nuclear do receptor ErbB3, liga-se ao promotor da ciclina D1 para ativar a proliferação celular, mas é controlado negativamente pelo p14ARF. Sinalização Celular, 24(5), 1074–1085.

Brand, T. M., Iida, M., Luthar, N., Wleklinski, M. J., Starr, M. M., & amp Wheeler, D. L. (2013). Mapeamento de domínios de transativação C-terminal do receptor tirosina quinase HER3 da família HER nuclear. PloS One, 8(8), e71518.

Koumakpayi, I.H., Diallo, J.S., Le Page, C., Lessard, L., Gleave, M., Begin, L.R., et al. (2006). Expressão e localização nuclear de ErbB3 no câncer de próstata. Pesquisa Clínica do Câncer, 12(9), 2730–2737.

Cheng, C. J., Ye, X. C., Vakar-Lopez, F., Kim, J., Tu, S. M., Chen, D. T., et al. (2007). O microambiente ósseo e o estado de andrógeno modulam a localização subcelular de ErbB3 em células de câncer de próstata. Molecular Cancer Research, 5(7), 675–684.

Harris, R. C., Chung, E., & amp Coffey, R. J. (2003). Ligantes do receptor de EGF. Experimental Cell Research, 284(1), 2–13.

Mill, C. P., Zordan, M. D., Rothenberg, S. M., Settleman, J., Leary, J. F., & amp Riese 2nd, D. J. (2011). ErbB2 é necessário para que os ligantes ErbB4 estimulem atividades oncogênicas em modelos de câncer de mama humano. Genes e Câncer, 2(8), 792–804.

Naresh, A., Long, W., Vidal, G. A., Wimley, W. C., Marrero, L., Sartor, C. I., et al. (2006). O domínio intracelular ERBB4 / HER4 4ICD é uma proteína apenas BH3 que promove a apoptose de células de câncer de mama. Cancer Research, 66(12), 6412–6420.

Uberall, I., Kolar, Z., Trojanec, R., Berkovcova, J., & amp Hajduch, M. (2008). O status e o papel dos receptores ErbB no câncer humano. Patologia Experimental e Molecular, 84(2), 79–89.

Tang, C. K., Concepcion, X. Z., Milan, M., Gong, X., Montgomery, E., & amp Lippman, M. E. (1999). A regulação negativa mediada por ribozima de ErbB-4 em células de câncer de mama positivas para receptor de estrogênio inibe a proliferação de ambos em vitro e na Vivo. Cancer Research, 59(20), 5315–5322.

Canfield, K., Li, J., Wilkins, O. M., Morrison, M. M., Ung, M., Wells, W., et al. (2015). O receptor tirosina quinase ERBB4 medeia a resistência adquirida aos inibidores ERBB2 em células de câncer de mama. Ciclo celular, 14(4), 648–655.

Kim, J. Y., Jung, H. H., Do, I. G., Bae, S., Lee, S. K., Kim, S. W., et al. (2016). Valor prognóstico da expressão de ERBB4 em pacientes com câncer de mama triplo negativo. BMC Cancer, 16, 138.

Gilbertson, R., Hernan, R., Pietsch, T., Pinto, L., Scotting, P., Allibone, R., et al. (2001). As novas variantes de splice por justamembrana ERBB4 são frequentemente expressas no meduloblastoma infantil. Genes, cromossomos e câncer, 31(3), 288–294.

Ding, L., Getz, G., Wheeler, D. A., Mardis, E. R., McLellan, M. D., Cibulskis, K., et al. (2008). As mutações somáticas afetam os caminhos-chave no adenocarcinoma pulmonar. Nature, 455(7216), 1069–1075.

Prickett, T. D., Agrawal, N. S., Wei, X., Yates, K. E., Lin, J. C., Wunderlich, J. R., et al. (2009). A análise do cinoma de tirosina no melanoma revela mutações recorrentes no ERBB4. Nature Genetics, 41(10), 1127–1132.

Kurppa, K. J., Denessiouk, K., Johnson, M. S., & amp Elenius, K. (2016). Ativando mutações ERBB4 em câncer de pulmão de células não pequenas. Oncogene, 35(10), 1283–1291.

Srinivasan, R., Gillett, C.E., Barnes, D. M., & amp Gullick, W. J. (2000). Nuclear expression of the c-erbB-4/HER-4 growth factor receptor in invasive breast cancers. Cancer Research, 60(6), 1483–1487.

Thompson, M., Lauderdale, S., Webster, M. J., Chong, V. Z., McClintock, B., Saunders, R., et al. (2007). Widespread expression of ErbB2, ErbB3 and ErbB4 in non-human primate brain. Brain Research, 1139, 95–109.

Icli, B., Bharti, A., Pentassuglia, L., Peng, X., & Sawyer, D. B. (2012). ErbB4 localization to cardiac myocyte nuclei, and its role in myocyte DNA damage response. Biochemical and Biophysical Research Communications, 418(1), 116–121.

Ni, C. Y., Murphy, M. P., Golde, T. E., & Carpenter, G. (2001). Gamma-secretase cleavage and nuclear localization of ErbB-4 receptor tyrosine kinase. Science, 294(5549), 2179–2181.

