TRAF4 Inhibits the Apoptosis and Promotes the Proliferation of Breast Cancer Cells by Inhibiting the Ubiquitination of Spindle Assembly-Associated Protein Eg5.

Tumor necrosis factor receptor associated factor 4 (TRAF4) is a RING domain E3 ubiquitin ligase that mediates the ubiquitination of various proteins and plays an important role in driving tumor progression. By studying the relationship between TRAF4 and Eg5, a member of the kinesin family that plays a critical role in spindle assembly, we demonstrated that TRAF4 regulated Eg5 ubiquitination and contributed to Eg5-mediated breast cancer proliferation and inhibited breast cancer apoptosis. TRAF4 and Eg5 were both highly expressed in breast cancer and their protein level was positively correlated. Relying on its Zinc fingers domain, TRAF4 interacted with Eg5 in the cytoplasm of breast cancer cells. TRAF4 was a mitosis-related protein, and by up-regulating the protein level of Eg5 TRAF4 participated in spindle assembly. Loss of TRAF4 resulted in monopolar spindles formation, but loss of function could be rescued by Eg5. Relying on its RING domain, TRAF4 up-regulated Eg5 protein levels by inhibition of Eg5 ubiquitination, thus stabilizing Eg5 protein level during mitosis. Furthermore, we found that Smurf2, a TRAF4-targeted ubiquitination substrate, mediated the regulation of Eg5 ubiquitination by TRAF4. TRAF4 inhibited the interaction between Smurf2 and Eg5, and down-regulated the protein level of Smurf2 by promoting its ubiquitination, thereby inhibited the Smurf2-catalyzed ubiquitination of Eg5 and up-regulated Eg5 protein levels. We also demonstrate that TRAF4 plays an important role in promoting cell proliferation and in inhibiting cell apoptosis induced by Eg5. In summary, our study suggests a new direction for investigating the role of TRAF4 in driving breast cancer progression.

Multiomics Landscape Uncovers the Molecular Mechanism of the Malignant Evolution of Lung Adenocarcinoma Cells to Chronic Low Dose Cadmium Exposure.

Cadmium (Cd) from cigarette smoke and polluted air can lead to lung adenocarcinoma after long-term inhalation. However, most studies are based on short-term exposure to this toxic metal at high concentrations. Here, we investigate the effects of long-term exposure of A549 cells (lung adenocarcinoma) to cadmium at low concentrations using morphological and multiomics analyses. First, we treated A549 cells continuously with CdCl2 at 1µM for 8 months and found that CdCl2 promoted cellular migration and invasion. After that, we applied transmission electron and fluorescence microscopies and did not observe significant morphological changes in Golgi apparatus, endoplasmic reticulum, lysosomes, or mitochondria on Cd treated cells; microfilaments, in contrast, accumulated in lamellipodium and adhesion plaques, which suggested that Cd enhanced cellular activity. Second, by using whole-exome sequencing (WES) we detected 4222 unique SNPs in Cd-treated cells, which included 382 unique non-synonymous mutation sites. The corresponding mutated genes, after GO and KEGG enrichments, were involved mainly in cell adhesion, movement, and metabolic pathways. Third, by RNA-seq analysis, we showed that 1250 genes (784 up and 466 down), 1623 mRNAs (1023 up and 591 down), and 679 lncRNAs (375 up and 304 down) were expressed differently. Furthermore, GO enrichment of these RNA-seq results suggested that most differentially expressed genes were related to cell adhesion and organization of the extracellular matrix in biological process terms; KEGG enrichment revealed that the differentially expressed genes took part in 26 pathways, among which the metabolic pathway was the most significant. These findings could be important for unveiling mechanisms of Cd-related cancers and for developing cancer therapies in the future.

Clinical comparison of tenosynovial giant cell tumors, synovial chondromatosis, and synovial sarcoma: analysis and report of 53 cases.

