RESUMO
Therapy-related leukemia carries a poor prognosis, and leukemia after chemotherapy is a growing risk in clinic, whose mechanism is still not well understood. Ikaros transcription factor is an important regulator in hematopoietic cells development and differentiation. In the absence of Ikaros, lymphoid cell differentiation is blocked at an extremely early stage, and myeloid cell differentiation is also significantly affected. In this work, we showed that chemotherapeutic drug etoposide reduced the protein levels of several isoforms of Ikaros including IK1, IK2 and IK4, but not IK6 or IK7, by accelerating protein degradation, in leukemic cells. To investigate the molecular mechanism of Ikaros degradation induced by etoposide, immunoprecipitation coupled with LC-MS/MS analysis was conducted to identify changes in protein interaction with Ikaros before and after etoposide treatment, which uncovered KCTD5 protein. Our further study demonstrates that KCTD5 is the key stabilizing factor of Ikaros and chemotherapeutic drug etoposide induces Ikaros protein degradation through decreasing the interaction of Ikaros with KCTD5. These results suggest that etoposide may induce leukemic transformation by downregulating Ikaros via KCTD5, and our work may provide insights to attenuate the negative impact of chemotherapy on hematopoiesis.
Assuntos
Etoposídeo , Fator de Transcrição Ikaros , Fator de Transcrição Ikaros/metabolismo , Etoposídeo/farmacologia , Humanos , Proteólise/efeitos dos fármacos , Antineoplásicos Fitogênicos/farmacologiaRESUMO
Immune checkpoint blockade therapy has been successful in treating some types of cancer but has not shown clinical benefits for treating leukaemia1. This result suggests that leukaemia uses unique mechanisms to evade this therapy. Certain immune inhibitory receptors that are expressed by normal immune cells are also present on leukaemia cells. Whether these receptors can initiate immune-related primary signalling in tumour cells remains unknown. Here we use mouse models and human cells to show that LILRB4, an immunoreceptor tyrosine-based inhibition motif-containing receptor and a marker of monocytic leukaemia, supports tumour cell infiltration into tissues and suppresses T cell activity via a signalling pathway that involves APOE, LILRB4, SHP-2, uPAR and ARG1 in acute myeloid leukaemia (AML) cells. Deletion of LILRB4 or the use of antibodies to block LILRB4 signalling impeded AML development. Thus, LILRB4 orchestrates tumour invasion pathways in monocytic leukaemia cells by creating an immunosuppressive microenvironment. LILRB4 represents a compelling target for the treatment of monocytic AML.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/patologia , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Evasão Tumoral/imunologia , Animais , Apolipoproteínas E/metabolismo , Arginase/metabolismo , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Movimento Celular , Proliferação de Células , Feminino , Humanos , Tolerância Imunológica/imunologia , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Masculino , Glicoproteínas de Membrana , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Ligação Proteica , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Receptores de Superfície Celular/deficiência , Receptores de Superfície Celular/genética , Receptores Imunológicos , Receptores de Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Evasão Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Breast cancer is the leading cause of cancer death for women worldwide. Most of the breast cancer death are due to disease recurrence and metastasis. Increasingly accumulating evidence indicates that mitochondria play key roles in cancer progression and metastasis. Our recent study revealed that transmembrane protein PRRG4 promotes the metastasis of breast cancer. However, it is not clear whether PRRG4 can affect the migration and invasion of breast cancer cells through regulating mitochondria function. METHODS: RNA-seq analyses were performed on breast cancer cells expressing control and PRRG4 shRNAs. Quantitative PCR analysis and measurements of mitochondrial ATP content and oxygen consumption were carried out to explore the roles of PRRG4 in regulating mitochondrial function. Luciferase reporter plasmids containing different lengths of promoter fragments were constructed. Luciferase activities in breast cancer cells transiently transfected with these reporter plasmids were analyzed to examine the effects of PRRG4 overexpression on promoter activity. Transwell assays were performed to determine the effects of PRRG4-regulated pathway on migratory behaviors of breast cancer cells. RESULTS: Analysis of the RNA-seq data revealed that PRRG4 knockdown decreased the transcript levels of all the mitochondrial protein-encoding genes. Subsequently, studies with PRRG4 knockdown and overexpression showed that PRRG4 expression increased mitochondrial DNA (mtDNA) content. Mechanistically, PRRG4 via Src activated STAT3 in breast cancer cells. Activated STAT3 in turn promoted the transcription of mtDNA polymerase POLG through a STAT3 DNA binding site present in the POLG promoter region, and increased mtDNA content as well as mitochondrial ATP production and oxygen consumption. In addition, PRRG4-mediated activation of STAT3 also enhanced filopodia formation, migration, and invasion of breast cancer cells. Moreover, PRRG4 elevated migratory behaviors and mitochondrial function of breast cancer cells through POLG. CONCLUSION: Our results indicate that PRRG4 via the Src-STAT3-POLG axis enhances mitochondrial function and promotes migratory behaviors of breast cancer cells.
