PlexinA1 promotes gastric cancer migration through preventing MICAL1 protein ubiquitin/proteasome-mediated degradation in a Rac1-dependent manner.

Metastasis promotes the development of tumors and is a significant cause of gastric cancer death. For metastasis to proceed, tumor cells must become mobile by modulating their cytoskeleton. MICAL1 (Molecule Interacting with CasL1) is known as an actin cytoskeleton regulator, but the mechanisms by which it drives gastric cancer cell migration are still unclear. Analysis of gastric cancer tissues revealed that MICAL1 expression is dramatically upregulated in stomach adenocarcinoma (STAD) samples as compared to noncancerous stomach tissues. Patients with high MICAL1 expression had shorter overall survival (OS), post-progression survival (PPS) and first-progression survival (FPS) compared with patients with low MICAL1 expression. RNAi-mediated silencing of MICAL1 inhibited the expression of Vimentin, a protein involved in epithelial-mesenchymal transition. This effect correlates with a significant reduction in gastric cancer cell migration. MICAL1 overexpression reversed these preventive effects. Immunoprecipitation experiments and immunofluorescence assays revealed that PlexinA1 forms a complex with MICAL1. Importantly, specific inhibition of PlexinA1 blocked the Rac1 activation and ROS production, which, in turn, impaired MICAL1 protein stability by accelerating MICAL1 ubiquitin/proteasome-dependent degradation. Overexpression of PlexinA1 enhanced Rac1 activation, ROS production, MICAL1 and Vimentin expressions, and favored cell migration. In conclusion, this study identified MICAL1 as an important facilitator of gastric cancer cell migration, at least in part, by affecting Vimentin expression and PlexinA1 promotes gastric cancer cell migration by binding to and suppressing MICAL1 degradation in a Rac1/ROS-dependent manner.

Association between CD10 expression and phyllodes tumor: A retrospective case control study.

The present study aimed to explore the association between immunohistochemical markers and phyllodes tumor (PT). The retrospective case control study included biopsies from patients with PT who underwent surgical treatment, and patients with fibronenoma (FA), diagnosed in our hospital from October 2014 to May 2021. Differences in microscopic histopathological characteristics and expressions of common immunohistochemical markers (CD10, cluster of differentiation 117 marker, cluster of differentiation 34 marker, tumor protein P53, cell proliferation antigen) for different grades of PT and FA were analyzed. A total of 69 patients were enrolled, of them 34 with PT (12 with benign PT, 13 with borderline PT, and 9 with malignant PT) and 35 with FA. With the increase of tumor malignancy, significant enlargement trend was noted; for FA, most tumor boundaries were well-defined, the stromal distribution was homogeneous, the stromal cellularity was small. In contrast for PT, as the degree of malignancy increased, tumor boundary gradually became ill-defined and the stromal distribution was heterogeneous; stromal cellularity and stromal overgrowth had increased significantly (All P < .05). Multivariate analysis showed that among other markers only CD10 expression (OR = 0.67, 95%CI: -0.88, 2.22, P < .05) was independently associated with PT. The study showed that in addition to histological features, CD10 expression was independently associated with PT and has a potential to be used as a differentiation marker.

Population pharmacokinetic modeling and simulation for nirmatrelvir exposure assessment in Chinese older patients with COVID-19 infection.

Nirmatrelvir is an effective component of Paxlovid, the first oral antiviral drug granted emergency use authorization by the FDA. Nirmatrelvir is prescribed extensively in older adult patients to treat the coronavirus disease 2019 (COVID-19) infection. In this study, population pharmacokinetic modeling with clinical study data was employed to explore the pharmacokinetic profile of nirmatrelvir in older adult Chinese patients with COVID-19 infection. The result suggests that the pharmacokinetic profile of nirmatrelvir can be described by a one-compartment model with first-order absorption and elimination in this study population. The calculated apparent clearance (CL/F), apparent volumes of distribution (V/F), and absorption rate constant (ka) for the typical patient were 4.16 L/h, 39.1 L, and 0.776, respectively. The area under the curve (AUC) of nirmatrelvir in the typical Chinese older adult was approximately three-fold higher than the AUCs in Chinese and Western young adult volunteers. At the same doses, the simulated AUCs were increased by 26%, 43%, 72%, and 135% in virtual populations with creatinine clearances of 60, 45, 30, and 15 mL/min, respectively. Our research provides an instructive reference for nirmatrelvir dose selection in older Chinese adults.