BACKGROUND: Tenosynovial giant cell tumors (TGCTs), synovial chondromatosis (SC), and synovial sarcoma (SS) exhibit similarities in clinical features and histochemical characteristics, and differential diagnosis remains challenging in clinical practice. METHODS: Data were collected from the pathology database of Shanghai Ninth People's Hospital regarding patients who underwent surgery from 2010 to 2019 with histologically confirmed TGCTs, SC, and SS. Demographic and clinicopathological data of these patients were reviewed. Immunohistochemistry staining of 14 different markers was performed. Correlation analyses of the prognoses were evaluated. RESULTS: A total of 26 patients with TGCTs (8 diffuse TGCTs and 18 localized TGCTs), 16 with SC, and 11 with SS were identified. Pain was the main symptom of patients with both TGCTs and SC, while a palpable mass was the most common symptom for patients with SS. In addition to clinical features, we identified vital risk factors for disease recurrence. The mean follow-up periods were 51, 39, and 14 months for TGCTs, SC, and SS, respectively. Younger patients with diffuse TGCTs or patients with a higher neutrophil/lymphocyte ratio (NLR) displayed a significantly higher frequency of recurrence. We also plotted receiver operating characteristic (ROC) curve analysis for age and NLR. The area under the ROC curve (AUC) was calculated and demonstrated the ability to distinguish recurrent from nonrecurrent cases. In addition, higher CD163 expression was linked to recurrent diffuse TGCT cases. CONCLUSIONS: These data indicated possible characteristics of different aspects of TGCTs, SC, and SS. Further clarification and understanding of these factors will help with differential clinical diagnosis and recurrent risk assessment.

The miR-5694/AF9/Snail Axis Provides Metastatic Advantages and a Therapeutic Target in Basal-like Breast Cancer.

Epigenetic deregulation, especially mutagenesis or the abnormal expression of epigenetic regulatory factors (ERFs), plays an important role in malignant tumorigenesis. To screen natural inhibitors of breast cancer metastasis, we adopted small interfering RNAs (siRNAs) to transiently knock down 591 ERF-coding genes in luminal breast cancer MCF-7 cells and found that depletion of AF9 significantly promoted MCF-7 cell invasion and migration. A mouse model of metastasis further confirmed the suppressive role of AF9 in breast cancer metastasis. RNA profiling revealed enrichment of AF9 targets genes in the epithelial-mesenchymal transition (EMT). Mechanistically, tandem mass spectrometry showed that AF9 interacts with Snail, which hampers Snail transcriptional activity in basal-like breast cancer (BLBC) cells. AF9 reconstitutes an activated state on the promoter of Snail, which is a master regulator of EMT, and derepresses genes by recruiting CBP or GCN5. Additionally, microRNA-5694 (miR-5694) targeted and degraded AF9 messenger RNA (mRNA) in BLBC cells, further enhancing cell invasion and migration. Notably, AF9 and miR-5694 expression in BLBC clinical samples correlated inversely. Hence, miR-5694 mediates downregulation of AF9 and provides metastatic advantages in BLBC. Restoring expression of the metastasis suppressor AF9 is a possible therapeutic strategy against metastatic breast cancer.

The Role of Tc-99m DTPA Renal Dynamic Scintigraphy in Retroperitoneal Liposarcoma.

