Association between serum calcium levels and first stroke: A community-based nested case-control study.

Background: Evidence from epidemiologic studies has been limited and inconsistent regarding the role of serum calcium in stroke incidence risk. We aimed to evaluate the association between serum albumin-corrected calcium and the risk of the first stroke in the Chinese community-dwelling population. Methods: The study sample population was drawn from the "H-type Hypertension and Stroke Prevention and Control Project." Using a nested case-control study, a total of 1,255 first-stroke cases and 1,255 controls matched for age, sex, and village were included in the final data analysis. We measured the serum calcium by inductively coupled plasma mass spectrometry and assessed the associations between serum albumin-corrected calcium and first stroke using conditional logistic regression. Results: The overall mean (SD) serum albumin-corrected calcium was 8.9 (0.6) mg/dl. Compared with the middle tertile (8.7-9.1 mg/dl), the multivariate-adjusted odds ratios (95% CIs) of first total stroke associated with the lowest tertile and the highest tertile of serum albumin-corrected calcium were 1.37 (1.10, 1.70) and 1.30 (1.04, 1.62), respectively. Similar trends were observed for the first ischemic stroke. Consistently, restricted cubic spline showed a U-shaped association between serum albumin-corrected calcium and risk of total stroke and ischemic stroke. However, serum albumin-corrected calcium had no significant effect on first hemorrhagic stroke. No significant effect modification was observed in the subgroup analysis. Conclusions: Our results suggested a U-shaped association between serum calcium and first stroke; both low and high serum calcium levels were associated with an increased risk of the first stroke in the Chinese population.

Plasma 25-hydroxyvitamin D3 concentrations and incident risk of ischemic stroke in rural Chinese adults: New insight on ceiling effect.

OBJECTIVE: The aim of this study was to investigate the relationship between 25-hydroxyvitamin D3 (25[OH]D3) and ischemic stroke and its potential modifying factors in rural Chinese adults. METHODS: This nested case-control study was drawn from the H-type Hypertension and Stroke Prevention and Control Project, a community-based, prospective, observational study. Plasma 25(OH)D3 was measured by liquid chromatography with tandem quadrupole mass spectrometry. All stroke records came from the Chinese Center for Disease Control and Prevention. Multiple logistic regression models were used to evaluate the association between 25(OH)D3 and risk of ischemic stroke. RESULTS: We included 1079 participants with ischemic stroke and 1079 matched controls. Due to a non-linear relationship, the analyses were stratified by 25(OH)D3. For those with 25(OH)D3 < 20 ng/mL, there was a 15% reduction in the risk of ischemic stroke for each SD increment in 25(OH)D3 (odds ratio, 0.85; 95% confidence interval, 0.73-0.99). Compared with the lowest-tertile group, the risk of ischemic stroke decreased by 39% (odds ratio, 0.61; 95% confidence interval, 0.41-0.89) in the highest-tertile group. Furthermore, two effect modifiers were identified: diabetes and homocysteine level. Although participants with 25(OH)D3 ≥ 20 ng/mL had the lowest risk of ischemic stroke overall, there was no dose-response association within that range. CONCLUSIONS: An inverse dose-response association between 25(OH)D3 and incident risk of ischemic stroke in rural Chinese adults was only observed in those with 25(OH)D3 < 20 ng/mL, along with two effect modifiers. Higher levels of 25(OH)D3 did not confer additional benefit.

Association between baseline brachial-ankle pulse wave velocity and short-term risk of first stroke among Chinese hypertensive adults.

