Sofia Nascimento dos Santos

Galectin-3 disruption impaired tumoral angiogenesis by reducing VEGF secretion from TGFβ1-induced macrophages.

In order to study the role of galectin‐3 in tumor angiogenesis associated with tumor‐associated macrophages (TAM) and tumor parenchyma, the galectin‐3 expression was reconstituted in Tm1 melanoma cell line that lacks this protein. Galectin‐3‐expressing cells (Tm1G3) and mock‐vector transfected cells (Tm1N3) were injected into wild‐type (WT) and galectin‐3 knockout (KO) C57Bl/6 mice. Tumors originated from Tm1G3 were larger in tumor volume with enlarged functional vessels, decreased necrotic areas, and increased vascular endothelial growth factor (VEGF) protein levels. Galectin‐3‐nonexpressing‐cells injected into WT and KO showed increased levels of transforming growth factor beta 1 (TGFβ1) and, in WT animals this feature was also accompanied by increased VEGFR2 expression and its phosphorylation. In KO animals, tumors derived from galectin‐3‐expressing cells were infiltrated by CD68+‐cells, whereas in tumors derived from galectin‐3‐nonexpressing‐cells, CD68+ cells failed to infiltrate tumors and accumulated in the periphery of the tumor mass. In vitro studies showed that Tm1G3 secreted more VEGF than Tm1N3 cells. In the latter case, TGFβ1 induced VEGF production. Basal secretion of VEGF was higher in WT‐bone marrow‐derived macrophages (BMDM) than in KO‐BMDM. TGFβ1 induced secretion of VEGF only in WT‐BMDM. Tm1G3‐induced tumors had the Arginase I mRNA increased, which upregulated alternative macrophage (M2)/TAM induction. M2 stimuli, such as interleukin‐4 (IL4) and TGFβ1, increased Arginase I protein levels and galectin‐3 expression in WT‐ BMDM, but not in cells from KO mice. Hence, we report that galectin‐3 disruption in tumor stroma and parenchyma decreases angiogenesis through interfering with the responses of macrophages to the interdependent VEGF and TGFβ1 signaling pathways.

Galectin-3 acts as an angiogenic switch to induce tumor angiogenesis via Jagged-1/Notch activation

Angiogenesis is a coordinated process tightly regulated by the balance between Delta-like-4 (DLL4) and Jagged-1 (JAG1) in endothelial cells. Here we show that galectin-3 (gal-3), a glycan-binding protein secreted by cancer cells under hypoxic conditions, triggers sprouting angiogenesis, assisted by hypoxic changes in the glycosylation status of endothelial cells that enhance binding to gal-3. Galectin-3′s proangiogenic functions were found to be predominantly dependent on the Notch ligand JAG1. Differential direct binding to JAG1 was shown by surface plasmon resonance assay. Upon binding to Notch ligands, gal-3 preferentially increased JAG1 protein half-life over DLL4 and preferentially activated JAG1/Notch-1 signaling in endothelial cells. JAG1 overexpression in Lewis lung carcinoma cells accelerated tumor growth in vivo, but this effect was prevented in Lgals3−/− mice. Our findings establish gal-3 as a molecular regulator of the JAG1/Notch-1 signaling pathway and have direct implications for the development of strategies aimed at controlling tumor angiogenesis.

Lack of galectin-3 increases Jagged1/Notch activation in bone marrow-derived dendritic cells and promotes dysregulation of T helper cell polarization.

Galectin-3, an endogenous glycan-binding protein, is abundantly expressed at sites of inflammation and immune cell activation. Although this lectin has been implicated in the control of T helper (Th) polarization, the mechanisms underlying this effect are not well understood. Here, we investigated the role of endogenous galectin-3 during the course of experimental Leishmania major infection using galectin-3-deficient (Lgals3(-/-)) mice in a BALB/c background and the involvement of Notch signaling pathway in this process. Lgals3(-/-) mice displayed an augmented, although mixed Th1/Th2 responses compared with wild-type (WT) mice. Concomitantly, lymph node and footpad lesion cells from infected Lgals3(-/-) mice showed enhanced levels of Notch signaling components (Notch-1, Jagged1, Jagged2 and Notch target gene Hes-1). Bone marrow-derived dendritic cells (BMDCs) from uninfected Lgals3(-/-) mice also displayed increased expression of the Notch ligands Delta-like-4 and Jagged1 and pro-inflammatory cytokines. In addition, activation of Notch signaling in BMDCs upon stimulation with Jagged1 was more pronounced in Lgals3(-/-) BMDCs compared to WT BMDCs; this condition resulted in increased production of IL-6 by Lgals3(-/-) BMDCs. Finally, addition of exogenous galectin-3 to Lgals3(-/-) BMDCs partially reverted the increased sensitivity to Jagged1 stimulation. Our results suggest that endogenous galectin-3 regulates Notch signaling activation in BMDCs and influences polarization of T helper responses, thus increasing susceptibility to L. major infection.

