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EphA2 Is a Therapy Target in EphA2-Positive Leukemias but Is Not Essential for Normal Hematopoiesis or Leukemia.

Charmsaz S, Beckett K, Smith FM, Bruedigam C, Moore AS, Al-Ejeh F, Lane SW, Boyd AW - PLoS ONE (2015)

Bottom Line: These studies showed that EphA2 does not have an obligatory role in normal hematopoiesis.We showed that treatment with EphA2 monoclonal antibody IF7 alone had no effect on tumorigenicity and latency of the MLL-AF9 leukemias, while targeting of EphA2 using EphA2 monoclonal antibody with a radioactive payload significantly impaired the leukemic process.Altogether, these results identify EphA2 as a potential radio-therapeutic target in leukemias with MLL translocation.

View Article: PubMed Central - PubMed

Affiliation: QIMR Berghofer Medical Research Institute, Brisbane, Australia.

ABSTRACT
Members of the Eph family of receptor tyrosine kinases and their membrane bound ephrin ligands have been shown to play critical roles in many developmental processes and more recently have been implicated in both normal and pathological processes in post-embryonic tissues. In particular, expression studies of Eph receptors and limited functional studies have demonstrated a role for the Eph/ephrin system in hematopoiesis and leukemogenesis. In particular, EphA2 was reported on hematopoietic stem cells and stromal cells. There are also reports of EphA2 expression in many different types of malignancies including leukemia, however there is a lack of knowledge in understanding the role of EphA2 in hematopoiesis and leukemogenesis. We explored the role of EphA2 in hematopoiesis by analyzing wild type and EphA2 knockout mice. Mature, differentiated cells, progenitors and hematopoietic stem cells derived from knockout and control mice were analyzed and no significant abnormality was detected. These studies showed that EphA2 does not have an obligatory role in normal hematopoiesis. Comparative studies using EphA2-negative MLL-AF9 leukemias derived from EphA2-knockout animals showed that there was no detectable functional role for EphA2 in the initiation or progression of the leukemic process. However, expression of EphA2 in leukemias initiated by MLL-AF9 suggested that this protein might be a possible therapy target in this type of leukemia. We showed that treatment with EphA2 monoclonal antibody IF7 alone had no effect on tumorigenicity and latency of the MLL-AF9 leukemias, while targeting of EphA2 using EphA2 monoclonal antibody with a radioactive payload significantly impaired the leukemic process. Altogether, these results identify EphA2 as a potential radio-therapeutic target in leukemias with MLL translocation.

No MeSH data available.


Related in: MedlinePlus

Stem/progenitor cell populations in EphA2 knockout mice compared to wild type littermates.(A) Gating for Progenitors (lineagelowcKithighSca-1-), LKS+ cells (lineagelowcKithighSca-1+ enriched for hematopoietic stem cells), CMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIlow), MEP (lineagelowcKithighSca-1-CD34-FCGRII/III-) and GMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIhigh) presented as percentage of Lineagelow cells. CD34/CD135 gating for MPP (LKS+CD34+CD135+), ST-HSC (LKS+CD34+CD135-) and LT-HSC (LKS+CD34-CD135-). CD48/CD150 gating for LT-HSC (LKS+CD150+CD48-) and MPP (LKS+CD150-CD48+). (B) There were no significant differences in progenitors, LKS+, CMP, MEP or GMP population in the EphA2 knockout mice compared to wild type control mice (n = 15, 3 independent experiments). (C) There were no significant differences in LT-HSC and MPP population gated by CD34/CD135 antigens in the EphA2 knockout mice compared to wild type control. There were significantly more (P = 0.0012) ST-HSCs in EphA2 knockout bone marrow compared to wild type control (n = 15, 3 independent experiments). (D) There were no significant differences in LT-HSC and MPP population gated using CD48/CD150 antigens in EphA2 knockout mice compared to wild type control (n = 15, 3 independent experiments). Each dot corresponds to one individual mouse. The data represent mean ± SEM. Unpaired t test was performed for statistical analyses.
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pone.0130692.g002: Stem/progenitor cell populations in EphA2 knockout mice compared to wild type littermates.(A) Gating for Progenitors (lineagelowcKithighSca-1-), LKS+ cells (lineagelowcKithighSca-1+ enriched for hematopoietic stem cells), CMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIlow), MEP (lineagelowcKithighSca-1-CD34-FCGRII/III-) and GMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIhigh) presented as percentage of Lineagelow cells. CD34/CD135 gating for MPP (LKS+CD34+CD135+), ST-HSC (LKS+CD34+CD135-) and LT-HSC (LKS+CD34-CD135-). CD48/CD150 gating for LT-HSC (LKS+CD150+CD48-) and MPP (LKS+CD150-CD48+). (B) There were no significant differences in progenitors, LKS+, CMP, MEP or GMP population in the EphA2 knockout mice compared to wild type control mice (n = 15, 3 independent experiments). (C) There were no significant differences in LT-HSC and MPP population gated by CD34/CD135 antigens in the EphA2 knockout mice compared to wild type control. There were significantly more (P = 0.0012) ST-HSCs in EphA2 knockout bone marrow compared to wild type control (n = 15, 3 independent experiments). (D) There were no significant differences in LT-HSC and MPP population gated using CD48/CD150 antigens in EphA2 knockout mice compared to wild type control (n = 15, 3 independent experiments). Each dot corresponds to one individual mouse. The data represent mean ± SEM. Unpaired t test was performed for statistical analyses.

