Inhibition of nuclear factor-kappa B differentially affects thyroid cancer cell growth, apoptosis, and invasion

<p>Abstract</p> <p>Background</p> <p>Nuclear factor-κB (NF-κB) is constitutively activated in many cancers and plays a key role in promoting cell proliferation, survival, and invasion. Our understanding of NF-κB signaling in thyroid cancer, however, is limited. In this study, we have investigated the role of NF-κB signaling in thyroid cancer cell proliferation, invasion, and apoptosis using selective genetic inhibition of NF-κB in advanced thyroid cancer cell lines.</p> <p>Results</p> <p>Three pharmacologic inhibitors of NF-κB differentially inhibited growth in a panel of advanced thyroid cancer cell lines, suggesting that these NF-κB inhibitors may have off-target effects. We therefore used a selective genetic approach to inhibit NF-κB signaling by overexpression of a dominant-negative IκBα (mIκBα). These studies revealed decreased cell growth in only one of five thyroid cancer cell lines (8505C), which occurred through a block in the S-G2/M transition. Resistance to TNFα-induced apoptosis was observed in all cell lines, likely through an NF-κB-dependent mechanism. Inhibition of NF-κB by mIκBα sensitized a subset of cell lines to TNFα-induced apoptosis. Sensitive cell lines displayed sustained activation of the stress-activated protein kinase/c-Jun NH2-terminal kinase (SAPK/JNK) pathway, defining a potential mechanism of response. Finally, NF-κB inhibition by mIκBα expression differentially reduced thyroid cancer cell invasion in these thyroid cancer cell lines. Sensitive cell lines demonstrated approximately a two-fold decrease in invasion, which was associated with differential expression of MMP-13. MMP-9 was reduced by mIκBα expression in all cell lines tested.</p> <p>Conclusions</p> <p>These data indicate that selective inhibition of NF-κB represents an attractive therapeutic target for the treatment of advanced thyroid. However, it is apparent that global regulation of thyroid cancer cell growth and invasion is not achieved by NF-κB signaling alone. Instead, our findings suggest that other important molecular processes play a critical role in defining the extent of NF-κB function within cancer cells.</p

Fibronectin Contributes to a Braf Inhibitor-Driven Invasive Phenotype in Thyroid Cancer Through EGR1, Which Can Be Blocked by Inhibition of ERK1/2

Mutations in BRAF are common in advanced papillary and anaplastic thyroid cancer (PTC and ATC). However, patients with BRAF-mutant PTC currently lack therapies targeting this pathway. Despite the approved combination of BRAF and MEK1/2 inhibition for patients with BRAF-mutant ATC, these patients often progress. Thus, we screened a panel of BRAF-mutant thyroid cancer cell lines to identify new therapeutic strategies. We showed that thyroid cancer cells resistant to BRAF inhibition (BRAFi) exhibit an increase in invasion and a proinvasive secretome in response to BRAFi. Using reverse-phase protein array (RPPA), we identified a nearly 2-fold increase in expression of the extracellular matrix protein, fibronectin, in response to BRAFi treatment, and a corresponding 1.8- to 3.0-fold increase in fibronectin secretion. Accordingly, the addition of exogenous fibronectin phenocopied the BRAFi-induced increase in invasion while depletion of fibronectin in resistant cells resulted in loss of increased invasion. We further showed that BRAFi-induced invasion can be blocked by inhibition of ERK1/2. In a BRAFi-resistant patient-derived xenograft model, we found that dual inhibition of BRAF and ERK1/2 slowed tumor growth and decreased circulating fibronectin. Using RNA sequencing, we identified EGR1 as a top downregulated gene in response to combined BRAF/ERK1/2 inhibition, and we further showed that EGR1 is necessary for a BRAFi-induced increase in invasion and for induction of fibronectin in response to BRAFi. Implications: Together, these data show that increased invasion represents a new mechanism of resistance to BRAF inhibition in thyroid cancer that can be targeted with an ERK1/2 inhibitor

Thyroid cancer cell lines: Critical models to study thyroid cancer biology and new therapeutic targets

