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1.
Sci Signal ; 13(621)2020 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-32127496

RESUMO

Non-small cell lung cancer (NSCLC) is often characterized by mutually exclusive mutations in the epidermal growth factor receptor (EGFR) or the guanosine triphosphatase KRAS. We hypothesized that blocking EGFR palmitoylation, previously shown to inhibit EGFR activity, might alter downstream signaling in the KRAS-mutant setting. Here, we found that blocking EGFR palmitoylation, by either knocking down the palmitoyltransferase DHHC20 or expressing a palmitoylation-resistant EGFR mutant, reduced activation of the kinase PI3K, the abundance of the transcription factor MYC, and the proliferation of cells in culture, as well as reduced tumor growth in a mouse model of KRAS-mutant lung adenocarcinoma. Knocking down DHHC20 reduced the growth of existing tumors derived from human KRAS-mutant lung cancer cells and increased the sensitivity of these cells to a PI3K inhibitor. Palmitoylated EGFR interacted with the PI3K regulatory subunit PIK3R1 (p85) and increased the recruitment of the PI3K heterodimer to the plasma membrane. Alternatively, blocking palmitoylation increased the association of EGFR with the MAPK adaptor Grb2 and decreased that with p85. This binary switching between MAPK and PI3K signaling, modulated by EGFR palmitoylation, was only observed in the presence of oncogenic KRAS. These findings suggest a mechanism whereby oncogenic KRAS saturates signaling through unpalmitoylated EGFR, reducing formation of the PI3K signaling complex. Future development of DHHC20 inhibitors to reduce EGFR-PI3K signaling could be beneficial to patients with KRAS-mutant tumors.


Assuntos
Carcinogênese/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Lipoilação , Neoplasias Pulmonares/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Transdução de Sinais , Aciltransferases/genética , Aciltransferases/metabolismo , Animais , Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Receptores ErbB/genética , Receptores ErbB/metabolismo , Neoplasias Pulmonares/genética , Camundongos , Células NIH 3T3 , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas p21(ras)/genética
2.
J Cell Mol Med ; 22(8): 3887-3898, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29761849

RESUMO

Acute myeloid leukaemia (AML) is an aggressive haematological malignancy with an unmet need for improved therapies. Responses to standard cytotoxic therapy in AML are often transient because of the emergence of chemotherapy-resistant disease. The MUC1-C oncoprotein governs critical pathways of tumorigenesis, including self-renewal and survival, and is aberrantly expressed in AML blasts and leukaemia stem cells (LSCs). However, a role for MUC1-C in linking leukaemogenesis and resistance to treatment has not been described. In this study, we demonstrate that MUC1-C overexpression is associated with increased leukaemia initiating capacity in an NSG mouse model. In concert with those results, MUC1-C silencing in multiple AML cell lines significantly reduced the establishment of AML in vivo. In addition, targeting MUC1-C with silencing or pharmacologic inhibition with GO-203 led to a decrease in active ß-catenin levels and, in-turn, down-regulation of survivin, a critical mediator of leukaemia cell survival. Targeting MUC1-C was also associated with increased sensitivity of AML cells to Cytarabine (Ara-C) treatment by a survivin-dependent mechanism. Notably, low MUC1 and survivin gene expression were associated with better clinical outcomes in patients with AML. These findings emphasize the importance of MUC1-C to myeloid leukaemogenesis and resistance to treatment by driving survivin expression. Our findings also highlight the potential translational relevance of combining GO-203 with Ara-C for the treatment of patients with AML.

