The Transforming Growth Factor-β Family Members Bone Morphogenetic Protein-2 and Macrophage Inhibitory Cytokine-1 as Mediators of the Antiangiogenic Activity of N-(4-Hydroxyphenyl)Retinamide

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Abstract
Purpose: Tumor growth appears to be an angiogenesis-dependent process. N-(4-hydroxyphenyl)retinamide (fenretinide; 4HPR) has been found to inhibit and/or prevent tumor growth under diverse conditions. Although 4HPR is antiangiogenic, the molecular mechanisms of this effect remain largely unknown. Experimental Design: Endothelial cells were treated with 4HPR in vitro to study the effects on migration, invasion, and organization, as well as gene expression by microarray and quantitative PCR studies. In vivo angiogenesis was evaluated in the Matrigel model. Results: 4HPR treatment substantially modified the biological activities of endothelial cells, repressing their capacity to migrate, invade, and organize into capillary-like structures. The inhibition of invasion induced by 4HPR was also associated with decreased activities of the metalloproteases matrix metalloproteinase-2 and CD13/APN. Using oligonucleotide microarrays, we observed that bone morphogenetic protein-2 and macrophage inhibitory cytokine-1, two multifunctional cytokines of the transforming growth factor-β family that regulate the growth, differentiation, apoptosis, and matrix accumulation of a variety of cells, are up-regulated in vitro by 4HPR. Both these molecules specifically inhibited endothelial cell growth, migration, and invasion in vitro and suppressed angiogenesis in the Matrigel plug assay in vivo. Blocking antibodies to bone morphogenetic protein-2 were able to reverse the suppressive effects of 4HPR in vitro and in vivo. Conclusions: These data support the conclusion that 4HPR inhibits tumor growth by repression of new vessel growth and identify novel points of regulation of angiogenesis in transforming growth factor-β family proteins.