Technical Notes
Her2/ Neu gene amplification
Clinical Significance:
HER-2/neu is a proto-oncogene localized on chromosome 17 that encodes a transmembrane tyrosine kinase human
epidermal growth factor receptor protein. Overexpression of HER-2/neu protein or HER-2/neu gene amplification
is seen in 10-33% of breast cancers and at a lower frequency in a number of other solid tumor types
(1, 2).
Ligand binding to the receptor complex on the cell surface leads to activation of intrinsic protein tyrosine kinase activity. This triggers a cascade of events leading to gene activation resulting in mitogenic stimulation. The HER-2/neu receptor, therefore, plays a key role in cellular growth. It is generally accepted that there is a significant correlation between a positive HER-2/neu status and an adverse clinical outcome in node positive tumors (3).
There is still controversy concerning its prognostic significance in node negative patients. Clinical studies have shown an apparent resistance of tumors with a positive HER-2/neu status to hormone therapy alone and a strong interaction between HER-2/neu overexpression and chemosensitivity to some chemotherapy regimes (4).
Methodology:
Minor groove binder (MGB) with Polymerase Chain Reaction (PCR) has been utilized to develop a quantitative,
Real Time PCR based assay for the ultrasensitive detection and quantitation of HER-2/ Neu gene amplification
(5).
Patient DNA would be isolated, purified, and subjected to PCR amplification using oligonucleotide primers and Taqman probe. An additional PCR amplification directed at a GAPDH gene segment would be performed as a control for sample DNA quality.
The assay has been validated by a positive control reaction using DNA from breast cancer cell lines and from breast carcinomas.
The assay has been validated by a positive control reaction using RNA of confirmed AML-M3 patients.
References:
1) Gelmini S., Orlando C., Sestini R. et al (1997) Clin. Chem. 43 (5), 752.2) Chiang P.W., Beer D.G., Wei W.L. et al (1999) Clin. Cancer Res. Jun 5 (6), 1381.
3) Bieche I, Olivi M., Champeme M.H. et al (1998) Int J. Cancer 78 (5), 661.
4) Bieche I., Onody P., Laurendeau I. et al (1999) Clin. Chem. Aug 45 (8), 1148.
5) Lehmann U., Glockner S., Kleeberger W. et al (2000) Am. J. Pathol. 156 (6), 1855.