Clinical relevance and outcome of familial papillary thyroid cancer: a single institution study of 626 familial cases.

Background: Whether familial thyroid cancer is more aggressive than sporadic thyroid cancer remains controversial. Additionally, whether the number of affected family members affects the prognosis is unknown. This study focused mainly on the comparison of the clinicopathological characteristics and prognoses between papillary thyroid cancer (PTC) patients with and without family history. Methods: A total of 626 familial papillary thyroid cancer (FPTC) and 1252 sporadic papillary thyroid cancer (SPTC) patients were included in our study. The clinical information associated with FPTC and SPTC was recorded and analyzed by univariate analysis. Results: Patients in the FPTC group had a higher rate of multifocality (p=0.001), bilaterality (p=0.000), extrathyroidal invasion (p=0.000), distant metastasis (p=0.012), lymph node metastasis (p=0.000), recurrence (p=0.000), a larger tumor size (p=0.000) and more malignant lymph nodes involved (central: p=0.000; lateral: p=0.000). In addition, our subgroup analysis showed no significant difference (p>0.05) between patients with only one affected family member and those with two of more group in all clinicopathological characteristics. In papillary thyroid microcarcinoma (PTMC) subgroup analysis, we found that FPTMC patients harbored significantly larger tumors (p=0.000), higher rates of multifocality (p=0.014), bilaterality (p=0.000), distant metastasis (p=0.038), lymph node metastasis (p=0.003), greater numbers of malignant lymph nodes (central: p=0.002; lateral: p=0.044), higher rates of I-131 treatment (p=0.000) and recurrence (p=0.000) than SPTMC patients. Conclusion: Our results indicated that PTC and PTMC patients with a positive family history had more aggressive clinicopathological behaviors, suggesting that more vigilant screening and management for FPTC may be helpful.

Screening tumor stage-specific candidate neoantigens in thyroid adenocarcinoma using integrated exome and transcriptome sequencing.

Background: The incidence of thyroid carcinoma (THCA), the most common endocrine tumor, is continuously increasing worldwide. Although the overall prognosis of THCA is good, patients with distant metastases exhibit a mortality rate of 5-20%. Methods: To improve the diagnosis and overall prognosis of patients with thyroid cancer, we screened specific candidate neoantigen genes in early- and late-stage THCA by analyzing the transcriptome and somatic cell mutations in this study. Results: The top five early-stage neoantigen-related genes (NRGs) were G protein-coupled receptor 4 [GPR4], chondroitin sulfate proteoglycan 4 [CSPG4], teneurin transmembrane protein 1 [TENM1], protein S 1 [PROS1], and thymidine kinase 1 [TK1], whereas the top five late-stage NRGs were cadherin 6 [CDH6], semaphorin 6B [SEMA6B], dysferlin [DYSF], xenotropic and polytropic retrovirus receptor 1 [XPR1], and ABR activator of RhoGEF and GTPase [ABR]. Subsequently, we used machine learning models to verify their ability to screen NRGs and analyze the correlations among NRGs, immune cell types, and immune checkpoint regulators. The use of candidate antigen genes resulted in a better diagnostic model (the area under the curve [AUC] value of the early-stage group [0.979] was higher than that of the late-stage group [0.959]). Then, a prognostic model was constructed to predict NRG survival, and the 1-, 3- and 5-year AUC values were 0.83, 0.87, and 0.86, respectively, which were closely related to different immune cell types. Comparison of the expression trends and mutation frequencies of NRGs in multiple tumors revealed their potential for the development of broad spectrum therapeutic drugs. Conclusion: In conclusion, the candidate NRGs identified in this study could potentially be used as therapeutic targets and diagnostic biomarkers for the development of novel broad spectrum therapeutic agents.

Pleckstrin homology and RhoGEF domain containing G4 (PLEKHG4) leads to the activation of RhoGTPases promoting the malignant phenotypes of thyroid cancer.