PURPOSE: Technetium-99m diethylene triamine pentaacetic acid (Tc-99m DTPA) renal dynamic scintigraphy is a widely used imaging technique that evaluates renal function of patients with extrarenal abnormalities, but its clinical value in potentially offering us information on proliferation of liposarcoma has not yet been reported. METHODS: We retrospectively reviewed 7 patients with histopathologically confirmed retroperitoneal liposarcoma who underwent Tc-99m DTPA renal dynamic scintigraphy. The clinical data, histopathological findings, Glomerular Filtration Rate (GFR), and Tc-99m DTPA uptake were recorded. RESULTS: Dedifferentiated liposarcoma and well-differentiated liposarcoma showed dissimilar degrees of Tc-99m DTPA uptake, an observation that correlated with Ki-67 expression (p < 0.01). 4 of the 7 patients were diagnosed with dedifferentiated liposarcoma, showing a moderate uptake of Tc-99m DTPA and greater than 20% Ki-67 expression on histological slides. Meanwhile, the remaining 3 patients, diagnosed with well-differentiated liposarcoma, showed no uptake of Tc-99m DTPA and Ki-67 expression of less than 5%. CONCLUSIONS: This study suggests that Tc-99m DTPA renal dynamic scintigraphy provides diagnostic value in patients with retroperitoneal liposarcoma, not only in evaluating renal function but also in visualizing lesion-related radionuclide uptake, which may potentially offer further clinical insights into tumor proliferation and provide prognostic value for clinical outcomes in patients with retroperitoneal liposarcoma.

Short article: Sequence variations of PKHD1 underlie congenital hepatic fibrosis in a Chinese family.

OBJECTIVE: Congenital hepatic fibrosis (CHF) is a developmental disorder of the portobiliary system characterized by hepatic fibrosis, portal hypertension, and renal cystic disease. The aim of our study was to identify the disease-causing gene of a Chinese family with CHF. PATIENTS AND METHODS: Whole-exome sequencing was performed in the family with CHF and variants were confirmed by Sanger sequencing. Online bioinformatics tools were used to evaluate the pathogenicity of the missense variants. Liver specimens were reviewed to confirm the histopathological diagnosis. RESULTS: The compound heterozygous variants c.7994T>C, p.(Leu2665Pro) and c.8518C>T, p.(Arg2840Cys) in PKHD1 were identified in a Chinese family with CHF by whole-exome sequencing. Liver histomorphology was reviewed to confirm the diagnosis of CHF. CONCLUSION: We have identified variations in PKHD1 in a Chinese family with CHF. Our study extends the mutation spectrum of CHF and provides information for genetic counseling of patients' family members.

The association between the TP53 Arg72Pro polymorphism and colorectal cancer: An updated meta-analysis based on 32 studies.

Several previous studies evaluated the association between the Arg72Pro (rs1042522) polymorphism in the TP53 tumor suppressor gene and colorectal cancer (CRC). However, the results are conflicting. This meta-analysis aimed to shed new light on the precise association between TP53 variants and CRC. We analyzed 32 published case-control studies involving 8,586 cases and 10,275 controls using crude odd ratios (ORs) with 95% confidence intervals (CIs). The meta-analysis was performed using a fixed-effect or random-effects model, as appropriate. We found that the TP53 Arg72Pro polymorphism was not significantly associated with CRC risk in the overall population. However, subgroup analysis based on ethnicity revealed an increased risk of CRC among Asians (CC vs. GC+GG: OR=1.22, 95% CI: 1.02-1.45), and similar results were found for rectal cancer (CC vs. GC+GG: OR=1.34, 95% CI: 1.120-1.62). These results suggest that the TP53 Arg72Pro polymorphism CC genotype may contribute to an increased risk of CRC, especially for rectal cancer and among Asians.

Inhibition of MDM2 Re-Sensitizes Rapamycin Resistant Renal Cancer Cells via the Activation of p53.

