Chronic myelogenous leukemia (CML) is an indolent hematological malignancy that affects approximately 4300 new patients each year in the United States. A 9:22 chromosomal translocation that results in juxtaposition of the bcr and abl genes is detected in more than 95% of these cases. Allogeneic bone marrow transplantation (BMT) is curative; but the majority of CML patients are not candidates for this treatment. High dose chemotherapy followed by rescue with autologous stem cells (autoBMT) appears to improve survival but is associated with a high rate of relapse. CML cells that persist in the autograft have been shown to contribute to this relapse. Recent studies have identified the tyrphostin AG957 and its more potent analog NSC 680410 as inhibitors of the p210bcr/abl kinase. Unlike agents that compete for the ATP binding site of the kinase, these agents mimic the polypeptide substrates of bcr/abl and are non-competitive with respect to ATP. Preliminary studies from the applicant's laboratory indicate that these agents cause a sequential decrease in tyrosine phosphorylation of the p210/bcr/abl kinase, degradation of the kinase, and activation of apoptotic pathways in the CML-derived K562 leukemia cell line. In addition, AG957 and NSC 680410 selectively inhibit proliferation of committed myeloid progenitors from CML patients as compared to normal controls. The cytotoxicity of these agents, combined with their selectivity for CML as opposed to normal committed progenitors, raises the possibility that these agents might be suitable for ex vivo purging of CML marrow in the setting of autologous BMT. To assess this possibility in greater detail, the applicant proposes to 1) examine the effects of a 24 hr. incubation with NSC 680410 (analogous to a potential purging regimen) on survival of committed progenitors from CML patients and normal volunteers; 2) examine the effects of a 24 hr. incubation with NSC 680410 on earlier CML and normal marrow progenitors using SCID mice implanted with bone marrow fragments; 3) determine the effect of combining NSC 680410 with inhibitors of other signal transduction pathways thought to be activated by bcr/abl; 4) identify the protease(s) responsible for NSC 680410-induced p210/bcr/abl degradation; and 5) attempt to identify the p210/bcr/abl-activated signal transduction pathway whose interruption by NSC 680410 plays a critical role in the cytotoxicity of this agent. Collectively, these studies will elucidate the mechanism of action of this class of agents and evaluate the feasibility of a subsequent trial of NSC 680410 as a potential CML-selective purging agent for use in autoBMT.