Thyroid cancer (TC) is one of the most common endocrine system cancers, and its incidence is elevating. There is an urgent need to develop a deeper understanding of TC pathogenesis and explore new therapeutic target for its treatment. This study aimed to investigate the effects of pleckstrin homology and RhoGEF domain containing G4 (PLEKHG4) on the progression of TC. Herein, 29 pairs of TC and adjacent tissues were used to assess the expression of PLEKHG4. A xenograft model of mouse was established by subcutaneously injected with TC cells. Lung metastasis model was established through left ventricular injection. The results revealed that PLEKHG4 was up-regulated in human TC tissues. PLEKHG4 level was correlated with clinicopathological parameters of TC patients. In vitro assays revealed that PLEKHG4 promoted TC cell proliferation, migration, invasion, and epithelial-mesenchymal transformation. Knockdown of PLEKHG4 led to the opposite effects, and the loss of PLEKHG4 enhanced the apoptosis ability and inhibited the stemness properties of TC cells. These findings were further confirmed by the in vivo growth and lung metastasis of TC tumor. Mechanistically, PLEKHG4 promoted the activation of RhoGTPases RhoA, Cdc42, and Rac1. The inhibitors of these RhoGTPases reversed the PLEKHG4-induced malignant phenotypes. Additionally, ubiquitin-conjugating enzyme E2O (UBE2O), a large E2 ubiquitin-conjugating enzyme acted as an ubiquitin enzyme of PLEKHG4, facilitated its ubiquitination and degradation. In conclusion, PLEKHG4, regulated by UBE2O, promoted the thyroid cancer progression via activating the RhoGTPases pathway. UBE2O/PLEKHG4/RhoGTPases axis is expected to be a novel a therapeutic target for TC treatment.

Hypermethylation of microRNA-497-3p contributes to progression of thyroid cancer through activation of PAK1/ß-catenin.

MicroRNA-497 (miR-497) has been reported to be a tumor-suppressive miRNA in thyroid cancer (TC), yet the mechanism is not clearly defined. In this study, we aim to determine the mechanism by which miR-497-3p affects the progression of TC. After characterization of low miR-497-3p expression pattern in TC and normal tissues, we assessed the correlation between miR-497-3p expression and clinicopathological features of TC patients. Its low expression shared associations with advanced tumor stage and lymph node metastasis. ChIP and methylation-specific PCR provided data showing that downregulation of miR-497-3p in TC tissues was induced by DNA methyltransferase-mediated hypermethylation. By performing dual-luciferase reporter assay, we identified that miR-497-3p targeted PAK1 while PAK1 could inhibit ß-catenin expression. Through this mechanism, miR-497-3p exerted the anti-proliferative, anti-invasive, pro-apoptotic, and anti-tumorigenic effects on TC cells on the strength of the results from gain-of-function and rescue experiments. This study suggested that hypermethylation of miR-497-3p resulted in upregulation of ß-catenin dependent on PAK1 and contributed to cancer progression in TC, which highlighted one of miR-mediated tumorigenic mechanism.

A Nomogram Predicting the Overall Survival and Cancer-Specific Survival in Patients with Parathyroid Cancer: A Retrospective Study.

Purpose: This study aimed to explore a visual model for predicting the prognosis of patients with parathyroid carcinoma (PC) and analyze related biochemistries in different groups of stage. Methods: The training dataset of 342 patients with PC was obtained from the Surveillance, Epidemiology, and End Results (SEER) database, and the validation dataset included 59 patients from The First Affiliated Hospital of Zhengzhou University. Univariate and multivariate Cox regression analyses were performed to evaluate significant independent prognostic factors. Based on those factors, nomograms and Web-based probability calculators were constructed to evaluate the overall survival (OS) and the cancer-specific survival (CSS) at 3, 5, and 8 years. The concordance index (C-index), receiver operating characteristic (ROC) curve, calibration curve, and decision curve analysis (DCA) were used to evaluate the nomogram in the training set and validation set. Moreover, biochemistries from the validation set were retrospectively analyzed in different groups of stage by Kruskal-Wallis test. Results: Age, marital status, tumor size, stage, lymph node status, and radiation were identified as prognostic factors of OS. In contrast, only tumor size and stage were predictive for CSS. The nomogram was developed based on these independent factors. The C-index, ROC curve, calibration curve, and DCA of the nomogram in both training and validation sets showed that the nomogram had good predictive value, stability, and clinical benefit in predicting 3-, 5-, and 8-year OS and CSS in PC patients. Among the 59 PC patients from our hospital, lower albumin (ALB) levels and higher postoperative parathyroid hormone (PTH) levels were found in patients with distant metastasis (Distant vs. Regional ALB levels: p = 0.037; Distant vs. Local ALB levels: p = 0.046; Distant vs. Regional postoperative PTH levels: p = 0.002; Distant vs. Local postoperative PTH: p = 0.002). Conclusion: The established nomogram application can provide accurate prognostics for patients with PC in the Chinese population, but it must be validated on prospectively collected real-world data.

