Dual PI3K-BRD4 Inhibitor SF1126 Inhibits Colorectal Cancer Cell Growth in Vitro and in Vivo.

BACKGROUND/AIMS: Bromodomain-containing protein 4 (BRD4) and phosphatidylinositol 3-kinase (PI3K) are key oncogenic cascades in colorectal cancer (CRC). SF1126 is a novel and potent PI3K-BRD4 dual inhibitor. METHODS: CRC cells and human colon epithelial cells were treated with SF1126. Cell survival was tested by MTT and soft agar colony formation assays. Cell proliferation was tested by BrdU ELISA method. Cell apoptosis was tested by a TUNEL staining method and Histone DNA ELISA. Western blotting was utilized to test the signaling proteins. A HT-29 xenograft mice model was established to study the anti-tumor activity of SF1126 in vivo. RESULTS: SF1126 potently inhibited the survival, proliferation, and progression of the cell cycle in an established CRC cell line (HT-29) and primary human colon cancer cells. Significant activation of apoptosis was detected in SF1126-treated CRC cells. In CRC cells, SF1126 blocked Akt-mammalian target of rapamycin (mTOR) complex1/2 signaling and downregulated BRD4 target proteins (Myc and cyclin D1). Further studies showed that SF1126 activated p38 signaling in CRC cells. In contrast, the p38 inhibitors or p38 short hairpin RNA inhibited SF1126-induced cytotoxicity and apoptosis in CRC cells. In vivo, subcutaneous administration of SF1126 significantly inhibited HT-29 xenograft tumor growth in nude mice. CONCLUSION: SF1126 inhibits CRC cell growth possibly by targeting PI3K-Akt-mTOR, BRD4, and p38 signaling.

Long Non-coding RNA RP11-395G23.3 Acts as a Competing Endogenous RNA of miR-124-3p to Regulate ROR1 in Anaplastic Thyroid Carcinoma.

Anaplastic thyroid carcinoma (ATC) is one of the most aggressive human malignancies with poor prognosis. However, the underlying mechanisms of ATC remain to be elucidated. Recently, increasing studies have focused on competitive endogenous RNA (ceRNA) to discover valuable biomarkers for the diagnosis of ATC. The present study identified 705 differentially expressed mRNAs and 47 differentially expressed lncRNAs. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were also conducted. Additionally, an lncRNA/miRNA/mRNA network was constructed which included 1103 regulatory relations. The upregulation of RP11-395G23.3 in ATC cells was confirmed by quantitative reverse transcription polymerase chain reaction (qRT-PCR). In the loss of function assays, results suggested silencing of RP11-395G23.3 inhibited cell proliferation and induced cell apoptosis. Mechanically, RP11-395G23.3 could increase ROR1 via sponging miR-124-3p as a ceRNA. Moreover, ROR1 expression was decreased with the downregulation of RP11-395G23.3, but was rescued by the co-transfection of the miR-124-3p inhibitor in ATC cells. Our research suggested that the RP11-395G23.3/miR-124-3p/ROR1 axis potentially acted as a potential target for the diagnosis of ATC.

Papillary Thyroid Microcarcinoma: A Nomogram Based on Clinical and Ultrasound Features to Improve the Prediction of Lymph Node Metastases in the Central Compartment.