Williams, C. C., Allison, J. G., Vidal, G. A., Burow, M. E., Beckman, B. S., Marrero, L., et al. (2004). The ERBB4/HER4 receptor tyrosine kinase regulates gene expression by functioning as a STAT5A nuclear chaperone. The Journal of Cell Biology, 167(3), 469–478.

Linggi, B., & Carpenter, G. (2006). ErbB-4 s80 intracellular domain abrogates ETO2-dependent transcriptional repression. The Journal of Biological Chemistry, 281(35), 25373–25380.

Arasada, R. R., & Carpenter, G. (2005). Secretase-dependent tyrosine phosphorylation of Mdm2 by the ErbB-4 intracellular domain fragment. The Journal of Biological Chemistry, 280(35), 30783–30787.

Junttila, T. T., Sundvall, M., Lundin, M., Lundin, J., Tanner, M., Harkonen, P., et al. (2005). Cleavable ErbB4 isoform in estrogen receptor-regulated growth of breast cancer cells. Cancer Research, 65(4), 1384–1393.

Naresh, A., Thor, A. D., Edgerton, S. M., Torkko, K. C., Kumar, R., & Jones, F. E. (2008). The HER4/4ICD estrogen receptor coactivator and BH3-only protein is an effector of tamoxifen-induced apoptosis. Cancer Research, 68(15), 6387–6395.

Trusolino, L., Bertotti, A., & Comoglio, P. M. (2010). MET signalling: principles and functions in development, organ regeneration and cancer. Nature Reviews. Molecular Cell Biology, 11(12), 834–848.

Lai, A. Z., Abella, J. V., & Park, M. (2009). Crosstalk in met receptor oncogenesis. Trends in Cell Biology, 19(10), 542–551.

Ho-Yen, C. M., Jones, J. L., & Kermorgant, S. (2015). The clinical and functional significance of c-met in breast cancer: a review. Breast Cancer Research, 17, 52.

Engelman, J. A., Zejnullahu, K., Mitsudomi, T., Song, Y., Hyland, C., Park, J. O., et al. (2007). MET amplification leads to gefitinib resistance in lung cancer by activating ERBB3 signaling. Science, 316(5827), 1039–1043.

Ghoussoub, R. A., Dillon, D. A., D’Aquila, T., Rimm, E. B., Fearon, E. R., & Rimm, D. L. (1998). Expression of c-met is a strong independent prognostic factor in breast carcinoma. Cancer, 82(8), 1513–1520.

Lengyel, E., Prechtel, D., Resau, J. H., Gauger, K., Welk, A., Lindemann, K., et al. (2005). C-met overexpression in node-positive breast cancer identifies patients with poor clinical outcome independent of Her2/neu. International Journal of Cancer, 113(4), 678–682.

Lee, W. Y., Chen, H. H., Chow, N. H., Su, W. C., Lin, P. W., & Guo, H. R. (2005). Prognostic significance of co-expression of RON and MET receptors in node-negative breast cancer patients. Clinical Cancer Research, 11(6), 2222–2228.

Minuti, G., Cappuzzo, F., Duchnowska, R., Jassem, J., Fabi, A., O’Brien, T., et al. (2012). Increased MET and HGF gene copy numbers are associated with trastuzumab failure in HER2-positive metastatic breast cancer. British Journal of Cancer, 107(5), 793–799.

Kim, Y. J., Choi, J. S., Seo, J., Song, J. Y., Lee, S. E., Kwon, M. J., et al. (2014). MET is a potential target for use in combination therapy with EGFR inhibition in triple-negative/basal-like breast cancer. International Journal of Cancer, 134(10), 2424–2436.

Hsu, Y. H., Yao, J., Chan, L. C., Wu, T. J., Hsu, J. L., Fang, Y. F., et al. (2014). Definition of PKC-alpha, CDK6, and MET as therapeutic targets in triple-negative breast cancer. Cancer Research, 74(17), 4822–4835.

Du, Y., Yamaguchi, H., Wei, Y., Hsu, J. L., Wang, H. L., Hsu, Y. H., et al. (2016). Blocking c-met-mediated PARP1 phosphorylation enhances anti-tumor effects of PARP inhibitors. Nature Medicine, 22(2), 194–201.

Orton, T. C., Doughty, S. E., Kalinowski, A. E., Lord, P. G., & Wadsworth, P. F. (1996). Expression of growth factors and growth factor receptors in the liver of C57BL/10J mice following administration of phenobarbitone. Carcinogenesis, 17(5), 973–981.

Pozner-Moulis, S., Pappas, D. J., & Rimm, D. L. (2006). Met, the hepatocyte growth factor receptor, localizes to the nucleus in cells at low density. Cancer Research, 66(16), 7976–7982.

Matteucci, E., Bendinelli, P., & Desiderio, M. A. (2009). Nuclear localization of active HGF receptor met in aggressive MDA-MB231 breast carcinoma cells. Carcinogenesis, 30(6), 937–945.

Gomes, D. A., Rodrigues, M. A., Leite, M. F., Gomez, M. V., Varnai, P., Balla, T., et al. (2008). C-met must translocate to the nucleus to initiate calcium signals. The Journal of Biological Chemistry, 283(7), 4344–4351.

