Cardiovascular outcomes associated with SGLT2 inhibitor therapy in patients with type 2 diabetes mellitus and cancer: a systematic review and meta-analysis.

BACKGROUND: Cancer patients with diabetes are at increased risk for cardiovascular diseases due to common risk factors and well-documented drug-associated cardiotoxicity. Sodium-glucose cotransporter-2 (SGLT2) inhibitors have shown cardiovascular benefits in patients with diabetes, but their effects on cancer patients remain unclear. This study aimed to evaluate the cardiovascular outcomes associated with SGLT2 inhibitor therapy in patients with concomitant diabetes and cancer. METHODS: We conducted a systematic review and meta-analysis of cohort studies comparing cardiovascular outcomes between cancer patients with diabetes receiving SGLT2 inhibitors and those not receiving SGLT2 inhibitors. PubMed, Embase, and the Cochrane Library were searched from inception to February 29, 2024. The primary outcome was all-cause mortality, and the secondary outcomes were heart failure hospitalization, and adverse events. Random-effect models were used to calculate pooled risk ratios (RR) with 95% confidence intervals (CI). Subgroup and sensitivity analyses were conducted to identify potential sources of heterogeneity and explore the effect of SGLT2 inhibitors on mitigating cardiotoxicity. RESULTS: Nine cohort studies involving 82,654 patients were included. SGLT2 inhibitor use was associated with a significantly lower risk of all-cause mortality (RR 0.46, 95% CI 0.31-0.68, P < 0.0001; I2 = 98%) and heart failure hospitalization (RR 0.49, 95% CI 0.30-0.81, P = 0.006; I2 = 21%) compared to non-use. The mortality benefit remained significant in patients receiving anthracycline chemotherapy (RR 0.50, 95% CI 0.28-0.89, P = 0.02; I2 = 71%). SGLT2 inhibitor use was also associated with a lower risk of sepsis (RR 0.32, 95% CI 0.23-0.44, P < 0.00001; I2 = 0%) and no increased risk of diabetic ketoacidosis (RR 0.66, 95% CI 0.20-2.16, P = 0.49; I2 = 0%). CONCLUSIONS: SGLT2 inhibitor therapy is associated with lower risks of all-cause mortality and heart failure hospitalization in patients with concomitant diabetes and cancer. These findings suggest that SGLT2 inhibitors may offer cardiovascular benefits in this high-risk population. Randomized controlled trials are needed to validate these findings and evaluate the safety and efficacy of SGLT2 inhibitors in specific cancer types and treatment regimens.

Iron deficiency in Taiwanese patients with heart failure and reduced ejection fraction.

BACKGROUND: Iron deficiency (ID) is a common comorbidity among patients with heart failure and reduced ejection fraction (HFrEF), and is associated with poorer outcomes independent of anemia. This study aimed to evaluate the prevalence and prognostic significance of ID in Taiwanese patients with HFrEF. METHODS: We included HFrEF patients from two multicenter cohorts at different periods. The multivariate Cox regression analysis was applied to assess the risk of outcomes associated with ID, accounting for the varying risk of death. RESULTS: Of the 3612 patients with HFrEF registered from 2013 to 2018, 665 patients (18.4%) had available baseline iron profile measurements. Of these, 290 patients (43.6%) were iron deficient; 20.2% had ID+/anemia+, 23.4% ID+/anemia-, 21.5% ID-/anemia+, and 34.9% ID-/anemia-. Regardless of anemia status, patients with coexisting ID had a higher risk than those without ID (all-cause mortality: 14.3 vs 9.5 per 100 patient-years, adjusted hazard ratio [HR] 1.33; 95% confidence interval [CI], 0.96-1.85; p = 0.091; cardiovascular mortality: 10.5 per 100 patient-years vs 6.1, adjusted HR 1.54 [95% CI, 1.03-2.30; p = 0.037]; cardiovascular mortality or first unplanned hospitalization for HF: 36.7 vs 19.7 per 100 patient-years, adjusted HR 1.57 [95% CI, 1.22-2.01; p < 0.001]). Among patients eligible for treatment in the IRONMAN trial design (43.9%), parenteral iron therapy was estimated to reduce heart failure hospitalizations and cardiovascular deaths by 13.7 per 100 patient-years. CONCLUSION: Iron profiles were tested in less than one-fifth of the Taiwanese HFrEF cohort. ID was present in 43.6% of tested patients and was independently associated with poor prognosis in these patients.

