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ABSTRACT Background xCT, also known as SLC7A11 (solute carrier Family 7 Member 11), is a cystine/glutamate antiporter protein that mediates regulated cell death and antioxidant defense. The aim of this study was to investigate the effect of xCT on the outcome of patients diagnosed with new head and neck squamous cell carcinoma (HNSCC). Methods This retrospective cohort study utilized a population‐based dataset, comprising all patients (n = 1033) diagnosed with new HNSCC during 2005–2015 in a population of 697,000 people. All patients (n = 585) with a tumor tissue sample available for immunohistochemical (IHC) staining were included. The follow‐up rates were 97% and 81% at 3 and 5 years, respectively. Also, the specificity of the anti‐xCT antibody was validated. Results The expression level and prognostic significance of xCT were strongly dependent on tumor location. In oropharyngeal squamous cell carcinoma (OPSCC) patients, xCT expression was a significant prognostic factor for 5‐year overall survival (OAS) (HR: 2.71; 95% CI 1.67–4.39; p

Evaluating an impairment in an individual's capacity to maintain, achieve, or restore balance suggests a deficiency in postural control. For effective identification of individuals at risk for falls, balance assessment should exhibit reliability, validity, and ease of use. This systematic review concentrated on dynamic and functional balance assessment methods and their validity in healthy adults aged 19-54. The objective was to clarify the tools that health professionals can utilize to assess balance in this healthy population. # Methods A systematic literature search conducted in August 2019 yielded nine articles meeting predefined selection criteria. Inclusion criteria required studies featuring healthy adult participants aged 19-54, published in English, and focusing on dynamic and functional balance testing. Exclusion criteria excluded studies involving participants with chronic diseases or musculoskeletal disorders, systematic reviews, professional athletes, and those lacking specific participant age information. The quality of the studies was evaluated using a modified PEDro scale. # Results This review analyzed ten distinct postural balance tests. The Star Excursion Balance Test and Y-Balance Test exhibited moderate to high reliability, establishing them as dependable measures of dynamic balance. The Nintendo Wii Balance Board, Clever Balance Board, and Posturomed device also displayed excellent reliability for assessing dynamic postural balance. Comparing one-arm and two-arm functional reach tests, the one-arm reach test emerged as a more suitable option for evaluating dynamic balance among young adults. Moreover, an investigation comparing three dynamic balance tests (one-leg jump landing, Posturomed device, and stimulated forward fall) revealed a low correlation among these tests, indicating a measurement of different balance constructs. # Discussion In conclusion, the Y-Balance Test stands out as the most practical dynamic balance assessment for clinical use, characterized by a standardized protocol, good repeatability, affordability, and ease of application. The Nintendo Wii Balance Board also presents itself as a cost-effective and reliable tool for dynamic balance evaluation in clinical settings. It is crucial to recognize that these tests appraise discrete postural skills, preventing direct comparisons between test outcomes. This review equips healthcare professionals with valuable insights into optimal balance assessment methods for the healthy, 19 to 54 aged population. # Levels of evidence Level 3

BackgroundTreatment resistance and relapse are common problems in head and neck squamous cell carcinoma (HNSCC). Except for p16, no clinically accepted prognostic biomarkers are available for HNSCC. New biomarkers predictive of recurrence and survival are crucial for optimal treatment planning and patient outcome. High translocator protein (TSPO) levels have been associated with poor survival in cancer, but the role of TSPO has not been extensively evaluated in HNSCC.Materials and methodsTSPO expression was determined in a large population-based tissue microarray cohort including 611 patients with HNSCC and evaluated for survival in several clinicopathological subgroups. A TCGA HNSCC cohort was used to further analyze the role of TSPO in HNSCC.ResultsTSPO expression was downregulated in more aggressive tumors. Low TSPO expression associated with worse 5-year survival and was an independent prognostic factor for disease-specific survival. Subgroup analyses showed that low TSPO expression associated with worse survival particularly in p16-positive oropharyngeal cancer. In silico analyses supported the prognostic role of TSPO. Cellular respiration had the highest significance in pathway analyses for genes expressed positively with TSPO.ConclusionDecreased TSPO expression associates with poor prognosis in HNSCC. TSPO is a prognostic biomarker in HNSCC to potentially guide treatment stratification especially in p16-positive oropharyngeal cancer.