Pollak, M. (2008). Insulin and insulin-like growth factor signalling in neoplasia. Nature Reviews. Cancer, 8(12), 915–928.

Saldana, S. M., Lee, H. H., Lowery, F. J., Khotskaya, Y. B., Xia, W., Zhang, C., et al. (2013). Inhibition of type I insulin-like growth factor receptor signaling attenuates the development of breast cancer brain metastasis. PloS One, 8(9), e73406.

Farabaugh, S. M., Boone, D. N., & Lee, A. V. (2015). Role of IGF1R in breast cancer subtypes, Stemness, and lineage differentiation. Front Endocrinol (Lausanne), 6, 59.

Beckwith, H., & Yee, D. (2015). Minireview: were the IGF signaling inhibitors all bad? Molecular Endocrinology, 29(11), 1549–1557.

Aleksic, T., Chitnis, M. M., Perestenko, O. V., Gao, S., Thomas, P. H., Turner, G. D., et al. (2010). Type 1 insulin-like growth factor receptor translocates to the nucleus of human tumor cells. Cancer Research, 70(16), 6412–6419.

Warsito, D., Sjostrom, S., Andersson, S., Larsson, O., & Sehat, B. (2012). Nuclear IGF1R is a transcriptional co-activator of LEF1/TCF. EMBO Reports, 13(3), 244–250.

Warsito, D., Lin, Y., Gnirck, A. C., Sehat, B., Larsson, O. 2016. Nuclearly translocated insulin-like growth factor 1 receptor phosphorylates histone H3 at tyrosine 41 and induces SNAI2 expression via Brg1 chromatin remodeling protein. Oncotarget.

Bodzin, A. S., Wei, Z., Hurtt, R., Gu, T., & Doria, C. (2012). Gefitinib resistance in HCC mahlavu cells: upregulation of CD133 expression, activation of IGF-1R signaling pathway, and enhancement of IGF-1R nuclear translocation. Journal of Cellular Physiology, 227(7), 2947–2952.

Murray, P. B., Lax, I., Reshetnyak, A., Ligon, G. F., Lillquist, J. S., Natoli Jr., E. J., et al. (2015). Heparin is an activating ligand of the orphan receptor tyrosine kinase ALK. Science Signaling, 8(360), ra6.

Hallberg, B., & Palmer, R. H. (2013). Mechanistic insight into ALK receptor tyrosine kinase in human cancer biology. Nature Reviews. Cancer, 13(10), 685–700.

Lin, E., Li, L., Guan, Y., Soriano, R., Rivers, C. S., Mohan, S., et al. (2009). Exon array profiling detects EML4-ALK fusion in breast, colorectal, and non-small cell lung cancers. Molecular Cancer Research, 7(9), 1466–1476.

Barreca, A., Lasorsa, E., Riera, L., Machiorlatti, R., Piva, R., Ponzoni, M., et al. (2011). Anaplastic lymphoma kinase in human cancer. Journal of Molecular Endocrinology, 47(1), R11–R23.

Robertson, F. M., Petricoin Iii, E. F., Van Laere, S. J., Bertucci, F., Chu, K., Fernandez, S. V., et al. (2013). Presence of anaplastic lymphoma kinase in inflammatory breast cancer. Springerplus., 2, 497.

Siraj, A. K., Beg, S., Jehan, Z., Prabhakaran, S., Ahmed, M., RH, A., et al. (2015). ALK alteration is a frequent event in aggressive breast cancers. Breast Cancer Research, 17, 127.

Choi, Y. L., Soda, M., Yamashita, Y., Ueno, T., Takashima, J., Nakajima, T., et al. (2010). EML4-ALK mutations in lung cancer that confer resistance to ALK inhibitors. The New England Journal of Medicine, 363(18), 1734–1739.

Shaw, A. T., Friboulet, L., Leshchiner, I., Gainor, J. F., Bergqvist, S., Brooun, A., et al. (2016). Resensitization to crizotinib by the lorlatinib ALK resistance mutation L1198F. The New England Journal of Medicine, 374(1), 54–61.

Xiong, Q., Chan, J. L., Zong, C. S., & Wang, L. H. (1996). Two chimeric receptors of epidermal growth factor receptor and c-Ros that differ in their transmembrane domains have opposite effects on cell growth. Molecular and Cellular Biology, 16(4), 1509–1518.

Davies, K. D., & Doebele, R. C. (2013). Molecular pathways: ROS1 fusion proteins in cancer. Clinical Cancer Research, 19(15), 4040–4045.

Solomon, B. (2015). Validating ROS1 rearrangements as a therapeutic target in non-small-cell lung cancer. Journal of Clinical Oncology, 33(9), 972–974.

Shaw, A. T., Ou, S. H., Bang, Y. J., Camidge, D. R., Solomon, B. J., Salgia, R., et al. (2014). Crizotinib in ROS1-rearranged non-small-cell lung cancer. The New England Journal of Medicine, 371(21), 1963–1971.