BACKGROUND/AIMS: Rapamycin is a potential anti-cancer agent, which modulates the activity of mTOR, a key regulator of cell growth and proliferation. However, several types of cancer cells are resistant to the anti-proliferative effects of rapamycin. In this study, we report a MDM2/p53-mediated rapamycin resistance in human renal cancer cells. METHODS: Trypan blue exclusion tests were used to determine the cell viability. Changes in mRNA and protein expression were measured using real-time PCR and western blot, respectively. Xenograft models were established to evaluate the in vivo effects of rapamycin combined with a MDM2 inhibitor. RESULTS: Rapamycin treatment suppresses the expression of MDM2 and exogenous overexpression of MDM2 in A498 cells contributes to rapamycin resistance. By establishing a rapamycin resistant cell line, we observed that MDM2 was significantly upregulated in rapamycin resistant cells than that in rapamycin sensitive cells. Importantly, the rapamycin resistant cells demonstrated attenuated accumulation of p53 in the nucleus in response to rapamycin treatment. Moreover, the inhibition of MDM2 by siMDM2 sensitizes A498 cells to rapamycin through the activation of p53. In both in vitro and in vivo models, the combination of rapamycin with the MDM2 inhibitor, MI-319, demonstrated a synergistic inhibitory effect on rapamycin resistant cells. CONCLUSION: Our study reports a novel mechanism for rapamycin resistance in human renal cancer and provides a new perspective for the development of anti-cancer drugs.

Lentivirus-mediated knockdown of NLK inhibits small-cell lung cancer growth and metastasis.

Nemo-like kinase (NLK), an evolutionarily conserved serine/threonine kinase, has been recognized as a critical regulator of various cancers. In this study, we investigated the role of NLK in human small-cell lung cancer (SCLC), which is the most aggressive form of lung cancer. NLK expression was evaluated by quantitative real-time polymerase chain reaction in 20 paired fresh SCLC tissue samples and found to be noticeably elevated in tumor tissues. Lentivirus-mediated RNAi efficiently suppressed NLK expression in NCI-H446 cells, resulting in a significant reduction in cell viability and proliferation in vitro. Moreover, knockdown of NLK led to cell cycle arrest at the S-phase via suppression of Cyclin A, CDK2, and CDC25A, which could contribute to cell growth inhibition. Furthermore, knockdown of NLK decreased the migration of NCI-H446 cells and downregulated matrix metalloproteinase 9. Treatment with NLK short hairpin RNA significantly reduced SCLC tumor growth in vivo. In conclusion, this study suggests that NLK plays an important role in the growth and metastasis of SCLC and may serve as a potential therapeutic target for the treatment of SCLC.

Association between TP53 Arg72Pro polymorphism and leukemia risk: a meta-analysis of 14 case-control studies.

The relationship between the TP53 Arg72Pro polymorphism (rs1042522) and the risk of leukemia remains controversial. Consequently, we performed a meta-analysis to accurately evaluate the association between TP53 Arg72Pro polymorphism and leukemia risk. A comprehensive search was conducted to find all eligible studies of TP53 Arg72Pro polymorphism and leukemia risk. Fourteen case-control studies, with 2,506 cases and 4,386 controls, were selected for analysis. The overall data failed to indicate a significant association between TP53 Arg72Pro polymorphism and the risk of leukemia (C vs. G: OR = 1.09, 95% CI = 0.93-1.26; CC vs. GC + GG: OR = 1.23, 95% CI = 0.96-1.57). In a subgroup analysis of clinical types, an increased risk was observed in the acute lymphocytic leukemia (ALL) subgroup (CC vs. GC + GG: OR = 1.73; 95% CI = 1.07-2.81) but not in the acute myeloid leukemia (AML) subgroup. In the subgroup analysis, no significant associations with ethnicity and the source of the controls were observed. In conclusion, the results suggest that there is no association between TP53 Arg72Pro polymorphism and the risk of leukemia, but the CC genotype may increase the risk of ALL TP53 Arg72Pro polymorphism CC genotype may increase the risk of ALL but is not associated with AML. Further large-scale, well-designed studies are needed to confirm our results.

Neurocytoma arising from a mature ovary teratoma: a case report.