Abstract Background A low tissue oxygen level,

Abstract CD73 is a cell surface ecto-5′-nucleotidase, which converts extracellular adenosine monophosphate to adenosine. High tumor CD73 expression is associated with poor outcome among triple-negative breast cancer (TNBC) patients. Here we investigated the mechanisms by which CD73 might contribute to TNBC progression. This was done by inhibiting CD73 with adenosine 5′-(α, β-methylene) diphosphate (APCP) in MDA-MB-231 or 4T1 TNBC cells or through shRNA-silencing (sh-CD73). Effects of such inhibition on cell behavior was then studied in normoxia and hypoxia in vitro and in an orthotopic mouse model in vivo. CD73 inhibition, through shRNA or APCP significantly decreased cellular viability and migration in normoxia. Inhibition of CD73 also resulted in suppression of hypoxia-induced increase in viability and prevented cell protrusion elongation in both normoxia and hypoxia in cancer cells. Sh-CD73 4T1 cells formed significantly smaller and less invasive 3D organoids in vitro, and significantly smaller orthotopic tumors and less lung metastases than control shRNA cells in vivo. CD73 suppression increased E-cadherin and decreased vimentin expression in vitro and in vivo, proposing maintenance of a more epithelial phenotype. In conclusion, our results suggest that CD73 may promote early steps of tumor progression, possibly through facilitating epithelial–mesenchymal transition.

Wild lingonberries are a traditional source of food in the Nordic countries and an important contributor to economic activity of non-wood forest products in the region. Lingonberries are a rich source of bioactive compounds and can be a valuable contributor to a healthy diet. However, there are few studies available on how the bioactive compounds in lingonberries develop as they ripen. In this investigation, we examined the content of 27 phenolic compounds, three sugars, four organic acids, and 71 volatile organic compounds at five ripening stages. The study showed that, while the highest content of phenolic compounds was found early in the development, the organoleptic quality of the fruits improved as they ripened. From the first to the last stage of development, anthocyanins went from being nearly absent to 100 mg/100 g fw, and there was an increased content of sugars from 2.7 to 7.2 g/100 g fw, whereas the content of organic acids decreased from 4.9 to 2.7 g/100 g fw, and there were several changes in the profile of volatiles. The contents of flavonols, cinnamic acid derivatives, flavan-3-ols, and the total concentration of phenolic compounds were significantly lower in the fully ripe berries compared to berries in the early green stage. In addition to the changes occurring due to ripening, there was observed variation in the profile of both phenolic compounds and volatiles, depending on the growth location of the berries. The present data are useful for the assessment of harvest time to obtain the desired quality of lingonberries.

Background: Community-based health insurance (CBHI) has emerged as a more efficient and equitable approach to healthcare financing. It was designed to ensure that sufficient resources are made available for members to access effective healthcare. This study assessed the willingness to pay (WTP) for CBHI among artisans in a town in Ekiti State, South West Nigeria. Methods: This was a cross-sectional survey conducted among 416 artisans in a town in Ekiti State. A semi-structured interviewer-administered questionnaire was designed and used for data collection on sociodemographic data and WTP for CBHIS. Data entry and analysis was done using IBM SPSS software version 25.0. Results: The mean age and standard deviation of the respondents was 29.7±10.9 years with male to female ratio of 1:1.4. Most of the respondents were willing to pay (86.3%) and willing to enroll other household members (73.6%) in the CBHI. A large percentage (44.3%) of those willing to pay were ready to pay between ₦1,000-₦5,000 (US$2.63–US$13.16) per year while 39.6% preferred frequency of payment to be annually. Positive predictors of WTP for CBHI were age groups ≥50 years and 40-49 years than