RESUMO
BACKGROUND: Pancreatic cancer is one of the most aggressive malignancies without effective targeted therapies. MUC1 has emerged as a potential common target for cancer therapy because it is overexpressed in a variety of different cancers including the majority of pancreatic cancer. However, there are still no approved monoclonal antibody drugs targeting MUC1 have been reported. Recently, we generated a humanized MUC1 antibody (HzMUC1) specific to the interaction region between MUC1-N and MUC1-C. In this study, we generated the antibody drug conjugate (ADC) by conjugating HzMUC1 with monomethyl auristatin (MMAE), and examined the efficacy of HzMUC1-MMAE against the MUC1-positive pancreatic cancer in vitro and in vivo. METHODS: Western blot and immunoprecipitation were used to detect MUC1 in pancreatic cancer cells. MUC1 localization in pancreatic cancer cells was determined by confocal microscopy. HzMUC1 was conjugated with the monomethyl auristatin (MMAE), generating the HzMUC1-MMAE ADC. Colony formation assay and flow cytometry were used to assess the effects of the HzMUC1-MMAE cell viability, cell cycle progression and apoptosis. Capan-2 and CFPAC-1 xenograft model were used to test the efficacy of HzMUC1-MMAE against pancreatic cancer. RESULTS: HzMUC1 antibody binds to MUC1 on the cell surface of pancreatic cancer cells. HzMUC1-MMAE significantly inhibited cell growth by inducing G2/M cell cycle arrest and apoptosis in pancreatic cancer cells. Importantly, HzMUC1-MMAE significantly reduced the growth of pancreatic xenograft tumors by inhibiting cell proliferation and enhancing cell death. CONCLUSION: Our results indicate that HzMUC1-ADC is a promising novel targeted therapy for pancreatic cancer. HzMUC1-ADC should also be an effective drug for the treatment of different MUC1-positive cancers.
RESUMO
Mucin 1 (MUC1) has been regarded as an ideal target for cancer treatment, since it is overexpressed in a variety of different cancers including the majority of breast cancer. However, there are still no approved monoclonal antibody drugs targeting MUC1. In this study, we generated a humanized MUC1 (HzMUC1) antibody from our previously developed MUC1 mouse monoclonal antibody that only recognizes MUC1 on the surface of tumor cells. Furthermore, an antibody-drug conjugate (ADC) was generated by conjugating HzMUC1 with monomethyl auristatin (MMAE), and the efficacy of HzMUC1-MMAE on the MUC1-positive HER2+ breast cancer in vitro and in 'Xenograft' model was tested. Results from western blot analysis and immunoprecipitation revealed that the HzMUC1 antibody did not recognize cell-free MUC1-N in sera from breast cancer patients. Confocal microscopy analysis showed that HzMUC1 antibody bound to MUC1 on the surface of breast cancer cells. Results from mapping experiments suggested that HzMUC1 may recognize an epitope present in the interaction region between MUC1-N and MUC1-C. Results from colony formation assay and flow cytometry demonstrated that HzMUC1-MMAE significantly inhibited cell growth by inducing G2/M cell cycle arrest and apoptosis in trastuzumab-resistant HER2-positive breast cancer cells. Meanwhile, HzMUC1-MMAE significantly reduced the growth of HCC1954 xenograft tumors by inhibiting cell proliferation and enhancing cell death. In conclusion, our results indicate that HzMUC1-ADC is a novel therapeutic drug that can overcome trastuzumab resistance of breast cancer. HzMUC1-ADC should also be an effective therapeutic drug for the treatment of different MUC1-positive cancers in clinic.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Antineoplásicos Imunológicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Imunoconjugados/farmacologia , Mucina-1/metabolismo , Trastuzumab/farmacologia , Animais , Anticorpos Monoclonais Humanizados/metabolismo , Anticorpos Monoclonais Humanizados/uso terapêutico , Antineoplásicos Imunológicos/metabolismo , Antineoplásicos Imunológicos/uso terapêutico , Apoptose/efeitos dos fármacos , Neoplasias da Mama/sangue , Neoplasias da Mama/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/imunologia , Epitopos , Humanos , Imunoconjugados/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mucina-1/sangue , Mucina-1/química , Mucina-1/imunologia , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Oligopeptídeos/uso terapêutico , Receptor ErbB-2/imunologia , Receptor ErbB-2/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cholesterol is a risk factor for breast cancer. However, it is still unclear whether the cholesterol biosynthesis pathway plays any significant role in breast carcinogenesis. 