Enhanced expression of the antiapoptotic gene BCL-2 may participate in chemoresistance. To ascertain if multiple myeloma cells surviving exposure to chemotherapy alter their BCL-2 expression, we treated the myeloma cell lines 8226, IM-9, and U266 as well as a primary myeloma cell culture with various injurious agents. Doxorubicin, etoposide, and hydrogen peroxide consistently induced a concentration- and time-dependent upregulation of BCL-2 expression in all myeloma target cell types assayed by flow cytometry and Western blot analysis. In contrast, serum starvation, dexamethasone, and anti-fas antibodies had no effect on expression. Enhanced expression of BCL-2 was relatively selective as treatments had no effect on expression of Ig light chains, BCL-X, or actin. An reverse transcriptase-polymerase chain reaction assay showed increased levels of BCL-2 RNA in 8226 cells as early as 4 hours after treatment with doxorubicin at a time when cell recoveries were not decreased. Thus, doxorubicin stimulates BCL-2 expression in individual 8226 cells rather than simply allowing a selected survival of high BCL-2-expressing cells in culture. Doxorubicin-treated 8226 cells with upregulated BCL-2 expression were relatively resistant to a second exposure of doxorubicin. In addition, BCL-2-transfected IM-9 cells, with enhanced expression of BCL-2 which was comparable to that achieved by initial exposure to doxorubicin, were resistant to doxorubicin and etoposide cytotoxicity. These data suggest that exposure to chemotherapeutic agents may enhance BCL-2 expression in surviving myeloma cells and contribute to acquired chemoresistance.