Background: Diabetes Mellitus is a non-communicable disease and a leading cause of morbidity and mortality in the world. The estimated prevalence of diabetes in Africa is 1% in rural areas and ranges from 5% to 7% in urban sub-Saharan Africa. 3 Nigeria is the most populous country in African and she contributes about one sixth of the Africa’s diabetic population. This study is aimed at determining the prevalence of diabetes mellitus and accessing its risk factors among public secondary school teachers in Ekiti State. Methodology: This survey is a cross-sectional study carried out among 357 public school teachers in Ekiti State using a multi stage sampling technique. Data was gathered using a semi structured questionnaire and Accu Check Glucometer machine for Fasting Blood Sugar. Analysis was done using SPSS version 23 and level of significance was taken as P = 0.05. Result: Mean age was 42.9±9.2 years. Prevalence of Diabetes Mellitus was 5.6%. Family history (15%), Alcohol intake (14.0%) and smoking (11.2%) were major risk factors in the respondents. Poor vision is the main complication being experienced by the respondents (40.3%). Predictors of Diabetes Mellitus as found by this study include increasing age (starting from above 40years), cigarette smoking, alcohol intake and positive family history. Conclusion: Prevalence of Diabetes Mellitus is high compared to what it was before. Alcohol intake, smoking and family history were major risk factors while poor vision is the commonest complication. It is recommended that all efforts must be made to put in mechanism that will halt this undesired progression through control of risk factors.

Immune checkpoint inhibitors (ICI) have provided significant improvement in clinical outcomes for some patients with solid tumors. However, for patients with head and neck cancer, the response rate to ICI monotherapy remains low, leading to the exploration of combinatorial treatment strategies. In this preclinical study, we use an oncolytic adenovirus (Ad5/3) encoding hTNFα and hIL-2 and non-replicate adenoviruses (Ad5) encoding mTNFα and mIL-2 with ICI to achieve superior tumor growth control and improved survival outcomes. The in vitro effect of Ad5/3-E2F-D24-hTNFa-IRES-hIL-2 was characterized through analyses of virus replication, transgene expression and lytic activity using head and neck cancer patient derived cell lines. Mouse models of ICI naïve and refractory oral cavity squamous cell carcinoma were established to evaluate the local and systemic anti-tumor immune response upon ICI treatment with or without the non-replicative adenovirus encoding mTNFα and mIL-2. We delineated the mechanism of action by measuring the metabolic activity and effector function of CD3+ tumor infiltrating lymphocytes (TIL) and transcriptomic profile of the CD45+ tumor immune compartment. Ad5/3-E2F-D24-hTNFa-IRES-hIL-2 demonstrated robust replicative capability in vitro across all head and neck cell lines screened through potent lytic activity, E1a and transgene expression. In vivo, in both ICI naïve and refractory models, we observed improvement to tumor growth control and long-term survival when combining anti-PD-1 or anti-PD-L1 with the non-replicative adenovirus encoding mTNFα and mIL-2 compared to monotherapies. This observation was verified by striking CD3+ TIL derived mGranzyme b and interferon gamma production complemented by increased T cell bioenergetics. Notably, interrogation of the tumor immune transcriptome revealed the upregulation of a gene signature distinctive of tertiary lymphoid structure formation upon treatment of murine anti-PD-L1 refractory tumors with non-replicative adenovirus encoding mTNFα and mIL-2. In addition, we detected an increase in anti-tumor antibody production and expansion of the memory T cell compartment in the secondary lymphoid organs. In summary, a non-replicative adenovirus encoding mTNFα and mIL-2 potentiates ICI therapy, demonstrated by improved tumor growth control and survival in head and neck tumor-bearing mice. Moreover, the data reveals a potential approach for inducing tertiary lymphoid structure formation. Altogether our results support the clinical potential of combining this adenovirotherapy with anti-PD-1 or anti-PD-L1.

Abstract Background Sheep is important in the socio-economic lives of people around the world. It is estimated that more than half of our once common livestock breeds are now endangered. Since genetic characterization of Nigerian sheep is still lacking, we analyzed ten morphological traits on 402 animals and 15 microsatellite DNA markers in 384 animals of the 4 Nigerian sheep breeds to better understand genetic diversity for breeding management and germplasm conservation. Results Morphological traits of Uda and Balami were significantly (P FST, FIT and FIS statistics across all loci were 0.088, 0.394 and 0.336 respectively. Yankasa and Balami are the most closely related breeds (DA = 0.184) while WAD and Balami are the farthest apart breeds (DA = 0.665), which is coincident with distance based on morphological analysis and population structure assessed by STRUCTURE. Conclusions These results suggest that within-breed genetic variation in Nigerian sheep is higher than between-breeds and may be a valuable tool for genetic improvement and conservation. The higher genetic variability in Yankasa suggests the presence of unique ancestral alleles reflecting the presence of certain functional genes which may result in better adaptability in more agro-ecological zones of Nigeria. These genetic characteristics are potentially useful in planning improvement and conservation strategies in Nigerian indigenous sheep.

