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author:("mile, L")
1.  Crosstalk between PKCα and Notch-4 in endocrine-resistant breast cancer cells 
Oncogenesis  2013;2(8):e60-.
The Notch pathway is functionally important in breast cancer. Notch-1 has been reported to maintain an estrogen-independent phenotype in estrogen receptor α (ERα)+ breast cancer cells. Notch-4 expression correlates with Ki67. Notch-4 also plays a key role in breast cancer stem-like cells. Estrogen-independent breast cancer cell lines have higher Notch activity than estrogen-dependent lines. Protein kinase Cα (PKCα) overexpression is common in endocrine-resistant breast cancers and promotes tamoxifen (TAM)-resistant growth in breast cancer cell lines. We tested whether PKCα overexpression affects Notch activity and whether Notch signaling contributes to endocrine resistance in PKCα-overexpressing breast cancer cells.Analysis of published microarray data from ERα+ breast carcinomas shows that PKCα expression correlates strongly with Notch-4. Real-time reverse transcription PCR and immunohistochemistry on archival specimens confirmed this finding. In a PKCα-overexpressing, TAM-resistant T47D model, PKCα selectively increases Notch-4, but not Notch-1, expression in vitro and in vivo. This effect is mediated by activator protein-1 (AP-1) occupancy of the Notch-4 promoter. Notch-4 knockdown inhibits estrogen-independent growth of PKCα-overexpressing T47D cells, whereas Notch-4IC expression stimulates it. Gene expression profiling shows that multiple genes and pathways associated with endocrine resistance are induced in Notch-4IC- and PKCα-expressing T47D cells. In PKCα-overexpressing T47D xenografts, an orally active γ-secretase inhibitor at clinically relevant doses significantly decreased estrogen-independent tumor growth, alone and in combination with TAM. In conclusion, PKCα overexpression induces Notch-4 through AP-1. Notch-4 promotes estrogen-independent, TAM-resistant growth and activates multiple pathways connected with endocrine resistance and chemoresistance. Notch inhibitors should be clinically evaluated in PKCα- and Notch-4-overexpressing, endocrine-resistant breast cancers.
PMCID: PMC3759125  PMID: 23917222
Notch; breast cancer; endocrine therapy; PKCα
2.  Targeting both Notch and ErbB-2 signalling pathways is required for prevention of ErbB-2-positive breast tumour recurrence 
British Journal of Cancer  2011;105(6):796-806.
We reported that Notch-1, a potent breast oncogene, is activated in response to trastuzumab and contributes to trastuzumab resistance in vitro. We sought to determine the preclinical benefit of combining a Notch inhibitor (γ-secretase inhibitor (GSI)) and trastuzumab in both trastuzumab-sensitive and trastuzumab-resistant, ErbB-2-positive, BT474 breast tumours in vivo. We also studied if the combination therapy of lapatinib plus GSI can induce tumour regression of ErbB-2-positive breast cancer.
We generated orthotopic breast tumour xenografts from trastuzumab- or lapatinib-sensitive and trastuzumab-resistant BT474 cells. We investigated the antitumour activities of two distinct GSIs, LY 411 575 and MRK-003, in vivo.
Our findings showed that combining trastuzumab plus a GSI completely prevented (MRK-003 GSI) or significantly reduced (LY 411 575 GSI) breast tumour recurrence post-trastuzumab treatment in sensitive tumours. Moreover, combining lapatinib plus MRK-003 GSI showed significant reduction of tumour growth. Furthermore, a GSI partially reversed trastuzumab resistance in resistant tumours.
Our data suggest that a combined inhibition of Notch and ErbB-2 signalling pathways could decrease recurrence rates for ErbB-2-positive breast tumours and may be beneficial in the treatment of recurrent trastuzumab-resistant disease.
PMCID: PMC3171020  PMID: 21847123
ErbB-2; trastuzumab; Notch-1; GSI; recurrence; resistance
3.  Inhibition of Notch3 signalling induces rhabdomyosarcoma cell differentiation promoting p38 phosphorylation and p21Cip1 expression and hampers tumour cell growth in vitro and in vivo 
Cell Death and Differentiation  2011;19(5):871-881.
