Original article
Amplification and expression of genes from the 17q11∼q12 amplicon in breast cancer cells

https://doi.org/10.1016/S0165-4608(01)00657-4Get rights and content

Abstract

Amplification of a portion of 17q11∼q12 involving the human epidermal growth factor receptor-2 (HER-2/neu) gene is common in solid tumors. HER-2/neu amplification in breast cancer is associated with a poor prognosis and may predict response to therapeutic interventions. The molecular complexity and informative content of the 17q11∼q12 amplified DNA remain largely unknown. Study of available sequence information in public database allowed us to construct a contig in bacterial artificial chromosome that covers this region. We have identified a several hundred kilobase core segment from this region that is amplified in three out of three breast cancer cell lines with HER-2/neu amplification. There is striking correlation between amplification and overexpression of genes from this region. Amplified and overexpressed genes, in addition to HER-2/neu, may play functional roles in the pathogenesis of breast cancer and may serve as additional targets for therapy. Published by Elsevier Science Inc.

Introduction

Gene amplification is common in solid tumors. It is typically associated with tumor progression and development of drug resistance [1]. Amplification of a segment of chromosome 17q11∼q12 involving the human epidermal growth factor receptor-2 (HER-2/neu) gene is present in a variety of solid tumors including breast cancer [2], ovarian cancer [3], endometrial cancer [4], prostate cancer [5], head and neck cancer [6], gastric, and esophageal cancer 7, 8, 9, 10, 11. Breast cancer with HER-2/neu amplification, found in 25–30% of the cases, is a more aggressive disease as it predicts shortened disease free and overall survival 12, 13, 14. Breast cancer with HER-2/neu amplification may respond differently to chemotherapy 12, 13, 14. Despite this obvious clinical implication, the informative content or the expression profile for genes in the chromosome 17q11∼q12 amplified region has not been well characterized. Overexpression of the HER-2/neu gene is thought to be responsible for many of the phenotypes associated with 17q11∼q12 amplification. Additionally, coamplified genes may contribute as well. Recent reports show that breast cancers with both HER-2/neu overexpression and amplification exhibit more aggressive biological characteristics than those with overexpression only 15, 16. Though other explanation is possible, these findings support the notion that additional genes in 17q11∼q12 contribute to the biological behavior of HER-2/neu amplified breast cancer. The topoisomerase 2A (Topo-2A) gene, which is amplified and overexpressed in a subset of breast cancer with HER-2/neu amplification, may be one such example [17]. Topo-2A is a target for anthracycline. Overexpression of Topo-2A may be the basis for increased sensitivity of 17q11∼q12 amplified breast cancer to anthracycline.

In this study, we report the assembly of 17q11∼q12 amplicon in an overlapping set of bacterial artificial chromosomes (BAC) [18]. Southern blot analysis was employed to identify the core sequence from this region that was commonly amplified from different breast cancer cell lines. The expression profile of resident genes from the commonly amplified region shared among different breast cancer cell lines was systematically examined as well.

Section snippets

BAC and plasmid constructs

BAC and plasmid clones used in this study are obtained from Research Genetics (Huntsville, AL, USA). BAC#131 has been sequenced in its entirety (Genbank accession number AC005288). The following BAC have only been sequenced in draft: BAC#413 (AC007776), BAC#387 (AC019095), BAC#94 (AC079199), BAC#62 (AC025531), BAC#2019 (AC040933), and BAC#390 (AC009283). Two BAC have been end sequenced: BAC#1044 (AQ663094 and AQ663011), and BAC#98 (AQ320284 and AQ320287). The identities and overlap of BAC have

Assembly of a 17q11∼q12 contig in overlapping BAC

Study of the sequence information available in the public database allows us to obtain nine BAC to assemble the 17q11∼q12 contig (Fig. 1A). These BAC, have been previously sequenced in their entirety or partially. BAC#413, #387, #94, #2019 and #62 share large stretches of sequences in common. The same applies to BAC#390 and #131. The two blocks of sequences are bridged together by BAC#98 that has been end sequenced. The alignment of these BAC has been verified by multiple PCR assays based upon

Discussion

We have performed a systematic analysis of the chromosome 17q11∼q12 amplified DNA in breast cancer cell lines. We identified a commonly amplified region that was several hundred kbs in size (>300 kb) shared among different breast cancer cell lines. Overexpression of amplified genes from this region was demonstrated by multiplex RT-PCR. Our observation supports the notion that multiple amplified genes, in addition to HER-2/neu, may contribute to biological behaviors of breast cancers and confer

Acknowledgements

Supported in part by Women's Health Program Pilot Grant Awards, University Research Council Faculty Research Support Program, and Ruth Lyons/Research Challenge Fund at University of Cincinnati.

The author thanks Dr. Dennis J. Slamon for his mentorship, Drs. David C. Page, John C. Winkelmann and Jeff Knauf for review of the manuscript, Dr. Ling-Pai Ting for her continued support during the study and Rashimi Tripathi for technical assistance.

References (31)

  • M.F. Press et al.

    Amplification and overexpression of HER-2/neu in carcinomas of the salivary glandcorrelation with poor prognosis

    Cancer Res

    (1994)
  • T. Ishikawa et al.

    Amplification of the c-erbB-2 (HER-2/neu) gene in gastric cancer cells. Detection by fluorescence in situ hybridization

    Am J Pathol

    (1997)
  • M. Nakajima et al.

    The prognostic significance of amplification and overexpression of c-met and c-erb B-2 in human gastric carcinomas

    Cancer

    (1999)
  • M. Nessling et al.

    Mapping of chromosomal imbalances in gastric adenocarcinoma revealed amplified protooncogenes MYCN, MET, WNT2, and ERBB2

    Genes Chromosomes Cancer

    (1998)
  • H. Yamauchi et al.

    When is a tumor marker ready for prime time? A case study of c-erbB-2 as a predictive factor in breast cancer

    J Clin Oncol

    (2001)
  • Cited by (0)

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