3.
Biochem Biophys Res Commun ; 493(1): 213-219, 2017 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-28899783

RESUMO

Currently, there are no effective therapeutic strategies targeting Kras driven cancers, and therefore, identifying new targeted therapies and overcoming drug resistance have become paramount for effective long-term cancer therapy. We have found that reducing expression of the palmitoyl transferase DHHC20 increases cell death induced by the EGFR inhibitor gefitinib in Kras and EGFR mutant cell lines, but not MCF7 cells harboring wildtype Kras. We show that the increased gefitinib sensitivity in cancer cells induced by DHHC20 inhibition is mediated directly through loss of palmitoylation on a previously identified cysteine residue in the C-terminal tail of EGFR. We utilized an EGFR point mutant in which the palmitoylated cysteine 1025 is mutated to alanine (EGFRC1025A), that results in receptor activation. Expression of the EGFR mutant alone in NIH3T3 cells does not increase sensitivity to gefitinib-induced cell death. However, when EGFRC1025A is expressed in cells expressing activated KrasG12V, EGFR inhibitor induced cell death is increased. Surprisingly, lung cancer cells harboring the EGFR inhibitor resistant mutation, T790M, become sensitive to EGFR inhibitor treatment when DHHC20 is inhibited. Finally, the small molecule, 2-bromopalmitate, which has been shown to inhibit palmitoyl transferases, acts synergistically with gefitinib to induce cell death in the gefitinib resistant cell line NCI-H1975.


Assuntos
Cisteína/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Neoplasias Experimentais/fisiopatologia , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Cocarcinogênese , Cisteína/metabolismo , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos , Receptores ErbB/metabolismo , Gefitinibe , Humanos , Lipoilação/efeitos dos fármacos , Lipoilação/genética , Células MCF-7 , Proteínas de Membrana , Mutação , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Quinazolinas/administração & dosagem , Quinazolinas/farmacologia
4.
J Clin Invest ; 127(6): 2378-2391, 2017 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-28481227

RESUMO

The esophageal lumen is lined by a stratified squamous epithelium comprised of proliferative basal cells that differentiate while migrating toward the luminal surface and eventually desquamate. Rapid epithelial renewal occurs, but the specific cell of origin that supports this high proliferative demand remains unknown. Herein, we have described a long-lived progenitor cell population in the mouse esophageal epithelium that is characterized by expression of keratin 15 (Krt15). Genetic in vivo lineage tracing revealed that the Krt15 promoter marks a long-lived basal cell population able to self-renew, proliferate, and generate differentiated cells, consistent with a progenitor/stem cell population. Transcriptional profiling demonstrated that Krt15+ basal cells are molecularly distinct from Krt15- basal cells. Depletion of Krt15-derived cells resulted in decreased proliferation, thereby leading to atrophy of the esophageal epithelium. Further, Krt15+ cells were radioresistant and contributed to esophageal epithelial regeneration following radiation-induced injury. These results establish the presence of a long-lived and indispensable Krt15+ progenitor cell population that provides additional perspective on esophageal epithelial biology and the widely prevalent diseases that afflict this epithelium.


Assuntos
Esôfago/citologia , Queratina-15/metabolismo , Células-Tronco/fisiologia , Animais , Diferenciação Celular , Movimento Celular , Proliferação de Células , Sobrevivência Celular/efeitos da radiação , Esôfago/fisiologia , Esôfago/efeitos da radiação , Homeostase , Humanos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mucosa/citologia , Regiões Promotoras Genéticas , Lesões Experimentais por Radiação/fisiopatologia , Regeneração , Células-Tronco/efeitos da radiação , Ativação Transcricional
5.
Mol Cell ; 62(3): 385-396, 2016 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-27153536

RESUMO

Inappropriate activation of the receptor tyrosine kinase EGFR contributes to a variety of human malignancies. Here we show a mechanism to induce vulnerability to an existing first line treatment for EGFR-driven cancers. We find that inhibiting the palmitoyltransferase DHHC20 creates a dependence on EGFR signaling for cancer cell survival. The loss of palmitoylation increases sustained EGFR signal activation and sensitizes cells to EGFR tyrosine kinase inhibition. Our work shows that the reversible modification of EGFR with palmitate "pins" the unstructured C-terminal tail to the plasma membrane, impeding EGFR activation. We identify by mass spectrometry palmitoylated cysteine residues within the C-terminal tail where mutation of the cysteine residues to alanine is sufficient to activate EGFR signaling promoting cell migration and transformation. Our results reveal that the targeting of a peripheral modulator of EGFR signaling, DHHC20, causes a loss of signal regulation and susceptibility to EGFR inhibitor-induced cell death.