Prognostic impact of MICALL1 and associates with immune infiltration in liver hepatocellular carcinoma patients.

BACKGROUND: Liver hepatocellular carcinoma (LIHC) is one of the most malignancy over the world. Previous studies have proven that Molecules Interacting with CasL-Like 1 (MICALL1) participated in cellular trafficking cascades, while there has no study to explore the function and carcinogenic mechanism MICALL1 in LIHC. METHODS: We aimed to investigate the relationship between MICALL1 mRNA expression and LIHC using TCGA database. The expression of MICALL1 protein in clinic samples were examined by UALCAN database. Kaplan-Meier method was used for survival analysis. Logistic regression and Cox regression were performed to evaluate the prognostic significance of MICALL1. The MICALL1-binding protein were built by the STRING tool. Enrichment analysis by GO, KEGG and GSEA was used to explore possible function of MICALL1. The ssGSEA method was used to investigate the association between MICALL1 expression and the immune infiltration level in LIHC. RESULTS: The expression and prognostic value of different MICAL family members in LIHC were evaluated. The expression of MICALL1 was significantly increased at both the transcript and protein levels in LIHC tissues. Further, the LIHC patients with high MICALL1 levels showed a worse OS, DSS and PFI. Some clinicopathologic features were identified to be related to MICALL1 expression in LIHC included clinical T stage, pathologic stage, histologic grade and AFP concentration. Univariate and multivariate survival analysis showed that MICALL1 was an independent prognostic marker for OS and DSS. Further enrichment analysis revealed that the K-RAS, TNFα/NF-κB and inflammatory response were significantly enriched in the high MICALL1 expression group. Immune infiltration analysis showed that high MICALL1 expression was correlated with infiltration level of macrophage cells, Th2 cells and some other immune cell types, including TFH. CONCLUSIONS: MICALL1 expression was significantly associated with immune cell infiltration and may regarded as a promising prognostic biomarker for LIHC patients.

Cytoplasmic localization of IRF5 induces Wnt5a/E-cadherin degradation and promotes gastric cancer cells metastasis.

IRF5, a nucleoplasm shuttling protein, is a pivotal transcription factor regulating immune system activity. It's well known that immunosuppression is involved in the development of gastric cancer. However, no data exist for the expression and function of IRF5 in gastric cancer. This study demonstrated that IRF5 was cytoplasm-enriched in gastric cancer cells. IRF5 promoted gastric cancer cell migration, which involved the inhibition of Wnt5a and E-cadherin proteins expression. IRF5 (LA) localized in nucleus had no significant effect on Wnt5a and E-cadherin expressions, while mutation of IRF5 (ΔNLS), which prevents IRF5 nuclear translocation, had more impact on these inhibitory effects. In addition, degradation rates of both Wnt5a and E-cadherin were enhanced by resiquimod, an IRF5 agonist. Further in vivo experiments indicated that IRF5 knockout of gastric cancer cells repressed their pulmonary metastasis in nude mice. Finally, the expression and clinical significance of IRF5 were analyzed using gastric cancer tissue microarrays, which suggested that the expression of IRF5 varied procedurally in different progressive stages of gastric cancer. Our data revealed that IRF5 cytoplasmic localization were associated with Wnt5a and E-cadherin degradation and gastric cancer cell metastasis. Inhibiting IRF5 expression and/or its cytoplasmic localization may provide a novel target for gastric cancer therapy.

Outcome of antithrombotic therapy in cancer patients with catheter-related thrombosis: a systematic review.

