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Background: Pharmacologic targeting of components of the MAPK/ERK pathway in differentiated thyroid carcinoma (DTC) is often limited due to the development of adaptive resistance. However, the detailed mechanism of MEK inhibitor (MEKi) resistance is not fully understood. Methods: RNA-seq of the constructed MEKi-resistant DTC cell line BCPAP-R (compared with the BCPAP line) was performed to investigate the intrinsic mechanism of drug resistance. Colony formation assays, cell viability assays, and cell cycle analysis were performed, and various murine models, including xenograft models, long-term MEKi-treated models, and transgenic model were used to evaluate the treatment effect of combination therapy (SHP099 and selumetinib). Findings: MEKi-resistant models were constructed successfully, in which multiple receptor tyrosine kinases (RTKs) signaling pathways and Src-homology 2 domain-containing phosphatase 2 (SHP2) were activated in MEKi-resistant cells. Given the physiological role of SHP2 as the downstream target of many RTKs, we first found that blockade of SHP2 enhanced the sensitivity to MEKi in constructed MEKi-resistant thyroid cancer models (including constructed cell lines and a long-term MEKi-treated murine model). Interestingly, we also found that compared with MEKi treatment alone, MEKi in combination with an SHP2 inhibitor markedly suppressed the reactivation of the MEK/ERK pathway; thus, the addition of the SHP2 inhibitor significantly improved the anti-tumor effects of MEKi. The synergistic suppression of DTC upon treatment with both inhibitors was futher confirmed in xenograft models and transgenic models. Interpretation: RTKs activation leads to reactivation of the MAPK pathway and resistance to MEKi in DTC, which is reversed by SHP2 blockade. As a novel active inhibitor of SHP2, SHP099 in combination with selumetinib is a promising therapeutic approach for advanced DTC and MEKi-resistant one. Funding Statement: This work was partially supported by grants from the National Natural Science Foundation of China (GRANT no. 81872169), Tianjin Key Research and Development Program Science and Technology Support Key Projects (grant no. 17YFZCSY00690), and Tianjin Municipal Science and Technology Project (grant no. 19JCYBJC27400). Declaration of Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Ethics Approval Statement: All animal experiments were approved by the Tianjin Medical University Cancer Institute and the Hospital Animal Care and Use Committee and performed according to the IACUC protocol.

Thyroid cancer (THCA) is a heterogeneous disease with multiple clinical outcomes Immune cells regulate its progression. Three immunomolecular subtypes (C1, C2, C3) were identified in gene expression data sets from TCGA and GEO databases. Among them, subtype C3 had highest frequency of BRAF mutations, lowest frequency of RAS mutations, highest mutation load and shorter progression-free survival. Functional enrichment analysis for the genes revealed that C1 was up-regulated in the ERK cascade pathway, C2 was up-regulated in cell migration and proliferation pathways, while C3 was enriched in body fluid, protein regulation and response to steroid hormones functions. Notably, the three molecular subtypes exhibit differences in immune microenvironments as shown by timer database and analysis of immune expression signatures. The abundance of B_cell, CD4_Tcell, Neutrophil, Macrophage and Dendritic cells in C2 subtype were lower than in C1 and C3 subtypes Leukocyte fraction, proliferation macrophage regulation, lymphocyte infiltration, IFN gamma response and TGF beta response scores were significantly higher in C3 compared with C1 and C2 subtypes. Unlike C3 subtype, it was observed that C1 and C2 subtypes were significantly negatively correlated with most immune checkpoint genes in two different cohorts. The characteristic genes were differentially expressed between cancer cells, adjacent tissues, and metastatic tissues in different cohorts. In summary, THCA can be subclassified into three molecular subtypes with distinct histological types, genetic and transcriptional phenotypes, all of which have potential clinical implications.

Background Celastrol, a triterpene present in the traditional Chinese medicine (TCM) Triptergium wilfordii, has been demonstrated to have remarkable anticancer activity. However, its specific mechanism on papillary thyroid carcinoma (PTC) remains to be elucidated. Methods Potential targets of celastrol were screened from public databases. Through the Gene Expression Omnibus (GEO) online database, we obtained the bioinformatics analysis profile of PTC, GSE33630, and analyzed the differentially expressed genes (DEGs). Then, a protein-protein interaction (PPI) network was constructed by utilizing the STRING database. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were conducted. Finally, drug interactions between hub genes and celastrol were verified by molecular docking. Results Four core nodes (MMP9, JUN, ICAM1, and VCAM1) were discerned via constructing a PPI network of 47 common targets. Through functional enrichment analysis, it was confirmed that the above target genes were basically enriched in the interleukin-17 (IL-17), nuclear factor kappa-B (NF-κB), and tumor necrosis factor (TNF) signaling pathways, which are involved in the inflammatory microenvironment to inhibit the development and progression of tumors. Molecular docking results demonstrated that celastrol has a strong binding efficiency with the 4 key proteins. Conclusions In this research, it was demonstrated that celastrol can regulate a variety of proteins and signaling pathways against PTC, providing a theoretical basis for future clinical applications.

The aberrant proliferation of nucleus pulposus (NP) cells has been reported to be implicated in the pathogenesis of intervertebral disc degeneration (IDD). Previous studies have demonstrated that microRNAs (miRNAs), which are a group of small noncoding RNAs, are critical regulators of cell proliferation in various pathologies. However, the role of miRNA‑96 (miR‑96) in the proliferation of NP cells remains to be determined. In the present study, reverse transcription‑quantitative polymerase chain reaction was used to investigate the expression of miR‑96 in NP tissues from patients with IDD and healthy tissues from patients with traumatic lumbar fracture as the control. A dual‑luciferase reporter assay was used to investigate whether AT‑rich interaction domain 2 (ARID2) may be a direct target gene for miR‑96. Furthermore, isolated NP cells from patients with IDD were transfected with miR‑96 mimics and ARID2‑targeting small interfering RNAs; cell proliferation, and the protein expression of Akt, phosphorylated Akt and ARID2 were examined, whereas the effects of an Akt inhibitor on NP cell proliferation were also evaluated. The present results demonstrated that miR‑96 expression was significantly upregulated in IDD samples, and the level of miR‑96 expression was positively associated with disc degeneration grade, which was evaluated by a modified Pfirrmann grading system. In addition, the current study identified ARID2 as a direct gene target of miR‑96. Furthermore, it was demonstrated that ARID2 mRNA expression was inversely correlated with the expression of miR‑96 in NP tissues. In addition, miR‑96 overexpression promoted NP cell proliferation and induced Akt phosphorylation, which led to increased cyclin D1 translation. Notably, overexpression of ARID2 or treatment with an Akt inhibitor decreased the effect of miR‑96 on NP cell proliferation. In conclusion, the results of the present study indicate that miR‑96 may promote the proliferation of human degenerated NP cells by targeting ARID2 via activation of the Akt pathway, and potentially serves as a therapeutic target for IDD.