Both homo-and hetero-dimers of ErbB receptor tyrosine kinases mediate signaling by a large group of epidermal growth factor (EGF)-like ligands. However, some ligands are more potent than others, although they bind to the same direct receptor. In addition, signaling by receptor heterodimers is superior to homodimers. We addressed the mechanism underlying these two features of signal tuning by using three ligands: EGF; transforming growth factor α (TGFα); and their chimera, denoted E4T, which act on cells singly expressing ErbB-1 as a weak, a strong, and a very strong agonist, respectively. Co-expression of ErbB-2, a developmentally important co-receptor whose expression is frequently elevated in human cancers, specifically potentiated EGF signaling to the level achieved by TGFα, an effect that was partially mimicked by ErbB-3. Analysis of the mechanism underlying this trans-potentiation implied that EGF-driven homodimers of ErbB-1 are destined for intracellular degradation, whereas the corresponding heterodimers with ErbB-2 or with ErbB-3, dissociate in the early endosome. As a consequence, in the presence of either co-receptor, ErbB-1 is recycled to the cell surface and its signaling is enhanced. This latter route is followed by TGFα-driven homodimers of ErbB-1, and also by E4T-bound receptors, whose signaling is further enhanced by repeated cycles of binding and dissociation from the receptors. We conclude that alternative endocytic routes of homo-and hetero-dimeric receptor complexes may contribute to tuning and diversification of signal transduction. In addition, the ability of ErbB-2 to shunt ligand-activated receptors to recycling may explain, in part, its oncogenic potential.

The combination of heat and chemotherapy was studied in an intraperitoneal tumour model. Rats bearing peritoneal CC531 tumours (2-6 mm) were treated i.p. with cDDP or CBDCA [maximal tolerated dose (MTD)] in combination with regional hyperthermia (41.5 degrees C, 1 h) of the peritoneal cavity. The addition of hyperthermia to the i.p. treatment led to a decrease in the MTD of cDDP by 33.3% at 41.5 degrees C. This was due to increased nephrotoxicity. The MTD of CBDCA did not change as a result of hyperthermia treatment. The chemo-hyperthermia treatment resulted in more cDDP or CBDCA DNA adducts in peritoneal tumours after the combined treatment than after chemotherapy alone. The increased tumour platinum concentrations, rising from 1.3 micrograms Pt g-1 tumour at 37 degrees C to 5.4 micrograms Pt g-1 tumour at 41.5 degrees C for cDDP and from 0.2 microgram Pt g-1 tumor to 0.7 microgram Pt g-1 tumour at 41.5 degrees C for CBDCA, contributed considerably to the enhanced numbers of cDDP or CBDCA DNA adducts. As a result of the latter, i.p. chemotherapy combined with regional hyperthermia led to an increase in tumour growth delay (TGD) after increasing the temperature to 41.5 degrees C for cDDP and CBDCA (by 40 days for cDDP, 22 days for CBDCA). These data were in agreement with the in vitro findings, i.e. that higher temperatures led to increased cytotoxicity.

Human epidermal growth factor (EGF) and human transforming growth factor alpha (TGF-alpha) are structurally related polypeptide growth factors that exert their mitogenic activity through interaction with a common cell-surface receptor, the epidermal growth factor receptor (EGFR). The biological effect induced by these two ligands is quantitatively similar in most cases; in some test systems, however, TGF-alpha functions as a more potent form of EGF. In this study, we have compared EGF, TGF-alpha and ten previously described chimaeras of these two ligands in terms of their ability to generate a mitogenic response in cells carrying the human EGFR, and observed that three of the mutant growth factors (E3T, E4T and T3E4T) are mitogenic at concentrations 10-fold lower than that of either wild-type EGF or TGF-alpha. No difference in tyrosine kinase activity of the receptor towards an external substrate was observed after binding of the various mutants. It has been established before [Ebner and Derynck (1991) Cell Regulation 2, 599-612] that EGF and TGF-alpha differ in the processing of the receptor-ligand complex after internalization, as a result of their different pH sensitivities of receptor binding. Similar measurements on our chimaeric mutants revealed that the above superagonists show an enhanced pH dependence of binding in comparison with EGF. Furthermore, induction of receptor recycling by these superagonists is largely comparable with that induced by TGF-alpha. No superagonistic behaviour was observed on a cell-line containing an EGFR/erbB-2 chimaera which does not show ligand-induced internalization. These data show that EGF/TGFalpha chimaeras can be more active than the naturally occurring ligands, and that receptor recycling after ligand-induced internalization seems to be a prerequisite for this phenomenon.

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