Increased Bone Marrow Microvascular Density in Hematologic Malignancies

Abstract

Background: Angiogenesis is a potential target for therapy, both in solid tumors and in hematologic malignancies. However, the regulation of angiogenesis is complex, different cancers are inherently heterogeneous, and these drugs target different angiogenic regulators. Therefore, a detailed characterization of angiogenesis in individual cancers is needed.Materials and methods: The study cohort consisted of 93 consecutive patients with hematologic neoplasia; i.e. acute myeloid leukemia (n=20), chronic lymphatic leukemia (n=14), multiple myeloma (n=11), and non-Hodgkin's lymphoma (NHL, n=48). We determined the microvessel density (MVD) in bone marrow biopsy specimens by immunohistochemical staining for CD34. In addition, we measured the plasma concentrations of eight putative angiogenesis regulators using Luminex multiplex assay, and the expression in peripheral blood mononuclear cells (PBMNC) of 40 angiogenesisrelated mRNAs using quantitative RT-PCR.Results: Untreated patients had increased bone marrow MVD (mean 39 microvessels/mm2) compared to healthy subjects (mean 20 microvessels/mm2; p<0.001). In particular, patients with advanced stage disease had increased bone marrow MVD (mean 45 microvessels/mm2) compared to patients with lower stage disease (mean 35 microvessels/mm2, p=0.030). A novel finding was that patients with NHL also had significantly increased bone marrow MVD (mean 38 microvessels/mm2). The patients had increased plasma levels of vascular endothelial growth factor (VEGF; mean 31 vs 19 pg/ml, p<0.001), interleukin (IL)-6 (median 4.6 vs 2.3 pg/ml, p=0.034) and IL-8 (median 7.2 vs 3.9 pg/ml, p=0.001). Plasma levels of fibroblast growth factor 2 (FGF2), tumor necrosis factor α, and angiogenin were not significantly different in patients compared to controls. Pre-treatment plasma levels of VEGF and FGF2 were significantly higher in those who did not achieve complete remission after cancer therapy. Patients with advanced stage disease had significantly higher plasma levels of IL-6 and -8. The mRNA most highly expressed in PBMNC was IL8, which was 15-fold up-regulated. The mRNA expression of VEGFA in PBMNC was strongly correlated to the expressions of hypoxia-inducible factors 1α and 2α (r=0.7 and p<0.001 for both). There were no differences in bone marrow MVD when comparing the different diagnoses. However, the majority of the putative regulators of angiogenesis studied were significantly different between the diagnoses; indicating that bone marrow MVD in hematologic malignancies was differentially regulated.Conclusions: Our data show that patients with hematologic malignancies have increased bone marrow MVD, supporting the notion that bone marrow angiogenesis plays a role in the pathogenesis and progression of these cancers. VEGF, IL-6 and IL-8 seem to contribute to the malignant phenotype.