Background: Currently, <10% of Nigerians are insured by the National Health Insurance Scheme (NHIS) and this among other things has led to the signing of the National Health Insurance Authority (NHIA) Act in May 2022, which aims at ensuring the effective implementation of a national health insurance policy and attainment of Universal Health Coverage (UHC) in Nigeria., Objective: To highlight the new features of the NHIA Act and its policy implications for the Nigerian health system., Methods: A modified Delphi method was used for extracting the differences in the two Acts. A total of three rounds of reviews were carried out among 5 reviewers within three weeks. Differences were tabulated and also presented in prose., Findings: The NHIA Act makes health insurance mandatory for all residents of Nigeria with the introduction of the vulnerable group fund and implementation of the Basic Health Care Provision Fund through the established State Health Insurance Schemes. Unlike the NHIS which is a Scheme, the NHIA is an Authority and has an expanded function to regulate, promote, manage and integrate all health insurance schemes and practices in Nigeria. Also, funds management has been transferred from Health Maintenance Organizations to the State Health Insurance Schemes and the Health Maintenance Organizations are now excluded from the Governing Council., Conclusion: Certainly, the journey towards UHC could be safer and more equitable with health insurance now mandatory for all Nigerians and the introduction of vulnerable group funds in the new Act. This will eliminate the catastrophic expenses of poor Nigerians if the Act is correctly implemented., Competing Interests: The Authors declare that no competing interest exists, (Copyright © 2023 by West African Journal of Medicine.)

Background: Every minute, six indigenous Africans develop new strokes. Patient-level and system-level contributors to early stroke fatality in this region are yet to be delineated. We aimed to identify and quantify the contributions of patient-level and system-level determinants of inpatient stroke fatality across 16 hospitals in Ghana and Nigeria., Methods: The Stroke Investigative Research and Educational Network (SIREN) is a multicentre study involving 16 sites in Ghana and Nigeria. Cases include adults (aged ≥18 years) with clinical and radiological evidence of an acute stroke. Data on stroke services and resources available at each study site were collected and analysed as system-level factors. A host of demographic and clinical variables of cases were analysed as patient-level factors. A mixed effect log-binomial model including both patient-level and system-level covariates was fitted. Results are presented as adjusted risk ratios (aRRs) with respective 95% CIs., Findings: Overall, 814 (21·8%) of the 3739 patients admitted with stroke died as inpatients: 476 (18·1%) of 2635 with ischaemic stroke and 338 (30·6%) of 1104 with intracerebral haemorrhage. The variability in the odds of stroke fatality that could be attributed to the system-level factors across study sites assessed using model intracluster correlation coefficient was substantial at 7·3% (above a 5% threshold). Stroke units were available at only five of 16 centres. The aRRs of six patient-level factors associated with stroke fatality were: low vegetable consumption, 1·19 (95% CI 1·07-1·33); systolic blood pressure, 1·02 (1·01-1·04) for each 10 mm Hg rise; stroke lesion volume more than 30 cm 3 , 1·48 (1·22-1·79); National Institutes of Health Stroke Scale (NIHSS) score, 1·20 (1·13-1·26) for each 5-unit rise; elevated intracranial pressure, 1·75 (1·31-2·33); and aspiration pneumonia, 1·79 (1·16-2·77)., Interpretation: Studies are needed to assess the efficacy of interventions targeting patient-level factors such as aspiration pneumonia in reducing acute stroke fatality in this region. Policy directives to improve stroke unit access are warranted., Funding: US National Institutes of Health., Translations: For the Twi, Yoruba and Hausa translations of the abstract see Supplementary Materials section., Competing Interests: Declaration of interests We declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY-NC-ND 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: We evaluated the characteristics of carotid and vertebral atherosclerosis in indigenous West Africans with stroke., Methodology: Of the 3778stroke patients recruited between 01/2014 and 08/2017, 1070 (28.3%) received carotid and vertebral artery evaluation with B-mode Ultrasound. Carotid and vertebral intima-media thickness (IMT) using multiple site technique were measured bilaterally and plaque frequency was determined. Descriptive and comparative analyses between stroke types and vessels were carried out., Results: There were 809 (75.6%) patients with ischemic stroke. The prevalence of intima-media thickening in the study population was 84.0% (898/1070) [95% CI: 81.7-86.1], being higher in the ischemic stroke (688/809, 85.0%) [95% CI: 82.4-87.3] than in the hemorrhagic stroke group (211/261, 80.8%) [95% CI: 75.6-85.2]. Overall prevalence of plaques which was 26.1% [95% CI: 23.5-28.8], was found also to be higher in ischemic than hemorrhagic stroke (29.8%[95% CI: 26.7-33.0] vs. 14.6% [95% CI: 10.8-19.4], p < 0.05). The mean IMT (carotids: 2.01+1.33 mm; vertebrals: 0.96+0.54mm, p<0.001) and prevalence of plaques (carotids: 8.8%; vertebrals: 1.7%,p<0.001) were higher in carotid than vertebral arteries. Age, hypertension, level of formal education, history of smoking, average monthly income, and family histories of hypertension and stroke were associated with intima-media thickening in the carotids (all p< 0.05) in the ischemic stroke patients while family history of hypertension, diabetes mellitus, and level of formal education were independently associated with intima-media thickening in the carotids (all p< 0.05) in the hemorrhagic stroke patients. No CVRF showed an independent association with the presence of plaque in the carotid and vertebral arteries both stroke types., Conclusions: One off our stroke patients in our cohort had atherosclerotic plaques, with ischemic patients being twice as likely to have this burden compared to hemorrhagic patients, and carotid atherosclerosis being five times as frequent as vertebral atherosclerosis., (Copyright © 2022 Nigerian Medical Association.)