Little information is available on the association between brachial-ankle pulse wave velocity (baPWV) and the risk of stroke in Chinese H-type hypertension patients. Therefore, our study aimed to assess this association between baseline baPWV and short-term risk of first stroke and to propose a cutoff value of baPWV that could predict near cerebrovascular events. A total of 9787 hypertension patients without preexisting stroke who underwent baPWV measurement were included. The primary end points were first symptomatic stroke. Secondary end points were first ischemic stroke and first hemorrhagic stroke. During a median follow-up of 20.8 months, there was a total of 138 first strokes including 123 first ischemic strokes and 15 first hemorrhagic strokes. When baPWV was categorized in quartiles, the higher risks of first stroke (HR = 1.52; 95% CI: 1.05-2.21) and first ischemic stroke (HR = 1.53; 95% CI: 1.03-2.26) were found in participants in quartile 4 (≥21.31 m/s), compared with those in quartile 1-3 (<21.31 m/s). In receiver operating characteristic curve analysis, the best cutoff value of baPWV that could predict first stroke was 21.43 m/s. Higher baPWV (≥21.43 m/s) was significantly associated with increased risk of first stroke (HR = 1.60; 95% CI: 1.10-2.32) and first ischemic stroke (HR = 1.60; 95% CI: 1.08-2.37). In conclusion, higher baPWV levels were associated with an increased risk of first stroke among Chinese H-type hypertensive patients. In addition, a cutoff value of 21.43 m/s of baPWV was proposed that could predict the next two years' cerebrovascular events.

Association between plasma copper levels and first stroke: a community-based nested case-control study.

BACKGROUND AND AIMS: Uncertainty remains regarding the association between the risk of stroke and plasma copper levels in population with copper mostly in normal range due to limited data. We examined the association between baseline plasma copper and risk of first stroke in Chinese community-dwelling population. METHODS: We conducted a nested case control study from 'H-type Hypertension and Stroke Prevention and Control Project'. A total of 1255 first stroke cases and 1255 controls matched for age, sex and study site were included in the analysis. Conditional logistic regression analyses were performed to evaluate the association between plasma copper and first stroke. RESULTS: The overall mean of copper was 15.90 (2.66) µmol/L. In total, 94.26% participants' copper concentration was in the normal range by Mayo Clinic laboratory reference values. Smoothing curve showed that the associations of plasma copper with first stroke and its subtypes were linear. Each standard deviation (SD) increment of plasma copper was independently and positively associated with risk of first stroke [odds ratio (OR): 1.17, 95% confidence interval (CI): 1.07-1.28]. The multivariable ORs with 95% CIs for total stroke, ischemic stroke and hemorrhagic stroke in the highest versus the lowest quartile of plasma copper were 1.49 (1.16-1.90; P-trend = 0.001), 1.46 (1.12-1.92; P-trend = 0.004) and 2.05 (0.95-4.38; P-trend = 0.050), respectively. CONCLUSIONS: Baseline plasma copper was positively associated with risk of first ischemic stroke in an approximately linear fashion among Chinese community population (80.32% hypertensives), although their copper levels were mostly within the normal range according to current reference values. Our findings warrant additional investigation.

High-Density Lipoprotein Cholesterol and the Risk of First Ischemic Stroke in a Chinese Hypertensive Population.

BACKGROUND AND PURPOSE: Elevated high-density lipoprotein cholesterol (HDL-C) levels have displayed protection against cardiovascular disease. However, the association between specific lipoprotein classes and first ischemic stroke (IS) has not been well defined, particularly in higher-risk hypertensive populations. Our study evaluated the associations of HDL-C with first IS in a Chinese hypertensive population. METHODS: The study population was obtained from a community-based cohort study of hypertension in Lianyungang and Rongcheng, China. A nested case-control design was used that included 2463 identified first IS cases and 2463 controls matched by age ± 1 year, sex, and region. RESULTS: After adjusting for potential confounders, HDL-C was inversely associated with first IS (adjusted odds ratio [aOR]: 0.91; 95% confidence interval [CI]: 0.85-0.98). HDL-C levels of at least 65.4 mg/dL displayed a significant protective effect for first IS (aOR: 0.82; 95% CI: 0.69-0.98). Conversely, adverse effects of first IS were observed for low-density lipoprotein cholesterol (LDL-C) levels ≥138.1 mg/dL (aOR: 1.20; 95% CI: 1.02-1.42) and triglyceride (TG) levels ≥140.8 mg/dL (aOR: 1.27; 95% CI: 1.09-1.49). The risk associations of LDL-C and TG with first IS were attenuated in the presence of high HDL-C (≥53.0 mg/dL); an increased risk of first IS was only found in the presence of low HDL-C (<53.0 mg/dL) when LDL-C (aOR: 1.66; 95% CI: 1.19-2.31) and TG (aOR: 1.47; 95% CI: 1.17-1.84) were combined with HDL-C for analysis. CONCLUSION: In this community-based Chinese hypertensive population, higher HDL-C was a significant protective factor of first IS. These data add to the evidence describing the relationship between lipids and IS and suggest that HDL-C maybe is a marker of IS risk in Chinses hypertensive population.