Background: Accurate preoperative identification of central lymph node metastasis (CLNM) is essential for surgical protocol establishment for patients with papillary thyroid microcarcinoma (PTMC). We aimed to develop a clinical and ultrasound characteristics-based nomogram for predicting CLNM. Methods: Our study included 399 patients who were pathologically diagnosed with PTMC between January 2011 and June 2018. Clinical and ultrasound features were collected for univariate and multivariate analyses to determine risk factors of CLNM. A nomogram comprising the prognostic model to predict the CLNM was established, and internal validation in the cohort was performed. The Cox regression model was used to determine the risk factors for recurrence-free survival (RFS) and cumulative hazard was calculated to predict prognosis. Results: Three variables of clinical and US features as potential predictors including sex (odd ratio [OR] = 1.888, 95% confidence interval [CI], 1.160-3.075; P =0.011), tumor size (OR = 1.933, 95% CI, 1.250-2.990; P =0.003) and ETE (OR = 6.829, 95% CI, 3.250-14.350; P <0.001) were taken into account. The predictive nomogram was established by involving all the factors above used for preoperative prediction of CLNM in patients with PTMC. The nomogram showed excellent calibration in predicting CLNM, with area under curves (AUC) of 0.684 (95% CI, 0.635 to 0.774). Furthermore, tumor size, multifocality, presence of ETE, vascular invasion, and CLNM were the significant factors related to the RFS. Conclusion: Through this easy-to-use nomogram by combining clinical and US risk factor, the possibility of CLNM can be objectively quantified preoperatively. This prediction model may serve as a useful clinical tool to help clinicians determine an individual's risk of CLNM in PTMC, thus make individualized treatment plans accordingly.

Significance of multifocality in papillary thyroid carcinoma.

BACKGROUND: Although multifocality is often observed in papillary thyroid carcinoma (PTC), the associations with clinicopathologic factors and the prognostic value are still controversial. We aimed to identify the risk factors for multifocality and bilaterality, and investigate the significance of multifocality on prognosis in all PTC, papillary thyroid microcarcinoma (PTMC) and non-PTMC. METHODS: Data from 635 patients who underwent lobectomy/total thyroidectomy plus cervical lymph node dissection for PTC were retrospectively analyzed. Clinicopathological factors associated with multifocal PTC and bilateral PTC were investigated by univariate analysis. Multivariate Cox regression analyses was used to identify the clinicopathological prognostic factors for recurrence-free survival. RESULTS: Multifocal and bilateral PTC were observed in 157 (24.7%) and 99 (15.6%) patients, respectively. The frequency of large diameter (>1.0 cm), extrathyroidal extension (ETE), vascular invasion and central lymph node metastases (CLNM) was higher in multifocal PTC than that of solitary PTC. Moreover, ETE, vascular invasion, CLNM and lateral lymph node metastasis (LLNM) were more frequent in patients with 3 or more tumor foci compared to those with 2 tumor foci and 1 tumor lesion. Bilateral PTC had higher rates of ETE and CLNM. Multifocality was found to be the predictor of recurrence in all PTC, PTMC, and non-PTMC. In addition, the risk of recurrence increased with increasing number of tumor foci in all multifocal PTC patients and multifocal non-PTMC patients. CONCLUSION: Although multifocality and bilaterality had more aggressive features in PTC, only multifocality was associated with the increased risk of recurrence. An increase in the number of tumors was associated with an increased risk of ETE, vascular invasion, CLNM and LLNM. The prognostic value of multifocality is particularly significant in non-PTMC patients.

Predictive Factors for Lateral Lymph Node Metastasis and Skip Metastasis in Papillary Thyroid Carcinoma.

In papillary thyroid cancer (PTC) patients, cervical lymph node metastases are common, which disseminate sequentially from the central neck to the lateral neck. However, there is also a chaotic pattern of lymph node metastasis occasionally. In this study, we summarized 653 PTC patients who underwent thyroidectomy and central lymph node dissection with or without lateral lymph node dissection from two hospitals to investigate the pattern and risk factors of lateral lymph node metastasis (LLNM) and skip metastasis. LLNM was significantly associated tumor size > 1 cm, presence of extrathyroidal extension, tumors in the upper-lateral pole, and the number of metastatic lymph nodes in the central compartment. The frequency of skip metastasis was 22.5% (20 of 89 patients). Multivariate analyses showed tumor size ≤ 1 cm, and tumors in the upper-lateral pole were separately and independently associated with the risk of skip metastasis. Presence of LLNM affected the recurrence-free survival (RFS). RFS did not show the significantly difference between patients with LLNM and skip metastasis. Despite the low incidence of skip metastasis, attention should be paid to the possibility of LLNM even in the absence of central lymph node metastases. Besides, for patients with risk factors of LLNM or skip metastasis, detailed preoperative examination for the lateral compartment, especially the level III, is essential.