LncRNA HCG11 Facilitates Nasopharyngeal Carcinoma Progression Through Regulating miRNA-490-3p/MAP3K9 Axis.

Purpose: Long noncoding RNAs (LncRNAs) play complex but important roles in the progression of various tumors. This study aimed to elucidate the functional mechanisms of the HLA complex group 11 (HCG11) in nasopharyngeal carcinoma (NPC). Patients and Methods: HCG11 levels in NPC specimens were determined by fluorescence in situ hybridization (FISH) and qPCR. Proliferation, apoptosis, and metastasis of NPC cells were determined using CCK8, colony formation, annexin V-PI, and transwell assays. A murine tumor xenograft model was used to investigate the regulatory function of HCG11 in NPC in vivo, and immunohistochemical staining was used to determine the Ki-67 level in tumors. The target relationships between HCG11, microRNA miR-490-3p, and MAPK kinase kinase 9 (MAP3K9) were detected using bioinformatics, qPCR, western blotting, and luciferase reporter assays. Results: HCG11 was highly expressed in NPC tissues and was positively associated with tumor stage, lymphatic metastasis, and poor prognosis. Functionally, HCG11 knockdown inhibited proliferation and migration and induced apoptosis of NPC cells. Mechanistically, miR-490-3p is a direct target of HCG11, oncogenic functions of HCG11 in NPC cell proliferation and migration can be partially reversed by the miR-490-3p inhibitor. HCG11 significantly increased mitogen-activated protein kinase MAPK kinase 9 (MAP3K9) levels by inhibiting miR-490-3p. Conclusion: HCG11 facilitates NPC progression via MAP3K9 signaling by sponging miRNA-490-3p, which may contribute to new prognostic markers and promising therapeutic targets.

Combination of Stimulated Thyroglobulin and Antithyroglobulin Antibody Predicts the Efficacy and Prognosis of 131I Therapy in Patients With Differentiated Thyroid Cancer Following Total Thyroidectomy: A Retrospective Study.

Background and Purpose: This study aimed to analyze the diagnostic ability of the combination of stimulated thyroglobulin (sTg) and antithyroglobulin antibody (TgAb) in predicting the efficacy and prognosis of radioactive iodine (131I) therapy (RAIT) in patients with differentiated thyroid carcinomas (DTCs) after total thyroidectomy (TT). Methods: This retrospective study comprised 409 DTC patients who underwent 131I treatment following TT in the First Affiliated Hospital of Zhengzhou University from January 2019 to August 2020, and they were followed up to November 2021. Patients were divided into the successful ablation and the unsuccessful ablation group based on the classification of the efficacy of RAIT in the 2015 American Thyroid Association guidelines. The clinical characteristics and the efficacy of the initial RAIT were evaluated. The cutoffs of preablation sTg, sTg/thyroid-stimulating hormone (TSH) ratio, and sTg×TgAb product were calculated to predict the efficacy of RAIT. Univariate and multivariate logistic regression analyses were used to identify the independent risk factors for unsuccessful ablation. Kaplan-Meier curves were used to estimate the prognostic value of sTg×TgAb product affecting progression-free survival (PFS). Results: The cohort consisted of 222 cases in the successful ablation group and 187 cases in the unsuccessful ablation group. Between the two groups, preablation sTg, sTg/TSH ratio, and sTg×TgAb product were significantly higher in the unsuccessful ablation group. The area under the curve (AUC) of the sTg×TgAb product was the highest among the above three factors. The cutoffs for the worse therapeutic effect of the initial RAIT in sTg, sTg/TSH ratio, and sTg×TgAb were >2.99 ng/ml, >0.029 mg/IU, and >34.18, respectively. STg >2.99 ng/ml and sTg×TgAb product >34.18 were independent risk factors for unsuccessful ablation. Patients with sTg×TgAb product >34.18 had shorter PFS than that of patients with sTg×TgAb product ≤34.18. In separate analyses of TgAb-negative and TgAb-positive subgroups, higher sTg×TgAb was both associated with a lower success rate of RAIT and a shorter PFS. Conclusion: STg×TgAb product predicted the efficacy and prognosis of 131I therapy for both TgAb-negative and TgAb-positive DTC patients before the initial 131I treatment following TT. Thus, it can be used as a clinical reference indicator for the surveillance of DTC patients.