24-Dehydrocholesterol reductase (DHCR24) is a key enzyme in the cholesterol synthesis pathway. Although DHCR24 is reported to have different functions in different cancers, it is not clear whether DHCR24 is involved in breast cancer. In this study, we found that DHCR24 expression was higher in breast cancer especially in luminal and HER2 positive breast cancer tissues compared with normal breast. Changes in DHCR24 expression altered cellular cholesterol content without affecting the adherent growth of breast cancer cells. However, DHCR24 knockdown reduced whereas DHCR24 overexpression enhanced breast cancer stem-like cell populations such as mammosphere and aldehyde dehydrogenase positive cell numbers. In addition, DHCR24 overexpression increased the expression of the Hedgehog pathway-regulated genes. Treating DHCR24 overexpressing breast cancer cell lines with the Hedgehog pathway inhibitor GANT61 blocked DHCR24-induced mammosphere growth and increased mRNA levels of the Hedgehog regulated genes. Furthermore, expression of a constitutively activated mutant of Smoothened, a key hedgehog signal transducer, rescued the decreases in mammosphere growth and Hedgehog regulated gene expression induced by knockdown of DHCR24. These results indicate that DHCR24 promotes the growth of breast cancer stem-like cells in part through enhancing the Hedgehog signaling pathway. Our data suggest that cholesterol contribute to breast carcinogenesis by enhancing Hedgehog signaling and cancer stem-like cell populations. Enzymes including DHCR24 involved in cholesterol biosynthesis should be considered as potential treatment targets for breast cancer.
Assuntos
Neoplasias da Mama/genética , Proteínas Hedgehog/genética , Células-Tronco Neoplásicas/patologia , Proteínas do Tecido Nervoso/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/genética , Transdução de Sinais/genética , Antineoplásicos/farmacologia , Mama/efeitos dos fármacos , Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Colesterol/genética , Feminino , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/genética , Células HEK293 , Humanos , Células MCF-7 , Células-Tronco Neoplásicas/efeitos dos fármacos , Piridinas/farmacologia , Pirimidinas/farmacologia , RNA Mensageiro/genética , Transdução de Sinais/efeitos dos fármacosRESUMO
Scaffolding adaptor Gab2 is overexpressed in a subset of high-grade ovarian cancer. Our published work shows that Gab2 via PI3K enhances migratory behaviors and epithelial to mesenchymal transition (EMT) features of ovarian cancer cells in vitro. However, it is still unclear how Gab2/PI3K pathway reuglates EMT characteristics and whether Gab2 promotes the growth of ovarian cancer stem cell (CSC)-like population and metastatic growth. In this study, we examined the effects of Gab2 expression on CSC-like cell growth using Aldefluor and tumorshpere assays commonly used for assessing ovarian cancer cells with CSC properties. Gab2 overexpression increased the number of ALDH+ cells and tumorsphere formation in two different ovarian cancer cell lines OVCAR5 and OVCAR8, whereas knockdown of Gab2 decreased the number of ALDH+ cells and tumorsphere formation in Caov-3â¯cells. Furthermore, Gab2 promoted metastatic tumor growth of OVCAR5 in nude mice. Mechanistically, we uncovered that Gab2 via PI3K specifically inhibited miR-200c expression. miR-200c downregulation contributed to the Gab2-enhanced cell migratory behaviors, EMT properties, and the expansion of ALDH+ cells and tumorspheres. Furthermore, Gab2 promoted CD44 expression and cell migration/invasion through miR-200c downregulation. Our findings support a model that Gab2-PI3K pathway via miR-200c downregulation promotes CD44 expression, EMT characteristics, and CSC-like cell growth. Therapies involving miR-200c or targeting CD44 should help treat ovarian cancer with high Gab2 expression.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Regulação Neoplásica da Expressão Gênica , MicroRNAs/fisiologia , Proteínas de Neoplasias/fisiologia , Neoplasias Ovarianas/genética , RNA Neoplásico/fisiologia , Animais , Movimento Celular , Regulação para Baixo , Feminino , Xenoenxertos , Humanos , Receptores de Hialuronatos/antagonistas & inibidores , Receptores de Hialuronatos/biossíntese , Receptores de Hialuronatos/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Invasividade Neoplásica , Metástase Neoplásica , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Fosfatidilinositol 3-Quinases/fisiologia , Interferência de RNA , RNA Neoplásico/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologiaRESUMO
Acute myeloid leukemia (AML) is one of the most common types of acute leukemia in adults with the lowest survival rate of all leukemia. Resistance to cytarabine and anthracycline-based chemotherapy is a major cause of treatment failure. Thus, finding new drugs with anti-leukemia activities and minimal side effect is urgently needed. Here through screening more than 1000 drugs approved by the Food and Drug Administration (FDA) of United States, the anthelmintic drug mebendazole (MBZ) was found to inhibit the growth of AML cell lines (THP-1, U937, NB4 and K562) and bone marrow mononuclear cells (BM-MNCs) from AML patients at pharmacologically achievable concentrations. In contrast, similar concentration of MBZ had little inhibitory effect on the growth of normal peripheral blood mononuclear cells (PBMC) or human umbilical vein endothelial cells (HUVEC). In addition, MBZ induced mitotic arrest and mitotic catastrophe in AML cells based on nuclear morphology, cell cycle distribution, mitotic marker analyses and the number of multinucleated cells and apoptotic cells. Furthermore, MBZ treatment inhibited activation of Akt and Erk in AML leukemic cells. Finally, MBZ repressed the progression of leukemic cells in vivo and prolonged survival in AML xenograft mouse model. Taken together, our results suggest that MBZ could be a potential new therapeutic agent for the treatment of AML patients.
Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Leucemia Mieloide Aguda/patologia , Mebendazol/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Células HEK293 , Humanos , Células K562 , Leucócitos Mononucleares/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Células U937 , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Acute myeloid leukemia (AML) is a heterogeneous group of hematopoietic malignancies due to sophisticated genetic mutations and epigenetic dysregulation. MicroRNAs (miRNAs), a class of small non-coding RNAs, are important regulators of gene expression in all biological processes, including leukemogenesis. Recently, miR-375 has been reported to be a suppressive miRNA in multiple types of cancers, but its underlying anti-leukemia activity in AML is largely unknown. METHODS: Quantitative reverse transcriptase PCR (qRT-PCR) was used to measure the expression of miR-375 and HOXB3 in leukemic cells and normal controls. Targets of miR-375 were confirmed by western blot and luciferase assay. Phenotypic effects of miR-375 overexpression and HOXB3 knockdown were assessed using viability (trypan blue exclusion assay), colony formation/replating, as well as tumor xenograft assays in vivo. RESULTS: The expression of miR-375 was substantially decreased in leukemic cell lines and primary AML blasts compared with normal controls, because DNA hypermethylation of precursor-miR-375 (pre-miR-375) promoter was discovered in leukemic cells but not in normal controls. Lower expression of miR-375 predicted poor outcome in AML patients. Furthermore, forced expression of miR-375 not only decreased proliferation and colony formation in leukemic cells but also reduced xenograft tumor size and prolonged the survival time in a leukemia xenograft mouse model. Mechanistically, overexpression of miR-375 reduced HOXB3 expression and repressed the activity of a luciferase reporter through binding 3'-untranslated regions (3'-UTR) of HOXB3 mRNA. Overexpression of HOXB3 partially blocked miR-375-induced arrest of proliferation and reduction of colony number, suggesting that HOXB3 plays an important role in miR-375-induced anti-leukemia activity. Knockdown of HOXB3 by short hairpin RNAs reduced the expression of cell division cycle associated 3 (CDCA3), which decreased cell proliferation. Furthermore, HOXB3 induced DNA methyltransferase 3B (DNMT3B) expression to bind in the pre-miR-375 promoter and enhanced DNA hypermethylation of pre-miR-375, leading to the lower expression of miR-375. CONCLUSIONS: Collectively, we have identified a miR-375-HOXB3-CDCA3/DNMT3B regulatory circuitry which contributes to leukemogenesis and suggests a therapeutic strategy of restoring miR-375 expression in AML.