Risks and outcomes of critical limb ischemia in hemodialysis patients: a prospective cohort study.

Background: Peripheral arterial disease (PAD) is more common in patients receiving maintenance hemodialysis than in the general population. Critical limb ischemia (CLI), the most severe form of PAD, is associated with high amputation and mortality risk. However, few prospective studies are available evaluating this disease's presentation, risk factors and outcomes for patients receiving hemodialysis. Methods: The Hsinchu VA study, a prospective multicentre study, investigated the impact of clinical factors on cardiovascular outcomes of patients receiving maintenance hemodialysis from January 2008 until December 2021. We evaluated the presentations and outcomes of patients with newly diagnosed PAD and the correlations of clinical variables with newly diagnosed CLI. Results: Of 1136 study participants, 1038 had no PAD on enrolment. After a median follow-up period of 3.3 years, 128 had newly diagnosed PAD. Of these, 65 presented with CLI, and 25 underwent amputation or died from PAD. Patients presenting with CLI had more below-the-knee (52%) and multi-level (41%) disease, and completely occluded segments (41%), and higher risk for amputation or PAD-related death compared with patients without CLI (27.7% vs 9.5%, P = .01). After multivariate adjustment, disability, diabetes mellitus, current smoking and atrial fibrillation were significantly associated with newly diagnosed CLI. Conclusions: Patients undergoing hemodialysis had higher rates of newly diagnosed CLI than the general population. Those with disabilities, diabetes mellitus, smoking and atrial fibrillation may require careful examination for PAD. Trial registration: Hsinchu VA study, ClinicalTrials.gov identifier: NCT04692636.

Treatment with angiotensin receptor neprilysin inhibitor for Taiwan heart failure patients: Rationale and baseline characteristics of the TAROT-HF study.

BACKGROUND: This study used a real-world database to investigate the prescription patterns of sacubitril/valsartan (Sac/Val) among Taiwanese patients with heart failure with reduced ejection fraction (HFrEF). METHODS: The Treatment with Angiotensin Receptor neprilysin inhibitor fOr Taiwan Heart Failure patients (TAROT-HF) study is a principal investigator-initiated, multicenter, observational, retrospective study on Taiwanese HFrEF patients. A total of 1772 patients with HFrEF (mean age 62.5 years, 75.3% male, mean left ventricular ejection fraction [LVEF] 29.3%) who received Sac/Val at 10 hospitals between 2017 and 2018 were enrolled at the date of Sac/Val initiation. Among these patients, 585 (33%) initially received Sac/Val during acute decompensated heart failure (HF) hospitalization (TAROT-AHF arm), whereas 1187 (67%) initially received the same at the outpatient clinic (TAROT-CHF arm). RESULTS: A total of 1343 (75.8%) patients received an initial dose of 50 mg twice daily or fewer, whereas 422 (23.8%) received the standard initiation dose (100 mg twice daily). During outpatient Sac/Val initiation, the mean dosages were significantly higher than that following hospitalization (117 ± 55 mg vs 109 ± 57 mg; p = 0.014). Multivariate analysis identified younger age, higher systolic blood pressure, higher LVEF, prior use of renin-angiotensin system inhibitors, use of ivabradine, and a history of diabetes mellitus as independent factors for initiating a standard Sac/Val dose. Over a follow-up period of 18 months, incidences of cardiovascular death or first unplanned HF hospitalization were 18.69 and 33.11 per 100-person years for the TAROT-CHF and TAROT-AHF arms, respectively. CONCLUSION: The TAROT-HF study provided an opportunity to describe the clinical features of patients with HFrEF who received Sac/Val, assess the real-world utilization and efficacy of Sac/Val, and compare these patients with those included in prior registries.

Cytoplasmic, but not nuclear Nrf2 expression, is associated with inferior survival and relapse rate and response to platinum-based chemotherapy in non-small cell lung cancer.