Central neurocytoma/extraventricular neurocytoma is a central nervous system (CNS) tumor composed of uniform round cells with neuronal differentiation. The typical lesions of central neurocytoma/extraventricular neurocytoma are at the interventricular foramen of the lateral ventricles (central neurocytoma) or brain parenchyma (extraventricular neurocytoma). Mature teratoma is a benign germ cell tumor commonly found in young women. Herein, we report a 24-year-old female with neurocytoma in a mature teratoma of the right ovary. The histological examinations showed mature epidermis, skin appendages, adipose and bone tissues in the tumor; microscopic foci of immature cartilage tissues were also found in some parts. In addition, massive solid sheets and uniform round tumor cells were found in the neuroectodermal tissues, with the formation of neuropil-like islands. Immunohistochemical examinations showed that the tumor cells were synaptophysin- and NeuN-positive but GFAP-negative. Based on these findings, the woman was diagnosed with neurocytoma arising from mature ovary teratoma, with microscopic foci of immature cartilage tissues. This is the fourth case report of neurocytoma outside the CNS to date.

Overexpression of δ-catenin is associated with a malignant phenotype and poor prognosis in colorectal cancer.

Little is known regarding the expression or clinical significance of δ-catenin, a member of the catenin family, in colorectal cancer (CRC). The present study examined the expression of δ-catenin using immunohistochemistry in 110 cases of CRC, including 70 cases with complete follow­up records and 40 cases with paired lymph node metastases. In addition, δ­catenin mRNA and protein expression were compared in 30 pairs of matched CRC and normal colorectal tissues by reverse transcription quantitative polymerase chain reaction and western blot analysis. δ­Catenin was weakly expressed or absent in the cytoplasm of normal intestinal epithelial cells, whereas positive δ­catenin expression localized to the cytoplasm was observed in CRC cells. The rate of positive δ­catenin expression in CRC (68.18%; 75/110) was significantly higher than that in normal colorectal tissues (36.7%; 11/30; P<0.001). In addition, δ­catenin mRNA and protein expression were significantly increased in CRC tissues compared to those in their matched normal tissues (all P<0.05). The expression of δ­catenin in stage III­IV CRC was higher than that in stage I­II CRC, and the expression of δ­catenin in the tumors of patients with lymph node metastases was higher than that in patients without lymph node metastases. Kaplan­Meier survival curves demonstrated that the survival time of patients with positive δ­catenin expression was shorter than that of patients with negative δ­catenin expression (P=0.005). Furthermore, Cox multivariate analysis indicated that the tumor, nodes and metastasis stage (P=0.02) and positive δ-catenin expression (P=0.033) were independent prognostic factors in CRC. The present study therefore indicated that δ-catenin may be a suitable independent prognostic factor for CRC.

F-box protein FBXO22 mediates polyubiquitination and degradation of KLF4 to promote hepatocellular carcinoma progression.

Kruppel-like factor 4 (KLF4), a member of the KLF family of transcription factors, has been considered as a crucial tumor suppressor in hepatocellular carcinoma (HCC). Using affinity purifications and mass spectrometry, we identified FBXO22, Cullin1 and SKP1 as interacting proteins of KLF4. We demonstrate that F-box only protein 22 (FBXO22) interacts with and thereby destabilizes KLF4 via polyubiquitination. As a result, FBXO22 could promote HCC cells proliferation both in vitro and in vivo. However, KLF4 deficiency largely blocked the proliferative roles of FBXO22. Importantly, FBXO22 expression was markedly increased in human HCC tissues, which was correlated with down-regulation of KLF4. Therefore, our results suggest that FBXO22 might be a major regulator of HCC development through direct degradation of KLF4.

Bufalin Induces Mitochondria-Dependent Apoptosis in Pancreatic and Oral Cancer Cells by Downregulating hTERT Expression via Activation of the JNK/p38 Pathway.

