The Hypoxic Microenvironment in Acute Myelogenous Leukemia: Critical Role of CXCR4 in the Induction of HIF-1α.

The bone marrow microenvironment provides functional and structural support for both normal and leukemic hematopoietic stem cells. Importantly, the marrow microenvironment is known to be hypoxic. Given the many critical functions of HIF-1α, we investigated HIF-1α levels in leukemic cell lines and primary AML blasts. Surprisingly, HIF-1^α expression was non-detectable in AML cells grown in suspension cultures under normoxic (21% O₂) or hypoxic (1% O₂) conditions. Normoxic co-cultures with bone marrow-derived mesenchymal stromal (MSC) or MS5 cells likewise did not induce HIF-1α, but hypoxic co-culture conditions induced HIF-1α protein without changes in HIF-1α mRNA levels, suggesting post-transcriptional regulation. Functionality of HIF-1α was confirmed by concomitant increase in the levels of glucose transporter glut-1, the HIF-1α downstream target. Inhibition of stroma-leukemia cell interactions with the small molecule CXCR4 inhibitor AMD3465 (Genzyme/Anormed) at 100 nM completely abrogated the induction of HIF-1α in HL-60 and MOLM13 AML cells. While SDF-1 was unable to induced HIF-1α under normoxic conditions, it did so under both physical (1% O₂) and chemical (COCl₂) hypoxic conditions, in two different cell lines. Inhibition of P13K (with LY294002) or MEK/ERK signaling (with CI-1040) abrogated HIF-1α induction under hypoxic conditions. Immunohistochemical staining of bone marrow samples from primary AML confirmed the presence of HIF-1α in leukemic cells localized adjacent to bone-lining stromal elements. Results suggest that

1. the bone marrow microenvironment of AML is hypoxic in vivo;

2. in leukemia cells HIF-1α induction under low oxygen tension depends on the presence of stromal cells;

3. HIF-1α induction is dependent on SDF-1/CXCR4 and is mediated by activation of P13K and MAPK signaling.

Altogether these findings suggest that SDF-1α/CXCR4 interactions contribute to the survival of leukemic cells via specific induction of HIF-1α signaling by the bone marrow microenvironment. Disruption of these interactions via CXCR4 inhibition strategies may suppress multiple pro-survival HIF-1α targets in leukemic cells.