Assuntos
DNA (Citosina-5-)-Metiltransferases/genética , Regulação Leucêmica da Expressão Gênica , Proteínas de Homeodomínio/genética , Leucemia Mieloide/genética , MicroRNAs/genética , Regiões 3' não Traduzidas/genética , Doença Aguda , Adulto , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , DNA (Citosina-5-)-Metiltransferases/metabolismo , Feminino , Células HL-60 , Proteínas de Homeodomínio/metabolismo , Humanos , Células K562 , Estimativa de Kaplan-Meier , Leucemia Mieloide/metabolismo , Leucemia Mieloide/patologia , Masculino , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Nus , Camundongos SCID , Pessoa de Meia-Idade , Transplante Heterólogo , Adulto Jovem , DNA Metiltransferase 3BRESUMO
SH2-domain containing protein tyrosine phosphatase 1 (Shp1/PTPN6) is mainly expressed in hematopoietic cells and acts a negative signaling regulator. Although Shp1 is also expressed in epithelial cells, the function of shp1 in normal epithelial is still less well understood, especially in regulating the growth of epithelial cells. In this study, different shRNAs and siRNAs against Shp1 were used to knockdown Shp1 expression in MCF10A, an immortalized mammary epithelial cell line. Shp1 knockdown resulted in inhibited cell growth in part due to lower percentage of MCF10A cells entering into S phase and reduced cyclin D1 expression. Accordingly, EGF-induced tyrosyl phosphorylation of EGFR and Stat5 was significantly inhibited in cells with Shp1 knockdown compared with control whereas EGF-induced Akt and Erk phosphorylation was not affected by Shp1 knockdown. Further analysis revealed that Shp1 knockdown lead to decreased EGFR protein expression without affecting EGFR mRNA expression or increasing EGFR protein degradation. Our data indicate that Shp1 functions as a positive regulator and acts in a novel mechanism through promoting EGFR protein expression in mammary epithelial cells.
Assuntos
Receptores ErbB/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 6/metabolismo , Transdução de Sinais , Células Cultivadas , Receptores ErbB/genética , Perfilação da Expressão Gênica , HumanosRESUMO
Honokiol, a constituent of Magnolia officinalis, has been reported to possess potent anti-cancer activity through targeting multiple signaling pathways in numerous malignancies including acute myeloid leukemia (AML). However, the underlying mechanisms remain to be defined. Here, we report that honokiol effectively decreased enzyme activity of histone deacetylases (HDACs) and reduced the protein expression of class I HDACs in leukemic cells. Moreover, treatment with proteasome inhibitor MG132 prevented honokiol-induced degradation of class I HDACs. Importantly, honokiol increased the levels of p21/waf1 and Bax via triggering acetylation of histone in the regions of p21/waf1 and Bax promoter. Honokiol induced apoptosis, decreased activity of HDACs, and significantly inhibited the clonogenic activity of hematopoietic progenitors in bone marrow mononuclear cells from patients with AML. However, honokiol did not decrease the activity of HDACs and induce apoptosis in normal hematopoietic progenitors from unbilicial cord blood. Finally, honokiol dramatically reduced tumorigenicity in a xenograft leukemia model. Collectively, our findings demonstrate that honokiol has anti-leukemia activity through inhibiting HDACs. Thus, being a relative non-toxic agent, honokiol may serve as a novel natural agent for cancer prevention and therapy in leukemia.