Mazieres, J., Zalcman, G., Crino, L., Biondani, P., Barlesi, F., Filleron, T., et al. (2015). Crizotinib therapy for advanced lung adenocarcinoma and a ROS1 rearrangement: results from the EUROS1 cohort. Journal of Clinical Oncology, 33(9), 992–999.

Eom, M., Lkhagvadorj, S., Oh, S. S., Han, A., & Park, K. H. (2013). ROS1 expression in invasive ductal carcinoma of the breast related to proliferation activity. Yonsei Medical Journal, 54(3), 650–657.

Stransky, N., Cerami, E., Schalm, S., Kim, J. L., & Lengauer, C. (2014). The landscape of kinase fusions in cancer. Nature Communications, 5, 4846.

Halford, M. M., & Stacker, S. A. (2001). Revelations of the RYK receptor. BioEssays, 23(1), 34–45.

Cadigan, K. M., & Liu, Y. I. (2006). Wnt signaling: complexity at the surface. Journal of Cell Science, 119(Pt 3), 395–402.

Lyu, J., Yamamoto, V., & Lu, W. (2008). Cleavage of the Wnt receptor Ryk regulates neuronal differentiation during cortical neurogenesis. Developmental Cell, 15(5), 773–780.

Lyu, J., Wesselschmidt, R. L., & Lu, W. (2009). Cdc37 regulates Ryk signaling by stabilizing the cleaved Ryk intracellular domain. The Journal of Biological Chemistry, 284(19), 12940–12948.

Zhong, J., Kim, H. T., Lyu, J., Yoshikawa, K., Nakafuku, M., & Lu, W. (2011). The Wnt receptor Ryk controls specification of GABAergic neurons versus oligodendrocytes during telencephalon development. Development, 138(3), 409–419.

Katso, R. M., Manek, S., Ganjavi, H., Biddolph, S., Charnock, M. F., Bradburn, M., et al. (2000). Overexpression of H-Ryk in epithelial ovarian cancer: prognostic significance of receptor expression. Clinical Cancer Research, 6(8), 3271–3281.

Alvarez-Zavala, M., Riveros-Magana, A. R., Garcia-Castro, B., Barrera-Chairez, E., Rubio-Jurado, B., Garces-Ruiz, O. M., et al. (2016). WNT receptors profile expression in mature blood cells and immature leukemic cells: RYK emerges as a hallmark receptor of acute leukemia. European Journal of Haematology, 97(2), 155–165.

Carpenter, G., Lembach, K. J., Morrison, M. M., & Cohen, S. (1975). Characterization of the binding of 125-I-labeled epidermal growth factor to human fibroblasts. The Journal of Biological Chemistry, 250(11), 4297–4304.

Carpenter, G., King Jr., L., & Cohen, S. (1978). Epidermal growth factor stimulates phosphorylation in membrane preparations em vitro. Nature, 276(5686), 409–410.

Ullrich, A., Coussens, L., Hayflick, J. S., Dull, T. J., Gray, A., Tam, A. W., et al. (1984). Human epidermal growth factor receptor cDNA sequence and aberrant expression of the amplified gene in A431 epidermoid carcinoma cells. Nature, 309(5967), 418–425.

Liao, H. W., Hsu, J. M., Xia, W., Wang, H. L., Wang, Y. N., Chang, W. C., et al. (2015). PRMT1-mediated methylation of the EGF receptor regulates signaling and cetuximab response. The Journal of clinical investigation., 125(12), 4529–4543.

Mai, A., Cheng, D., Bedford, M. T., Valente, S., Nebbioso, A., Perrone, A., et al. (2008). Epigenetic multiple ligands: mixed histone/protein methyltransferase, acetyltransferase, and class III deacetylase (sirtuin) inhibitors. Journal of medicinal chemistry., 51(7), 2279–2290.

Sharma, P., & Allison, J. P. (2015). The future of immune checkpoint therapy. Science, 348(6230), 56–61.

Sharma, P., & Allison, J. P. (2015). Immune checkpoint targeting in cancer therapy: toward combination strategies with curative potential. Cell, 161(2), 205–214.

Li, C. W., Lim, S. O., Xia, W., Lee, H. H., Chan, L. C., Kuo, C. W., et al. (2016). Glycosylation and stabilization of programmed death ligand-1 suppresses T-cell activity. Nature Communications, 7, 12632.

Yang, X., Zhang, X., Mortenson, E. D., Radkevich-Brown, O., Wang, Y., & Fu, Y. X. (2013). Cetuximab-mediated tumor regression depends on innate and adaptive immune responses. Molecular therapy : the journal of the American Society of Gene Therapy., 21(1), 91–100.