Assuntos
Aciltransferases/metabolismo , Neoplasias da Mama/enzimologia , Receptores ErbB/metabolismo , Processamento de Proteína Pós-Traducional , Transdução de Sinais , Aciltransferases/genética , Animais , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Morte Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Cisteína , Endocitose , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/química , Receptores ErbB/efeitos dos fármacos , Receptores ErbB/genética , Proteína Adaptadora GRB2/metabolismo , Gefitinibe , Células HEK293 , Humanos , Lipoilação , Espectrometria de Massas , Camundongos , Mutação , Células NIH 3T3 , Conformação Proteica , Inibidores de Proteínas Quinases/farmacologia , Transporte Proteico , Proteólise , Quinazolinas/farmacologia , Interferência de RNA , Transdução de Sinais/efeitos dos fármacos , Relação Estrutura-Atividade , Transfecção
6.
Cancer Res ; 76(6): 1538-48, 2016 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-26833129

RESUMO

Dysregulation of MYC expression is a hallmark of cancer, but the development of agents that target MYC has remained challenging. The oncogenic MUC1-C transmembrane protein is, like MYC, aberrantly expressed in diverse human cancers. The present studies demonstrate that MUC1-C induces MYC expression in KRAS mutant non-small cell lung cancer (NSCLC) cells, an effect that can be suppressed by targeting MUC1-C via shRNA silencing, CRISPR editing, or pharmacologic inhibition with GO-203. MUC1-C activated the WNT/ß-catenin (CTNNB1) pathway and promoted occupancy of MUC1-C/ß-catenin/TCF4 complexes on the MYC promoter. MUC1-C also promoted the recruitment of the p300 histone acetylase (EP300) and, in turn, induced histone H3 acetylation and activation of MYC gene transcription. We also show that targeting MUC1-C decreased the expression of key MYC target genes essential for the growth and survival of NSCLC cells, such as TERT and CDK4. Based on these results, we found that the combination of GO-203 and the BET bromodomain inhibitor JQ1, which targets MYC transcription, synergistically suppressed MYC expression and cell survival in vitro as well as tumor xenograft growth. Furthermore, MUC1 expression significantly correlated with that of MYC and its target genes in human KRAS mutant NSCLC tumors. Taken together, these findings suggest a therapeutic approach for targeting MYC-dependent cancers and provide the framework for the ongoing clinical studies addressing the efficacy of MUC1-C inhibition in solid tumors.


Assuntos
Adenocarcinoma/genética , Neoplasias Pulmonares/genética , Mucina-1/genética , Mutação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma de Pulmão , Animais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Proteína p300 Associada a E1A/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Células HEK293 , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Camundongos Nus , Mutação/genética , Peptídeos/farmacologia , RNA Interferente Pequeno/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Proteínas Wnt/genética , beta Catenina/genética
7.
Clin Cancer Res ; 21(10): 2338-47, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25712682

RESUMO

PURPOSE: The MUC1-C oncoprotein is an intracellular target that is druggable with cell-penetrating peptide inhibitors. However, development of peptidyl drugs for treating cancer has been a challenge because of unfavorable pharmacokinetic parameters and limited cell-penetrating capabilities. EXPERIMENTAL DESIGN: Encapsulation of the MUC1-C inhibitor GO-203 in novel polymeric nanoparticles was studied for effects on intracellular targeting of MUC1-C signaling and function. RESULTS: Our results show that loading GO-203 into tetrablock polylactic acid (PLA)-polyethylene glycol (PEG)-polypropylene glycol (PPG)-PEG copolymers is achievable and, notably, is enhanced by increasing PEG chain length. In addition, we found that release of GO-203 from these nanoparticles is controllable over at least 7 days. GO-203/nanoparticle treatment of MUC1-C-positive breast and lung cancer cells in vitro was more active with less frequent dosing than that achieved with nonencapsulated GO-203. Moreover, treatment with GO-203/nanoparticles blocked MUC1-C homodimerization, consistent with on-target effects. GO-203/nanoparticle treatment was also effective in downregulating TIGAR, disrupting redox balance, and inhibiting the self-renewal capacity of cancer cells. Significantly, weekly administration of GO-203/nanoparticles to mice bearing syngeneic or xenograft tumors was associated with regressions that were comparable with those found when dosing on a daily basis with GO-203. CONCLUSIONS: These findings thus define an effective approach for (i) sustained administration of GO-203 in polymeric PLA-(PEG-PPG-PEG) nanoparticles to target MUC1-C in cancer cells and (ii) the potential delivery of other anticancer peptide drugs.