Sex difference in the association between plasma selenium and first stroke: a community-based nested case-control study.

BACKGROUND: To date, there is no clearly defined association between plasma selenium levels and first stroke. We aimed to investigate the association between baseline plasma selenium and first stroke risk in a community-based Chinese population. METHODS: Using a nested case-control study design, a total of 1255 first stroke cases and 1255 matched controls were analyzed. Participant plasma selenium concentrations were measured by inductively coupled plasma mass spectrometry (ICP-MS), and the association of plasma selenium with first stroke risk was estimated by conditional logistic regression models. RESULTS: Overall, a non-linear negative association between plasma selenium and first total stroke and first ischemic stroke risks was found in males but not in females. Compared with participants with lower selenium levels (tertile 1-2, < 94.1 ng/mL), participants with higher selenium levels (tertile 3, ≥ 94.1 ng/mL) had significantly lower risks of first total stroke (OR 0.63; 95% CI 0.48, 0.83) and first ischemic stroke (OR 0.61; 95% CI 0.45, 0.83) in males but not in females with first total stroke (OR 0.92; 95% CI 0.69, 1.22) and first ischemic stroke (OR 0.89; 95% CI 0.65, 1.22). Furthermore, a stronger association between plasma selenium and first total stroke was found in males with higher vitamin E levels (≥ 13.5 µg/mL vs. < 13.5 µg/mL P-interaction = 0.007). No significant association was observed between plasma selenium and first hemorrhagic stroke risk in either males or females. CONCLUSION: Our study indicated a significant, non-linear, negative association between plasma selenium and first stroke in males but not in females. TRIAL REGISTRATION: ChiCTR1800017274 .

Plasma Magnesium Concentrations and Risk of Incident Cancer in Adults with Hypertension: A Nested Case-Control Study.

OBJECTIVE: The association between plasma magnesium and risk of incident cancer remains inconclusive in previous studies. We aimed to investigate the prospective relationship of baseline plasma magnesium concentrations with the risk of incident cancer and to examine possible effect modifiers. METHODS: A nested case-control study with 228 incident cancer cases and 228 matched controls was conducted using data from the China Stroke Primary Prevention Trial (CSPPT), a randomized, double-blind, controlled trial, conducted from May 2008 to August 2013. Study outcomes included incident cancer and its subtypes. RESULTS: When plasma magnesium concentrations were assessed as quartiles, a significantly higher incident risk of total cancer was found in participants in quartile 1 (<0.76 mmol/L; odds ratio [OR] = 2.70; 95% CI: 1.33-5.49) and quartile 4 (≥0.89 mmol/L; OR = 2.05; 95% CI: 1.12-3.76), compared with those in quartile 3 (0.83 to <0.89 mmol/L). In cancer site-specific analyses, similar trends were found for gastrointestinal cancer, esophageal cancer, gastric cancer, breast cancer, lung cancer, and other cancers. Furthermore, none of the variables, including age, sex, current smoking status, current alcohol intake, BMI, systolic blood pressure, and total cholesterol levels at baseline significantly modified the association between plasma magnesium and cancer risk. CONCLUSIONS: Both low and high plasma magnesium concentrations were significantly associated with an increased incident risk of cancer, compared with the reference concentrations of 0.83 to <0.89 mmol/L among hypertensive adults.

Serum folate modified the association between low-density lipoprotein cholesterol and carotid intima-media thickness in Chinese hypertensive adults.