BACKGROUND: Several studies have previously indicated that nuclear factor erythroid 2-related factor 2 (Nrf2) expression may promote tumor progression when the Keap1/Nrf2 pathway is activated, but few reports have demonstrated the role of cytoplasmic Nrf2 on tumorigenesis. METHODS: Immunohistochemistry was conducted to evaluate Nrf2 expression in 167 tumors from surgically-resected patients with non-small cell lung cancer (NSCLC). Univariate and multivariate analyses were performed to examine the association of Nrf2 expression with patients' prognosis. This study was conducted to examine the association of Nrf2 expression with tumor response to cisplatin-based chemotherapy. RESULTS: Among these tumors, 56 and 32 of 167 tumors expressed Nrf2 in the cytoplasm (34% for C+/N-) and in the cytoplasm/nucleus (19% for C+/N+), but not in the nucleus of tumor cells. Nrf2 was negatively expressed in the remainder of the tumor samples (C-/N-, 79 of 167, 47%). Univariate analysis indicated that patients with Nrf2 positive tumors (C+/N- plus C+/N+) had worse overall survival (OS), but not relapse-free survival (RFS) than with Nrf2 negative tumors (C-/N-). However, patients with C+/N- tumors possessed worse OS and RFS than those with Nrf2 negative tumors (C-/N-). Multivariate analysis further confirmed the prognostic significance of patients with Nrf2 positive and C+/N- tumors on OS and RFS, but not on RFS for patients with Nrf2 positive tumors. Patients with Nrf2 positive and C+/N- tumors were determined to more frequently have an unfavorable response to cisplatin-based chemotherapy than those with Nrf2 negative tumors. CONCLUSIONS: Cytoplasmic Nrf2 expression might potentially be used to predict poor prognosis and unfavorable response to cisplatin-based chemotherapy in patients with NSCLC. KEY POINTS: The expression of cytoplasmic Nrf2 showed a significant relationship with patients' response to cisplatin-based chemotherapy and influenced NSCLC prognosis. A proteasomal inhibitor such as carfilzomib might be used to improve the outcomes and therapeutic response to cisplatin-based chemotherapy in patients with tumors showing cytoplasmic Nrf2 expression.

Effectiveness of a Non-Taped Compression Dress in Patients Receiving Cardiac Implantable Electronic Devices.

BACKGROUND: Hematoma and skin damage are not uncommon after cardiac implantable electronic device (CIED) placement. The use of conventional hemostatic gauze and tape seems to be suboptimal in controlling these complications. This study aimed to evaluate the impact of a novel compression dress with a special pad and elastic bands for postoperative care. METHODS: A total of 175 CIED recipients were randomly divided into two groups: an experimental group with 85 patients who used a non-taped compression dress and a control group with 90 patients who used conventional gauze ball and elastic tapes. Skin integrity, hematoma, and oozing were compared between these two groups within 7 days after surgery. RESULTS: The mean age of the patients was 71.2 ± 13.3 years, and 83 (47.4%) were male. The results of the experimental vs. control group were as follows: skin integrity - 96.5% vs. 86.7% (p < 0.05); hematoma - 0% vs. 7.8% (p < 0.05); and oozing - 1.2% vs. 7.8% (p < 0.05). All observed endpoints were better in the experimental group. CONCLUSIONS: The use of a non-taped compression dress was associated with less unfavorable outcomes in terms of skin integrity and hemostasis.

RV-59 suppresses cytoplasmic Nrf2-mediated 5-fluorouracil resistance and tumor growth in colorectal cancer.

