Transcriptomics

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HER2-positive breast cancer cells resistant to trastuzumab and lapatinib lose reliance upon HER2 and are sensitive to the multitargeted kinase inhibitor sorafenib


ABSTRACT: HER2 targeting with trastuzumab has changed the prognosis of breast cancer patients carrying amplification and/or overexpression of this oncogene. Despite this progress, however, resistance to trastuzumab occurs in the vast majority of patients. Newer anti-HER2 therapies, like the dual tyrosine-kinase inhibitor (TKI) lapatinib, show antitumor activity in a limited proportion of patients, indicating that HER2 can be still exploited as a target after trastuzumab failure. However, due to the high proportion of patients that fail to respond to these alternative strategies, it is reasonable to assume that cells escaping HER2 targeting may rely on alternative pathways not involving HER2 to sustain their growth. Their knowledge is relevant for exploiting new therapies. To investigate this hypothesis we generated a human HER2 overexpressing breast cancer cell line resistant to trastuzumab and lapatinib (T100) and we have characterized it genetically and molecularly. In T100 cells, the previously proposed mechanisms of resistance to HER2 inhibitors did not explain cell escape. Notably, silencing HER2 by shRNA did not affect the growth of T100 cells, suggesting loss of reliance upon HER2. Among the alternative pathways that we explored, altered levels of the antiapoptoptic proteins Mcl-1 and Survivin were observed. This suggested the possibility of targeting resistant cells with the multitarget inhibitor sorafenib. Moreover, sorafenib, nearly inactive in parental cells, strongly inhibited the in vitro growth of T100 cells. In conclusion, we provide a new model where the activation of HER2 independent mechanisms sustains the growth of tumor cells, significantly increasing the sensitivity to sorafenib. Keywords: HER2, breast cancer, resistance, trastuzumab, lapatinib, sorafenib Total RNA was isolated using TRIZOL reagent (Life Technologies, Gathersburg, MD) and quantified by Bioanalyzer 2100 (Agilent Technologies); following the isolation procedure, mRNA was amplified starting from 1μg using MessageAmp II aRNA Amplification kit (Ambion Inc. Austin TX, USA). Ammino-allyl modified nucleotides were incorporated in in vitro transcription step according to the manufacturer’s protocol. Labeling was performed using NHS (N-hydroxysuccinimidyl) ester Cy3 or Cy5 dies (GE Healthcare Europe GMBH, Upsala - Sweden) able to react with the modified RNA. At least 5 μg of mRNA, for each sample, were labeled and purified with columns respectively. The Dye-Swap replication procedure was applied, in order to increase the accuracy of results. Samples were hybridized on 44K glass arrays (Agilent Technologies). Arrays were scanned and the images obtained were analyzed by the Feature Extraction software Agilent (version 9.5) and the text files were then processed using the Bioconductor package Limma (Linear models for microarray analysis).

ORGANISM(S): Homo sapiens  

SUBMITTER: Giorgio Valabrega  caterina Peraldo Neia   Andrea Milani   Filippo Montemurro   Giorgia Migliardi   Giuliana Cavalloni   Caterina Peraldo-Neia   Annalisa Petrelli   Gianluca Zaccarello   Loretta Gammaitoni   Aldo Moggio   Francesco Favero   Sonia Capellero   Massimo Aglietta   Silvia Giordano    

PROVIDER: E-GEOD-17630 | ArrayExpress | 2010-12-15

SECONDARY ACCESSION(S): GSE17630PRJNA118621

REPOSITORIES: GEO, ArrayExpress

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HER2-positive breast cancer cells resistant to trastuzumab and lapatinib lose reliance upon HER2 and are sensitive to the multitargeted kinase inhibitor sorafenib.

Valabrega Giorgio G   Capellero Sonia S   Cavalloni Giuliana G   Zaccarello Gianluca G   Petrelli Annalisa A   Migliardi Giorgia G   Milani Andrea A   Peraldo-Neia Caterina C   Gammaitoni Loretta L   Sapino Anna A   Pecchioni Carla C   Moggio Aldo A   Giordano Silvia S   Aglietta Massimo M   Montemurro Filippo F  

Breast cancer research and treatment 20101209 1


Trastuzumab has changed the prognosis of HER2 positive breast cancers. Despite this progress, resistance to trastuzumab occurs in most patients. Newer anti-HER2 therapies, like the dual tyrosine-kinase inhibitor (TKI) lapatinib, show significant antitumor activity, indicating that HER2 can be still exploited as a target after trastuzumab failure. However, since a high proportion of patients fail to respond to these alternative strategies, it is possible that cell escape from HER2 targeting may r  ...[more]

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