Introduction: The guidelines' recommendations for anticoagulation in cancer patients with catheter-related thrombosis are unclear. The aim of this systematic review was to assess anticoagulation management in cancer patients with catheter-related thrombosis (CRT) based on previously published studies. Methods: As of June 10, 2023,we searched databases including PubMed, Embase, and Cochrane and included 11 observational studies that met the criteria. We evaluated 770 adults with active cancer and objectively confirmed patients with CRT who were using drugs including warfarin, LMWH, and new oral anticoagulants as antithrombotic therapy. Results: We extracted outcome data, including thrombosis recurrence, catheter dysfunction, major bleeding, and death, and performed a meta-analysis. Discussion: In this study we found that the risk of VTE recurrence was higher with rivaroxaban, the risk of bleeding and death appeared to be greater with warfarin, and although the risk of catheter dysfunction due to LMWH is a concern, it is still a more reasonable option for cancer patients with catheter-related thrombosis. Systematic Review Registration: http://www.clinicaltrials.gov, identifier (CRD42022367979).

Andrographolide elevates tumor necrosis factor-related apoptosis-inducing ligand lethality through reactive oxygen species accumulation and gasdermin E cleavage in breast cancer cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is selectively lethal to cancer cells and harmless to normal cells, making it a potential agent for cancer therapy. However, some breast cancer cells are resistant to TRAIL. This study revealed that andrographolide (Andro), an extract from Andrographis paniculate, a natural compound, sensitized breast cancer cells to TRAIL-induced tumor suppression; it identified apoptosis-associated protein regulation, reactive oxygen species accumulation, mitochondria membrane potential disruption, caspase cascade activation, and gasdermin-E cleavage to be involved in the tumor lethality mediated by Andro combined with TRAIL treatment. The flow cytometry results showed the combination of Andro and TRAIL repressed breast cancer cells by cell death induction, and the assessment of combined index indicated that the combined treatment with Andro and TRAIL repressed breast cancer cells synergistically. Blotting results displayed Andro and TRAIL combination elevated TRAIL-associated receptors, death receptors 4 and 5, at protein levels. These results provided critical insight into breast cancer patients' therapy and exploration direction for TRAIL clinical application.

Elevation of effective p53 expression sensitizes wild-type p53 breast cancer cells to CDK7 inhibitor THZ1.

BACKGROUND: The cyclin-dependent kinase 7 (CDK7) inhibitor THZ1 represses multiple cancer cells. However, its tumor-repressive efficiency in wild-type p53 breast cancer cells remains controversial. METHODS: We conducted various assays, including CCK8, colony formation, flow cytometry, western blotting, and lactate dehydrogenase release detection, to clarify whether p53 elevation sensitizes breast cancer cells to THZ1. RESULTS: We found that upregulating functional p53 contributes to the increased sensitivity of breast cancer cells to THZ1. Increased THZ1 sensitivity requires active p53 and an intact p53 pathway, which was confirmed by introducing exogenous wild-type p53 and the subsequent elevation of THZ1-mediated tumor suppression in breast cancer cells carrying mutant p53. We confirmed that p53 accumulates in the nucleus and mitochondria during cell death. Furthermore, we identified extensive transcriptional disruption, rather than solely CDK7 inhibition, as the mechanism underlying the nutlin-3 and THZ1-induced death of breast cancer cells. Finally, we observed the combined nutlin-3 and THZ1 treatment amplified gasdermin E cleavage. CONCLUSION: Enhanced sensitivity of breast cancer cells to THZ1 can be achieved by increasing effective p53 expression. Our approach may serve as a potential treatment for patients with breast cancer resistant to regular therapies. Video Abstract.

MICAL2 contributes to gastric cancer cell migration via Cdc42-dependent activation of E-cadherin/ß-catenin signaling pathway.

BACKGROUND: Gastric cancer is a common and lethal human malignancy worldwide and cancer cell metastasis is the leading cause of cancer-related mortality. MICAL2, a flavoprotein monooxygenase, is an important regulator of epithelial-to-mesenchymal transition. The aim of this study was to explore the effects of MICAL2 on gastric cancer cell migration and determine the underlying molecular mechanisms. METHODS: Cell migration was examined by wound healing and transwell assays. Changes in E-cadherin/ß-catenin signaling were determined by qPCR and analysis of cytoplasmic and nuclear protein fractions. E-cadherin/ß-catenin binding was determined by co-immunoprecipitation assays. Cdc42 activity was examined by pulldown assay. RESULTS: MICAL2 was highly expressed in gastric cancer tissues. The knockdown of MICAL2 significantly attenuated migratory ability and ß-catenin nuclear translocation in gastric cancer cells while LiCl treatment, an inhibitor of GSK3ß, reversed these MICAL2 knockdown-induced effects. Meanwhile, E-cadherin expression was markedly enhanced in MICAL2-depleted cells. MICAL2 knockdown led to a significant attenuation of E-cadherin ubiquitination and degradation in a Cdc42-dependent manner, then enhanced E-cadherin/ß-catenin binding, and reduced ß-catenin nuclear translocation. CONCLUSIONS: Together, our results indicated that MICAL2 promotes E-cadherin ubiquitination and degradation, leading to enhanced ß-catenin signaling via the disruption of the E-cadherin/ß-catenin complex and, consequently, the promotion of gastric cell migration. Video Abstract.