Assuntos
Apoptose/efeitos dos fármacos , Compostos de Bifenilo/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Histona Desacetilases/metabolismo , Leucemia Mieloide/tratamento farmacológico , Lignanas/farmacologia , Doença Aguda , Adulto , Idoso , Animais , Biocatálise/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Humanos , Células K562 , Leucemia Mieloide/genética , Leucemia Mieloide/metabolismo , Masculino , Camundongos Nus , Pessoa de Meia-Idade , Complexo de Endopeptidases do Proteassoma/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
Peroxiredoxins (Prxs) are potential therapeutic targets for major diseases such as cancers. However, isotype-specific inhibitors remain to be developed. We report that adenanthin, a diterpenoid isolated from the leaves of Rabdosia adenantha, induces differentiation of acute promyelocytic leukemia (APL) cells. We show that adenanthin directly targets the conserved resolving cysteines of Prx I and Prx II and inhibits their peroxidase activities. Consequently, cellular H(2)O(2) is elevated, leading to the activation of extracellular signal-regulated kinases and increased transcription of CCAAT/enhancer-binding protein ß, which contributes to adenanthin-induced differentiation. Adenanthin induces APL-like cell differentiation, represses tumor growth in vivo and prolongs the survival of mouse APL models that are sensitive and resistant to retinoic acid. Thus, adenanthin can serve as what is to our knowledge the first lead natural compound for the development of Prx I- and Prx II-targeted therapeutic agents, which may represent a promising approach to inducing differentiation of APL cells.
Assuntos
Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Diterpenos do Tipo Caurano/farmacologia , Diterpenos/farmacologia , Leucemia Promielocítica Aguda/tratamento farmacológico , Peroxirredoxinas/antagonistas & inibidores , Animais , Antineoplásicos/química , Proteína beta Intensificadora de Ligação a CCAAT/biossíntese , Cisteína/química , Diterpenos/química , Diterpenos do Tipo Caurano/química , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Peróxido de Hidrogênio/análise , Camundongos , Peroxirredoxinas/química , Tretinoína/farmacologia , Células Tumorais CultivadasRESUMO
Breast cancer is the most common malignant cancer in women. Triple-negative breast cancer (TNBC) has a poorer prognosis than other subtypes and is challenging to treat. MUC1 is a therapeutic target in breast and pancreatic cancer. We developed a novel humanized antibody that specifically binds MUC1 expressed in breast cancer cells and conjugated a humanized MUC1 (HzMUC1) antibody to monomethyl auristatin (MMAE). HzMUC1-MMAE showed an anti-proliferative effect on HER2 positive trastuzumab-resistant breast cancer. Immunoprecipitation indicated that HzMUC1 recognized native MUC1 in TNBC cells. Confocal microscopy showed that HzMUC1 bound MUC1 on the surface of TNBC cells, and the conjugates exhibited the same binding ability to HCC70 as unconjugated HzMUC1 by cell-based ELISA. Treatment of TNBC cells with HzMUC1-MMAE reduced growth of MUC1-positive cells and induced G2/M cell cycle arrest and apoptosis. In a mouse model of breast cancer, HzMUC1-MMAE significantly reduced the growth of tumors established by subcutaneous injection of HCC70 TNBC cells. Therefore, HzMUC1-ADC has therapeutic potential for TNBC.
RESUMO
Background: Cytogenetic abnormalities are considered initiating events in the pathogenesis of multiple myeloma (MM) and are assumed to be of clinical significance. Methods: Fluorescence in situ hybridization (FISH) was used to analyze chromosomal architecture in 101 patients with MM. We evaluated overall patient survival and assessed the cytotoxicity of imatinib against MM cells using a CCK8 assay. Results: ABL gene amplification was detected in 67 patients (66.3%). However, ABL gene amplification was not associated with clinical features, cytogenetic abnormalities (c-Myc amplification, IGH rearrangement, RB1 deletion, p53 deletion, or 1q21 amplification), or overall survival. ABL amplification in MM cell lines (LP-1 and U266) was revealed by FISH. Furthermore, the ABL protein was easily detectable in MM cell lines and some tumor cells by western blotting. A CCK8 assay indicated limited cytotoxicity of imatinib against MM cells. Conclusions: Our study firstly discussed ABL gene amplification was prevalent in MM cells, and we believe that the ABL gene would potentially be a useful target in the treatment of combination strategy for MM with ABL amplification in the future.