The proportionate use of energy represents economic activity as well as environmental degradation. This study intends to examine the volatility spillover of environmental fluctuations (energy prices) to the stock markets of south Asian countries (i.e., Bangladesh, India, and Pakistan). In this regard, the data have been gathered from the Thomson Reuters DataStream from 2013 to 2021. This study has applied the Granger causality test and ARCH-GARCH (1, 1). It concludes that the bidirectional causality exists between the environmental prices (i.e., energy market) and Bangladesh, Pakistan, and India stock markets (BSE-100, DSE-30, and KSE-100, respectively). The empirical findings of this study show that there are volatility spillovers from the energy to the stock markets of Pakistan and India. On the other hand, no volatility spillover is observed from the energy to the stock market of Bangladesh. Moreover, the study implies that investors should invest in these stock markets to reduce the risk involved with diversification., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2022 R. M. Ammar Zahid et al.)

Background and Purpose: To identify the qualitative and quantitative contributions of conventional risk factors for occurrence of ischemic stroke and its key pathophysiologic subtypes among West Africans., Methods: The SIREN (Stroke Investigative Research and Educational Network) is a multicenter, case-control study involving 15 sites in Ghana and Nigeria. Cases include adults aged ≥18 years with ischemic stroke who were etiologically subtyped using the A-S-C-O-D classification into atherosclerosis, small-vessel occlusion, cardiac pathology, other causes, and dissection. Controls were age- and gender-matched stroke-free adults. Detailed evaluations for vascular, lifestyle, and psychosocial factors were performed. We used conditional logistic regression to estimate adjusted odds ratios with 95% CI., Results: There were 2431 ischemic stroke case and stroke-free control pairs with respective mean ages of 62.2±14.0 versus 60.9±13.7 years. There were 1024 (42.1%) small vessel occlusions, 427 (17.6%) large-artery atherosclerosis, 258 (10.6%) cardio-embolic, 3 (0.1%) carotid dissections, and 719 (29.6%) undetermined/other causes. The adjusted odds ratio (95% CI) for the 8 dominant risk factors for ischemic stroke were hypertension, 10.34 (6.91-15.45); dyslipidemia, 5.16 (3.78-7.03); diabetes, 3.44 (2.60-4.56); low green vegetable consumption, 1.89 (1.45-2.46); red meat consumption, 1.89 (1.45-2.46); cardiac disease, 1.88 (1.22-2.90); monthly income $100 or more, 1.72 (1.24-2.39); and psychosocial stress, 1.62 (1.18-2.21). Hypertension, dyslipidemia, diabetes were confluent factors shared by small-vessel, large-vessel and cardio-embolic subtypes. Stroke cases and stroke-free controls had a mean of 5.3±1.5 versus 3.2±1.0 adverse cardio-metabolic risk factors respectively ( P <0.0001)., Conclusions: Traditional vascular risk factors demonstrate important differential effect sizes with pathophysiologic, clinical and preventative implications on the occurrence of ischemic stroke among indigenous West Africans.

