Mechanisms of cell death of chronic lymphocytic leukemia lymphocytes by RNA-directed agent, 8-NH2-adenosine

Purpose: To determine if RNA-directed nucleoside analogue, 8-NH2-adenosine, induces cell death and if that is accompanied with transcription inhibition of the key survival factors of chronic lymphocytic leukemia (CLL) cells. Experimental Design: Primary lymphocytes from CLL patients were incubated with 10 mu mol/L 8-NH2-adenosine for 2, 4, and 6 or 8 hours. The accumulation of analogue triphosphate and the decline in endogenous ATP pool were analyzed by high-performance liquid chromatography. Inhibition of global RNA and protein synthesis was measured and correlated with specific decline in transcript and protein levels of MCL-1, XIAP, and BCL-2, the key survival factors of CLL. These biochemical and molecular end points were related to cell death of these quiescent lymphocytes. Results: In vitro incubations of CLL lymphocytes with 8-NH2-adenosine resulted in rapid but heterogeneous accumulation of 8-NH2-ATP (390-680 mu mol/L), with a concomitant decline in endogenous ATP (median, > 50% by 4 hour). Global RNA synthesis was decreased in all samples and was associated with a decline in MCL-1, XIAP, and BCL-2 transcripts. There was a parallel decrease in the protein level of MCL-1 and XIAP but hot BCL-2. These biochemical changes were accompanied by apoptosis. Conclusion: The evidence of CLL cell death with complementary changes in the expression of survival proteins provides a molecular rationale for using 8-NH2-adenosine as a therapeutic agent for this indolent leukemia.