Page 73 - Haematologica-April 2018

Gfi1b in AML and MDS
Kras. RAS mutations are found in 5-10% of AML patients.2 These mice harbor a transcriptional stop codon flanked by loxP sites upstream of a mutated Kras allele and, after removal of the stop codon, develop myelopro- liferative disorder.35 We crossed these mice with Gfi1bfl/flMxCretg or Gfi1bwt/wtMxCretg mice, and after poly(I:C) administration, we observed mice for the emer- gence of disease (Figure 4A).
While Gfi1bwt/wtMxCretgKras+/fl mice developed a lethal myeloproliferative disorder with a median survival of approximately 25 days, loss of Gfi1b significantly short- ened the latency period of the disease to a median sur- vival of approximately seven days (Figure 4B). There was no difference with regard to cytological appearance, num- ber of myeloid cells or level of apoptosis (Figure 4C and D and Online Supplementary Figure S5A-D). We also did not observe any significant difference with regard to white blood counts, platelet counts or spleen size but a significant difference in hemoglobin and red blood cells between Gfi1bfl/flMxCretg and Gfi1bwt/wtMxCretg animals (Online Supplementary Figure S5E-H), which might be due to the role of Gfi1b in erythropoiesis.6,9
Loss of Gfi1b promotes the progression of AML initiated by retroviral MLL-AF9 expression
The Mixed Lineage Leukemia (MLL) gene is a common target for chromosomal translocations.2 MLL-AF9 is a fusion protein frequently occurring in a subset of AML patients,2 and its expression in hematopoietic progenitors has been linked to the induction of AML in mice.36 As a third AML mouse model, we thus used mice that devel- oped AML through the induction of MLL-AF9 expression, the product of the t(9;11)(q22;p23) translocation. Lin- BM cells derived from Gfi1bwt/wtMxCretg or Gfi1bfl/flMxCretg mice were transduced with a retrovirus expressing MLL-AF9 and transplanted into lethally irradiated C57BL/6J mice. For Cre-mediated excision of Gfi1b in the transplanted cells, mice were injected with poly(I:C) three weeks after transplantation (Figure 4E). Poly(I:C)-injected mice with MLL-AF9-transduced Gfi1bfl/flMxCretg (Gfi1b-deficient) cells succumbed faster to leukemia than mice injected with poly(I:C) and transplanted with MLL-AF9-transduced Gfi1bwt/wtMxCretg (Gfi1b-expressing) cells (Figure 4F). However, there were no major qualitative differences con- cerning cytological findings and or blood parameters
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Figure 2. Different Gfi1B levels are indica- tive of prognosis of myelodysplastic syn- drome (MDS) or acute myeloid leukemia (AML) patients. (A) High expression of GFI1B in human AML cells (n=116) com- pared to lower expression of GFI1B (0-15%) in AML cells (n=394) based on the patient cohort published by Verhaak et al.22 (B) Overall survival (OS) from patients described in Verhaak et al.22 with regard to GFI1B expression (P=0.0443). (C) Overall survival (OS) from patients described in Verhaak et al.22 (restricted to cytogenetically normal patients) with regard to GFI1B expression; P=0.0407. (D) Same as in (C) but with regard to event-free survival (EFS); P=0.0350. (E) Analysis of signaling pathways with low GFI1B expression (the lowest 5% com- pared with the highest 20% of expression levels) in a bigger dataset from Verhaak et al.22 Analysis was performed by Signaling Pathway Enrichment using Experimental Datasets (SPEED) analysis. Pathways such as reactive oxygen species (ROS; H2O2), MAPK, JAK, TGFB and TLR are highly significant. (F) High expression of GFI1B (31-100%) in human MDS patients (n=32) compared to lower expression of GFI1B (0-30%) in MDS patients (n=85) based on the patient cohort published by Papaemmanuil et al.23 (G) EFS of patients described in Papaemmanuil et al.23 with regard to GFI1B expression; P=0.032.
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