BACKGROUND AND AIMS: While folate is known for its importance in cardiovascular health, it is unknown whether folate status can modify the association between low-density lipoprotein cholesterol (LDL-C) and carotid intima-media thickness (CIMT). We aimed to investigate this question in a Chinese hypertensive population, who are at high-risk of low folate and atherosclerosis. METHODS AND RESULTS: This report included 14,970 hypertensive adults (mean age 64.5 years; 40.3% male) from the China Stroke Primary Prevention Trial (CSPPT) and analyzed the fasting serum LDL-C and folate, and CIMT data obtained at the last follow-up visit. LDL-C was calculated using the Friedewald equation. Serum folate levels were measured by chemiluminescent immunoassay. CIMT was measured by ultrasound. Non-parametric smoothing plots, multivariate linear regression analysis, subgroup analyses and interaction testing were performed to examine the LDL-C-CIMI relationship and effect modification by folate. Consistent with graphic plots, multivariate linear regression showed that LDL-C levels were independently and positively associated with CIMT (ß = 7.69, 95%CI: 5.76-9.62). More importantly, the relationship between LDL-C and CIMT was significantly attenuated with increasing serum folate levels (1st tertile: ß = 10.06, 95%CI: 6.67-13.46; 2nd tertile: ß = 6.81, 95%CI: 3.55-10.07; 3rd tertile: ß = 5.96, 95%CI: 2.55-9.36; P-interaction = 0.045). Subgroup analyses showed the association between LDL-C and CIMT across serum folate tertiles was robust among various strata (all P-interaction >0.05). CONCLUSIONS: Among Chinese hypertensive adults, the serum folate levels could modify the association between LDL-C and CIMT. Our findings, if further confirmed, have important clinical implications.

Achieving blood pressure control targets in hypertensive patients of rural China - a pilot randomized trial.

BACKGROUND: This study aimed to test the feasibility and titration methods used to achieve specific blood pressure (BP) control targets in hypertensive patients of rural China. METHODS: A randomized, controlled, open-label trial was conducted in Rongcheng, China. We enrolled 105 hypertensive participants aged over 60 years, and who had no history of stroke or cardiovascular disease. The patients were randomly assigned to one of three systolic-BP target groups: standard: 140 to < 150 mmHg; moderately intensive: 130 to < 140 mmHg; and intensive: < 130 mmHg. The patients were followed for 6 months. DISCUSSION: The optimal target for systolic blood pressure (SBP) lowering is still uncertain worldwide and such information is critically needed, especially in China. However, in China the rates of awareness, treatment and control are only 46.9%, 40.7%, and 15.3%, respectively. It is challenging to achieve BP control in the real world and it is very important to develop population-specific BP-control protocols that fully consider the population's characteristics, such as age, sex, socio-economic status, compliance with medication, education level, and lifestyle. This randomized trial showed the feasibility and safety of the titration protocol to achieve desirable SBP targets (< 150, < 140, and < 130 mmHg) in a sample of rural, Chinese hypertensive patients. The three BP target groups had similar baseline characteristics. After 6 months of treatment, the mean SBP measured at an office visit was 137.2 mmHg, 131.1 mmHg, and 124.2 mmHg, respectively, in the three groups. Home BP and central aortic BP measurements were also obtained. At 6 months, home BP measurements (2 h after drug administration) showed a mean SBP of 130.9 mmHg in the standard group, 124.9 mmHg in the moderately intensive group, and 119.7 mmHg in the intensive group. No serious adverse events were recorded over the 6-month study period. Rates of adverse events, including dry cough, palpitations, and arthralgia, were low and showed no significant differences between the three groups. This trial provided real-world experience and laid the foundation for a future, large-scale, BP target study. TRIAL REGISTRATION: Feasibility Study of the Intensive Systolic Blood Pressure Control; ClinicalTrials.gov, ID: NCT02817503. Registered retrospectively on 29 June 2016.