Regulation of Epidermal Growth Factor Receptor Signaling by Endocytosis and Intracellular Trafficking

Ligand activation of the epidermal growth factor receptor (EGFR) leads to its rapid internalization and eventual delivery to lysosomes. This process is thought to be a mechanism to attenuate signaling, but signals could potentially be generated after endocytosis. To directly evaluate EGFR signaling during receptor trafficking, we developed a technique to rapidly and selectively isolate internalized EGFR and associated molecules with the use of reversibly biotinylated anti-EGFR antibodies. In addition, we developed antibodies specific to tyrosine-phosphorylated EGFR. With the use of a combination of fluorescence imaging and affinity precipitation approaches, we evaluated the state of EGFR activation and substrate association during trafficking in epithelial cells. We found that after internalization, EGFR remained active in the early endosomes. However, receptors were inactivated before degradation, apparently due to ligand removal from endosomes. Adapter molecules, such as Shc, were associated with EGFR both at the cell surface and within endosomes. Some molecules, such as Grb2, were primarily found associated with surface EGFR, whereas others, such as Eps8, were found only with intracellular receptors. During the inactivation phase, c-Cbl became EGFR associated, consistent with its postulated role in receptor attenuation. We conclude that the association of the EGFR with different proteins is compartment specific. In addition, ligand loss is the proximal cause of EGFR inactivation. Thus, regulated trafficking could potentially influence the pattern as well as the duration of signal transduction.


Conteúdo

TrkC is the high affinity catalytic receptor for the neurotrophin-3 (also known as NTF3 or NT-3). Similar to other NTRK receptors and receptor tyrosine kinases in general, ligand binding induces receptor dimerization followed by trans-autophosphorylation on conserved tyrosine in the intracellular (cytoplasmic) domain of the receptor. These conserved tyrosine serve as docking sites for adaptor proteins that trigger downstream signaling cascades. Signaling through PLCG1, PI3K and RAS [ disambiguation needed ] , downstream of activated NTRK3, regulates cell survival, proliferation and motility [7]

Moreover, TrkC has been identified as a novel synaptogenic adhesion molecule responsible for excitatory synapse development. [8]

The TrkC locus encodes at least eight isoforms including forms without the kinase domain or with kinase insertions adjacent to the major autophosphorylation site. These forms arise by alternative splicing events and are expressed in different tissues and cell types. [9] NT-3 activation of catalytic TrkC isoform promotes both proliferation of neural crest cells and neuronal differentiation. On the other hand, the binding of NT-3 to the non-catalytic TrkC isoform induces neuronal differentiation, but nor neuronal proliferation [10]

Tropomyosin receptor kinases, also known as neurotrophic tyrosine kinase receptors (Trk) play an essential role in the biology of neurons by mediating Neurotrophin-activated signaling. There are three transmembrane receptors TrkA, TrkB and TrkC (encoded by the genes NTRK1, NTRK2 and NTRK3 respectively) make up the Trk receptor family. [11] This family of receptors are all activated by neurotrophins, including NGF (for Nerve Growth Factor), BDNF (for Brain Derived Neurotrophic Factor), NT-4 (for Neurotrophin-4) and NT-3 (for Neurotrophin-3). While TrkA mediated the effects of NGF, TrkB is bound and activated by BDNF , NT-4 and NT-3. Further, TrkC binds and is activated by NT-3. [12] TrkB binds BDNF and NT-4 more strongly than it binds NT-3. TrkC binds NT-3 more strongly than TrkB does.

There is one other NT-3 receptor family besides the Trks (TrkC & TrkB), called the "LNGFR" (for "low affinity nerve growth factor receptor"). As opposed to TrkC, the LNGFR plays a somewhat less clear role in NT-3 biology. Some researchers have shown the LNGFR binds and serves as a "sink" for neurotrophins. Cells which express both the LNGFR and the Trk receptors might therefore have a greater activity - since they have a higher "microconcentration" of the neurotrophin. It has also been shown, however, that the LNGFR may signal a cell to die via apoptosis - so therefore cells expressing the LNGFR in the absence of Trk receptors may die rather than live in the presence of a neurotrophin.

It has been demonstrated that NTRK3 is a dependence receptor, meaning that it can be capable of inducing proliferation when it binds to its ligand NT-3, however, the absence of the NT-3 will result in the induction of apoptosis by NTRK3. [13]

With the past of the years, lot of studies have shown that the lack or deregulation of TrkC or the complex TrkC:NT-3 can be associated with different diseases.

One study have demonstrated that mice defective for either NT-3 or TrkC display severe sensory defects. These mice have normal nociception, but they are defective in proprioception, the sensory activity responsible for localizing the limbs in space. [14]

The reduction of TrkC expression has been observed in neurodegenerative diseases, including Alzheimer's (AD), Parkinson's (PD), and Huntington's diseases (HD). [15] The role of NT-3 was also therapeutically studied in models of amyotrophic lateral sclerosis (ALS) with loss of spinal cord motor neurons that express TrkC [16]

Moreover, it has been shown that TrkC plays a role in cancer. The expression and function of Trk subtypes are dependent on the tumor type. For example, in neuroblastoma, TrkC expression correlates with a good prognosis, but in breast, prostate and pancreatic cancers, the expression of the same TrkC subtype is associated with cancer progression and metastasis. [17]

Although originally identified as an oncogenic fusion in 1982, [18] only recently has there been a renewed interest in the Trk family as it relates to its role in human cancers because of the identification of NTRK1 (TrkA), NTRK2 (TrkB) and NTRK3 (TrkC) gene fusions and other oncogenic alterations in a number of tumor types. A number of Trk inhibitors are (in 2015) in clinical trials and have shown early promise in shrinking human tumors. [19] Family of neurotrophin receptors including NTRK3 have been shown to induce a variety of pleiotorpic response in malignant cells, including enhanced tumor cell invasiveness and chemotoxis. [20] Increased NTRK3 expression has been demonstrated in neuroblastoma, [21] in medulloblastoma, [22] and in neuroectodermal brain tumors. [23]