MCM10 is a Prognostic Biomarker and Correlated With Immune Checkpoints in Ovarian Cancer.

Background: Microchromosome maintenance protein 10 (MCM10) is required for DNA replication in all eukaryotes, and it plays a key role in the development of many types of malignancies. However, we currently still do not know the relationship between MCM10 and ovarian cancer (OV) prognosis and immune checkpoints. Methods: The Gene Expression Profiling Interactive Analysis and Tumor Immunology Estimation Resource (TIMER) databases were used to investigate MCM10 expression in Fan cancer. The Kaplan-Meier Plotter and PrognoScan were used to assess the relationship between MCM10 and OV prognosis. The LinkedOmics database was used to analyze the MCM10 co-expression network and explore GO term annotation and the KEGG pathway. The relationship between MCM10 expression and immune infiltration in OV was investigated using the Tumor Immunology Estimation Resource database. cBioPortal database was used to explore the relationship between MCM10 expression and 25 immune checkpoints. Finally, quantitative real-time polymerase chain reaction (qRT-PCR) was performed to detect MCM10 expression. The prognosis was also analyzed by distinguishing between high and low expression groups based on median expression values. Results: The results of the three data sets (220,651_s_at, 222,962_s_at and 223,570_at) in KM Plotter all indicated that the overall survivalof the high MCM10 expression group was lower than that of the low expression group OV, and the results of GSE9891 also reached the same conclusion. The expression level of MCM10 was negatively correlated with B cells and CD8+T cells, and positively correlated with CD4+T Cells and Macrophages. GO term annotation and KEGG pathway analysis showed that the co-expressed genes of MCM10 were mainly enriched in cell cycle and DNA replication. The alterations in MCM10 coexisted statistically with the immune checkpoints CTLA4, TNFSF4, TNFSF18, CD80, ICOSLG, LILRB1 and CD200. PCR results displayed that MCM10 was highly expressed in OV tissues, and the increased expression of MCM10 was significantly associated with poor overall survival. Conclusion: These results demonstrated that high expression of MCM10 was associated with poor prognosis in OV and correlated with immune checkpoints.

MICAL2 Contributes to Gastric Cancer Cell Proliferation by Promoting YAP Dephosphorylation and Nuclear Translocation.

Dynamic cytoskeletal rearrangements underlie the changes that occur during cell division in proliferating cells. MICAL2 has been reported to possess reactive oxygen species- (ROS-) generating properties and act as an important regulator of cytoskeletal dynamics. However, whether it plays a role in gastric cancer cell proliferation is not known. In the present study, we found that MICAL2 was highly expressed in gastric cancer tissues, and this high expression level was associated with carcinogenesis and poor overall survival in gastric cancer patients. The knockdown of MICAL2 led to cell cycle arrest in the S phase and attenuated cell proliferation. Concomitant with S-phase arrest, a decrease in CDK6 and cyclin D protein levels was observed. Furthermore, MICAL2 knockdown attenuated intracellular ROS generation, while MICAL2 overexpression led to a decrease in the p-YAP/YAP ratio and promoted YAP nuclear localization and cell proliferation, effects that were reversed by pretreatment with the ROS scavenger N-acetyl-L-cysteine (NAC) and SOD-mimetic drug tempol. We further found that MICAL2 induced Cdc42 activation, and activated Cdc42 mediated the effect of MICAL2 on YAP dephosphorylation and nuclear translocation. Collectively, our results showed that MICAL2 has a promotive effect on gastric cancer cell proliferation through ROS generation and Cdc42 activation, both of which independently contribute to YAP dephosphorylation and its nuclear translocation.

p53-GSDME Elevation: A Path for CDK7 Inhibition to Suppress Breast Cancer Cell Survival.

