MMP-14 Mediates Migration of Acute Myelogenous Leukemia Cells

Matrix metalloproteinase (MMP)-14 expression correlates with progression and metastasis of multiple tumor cell types and is a major mediator of cell migration and invasion. MMP-14 possesses a trans-membrane domain that tethers the enzyme to the plasma membrane and not only activates proMMP-2 but also degrades extracellular matrix (ECM) by pericellular proteolysis and cleaves several non-ECM molecules, including adhesion molecules and chemokines. To assess the role of MMP-14 in leukemic dissemination we evaluated its expression in myeloid cell lines and primary acute myelogenous leukemic (AML) samples. Using RT-PCR, flow cytometry and Western blotting we found that MMP-14 is highly expressed in

• leukemic myeloid cell lines THP-1, U937, HEL and K562; and

• primary samples from 37 out of 40 patients (pts) diagnosed with AML (WHO classification, AML with recurrent cytogenetic translocations: 9 pts; AML with multilineage dysplasia: 4 pts; AML not otherwise categorized: 27 pts).

Moreover, primary AML blasts secreted the 72 kDa proenzyme form of MMP-2 into media (zymography) which became activated after co-culture of AML blasts with bone marrow (BM) stromal cells. This activation was inhibited by the potent MMP-14 inhibitor epigallocatechin-3-gallate (EGCG). We also found that

• migration of primary AML cells (trans-Matrigel invasion assay) was inhibited by EGCG; and

• silencing of MMP-14 by transfecting THP-1 cells and AML blasts with MMP-14 siRNA oligonucleotides resulted in reduced trans-Matrigel migration of these cells.

Given that AML blasts constitutively secrete TNF-α, we confirmed that

• AML blasts express mRNA for TNF-α as well as its receptors TNFR1 and TNFR2; and

• TNF-α levels are elevated in the plasma of AML patients.

However, we demonstrated that recombinant human (rh) TNF-α further upregulated MMP-14 expression in AML blasts (RT-PCR, Western blot) and increased its incorporation into membrane lipid rafts (confocal microscopy). Moreover, rh TNF-α- stimulated AML blasts

• were found to be more potent activators of pro-MMP-2 than unstimulated cells (zymography), indicating that activation may occur via upregulation of MMP-14;

• had significantly higher migration which was inhibited by EGCG and also by the TNF-α receptor inhibitor Enbrel; and

• rh TNF-α had no effect on the migration of MMP-14 siRNA-silenced AML cells.

Other factors tested, for example, SDF-1, IL-1 and TGF-β, had no effect on MMP-14 expression in AML cells (flow cytometry). In conclusion, we report here for the first time that MMP-14 is expressed in AML blasts and is modulated by TNF-α. We suggest that, by pericellular degradation of ECM and activation of latent MMP-2 secreted by AML blasts, MMP-14 may contribute to the highly proteolytic BM microenvironment in AML and the invasive phenotype of this malignancy.