The therapeutic effect of the BRD4-degrading PROTAC A1874 in human colon cancer cells.

A1874 is a novel BRD4-degrading proteolysis targeting chimera (PROTAC). In primary colon cancer cells and established HCT116 cells, A1874 potently inhibited cell viability, proliferation, cell cycle progression, as well as cell migration and invasion. The BRD4-degrading PROTAC was able to induce caspase and apoptosis activation in colon cancer cells. Furthermore, A1874-induced degradation of BRD4 protein and downregulated BRD-dependent genes (c-Myc, Bcl-2, and cyclin D1) in colon cancer cells. Significantly, A1874-induced anti-colon cancer cell activity was more potent than the known BRD4 inhibitors (JQ1, CPI203, and I-BET151). In BRD4-knockout colon cancer cells A1874 remained cytotoxic, indicating the existence of BRD4-independent mechanisms. In addition to BRD4 degradation, A1874 cytotoxicity in colon cancer cells was also associated with p53 protein stabilization and reactive oxygen species production. Importantly, the antioxidant N-acetyl-cysteine and the p53 inhibitor pifithrin-α attenuated A1874-induced cell death and apoptosis in colon cancer cells. In vivo, A1874 oral administration potently inhibited colon cancer xenograft growth in severe combined immuno-deficient mice. BRD4 degradation and p53 protein elevation, as well as apoptosis induction and oxidative stress were detected in A1874-treated colon cancer tissues. Together, A1874 inhibits colon cancer cell growth through both BRD4-dependent and -independent mechanisms.

Influence of Tumor Number on Clinicopathologic Features and Outcomes of Patients With Papillary Thyroid Carcinoma.

OBJECTIVES: The purpose of this study was to investigate the significance of tumor number on clinicopathologic factors and outcomes of patients with papillary thyroid carcinoma (PTC). METHODS: We retrospectively analyzed 667 patients with PTC. We compared clinicopathologic features of patients with a different tumor number. Cox proportional hazards model was used to analyze risk factors of recurrence. RESULTS: In papillary thyroid microcarcinoma (PTMC), the increase in the number of tumor foci was related to a higher risk of minimal extrathyroidal extension (ETE) and lymphovascular invasion (P < .05). Patients with PTMC with four or more foci had a significantly higher risk of central lymph node metastasis (CLNM) and lateral lymph node metastasis (LLNM) than patients with solitary tumors (P < .05). Patients with macro-PTC with four or more foci and with three foci had a higher risk of gross ETE and lymphovascular invasion than patients with solitary tumors (P < .05). The increase in the tumor number was related to a higher risk of CLNM in macro-PTC (P < .05). The number of foci was the independent predictor of recurrence in patients with macro-PTC (P < .05). CONCLUSIONS: An increase in the number of tumors was associated with an increased risk of aggressive clinicopathologic features in PTMC and macro-PTC. The number of tumor foci could influence risk of recurrence in macro-PTC.

Management of Clinically Solitary Papillary Thyroid Carcinoma Patients According to Risk-Scoring Model for Contralateral Occult Carcinoma.

Objective: The aim of this study was to investigate risk factors of occult carcinoma in clinically solitary papillary thyroid carcinoma (PTC) patients, and to put emphasis on the predictive value of risk-scoring model to determine the optimal scope of surgery. Methods: A total of 573 clinically solitary PTC patients who underwent total thyroidectomy (TT) from two hospitals were retrospectively analyzed. Clinicopathological features were collected, univariate and multivariate analyses were performed to determine risk factors of occult carcinoma. The Cox proportional hazards model was used to analyze the risk factors of recurrence. A scoring model was constructed according to independent risk factors of contralateral occult carcinoma. Results: 19.2% of clinically solitary PTC patients had occult carcinoma, among which 3.7% patients had ipsilateral occult carcinoma and 15.5% patients had contralateral occult carcinoma. Factors such as male, the presence of benign nodule, and vascular invasion increase the risk of ipsilateral occult carcinoma. Tumor size >1 cm, the presence of benign nodule, extrathyroidal extension, central lymph node metastasis, lateral lymph node metastasis are independent predictors of contralateral occult carcinoma. Contralateral occult carcinoma is the independent predictor of recurrence. A 10-point risk-scoring model was established to predict the contralateral occult carcinoma in clinically solitary PTC patients. Conclusion: Lobectomy is sufficient for clinically solitary PTC patients with risk factors of ipsilateral occult carcinoma. For clinically solitary PTC patients with score ≥4, careful preoperative evaluations are required to rule out the contralateral occult carcinoma. Even if contralateral occult carcinoma is not detected preoperatively, TT is recommended for high-risk patients.