Thyroid cancer is the most common endocrine malignancy and the incidence is rising. Currently, there are no effective treatments for patients with advanced forms of thyroid cancer. Anaplastic thyroid represents the most severe form of the disease with 95% mortality at 6 months. It is therefore critical to better understand the mechanisms involved in thyroid cancer development and progression in order to develop more effective therapeutic strategies. Cell lines derived from thyroid tumors represent a critical tool to understand the oncogenic mechanisms driving thyroid cancer, as well as preclinical tools to study the efficacy of new therapies in vitro and in vivo. For thyroid cancer, the development of new therapies has been hampered by the lack of thyroid cancer cell lines in the widely used NCI-60 panel which has been used to screen over 100,000 anti-cancer drugs. In addition, the recent discovery that ~20 out of 40 existing thyroid cancer cell lines are either redundant or misidentified with cell lines of other tissue lineages has further hampered progress in the field. Of the available cell lines, 23 were identified as unique and presumably of thyroid origin based on the expression of thyroid-specific genes. Thus, there is a great need for validated thyroid cancer cell lines representing different stages of disease in addition to distinct oncogenic mutations. New, authenticated thyroid cancer cell lines are beginning to be developed, adding to the tools available to study genes and pathways important for thyroid cancer pathogenesis. In summary, the use of validated thyroid cancer cell lines which closely recapitulate disease is critical for the discovery of new drug targets and ultimately new therapies

The effects of Aurora Kinase inhibition on thyroid cancer growth and sensitivity to MAPK-directed therapies

ABSTRACTThyroid cancer is one of the deadliest endocrine cancers, and its incidence has been increasing. While mutations in BRAF are common in thyroid cancer, advanced PTC patients currently lack therapeutic options targeting the MAPK pathway, and despite the approved combination of BRAF and MEK1/2 inhibition for BRAF-mutant ATC, resistance often occurs. Here, we assess growth and signaling responses to combined BRAF and MEK1/2 inhibition in a panel of BRAF-mutant thyroid cancer cell lines. We first showed that combined BRAF and MEK1/2 inhibition synergistically inhibits cell growth in four out of six of the ­BRAF-mutant thyroid cancer cell lines tested. Western blotting showed that the MAPK pathway was robustly inhibited in all cell lines. Therefore, to identify potential mechanisms of resistance, we performed RNA-sequencing in cells sensitive or resistant to MEK1/2 inhibition. In response to MEK1/2 inhibition, we identified a downregulation of Aurora Kinase B (AURKB) in sensitive but not resistant cells. We further demonstrated that combined MEK1/2 and AURKB inhibition slowed cell growth, which was phenocopied by inhibiting AURKB and ERK1/2. Finally, we show that combined AURKB and ERK1/2 inhibition induces apoptosis in BRAF-mutant thyroid cancer cell lines, together suggesting a potential combination therapy for BRAF-mutant thyroid cancer patients

Inhibition of Src with AZD0530 Reveals the Src-Focal Adhesion Kinase Complex as a Novel Therapeutic Target in Papillary and Anaplastic Thyroid Cancer

Context: Focal adhesion kinase (FAK) and Src are overexpressed and activated in many cancers and have been associated with tumor progression. The role of the Src-FAK complex has not been characterized in papillary and anaplastic thyroid cancer (PTC and ATC)

PPARγ Promotes Growth and Invasion of Thyroid Cancer Cells

Undifferentiated (anaplastic) thyroid cancer (ATC) is one of the most aggressive human malignancies and no effective therapy is currently available. We show here that PPARγ levels are elevated in cells derived from ATC. Depletion of PPARγ in HTh74 ATC cells resulted in decreased cell growth, cell cycle arrest and a reduction in pRb and cyclin A and B1 levels. We further showed that both flank and orthotopic thyroid tumors derived from PPARγ-depleted cells grew more slowly than PPARγ-expressing cells. When PPARγ was overexpressed in more differentiated thyroid cancer BCPAP cells which lack PPARγ, there was increased growth and raised pRb and cyclin A and B1 levels. Finally, PPARγ depletion in ATC cells decreased their invasive capacity whereas overexpression in PTC cells increased invasiveness. These data suggest that PPARγ may play a detrimental role in thyroid cancer and that targeting it therapeutically may lead to improved treatment of advanced thyroid cancer