NTRK3 methylation Edit

The promoter region of NTRK3 contains a dense CpG island located relatively adjacent to the transcription start site (TSS). Using HumanMethylation450 arrays, quantitative methylation-specific PCR (qMSP), and Methylight assays, it has been indicated that NTRK3 is methylated in all CRC cell lines and non of the normal epithelium samples. In light of its preferential methylation in CRCs and because of its role as a neurotrophin receptor, it has been suggested to have a functional role in colorectal cancer formation. [24] It has also been suggested that methylation status of NTRK3 promoter is capable of discriminating CRC tumor samples from normal adjacent tumor-free tissue. Hence it can be considered as a biomarker for molecular detection of CRC, specially in combination with other markers like SEPT9. [25] NTRK3 has also been indicated as one of the genes in the panel of nine CpG methylation probes located at promoter or exon 1 region of eight genes (including DDIT3, FES, FLT3, SEPT5, SEPT9, SOX1, SOX17, and NTRK3) for prognostic prediction in ESCC (esophageal squamous cell carcinoma) patients. [26]

TrkC (NTRK3 gene) inhibitors in development Edit

Entrectinib (formerly RXDX-101) is an investigational drug developed by Ignyta, Inc., which has potential antitumor activity. It is an oral pan-TRK, ALK and ROS1 inhibitor that has demonstrated its anti tumor activity in murine, human tumor cell lines, and patient-derived xenograft tumor models. In vitro, entrectinib inhibits the Trk family members TrkA, TrkB and TrkC at low nano molar concentrations. It is highly bound to plasma proteins (99,5%), and can readily diffuse across the blood-brain barrier (BBB). [27]

Entrectinib has been approved by the FDA on August 15, 2019 for the treatment of adult and pediatric patients 12 years of age and older with solid tumors that have a neurotrophic tyrosine kinase receptor gene fusion [28]

TrkC has been shown to interact [ disambiguation needed ] with:

  • SH2B2
  • SQSTM1
  • KIDINS220
  • PTPRS [29]
  • MAPK8IP3/JIP3 [30][31][32][33][34]
  • TβRII [35]
  • DOK5 [36]
  • BMPRII [37]
  • PLCG1 [38][39]

Small molecules peptidomimetics based on β-turn NT-3, with the rationale of targeting the extracellular domain of the TrkC receptor have shown to be agonist of TrkC. [40] Posterior studies, have shown that peptidomimetics with an organic backbone, and a pharmacophore based on β-turn NT-3 structure can also function as an antagonist of TrkC. [41]


Fluorescent kinase probes enabling identification and dynamic imaging of HER2(+) Cells

The human epidermal growth factor receptor, EGFR/ ERBB/HER, family of receptor tyrosine kinases is central to many signaling pathways and a validated chemotherapy target in multiple cancers. While EGFR/ERBB-targeted therapies, including monoclonal antibodies, e.g., trastuzumab, and small molecule kinase inhibitors, such as lapatinib, have been developed, rapid identification and classification of cancer cells is key to identifying the best treatment regime. We report ERBB2 (also HER2) targeting kinase probes that exhibit a "turn-on" emission response upon binding. These live cell compatible probes differentiate ERBB2(+) cells from low-level, ERBB2(-) cells by targeting the intracellular ATP-binding pocket of ERBB2 with therapeutic inhibitor-like specificity. Beyond kinase expression levels, probe signal is linked to the phosphotyrosine-correlated activation state of the ERBB2 population. Additionally, the rapid signaling capability of the probes can report changes in activation state in live cells providing a unique type of complementary information to immunohistochemical assays of receptor kinase populations.


INTRODUÇÃO

The structure and function of the epidermal growth factor receptor (EGFR) is evolutionarily conserved fromCaenorhabditis elegans para Homo sapiens(Aroian et al., 1990) and its activity regulates the proliferation, motility, and differentiation of many different cell types (Sibilia and Wagner, 1995 Threadgill et al., 1995). Binding of any one of at least five ligands activates the intrinsic tyrosine kinase domain of the EGFR (van der Geer et al., 1994), which phosphorylates itself and activates other members of the EGFR family, such as HER2 (Stern and Kamps, 1988 van der Geer et al., 1994). Receptor phosphotyrosine residues act as nucleation sites for additional proteins such as Shc, Grb2, mSOS, ras-GAP, phospholipase C-γ, Eps8, and c-Cbl (Rozakis-Adcock et al., 1992 Fazioli et al., 1993 van der Geer et al., 1994 Levkowitz et al., 1998). These receptor signaling partners are activated by allosteric effects or by tyrosine phosphorylation, leading to recruitment of additional signaling molecules (van der Geer et al., 1994). Downstream kinase cascades and specific protein-protein assemblages can, in turn, determine cell type-specific responses (Tan and Kim, 1999).