Assuntos
Antineoplásicos/administração & dosagem , Carcinoma de Ehrlich/tratamento farmacológico , Mucina-1/metabolismo , Nanocápsulas/administração & dosagem , Peptídeos/administração & dosagem , Sequência de Aminoácidos , Animais , Antineoplásicos/química , Carcinoma de Ehrlich/patologia , Linhagem Celular Tumoral , Autorrenovação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Composição de Medicamentos , Humanos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Terapia de Alvo Molecular , Nanocápsulas/química , Oxirredução , Peptídeos/química , Multimerização Proteica , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Oncotarget ; 5(19): 8893-905, 2014 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-25245423

RESUMO

Non-small cell lung cancers (NSCLCs) that harbor an oncogenic KRAS mutation are often associated with resistance to targeted therapies. The MUC1-C transmembrane protein is aberrantly overexpressed in NSCLCs and confers a poor outcome; however, the functional role for MUC1-C in mutant KRAS NSCLC cells has remained unclear. The present studies demonstrate that silencing MUC1-C in A549/KRAS(G12S) and H460/KRAS(Q61H) NSCLC cells is associated with downregulation of AKT signaling and inhibition of growth. Overexpression of a MUC1-C(CQC→AQA) mutant, which inhibits MUC1-C homodimerization and function, suppressed both AKT and MEK activation. Moreover, treatment with GO-203, an inhibitor of MUC1-C homodimerization, blocked AKT and MEK signaling and decreased cell survival. The results further demonstrate that targeting MUC1-C suppresses expression of the ZEB1 transcriptional repressor by an AKT-mediated mechanism, and in turn induces miR-200c. In concert with these effects on the ZEB1/miR-200c regulatory loop, targeting MUC1-C was associated with reversal of the epithelial-mesenchymal transition (EMT) and inhibition of self-renewal capacity. Loss of MUC1-C function also attenuated KRAS independence and inhibited growth of KRAS mutant NSCLC cells as tumors in mice. These findings support a model in which targeting MUC1-C inhibits mutant KRAS signaling in NSCLC cells and thereby reverses the EMT phenotype and decreases self-renewal.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Transição Epitelial-Mesenquimal/genética , Neoplasias Pulmonares/genética , Mucina-1/genética , Proteínas Proto-Oncogênicas/genética , Proteínas ras/genética , Animais , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Proteínas de Homeodomínio/biossíntese , Humanos , MAP Quinase Quinase 1/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Mucina-1/biossíntese , Transplante de Neoplasias , Oxidiazóis/farmacologia , Peptídeos/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas p21(ras) , Interferência de RNA , RNA Interferente Pequeno , Transdução de Sinais/efeitos dos fármacos , Esferoides Celulares , Fatores de Transcrição/biossíntese , Células Tumorais Cultivadas , Homeobox 1 de Ligação a E-box em Dedo de Zinco
9.
Clin Cancer Res ; 20(21): 5423-34, 2014 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-25189483

RESUMO

PURPOSE: Non-small cell lung cancers (NSCLC) that express EGF receptor with activating mutations frequently develop resistance to EGFR kinase inhibitors. The mucin 1 (MUC1) heterodimeric protein is aberrantly overexpressed in NSCLC cells and confers a poor prognosis; however, the functional involvement of MUC1 in mutant EGFR signaling is not known. EXPERIMENTAL DESIGN: Targeting the oncogenic MUC1 C-terminal subunit (MUC1-C) in NSCLC cells harboring mutant EGFR was studied for effects on signaling, growth, clonogenic survival, and tumorigenicity. RESULTS: Stable silencing of MUC1-C in H1975/EGFR(L858R/T790M) cells resulted in downregulation of AKT signaling and inhibition of growth, colony formation, and tumorigenicity. Similar findings were obtained when MUC1-C was silenced in gefitinib-resistant PC9GR cells expressing EGFR(delE746_A750/T790M). The results further show that expression of a MUC1-C(CQC → AQA) mutant, which blocks MUC1-C homodimerization, suppresses EGFR(T790M), AKT and MEK → ERK activation, colony formation, and tumorigenicity. In concert with these results, treatment of H1975 and PC9GR cells with GO-203, a cell-penetrating peptide that blocks MUC1-C homodimerization, resulted in inhibition of EGFR, AKT, and MEK → ERK signaling and in loss of survival. Combination studies of GO-203 and afatinib, an irreversible inhibitor of EGFR, further demonstrate that these agents are synergistic in inhibiting growth of NSCLC cells harboring the activating EGFR(T790M) or EGFR(delE746-A750) mutants. CONCLUSIONS: These findings indicate that targeting MUC1-C inhibits mutant EGFR signaling and survival, and thus represents a potential approach alone and in combination for the treatment of NSCLCs resistant to EGFR kinase inhibitors.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Sobrevivência Celular/efeitos dos fármacos , Receptores ErbB/metabolismo , Neoplasias Pulmonares/tratamento farmacológico , Mucina-1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Humanos , Neoplasias Pulmonares/genética , Oncogenes/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/genética
10.
J Biol Chem ; 288(43): 30892-903, 2013 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-24043631

