CYTOGENETIC AND MOLECULAR CHARACTERIZATION OF RANDOM CHROMOSOMAL REARRANGEMENTS ACTIVATING THE DRUG-RESISTANCE GENE, MDR1 P-GLYCOPROTEIN, IN DRUG-SELECTED CELL-LINES AND PATIENTS WITH DRUG-REFRACTORY ALL/

resistance, both primary and acquired, is a major obstacle to advances in cancer chemotherapy. In vitro, multidrug resistance can be mediate d by P-glycoprotein (PGY 1), a cell surface phosphoglycoprotein that a cts to efflux natural products from cells. PGY 1 is encoded by the MDR 1 gene located at 7q21.1. Overexpression of MDR1 has been demonstrated in many cancers, both in patient tumors and in cell lines selected wi th a variety of chemotherapeutic agents. Recent studies in drug-select ed cell lines and patients samples have identified hybrid mRNAs compri sed of an active, but apparently random, gene fused 5' to MDR1. This o bservation indicates that random chromosomal rearrangements, such as t ranslocations and inversions, leading to ''capture'' of MDR1 by consti tutively expressed genes may be a mechanism for activation of this gen e following drug exposure. In this study, fluorescence in situ hybridi zation (FISH) using whole chromosome paints (WCP) and bacterial artifi cial chromosome (BAG)-derived probes showed structural rearrangements involving 7q in metaphase and interphase cells, and comparative genomi c hybridization (CGH) revealed high levels of amplification at chromos omal breakpoints. In an adriamycin-selected resistant colon cancer lin e (S48-3s/Adr), WCP4/WCP7 revealed t(4;7)(q31;q21) and BAG-derived pro bes demonstrated that the breakpoint lay between MDR1 and sequences 50 0-1000 KB telomeric to it. Similarly, in a subline isolated following exposure to actinomycin D (S48-3s/ActD), a hybrid MDR1 gene composed o f heme oxygenase-2 sequences (at 16p13) fused to MDR1 was identified a nd a rearrangement confirmed with WCP7 and a subtelomeric 16p probe. L ikewise, in a paclitaxel-selected MCF-7 subline where GASP sequences ( at 7q22) were shown to be fused to MDR1, WCP7 showed an elongated chro mosome 7 with a homogeneously staining regions (hsr); BAG-derived prob es demonstrated that the hsr was composed of highly amplified MDR1 and GASP sequences. In all three selected cell lines, CGH demonstrated am plification at breakpoints involving MDR1 (at 7q21) and genes fused to MDR1 at 4q31, 7q22, and 16p13.3. Finally, in samples obtained from tw o patients with drug refractory ALL, BAG-derived probes applied to arc hived marrow cells demonstrated that a breakpoint occurred between MDR 1 and sequences 500-1000 KB telomeric to MDR1, consistent with a rando m chromosomal rearrangement. These results support the proposal that r andom chromosomal rearrangement leading to capture and activation of M DR1 is a mechanism of acquired drug resistance. (C) 1998 Wiley-Liss, I nc.