Preclinical Mouse Models of Thyroid Cancer

Division Of Basic Sciences - Nci

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Abstract

Discovered that attenuated MEK signaling curbs cancer stem-like cell activity in human ATC. Combined treatment of ATC patients, harboring the BRAFV600E mutation, with BRAF and MEK combined inhibitors, exhibited some efficacy. However, it is unknown whether cancer stem-like cells (CSC) activity could be inhibited by such treatment. Using ATC cells expressing a fluorescent CSC SORE6 reporter, we showed reduced CSC activity in the rank order of combined >trametinib>dabrafenib through in vitro and in vivo xenograft models. Molecular analyses showed that suppression of CSC activity by these drugs was in part mediated by attenuation of the transcription through dampening the RNA polymerase II activity. The suppression of CSC activity through the attenuating the MEK signaling via inhibitors (ie., trametinib and dabrafenib), could partially account for the efficacy of the combined treatment shown in ATC patients. However, our studies also showed that not all CSC activity was abolished, which may account for the recurrence observed in ATC patients. Our findings provide new insights into the molecular basis of efficacy and limitations of the combined treatment of trametinib and dabrafenib in ATC patients. Demonstrated that M2 polarization of macrophages is induced by combinatory actions of cytokines in anaplastic thyroid Cancer. It has been shown that tumor-associated macrophages (TAMs) contribute to the aggressiveness of ATC. However, stimulatory factors that could facilitate the induction and infiltration of TAMs in the ATC tumor microenvironment (TME) are not fully elucidated. In this study, we used a human leukemia monocytic cell line (THP-1) to study the differentiation of THP-1 into M2-like macrophages (M2) by conditioned media (CM) derived from each of the three human ATC cells: 8505C, THJ-11T (11T), and THJ-16T (16T). The capacity of CM to induce M2 was in the order of 16T>8505C>11T cells as determined by the expression of M2 markers (CD163, CD204, and CCL13). Cytokine arrays and ELISA assays revealed five commonly enriched cytokines (IL-6, IL-8, MCP-1, TIMP-1, and TGF-beta1) in the CM derived from each of the three ATC cells. These cytokines, individually, had weak activity, but together, they mimicked full CM activity in the induction of M2. Further, they collaboratively activated STAT3, ERK, and PI3K-AKT signaling to facilitate the induction of M2 as found in CM. Importantly, we found that the CM-induced M2 could secrete soluble growth factors to promote ATC cell proliferation as evidenced by the increased Ki-67, cMYC, and cyclin D1 protein levels. Our studies identified the major stimulatory cytokines which acted collaboratively to induce M2 in the TME. Importantly, the present studies indicate that when using inhibitors to targeting TAMs, combination therapies would be required for effective treatment of ATC. Elucidated the mechanisms by which p53 acts to lessen aggressiveness of ATC tumor progression. Mutations of the tumor suppressor, p53, are known to occur at a late stage of ATC development and are highly associated with lethal outcomes of ATC. We therefore evaluated whether wild type p53 (WTp53) mitigates ATC aggressive progression. We used human 8505C cells (from human ATC tumors) as a model, harboring BRAFV600E mutation and one allele of mutated p53Arg248Gly. We exogenously expressed WTp53 or mutant TP53C742G into 8505C cells (8505C-WTp53 or 8505C-MTp53, respectively). The expressed WTp53 inhibited cell proliferation, decreased cell migration, and induced apoptosis via induction of proapoptotic WTp53 target BAX and PUMA genes in vitro and in vivo xenograft mouse models. Single-cell RNA-sequencing of tumors induced by 8505C, 8505C-WTp53, and 8505C-MTp53 cells demonstrated differential expression gene (DEG) patterns between 8505C-WTp53 and 8505C tumors. DEG analysis identified alteration of multiple pathways, leading to attenuation of the oncogenic actions of mutant p53. The discovery of the suppression of TNFbeta via the NFkepaB pathway topped the pathways list, which resulted in subduing the deleterious inflammatory responses caused by mutant p53. Our finding that exogenously expressed WTp53 could counteract the oncogenic actions of p53 has heightened the feasibility of using CRISPR/Cas9 genome editing to modify the p53 alleles for potential treatment of ATC patients.