Higher cyclin-dependent kinase (CDK7) expression is a character of breast cancer and indicates poor prognosis. Inhibiting CDK7 exhibited effective cancer cell suppression which implies the potential of CDK7 inhibition to be a method for anti-cancer treatment. Our study aimed to explore a novel mechanism of CDK7 inhibition for suppressing breast cancer cell survival. Here, we proved inhibiting CDK7 repressed breast cancer cell proliferation and colony formation and increased the apoptotic cell rate, with p53 and GSDME protein level elevation. When p53 was suppressed in MCF-7 cells, the decline of GSDME expression and associated stronger proliferation and colony formation could be observed. Since downregulation of GSDME was of benefit to breast cancer cells, p53 inhibition blocked the elevation of GSDME induced by CDK7 inhibition and retrieved cells from the tumor suppressive effect of CDK7 inhibition. Therefore, CDK7 inhibition exerted a negative effect on breast cancer cell proliferation and colony formation in a p53-GSDME dependent manner. These results revealed the CDK7-p53-GSDME axis could be a pathway affecting breast cancer cell survival.

Gasdermin E-mediated programmed cell death: An unpaved path to tumor suppression.

Hearing loss-associated protein gasdermin E (GSDME), an effector of secondary necrosis, has been identified in a new pathway of programmed cell death (PCD). GSDME epigenetic silencing and mutations resulting in loss-of-function have been reported in cancer tissues. Additionally, GSDME upregulation inhibits tumor proliferation as well as colony forming ability, and reduces the incidence of lymphatic metastasis, demonstrating that GSDME may act as a tumor suppressor. Here, we have focused on the molecular mechanisms of GSDME-mediated PCD, and tried to reveal the crosstalk between this cell death pathway and apoptosis, autophagy, GSDMD-mediated pyroptosis. Moreover, we concluded the anti-cancer activity of GSDME include forming permeable membranes, and triggering anti-cancer immunity. Thus, GSDME was potential to be a novel target for cancer prevention and treatment.

Targeting Mutated p53 Dependency in Triple-Negative Breast Cancer Cells Through CDK7 Inhibition.

BACKGROUND: Cyclin-dependent kinase 7 (CDK7) is crucial for cell cycle progression and gene expression transcriptional regulation, which are often not assessed in cancer developing process. CDK7 inhibitors have emerged as promising drugs for treating diverse cancers, including breast cancer. However, the mechanism behind its anticancer effect has not been well investigated. Here, the possible mechanism of CDK7 inhibitors for treating human triple-negative breast cancer (TNBC) has been studied. METHODS: The effects of CDK7 inhibitors on breast cancer cells have been identified by measuring cell viability (Cell Counting Kit-8) and cell proliferation and calculating colony formation. The short hairpin RNA and short interfering RNA were used for the construction of knockdown cells. To assess the expression of associated proteins, western blot was used. RESULTS: This study confirmed that, compared to hormone receptor-positive breast cancer cells, TNBC cells were more sensitive to THZ1, a novel CDK7 inhibitor. THZ1 treatment specifically downregulated mutated p53 in a dose- and time-dependent manner in TNBC cells with p53 mutation. Another CDK7 inhibitor, LDC4297, also potently interfered with the expression of mutated p53. Furthermore, endogenous CDK7 expression was positively correlated with the levels of mutated p53 in TNBC cells with p53 mutation. Downregulating mutated p53 expression significantly suppressed the proliferation of TNBC cells with p53 mutation. CONCLUSION: Our findings demonstrated that targeting CDK7 was an effective approach for the treatment of TNBC with p53 mutation.

MICAL2 Facilitates Gastric Cancer Cell Migration via MRTF-A-Mediated CDC42 Activation.