RESUMO

Aldehyde dehydrogenase 1A1 (ALDH1A1) activity is used as a marker of breast cancer stem cells; however, little is known about the regulation of ALDH1A1 expression. Mucin 1 (MUC1) is a heterodimeric protein that is aberrantly overexpressed in most human breast cancers. In studies of breast cancer cells stably silenced for MUC1 or overexpressing the oncogenic MUC1-C subunit, we demonstrate that MUC1-C is sufficient for induction of MEK → ERK signaling and that treatment with a MUC1-C inhibitor suppresses ERK activation. In turn, MUC1-C induces ERK-mediated phosphorylation and activation of the CCAAT/enhancer-binding protein ß (C/EBPß) transcription factor. The results further show that MUC1-C and C/EBPß form a complex on the ALDH1A1 gene promoter and activate ALDH1A1 gene transcription. MUC1-C-induced up-regulation of ALDH1A1 expression is associated with increases in ALDH activity and is detectable in stem-like cells when expanded as mammospheres. These findings demonstrate that MUC1-C (i) activates a previously unrecognized ERK→C/EBPß→ALDH1A1 pathway, and (ii) promotes the induction of ALDH activity in breast cancer cells.


Assuntos
Aldeído Desidrogenase/biossíntese , Neoplasias da Mama/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Sistema de Sinalização das MAP Quinases , Mucina-1/metabolismo , Proteínas de Neoplasias/metabolismo , Aldeído Desidrogenase/genética , Família Aldeído Desidrogenase 1 , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteína beta Intensificadora de Ligação a CCAAT/genética , Linhagem Celular Tumoral , Feminino , Regulação Enzimológica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Mucina-1/genética , Proteínas de Neoplasias/genética , Subunidades Proteicas , Retinal Desidrogenase , Transcrição Gênica/genética
11.
Mol Cancer Res ; 11(7): 714-23, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23538857

RESUMO

Tamoxifen resistance of estrogen receptor-positive (ER+) breast cancer cells has been linked in part to activation of receptor tyrosine kinases, such as HER2, and the PI3K-AKT pathway. Mucin 1 (MUC1) is aberrantly overexpressed in about 90% of human breast cancers, and the oncogenic MUC1-C subunit is associated with ERα. The present studies using HER2 overexpressing BT-474 breast cancer cells, which are constitutively resistant to tamoxifen, demonstrate that silencing MUC1-C is associated with (i) downregulation of p-HER2 and (ii) sensitivity to tamoxifen-induced growth inhibition and loss of clonogenic survival. In contrast, overexpression of MUC1-C in tamoxifen-sensitive MCF-7 breast cancer cells resulted in upregulation of p-AKT and tamoxifen resistance. We show that MUC1-C forms complexes with ERα on the estrogen-responsive promoter of Rab31 and that MUC1-C blocks tamoxifen-induced decreases in ERα occupancy. MUC1-C also attenuated tamoxifen-induced decreases in (i) recruitment of the coactivator CREB binding protein, (ii) Rab31 promoter activation, and (iii) Rab31 mRNA and protein levels. The importance of MUC1-C is further supported by the demonstration that targeting MUC1-C with the cell-penetrating peptide inhibitor, GO-203, sensitized tamoxifen-resistant cells to tamoxifen treatment. Moreover, we show that targeting MUC1-C in combination with tamoxifen is highly synergistic in the treatment of tamoxifen-resistant breast cancer cells. Combined, these findings indicate that MUC1-C contributes to tamoxifen resistance.