Our previous studies indicated that tumor invasion and 5-flurouracil (5-FU) resistance in colorectal cancer (CRC) was more affected by cytoplasmic localization of expressed Nrf2 (cNrf2) than by nuclear localization (nNrf2), indicating a need for novel antitumor agents to overcome 5-FU resistance and improve outcomes in patients with CRC. In the present study, 20 nitrogen-substituted anthra[1,2-c][1,2,5] thiadiazole-6,11-dione derivatives were collected to verify the compound most able to suppress cell growth in nuclear location sequence (NLS)-mutated Nrf2-transfected shNrf2-HCT116 stable clones that have high cNrf2 expression. The MTT assay indicated that these high-cNrf2-expressing shNrf2-HCT116 stable clones exhibited the lowest percentage survival when treated with RV-59 than with the other 19 compounds. As expected, the high-cNrf2-expressing cells also showed a higher value for the inhibitory concentration of 50% cell survival (IC50) for 5-FU when compared with Nrf2-knockdown HCT116 stable clones (17.74 µM vs. 5.34 µM). Interestingly, a lower RV-59 IC50 value was seen in the high-cNrf2-expressing stable clones than in the Nrf2-knockdown stable clones (3.55 µM vs. 16.81 µM). A similar low RV-59 IC50 value was observed in high-cNrf2-expressing NLS-mutated Nrf2-transfected shNrf2-HCT116 stable clones and p53 null (-/-) HCT116 cells (4.2 µM vs. 4.4 µM), whereas the IC50 value was 17.6 µM in normal colon FHC epithelial cells. Colony-forming assays confirmed that RV-59 treatment inhibited colony formation in NLS-mutated Nrf2-transfected shNrf2-HCT116 stable clones and in p53-/- HCT116 cells. Annexin-V/PI staining showed an involvement of apoptosis in the inhibitory effect of RV-59 on cell viability. A nude mouse xenograft tumor model showed that RV-59 efficiently suppressed tumor growth induced by transplanted NLS-mutated Nrf2-transfected shNrf2-HCT116 stable clones without affecting the body weight of the nude mice over the 37 day experimental period. These results strongly suggest that RV-59 may be a novel antitumor agent for suppression of 5-FU resistance and may have therapeutic potential for improving outcomes in patients with cNrf2-expressing tumors.

PSMD4 is a novel therapeutic target in chemoresistant colorectal cancer activated by cytoplasmic localization of Nrf2.

Nuclear Nrf2 (nNrf2) binding to the antioxidant response element may promote chemoresistance in colorectal cancer. However, the shuttling of Nrf2 between cytoplasm and nucleus in colon cancer cells has revealed the possibility that cytoplasmic location of Nrf2 (cNrf2) may play a specific role in chemoresistance. Transfection of a nuclear location sequence (NLS)-wild-type or NLS-mutated Nrf2 expression vector into a stable shNrf2 HCT116 clone using the MTT assay to examine whether chemoresistance induced by cNrf2 may be greater than nNrf2. Different specific inhibitors and small hairpin (sh)RNAs of targeting genes were used to verify the mechanistic action of cNrf2 in chemoresistance and further confirmed by an animal model. The association of cNrf2 with chemotherapeutic response in patients with colorectal cancer was statistically analyzed. The MTT assay indicated that cNrf2 may play a more important role than nNrf2 in conferring 5-fluorouracil (5-FU) and oxaliplatin resistance in HCT116 cells. Mechanistically, cNrf2-induced PSMD4 expression was responsible for chemoresistance in the NLS-mutated Nrf2-tranfected shNrf2HCT116 clone via the NF-κB/AKT/ß-catenin/ZEB1 cascades. The tumor burden induced by the NLS-mutated Nrf2-transfected shNrf2HCT116 clone was completely suppressed by treatment with 5-FU in combination with carfilzomib. A higher prevalence of unfavorable chemotherapeutic response in colorectal cancer patients with cNrf2, PSMD4-positive, p-p65-positive, and nuclear ß-catenin tumors was observed when compared to their counterparts. cNrf2 may play a more important role than nNrf2 in the chemoresistance of colorectal cancer. Activation of the NF-κB/AKT/ß-catenin/ZEB1 cascade by PSMD4 may be responsible for cNrf2-mediated chemoresistance. CONDENSED ABSTRACT: CNrf2 may play a more important role than nNrf2 in conferring 5-FU and oxaliplatin resistance. This observation in patients seemed to support the findings of the cell and animal models and suggested that PSMD4 may be responsible cNrf2-mediated chemoresistance via the NF-κB/AKT/ß-catenin /ZEB1 cascades.

PD-L1 confers resistance to EGFR mutation-independent tyrosine kinase inhibitors in non-small cell lung cancer via upregulation of YAP1 expression.