VS-5584 mediates potent anti-myeloma activity via the upregulation of a class II tumor suppressor gene, RARRES3 and the activation of Bim.

The PI3K/mTOR/AKT pathway is an integral regulator of survival and drug resistance in multiple myeloma (MM). VS-5584 was synthesized with dual-specific and equipotent activity against mTORC1/2 and all four Class I PI3K isoforms so as to durably inhibit this pathway. We show that VS-5584 is highly efficacious against MM cell lines even in the presence of IL-6 and IGF-1 and that this growth inhibition is partially dependent on Bim. Importantly, VS-5584 triggers apoptosis in patient cells with a favorable therapeutic index. Gene expression profiling revealed a VS-5584-induced upregulation of RARRES3, a class II tumor suppressor gene. MM patient databases, UAMS and APEX, show that RARRES3 is under-expressed in 11q13 subsets which correlates with the reduced effectiveness of VS-5584 in 11q13 cell lines. Silencing RARRES3 expression significantly rescues VS-5584-induced cell death and increases cyclin D2 expression but not cyclin D1 or other cyclins implying a role for RARRES3 in cell cycle arrest. In vivo, VS-5584 significantly reduces the tumor burden of MM mouse xenografts. We further identified that VS-5584 synergised with Dexamethasone, Velcade, and exceptionally so with HDAC inhibitor, Panobinostat. Interestingly, this was consistently observed in several patient samples, proposing a promising novel clinical strategy for combination treatment especially in relapsed/refractory patients.

Inhibition of LIN28B impairs leukemia cell growth and metabolism in acute myeloid leukemia.

BACKGROUND: Current conventional chemotherapy for acute myeloid leukemia (AML) can achieve remission in over 70% of patients, but a majority of them will relapse within 5 years despite continued treatment. The relapse is postulated to be due to leukemia stem cells (LSCs), which are different from normal hematopoietic stem cells (HSCs). LIN28B is microRNA regulator and stem cell reprogramming factor. Overexpression of LIN28B has been associated with advance human malignancies and cancer stem cells (CSCs), including AML. However, the molecular mechanism by which LIN28B contributes to the development of AML remains largely elusive. METHODS: We modulated LIN28B expression in AML and non-leukemic cells and investigated functional consequences in cell proliferation, cell cycle, and colony-forming assays. We performed a microarray-based analysis for LIN28B-silencing cells and interrogated gene expression data with different bioinformatic tools. AML mouse xenograft model was used to examine the in vivo function of LIN28B. RESULTS: We demonstrated that targeting LIN28B in AML cells resulted in cell cycle arrest, inhibition of cell proliferation and colony formation, which was induced by de-repression of let-7a miRNA. On the other hand, overexpression of LIN28B promoted cell proliferation. Data point to a mechanism where that inhibition of LIN28B induces metabolic changes in AML cells. IGF2BP1 was confirmed to be a novel downstream target of LIN28B via let-7 miRNA in AML. Notably, ectopic expression of LIN28B increased tumorigenicity, while silencing LIN28B led to slow tumor growth in vivo. CONCLUSIONS: In sum, these results uncover a novel mechanism of an important regulatory signaling, LIN28B/let-7/IGF2BP1, in leukemogenesis and provide a rationale to target this pathway as effective therapeutic strategy.

Belinostat exerts antitumor cytotoxicity through the ubiquitin-proteasome pathway in lung squamous cell carcinoma.

There have been advances in personalized therapy directed by molecular profiles in lung adenocarcinoma, but not in lung squamous cell carcinoma (SCC). The lack of actionable driver oncogenes in SCC has restricted the use of small-molecule inhibitors. Here, we show that SCC cell lines displayed differential sensitivities to belinostat, a pan-histone deacetylase inhibitor. Phosphoproteomic analysis of belinostat-treated SCC cells revealed significant downregulation of the MAPK pathway, along with the induction of apoptosis. In cisplatin-resistant cells that demonstrated aberrant MAPK activation, combined treatment with belinostat significantly inhibited cisplatin-induced ERK phosphorylation and exhibited strong synergistic cytotoxicity. Furthermore, belinostat transcriptionally upregulated the F-box proteins FBXO3 and FBXW10, which directly targeted son of sevenless (SOS), an upstream regulator of the MAPK pathway, for proteasome-mediated degradation. Supporting this, suppression of SOS/ERK pathway by belinostat could be abrogated by inhibiting proteasomal activity either with bortezomib or with siRNA knockdown of FBXO3/FBXW10. Taken together, these preclinical data offer a novel understanding of the epigenetic mechanism by which belinostat exerts its cytotoxicity and supports the combination with cisplatin in clinical settings for chemorefractory SCC tumors.