Rhabdomyosarcoma (RMS) is a paediatric soft-tissue sarcoma arising from skeletal muscle precursors coexpressing markers of proliferation and differentiation. Inducers of myogenic differentiation suppress RMS tumourigenic phenotype. The Notch target gene HES1 is upregulated in RMS and prevents tumour cell differentiation in a Notch-dependent manner. However, Notch receptors regulating this phenomenon are unknown. In agreement with data in RMS primary tumours, we show here that the Notch3 receptor is overexpressed in RMS cell lines versus normal myoblasts. Notch3-targeted downregulation in RMS cells induces hyper-phosphorylation of p38 and Akt essential for myogenesis, resulting in the differentiation of tumour cells into multinucleated myotubes expressing Myosin Heavy Chain. These phenomena are associated to a marked decrease in HES1 expression, an increase in p21Cip1 level and the accumulation of RMS cells in the G1 phase. HES1-forced overexpression in RMS cells reverses, at least in part, the pro-differentiative effects of Notch3 downregulation. Notch3 depletion also reduces the tumourigenic potential of RMS cells both in vitro and in vivo. These results indicate that downregulation of Notch3 is sufficient to force RMS cells into completing a correct full myogenic program providing evidence that it contributes, partially through HES1 sustained expression, to their malignant phenotype. Moreover, they suggest Notch3 as a novel potential target in human RMS.
PMCID: PMC3321627  PMID: 22117196
Notch; rhabdomyosarcoma; differentiation; muscle cells; sarcoma
4.  Prognostic factors for survival in patients with early-intermediate hepatocellular carcinoma undergoing non-surgical therapy: comparison of Okuda, CLIP, and BCLC staging systems in a single Italian centre 
Gut  2005;54(3):411-418.
Background: Several prognostic models have been developed to stage hepatocellular carcinoma (HCC) but there is no general consensus on which is the most reliable. We compared three prognostic indices (Okuda, CLIP, and BCLC scoring systems) in a large series of cirrhotic patients with HCC undergoing non-surgical treatment in terms of their ability to classify patients into different risk groups
Methods: We retrospectively studied 268 Italian patients with HCC. A total of 146 patients were treated with ablation, 132 with percutaneous ethanol injection, and 14 with radiofrequency ablation; 103 underwent transcatheter arterial chemoembolisation and 19 had supportive care alone. Factors determining survival were analysed by univariate and multivariate analysis using the Kaplan-Meier method and Cox proportional hazard regression models. Okuda, CLIP, and BCLC scores evaluated before treatment were applied.
Results: Median survival was 25.7 months. In a multivariate analysis, portal vein thrombosis, α fetoprotein, total bilirubin, and tumour size were significant predictors of survival. Okuda, CLIP, and BCLC scores were all able to predict survival (p<0.001). They identified two, four, and six risk groups, respectively, with a median survival ranging from 27 to 19 months for Okuda, 30 to 5 months for CLIP, and 43 to 7 months for BCLC.
Conclusions: Both CLIP and BCLC scores were more effective than the Okuda score in stratifying patients into different risk groups with early-intermediate HCC. However, the BCLC scoring system gave a better prediction of prognosis in patients with disease diagnosis at a very early stage.
PMCID: PMC1774422  PMID: 15710992
hepatocellular carcinoma; prognostic score; CLIP score; BCLC staging system; Okuda staging; staging systems; liver cirrhosis; survival
5.  Is coeliac disease a confounding factor in the diagnosis of NASH? 
Gut  2001;49(4):596.
PMCID: PMC1728483  PMID: 11589191
7.  Sporadic HEV hepatitis in Italy 
Gut  2001;48(4):580.
PMCID: PMC1728220  PMID: 11288737
9.  Tissue-specific expression of the gene coding for human Clara cell 10-kD protein, a phospholipase A2-inhibitory protein. 
Journal of Clinical Investigation  1993;92(5):2099-2109.
Clara cell 10-kD protein (cc10kD), a secretory phospholipase A2 inhibitor, is suggested to be the human counterpart of rabbit uteroglobin (UG). Because cc10kD is expressed constitutively at a very high level in the human respiratory epithelium, the 5' region of its gene may be useful in achieving organ-specific expression of recombinant DNA in gene therapy of diseases such as cystic fibrosis. However, it is important to establish the tissue-specific expression of this gene before designing gene transfer experiments. Since the UG gene in the rabbit is expressed in many other organs besides the lung and the endometrium, we investigated the organ and tissue specificity of human cc10kD gene expression using polymerase chain reaction, nucleotide sequence analysis, immunofluorescence, and Northern blotting. Our results indicate that, in addition to the lung, cc10kD is expressed in several nonrespiratory organs, with a distribution pattern very similar, if not identical, to that of UG in the rabbit. These results underscore the necessity for more detailed analyses of the 5' region of the human cc10kD gene before its usefulness in gene therapy could be fully assessed. These data also suggest that cc10kD and UG may have similar physiological function(s).
PMCID: PMC288387  PMID: 8227325

Results 1-9 (9)