Programmed death ligand (PD-L1) expression was associated with tumor immune escape and subsequent poor prognosis in non-small cell lung cancer (NSCLC). This expression was higher in patients with EGFR-mutated NSCLC tumors than in those with EGFR-wild-type (WT) NSCLC tumors. We therefore hypothesized that poor prognosis mediated by higher PD-L1 may be partially through conferring resistance to tyrosine kinase inhibitor (TKI) in NSCLC regardless of EGFR mutation. The change in PD-L1 expression following gene manipulation corresponded with changes in expression of HIF-1α and YAP1. The expression of HIF-1α and YAP1 was concomitantly decreased by PD-L1 silencing or by ROS scavenger treatment (N-acetylcysteine, NAC); however, a ROS inducer treatment (pyocyanin) completely reversed the decreased expression of both genes in EGFR-mutated and -wild-type (WT) NSCLC cells. The MTT assay indicated that the inhibitory concentration of gefitinib yielding 50% cell viability (IC50) depended on PD-L1-mediated YAP1 expression. Mechanistic studies indicated that upregulation of YAP1 by PD-L1 might be responsible for EGFR mutation-independent TKI resistance via the ROS/HIF-1α axis. An unfavorable TKI response was more common in patient tumors with high PD-L1 or YAP1 mRNA expression than in patient tumors with low mRNA expression of these genes. In conclusion, PD-L1 might confer EGFR mutation-independent TKI resistance in NSCLC cells via upregulation of YAP1 expression.

Hepatitis B virus X protein represses LKB1 expression to promote tumor progression and poor postoperative outcome in hepatocellular carcinoma.

BACKGROUND: Hepatitis B virus X (HBx) protein plays critical roles in hepatitis B virus (HBV)-associated hepatocellular tumorigenesis through different molecular mechanisms, including inactivation of p53, a key transcription factor of liver kinase B1 (LKB1). We hypothesized that p53 inactivation by HBx protein could decrease LKB1 expression, thereby promoting tumor progression and poor outcomes in patients with HBV-associated hepatocellular carcinoma. METHODS: Manipulation strategies for HBx protein and/or p53 were used to verify that loss of LKB1 could promote colony formation and invasiveness in HepG2 and Hep3B cells. The expressions of HBx protein and LKB1 in 93 hepatocellular carcinomas (HCC) were also evaluated by immunohistochemistry. Kaplan-Meier and Cox regression models were used to assess the prognostic value of both HBx protein and LKB1 proteins in patients with hepatocellular carcinoma. RESULTS: Mechanistically, LKB1 expression was decreased at the transcriptional level after inactivation of p53 by HBx protein. Decreases in LKB1 expression were also associated with HBx protein-mediated colony formation and invasive capabilities. HBx protein, LKB1, and a combination of both proteins had prognostic significance for overall survival and relapse-free survival in our study population. CONCLUSION: The results from cell line experiments and evaluation of patient prognosis according to expression of HBx protein and LKB1 in their HCC strongly support the hypothesis that decreases in LKB1 expression by HBx protein-mediated p53 inactivation may play an important role in HBV-associated hepatocellular tumorigenesis.

An increase in BAG-1 by PD-L1 confers resistance to tyrosine kinase inhibitor in non-small cell lung cancer via persistent activation of ERK signalling.

High programmed cell death 1 ligand 1 (PD-L1) expression in tumour tissues was associated with poor outcomes in non-small cell lung cancer (NSCLC) due to evasion of tumour immune surveillance. However, the role of PD-L1 in tumour invasion and resistance to tyrosine kinase inhibitor (TKI) treatments is not fully understood. Here, we provide evidence to support the involvement of PD-L1 expression in the invasiveness and TKI resistance in NSCLC cells by increased Bcl-2-associated athanogene-1 (BAG-1) expression. The upregulation of BAG-1 transcription by PD-L1 was verified by constructing the BAG-1 promoters using the polymerase chain reaction (PCR) and deletion mutations for luciferase reporter assays. The results indicated that C/EBPß phosphorylation by extracellular signal-regulated kinase (ERK) signalling was responsible for PD-L1-mediated BAG-1 transcription. Mechanistically, the PD-L1-induced BAG-1 expression reciprocally increased PD-L1 expression due to persistent activation of ERK signalling, and it consequently conferred TKI resistance in NSCLC cells. The mechanistic action of this cell model was further confirmed by an animal model, affirming that PD-L1 conferred tumour invasiveness and TKI resistance via persistent activation of ERK signalling by the PD-L1/BAG-1 axis. We therefore suggest a combination of an ERK inhibitor with a TKI as a potential strategy for conquering PD-L1-mediated tumour invasion and TKI resistance in NSCLC patients whose tumours harbour high PD-L1/high BAG-1 expression.