LIN28B Activation by PRL-3 Promotes Leukemogenesis and a Stem Cell-like Transcriptional Program in AML.

PRL-3 (PTP4A3), a metastasis-associated phosphatase, is also upregulated in patients with acute myeloid leukemia (AML) and is associated with poor prognosis, but the underlying molecular mechanism is unknown. Here, constitutive expression of PRL-3 in human AML cells sustains leukemogenesis in vitro and in vivo Furthermore, PRL-3 phosphatase activity dependently upregulates LIN28B, a stem cell reprogramming factor, which in turn represses the let-7 mRNA family, inducing a stem cell-like transcriptional program. Notably, elevated levels of LIN28B protein independently associate with worse survival in AML patients. Thus, these results establish a novel signaling axis involving PRL-3/LIN28B/let-7, which confers stem cell-like properties to leukemia cells that is important for leukemogenesis.Implications: The current study offers a rationale for targeting PRL-3 as a therapeutic approach for a subset of AML patients with poor prognosis. Mol Cancer Res; 15(3); 294-303. ©2016 AACR.

Exocytosis of polyubiquitinated proteins in bortezomib-resistant leukemia cells: a role for MARCKS in acquired resistance to proteasome inhibitors.

PSMB5 mutations and upregulation of the ß5 subunit of the proteasome represent key determinants of acquired resistance to the proteasome inhibitor bortezomib (BTZ) in leukemic cells in vitro. We here undertook a multi-modality (DNA, mRNA, miRNA) array-based analysis of human CCRF-CEM leukemia cells and BTZ-resistant subclones to determine whether or not complementary mechanisms contribute to BTZ resistance. These studies revealed signatures of markedly reduced expression of proteolytic stress related genes in drug resistant cells over a broad range of BTZ concentrations along with a high upregulation of myristoylated alanine-rich C-kinase substrate (MARCKS) gene expression. MARCKS upregulation was confirmed on protein level and also observed in other BTZ-resistant tumor cell lines as well as in leukemia cells with acquired resistance to other proteasome inhibitors. Moreover, when MARCKS protein expression was demonstrated in specimens derived from therapy-refractory pediatric leukemia patients (n = 44), higher MARCKS protein expression trended (p = 0.073) towards a dismal response to BTZ-containing chemotherapy. Mechanistically, we show a BTZ concentration-dependent association of MARCKS protein levels with the emergence of ubiquitin-containing vesicles in BTZ-resistant CEM cells. These vesicles were found to be extruded and taken up in co-cultures with proteasome-proficient acceptor cells. Consistent with these observations, MARCKS protein associated with ubiquitin-containing vesicles was also more prominent in clinical leukemic specimen with ex vivo BTZ resistance compared to BTZ-sensitive leukemia cells. Collectively, we propose a role for MARCKS in a novel mechanism of BTZ resistance via exocytosis of ubiquitinated proteins in BTZ-resistant cells leading to quenching of proteolytic stress.

PRIMA-1 targets the vulnerability of multiple myeloma of deregulated protein homeostasis through the perturbation of ER stress via p73 demethylation.