[Variations of the amount of sialic acids on hepatocellular carcinoma cell membrane].

OBJECTIVE: To observe the change in the amount of sialic acids on hepatocellular carcinoma (HCC) cell membrane. METHODS: Surgical specimens of HCC and liver cirrhosis tissues were obtained from 28 patients to prepare carcinoma cell and hepatocyte suspensions by collagenase digestion. For assay of α2, 3 and α2, 6-sialic acids, the cells were suspended in the staining buffer containing either fluorescein isothiocyanate-Maackia amurensis lectin (FITC-MAL) or fluorescein isothiocyanate-Sambucus nigra bark lectin (FITC-SNA) and incubated for 1 h, respectively. Flow cytometric analysis was carried out to measure the mean fluorescence intensity (MFI) on the cell surface. RESULTS: In both FITC-MAL- and FITC-SNA-incubated HCC cells, the MFI on the cell surface was greater than that of the hepatocytes. CONCLUSION: Both of α2, 3 and α2, 6- sialic acids increases significantly on the hepatocyte membrane after the carcinomatous change.

[Protocols for cloning human bone marrow-derived hepatic stem cells in vitro].

OBJECTIVE: To explore practical protocols for cloning bone marrow-derived hepatic stem cells in vitro. METHODS: The cell fraction rich in CD117(+) cells and CD184(+) cells was separated from fresh bone marrow by density gradient centrifugation and cultured for 0, 7 and 14 days in high-glucose DMEM supplemented with or without 10% autologous serum or in serum-free high-glucose DMEM. All the media were supplemented with different concentrations of hepatocyte growth promoting factors (HGPF), thrombopoietin (TPO) and interleukin-3 (IL-3). The quantitative changes of CD117(+) cells and CD184(+) cells were measured by flow cytometry. RESULTS: The optimal effect for cell cloning was achieved with high-glucose DMEM with 10% autologous serum group supplemented with 40 microg/ml HGPF, 50 ng/ml TPO, and 10 ng/ml IL-3. At day 7 of cell culture in this media, the quantity of CD117(+) cells and CD184(+) cells increased by 6.55 and 6.20 folds, and by 11.62 and 20.57 folds at day 14, respectively. CONCLUSION: It is practical for cloning bone marrow-derived hepatic stem cells in high-glucose DMEM with 10% autologous serum supplemented with 40 microg/ml HGPF, 50 ng/ml TPO, and 10 ng/ml IL-3.

[Intrahepatic transplantation of in vitro induced autologous bone marrow-derived liver stem cells in patients with posthepatitic cirrhosis].

OBJECTIVE: To evaluate the therapeutic effect of in vitro induced autologous bone marrow-derived liver stem cell transplantation for posthepatitic cirrhosis. METHODS: Between Jun 2008 and Mar 2009, 12 patients with posthepatitic cirrhosis and portal hypertensive underwent azygousportal disconnection and splenectomy in our department. The patients were then divided into two groups to receive autologous bone marrow-deprived liver stem cell infusion via the hepatic artery after in vitro induction for 7 days (n=6) or saline (n=6). The therapeutic effects of the operations on the liver functions and liver fibrosis index were evaluated. RESULTS: All the patients recovered uneventfully and no side effect of the operation was found. After the operation, the patients receiving bone marrow-deprived liver stem cell infusion showed better hepatic function improvement than those receiving saline infusion (P<0.05). CONCLUSION: Transplantation of in vitro induced autologous bone marrow-derived liver stem cell via the hepatic artery is safe and effective for treatment of posthepatitic cirrhosis.