Glalectin-1-glycan interactions, links Hif-1a-independent hypoxia and angiogenesis in Kaposi sarcoma

CROCI DO; SALATINO M; RUBINSTIEN N; ILARREGUI JM; TOSCANO MA; A. ALBINI; RABINOVICH GA

San Diego, California

Simposio; THE 12TH INTERNATIONAL SYMPOSIUM ON ANTI ANGIOGENIC AGENTS; 2010

Angiogenesis is critical for tumor progression. We previously demonstrated that galectin-1(Gal-1), controls tumor growth by favoring tumor-immune escape. The aim of this study was to investigate, using in vitro and in vivo strategies, the role of Gal-1 in the control of tumor angiogenesis and to analyze the mechanisms underlying this effect.To examine the contribution of Gal-1 to tumor angiogenesis in vivo we inhibited Gal-1 expression by silencing-gene strategies in human Kaposi sarcoma tumor cell lines. Targeted inhibition of gal-1 gene expression inhibited the formation of new blood vessels and suppressed tumor growth in vivo (p<0,05). Expression of Gal-1 in Kaposi sarcoma (KS) cells was found to be up-regulated under hypoxic conditions, both in vitro and in vivo. This over expression was dependent of NFkB but not  HIF-1a activity, This up-regulated expression was dependent on NF-kB but not on HIF-1a activity since inhibitor of IkB phosphorylation BAY 11-7082 (1.5 µM) but not Hif-1a (2µM) prevented hypoxia-induced expression of Gal-1 (p<0.05) . Moreover, conditioned medium from KS hypoxic cells was more effective at inducing tubulogenesis and invasion of endothelial cells (p<0.05). Interestingly, this effect was abrogated when KS hypoxic cells were infected with a retrovirus encoding shRNA to gal-1(p<0.01). This result was confirmed in vivo using a matrigel sponges model in which hypoxic MC-KS were more effective to induce hemoglobin accumulation in matrigel plugs (p<0.05 hypoxic MC-KS vs normoxic MC-KS).  We next addressed the effect of Gal-1 on endothelial cells. Recombinant Gal-1 bound preferentially N-glycan motives in human umbilical vein endothelial cells (HUVEC) in a dose-dependent manner (p<0.01), and promoted tubulogenesis, (p<0.01) proliferation (p<0.05) and migration/invasion (p<0.05) of HUVEC through binding to VEGFR2. A mechanistic analysis of the intracellular signals involved in this effects revealed a substantial increase in phosphorylated-Akt and Erk ½ following Gal-1 treatment (F.I>3). Pretreatment of HUVEC with either PI3K-inhibitor LY294.002 (2µM) or Erk1/2-inhibitor U0126 (5µM) but not NFkB, p38 or JNKs inhibitors, abolished the ability of Gal-1 to induce tubulogenesis (p<0.01), proliferation (p<0.05) and invasion (p<0.05).Our results suggest that hypoxia-regulated Gal-1 is a key modulator of tumor angiogenesis by promoting tubulogenesis, proliferation and invasion of endothelial cells. In addition, we propose a model in which Gal-1 regulated by hypoxia is secreted to the extracellular milieu to regulate angiogenesis and tumor-immune escape, thus influencing tumor progression.