TRIB2 Transformed GMP is the Myeloid Leukaemia Initiating Cell
2016
serine/threonine pseudokinases. When ectopically expressed in hematopoietic
stem and progenitor cell (HSPC) enriched bone marrow cells, TRIB2 was
shown to induce acute myeloid leukemia (AML) in a murine transplant model
dependent on C/EBPα degradation, showing it to be a potent myeloid oncogene.
The cell of origin or leukemia initiating cell (LIC) refers to the cell from
which a specific leukemia normally arises, and it is hypothesised that the LIC
may influence the progression, disease phenotype and response to therapy. It
remains unclear whether the LIC is a HSC or a more committed progenitor cell
in TRIB2-driven AML.
Aims: Our current study focuses on identifying the LIC in TRIB2-driven AML
and characterizing its role in disease potency and maintenance.
Methods: FACS sorted CD45.2+ stem and progenitor cell populations (HSC,
MPP, CMP, GMP and MEP) transduced with a lentiviral vector encoding TRIB2
were cultured in methocult supplemented with cytokines that support myeloid
cell growth and differentiation. Following the first plating (P1), GFP+ cells were
sorted and serially replated. Cells which still formed colonies by the third
replating (P3) implied acquisition of self-renewal ability and increased proliferation
characteristic of myeloid transformation. These CD45.2+ cells from P3
were transplanted into sublethally irradiated CD45.1+ C57BL/6 recipient mice
and chimeric animals were monitored for 1 year. Chemoresistance experiments
were performed on the bulk bone marrow population (CD45.2+ cells >95%) from the TRIB2 AML mice. Cells were treated with a range of concentrations
of daunorubicin (DNR), followed by trypan blue cell counts to assess
viability.
Results: Our study identified that while lentivirally transduced TRIB2 can transform
all stem and progenitor cell populations of the hematopoietic system with
variable efficiencies in vitro, the GMP subpopulation was identified as the LIC
of TRIB2-driven AML. TRIB2 transformed GMP cells generated a more potent
AML with complete penetrance and shortened latency compared to all other
HSPC populations analysed. Indeed, phenotypically different diseases were
propagated from TRIB2 expression in the HSC and GMP, with the former having
a weakly penetrant, longer latency AML with a mixed lineage phenotype, whereas
the later was a dominantly myeloid disease phenotype with a short latency.
We next addressed the chemotherapeutic response of TRIB2 positive AML
cells. We show that GMP-TRIB2 AML and bulk TRIB2 AML cells are chemoresistant.
TRIB2 overexpression decreases DNR induced apoptosis, and knockdown
of TRIB2 expression in AML cells leads to an increase in apoptotic gene
expression. Our studies illustrate that TRIB2 expression is key in mediating
the anti-apoptotic signals following DNR treatment.
Summary/Conclusions:We identify the GMP as the LIC in TRIB2 driven AML.
Our findings are further supported by our previous work showing that degradation
of C/EBPa is required for TRIB2-driven AML and the GMP population
expresses the highest level of C/EBPa in hematopoiesis. We provide evidence
for TRIB2 role in chemoresistance, and that the TRIB2 LIC is a highly chemoresistant
cell. Our findings provide insight into the molecular events contributing
to AML, and provide potential for novel avenues for therapeutic targeting.
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