RESUMO
Acute myeloid leukemia (AML) is a hematologic malignant disease largely affecting older adults with poor outcomes. Lack of effective targeted treatment is a major challenge in managing the disease in the clinic. Scaffolding adaptor Gab2 is amplified in a subset of AML. However, the causative role of Gab2 in AML remains to be explored. In this study, it was found that Gab2 was expressed at high levels in AML patient samples and AML cell lines. Experiments by knocking down Gab2 expression using shRNA showed that Gab2 promoted AML cell growth and migration in vitro and in vivo. Further studies using Gab2 mutants and pharmacological inhibitors revealed that Gab2 increased CREB phosphorylation via the SHP-2/Erk signaling pathway. CREB phosphorylation contributed to Gab2-induced cell migration by increasing MMP2 and MMP9 expression. This research indicates that high Gab2 expression promotes AML progression through the SHP2-Erk-CREB signaling pathway. CREB suppression may help treat AML with high Gab2 expression.
Assuntos
Leucemia Mieloide Aguda , Metaloproteinase 2 da Matriz , Proteínas Adaptadoras de Transdução de Sinal , Idoso , Humanos , Leucemia Mieloide Aguda/genética , Metaloproteinase 9 da Matriz , RNA Interferente Pequeno/genética , Transdução de SinaisRESUMO
Ikaros is an important transcription factor involved in the development and differentiation of hematopoietic cells. In this work, we found that chemotherapeutic drugs or ultraviolet radiation (UV) treatment could reduce the expression of full-length Ikaros (IK1) protein in less than 3h in leukemic NB4, Kasumi-1 and Jurkat cells, prior to the activation of caspase-3. Etoposide treatment could not alter the mRNA level of IK1 but it could shorten the half-life of IK1. Co-treatment with the proteasome inhibitor MG132 or epoxomicin but not calpain inhibitor calpeptin inhibited etoposide-induced Ikaros downregulation. Overexpression of IK1 could accelerate etoposide-induced apoptosis in NB4 cells, as evidenced by the increase of Annexin V positive cells and the more early activation of caspase 3. To our knowledge, this is the first report to show that upon chemotherapy drugs or UV treatment, IK1 could be degraded via the proteasome system in the early phase of apoptosis induction. These data might shed new insight on the role of IK1 in apoptosis and the post-translational regulation of IK1.
Assuntos
Apoptose , Fator de Transcrição Ikaros/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Caspase 3/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo , Etoposídeo/farmacologia , Humanos , Células Jurkat , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/efeitos da radiação , Ubiquitina/metabolismo , Raios UltravioletaRESUMO
Although endocrine therapies targeting estrogen receptor α (ERα) are effective in managing ER positive (+) breast cancer, many patients have primary resistance or develop resistance to endocrine therapies. In addition, ER+ breast cancer with PIK3CA activating mutations and 11q13-14 amplification have poor survival with unclear mechanism. We uncovered that higher expression of deubiquitinase USP35, located in 11q14.1, was associated with ER+ breast cancer and poor survival. Estrogen enhanced USP35 protein levels by downregulating USP35-targeting miRNA-140-3p and miRNA-26a-5p. USP35 promoted the growth of ER+ breast cancer in vitro and in vivo, and reduced the sensitivity of ER+ breast cancer cells to endocrine therapies such as tamoxifen and fulvestrant. Mechanistically, USP35 enhanced ERα stability by interacting and deubiquitinating ERα, and transcriptional activity of ERα by interacting with ERα in DNA regions containing estrogen response element. In addition, AKT, a key effector of PI3K, phosphorylated USP35 at Serine613, which promoted USP35 nuclear translocation, ERα transcriptional activity, and the growth of ER+ breast cancer cells. Our data indicate that USP35 and ERα form a positive feedback loop in promoting the growth of ER+ breast cancer. USP35 may be a treatment target for ER+ breast cancer with endocrine resistance or with PIK3CA mutations or hyperactivation of the PI3K pathway.