Background: It is crucial to assess genomic literacy related to stroke among Africans in preparation for the ethical, legal and societal implications of the genetic revolution which has begun in Africa., Objective: To assess the knowledge, attitudes and practices (KAP) of West Africans about stroke genetic studies., Methods: A comparative cross-sectional study was conducted among stroke patients and stroke-free controls recruited across 15 sites in Ghana and Nigeria. Participants' knowledge of heritability of stroke, willingness to undergo genetic testing and perception of the potential benefits of stroke genetic research were assessed using interviewer-administered questionnaire. Descriptive, frequency distribution and multiple regression analyses were performed., Results: Only 49% of 2029 stroke patients and 57% of 2603 stroke-free individuals knew that stroke was a heritable disorder. Among those who knew, 90% were willing to undergo genetic testing. Knowledge of stroke heritability was associated with having at least post-secondary education (OR 1.51, 1.25-1.81) and a family history of stroke (OR 1.20, 1.03-1.39) while Islamic religion (OR=0.82, CI: 0.72-0.94), being currently unmarried (OR = 0.81, CI: 0.70-0.92), and alcohol use (OR = 0.78, CI: 0.67-0.91) were associated with lower odds of awareness of stroke as a heritable disorder. Willingness to undergo genetic testing for stroke was associated with having a family history of stroke (OR 1.34, 1.03-1.74) but inversely associated with a medical history of high blood pressure (OR = 0.79, 0.65-0.96)., Conclusion: To further improve knowledge of stroke heritability and willingness to embrace genetic testing for stroke, individuals with less formal education, history of high blood pressure and no family history of stroke require targeted interventions.

Introduction: Stroke is a prominent cause of death, disability, and dementia in sub-Saharan Africa (SSA). The Stroke Investigative Research and Education Network works collaboratively with stroke survivors and individuals serving as community controls to comprehensively characterize the genomic, sociocultural, economic and behavioral risk factors for stroke in SSA., Purpose: In this paper, we aim to: i) explore the attitudes, beliefs, and practices related to stroke in Ghana and Nigeria using the process of qualitative description; and ii) propose actions for future research and community-based participation and education., Methods: Stroke survivors, their caregivers, health care professionals, and community representatives and faith-based leaders participated in one of twenty-six focus groups, which qualitatively explored community beliefs, attitudes and practices related to stroke in Ghana and Nigeria. Arthur Kleinman's Explanatory Model of Illness and the Social Ecological Model guided the questions and/or thematic analysis of the qualitative data. We hereby describe our focus group methods and analyses of qualitative data, as well as the findings and suggestions for improving stroke outcomes., Results and Discussion: The major findings illustrate the fears, causes, chief problems, treatment, and recommendations related to stroke through the views of the participants, as well as recommendations for working effectively with the SIREN communities. Findings are compared to SIREN quantitative data and other qualitative studies in Africa. As far as we are aware, this is the first paper to qualitatively explore and contrast community beliefs, attitudes, and practices among stroke survivors and their caregivers, community and faith-based leaders, and health professionals in multiple communities within Nigeria and Ghana., Competing Interests: The authors have declared that no competing interests exist.

Purpose: The hypoxia-inducible factor-1 alpha (HIF-alpha) is a key regulator of tumor angiogenesis. Mammalian target of rapamycin (mTOR) and histone deacetylase (HDAC) inhibitors suppress tumor-induced angiogenesis by reducing tumor HIF-1 alpha protein expression. Thus, we hypothesized that combination treatment of rapamycin and the HDAC inhibitor LBH589 has greater antiangiogenic and antitumor activity compared with single agents., Experimental Design: To evaluate the effect of LBH589 and rapamycin on HIF-1 alpha in human prostate PC3, renal C2 carcinoma cell lines, and endothelial cells (human umbilical vein endothelial cells), we did Western blot analysis. To determine the antitumor activity of LBH589 and rapamycin, cell proliferation assays and xenograft experiments were conducted., Results: Western blotting showed that combination treatment of human umbilical vein endothelial cells, C2 and PC3, significantly reduced HIF-1 alpha protein expression compared with single agents. Treatment with rapamycin resulted in inhibition of the downstream signals of the mTOR pathway and increased phosphorylation of Akt in C2 cells, whereas the constitutively activated Akt in PC3 cells was not modulated. LBH589 decreased both constitutively expressed and rapamycin-induced phosphorylated Akt levels in PC3 and C2 cell lines. In clonogenic assays, the combination treatment had a greater inhibitory effect in PC3 cells (93 +/- 1.4%) compared with single agents (66 +/- 9% rapamycin and 43 +/- 4% LBH589). Combination of rapamycin and LBH589 significantly inhibited PC3 and C2 in vivo tumor growth and angiogenesis as measured by tumor weight and microvessel density., Conclusions: Combination treatment of mTOR and HDAC inhibitors represents a rational therapeutic strategy targeting HIF-1 alpha that warrants clinical testing.

