RATIO OF BCL-XSHORT TO BCL-XLONG IS DIFFERENT IN GOOD-PROGNOSIS AND POOR-PROGNOSIS SUBSETS OF ACUTE MYELOID-LEUKEMIA

Background: Acute myeloid leukemia (AML) is a heterogeneous collection of leukemic disorders ranging from chemotherapy-sensitive subsets [in version 16 and t(8;21)], which often can he cured with cytosine arabin oside alone, to the most resistant subsets, which can survive even sup ralethal levels of combination alkylator chemotherapy (cytogenetic sub sets monosomy 5 and monosomy 7). Materials and Methods: To analyze the expression of BCL-2 family genes, which are expressed in these subset s of AML, we used PCR sequence amplification reactions that are depend ent on oligonucleotide primers representing the BH1 and BH2 homology d omains to generate the unique regions between BH1 and BH2. These prime rs are conserved among all members of the BCL-2 gene family and are se parated by a 150 nucleotide region sequence between the BH1 and BH2 do mains. The PCR products unique to each BCL-2 family member were cloned directionally into sequencing vectors. The identity of the insert of each clone was determined by slot-blots of the DNA amplified from indi vidual colonies and by hybridization with radioactive probes specific to the bcl-2, bcl-x, or bax genes. Results: We found that bcl-2 is the predominant member expressed in AML samples with a poor prognosis (-5 , -7), whereas the transcripts of bcl-x are higher than those of bcl-2 in the AML samples with a good prognosis [invl6, t(8;21)]. No signifi cant difference in bax expression was detected between AML subsets of good and bad prognosis. The ratio of bcl-xlong, which inhibits apoptos is, to bcl-xshort, which promotes apoptosis, was determined by amplifi cation with a pair of primers specific to bcl-x followed by separation of the PCR product on agarose gels. Bcl-xlong and bcl-xshort appeared as bands of different molecular mass on a molecular weight gel and we re visualized by ethidium bromide staining or Southern blot analysis w ith a bcl-x-speclfic probe. Conclusions: We found that the ratio of bc l-x long to bcl-x short was higher in the AML patients with a poor pro gnosis. These experiments showed that the levels of BCL-2 family membe rs in the leukemia cells of good-and poor-prognosis subsets are differ ent. In addition, novel members of the BCL-2 family were isolated from the cells of AML patients of either prognosis.