Malignant tumors exhibit increased dependence on glycolysis, resulting in abundant export of lactic acid, a hypothesized key step in tumorigenesis. Lactic acid is mainly transported by two H + /lactate symporters, MCT1/MCT4, that require the ancillary protein CD147/Basigin for their functionality. First, we showed that blocking MCT1/2 in Ras-transformed fibroblasts with AR-C155858 suppressed lactate export, glycolysis, and tumor growth, whereas ectopic expression of MCT4 in these cells conferred resistance to MCT1/2 inhibition and reestablished tumorigenicty. A mutant-derivative, deficient in respiration (res − ) and exclusively relying on glycolysis for energy, displayed low tumorigenicity. These res − cells could develop resistance to MCT1/2 inhibition and became highly tumorigenic by reactivating their endogenous mct4 gene, highlighting that MCT4, the hypoxia-inducible and tumor-associated lactate/H + symporter, drives tumorigenicity. Second, in the human colon adenocarcinoma cell line (LS174T), we showed that combined silencing of MCT1/MCT4 via inducible shRNA, or silencing of CD147/Basigin alone, significantly reduced glycolytic flux and tumor growth. However, both silencing approaches, which reduced tumor growth, displayed a low level of CD147/Basigin, a multifunctional protumoral protein. To gain insight into CD147/Basigin function, we designed experiments, via zinc finger nuclease-mediated mct4 and basigin knockouts, to uncouple MCTs from Basigin expression. Inhibition of MCT1 in MCT4-null, Basigin high cells suppressed tumor growth. Conversely, in Basigin-null cells, in which MCT activity had been maintained, tumorigenicity was not affected. Collectively, these findings highlight that the major protumoral action of CD147/Basigin is to control the energetics of glycolytic tumors via MCT1/MCT4 activity and that blocking lactic acid export provides an efficient anticancer strategy.

HACE1 is an E3 ubiquitin ligase described as a tumour suppressor because HACE1-knockout mice develop multi-organ, late-onset cancers and because HACE1 expression is lost in several neoplasms, such as Wilms' tumours and colorectal cancer. However, a search of public databases indicated that HACE1 expression is maintained in melanomas. We demonstrated that HACE1 promoted melanoma cell migration and adhesion in vitro and was required for mouse lung colonisation by melanoma cells in vivo. Transcriptomic analysis of HACE1-depleted melanoma cells revealed an inhibition of ITGAV and ITGB1 as well changes in other genes involved in cell migration. We revealed that HACE1 promoted the K27 ubiquitination of fibronectin and regulated its secretion. Secreted fibronectin regulated ITGAV and ITGB1 expression, as well as melanoma cell adhesion and migration. Our findings disclose a novel molecular cascade involved in the regulation of fibronectin secretion, integrin expression and melanoma cell adhesion. By controlling this cascade, HACE1 displays pro-tumoural properties and is an important regulator of melanoma cell invasive properties.

Filaggrin is an epidermal protein involved in skin barrier formation and hydration, whose expression is altered in canine atopic dermatitis (CAD). CAD patients also present an abnormal immune response with an altered expression of antimicrobial peptides (AMPs), such as β-defensins and cathelicidins. Sphingolipids and glycosaminoglycans (GAGs) have been reported to improve the skin barrier in several animal species, including dogs. Our objective was to evaluate the in vitro effects of a sphingomyelin-rich lipid extract (LE), a hyaluronic acid-rich GAG matrix, and their combination, on the expression of filaggrin and human β-defensin 2 (hBD-2). Filaggrin expression was quantified in a reconstructed human epidermis (RHE), and hBD-2 in normal human epidermal keratinocyte (NHEK) cultures. LE and GAGs were tested at 0.02 mg/mL, with or without adding a cytokine mix. A significant increase in mean hBD-2, compared to the control (99 pg/mL) was achieved with LE (138 pg/mL) and LE+GAGs (165 pg/mL). Filaggrin increased with GAGs (202% ± 83) and LE (193% ± 44) vs. the stimulated control, but this difference was statistically significant (p < 0.05) only with LE+GAGs (210% ± 39). In conclusion, the tested GAGs and LE enhance filaggrin and AMP expression in vitro, which might benefit CAD patients if applied in vivo.

Intense conversion of glucose to lactic acid via glycolysis is a feature of rapidly growing cells often encountered in hypoxic tumor microenvironments. To survive and expand, tumor cells must efficiently export lactic acid to maintain intracellular pH. Cells possess several systems for lactic acid extrusion. A family of H+-linked MonoCarboxylate Transporters (MCTs) is represented by the ubiquitously expressed MCT1, a H+/lactate symporter that operates in both directions. MCT4, a close relative of MCT1, is up-regulated by HIF-1 and is highly expressed in aggressive malignant tumors. In addition, the functional expression of MCT1/MCT4 requires the interaction with the glycoprotein CD147/Basigin also known as EMMPRIN, a protumoral protein involved in invasion. Objectives: 1) Demonstrate that tumor growth is dependent on lactic acid export and that both transporters MCT1/MCT4 represent key anticancer targets. 2) Demonstrate whether the protumoral function of Basigin/CD147 is primary linked to lactic acid export or to other invasive functions. 3) Demonstrate whether Metformin, an inhibitor of mitochondrial complex I, sensitizes glycolytic tumor cells to MCTs inhibitors. Methods: We exploited two tumoral models. i) Ras transformed fibroblasts expressing only MCT1/MCT2 and ii) the human colon adenocarcinoma cell line LS174T expressing MCT1 and MCT4. We inhibited MCT1/MCT2 with the specific astraZeneca compound AR-C155858 and knocked-down MCT1, MCT4 and CD147 with inducible shRNAs. In addition we knocked-out mct4 or basigin with Zinc Finger Nucleases in LS174T cells. Results: First we demonstrated that silencing or pharmacological blockage of MCTs, reduced pHi, the rate of glycolysis and tumor growth in mice xenografts. This tumor growth inhibition was recapitulated by a single silencing or knock out of basigin/CD147 gene that also reduced the plasma membrane expression of MCT1 and MCT4 and lactate transport up to 10-fold. Second, to gain insight into CD147/Basigin function, we uncoupled MCTs from Basigin expression. Inhibition of MCT1 in MCT4-null, Basigin-high expressors, suppressed tumor growth. Conversely in Basigin-null cells, in which MCT activity had been maintained, tumorigenicity was not affected. Thirdly, we showed that tumor cells lacking Basigin or MCT4 become highly sensitive to MCT1 inhibition when treated with Metformin. This synthetic lethality demonstrated in vitro and currently being tested in vivo will be discussed. Conclusions: These findings highlight that a major protumoral action of CD147/Basigin is to control the energetics of glycolytic tumors via MCT1/MCT4 activity and that blocking lactic acid export provides an efficient anticancer approach. Furthermore, we reveal that Metformin, by sensitizing normoxic cells to inhibitors of lactic export (MCTs), could offer an interesting novel anticancer strategy for rapidly growing tumors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3225. doi:1538-7445.AM2012-3225