Microbead arrays for the analysis of erbb receptor tyrosine kinase activation and dimerization in breast cancer cells

Imran H. Khan, Jing Zhao, Paramita Ghosh, Melanie Ziman, Colleen Sweeney, Hsing Jien Kung, Paul A. Luciw

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Receptor tyrosine kinases (RTKs) in the ErbB family (EGFR, ErbB2, ErbB3, and ErbB4) are implicated in a variety of human malignancies. Accordingly, determination of both expression and activation (dimerization/heterodimerization and phosphorylation) of ErbB proteins is critical in defining their functional role in cancer. Efficient and comprehensive methods to study molecular functions of ErbB family of RTKs are needed not only for improvements in diagnostics but also for early screening of targeted drugs (eg, small molecule inhibitors and therapeutic antibodies). We report development of 3 multiplex microbead immunoassays for simultaneous detection of expression, protein-protein interactions, and phosphorylation of these RTKs. These novel multiplex immunoassays were used to study ErbB RTKs under different cell activation conditions in 2 breast cancer cell lines (MDA-MB-453 and MDA-MB-468) and an epidermoid cancer cell line (A431). The results were confirmed by immunoprecipitation/western blot. Importantly, the multiplex immunoassay facilitated time-course studies in these cell lines after cell activation with EGF and neuregulin, revealing the kinetics of phosphorylation of the ErbB family RTKs. This study demonstrates the utility of the Luminex® multiplex system as an efficient and comprehensive approach to study different aspects of molecular roles of these RTKs. Importantly, the study provides proof-of-concept for the utility of the multiplex microbead immunoassay approach for potential use in efficient, robust, and rapid screening of drugs, particularly those targeting functional aspects of these potent signaling molecules. In addition, the assays described here may be useful for cancer diagnostics and monitoring efficacy of therapy targeting the ErbB family of RTKs.

Original languageEnglish
Pages (from-to)27-36
Number of pages10
JournalAssay and Drug Development Technologies
Volume8
Issue number1
DOIs
Publication statusPublished - Feb 1 2010
Externally publishedYes

Fingerprint

Dimerization
Microspheres
Immunoassay
Protein-Tyrosine Kinases
Receptor Protein-Tyrosine Kinases
Breast Neoplasms
Preclinical Drug Evaluations
Phosphorylation
Cell Line
Neoplasms
Neuregulins
Proteins
Immunoprecipitation
Epidermal Growth Factor
Western Blotting
ErbB Receptors
Antibodies
Therapeutics

ASJC Scopus subject areas

  • Molecular Medicine
  • Drug Discovery

Cite this

Microbead arrays for the analysis of erbb receptor tyrosine kinase activation and dimerization in breast cancer cells. / Khan, Imran H.; Zhao, Jing; Ghosh, Paramita; Ziman, Melanie; Sweeney, Colleen; Kung, Hsing Jien; Luciw, Paul A.

In: Assay and Drug Development Technologies, Vol. 8, No. 1, 01.02.2010, p. 27-36.

Research output: Contribution to journalArticle

Khan, Imran H. ; Zhao, Jing ; Ghosh, Paramita ; Ziman, Melanie ; Sweeney, Colleen ; Kung, Hsing Jien ; Luciw, Paul A. / Microbead arrays for the analysis of erbb receptor tyrosine kinase activation and dimerization in breast cancer cells. In: Assay and Drug Development Technologies. 2010 ; Vol. 8, No. 1. pp. 27-36.
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