Long non-coding ROR promotes the progression of papillary thyroid carcinoma through regulation of the TESC/ALDH1A1/TUBB3/PTEN axis.

Papillary thyroidal carcinoma (PTC) is a common endocrine cancer that plagues people across the world. The potential roles of long non-coding RNAs (lncRNAs) in PTC have gained increasing attention. In this study, we aimed to explore whether lncRNA ROR affects the progression of PTC, with the involvement of tescalcin (TESC)/aldehyde dehydrogenase isoform 1A1 (ALDH1A1)/ßIII-tubulin (TUBB3)/tensin homolog (PTEN) axis. PTC tumor and adjacent tissues were obtained, followed by measurement of lncRNA ROR and TESC, ALDH1A1, and TUBB3 expression. Interactions among lncRNA ROR, TESC, ALDH1A1, TUBB3, and PTEN were evaluated by ChIP assay, RT-qPCR, or western blot analysis. After ectopic expression and depletion experiments in PTC cells, MTT and colony formation assay, Transwell assay, and flow cytometry were performed to detect cell viability and colony formation, cell migration and invasion, and apoptosis, respectively. In addition, xenograft in nude mice was performed to test the effects of lncRNA ROR and PTEN on tumor growth in PTC in vivo. LncRNA ROR, TESC, ALDH1A1, and TUBB3 were highly expressed in PTC tissues and cells. Overexpression of lncRNA ROR activated TESC by inhibiting the G9a recruitment on the promoter of TESC and histone H3-lysine 9me methylation. Moreover, TESC upregulated ALDH1A1 expression to increase TUBB3 expression, which then reduced PTEN expression. Overexpression of lncRNA ROR, TESC, ALDH1A1 or TUBB3 and silencing of PTEN promoted PTC cell viability, colony formation, migration, and invasion while suppressing apoptosis. Moreover, overexpression of lncRNA ROR increased tumor growth by inhibiting PTEN in vivo. Taken together, the current study demonstrated that lncRNA ROR mediated TESC/ALDH1A1/TUBB3/PTEN axis, thereby facilitating the development of PTC.

Silencing of circARHGAP12 inhibits the progression of atherosclerosis via miR-630/EZH2/TIMP2 signal axis.

Atherosclerosis (AS) is a common disease that seriously threatens human health. So far, the pathogenesis of AS has not been fully understood. This project investigates the effects of circARHGAP12 on AS and its regulatory mechanism. ApoE-/- knockout mice (ApoE) were adopted and reared with a high-fat diet to construct an AS model. Lentivirus was established to knock down the expression of circARHGAP12 in mice. After 12 weeks, the aorta was removed and the expression of circARHGAP12 was detected. Vascular oil red O staining was used to detect the degree of AS. The expression of inflammatory factors was detected by ELISA. Aortic smooth muscle cells (MASMCs) were cultured to evaluate the effects of circARHGAP12 on the phenotype of MASMCs. RNA pull-down and luciferase assay were used to verify the downstream target genes of circARHGAP12. In addition, the effects of circARHGAP12 on MASMCs proliferation and migration were detected by MTT and transwell assay. Compared with the normal group, the expression of circARHGAP12 in the MASMCs under ox-LDL treatment was elevated, and circARHGAP12 silencing could inhibit AS in vitro and in vivo. The results of the mechanism study showed that circARHGAP12 can directly bind with miR-630. In addition, miR-630 can also target EZH2 to modulate the transcription of TIMP2 and to influence the migration of MASMCs. circARHGAP12 is upregulated in AS. CircARHGAP12 knockdown can inhibit the progression of AS. This study expands on the role of circRNA in AS and provides potential targets for the treatment of AS.

Ultrasound images-based deep learning radiomics nomogram for preoperative prediction of RET rearrangement in papillary thyroid carcinoma.

