Galectins as pivotal components in oncogenesis and immune exclusion in human malignancies.

Galectins are galactoside-binding proteins, exerting numerous functions inside and outside the cell, particularly conferring adaptation to stress factors. For most of them, aberrant expression profiles have been reported in the context of cancer. Albeit not being oncogenic drivers, galectins can be harnessed to exacerbate the malignant phenotype. Their impact on disease establishment and progression is not limited to making cancer cells resistant to apoptosis, but is prominent in the context of the tumor microenvironment, where it fosters angiogenesis, immune escape and exclusion. This review focuses mainly on Gal-1, Gal-3 and Gal-9 for which the involvement in cancer biology is best known. It presents the types of galectin dysregulations, attempts to explain the mechanisms behind them and analyzes the different ways in which they favor tumour growth. In an era where tumour resistance to immunotherapy appears as a major challenge, we highlight the crucial immunosuppressive roles of galectins and the potential therapeutic benefits of combinatorial approaches including galectin inhibition.

Advanced microRNA-based cancer diagnostics using amplified time-gated FRET.

MicroRNAs (miRNAs) play an important role in cellular functions and in the development and progression of cancer. Precise quantification of endogenous miRNAs from different clinical patient and control samples combined with a one-to-one comparison to standard technologies is a challenging but necessary endeavor that is largely neglected by many emerging fluorescence technologies. Here, we present a simple, precise, sensitive, and specific ratiometric assay for absolute quantification of miRNAs. Isothermally amplified time-gated Förster resonance energy transfer (TG-FRET) between Tb donors and dye acceptors resulted in miRNA assays with single-nucleotide variant specificity and detection limits down to 4.2 ± 0.5 attomoles. Quantification of miR-21 from human tissues and plasma samples revealed the relevance for breast and ovarian cancer diagnostics. Analysis of miR-132 and miR-146a from acute monocytic leukemia cells (THP-1) demonstrated the broad applicability to different miRNAs and other types of clinical samples. Direct comparison to the gold standard RT-qPCR showed advantages of amplified TG-FRET concerning precision and specificity when quantifying low concentrations of miRNAs as required for diagnostic applications. Our results demonstrate that a careful implementation of rolling circle amplification and TG-FRET into one straightforward nucleic acid detection method can significantly advance the possibilities of miRNA-based cancer diagnostics and research.

Interferon beta induces apoptosis in nasopharyngeal carcinoma cells via the TRAIL-signaling pathway.

The combination of neoadjuvant chemotherapy, radiochemotherapy, and maintenance therapy with interferon beta (IFNß) has led to superior results in the treatment of children and adolescents with nasopharyngeal carcinoma (NPC). However, nothing is known about the mechanism of the antitumor activity of IFNß in NPC. Here, we investigate the role of IFNß on apoptosis in NPC cells. Six NPC cell lines, one patient-derived NPC xenograft (PDX) and one SV40-transformed nasoepithelial cell line were used. Induction of apoptosis by IFNß was measured by flow cytometric analysis of subG1-DNA-content, Hoechst 33258 staining and activation of caspase-3. Dissection of death ligand signaling pathways included measuring surface expression of its components by flow cytometry, activation by death ligands and neutralization with specific antibodies and siRNA. IFNß induced apoptosis at concentrations achievable in humans in five of six NPC cell lines and in PDX cells but not in nasoepithelial cells. Inhibition of caspases-3 and -8 abrogated this effect suggesting IFNß promoted apoptosis through the extrinsic pathway. IFNß induced surface expression of TRAIL and TRAIL-R2 and the addition of an anti-TRAIL-antibody or transfection with TRAIL-siRNA blocked IFNß-induced apoptosis. No induction of TRAIL-expression was noted in the IFNß-resistant cell line. In conclusion, IFNß leads to apoptosis in NPC cells in an autocrine way via the induction of TRAIL expression and subsequent activation of the TRAIL-signaling pathway. The mechanism described could at least partly explain the clinical benefit of IFNß in the treatment of NPC. Further studies in a mouse-xenograft model are warranted to substantiate this effect in vivo.

Stimulation of the toll-like receptor 3 promotes metabolic reprogramming in head and neck carcinoma cells.

In this study, a possible link between the innate immune recognition receptor TLR3 and metabolic reprogramming in Head and Neck carcinoma (HNC) cells was investigated. The effects of TLR3 stimulation/knock-down were assessed under several culture conditions in 4 HNC cell-lines by cell growth assays, targeted metabolomics, and glycolysis assays based on time-resolved analysis of proton release (Seahorse analyzer). The stimulation of TLR3 by its synthetic agonist Poly(A:U) resulted in a faster growth of HNC cells under low foetal calf serum conditions. Targeted analysis of glucose metabolism pathways demonstrated a tendency towards a shift from tricarboxylic acid cycle (Krebs cycle) to glycolysis and anabolic reactions in cells treated with Poly(A:U). Glycolysis assays confirmed that TLR3 stimulation enhanced the capacity of malignant cells to switch from oxidative phosphorylation to extra-mitochondrial glycolysis. We found evidence that HIF-1α is involved in this process: addition of the TLR3 agonist resulted in a higher cell concentration of the HIF-1α protein, even in normoxia, whereas knocking-down TLR3 resulted in a lower concentration, even in hypoxia. Finally, we assessed TLR3 expression by immunohistochemistry in a series of 7 HNSCC specimens and found that TLR3 was detected at higher levels in tumors displaying a hypoxic staining pattern. Overall, our results demonstrate that TLR3 stimulation induces the Warburg effect in HNC cells in vitro, and suggest that TLR3 may play a role in tumor adaptation to hypoxia.

Plasma miR-200b in ovarian carcinoma patients: distinct pattern of pre/post-treatment variation compared to CA-125 and potential for prediction of progression-free survival.

Ovarian carcinomas (OvCa) are highly heterogeneous malignancies. We investigated four circulating plasma microRNAs (miR-21, miR-34a, miR-200b and miR-205) as candidate biomarkers. Using qPCR, we assessed the plasma concentration of these markers in 101 women, including 51 previously untreated OvCa patients, 25 healthy women and 25 patients bearing benign pelvic lesions. For a subset of 33 OvCa patients, the assay was repeated at the end of the primary treatment. The pattern of variations (post- minus pre-treatment) of concentration was compared to that of CA-125. A Cox regression model was used to study the association between variations and the progression-free survival (PFS). Plasma miR-200b proved to have a greater average concentration in OvCa samples (median 2-ΔΔCt = 15.18) than in samples linked to non-malignant lesions (median 2-ΔΔCt = 1.26, p-value = 0.0004). Its concentration was highly heterogeneous among OvCa patients, without any correlations with the FIGO stage and the pre-treatment CA-125 level. The decrease in CA-125 concentration was constant and often dramatic, while the variations of miR-200b concentration were much more diverse. The variation of miR-200b was marginally associated with the PFS (hazard ratio=2.95 95%CI=[0.94; 9.28], p=0.06) while miR-200b as a continuous time-dependent variable was significantly associated (HR=1.06 [1.02; 1.10], p=0.003). This study is the first direct empirical evidence that miR-200b can provide additional information, independent of CA-125 in OvCa patients.