Activated EGFR are rapidly internalized by coated pits, sorted through early endosomes, and ultimately degraded in lysosomes by a process generally known as receptor down-regulation (Wiley et al., 1991 Sorkin and Waters, 1993). G-protein coupled receptors, as well as other receptor tyrosine kinases, are also down-regulated after ligand activation (Sorkin and Waters, 1993 Kallal et al., 1998). Although degradation is the ultimate fate of internalized receptors, the rate of receptor degradation is much slower than their rate of internalization. Thus, substantial intracellular pools of receptors and ligands can accumulate (Wiley et al., 1985). It is clear that receptors are initially activated at the plasma membrane, but it is much less certain whether internalized receptors remain active until they are degraded. It is also unknown whether signals from internalized receptors are qualitatively different from those generated at the cell surface.

For more than a decade, investigators have debated the existence of “signaling endosomes.” Experiments with rat liver have demonstrated that, after the administration of a bolus of EGF, intracellular EGFR are associated with Shc, Grb2, and mSOS (Di Guglielmo et al., 1994). These signaling cofactors are thought to be responsible for initiating signals at the cell surface (van der Geeret al., 1994). Additionally, other receptor substrates, such as c-src and rho-B, are enriched in endosomes (Adamson et al., 1992 Kaplan et al., 1992). The strongest evidence supporting the signaling endosome hypothesis comes from recent genetic and biochemical experiments with the EGFR and the β-adrenergic receptor. Schmid and colleagues used a conditional dynamin mutant to block EGFR endocytosis, resulting in specific signal transduction pathways being up-regulated and others being attenuated (Vieiraet al., 1996). In similar experiments with the β-adrenergic receptor, endocytosis was inhibited with the use of both the nonspecific conditional dynamin mutation and a specific mutation in β-arrestin. This resulted in inhibition of mitogen-activated protein kinase activation (Daaka et al., 1998 Ahn et al., 1999). Together, these data suggest that specific signals can arise from the endosomal compartment.

Despite the positive evidence, it has been argued that EGFR signal transduction is primarily restricted to the cell surface (Fiore and Gill, 1999). To a large extent, this idea is based on the correlation between low rates of EGFR internalization and cell transformation (Wells et al., 1990 Huang et al., 1997). Supporting this argument is the observation that v-Cbl transforms cells at least in part by shunting EGFR back to the cell surface (Levkowitzet al., 1998). These data, however, do not directly rule out the possibility that signal transduction can arise from endosomes nor do they separate the effects of inhibiting receptor endocytosis from the effects of inhibiting ligand or receptor degradation. Endosomes could still make up an important signaling compartment.

A major difficulty in evaluating the role of endosomal signaling is the low sensitivity of current techniques. In general, one must isolate endosomal compartments at different times after ligand stimulation and evaluate their composition (Wada et al., 1992). Because of the low yield and time-consuming nature of this approach, previous studies have been restricted to abundant tissues, such as rat liver, or transformed cells that overexpress receptors or specific signaling components (Levkowitz et al., 1998 Xue and Lucocq, 1998). Although these studies have been informative, they have necessary limitations. Rat liver is not a physiologically important target of EGFR action and overexpression of receptors or signaling molecules can lead to altered trafficking or function. These technical issues have made it difficult to determine whether endosomal signaling is a normal consequence of EGFR activation or is restricted to specific experimental systems.

To investigate the role of EGFR trafficking in its biological actions, we have used responsive human mammary epithelial cells (HMEC). Genetic and biochemical studies in mice have shown that normal EGFR function is critical for the development of the mammary epithelium (Fowler et al., 1995 Xie et al., 1997). In vitro, blocking the EGFR in HMEC leads to cell cycle arrest as well as inhibition of cell migration and organization (Stampfer et al., 1993 Wileyet al., 1998 Dong et al., 1999). Importantly, HMEC normally express high levels of EGFR, facilitating biochemical studies (Bates et al., 1990 Burke and Wiley, 1999). To investigate EGFR trafficking, we developed a new biochemical technique to isolate activated EGFR within endosomes with the use of a reversibly biotinylated nonantagonistic anti-EGFR antibody. In addition, we developed antibodies specific to tyrosine-phosphorylated EGFR to follow activated EGFR by immunofluorescence techniques. With the use of these approaches, we observed that the pattern of EGFR association with substrates and adaptor proteins changed as the EGFR moved from the cell surface through the endosomal compartment. In addition, we found that internalized EGFR lost both phosphotyrosine and associated ligand before degradation. Our results suggest that endosomes make up a major site of regulated EGFR signaling in responsive cells and that ligand loss is the proximal cause of attenuated receptor signaling.