Purpose: To create an ultrasound -based deep learning radiomics nomogram (DLRN) for preoperatively predicting the presence of RET rearrangement among patients with papillary thyroid carcinoma (PTC). Methods: We retrospectively enrolled 650 patients with PTC. Patients were divided into the RET/PTC rearrangement group (n = 103) and the non-RET/PTC rearrangement group (n = 547). Radiomics features were extracted based on hand-crafted features from the ultrasound images, and deep learning networks were used to extract deep transfer learning features. The least absolute shrinkage and selection operator regression was applied to select the features of nonzero coefficients from radiomics and deep transfer learning features; then, we established the deep learning radiomics signature. DLRN was constructed using a logistic regression algorithm by combining clinical and deep learning radiomics signatures. The prediction performance was evaluated using the receiver operating characteristic curve, calibration curve, and decision curve analysis. Results: Comparing the effectiveness of the models by linking the area under the receiver operating characteristic curve of each model, we found that the area under the curve of DLRN could reach 0.9545 (95% confidence interval: 0.9133-0.9558) in the test cohort and 0.9396 (95% confidence interval: 0.9185-0.9607) in the training cohort, indicating that the model has an excellent performance in predicting RET rearrangement in PTC. The decision curve analysis demonstrated that the combined model was clinically useful. Conclusion: The novel ultrasonic-based DLRN has an important clinical value for predicting RET rearrangement in PTC. It can provide physicians with a preoperative non-invasive primary screening method for RET rearrangement diagnosis, thus facilitating targeted patients with purposeful molecular sequencing to avoid unnecessary medical investment and improve treatment outcomes.

IGF2BP2 knockdown suppresses thyroid cancer progression by reducing the expression of long non-coding RNA HAGLR.

BACKGROUND: N6-methyladenosine (m6A), a common internal modification on RNAs, has been found to be closely linked with RNA biosynthesis/metabolism and cancer development. In this text, the roles and molecular mechanisms of m6A-bind protein IGF2BP2 in the development of thyroid cancer (TC) were investigated in vitro. METHODS: IGF2BP2 and lncRNA HAGLR were screened out through multiple public databases such as TCGA, Ualcan, POSTAR2, Starbase, and GEPIA. Cell proliferative, migratory and invasive abilities were assessed by CCK-8, Transwell migration and invasion assays, respectively. Cell cycle distribution and cell apoptotic patterns were measured by flow cytometry. The interaction between HAGLR and IGF2BP2 was examined by RIP, RNA pull-down and luciferase assays and bioinformatics analysis. The effect of IGF2BP2 knockdown on the m6A level of HAGLR was explored by meRIP assay. RESULTS: IGF2BP2 was highly expressed in TC tumor tissues. IGF2BP2 knockdown weakened cell proliferative, migratory, and invasive abilities, and induced cell cycle arrest and cell apoptosis in TC cells. LncRNA HAGLR expression was markedly upregulated and positively associated with IGF2BP2 expression in TC tissues. IGF2BP2 knockdown reduced HAGLR expression and transcript stability in TC cells. IGF2BP2 regulated HAGLR expression in an m6A-dependent manner. HAGLR overexpression weakened the effects of IGF2BP2 loss on cell proliferation, migration, invasion, apoptosis, and cell cycle progression in TC cells. CONCLUSION: IGF2BP2 loss inhibited cell proliferation, migration and invasion, and induced cell apoptosis and cell cycle arrest by down-regulating HAGLR expression in an m6A-dependent manner in TC cells, providing some potential diagnostic and therapeutic targets for TC.

LncRNA UCA1 attenuated the killing effect of cytotoxic CD8 + T cells on anaplastic thyroid carcinoma via miR-148a/PD-L1 pathway.

BACKGROUND: LncRNAs play an important role in the regulation of the killing effect of cytotoxic CD8 + T cells in various cancers. However, the role and underlying mechanisms of UCA1 in the killing effect of cytotoxic CD8 + T cells in anaplastic thyroid carcinoma (ATC) are not clear. METHODS: UCA1, miR-148a, and PD-L1 expression were detected by quantitative real-time PCR and/or Western blot. The ratio of PD-L1+ATC cells/ATC cells was determined using flow cytometry. The ability of CD8 + T cells to kill target ATC cells was detected by Chromium-51 (51Cr) release assay. The targeted relationship between UCA1 and miR-148a was confirmed by dual-luciferase reporter gene assay. RESULTS: UCA1 and PD-L1 expression levels were elevated in ATC tissues and cells. Silencing UCA1 and PD-L1 enhanced the killing effect of cytotoxic CD8 + T cells on ATC cells. UCA1 negatively regulated the expression of miR-148a, and miR-148a targeted PD-L1 to down-regulate its expression. Besides, we found that UCA1 attenuated the killing effect of cytotoxic CD8 + T cells and reduced cytokine secretion through PD-L1 and miR-148a. Finally, silencing UCA1 or PD-L1 in ATC cells restored the suppression of the killing effect of CD8 + T cells in vivo. CONCLUSION: UCA1 attenuated the killing effect of cytotoxic CD8 + T cells on ATC cells through the miR-148a/PD-L1 pathway.