Mentions: We next analyzed the numbers of immunophenotype-defined hematopoietic stem and progenitor cells (HSPCs) in the EphA2 knockout mice compared to wild type littermates (Fig 2A). This analysis showed no significant differences in the percentage of Lineage-c-Kit+Sca-1+ (LKS+), progenitors and mature myeloid progenitor cells, including common myeloid progenitors (CMPs), megakaryocyte-erythrocyte progenitors (MEPs), and granulocyte-monocyte progenitors (GMPs) (Fig 2B). To further analyze HSCs, CD34 and FLK2 (CD135) markers were utilized to isolate LT-HSCs (LKS+CD34-Flk2-), ST-HSCs (LKS+CD34+Flk2) or MPP, (LKS+CD34+Flk2+) population. A significant increase in the frequency of ST-HSCs (P = 0.0012) was observed in the EphA2 knockout bone marrow; however no significant differences in either the MPPs or the LT-HSCs were evident (Fig 2C). CD150 and CD48 markers were also used as another method to fractionate HSCs, with (LKS+CD150+CD48-) cell population representing LT-HSCs and (LKS+CD150-CD48+) representing MPPs. Similar to the previous results, there were no significant differences in the frequency of LT-HSCs or MPPs in EphA2 knockout mice compared to wild type control mice (Fig 2D). Altogether, these data suggest that EphA2 is not essential for normal blood development.


EphA2 Is a Therapy Target in EphA2-Positive Leukemias but Is Not Essential for Normal Hematopoiesis or Leukemia.

Charmsaz S, Beckett K, Smith FM, Bruedigam C, Moore AS, Al-Ejeh F, Lane SW, Boyd AW - PLoS ONE (2015)

Stem/progenitor cell populations in EphA2 knockout mice compared to wild type littermates.(A) Gating for Progenitors (lineagelowcKithighSca-1-), LKS+ cells (lineagelowcKithighSca-1+ enriched for hematopoietic stem cells), CMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIlow), MEP (lineagelowcKithighSca-1-CD34-FCGRII/III-) and GMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIhigh) presented as percentage of Lineagelow cells. CD34/CD135 gating for MPP (LKS+CD34+CD135+), ST-HSC (LKS+CD34+CD135-) and LT-HSC (LKS+CD34-CD135-). CD48/CD150 gating for LT-HSC (LKS+CD150+CD48-) and MPP (LKS+CD150-CD48+). (B) There were no significant differences in progenitors, LKS+, CMP, MEP or GMP population in the EphA2 knockout mice compared to wild type control mice (n = 15, 3 independent experiments). (C) There were no significant differences in LT-HSC and MPP population gated by CD34/CD135 antigens in the EphA2 knockout mice compared to wild type control. There were significantly more (P = 0.0012) ST-HSCs in EphA2 knockout bone marrow compared to wild type control (n = 15, 3 independent experiments). (D) There were no significant differences in LT-HSC and MPP population gated using CD48/CD150 antigens in EphA2 knockout mice compared to wild type control (n = 15, 3 independent experiments). Each dot corresponds to one individual mouse. The data represent mean ± SEM. Unpaired t test was performed for statistical analyses.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4470658&req=5