Assuntos
Neoplasias da Mama/genética , Endopeptidases/fisiologia , Estradiol/farmacologia , Receptor alfa de Estrogênio/genética , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Carcinogênese/efeitos dos fármacos , Carcinogênese/genética , Carcinogênese/metabolismo , Endopeptidases/genética , Receptor alfa de Estrogênio/efeitos dos fármacos , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Células MCF-7 , Camundongos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Estabilidade Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
As a subtype of acute myeloid leukemia (AML), acute promyelocytic leukemia (APL) is characterized by a chromosomal translocation, most of which result in the production of a PML-RAR alpha fusion protein. Although the overall survival rate of APL patients has improved dramatically due to all-trans retinoic acid (ATRA) treatment, ATRA-resistance remains a clinical challenge in the management of APL. Therefore, alternative agents should be considered for ATRA-resistant APL patients. Here, we report that antimalaria drug primaquine phosphate (PRQ) exhibits an anti-leukemia effect on both ATRA-sensitive cell line NB4 and ATRA-resistant APL cell lines, NB4-LR2, NB4-LR1, and NB4-MR2. Moreover, PRQ significantly inhibited primary colony formation of untreated or relapsed APL patients. Further study showed that PRQ could induce the apoptosis of APL cells by inhibiting NF-κB signaling pathway. The in vivo study showed that PRQ significantly inhibited NB4-LR2 xenograft tumors growth. These results suggest that PRQ is a potential therapeutic agent for ATRA-resistant APL patients.
Assuntos
Apoptose/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Leucemia Promielocítica Aguda/patologia , NF-kappa B/metabolismo , Primaquina/farmacologia , Transdução de Sinais , Tretinoína/farmacologia , Adulto , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Primaquina/química , Transdução de Sinais/efeitos dos fármacos , Ensaio Tumoral de Célula-Tronco , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
We recently demonstrated that leukocyte Ig-like receptor 4 (LILRB4) expressed by monocytic acute myeloid leukemia (AML) cells mediates T-cell inhibition and leukemia cell infiltration via its intracellular domain. The cytoplasmic domain of LILRB4 contains three immunoreceptor tyrosine-based inhibitory motifs (ITIMs); the tyrosines at positions 360, 412, and 442 are phosphorylation sites. Here, we analyzed how the ITIMs of LILRB4 in AML cells mediate its function. Our in vitro and in vivo data show that Y412 and Y442, but not Y360, of LILRB4 are required for T-cell inhibition, and all three ITIMs are needed for leukemia cell infiltration. We constructed chimeric proteins containing the extracellular domain of LILRB4 and the intracellular domain of LILRB1 and vice versa. The intracellular domain of LILRB4, but not that of LILRB1, mediates T-cell suppression and AML cell migration. Our studies thus defined the unique signaling roles of LILRB4 ITIMs in AML cells.
Assuntos
Movimento Celular/imunologia , Tolerância Imunológica , Glicoproteínas de Membrana/imunologia , Proteínas de Neoplasias/imunologia , Receptores Imunológicos/imunologia , Linfócitos T/imunologia , Motivos de Aminoácidos , Animais , Movimento Celular/genética , Humanos , Leucemia Mieloide Aguda , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Proteínas de Neoplasias/genética , Receptores Imunológicos/genética , Linfócitos T/patologia , Células THP-1RESUMO
Metastasis is responsible for the death of most breast cancer patients. Robo1 has been implicated as a tumor suppressor for various cancers including breast cancer. However, it is not well understood how Robo1 expression is regulated during tumorigenesis. In this study, we uncovered that the transmembrane proline rich γ-carboxyglutamic acid protein 4 (PRRG4) promotes breast cancer metastasis by downregulating Robo1. Analysis of mRNA expression data in The Cancer Genome Atlas and immunohistochemistry assay on breast tumor samples showed that PRRG4 expression was higher in breast tumors than in normal breast tissues. Experiments with PRRG4 knockdown and overexpression revealed that PRRG4 promoted migration and invasion of breast cancer cells, and enhanced metastasis in an experimental metastasis model. Mechanistically, we found that PRRG4 via its LPSY and PPPY motifs recruited the E3 ubiquitin ligase NEDD4, which induced ubiquitination and degradation of Robo1, thus contributing to migration and invasion of breast cancer cells. In addition, PRRG4 interacted with and enhanced protein tyrosine kinase Src and FAK activation. Overall, our data support a model that PRRG4 via NEDD4 downregulates the Robo1, resulting in the activation of Src and FAK and promoting breast cancer metastasis. PRRG4 may be a novel target for treating metastatic breast cancer.