Bufalin, a digoxin-like active component of the traditional Chinese medicine Chan Su, exhibits potent antitumor activities in many human cancers. Bufalin induces mitochondria-dependent apoptosis in cancer cells, but the detailed molecular mechanisms are largely unknown. hTERT, the catalytic subunit of telomerase, protects against mitochondrial damage by binding to mitochondrial DNA and reducing mitochondrial ROS production. In the present study, we investigated the effects of bufalin on the cell viability, ROS production, DNA damage, and apoptosis of CAPAN-2 human pancreatic and CAL-27 human oral cancer cells. Bufalin reduced CAPAN-2 and CAL-27 cell viability with IC50 values of 159.2 nM and 122.6 nM, respectively. The reduced cell viability was accompanied by increased ROS production, DNA damage, and apoptosis and decreased expression of hTERT. hTERT silencing in CAPAN-2 and CAL-27 cells by siRNA resulted in increased caspase-9/-3 cleavage and DNA damage and decreased cell viability. Collectively, these data suggest that bufalin downregulates hTERT to induce mitochondria-dependent apoptosis in CAPAN-2 and CAL-27 cells. Moreover, bufalin increased the phosphorylation of JNK and p38-MAPK in CAPAN-2 and CAL-27 cells, and blocking the JNK/p38-MAPK pathway using the JNK inhibitor SP600125 or the p38-MAPK inhibitor SB203580 reversed bufalin-induced hTERT downregulation. Thus, the JNK/p38 pathway is involved in bufalin-induced hTERT downregulation and subsequent induction of apoptosis by the mitochondrial pathway.

Co-expression of delta-catenin and RhoA is significantly associated with a malignant lung cancer phenotype.

Delta-catenin, a member of the p120-catenin subfamily, and the Rho GTPase RhoA both have roles in the regulation of the cytoskeleton. In this study, we found that delta-catenin positive expression and RhoA over-expression is consistently found in non-small cell lung cancer, but not in normal lung tissue, and that their co-expression was significantly associated with histological type, differentiation, pTNM stage, lymphatic metastasis and a poor prognosis. We also demonstrate that delta-catenin can directly interact with RhoA and regulate its activity, which in turn mediates tumor invasion and metastasis.

Malignant solitary fibrous tumor of breast: a rare case report.

Solitary fibrous tumor (SFT) is rare mesenchymal neoplasm that has been originally and most often documented in the pleura. Recently, the ubiquitous nature of the SFT has been recognized with reports of involvement of numerous sites all over the body such as: upper respiratory tract, somatic tissue, mediastinum, head, and neck. Less than 10 cases SFT of breast have been reported. Herein, we presented a 52-year-old Asian female with SFT of breast, this tumor showed predominant malignant features. To our knowledge, SFT of breast with such malignant evidence is extremely rare.

Delta-catenin promotes the proliferation and invasion of colorectal cancer cells by binding to E-cadherin in a competitive manner with p120 catenin.

δ-Catenin is the only member of the p120 catenin (p120ctn) subfamily whose normal pattern of expression is restricted to the brain. Similar to p120ctn, δ-catenin can bind to the juxtamembrane domain of E-cadherin. We examined the expression of δ-catenin, p120ctn, and E-cadherin using immunohistochemistry in 95 cases of colorectal cancer (CRC) and 15 normal colon tissues. Co-immunoprecipitation was used to examine whether δ-catenin competed with p120ctn to bind E-cadherin in CRC cells. The effects of δ-catenin overexpression or siRNA-mediated knockdown on the proliferation and invasive ability of CRC cells were investigated using the MTT and Matrigel invasion assays. The results showed that positive δ-catenin expression was significantly more frequent in CRC compared to normal colon tissues and associated with poor differentiation, stage III-IV disease, and lymph node metastasis in CRC (all P < 0.05). In two CRC cell lines, δ-catenin bound to E-cadherin in competition with p120ctn. Overexpression of δ-catenin promoted the proliferation and invasion of CRC cells; knockdown of δ-catenin reduced CRC cell proliferation and invasion. In conclusion, we speculate that overexpression of δ-catenin reduces the expression of E-cadherin and alters the balance between E-cadherin and p120ctn, which in turn affects the formation of intercellular adhesions and promotes invasion and metastasis in CRC.