Demonstration of ligand-dependent signaling by the erbB-3 tyrosine kinase and its constitutive activation in human breast tumor cells.

Abstract

The predicted human erbB-3 gene product is closely related to epidermal growth factor receptor (EGFR) and erbB-2, which have been implicated as oncogenes in model systems and human neoplasia. We expressed the erbB-3 coding sequence in NIH 3T3 fibroblasts and identified its product as a 180-kDa glycoprotein, gplsOerbB-3. Tunicamycin and pulsechase experiments revealed that the mature protein was processed by N-linked glycosylation of a 145-kDa erbB-3 core polypeptide. The intrinsic catalytic function of gpl8OerbB3 was shown by its ability to autophosphorylate in vitro. Liganddependent signaling of its cytoplasmic domain was established employing transfectants that express a chimeric EGFR/erbB-3 protein, gplSOI FR/erbB-3* EGF induced tyrosine phosphorylation of the chimera and promoted soft agar colony formation of such transfectants. These fimdings combined with the detection of constitutive tyrosine phosphorylation of gpl8OerbB3 in 4 of 12 human mammary tumor cell lines implicate the activated erbB-3 product in the pathogenesis of some human malignancies.

c-erbB3 and c-erbB4 expression is a feature of the endocrine responsive phenotype in clinical breast cancer

“…As for the mRNA, status levels of c-erbB3 protein were found to be in agreement with those reported in the literature as positive (ZR75.1, MCF-7, MDA 453; Lemoine et al, 1992;Kraus et al, 1993) or negative (MDA 231, A431 Rajkumar and Gullick, 1994) respectively for c-erbB3 protein (not illustrated). Having thus standardized the conditions of the assay, an immunocytochemical analysis of c-erbB3 in group 2 primary breast cancer specimens revealed membrane immunostaining in 84% specimens.…”

“…The developed RT ± PCR procedure for the detection of c-erbB3 and c-erbB4 mRNA was tested further using a small sub-group of breast tumour samples and was found to be highly reproducible, with a mean densitometry variance of only *4% (not illustrated). Furthermore, analysis of c-erbB3 mRNA expression by RT ± PCR for a series of breast cancer cell lines was in concurrence with the literature, both for those reported as positive (ZR75.1, MCF-7, MDA 453; Lemoine et al, 1992;Kraus et al, 1993;Chan et al, 1995) and negative (MDA 231, A431;Rajkumar and Gullick, 1994) respectively for c-erbB3 mRNA and protein (not illustrated).…”

Immune responses to all ErbB family receptors detectable in serum of cancer patients

“…These transfectants have previously been characterized and are known to express individual ErbB receptors in the range of 0.6 ± 3610 6 molecules/cell (Di Fiore et al, 1987b;Kraus et al, 1993;Fedi et al, 1994;. A recombinant model system expressing high receptor levels was utilized, to facilitate sensitive and speci®c detection of single ErbB family members, as frequent co-expression of ErbB receptors in human tumor cell lines may confound de®nition of a receptorspeci®c response.…”

“…A recombinant model system expressing high receptor levels was utilized, to facilitate sensitive and speci®c detection of single ErbB family members, as frequent co-expression of ErbB receptors in human tumor cell lines may confound de®nition of a receptorspeci®c response. Figure 1 illustrates detection of EGFR, ErbB2, ErbB3 or ErbB4 in the 170-185 kDa region of the corresponding transfectant by receptorspeci®c polyclonal rabbit antisera raised against carboxyl-terminal epitopes of single ErbB receptors (Fedi et al, 1994;Alimandi et al, 1995;Kraus et al, 1993;Baulida et al, 1996). Overexpression of the respective recombinant human ErbB receptor was validated in LTR-EGFR, LTR-ErbB2, LTR-ErbB3, or LTR-ErbB4 when compared with NIH3T3 controls (Figure 1).…”

Activation of Src in human breast tumor cell lines: elevated levels of phosphotyrosine phosphatase activity that preferentially recognizes the Src carboxy terminal negative regulatory tyrosine 530

“…Src has also been shown to associate in vitro and in vivo with activated forms of the epidermal growth factor receptor (EGFR) and Neu/HER2 receptor (Luttrell et al, 1994;Maa et al, 1995;Oude Weernink et al, 1994;Stover et al, 1995), both of which have been correlated with poor prognosis in the 20 ± 30% of breast cancer cases in which these receptors are found to be ampli®ed or overexpressed (Berger et al, 1988;Kraus et al, 1993;Slamon et al, 1987Slamon et al, , 1989Varley et al, 1987). Overexpression of the neu proto-oncogene in the mammary glands of transgenic mice has been shown to cause tumors, following a latency period of 200 ± 400 days (Guy et al, 1992).…”