Lack of galectin-3 modifies differentially Notch ligands in bone marrow and spleen stromal cells interfering with B cell differentiation

Galectin-3 (Gal-3) is a β-galactoside binding protein that controls cell-cell and cell-extracellular matrix interactions. In lymphoid organs, gal-3 inhibits B cell differentiation by mechanisms poorly understood. The B cell development is dependent on tissue organization and stromal cell signaling, including IL-7 and Notch pathways. Here, we investigate possible mechanisms that gal-3 interferes during B lymphocyte differentiation in the bone marrow (BM) and spleen. The BM of gal-3-deficient mice (Lgals3−/− mice) was evidenced by elevated numbers of B220+CD19+c-Kit+IL-7R+ progenitor B cells. In parallel, CD45− bone marrow stromal cells expressed high levels of mRNA IL-7, Notch ligands (Jagged-1 and Delta-like 4), and transcription factors (Hes-1, Hey-1, Hey-2 and Hey-L). The spleen of Lgals3−/− mice was hallmarked by marginal zone disorganization, high number of IgM+IgD+ B cells and CD138+ plasma cells, overexpression of Notch ligands (Jagged-1, Delta-like 1 and Delta-like 4) by stromal cells and Hey-1. Morever, IgM+IgD+ B cells and B220+CD138+ CXCR4+ plasmablasts were significantly increased in the BM and blood of Lgals3−/− mice. For the first time, we demonstrated that gal-3 inhibits Notch signaling activation in lymphoid organs regulating earlier and terminal events of B cell differentiation.

Magnetic core mesoporous silica nanoparticles doped with dacarbazine and labelled with 99mTc for early and differential detection of metastatic melanoma by single photon emission computed tomography

Abstract Cancer is responsible for more than 12% of all causes of death in the world, with an annual death rate of more than 7 million people. In this scenario melanoma is one of the most aggressive ones with serious limitation in early detection and therapy. In this direction we developed, characterized and tested in vivo a new drug delivery system based on magnetic core-mesoporous silica nanoparticle that has been doped with dacarbazine and labelled with technetium 99 m to be used as nano-imaging agent (nanoradiopharmaceutical) for early and differential diagnosis and melanoma by single photon emission computed tomography. The results demonstrated the ability of the magnetic core-mesoporous silica to be efficiently (>98%) doped with dacarbazine and also efficiently labelled with 99mTc (technetium 99 m) (>99%). The in vivo test, using inducted mice with melanoma, demonstrated the EPR effect of the magnetic core-mesoporous silica nanoparticles doped with dacarbazine and labelled with technetium 99 metastable when injected intratumorally and the possibility to be used as systemic injection too. In both cases, magnetic core-mesoporous silica nanoparticles doped with dacarbazine and labelled with technetium 99 metastable showed to be a reliable and efficient nano-imaging agent for melanoma. Graphical Abstract

O-glycan sialylation alters galectin-3 subcellular localization and decreases chemotherapy sensitivity in gastric cancer

ST6GalNAc-I, the sialyltransferase responsible for sialyl-Tn (sTn) synthesis, has been previously reported to be positively associated with cancer aggressiveness. Here we describe a novel sTn-dependent mechanism for chemotherapeutic resistance. We show that sTn protects cancer cells against chemotherapeutic-induced cell death by decreasing the interaction of cell surface glycan receptors with galectin-3 and increasing its intracellular accumulation. Moreover, exogenously added galectin-3 potentiated the chemotherapeutics-induced cytotoxicity in sTn non-expressing cells, while sTn overexpressing cells were protected. We also found that the expression of sTn was associated with a reduction in galectin-3-binding sites in human gastric samples tumors. ST6GalNAc-I knockdown restored galectin-3-binding sites on the cell surface and chemotherapeutics sensibility. Our results clearly demonstrate that an interruption of O-glycans extension caused by ST6GalNAc-I enzymatic activity leads to tumor cells resistance to chemotherapeutic drugs, highlighting the need for the development of novel strategies to target galectin-3 and/or ST6GalNAc-I.

Diagnosing lung cancer using etoposide microparticles labeled with 99mTc

Abstract The diagnosis of lung cancer mostly occurs when the cancer is already in an advanced stage. In this situation, there are few options for the treatment and most of them have few chances of success. In this study, we developed and tested etoposide microparticles as a diagnostic agent for imaging lung cancer at early stages of development. We tested etoposide microparticles labeled with technetium 99m in inducted mice. The results demonstrated that over 10% of the total dose used was uptake by the tumor site. Also, the results showed that the microparticles had a good renal clearance and low uptake by liver and spleen. The data suggest that these micro-radiopharmaceuticals may be used for lung cancer imaging exam, especially single-photo emission computed tomography (SPECT).

Characterization of the mechanisms underlying the crosstalk between galectins and notch in gastric cancer

Background Gastric cancer is the fourth most common cancer and the second leading cause of cancer-related deaths worldwide. Galectins form a family of b-galactosides binding proteins that recognize a variety of glycan-containing proteins at the cell surface and are overexpressed in various tumors, including gastric cancer. Galectins overexpression as well as changes in their subcellular distribution has been associated with gastric cancer progression and poor prognosis. It is not well understood, however, how the interaction between galectins and glycosylated receptors modulates tumor development and growth. Since Notch receptors and ligands contain glycan structures known to bind galectins, we aim to demonstrate that galectins expression in the tumor microenvironment may interfere with Notch signaling activation during tumor development and progression.