A role for MEIS1 in MLL-fusion gene leukemia

Abstract

Leukemias with MLL rearrangements are characterized by high expression of the homeobox gene MEIS1. In these studies, we knocked down Meis1 expression by shRNA lentivirus transduction in murine Mll-AF9 leukemia cells. Meis1 knockdown resulted in decreased proliferation and survival of murine Mll-AF9 leukemia cells. We also observed reduced clonogenic capacity and increased monocytic differentiation. The establishment of leukemia in transplantation recipients was significantly delayed by Meis1 knockdown. Gene expression profiling of cells transduced with Meis1 shRNA showed reduced expression of genes associated with cell cycle entry and progression. shRNA-mediated knockdown of MEIS1 in human MLL-fusion gene leukemia cell lines resulted in reduced cell growth. These results show that MEIS1 expression is important for MLL-rearranged leukemias and suggest that MEIS1 promotes cell-cycle entry. Targeting MEIS1 may have therapeutic potential for treating leukemias expressing this transcription factor.

Figure 1

shRNA lentivirus-mediated Meis1 knockdown inhibits cell growth and promotes differentiation in murine Mll-AF9 leukemia. The 4166 cells were transduced for 5 days with the various lentivirus constructs and cultured in the presence of puromycin as described. (A) RT-PCR for Meis1 and Gapdh (housekeeping gene). (B) Bar graph depicting cell growth after Meis1 shRNA transduction compared with vector control, the latter being 100%. (C) By light microscopy, Meis1 knockdown resulted in monocytic differentiation. Staining for the monocyte-specific marker F4/80 showed increased uptake with Meis1 knockdown. Slides were viewed with a Nikon Eclipse 80i microscope (Nikon, Melville, NY) at a 10×/0.3 NA, WD 16.0 mm objective with fixed cells imaging medium. Images were acquired using a Nikon Digital Sight DS-5M-L1 camera and were processed using Nikon DS-L1 image acquisition software. Image adjustment was performed using Adobe Photoshop CS2 (Adobe Systems, San Jose, CA).

Figure 2

Meis1 knockdown induces cell-cycle arrest and apoptosis and delays establishment of leukemia in vivo. The 4166 cells were transduced with M26 or control virus as described and cultured for 48 hours without puromycin. (A) Top panel: Nuclei were extracted and stained with PI. Analysis of DNA content by flow cytometry showed an increase in the proportion of G₀/G₁ nuclei (left peak) in M26-transduced cells compared with control virus. Bottom panel: Increased apoptosis was evident by an increase in the uptake of PI and the pan-activated caspase maker CaspaTag. (B) Lethally irradiated mice were given 4166 cells that were either untreated or transduced with M26 or control virus as described. Mice were killed when showing signs of distress. The graph shows a survival analysis comparing the 3 groups (9 mice per group). Mice receiving M26-transduced cells had significantly prolonged survival (P < .001). (C) MEIS1 knockdown inhibits growth of human MLL-fusion gene leukemia cell lines. The cell lines indicated were transduced with MEIS1 shRNA lentivirus or control lentivirus for 3 to 5 days as described at MOI of 10 to 50. Bars represent relative cell growth and MEIS1 expression compared with controls (mean ± SE; n = 3; P < .05 except for HPB-NULL).