A Heparin-Based Inhibitor of Heparanase Blocks Myeloma Growth In Vivo by Targeting the Tumor Microenvironment.

Heparanase is an enzyme that cleaves heparan sulfate chains of proteoglycans and promotes the growth and metastasis of many types of human tumors. Our previous work demonstrates that enzymatically active heparanase

• is present in the bone marrow of myeloma patients and is associated with a poor prognosis,

• substantially enhances tumor growth and spontaneous metastasis to bone in an animal model of myeloma, and

• increases the synthesis and shedding of syndecan-1 by myeloma cells,

this in turn contributes to myeloma progression by elevating levels of syndecan-1 in the tumor microenvironment. Thus, we hypothesized that inhibitors of heparanase activity would have a dramatic impact on the growth of myeloma tumors. To test this we used a chemically modified form of heparin that is 100% N-acetylated and 25% glycol-split (designated ¹⁰⁰NA,RO-H). This form heparin is a potent inhibitor of heparanase enzyme activity but lacks anticoagulant activity thus enabling use of relatively high doses of the drug in vivo. Delivery of the ¹⁰⁰NA,RO-H to animals bearing established myeloma tumors dramatically blocked tumor growth and progression in a dose-dependent manner(P<0.04). To understand the mechanism of action of ¹⁰⁰NA,RO-H, a series of experiments were performed on tumor tissue harvested from the animals. Results demonstrate that the modified heparin significantly inhibits proliferation of cells within the tumor (33 ± 8/mm² Ki-67 positive cells in treated vs. 688 ± 164/mm² positive cells in controls, P = 0.002). ¹⁰⁰NA,RO-H also dramatically inhibits angiogenesis as compared to controls as assessed by CD34 staining of tumor tissue. This anti-angiogenic effect may be due at least in part to its regulation of hepatocyte growth factor and/or vascular endothelial growth factor, two angiogenic factors that are detected by immunohistochemistry at high levels in controls but virtually absent from tumors of animals treated with ¹⁰⁰NA,RO-H. In vitro studies demonstrate that ¹⁰⁰NA,RO-H blocks syndecan-1 shedding from cells, consistent with the role of heparanase in promoting syndecan-1 shedding. In addition, in contrast to our finding that ¹⁰⁰NA,RO-H blocks tumor growth in vivo, it only slightly inhibited proliferation, cell cycle progression and growth factor signaling in myeloma cells growing in vitro. Thus, the compound does not appear to have substantial direct effects on tumor cells. Although it is not yet clear if all of the effects of ¹⁰⁰NA,RO-H are due to its anti-heparanase activity, we conclude that this modified heparin disrupts the myeloma tumor microenvironment thereby blocking in vivo growth and progression of the cancer. Therefore, its use as a single agent or in combination with agents having direct anti-tumor cell activity may constitute a potent new anti-myeloma therapy.