Chlorambucil induced apoptosis in chronic lymphocytic leukemia (CLL) and its relationship to clinical efficacy

Chlorambucil induced apoptosis was measured in CLL cells treated with clinically achievable drug doses in vitro. While spontaneous apoptosis occurred in CLL cells incubated in vitro in the absence of drug, the level of apoptosis, as measured by the extent of DNA fragmentation, was greater in cells treated with chlorambucil. In addition, macrophages were shown to engulf drug treated CLL cells in vitro. To determine if chlorambucil can also induce apoptosis in vivo, CLL cells were isolated from patients before treatment and at intervals after clinical therapy with chlorambucil (0.9 mg/kg given over 3 days). Apoptosis was measured in these cells immediately after isolation and following incubation in vitro for 72 hr. No apoptotic changes were detected in cells immediately after isolation either before or after clinical treatment. In contrast, apoptosis was observed in cells that were incubated ex vivo for 72 hr, and the level of apoptosis was greater in cells that were isolated after chlorambucil treatment compared with cells obtained prior to therapy. The increased apoptosis observed in CLL cells ex vivo after therapy was related to the fall in the patient's lymphocyte count. In general, a large increase in apoptosis ex vivo after treatment was followed by a significant decrease in the patient's lymphocyte count. Thus, chlorambucil may produce its antitumor effect in CLL by inducing apoptosis-associated membrane changes that result in rapid clearance of the apoptotic cells by the immune system.