No abstract

RELA, RELB, CREL, NFKB1 and NFKB2, and the upstream regulators NEMO and NIK were knocked-down in lymph endothelial cells (LECs) and in MDA-MB231 breast cancer spheroids to study the contribution of NF-κB in vascular barrier breaching. Suppression of RELA, NFKB1 and NEMO inhibited “circular chemo-repellent induced defects” (CCIDs), which form when cancer cells cross the lymphatic vasculature, by ~20–30%. Suppression of RELB, NFKB2 and NIK inhibited CCIDs by only ~10–15%. In MDA-MB231 cells RELA and NFKB1 constituted MMP1 expression, which caused the activation of PAR1 in adjacent LECs. The knock-down of MMP1 in MDA-MB231 spheroids and pharmacological inhibition of PAR1 in LECs inhibited CCID formation by ~30%. Intracellular Ca2+ release in LECs, which was induced by recombinant MMP1, was suppressed by the PAR1 inhibitor SCH79797, thereby confirming a functional intercellular axis: RELA/NFKB1 – MMP1 (MDA-MB231) – PAR1 (LEC). Recombinant MMP1 induced PAR1-dependent phosphorylation of MLC2 and FAK in LECs, which is indicative for their activity and for directional cell migration such as observed during CCID formation. The combined knock-down of the NF-κB pathways in LECs and MDA-MB231 spheroids inhibited CCIDs significantly stronger than knock-down in either cell type alone. Also the knock-down of ICAM-1 in LECs (a NF-κB endpoint with relevance for CCID formation) and knock-down of MMP1 in MDA-MB231 augmented CCID inhibition. This evidences that in both cell types NF-κB significantly and independently contributes to tumour-mediated breaching of the lymphatic barrier. Hence, inflamed tumour tissue and/or vasculature pose an additional threat to cancer progression.

Acute myeloid leukemia (AML) is a heterogeneous disease with respect to its genetic and molecular basis and to patients´ outcome. Clinical, cytogenetic, and mutational data are used to classify patients into risk groups with different survival, however, within-group heterogeneity is still an issue. Here, we used a robust likelihood-based survival modeling approach and publicly available gene expression data to identify a minimal number of genes whose combined expression values were prognostic of overall survival. The resulting gene expression signature (4-GES) consisted of 4 genes ( SOCS2 , IL2RA , NPDC1 , PHGDH ), predicted patient survival as an independent prognostic parameter in several cohorts of AML patients (total, 1272 patients), and further refined prognostication based on the European Leukemia Net classification. An oncogenic role of the top scoring gene in this signature, SOCS2 , was investigated using MLL-AF9 and Flt3 -ITD/ NPM1c driven mouse models of AML. SOCS2 promoted leukemogenesis as well as the abundance, quiescence, and activity of AML stem cells. Overall, the 4-GES represents a highly discriminating prognostic parameter in AML, whose clinical applicability is greatly enhanced by its small number of genes. The newly established role of SOCS2 in leukemia aggressiveness and stemness raises the possibility that the signature might even be exploitable therapeutically.