GROWTH SUPPRESSION OF ACUTE PROMYELOCYTIC LEUKEMIA-CELLS HAVING INCREASED EXPRESSION OF THE NONREARRANGED ALLELES - RAR-ALPHA OR PML

The balanced t(15;17) rearrangement found in acute promyelocytic leuke mia (APL) cells fuses PML on chromosome 15 to the retinoic acid recept or alpha (RAR alpha) on chromosome 17, PML/RAR alpha is expressed in A PL cells with the non-rearranged alleles, PML and RAR alpha, Clinical remissions induced by all-trans-retinoic acid (RA) treatment of APL pa tients are linked to expression of PML/ RAR alpha, a transcription fac tor with reported dominant negative functions, The roles of PML and RA R alpha in the RA response of APL have not yet been fully explored, Th is study examines these roles by individually transfecting RAR alpha a nd PML into NB4 APL cells. NB4 is the sole APL cell Line containing th e t(15;17), RA treatment represses NB4 cell growth and induces a myelo id phentoype, Full length cDNAs for RAR alpha and PML were individuall y cloned into a CMV-driven expression vector containing the neomycin r esistance gene, Surprisingly, none of the obtained stable transfectant s expressed exogenous RAR alpha or PML mRNAs even when reverse transcr iption polymerase chain reaction (RT-PCR) detection assays were used. All clones expressed the neomycin resistance gene and were similar to parental NB4 cells in their growth and differentiation properties, An explanation explored for this lack of gene expression was that increas ed levels of RAR alpha or PML might suppress APL cell growth, To exami ne this possibility, transfection experiments were repeated using an e pisomal vector-based expression system containing an SV40 driven RAR a lpha or PML cDNA and the hygromycin B resistance gene, A new selection strategy augmented expression of the desired cDNAs, A control episoma l vector lacked a cDNA insert, Following electroporation and selection , exogenous RAR alpha expression was obtained. Compared to controls, t he growth of these transfectants was markedly inhibited before and aft er RA-treatment and these cells more prominently induced myeloid matur ation markers, In contrast, exogenous PML expression was transient sin ce these transfectants did not appear to propagate in culture, These f indings indicate: (1) a growth disadvantage for NB4 cells having incre ased expression of RAR alpha or PML and (2) increased RAR alpha expres sion augmented RA-mediated maturation of NB4 cells, This implicates a role for RAR alpha or PML in regulating the growth or differentiation of APL cells, It is hypothesized this occurs through antagonism of PML /RAR alpha actions in these leukemic cells.