Forkhead box K1 facilitates growth of papillary thyroid carcinoma cells by regulating connective tissue growth factor expression.

Forkhead box (FOX) proteins have been identified as key transcription factors in diverse biological processes involved in tumor progression. A large number of FOX proteins are implicated in tumorigenesis of papillary thyroid carcinoma (PTC). Here we investigated the role of Forkhead box K1 (FOXK1) in PTC progression. First, we found that FOXK1 was elevated in both PTC tissues (N = 68) and cell lines. Moreover, up-regulated FOXK1 was associated with shorter overall survival of PTC patients. Second, in vitro functional assays showed that FOXK1 promoted progression of PTC. Mechanistically, FOXK1 could bind to the promoter of cysteine-rich angiogenic inducer 61 (CYR61) and regulate connective tissue growth factor (CTGF) expression through CYR61. Notably, over-expression of CTGF weakened suppression of PTC progression induced by FOXK1 knockdown. Finally, in vivo xenotransplant tumor model indicated that knockdown of FOXK1 suppressed PTC growth. In conclusion, our results indicate that FOXK1 exerts oncogenic roles in PTC via CYR61/CTGF axis, which suggests FOXK1 might act as a potential therapeutic target.

LINC-PINT Suppresses the Aggressiveness of Thyroid Cancer by Downregulating miR-767-5p to Induce TET2 Expression.

Long noncoding RNA (lncRNA) long intergenic nonprotein-coding RNA, p53-induced transcript (LINC-PINT) has shown anti-invasive activity in lung and colon cancer cells. However, the role of LINC-PINT in thyroid cancer is unclear. In the present work, we explored the expression of LINC-PINT in 60 paired thyroid cancer and adjacent normal tissues. The clinical significance and biological function of LINC-PINT in thyroid cancer were determined. LINC-PINT expression was downregulated in thyroid cancer relative to adjacent normal tissues (p = 0.0002). Low expression of LINC-PINT was significantly associated with advanced tumor node metastasis (TNM) stage (p = 0.0306) and lymph node metastasis (p = 0.0359). Ectopic expression of LINC-PINT suppressed the proliferation, invasion, and tumorigenesis of thyroid cancer cells. Mechanistically, LINC-PINT associated with and downregulated microRNA (miR)-767-5p. Moreover, LINC-PINT overexpression relieved miR-767-5p-mediated repression of ten-eleven translocation 2 (TET2). miR-767-5p promoted aggressiveness of thyroid cancer, which was reversed by overexpression of TET2. Coexpression of miR-767-5p or depletion of TET2 rescued the inhibitory effect of LINC-PINT on thyroid cancer cell proliferation and invasion. In addition, there was a negative correlation between miR-767-5p and LINC-PINT in thyroid cancer (r = -0.34772, p = 0.01789). Taken together, LINC-PINT functions as a tumor suppressor in thyroid cancer via the miR-767-5p/TET2 axis, representing a potential therapeutic target for thyroid cancer.

Evaluation of immune infiltrating of thyroid cancer based on the intrinsic correlation between pair-wise immune genes.