2018 &ndash present Biomedical Scientist, Physical and Life Sciences, Lawrence Livermore National Laboratory

2015 &ndash 2018 Postdoctoral Fellow, Physical and Life Sciences, Lawrence Livermore National Laboratory

2010 - 2015 Graduate Student Research Associate, Radiation Oncology, University of California, Davis

2008 - 2010 Research Specialist, The NSF Center for Biophotonics, University of California, Davis

2001 - 2003 Project Scientist, State Key Laboratory of Genetic Engineering, Fudan University, Shanghai, China


Duolink ® Proximity Ligation Assay (PLA) Multicolor Kits for Sensitive Multiplex Detection

Protein-protein Interaction Events

Interacting proteins often function as key components in the initiation and cascade of inter-and intracellular communications and influence a large variety of cellular functions and behaviors. However, accurately identifying protein-protein interactions in fixed tissues and cells can be challenging, especially for low abundant proteins or weekly interacting protein complexes. Due to the discovery of a growing list of fluorophores and novel detection technologies, diverse fields of research have noted tremendous advancements due to the identification of intracellular protein interactions. As a result, the detection of these interactions has led to improvements in our understanding of protein function and have provided key insight into numerous signaling pathways in healthy and diseased tissues. Duolink ® PLA Multicolor technology allows for the visual detection and quantification of several distinct protein-protein interactions simultaneously in fixed tissues and cells. 1

Duolink ® PLA Multicolor Technology

Duolink ® PLA Multicolor Probemaker kits and reagent packs deliver an effective solution for visually identifying and quantifying up to four distinct protein-protein interactions within a variety of fixed cells and tissues. Two highly specific primary antibodies are required for the evaluation of each protein-protein interaction. The generation of a plus and minus PLA probe is accomplished using Probemaker technology and involves directly conjugating a single oligo to each primary antibody. If each plus and minus PLA probe pair are near each other (<40nM), DNA ligation followed by rolling circle amplification occurs with the addition of polymerase and produces a connected single-stranded concatemeric sequence with up to 1000 copies of the template. The addition of labeled oligos or detection probes hybridize to the complementary template sequences within the amplicon resulting in up to a 1000x protein signal amplification for each protein detection event. Multiplex Duolink ® PLA Multicolor technology allows for the detection of both stable and weakly interacting proteins for up to four protein events in detection colors consisting of green, orange, red, and far red (figura 1) Importantly, both imaging and data analysis are achievable with the use of standard high-content screening imagers, flow cytometry, or imaging flow cytometers.

Figura 1: Directly conjugated primary antibodies (PLA probes), generated with Probemaker technology, are used to identify and amplify signals for four unique protein events in green, orange, red, and far red detection colors.

Duolink ® PLA Multicolor Technology and Cellular Pathway Analyses

Investigating protein-protein interactions and cellular pathway analyses includes a variety of methods and technologies. Commonly used fluorophores and fluorescent microscopy may involve the use of recombinant proteins and the colocalization of signals within the cell or tissue of interest. However, the use of recombinant proteins and colocalization imaging techniques are limited and may not provide data for interacting proteins in their native state. Furthermore, studying various diseases where complex signaling pathways are involved may include the analysis of proteins with subtle post-translational modifications or the detection of phosphorylated and non-phosphorylated proteins. Duolink ® PLA Multicolor technology is effective at analyzing protein-protein interactions with potential post-translational or alternative modifications.

Investigation of the HER Family of Receptor Tyrosine Kinases Using Multiplex Duolink ® PLA Multicolor Technology

The HER family of receptor tyrosine kinases consists of four members: EGFR (HER1 or ErbB1), HER2 (ErbB2), HER3 (ErbB3), and HER4 (ErbB4). Importantly, these proteins are overexpressed or mutated in numerous types of cancer and are of great interests to researchers. 2 Provided the high specificity and the ability to investigate up to four protein-protein interactions simultaneously, Multiplex Duolink ® PLA Multicolor technology can be effectively used to visualize complex signaling cascades, including the EGFR pathway (Figura 2).

Figura 2: (A) Schematic of multiplex PLA detection and analysis of EGFR signaling of post-translational modifications and protein-protein interactions in SK-OV3 cells treated with EGF (+ EGF) or untreated cells. Multiplex PLA detection of the phosphorylation of EGFR (green), EGFR-HER2 interactions (orange), and the phosphorylation of HER2 (far red) were performed on adherent cells (B) or detached cells (C). Data collection was performed by fluorescence microscopy or flow cytometry, respectively. Nuclei were stained with DAPI (blue) in (B).

Refer to the Duolink ® PLA Multicolor Detection protocol for further detailed information on executing an experiment.


Resumo

Although a previously developed bump-hole approach has proven powerful in generating specific inhibitors for mapping functions of protein kinases, its application is limited by the intolerance of the large-to-small mutation by certain kinases and the inability to control two kinases separately in the same cells. Herein, we describe the development of an alternative chemical-genetic approach to overcome these limitations. Our approach features the use of an engineered cysteine residue at a particular position as a reactive feature to sensitize a kinase of interest to selective covalent blockade by electrophilic inhibitors and is thus termed the Ele-Cys abordagem. We successfully applied the Ele-Cys approach to identify selective covalent inhibitors of a receptor tyrosine kinase EphB1 and solved cocrystal structures to determine the mode of covalent binding. Importantly, the Ele-Cys and bump-hole approaches afforded orthogonal inhibition of two distinct kinases in the cell, opening the door to their combined use in the study of multikinase signaling pathways.


Assista o vídeo: Tirosina Kinasa- JAK STAT y Receptores Nucleares --Los Waffles (Dezembro 2022).