AF64A-INDUCED CHANGES IN N-MYC EXPRESSION IN THE LA-N-2 HUMAN NEUROBLASTOMA CELL-LINE ARE MODULATED BY CHOLINE AND HEMICHOLINIUM-3

Due to AF64A's structural similarity to choline, AF64A can selectively affect cholinergic neurons, which possess a high affinity choline tra nsport system for acetylcholine synthesis. The mechanism by which AF64 A selectively produces its cytotoxic effect is unknown. However, based on previous studies that demonstrate that DNA lesions produced by AF6 4A caused premature termination of N-myc transcription in vitro, it is possible that AF64A may affect the transcription of genes necessary f or developmental maintenance in cholinergic cells. Using the LA-N-2 ce lls as a model to study the effects of AF64A in a purely cholinergic s ystem, we investigated the effects of AF64A on the expression of the N -myc gene and monitored cell growth. AF64A produced a maximal decrease in N-myc mRNA with a return to steady state levels al later time poin ts. Moreover, a decrease in cell numbers in AF64A-treated cells was ob served, and these cells did not double in number at their respective d oubling time as compared to control. In other studies, a causal relati onship between a reduction in N-myc and an inhibition of cell growth a nd replication has been reported. While these studies do not allow us to conclude that AF64A is specific for N-myc, the data do, nevertheles s, suggest that AF64A affects cell growth and/or replication by down-r egulating the expression of N-myc which is involved in differentiation and cell growth in neuroblastomas. Presence of choline or hemicholini um-3 prevented the AF64A-induced decrease of N-myc levels by competing with, or inhibiting the choline transport mechanism by which AF64A en ters the cell, respectively.