A combination of anti-PD-L1 mAb plus Lm-LLO-E6 vaccine efficiently suppresses tumor growth and metastasis in HPV-infected cancers.

PD-1/PD-L1 immunotherapy is viewed as having clinical benefits in advanced cancers but is effective in only a few patients, suggesting that an efficient combination approach is needed to improve efficacy. Immunohistochemistry analysis indicated that PD-L1 expression was correlated with the E6 expression in tumors from 122 lung cancer patients. The poorest survival occurred in PD-L1-positive/E6-positive tumor. PD-L1 expression was increased by the expression of E6, but not the E7, oncoprotein in lung and cervical cancer cells. PD-L1 expression was responsible for E6-mediated colony formation and soft agar growth. Therefore, PD-L1 secreted from tumor cells may directly promote tumor progression, particularly in E6-positive tumors. Immune deficiency nude mice were used to test the possibility that combining anti-PD-L1 mAb with Lm-LLO-E6 vaccine could have a higher antitumor activity compared with anti-PD-L1 mAb or Lm-LLO-E6 vaccine alone. A greater antitumor activity was obtained with anti-PD-L1 mAb + Lm-LLO-E6 vaccine than with anti-PD-L1 mAb or Lm-LLO-E6 alone in subcutaneous and metastatic tumors induced by TL-1 and SiHa cells. The longest survival time for nude mice was observed in the anti-PD-L1 mAb + Lm-LLO-E6 vaccine group. In conclusion, an anti-PD-L1 mAb + Lm-LLO-E6 vaccine may be an efficient treatment for suppression of tumor growth and metastasis induced by HPV-infected cells.

The YAP1/SIX2 axis is required for DDX3-mediated tumor aggressiveness and cetuximab resistance in KRAS-wild-type colorectal cancer.

The mechanism underlying tumor aggressiveness and cetuximab (CTX) resistance in KRAS-wild-type (KRAS -WT) colorectal cancer remains obscure. We here provide evidence that DDX3 promoted soft agar growth and invasiveness of KRAS-WT cells, as already confirmed in KRAS-mutated cells. Mechanistically, increased KRAS expression induced ROS production, which elevated HIF-1α and YAP1 expression. Increased HIF-1α persistently promoted DDX3 expression via a KRAS/ROS/HIF-1α feedback loop. DDX3-mediated aggressiveness and CTX resistance were regulated by the YAP1/SIX2 axis in KRAS-WT cells and further confirmed in animal models. Kaplan-Meier and Cox regression analysis indicated that DDX3, KRAS, and YAP1 expression had prognostic value for OS and RFS in KRAS-WT and KRAS-mutated tumors, but SIX2 and YAP1/SIX2 were prognostic value only in KRAS-WT patients. The observation from patients seemed to support the mechanistic action of cell and animal models. We therefore suggest that combining YAP1 inhibitors with CTX may therefore suppress DDX3-mediated tumor aggressiveness and enhance CTX sensitivity in KRAS-WT colorectal cancer.

Fractional Flow Reserve Assessment of a Significant Coronary Stenosis Masked by a Downstream Serial Lesion.

Fractional flow reserve (FFR) has been recognized as an effective tool to determine functional significance in intermediate coronary lesions and FFR-guided percutaneous coronary intervention (PCI) improves clinical outcomes. However, hemodynamic interaction between serial stenoses within one coronary artery complicates the assessment of functional severity of each individual lesion. We present a case in which FFR measurement by intracoronary bolus injection of adenosine helps to make appropriate revascularization decision in serial stenoses when the procedures are performed systemically and properly.

Reduction of microRNA-184 by E6 oncoprotein confers cisplatin resistance in lung cancer via increasing Bcl-2.