pone.0130692.g002: Stem/progenitor cell populations in EphA2 knockout mice compared to wild type littermates.(A) Gating for Progenitors (lineagelowcKithighSca-1-), LKS+ cells (lineagelowcKithighSca-1+ enriched for hematopoietic stem cells), CMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIlow), MEP (lineagelowcKithighSca-1-CD34-FCGRII/III-) and GMP (lineagelowcKithighSca-1-CD34+FCGRII/IIIhigh) presented as percentage of Lineagelow cells. CD34/CD135 gating for MPP (LKS+CD34+CD135+), ST-HSC (LKS+CD34+CD135-) and LT-HSC (LKS+CD34-CD135-). CD48/CD150 gating for LT-HSC (LKS+CD150+CD48-) and MPP (LKS+CD150-CD48+). (B) There were no significant differences in progenitors, LKS+, CMP, MEP or GMP population in the EphA2 knockout mice compared to wild type control mice (n = 15, 3 independent experiments). (C) There were no significant differences in LT-HSC and MPP population gated by CD34/CD135 antigens in the EphA2 knockout mice compared to wild type control. There were significantly more (P = 0.0012) ST-HSCs in EphA2 knockout bone marrow compared to wild type control (n = 15, 3 independent experiments). (D) There were no significant differences in LT-HSC and MPP population gated using CD48/CD150 antigens in EphA2 knockout mice compared to wild type control (n = 15, 3 independent experiments). Each dot corresponds to one individual mouse. The data represent mean ± SEM. Unpaired t test was performed for statistical analyses.
Mentions: We next analyzed the numbers of immunophenotype-defined hematopoietic stem and progenitor cells (HSPCs) in the EphA2 knockout mice compared to wild type littermates (Fig 2A). This analysis showed no significant differences in the percentage of Lineage-c-Kit+Sca-1+ (LKS+), progenitors and mature myeloid progenitor cells, including common myeloid progenitors (CMPs), megakaryocyte-erythrocyte progenitors (MEPs), and granulocyte-monocyte progenitors (GMPs) (Fig 2B). To further analyze HSCs, CD34 and FLK2 (CD135) markers were utilized to isolate LT-HSCs (LKS+CD34-Flk2-), ST-HSCs (LKS+CD34+Flk2) or MPP, (LKS+CD34+Flk2+) population. A significant increase in the frequency of ST-HSCs (P = 0.0012) was observed in the EphA2 knockout bone marrow; however no significant differences in either the MPPs or the LT-HSCs were evident (Fig 2C). CD150 and CD48 markers were also used as another method to fractionate HSCs, with (LKS+CD150+CD48-) cell population representing LT-HSCs and (LKS+CD150-CD48+) representing MPPs. Similar to the previous results, there were no significant differences in the frequency of LT-HSCs or MPPs in EphA2 knockout mice compared to wild type control mice (Fig 2D). Altogether, these data suggest that EphA2 is not essential for normal blood development.

Bottom Line: These studies showed that EphA2 does not have an obligatory role in normal hematopoiesis.We showed that treatment with EphA2 monoclonal antibody IF7 alone had no effect on tumorigenicity and latency of the MLL-AF9 leukemias, while targeting of EphA2 using EphA2 monoclonal antibody with a radioactive payload significantly impaired the leukemic process.Altogether, these results identify EphA2 as a potential radio-therapeutic target in leukemias with MLL translocation.

View Article: PubMed Central - PubMed

Affiliation: QIMR Berghofer Medical Research Institute, Brisbane, Australia.

ABSTRACT
Members of the Eph family of receptor tyrosine kinases and their membrane bound ephrin ligands have been shown to play critical roles in many developmental processes and more recently have been implicated in both normal and pathological processes in post-embryonic tissues. In particular, expression studies of Eph receptors and limited functional studies have demonstrated a role for the Eph/ephrin system in hematopoiesis and leukemogenesis. In particular, EphA2 was reported on hematopoietic stem cells and stromal cells. There are also reports of EphA2 expression in many different types of malignancies including leukemia, however there is a lack of knowledge in understanding the role of EphA2 in hematopoiesis and leukemogenesis. We explored the role of EphA2 in hematopoiesis by analyzing wild type and EphA2 knockout mice. Mature, differentiated cells, progenitors and hematopoietic stem cells derived from knockout and control mice were analyzed and no significant abnormality was detected. These studies showed that EphA2 does not have an obligatory role in normal hematopoiesis. Comparative studies using EphA2-negative MLL-AF9 leukemias derived from EphA2-knockout animals showed that there was no detectable functional role for EphA2 in the initiation or progression of the leukemic process. However, expression of EphA2 in leukemias initiated by MLL-AF9 suggested that this protein might be a possible therapy target in this type of leukemia. We showed that treatment with EphA2 monoclonal antibody IF7 alone had no effect on tumorigenicity and latency of the MLL-AF9 leukemias, while targeting of EphA2 using EphA2 monoclonal antibody with a radioactive payload significantly impaired the leukemic process. Altogether, these results identify EphA2 as a potential radio-therapeutic target in leukemias with MLL translocation.

No MeSH data available.


Related in: MedlinePlus