INTRODUCTION: Unlike most mutation-driven cancers, thyroid cancer is thought to be highly dependent on changes in human hormone levels. It has become research hotspot using the change of gene expression level as a detection and diagnostic marker. The internal relationship between two genes and disease development is used to avoid the instability caused by single gene fluctuation. Aim It is possible to achieve early diagnosis in thyroid cancer during tumorigenesis and recurrence using IGPS (immune gene pairs). METHODS: We extracted thyroid cancer data from The Cancer Genome Atlas (TCGA), using CIBERSORT algorithm to infiltrate out 22 immune cells types. We screened out IGPS that differ significantly between different groups, then used LinearSVC model to learn and screen features, combined with deep learning neural network model to predict benign and malignant cancer as well as patients at different groups. KEY FINDINGS: There are significant differences of immune cell ratio in tumor stages and relapse samples. We screen out 42 and 64 IGPS for in normal-tumor and non-relapsed groups respectively, for example ASCC3-MAP3K7 and ATF2-SOCS5, have significant correlation in IGPS expression. Then we use the IGPS to train the tumor diagnostic classifier, obtain average AUC are both 0.99 after ten times cross-validation. SIGNIFICANCE: The IGPS gives us new insight to explore immune cell infiltration of thyroid cancer, deep learning model can be further used in early diagnosis of thyroid cancer and estimation of the risk of recurrence.

TMPO-AS1 promotes cell proliferation of thyroid cancer via sponging miR-498 to modulate TMPO.

BACKGROUND: Thyroid cancer (TC) is the most frequent endocrine malignancy. Long noncoding RNAs (lncRNAs) have been confirmed to act as significant roles in tumor development. The role of lncRNA TMPO-AS1 in TC is still unclear, so it remains to be explored. The aim of the research is to investigate the role and regulatory mechanism of TMPO-AS1 in TC. METHODS: TMPO-AS1 and TMPO expression in TC tumors and cells was detected by TCGA database and QRT-PCR assay respectively. CCK-8, EDU, TUNEL and western blot assays were conducted to identify the biological functions of TMPO-AS1 in TC. Luciferase reporter and RNA pull down assays were conducted to measure the interaction among TMPO-AS1, TMPO and miR-498. RESULTS: TMPO-AS1 was overexpressed in TC tissues and cell lines. Knockdown of TMPO-AS1 suppressed cell growth and accelerated cell apoptosis in TC. Furthermore, downregulation of TMPO-AS1 suppressed TMPO expression in TC. The data suggested that TMPO expression was upregulated in TC tissues and cell lines and was positively correlated with TMPO-AS1 expression in TC. Furthermore, the expression of miR-498 presented low expression in TC cells. And miR-498 expression was negatively regulated by TMPO-AS1, meanwhile, TMPO expression was negatively regulated by miR-498 in TC cells. Besides, it was confirmed that TMPO-AS1 could bind with miR-498 and TMPO in TC cells. In addition, it was validated that TMPO-AS1 elevated the levels of TMPO via sponging miR-498 in TC cells. CONCLUSIONS: TMPO-AS1 promotes cell proliferation in TC via sponging miR-498 to modulate TMPO.

A Novel Immune-Related Prognostic Signature for Thyroid Carcinoma.

BACKGROUND: Recent studies have confirmed that immune-associated genes perform a crucial function in recurrence and metastasis of thyroid carcinoma. A reliable immune-related prognostic signature for patients with thyroid cancer is needed. This study constructed a novel immune-related prognostic signature for thyroid cancer and evaluated its prognostic value by bioinformatics analysis. METHODS: In this study, we anatomized differentially expressed immune-associated genes from The Cancer Genome Atlas database. The samples from The Cancer Genome Atlas database were randomly divided into training set and test set. A novel immune-related prognostic signature for thyroid cancer was developed by least absolute shrinkage and selection operator and Cox regression analysis: Risk score = (0.6846 × expression value of C-X-C motif chemokine ligand 5 [CXCL5]) + (1.1556 × expression value of Azurocidin 1 [AZU1]) + (-0.3156 × expression value of nucleotide binding oligomerization domain containing 1 [NOD1] + (0.0542 × expression value of TNF Receptor Superfamily Member 11b [TNFRSF11B]) + (0.0952 × expression value of VGF nerve growth factor inducible [VGF]). The established prognostic signature was evaluated based on training set and test set by survival curves, receiver-operator characteristic curves, risk score, survival status, heatmap, and independent prognostic analysis. Meanwhile, we appraised the correlation between target immune-associated genes and clinical stage, tumor-infiltrating immune cells respectively. RESULTS: Five immune-associated genes were used for constructing an immune-related prognostic signature by least absolute shrinkage and selection operator, univariate, and multivariate analysis. Survival curves, receiver-operator characteristic curves, and independent prognostic analysis showed the signature had significant prediction value. Clinical and immune cell correlation analyses indicated that target immune-associated genes may participate in tumor immune infiltration and tumor progression. CONCLUSIONS: We constructed a novel 5 immune-associated genes signature for predicting the prognosis of patients with thyroid cancer, which may help clinical workers evaluate individualized therapy and prognosis.