Despite therapeutic advancement, multiple myeloma (MM) remains incurable with drug resistance being one of the main challenges in the clinic. Myeloma cells possess high protein secretory load, leading to increased intracellular endoplasmic reticulum (ER) stress. Hence, they are vulnerable to further perturbation to its protein homeostasis. In studying the therapeutic mechanism of PRIMA-1 (mutant-p53-reactivating-agent), we uncovered its novel p53-independent-mechanism that can be exploited for myeloma. Despite its inability in restoring the wild type-p53 protein conformation and transcriptional function in the mutant-p53-human-myeloma-cells, PRIMA-1 was efficacious against myeloma cells with differential p53 genotypes. Strikingly, cells without p53 expression demonstrated highest drug sensitivity. Genome-wide gene-expression analysis revealed the involvement of ER stress/UPR-pathway in inducing PRIMA-1-toxicity. UPR markers, HSP70, CHOP and GADD34, were significantly up-regulated, concomitantly with the induction of apoptosis. Furthermore, there was a global attenuation of protein synthesis, correlated with phospho-eIF2a up-regulation. Mechanistically, we identified that PRIMA-1 could cause the demethylation of TP73, through DNMT1 depletion, to subsequently enhance UPR. Of clinical significance, we observed that PRIMA-1 had additive therapeutic effects with another UPR-inducing-agent, bortezomib. Importantly, it can partially re-sensitize bortezomib-resistant cells to bortezomib. Given that MM is already stressed at the baseline in the ER, our results implicated that PRIMA-1 is a potential therapeutic option in MM by targeting its Achilles heel.

EZH2 phosphorylation by JAK3 mediates a switch to noncanonical function in natural killer/T-cell lymphoma.

The best-understood mechanism by which EZH2 exerts its oncogenic function is through polycomb repressive complex 2 (PRC2)-mediated gene repression, which requires its histone methyltransferase activity. However, small-molecule inhibitors of EZH2 that selectively target its enzymatic activity turn out to be potent only for lymphoma cells with EZH2-activating mutation. Intriguingly, recent discoveries, including ours, have placed EZH2 into the category of transcriptional coactivators and thus raised the possibility of noncanonical signaling pathways. However, it remains unclear how EZH2 switches to this catalytic independent function. In the current study, using natural killer/T-cell lymphoma (NKTL) as a disease model, we found that phosphorylation of EZH2 by JAK3 promotes the dissociation of the PRC2 complex leading to decreased global H3K27me3 levels, while it switches EZH2 to a transcriptional activator, conferring higher proliferative capacity of the affected cells. Gene expression data analysis also suggests that the noncanonical function of EZH2 as a transcriptional activator upregulates a set of genes involved in DNA replication, cell cycle, biosynthesis, stemness, and invasiveness. Consistently, JAK3 inhibitor was able to significantly reduce the growth of NKTL cells, in an EZH2 phosphorylation-dependent manner, whereas various compounds recently developed to inhibit EZH2 methyltransferase activity have no such effect. Thus, pharmacological inhibition of JAK3 activity may provide a promising treatment option for NKTL through the novel mechanism of suppressing noncanonical EZH2 activity.

Genome-wide pharmacologic unmasking identifies tumor suppressive microRNAs in multiple myeloma.

Epigenetic alterations have emerged as an important cause of microRNA (miRNA) deregulation. In Multiple Myeloma (MM), a few tumor suppressive miRNAs silenced by DNA hypermethylation have been reported, but so far there are few systemic investigations on epigenetically silenced miRNAs. We conducted genome-wide screening for tumor suppressive miRNAs epigenetically silenced in MM. Four Human MM Cell lines were treated with demethylating agent 5'azacytidine (5'aza). Consistently upregulated miRNAs include miR-155, miR-198, miR-135a*, miR-200c, miR-125a-3p, miR-188-5p, miR-483-5p, miR-663, and miR-630. Methylation array analysis revealed increased methylation at or near miRNA-associated CpG islands in MM patients. Ectopic restoration of miR-155, miR-198, miR-135a*, miR-200c, miR-663 and miR-483-5p significantly repressed MM cell proliferation, migration and colony formation. Furthermore, we derived a 33-gene signature from predicted miRNA target genes that were also upregulated in MM patients and associated with patient survival in three independent myeloma datasets. In summary, we have revealed important, epigenetically silenced tumor suppressive miRNAs by pharmacologic reversal of epigenetic silencing.