Purpose: Angiogenesis inhibitors have shown clinical benefit in patients with advanced renal cell cancer, but further therapeutic improvement is needed. Vascular endothelial growth factor (VEGF) Trap is a newly developed VEGF-blocking agent with stronger affinity and broader activity than the anti-VEGF antibody bevacizumab. In this study, we tested the activity of VEGF Trap in an orthotopic murine model of renal cancer with spontaneous lung metastases., Experimental Design: Murine syngeneic renal cell carcinoma cells (RENCA) transfected with a luciferase-expressing vector were injected into the renal capsule of BALB/c mice. I.p. treatment with VEGF Trap or control protein (10 or 25 mg/kg twice weekly) was started shortly after tumor injection to prevent tumor development (prevention model) or after established tumors were formed to inhibit tumor growth and metastasis formation (intervention model)., Results: In the prevention model, VEGF Trap inhibited tumor growth by 87 +/- 14% compared with control (P=0.007) and significantly prolonged survival. In the intervention model, VEGF Trap inhibited tumor growth by 74 +/- 9% (P<0.001) and the formation of lung metastases was inhibited by 98% (P<0.004). Microvascular density was reduced by 66% due to VEGF Trap treatment (P<0.001). In addition, VEGF Trap prevented fibrinogen leakage into the tumor microenvironment representative for reduced vascular leaking as shown by immunohistochemical staining., Conclusions: VEGF Trap is a potent inhibitor of RENCA tumor growth and metastasis formation and blocks the biological function of VEGF in vivo. These results support further clinical development of VEGF Trap for renal cell cancer and other cancer types.

Purpose: Angiogenesis is required for tumor progression and represents a rational target for therapeutic intervention. Histone deacetylase (HDAC) inhibitors have been shown to have activity against various tumor cell types by inhibiting proliferation and inducing apoptosis both in vitro and in vivo. HDAC inhibitors have also been reported to inhibit angiogenesis. The goal of this study was to characterize the antiangiogenic and antitumor activity of a recently developed HDAC inhibitor, the hydroxamic derivative LBH589., Materials and Methods: To evaluate the antiangiogenesis activity of LBH589, we did cell cycle analysis, cell proliferation, tube formation, invasion assays in vitro, and Matrigel plug assay in vivo. To determine the antitumor activity of LBH589, we established human prostate carcinoma cell PC-3 xenografts in vivo. To evaluate the effect of LBH589 on endothelial signaling pathways, gene expression, and protein acetylation, we did Western blots and reverse transcription-PCR in human umbilical vein endothelial cells (HUVEC). Immunohistochemical analysis was done to evaluate new blood vessel formation in vivo., Results: LBH589 induced acetylation of histone H3 and alpha-tubulin protein in HUVECs. Histone and nonhistone protein acetylation correlated with induction of G(2)-M cell cycle arrest, inhibition of HUVEC proliferation, and viability. Noncytotoxic concentrations of LBH589 inhibited endothelial tube formation, Matrigel invasion, AKT, extracellular signal-regulated kinase 1/2 phosphorylation, and chemokine receptor CXCR4 expression. In vivo dosing of mice with LBH589 (10 mg/kg/d) reduced angiogenesis and PC-3 tumor growth., Conclusion: This study provides evidence that LBH589 induces a wide range of effects on endothelial cells that lead to inhibition of tumor angiogenesis. These results support the role of HDAC inhibitors as a therapeutic strategy to target both the tumor and endothelial compartment and warrant the clinical development of these agents in combination with angiogenesis inhibitors.