Epidermal growth factor and transforming growth factor alpha mRNA expression in human breast cancer biopsies; analysis in relation to estradiol, progesterone and EGF receptor content.

Among the peptide growth factors active in breast glandular cell proliferation epidermal growth factor (EGF) and transforming growth factor alpha (TGF alpha) are thought to play a major role in tumour development. They operate through binding to and activation of a common membrane receptor, defined as EGF-R. Their production is modulated by hormones and local growth factors. After it was shown by previous investigation in this laboratory that EGF-R could be detected in 90% of the tumours, but was masked by endogenous ligand in 36% of them, the question was raised as to the level of the ligand's expression in tumour tissue biopsies. Therefore, we investigated the expression of EGF and TGF alpha mRNA in 146 breast cancer biopsies by slot blot analysis using specific 32P-labelled probes. The data were correlated with sex steroids and EGF receptor content. Our results showed that EGF and TGF alpha coexisted in all tumour samples, and that their level of mRNA expression was similar in half of the tumours. Northern blot and polymerase chain reaction (PCR) analysis validated these findings. A significant direct correlation was found between the level of TGF alpha/EGF mRNA expression and the ER/progesterone receptor (PGR) content. TGF alpha and EGF mRNA levels were significantly higher in ER+ (P = 0.0015 and P = 0.0001, respectively) and in PGR+ tumours (P < 0.005 and P = 0.0001) than in their negative counterparts. Moreover, TGF alpha mRNA expression negatively correlated with the number of EGF-R binding sites measured by the standard method (P = 0.02), and it was significantly related to the number of sites occupied by endogenous ligand. In conclusion, it was shown that TGF alpha and EGF mRNA were coexpressed in all the tumour biopsies tested and that their level was higher in the hormone receptor positive than in negative samples. The correlation between the presence of ER/PGR sites, high level of TGF alpha/EGF mRNA and EGF-R occupancy by endogenous ligand is in favour of ER mediated control of TGF alpha and EGF production.

Analysis of epidermal growth factor receptor mRNA expression by polymerase chain reaction assay in 94 human breast adenocarcinoma tumors.

It is well known that breast cancer cells can synthesize and secrete various growth factors that are able to stimulate tumor growth through autocrine and/or paracrine mechanisms. EGF is one of these growth factors involved in normal breast epithelial development and tumor proliferation. EGF and TGF alpha (EGF-like peptide) are produced in variable amounts and both bind to the EGF receptor (EGF-R). Previous investigation in the laboratory measuring free and occupied EGF-R sites by differential ligand binding assays had demonstrated that non-occupied and total binding sites were present in 54 and 90% of 216 breast tumor biopsies respectively. EGF-R appeared to be totally masked by endogenous ligand in 40 and 21% of estrogen receptor positive and negative tumors respectively. The aim of the present study was to check by a molecular method the expression of the EGF-R gene. The PCR method was applied to 94 tumor samples of the previous series. Total RNA was treated with 0.5 units of Rnase-free Dnase/mg of RNA to remove any contaminating DNA. We simultaneously reverse transcribed and amplified another transcript (beta-actin) as an internal standard. Both signals were present in 88 of the 94 samples while the presence of EGF-R was detected in 74 of them when assessed by radioligand assay. The findings indicate that 93% of the tumors analysed in this series expressed EGF-R mRNA, in agreement with our previous data on occupied EGF-R sites, i.e. two-fold more than by using the standard binding assay. No significant correlation was observed between the expression of the EGF-R gene and the estrogen receptor content.