MicroRNA-184 suppresses cell growth and survival via targeting c-Myc and Bcl- 2. We recently reported that miR-184 promotes tumor progression in non-small cell lung cancer via targeting CDC25A and c-Myc. We here hypothesized that miR-184 could be down-regulated by E6 oncoprotein to confer cisplatin resistance in NSCLC. Human papillomavirus (HPV) 16-positive lung cancer TL-1 and cervical cancer SiHa cells compared with HPV16-negative TL-10 and C33A cells were enrolled for E6 manipulation. MiR-184 expression levels were increased by E6-knockdown in TL-1 and SiHa cells, but decreased by E6-overexpression in TL-10 and C33A cells. The MTT assay showed that the inhibition concentration of cisplatin yielding for 50% cell viability was dependent on miR-184 levels. Bcl-2 de-targeted by E6-mediated miR- 184 reduction was responsible for cisplatin resistance. Luciferase reporter assay and real- time PCR analysis indicated that the miR-184 promoter activity and its expression were modulated by E6 and/or p53 manipulation. Chromatin immunoprecipitation (ChIP) assay confirmed that p53 was bound onto the miR-184 promoter and its binding activity was modulated by E6 and/or p53 manipulation. Among patients, high miR184 and high Bcl-2 mRNA expression was more commonly occurred in E6- positive tumors than in E6-negative tumors. Fifty-nine out of 136 patients receiving cisplatin-based chemotherapy were available for the retrospective study. Patients with low-mR-184, E6-positive, high-Bcl-2 tumors, and both combinations were more prevalently occurred unfavorable response to cisplatin-based chemotherapy than their counterparts. In conclusion, a decrease in miR-184 level by E6 oncoprotein may predict unfavorable response to cisplatin-based chemotherapy in HPV-infected NSCLC patients via increasing Bcl-2 expression.

Cytoplasmic localization of Nrf2 promotes colorectal cancer with more aggressive tumors via upregulation of PSMD4.

Differences in subcellular localization of Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) have been associated with poor outcomes in human cancers. However, the prognostic value of subcellular localization of Nrf2 in colorectal cancer and the underlying mechanism in tumor invasion remain unknown. We enrolled tumors from colorectal patients to evaluate Nrf2, NQO1, and HO-1 expression by immunohistochemistry. NQO1 and HO-1 positive tumors showed nearly complete expression of Nrf2 in the nucleus and/or showed partial expression in the nucleus/cytoplasm (nNrf2); however, tumors negative for NQO1 and HO-1 showed almost complete expression of Nrf2 in the cytoplasm and/or partial expression in the nucleus/cytoplasm (cNrf2). Kaplan-Meier and Cox regression analysis indicated poorer overall survival in patients with cNrf2 tumors than with nNrf2 tumors. Cell models provided evidence that cNrf2, rather than nNrf2, was responsible for cell invasion and soft agar growth triggered by activation of the NF-κB/AKT/ß-catenin cascade. Mechanistically, cNrf2 persistently increased PSMD4 expression by the HIF1α/ß-catenin axis, whereas PSMD4 reciprocally enhanced Nrf2 nuclear export by increasing CRM1 expression through p53 degradation. The mechanistic action of the cell model was further confirmed with a nude mouse animal model in which xenograft tumors induced by cNrf2 were nearly completely suppressed by the proteasomal inhibitor carfilzomib or the ß-catenin inhibitor XAV939. We therefore suggest that PSMD4 or ß-catenin might be potential targets for suppressing tumor aggressiveness, and consequently, improving outcomes in patients whose tumors express cNrf2.

DDX3 enhances oncogenic KRAS­induced tumor invasion in colorectal cancer via the ß­catenin/ZEB1 axis.

