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Nuclear factor-kappaB is constitutively active in C-cell carcinoma and required for RET-induced transformation
L Ludwig1, H Kessler, M Wagner
1Department of Internal Medicine I, University of Ulm, Robert-Koch-Street 8, D-89081 Ulm, Germany.
Abstract:
Specific point mutations of the RET proto-oncogene have been demonstrated to be responsible for multiple endocrine neoplasia (MEN) types 2A and 2B, for familial medullary thyroid carcinoma (MTC) syndromes, as well as for sporadic MTC. Here we show that nuclear factor (NF)-kappaB is activated in RET-associated C-cell carcinoma specimens. TT cells, a human MTC cell line expressing MEN 2A type RET, display transcriptionally active RelA(p65) in the nucleus. NF-kappaB activity in these cells is attributable to constitutive IkappaB kinase (IKK) activity and high turn over of IkappaBalpha. RET harboring the mutations C634R (MEN 2A) or M918T (MEN 2B), in contrast to wild-type RET, activates a NF-kappaB-dependent reporter construct upon transient transfection in HeLa cells. We show that the prototype RET mutation C634R enhances phosphorylation of IkappaBalpha by IKKbeta but not by IKKalpha. RET-induced NF-kappaB and IKKbeta activity requires Ras function but does neither involve the classical mitogen-activated protein kinase kinase/extracellular signal-regulated kinase nor the phosphoinositide 3-kinase/Akt pathways. In contrast, RET-induced NF-kappaB activity is dependent on Raf and MEKK1. Inhibition of constitutive NF-kappaB activity results in cell death of TT cells and blocks focus formation induced by oncogenic forms of RET in NIH 3T3 cells. These results suggest that RET-mediated carcinogenesis critically depends on IKK activity and subsequent NF-kappaB activation.
Insights
RET proto-oncogene mutations activate nuclear factor-kappaB (NF-kappaB) in medullary thyroid carcinoma (MTC). This NF-kappaB activation is crucial for RET-driven cancer development and cell survival.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Specific RET proto-oncogene mutations are linked to Multiple Endocrine Neoplasia (MEN) types 2A and 2B, familial medullary thyroid carcinoma (MTC), and sporadic MTC.
- Nuclear factor-kappaB (NF-kappaB) signaling is implicated in various cancers.
Purpose of the Study:
- To investigate the role of NF-kappaB activation in RET-associated C-cell carcinoma.
- To determine the mechanisms by which RET mutations activate NF-kappaB.
- To assess the therapeutic potential of inhibiting NF-kappaB in MTC.
Main Methods:
- Analysis of NF-kappaB activation in MTC specimens and cell lines (TT cells).
- Transient transfection assays in HeLa cells to assess RET mutant-induced NF-kappaB activation.
- Investigating the involvement of specific kinases (IKKalpha, IKKbeta, Ras, Raf, MEKK1, MAPK/ERK, PI3K/Akt) in RET-mediated NF-kappaB activation.
- Assessing the effect of NF-kappaB inhibition on MTC cell viability and focus formation.
Main Results:
- NF-kappaB is constitutively activated in RET-associated C-cell carcinoma specimens and TT cells.
- RET mutations (C634R and M918T) activate NF-kappaB-dependent reporter constructs.
- RET-induced NF-kappaB activation is dependent on IKKbeta, Ras, Raf, and MEKK1, but not MAPK/ERK or PI3K/Akt pathways.
- Inhibition of NF-kappaB leads to cell death in TT cells and blocks focus formation in NIH 3T3 cells.
Conclusions:
- Constitutive NF-kappaB activation is a key feature of RET-associated MTC.
- RET mutations drive MTC pathogenesis through IKK/NF-kappaB signaling.
- Targeting the IKK/NF-kappaB pathway represents a potential therapeutic strategy for MTC.