Novel RUNX1 isoforms determine the fate of acute myeloid leukemia cells by controlling CD56 expression

Gattenloehner S, Chuvpilo S, Langebrake C, Reinhardt D, Muller-Hermelink HK, Serfling E, Vincent A, Marx A
Source: Blood
Publication Date: (2007)
Issue: 110(6): 2027-33
Research Area:
Cancer Research/Cell Biology
Immunotherapy / Hematology
Cells used in publication:
Species: human
Tissue Origin: blood
Species: human
Tissue Origin: blood
Species: human
Tissue Origin: blood
Nucleofector® I/II/2b
For cell transfection, the RUNX1 isoforms p48, p38a, p30 and p24 as well as the CD56 isoforms CD56120kDa and CD56140kDa were cloned into the mammalian expression MIDGE-vector pMCV1.2. 2x10^6 K562, HL-60 and U937 cells were transfected with 1-3 µg of plasmid DNA using solution V and program T-019 (HL-60) and T-003 (K562). For controls, equal amounts of pmaxGFP expressing plasmid were used to nucleofect cells under identical conditions. Transfection efficieny was determined as the proportion of GFP positive cells assessed with a fluorescent microscope and ranged between 60% (HL-60) and 85% (K562). For RNAi, K562 and HL-60 cells were nucleofected with 50 nM RUNX1-exon 3, -exon 5a and -exon 5.4 as well as AlexaFluor488 labelled negative control siRNA duplexes. Transfection efficiency (~98%) was determined by fluorescence microscopy.
CD56(high) acute myeloid leukemias (AMLs) have a poor prognosis but it has been unclear how CD56 expression is controlled and how it relates to clinical aggressiveness. We show here that CD56 expression on AML cells correlates with an abnormal expression pattern of RUNX1 isoforms. Whereas full-length p48 RUNX1 (p48) upregulated CD56 in AML cells, three previously unknown shorter RUNX1 isoforms, p38a, p30 and p24, suppressed CD56 expression. Both p48 and CD56 induced nuclear translocation of NF-kappaB and increased bcl2L12 expression, and inhibition of this pathway by siRNA-mediated p48 knock-down or NF-kappaB blockade, substantially increased apoptosis in CD56(+) AML cell lines. These findings indicate the potential for new therapy of CD56(high) AML by suppression of the "overactive" RUNX1/CD56/NF-kappaB signalling pathway(s).