MEK Inhibition Overcomes Cisplatin Resistance Conferred by SOS/MAPK Pathway Activation in Squamous Cell Carcinoma.

Genomic analyses of squamous cell carcinoma (SCC) have yet to yield significant strategies against pathway activation to improve treatment. Platinum-based chemotherapy remains the mainstay of treatment for SCC of different histotypes either as a single-agent or alongside other chemotherapeutic drugs or radiotherapy; however, resistance inevitably emerges, which limits the duration of treatment response. To elucidate mechanisms that mediate resistance to cisplatin, we compared drug-induced perturbations to gene and protein expression between cisplatin-sensitive and -resistant SCC cells, and identified MAPK-ERK pathway upregulation and activation in drug-resistant cells. ERK-induced resistance appeared to be activated by Son of Sevenless (SOS) upstream, and mediated through Bim degradation downstream. Clinically, elevated p-ERK expression was associated with shorter disease-free survival in patients with locally advanced head and neck SCC treated with concurrent chemoradiation. Inhibition of MEK/ERK, but not that of EGFR or RAF, augmented cisplatin sensitivity in vitro and demonstrated efficacy and tolerability in vivo. Collectively, these findings suggest that inhibition of the activated SOS-MAPK-ERK pathway may augment patient responses to cisplatin treatment.

Phosphatase of regenerating liver-3 is regulated by signal transducer and activator of transcription 3 in acute myeloid leukemia.

Overexpression of protein-tyrosine phosphatase of regenerating liver 3 (PRL-3) has been identified in about 50% of patients with acute myeloid leukemia (AML). The mechanism of regulation of PRL-3 remains obscure. Signal transducer and activator of transcription 3 (STAT3), a latent transcriptional factor, has also been often found to be activated in AML. We first identified STAT3-consensus-binding sites in the promoter of PRL-3 genes. Then we experimentally validated the direct binding and transcriptional activation. We applied shRNA-mediated knockdown and overexpression approaches in STAT3(-/-) liver cells and leukemic cells to validate the functional regulation of PRL-3 by STAT3. A STAT3 core signature, derived through data mining from publicly available gene expression data, was employed to correlate PRL-3 expression in large AML patient samples. We discovered that STAT3 binds to the -201 to -210 region of PRL-3, which was conserved between human and mouse. Importantly, PRL-3 protein was significantly reduced in mouse STAT3-knockout liver cells compared with STAT3-wild type counterparts, and ectopic expression of STAT3 in these cells led to a pronounced increase in PRL-3 protein. We demonstrated that STAT3 functionally regulated PRL-3, and STAT3 core signature was enriched in AML with high PRL-3 expression. Targeting either STAT3 or PRL-3 reduced leukemic cell viability. Silencing PRL-3 impaired invasiveness and induced leukemic cell differentiation. In conclusion, PRL-3 was transcriptionally regulated by STAT3. The STAT3/PRL-3 regulatory loop contributes to the pathogenesis of AML, and it might represent an attractive therapeutic target for antileukemic therapy.

MicroRNA: important player in the pathobiology of multiple myeloma.

Recent studies have revealed a pivotal role played by a class of small, noncoding RNAs, microRNA (miRNA), in multiple myeloma (MM), a plasma cell (PC) malignancy causing significant morbidity and mortality. Deregulated miRNA expression in patient's PCs and plasma has been associated with tumor progression, molecular subtypes, clinical staging, prognosis, and drug response in MM. A number of important oncogenic and tumor suppressor miRNAs have been discovered to regulate important genes and pathways such as p53 and IL6-JAK-STAT signaling. miRNAs may also form complex regulatory circuitry with genetic and epigenetic machineries, the deregulation of which could lead to malignant transformation and progression. The translational potential of miRNAs in the clinic is being increasingly recognized that they could represent novel biomarkers and therapeutic targets. This review comprehensively summarizes current progress in delineating the roles of miRNAs in MM pathobiology and management.