Aims and Hypothesis: Cell migration is driven by the reorganization of the actin cytoskeleton. Although MICAL2 is known to mediate the oxidation of actin filaments to regulate F-actin dynamics, relatively few studies have investigated the potential role of MICAL2 during cancer cell migration. Methods: The migratory ability of gastric cancer cells was measured by wound healing and transwell assays. The relationship between MICAL2 expression and MRTF-A nuclear localization was analyzed using gene overexpression and knockdown strategies. The production of reactive oxygen species (ROS) was evaluated by DCFH-DA staining. mRNA and protein levels of MMP9 were measured using qPCR and immunoblotting analysis. The activities of CDC42 and RhoA were assessed using pulldown assays. Results: Depletion of MICAL2 markedly reduced gastric cancer cell migration. Mechanistically, silencing of MICAL2 inhibited the nuclear translocation of MRTF-A in response to EGF and serum stimulation, whereas the contents of MRTF-A remained unchanged. Further analysis showed that silencing of MICAL2 decreased the activation of CDC42 as well as mRNA and protein levels of MMP9. Ectopic expression of MICAL2 augmented MRTF-A levels in the nucleus, and promoted the activation of CDC42, MMP9 expression, and gastric cancer cell migration. Moreover, silencing of MRTF-A inhibited the CDC42 activation induced by overexpression of MICAL2. In addition, MICAL2-induced ROS generation contributed to the effect exerted by MICAL2 on MRTF-A nuclear translocation. Conclusion: Together, these results provide evidence that MICAL2 facilitates gastric cancer cell migration via positive regulation of nuclear translocation of MRTF-A and subsequent CDC42 activation and MMP9 expression.

Malignant adenomyoepithelioma of the breast: A case report.

RATIONALE: Adenomyoepithelioma (AME) is a rare biphasic tumor consisting of epithelial and Myoepithelial cell. Most of the AME is benign, and only a few will progress to malignancy, Here, we report a case of low-grade malignant adenomyoepithelioma, and review the related literature, in a bid to investigate its clinical and pathological features and thus, enhance our understanding of this tumor. PATIENT CONCERNS: A 64-year-old woman visited our hospital with a 1-year history of a painless mass in her left breast. Physical examination revealed a palpable painless mass, measuring approximately 4.5 cm, in the left breast. DIAGNOSIS: Histological examination confirmed the diagnosis of malignant adenomyoepithelioma. INTERVENTIONS: The patient underwent local excision of the mass, with frozen section analysis revealing ductal carcinoma in situ. Mastectomy and sentinel lymph node biopsy were then performed. OUTCOMES: We conducted a one-year follow-up, and relapse was not observed. LESSONS: Treatment of AME remains controversial owing to the lack of high volume data and absence of prospective studies. Simple mastectomy is an acceptable treatment of this tumor.

FLCN Regulates HIF2α Nuclear Import and Proliferation of Clear Cell Renal Cell Carcinoma.

Aims and Hypothesis: This study aims to explore the specific molecular mechanism of folliculin (FLCN)-induced proliferation, migration, and invasion in clear cell renal cell carcinoma (ccRCC) and to investigate the relationship of FLCN and HIF2α. Folliculin was identified as a tumor suppressor gene. Its deletions and mutations are associated with a potential risk of renal cancer. At present, the specific molecular mechanism of FLCN-induced proliferation, invasion, and migration in ccRCC remains elusive. Methods: Cell proliferation was measured by flow cytometry analysis, while cell migration and invasion were measured by wound healing assay and Matrigel invasion assay. The expression of FLCN, HIF2α, MMP9, and p-AKT was examined by Western blotting. The cells were transfected with plasmids or siRNA to upregulate or downregulate the expression of FLCN. Immunofluorescence microscopy was carried out to display the HIF2α location. We also determined the correlation of FLCN and HIF2α in human renal cancer samples. Results: FLCN was combined with HIF2α in renal tubular epithelial and cancer cells, and it effectively alleviates the deterioration of renal cancer cells by degrading HIF2α. The silencing of FLCN showed a promotion of HIF2α protein expression via PI3K/mTORC2 pathway, which in turn led to an increase in downstream target genes Cyclin D1 and MMP9. Moreover, interfering with siFLCN advanced the time of HIF2α entry into the nucleus. Conclusions: Our study illustrated that FLCN could be a new therapeutic target in ccRCC. FLCN combined with HIF2α and identified a novel PI3K/mTORC2/HIF2α signaling in ccRCC cells.

