ENHANCED SENSITIVITY TO TUMOR-NECROSIS-FACTOR-ALPHA IN DOXORUBICIN-RESISTANT TUMOR-CELL LINES DUE TO DOWN-REGULATED C-ERBB2

We have studied the relationship between tumor necrosis factor (TNF)-s ensitivity and doxorubicin-resistance in our doxorubicin-resistant cel l line panel consisting of the parental cell line GLC(4) plus GLC(4)-A dr(2x) and GLC(4)-Adr(350x) with respective resistance factors of 2 an d 350 compared with GLC(4). At the highest dose of 1000 ng/ml TNF, GLC (4) was almost completely resistant to TNF, whereas 37% and 68% growth inhibition was observed in GLC(4)-Adr(2x) and GLC(4)-Adr(350x), respe ctively. Sensitivity to TNF appeared to correlate inversely with the e xpression and gene copies of topoisomerase II alpha in these cell line s. The gene encoding for c-erbB2 is in the proximity of the topoisomer ase II alpha gene and its product is a known cause for TNF-resistance. We found that our doxorubicin-resistant cell lines with decreased top oisomerase II alpha gene copies have a simultaneous decrease in c-erbB 2 gene copies, probably due to linkage between these 2 genes. This red uced number of c-erbB2 gene copies resulted in decreased c-erbB2 expre ssion and subsequently in increased sensitivity to TNF. Additionally, we tried to establish some of the mechanisms associated with TNF-resis tance in GLC4 related to c-erbB2 overexpression. There was no differen ce in TNF-receptor-I expression between the cell lines. Compared with the TNF-sensitive GLC(4)-Adr(350x), GLC(4) appeared to have a decrease d activation of NF-kappa B after exposure to TNF that might indicate a reduced TNF-receptor function. In GLC4, increased Bcl-2 expression wa s found, a protein described to confer TNF-resistance. Moreover, it wa s demonstrated that combining TNF with the poly-ADP-ribose polymerase (PARP) inhibitors 6-aminonicotinamide and 3-aminobenzamide did not aff ect TNF-sensitivity in GLC(4) and GLC(4)-Adr(350x) excluding a pivotal role of PARP in TNF-resistance in these cell lines. In conclusion, ou r data show that doxorubicin-resistant tumor cell lines with decreased topoisomerase II alpha gene copies can become sensitive to TNF due to loss of c-erbB2 gene copies. We also found that several mechanisms as sociated with c-erbB2 overexpressing contribute to TNF-resistance in G LC(4). (C) 1998 Wiley-Liss, Inc.