Description
Recent studies suggest a minority population of primitive cells, termed tumor-initiating cells (TICs), resist chemotherapy and re-create heterogeneous tumors to cause relapse. Identifying the mechanisms that support this elusive population will be critical for preventing relapse. Our lab has identified a critical role for classical and alternative NF-κB transcription factors in supporting ovarian cancer progression. We have found that alternative transcription factor, RelB, maintains quiescent ovarian TICs, whereas classical transcription factor, RelA, maintains a more proliferative subpopulation that may depend on MAPK/ERK. Presumably, both populations are required for efficient relapse. Here, I investigate activation of the NF-κB pathway in TICs and non-TICs to distinguish its role in maintaining proliferation and whether this process requires co-activation of MAPK/ERK. Previous RNA-Sequencing studies showed that relative to TIC cultures, non-TIC cultures had increased proliferation pathways, which included MAPK genes ERK2 and MEKK that were significantly downregulated in RelA shRNA knockdown cells, implying RelA regulation. To further investigate RelA, RelB and MAPK/ERK activity in TICs and non-TICs, I cultured ovarian cancer cells in standard 2D conditions and in 3D spheroid forming conditions that enrich for TICs to assess growth, spheroid formation and gene and protein expression changes under these different growth conditions. I validated the role of NF-κB and MAPK using knockdowns of RelA and RelB and the MEK inhibitor selumentinib, respectively. I isolated CD117+ TICs and CD117- non-TICs to characterize NF-κB reporter activity and drug resistance. Lastly, I completed ChIP sequencing experiments to understand gene targets shared by MAPK and NF-κB. Altogether, my data suggest the MAPK/ERK pathway maintains growth of non-TICs while both NF-κB transcription factors maintain growth of TICs and these are being validated in relapse model of ovarian cancer. Although only quiescent TICs remain following chemotherapy, efficient relapse will also require a proliferative population dependent on MAPK/ERK. A better understanding of NF-κB and MAPK/ERK signaling in TICs and proliferative non-TICs will guide the design for more effective therapies to overcome chemoresistance and relapse.
