Despite their widespread use, little is known of either the mechanism of action of glucocorticoids in the treatment of multiple myeloma or why patients ultimately become resistant to their therapeutic effects. Here, we address these issues by examining the direct effects of the glucocorticoid dexamethasone (DEX) on a hormone-sensitive clone (MM.1S) of a human multiple myeloma line and compare them with those of its hormone-resistant counterpart (MM.1R). MM.1S expresses approximately 50,000 glucocorticoid receptors (GR) per cell, the full-length 7.1-kb GR mRNA at high levels, and is lysed by DEX. DEX-induced cytolysis is effectively blocked by the glucocorticoid antagonist, RU 486, indicating the specificity of this response for the GR. In contrast to MM.1S, MM.1R is not lysed by hormone, has little hormone-binding activity, and expresses the 7.1-kb GR mRNA at low levels. Interestingly, we have found that two distinct phenotypes emerge from MM.1R with increasing periods of growth in culture. The first or “early” form, MM.1Re, expresses high levels of a variant GR mRNA of 5.5 kb that has a deletion in its 3′ end. With further growth in the presence or absence of selective media, the expression of this transcript is repressed, resulting in the second or “late” phenotype characteristic of MM.1RL. No discernible differences in the organization of the genomic GR sequence in DEX-sensitive and -resistant cells were detectable by Southern analysis, suggesting that no gross deletions, rearrangements, or allelic variations in the genomic sequence account for the resistant phenotypes of MM.1R.