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Thyroid cancer (TC) is the most common endocrine tumor, amongst which anaplastic thyroid carcinoma (ATC) is the most deadly. Aurora-A usually functions as oncogenes, and its inhibitor Alisertib exerts a powerful antitumor effect in various tumors. However, the mechanism of Aurora-A in regulating TC cell energy supply remains unclear. In the present study, we demonstrated the antitumor effect of Alisertib and an association between high Aurora-A expression and shorter survival. Multi-omics data and in vitro validation data suggested that Aurora-A induced PFKFB3-mediated glycolysis to increase ATP supply, which significantly upregulated the phosphorylation of ERK and AKT. Furthermore, the combination of Alisertib and Sorafenib had a synergistic effect, further confirmed in xenograft models and in vitro. Collectively, our study provides compelling evidence of the prognostic value of Aurora-A expression and suggests that Aurora-A upregulates PFKFB3-mediated glycolysis to enhance ATP supply and promote TC progression. Combining Alisertib with Sorafenib has huge prospects for application in treating advanced thyroid carcinoma.

Additional file 2: Table S1, Table S2. qRT-PCR primer sequences in this study. Table S3. sgRNA sequences in this study. Table S4. siRNA sequences in this study. Table S5. shRNA sequences in this study. Table S6. Primer sequences in this study

Supplemental material, sj-docx-1-tam-10.1177_17588359221119318 for Efficacy and safety of selpercatinib in Chinese patients with advanced RET-altered thyroid cancers: results from the phase II LIBRETTO-321 study by Xiangqian Zheng, Qinghai Ji, Yuping Sun, Minghua Ge, Bin Zhang, Ying Cheng, Shangtong Lei, Feng Shi, Ye Guo, Linfa Li, Lu Chen, Jingxin Shao, Wanli Zhang and Ming Gao in Therapeutic Advances in Medical Oncology

Supplemental material, sj-docx-1-tam-10.1177_17588359221119318 for Efficacy and safety of selpercatinib in Chinese patients with advanced RET-altered thyroid cancers: results from the phase II LIBRETTO-321 study by Xiangqian Zheng, Qinghai Ji, Yuping Sun, Minghua Ge, Bin Zhang, Ying Cheng, Shangtong Lei, Feng Shi, Ye Guo, Linfa Li, Lu Chen, Jingxin Shao, Wanli Zhang and Ming Gao in Therapeutic Advances in Medical Oncology

Additional file 1: Figure S1. Expression level of AURKA in thyroid cancer cell lines. (A) Quantitative real-time PCR to measure the mRNA level of AURKA in normal thyroid cells and thyroid cancer cells. (B) The protein expression levels of AURKA were examined by western blotting in normal thyroid cells and thyroid cancer cells. Figure S2. AURKA plays oncoprotein roles in PTC. (A)Schematic of CRISPR Cas9 technology. (B) The protein level of AURKA was detected in AURKA knockdown IHH4 cells and AURKA-overexpressing K1 cells by western blotting.(C) The viability of shAURKA-transfected IHH4 cells or AURKA-overexpressing K1 cells compared with control cells was detected using the CCK-8 assay. (D) The ratio of EdU-labelled cells to total cells was observed in AURKA knockdown IHH4 cells and AURKA-overexpressing K1 cells compared with the control group (original magnification x20). (E) shAURKA-transfected IHH4 cells and AURKA overexpressing K1 cells were cultured for 10 days prior to crystal violet staining. (F) Transwell migration assays were used to detect the migration ability of AURKA knockdown IHH4 cells and AURKA-overexpressing K1 cells compared with control cells. (G) The expression levels of AURKA, Ki67 in xenografts of each group were assessed by immunohistochemical staining. The scale bar is 50 μm. Data are shown as the mean ± SD of three replicates (*P < 0.05, **P < 0.01, ***P < 0.001). Figure S3. AURKA affects the protein stability of SIN1. (A)Western blotting validation the P13K/AKT signaling pathway in AURKA knockdown IHH4 cells and AURKA-overexpressing K1 cells. (B) The expression levels of P-AKT in xenografts of each group were assessed by immunohistochemical staining. The scale bar is 50 μm. (C) In TPC1 cells, AURKA (green) was immunofluorescent with SIN1 (red), and AURKA was found to colocalize with SIN1 (yellow). (D) The protein expression levels of SIN1 were examined by western blotting in shAURKA-transfected IHH4 cells and AURKA-overexpressing K1 cells. (E) The expression levels of SIN1 in xenografts of each group were assessed by immunohistochemical staining. The scale bar is 50 μm. (F) The RNA expression levels of SIN1 were examined by quantitative real-time PCR in AURKA-overexpression plasmid-transfected K1 cells or KTC cells and shAURKA-transfected TPC-1 cells and IHH4 cells. Figure S4. CUL4B affects the protein stability of SIN1. (A) The RNA expression levels of Cullins were examined by quantitative real-time PCR in siRNA-transfected TPC-1 cells. (B) The protein expression of SIN1 in TPC-1 cells treated with 200 μM CHX for different durations was measured by western blotting analysis. (C) 293T cells were transfected with the indicated plasmids and siRNA, and the protein expression was examined by western blotting. MG132 (20 μM) was added 6 hrs prior to harvest.