Plasma miR-323 as a Biomarker for Screening Papillary Thyroid Cancer From Healthy Controls.

The present study aims to evaluate whether plasma miR-323 serves as a potential biomarker to screen patients with papillary thyroid cancer (PTC) from healthy controls. Real-time PCR was performed to evaluate miR-323 expression in healthy controls and benign thyroid nodule (BTN) and PTC patients. Receiver operating characteristic (ROC) curve analysis was used to evaluate whether plasma miR-323 could be used to screen PTC patients from BTN patients and healthy controls. Plasma miR-323 was significantly increased in PTC patients compared with that in BNT patients and healthy controls. Moreover, miR-323 in the thyroid tissue was significantly increased in PTC patients when compared to BNT patients. We further showed that plasma and tissue miR-323 levels were significantly increased in PTC patients with metastasis compared to those without metastasis. Plasma miR-323 was significantly increased in PTC patients with BRAF V600E mutation when compared to those with wild-type BRAF. Furthermore, plasma miR-323 was significantly increased in PTC patients with higher Tg-FNAB. ROC analysis showed that plasma miR-323 could distinguish PTC patients from BNT patients and healthy controls. The present study demonstrated that plasma miR-323 might be an effective noninvasive indicator for PTC progression and serve as a biomarker for the diagnosis of PTC.

Knockout of ASAP1 induces autophagy in papillary thyroid carcinoma by inhibiting the mTOR signaling pathway.

Due to lymph node metastasis and infiltration, surgery for PTC (papillary thyroid carcinoma) is a high-risk treatment strategy. Our work reports for the first time that ASAP1 (ArfGAP with SH3 Domain, Ankyrin Repeat and PH Domain 1) is highly expressed in PTC and that its high expression is related to autophagy. Autophagy and ASAP1 expression in 40 PTC tissues and normal tissues were detected by immunofluorescence. We used the lentiviral CRISPR/Cas9 nickase to generate stable cell lines. The difference in autophagy levels between the ASAP1 KO group and the control group was determined by Western blot and immunofluorescence analyses. We added chloroquine (CQ) to verify that ASAP1 increased the formation of autophagosomes rather than reducing their degradation. The expression of mTOR activity-related proteins (P-P70S6K, P-MTOR) was studied by Western blotting. ASAP1 was upregulated while autophagy was downregulated in PTC tissues compared to normal tissues. Knockout of ASAP1 induced autophagy in both MDA-T32 and MDA-T85 cells. Knockout of ASAP1 attenuated the activation of the mTOR signaling pathway. Our studies demonstrated that ASAP1 is upregulated while autophagy is reduced in PTC tissues. In addition, knockout of ASAP1 induces autophagy in PTC by inhibiting the mTOR signaling pathway.

miR-744-5p mediates lncRNA HOTTIP to regulate the proliferation and apoptosis of papillary thyroid carcinoma cells.

Papillary thyroid carcinoma (PTC) is the most common malignancy in thyroid. miR-744-5p plays an efficient role in various cancers, but its role in PTC remains unknown. In this work, we aimed to explore the function of miR-744-5p and the mechanism by which miR-744-5p acted in PTC. We observed that miR-744-5p expression was significantly declined in PTC tissues and cell lines. The high level of miR-744-5p is significantly associated with a better clinical picture of PTC patients. Overexpression of miR-744-5p inhibited the proliferation, arrested the cell cycle, and promoted the apoptosis in PTC cells. Oppositely, down-regulation of miR-744-5p reversed the above tendencies. We also found that miR-744-5p down-regulated its downstream genes c-myc and attenuated cell proliferation induced by c-myc. Long non-coding RNA (lncRNA) HOTTIP was found to be up-regulated and to act as an oncogene in PTC. In this study, miR-744-5p bound to HOTTIP and was negatively regulated by HOTTIP. In conclusion, miR-744-5p acts as a tumor suppressor to inhibit proliferation and promotes the apoptosis of PTC cells via targeting c-myc. Moreover, miR-744-5p expression interferes with lncRNA HOTTIP ability to promote proliferation and downregulate apoptosis in papillary thyroid carcinoma.