DDX3 plays a dual role in colorectal cancer; however, the role and underlying mechanism of DDX3 in colorectal tumorigenesis remains unclear. Here, we provide evidence that DDX3 enhances oncogenic KRAS transcription via an increase in SP1 binding to its promoter. Accelerating oncogenic KRAS expression by DDX3 promotes the invasion capability via the ERK/PTEN/AKT/ß-catenin cascade. Moreover, the ß-catenin/ZEB1 axis is responsible for DDX3-induced cell invasiveness and xenograft lung tumor nodule formation. The xenograft lung tumor nodules induced by DDX3-overexpressing T84 stable clone were nearly suppressed by the inhibitor of AKT (perifosine) or ß-catenin (XAV939). Among patients, high KRAS, positive nuclear ß-catenin expression and high ZEB1 were more commonly occurred in high-DDX3 tumors than in low-DDX3 tumors. High-DDX3, high-KRAS, positive nuclear ß-catenin tumors, and high-ZEB1 exhibited worse overall survival (OS) and relapse free survival (RFS) than their counterparts. In conclusion, DDX3 may play an oncogenic role to promote tumor growth and invasion in colon cancer cells via the ß-catenin/ZEB1 axis due to increasing KRAS transcription. We therefore suggest that AKT or ß-catenin may potentially act as a therapeutic target to improve tumor regression and outcomes in colorectal cancer patients who harbored high-DDX3 tumors.

DDX3 promotes tumor invasion in colorectal cancer via the CK1ε/Dvl2 axis.

DDX3, a subunit of CK1ε, phosphorylates Dvl2 to promote ß-catenin activation. Overexpression of the Dvl2 protein results in potent activation of ß-catenin/TCF signaling in colorectal cancer. Therefore, we hypothesized that DDX3 might promote tumor invasion via the CK1ε/Dvl2 axis due to ß-catenin/TCF activation. Western blotting showed that ß-catenin expression was decreased by DDX3 knockdown and increased by DDX3 overexpression in colorectal cancer cells. The TCF promoter activity and invasion capability were concomitantly increased and decreased by DDX3 manipulation in these cells. The invasion capability in colon cancer cells and xenograft lung tumor nodules induced by a DDX3-overexpressing T84 stable clone in tail-vein injection model were nearly suppressed by inhibitors of CK1ε (PF4800567) and ß-catenin/TCF signaling (XAV939). Among colorectal cancer patients, DDX3 expression was positively correlated with the expression of pDvl2 and nuclear ß-catenin in tumor tissues. The expression of pDvl2 occurred more frequently in high-nuclear than in low-nuclear ß-catenin tumors. A prognostic significance of DDX3, pDvl2, and nuclear ß-catenin on overall survival and relapse free survival was observed in this study population. We therefore suggest CK1ε or ß-catenin/TCF signaling as potential targets for improving tumor regression and outcomes in colorectal cancer, particularly tumors with high-DDX3/high-nuclear ß-catenin or high-DDX3/high-pDvl2/high-nuclear ß-catenin expression.

Mutant p53 confers chemoresistance in non-small cell lung cancer by upregulating Nrf2.

Nrf2 is a key transcription factor for genes coding for antioxidants, detoxification enzymes, and multiple drug resistance and it also confers resistance to anticancer drugs. Here, we hypothesized that mutant p53 could upregulate Nrf2 expression at the transcriptional level, thereby conferring cisplatin resistance in non-small cell lung cancer (NSCLC). Luciferase reporter assays and real-time PCR analysis indicated that the Nrf2 promoter activity and its mRNA levels were markedly suppressed by wild-type p53, but not by mutant p53. Chromatin immunoprecipitation (ChIP) further confirmed that wild-type p53 binds at the p53 putative binding site to block Sp1 binding to the Nrf2 promoter and consequently to suppress the Nrf2 promoter activity. The MTT assay indicated that an increase in Nrf2 expression by mutant p53 is responsible for cisplatin resistance. Among the Nrf2 downstream genes, Bcl-2 and Bcl-xL contribute more strongly to Nrf2-mediated cisplatin resistance when compared with heme oxygenase 1 (HO-1). Cox regression analysis showed that patients with high-Nrf2, high-Bcl-2, high-Bcl-xL mRNA tumors were more commonly occurred unfavorable response to cisplatin-based chemotherapy than their counterparts. The prognostic significance of Nrf2 mRNA levels on OS and RFS was also observed in patients who have received cisplatin-based chemotherapy, particularly in p53-mutant patients. Collectively, mutant p53 may confer cisplatin resistance via upregulation of Nrf2 expression, and Nrf2 mRNA level may predict chemotherapeutic response and outcomes in NSCLC.