MICAL-L2 Is Essential for c-Myc Deubiquitination and Stability in Non-small Cell Lung Cancer Cells.

Objectives: MICAL-L2, a member of the molecules interacting with the CasL (MICAL) family, was reported to be highly expressed in several types of cancers, however, the roles of MICAL-L2 in NSCLC pathogenesis remain to be explored. This study is designed to clarify the mechanisms by which MICAL-L2 participates in NSCLC cell proliferation. Materials and Methods: The expression levels of MICAL-L2 in human lung cancer samples were assessed by immunohistochemical staining. Cells were transfected with siRNA or plasmids to regulate MICAL-L2 expression. Cell proliferation was measured by EdU staining and CCK-8 assays. MICAL-L2 and phosphorylated/total c-Myc expression were examined by Western blotting analysis. Interaction between MICAL-L2 and c-Myc was assessed by immunofluorescence staining, Western blotting and co-immunoprecipitation assays. Western blotting, polyubiquitylation detection and protein stability assays were used to assess whether MICAL-L2 exerts its oncogenic effect via c-Myc. Results: We found that MICAL-L2 was highly expressed in human NSCLC. While overexpressing MICAL-L2 increased NSCLC cell proliferation, MICAL-L2 depletion decreased the proliferation of NSCLC cells, an effect that was linked to cell cycle arrest. MICAL-L2 physically interacted with the c-Myc protein and functioned to maintain nuclear c-Myc levels and prolonged its half-life. Knockdown of MICAL-L2 expression led to decreased c-Myc protein stability through accelerating polyubiquitylation of c-Myc and gave rise to c-Myc degradation. We further found that MICAL-L2 deubiquitinated c-Myc and blocked its degradation, presumably by inhibiting c-Myc phosphorylation at threonine residue 58. Conclusions: These results indicate that MICAL-L2 is a key regulator of c-Myc deubiquitination and stability in the nucleus, and this activity may be involved in promoting NSCLC cell proliferation.

SOX2 promotes hypoxia-induced breast cancer cell migration by inducing NEDD9 expression and subsequent activation of Rac1/HIF-1α signaling.

BACKGROUND: Hypoxia, a major condition associated with the tumor microenvironment, stimulates the migration of cancer cells. SOX2 is a powerful transcription factor that shows higher expression in several cancers, however, its role in hypoxia-induced breast cancer cell migration remains largely elusive. METHODS: The human breast cancer cell lines MDA-MB-231 and MDA-MB-468 were cultured under hypoxic conditions. The cell migration rate was determined using the wound-healing and transwell assays. The protein levels of SOX2, NEDD9 and HIF-1α were evaluated via western blotting analysis. The NEDD9 mRNA levels were evaluated using qPCR. The activation of Rac1 was detected with the pulldown assay. The binding of SOX2 to the NEDD9 promoter was checked using the luciferase reporter assay. We also transfected breast cancer cells with specific siRNA for SOX2, NEDD9 or the Rac1 inactive mutant (T17 N) to investigate the role of SOX2, NEDD9 and Rac1 in the response to hypoxia. RESULTS: Hypoxia markedly increased SOX2 protein levels in a time-dependent manner. SiRNA-mediated disruption of SOX2 inhibited cell migration under hypoxic conditions. Hypoxia also significantly augmented the NEDD9 mRNA and protein levels. Interestingly, SOX2 is a positive transcriptional regulator of NEDD9. Knockdown of SOX2 inhibited hypoxia-induced NEDD9 mRNA and protein expressions. Furthermore, hypoxia-induced upregulation of Rac1 activity and HIF-1α expression was attenuated by SOX2 or NEDD9 silencing, and Rac1-T17 N abolished HIF-1α expression as well as cell migration in cells subjected to hypoxia. CONCLUSIONS: Our results highlight the essential role of SOX2 in breast cancer cell motility. The upregulation of SOX2 under hypoxic conditions may facilitate NEDD9 transcription and expression, and subsequent activation of Rac1 and HIF-1α expression. This could accelerate breast cancer cell migration.