Assuntos
Neoplasias da Mama/patologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Mucina-1/metabolismo , Proteínas de Neoplasias/metabolismo , Tamoxifeno/farmacologia , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Receptor alfa de Estrogênio/metabolismo , Estrogênios/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Humanos , Peptídeos/farmacologia , Regiões Promotoras Genéticas/genética , Subunidades Proteicas/metabolismo , Receptor ErbB-2/metabolismo , Transcrição Gênica/efeitos dos fármacos , Proteínas rab de Ligação ao GTP/genética
12.
PLoS One ; 7(7): e39432, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22792175

RESUMO

Rab31 is a member of the Ras superfamily of small GTPases that has been linked to poor outcomes in patients with breast cancer. The MUC1-C oncoprotein is aberrantly overexpressed in most human breast cancers and also confers a poor prognosis. The present results demonstrate that MUC1-C induces Rab31 expression in estrogen receptor positive (ER+) breast cancer cells. We show that MUC1-C forms a complex with estrogen receptor α (ERα) on the Rab31 promoter and activates Rab31 gene transcription in an estrogen-dependent manner. In turn, Rab31 contributes to the upregulation of MUC1-C abundance in breast cancer cells by attenuating degradation of MUC1-C in lysosomes. Expression of an inactive Rab31(S20N) mutant in nonmalignant breast epithelial cells confirmed that Rab31 regulates MUC1-C expression. The functional significance of the MUC1-C/Rab31 interaction is supported by the demonstration that Rab31 confers the formation of mammospheres by a MUC1-C-dependent mechanism. Analysis of microarray databases further showed that (i) Rab31 is expressed at higher levels in breast cancers as compared to that in normal breast tissues, (ii) MUC1+ and ER+ breast cancers have increased levels of Rab31 expression, and (iii) patients with Rab31-positive breast tumors have a significantly decreased ten-year overall survival as compared to those with Rab31-negative tumors. These findings indicate that MUC1-C and Rab31 function in an autoinductive loop that contributes to overexpression of MUC1-C in breast cancer cells.


Assuntos
Neoplasias da Mama/metabolismo , Mucina-1/metabolismo , Receptores de Estrogênio/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Estradiol/farmacologia , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Mucina-1/genética , Regiões Promotoras Genéticas , Ligação Proteica , Esferoides Celulares , Ativação Transcricional , Células Tumorais Cultivadas , Proteínas rab de Ligação ao GTP/genética
13.
Hybridoma (Larchmt) ; 30(6): 531-5, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22149278

RESUMO

Mucin 1 (MUC1) is a heterodimeric protein that is aberrantly overexpressed in diverse human carcinomas and certain hematologic malignancies. The transmembrane MUC1-C subunit confers tumorigenicity and is a target for anti-cancer drug development. In this regard, the MUC1-C cytoplasmic domain interacts with multiple effectors that have been linked to transformation. Here we report on the generation of a mouse monoclonal antibody (MAb) against the human MUC1-C cytoplasmic domain (MUC1-CD). This IgG1 MAb, designated anti-MUC1-CD, reacts with the NYGQLDIFP epitope. We show that anti-MUC1-CD is useful in immunoblotting and immunoprecipitation experiments. In addition, anti-MUC1-CD can be used to detect expression of the MUC1-C subunit in formalin-fixed, paraffin-embedded tissues. The MUC1-C inhibitor has entered Phase I evaluation for patients with refractory solid tumors. The present results indicate that the anti-MUC1-CD antibody could be useful as a biomarker to identify patients with tumors that may be responsive to MUC1-C inhibitors.


Assuntos
Anticorpos Monoclonais Murinos/imunologia , Epitopos/imunologia , Mucina-1/imunologia , Sequência de Aminoácidos , Animais , Reações Antígeno-Anticorpo , Biomarcadores Tumorais/imunologia , Linhagem Celular Tumoral , Mapeamento de Epitopos , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Hibridomas/imunologia , Imunização , Immunoblotting , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imuno-Histoquímica , Imunoprecipitação , Isopropiltiogalactosídeo/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Mucina-1/genética , Estrutura Terciária de Proteína , Estômago/imunologia , Fixação de Tecidos/métodos
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