IKZF3 polymorphisms contribute to the increased risk of acute lymphoblastic leukemia in children.

BACKGROUND: Acute lymphoblastic leukemia (ALL) is the most common cancer in children. IKZF3 (IKAROS family zinc finger 3) is a hematopoietic-specific transcription factor, and it has been validated that it is involved in leukemia. However, the role of IKZF3 single-nucleotide polymorphisms (SNPs) remains unclear. In this case-control study, the authors investigated the association of IKZF3 SNPs with ALL in children. METHODS: Six IKZF3 reference SNPs (rs9635726, rs2060941, rs907092, rs12946510, rs1453559, and rs62066988) were genotyped in 692 patients who had ALL (cases) and in 926 controls. The associations between IKZF3 polymorphisms and ALL risk were determined using odds ratios (ORs) and 95% confidence intervals (CIs). The associations of rs9635726 and rs2060941 with the risk of ALL were further estimated by using false-positive report probability (FPRP) analysis. Functional analysis in silico was performed to evaluate the probability that rs9635726 and rs2060941 might influence the regulation of IKZF3. RESULTS: The authors observed that rs9635726C>T (adjusted OR, 1.49; 95% CI, 1.06-2.11; p = .023) and rs2060941G>T (adjusted OR, 1.51; 95% CI, 1.24-1.84; p = .001) were related to and increased risk of ALL in the recessive and dominant models, respectively. Furthermore, the associations of both rs9635726 (FPRP = .177) and rs2060941 (FPRP < .001) with ALL were noteworthy in the FPRP analysis. Functional analysis indicated that rs9635726 and rs2060941 might repress the transcription of IKZF3 by disrupting its binding to MLLT1, TAF1, POLR2A, and/or RAD21. CONCLUSIONS: This study revealed that IKZF3 polymorphisms were associated with increased ALL susceptibility in children and might influence the expression of IKZF3 by disrupting its binding to MLLT1, TAF1, POLR2A, and/or RAD21. IKZF3 polymorphisms were suggested as a biomarker for childhood ALL.

HOTAIR gene polymorphisms contribute to increased neuroblastoma susceptibility in Chinese children.

BACKGROUND: Neuroblastoma is the most frequently diagnosed extracranial solid tumor in children. Previous studies have shown that single-nucleotide polymorphisms in some genes are associated with the risk of multiple cancers, including neuroblastoma. Although Hox transcript antisense intergenic RNA (HOTAIR) gene polymorphisms have been investigated in a variety of cancers, to the authors' knowledge the relationships between HOTAIR gene polymorphisms and neuroblastoma susceptibility have not been reported to date. The objective of the current study was to evaluate the correlation between HOTAIR gene polymorphisms and neuroblastoma risk in Chinese children. METHODS: The authors genotyped 6 polymorphisms (rs920778 A>G, rs12826786 C>T, rs4759314 A>G, rs7958904 G>C, rs874945 C>T, and rs1899663 C>A) of the HOTAIR gene in 2 Chinese populations including 393 neuroblastoma cases and 812 healthy controls. The strength of the associations was evaluated using odds ratios and 95% confidence intervals. Further stratification analyses were conducted to explore the association between the HOTAIR gene polymorphisms rs12826786 C>T, rs874945 C>T, and rs1899663 C>A with neuroblastoma susceptibility in terms of age, sex, clinical stage of disease, and sites of origin. RESULTS: The authors found that the rs12826786 C>T (P =.013), rs874945 C>T (P =.020), and rs1899663 C>A (P =.029) polymorphisms were significantly associated with increased neuroblastoma risk. In stratification analyses, these associations were more predominant in females and among patients with tumor in the retroperitoneal region or mediastinum. The remaining 3 polymorphisms were not found to be related to neuroblastoma susceptibility. CONCLUSIONS: The results of the current study verified that HOTAIR gene polymorphisms are associated with increased neuroblastoma risk and suggest that HOTAIR gene polymorphisms might be a potential biomarker for neuroblastoma susceptibility. Cancer 2018;124:2599-606. © 2018 American Cancer Society.

Endothelin and vascular remodelling in colitis pathogenesis--appendicitis and appendectomy limit colitis by suppressing endothelin pathways.

PURPOSE: Appendicitis and appendectomy(AA), when done at a young age, offer protection against inflammatory bowel disease (IBD) development in later life. However, IBD pathogenesis involves both immunological and vascular abnormalities. Using the first murine model of AA (developed by us), we aimed to determine the role of AA in modulating vascular remodelling mediated by endothelin activity in IBD. METHODS: Mice with two laparotomies each served as controls (sham-sham or SS). Distal colons were harvested (four AA group colons, four SS group colons), and RNA extracted from each. The RNA was subjected to microarray analysis and RT-PCR validation. Gene set enrichment analysis (GSEA) software was used to further analyze the microarray data. RESULTS: Gene expression of seven genes closely associated with endothelin activity was examined in distal colons 3 days post-AA and 28 days post-AA. While there were no gene expression changes 3 days post-AA, the genes EDN1 (0.7-fold), EDN2 (0.8-fold) and ECE2 (0.8-fold) were downregulated (*p value <0.05) 28 days post-AA. However, EDN3 (1.3-fold) was upregulated 28 days post-AA (*p value <0.05). GSEA analysis showed downregulation of 11 gene sets (stringent cut-offs-false discovery rate <5 % and p value <0.001) associated with endothelin and endothelin-converting enzyme genes by AA, in contrast to only 1 being upregulated. CONCLUSIONS: AA induces a delayed but significant suppression of genes pertaining to endothelin activity. Elucidating the pathways involved in suppression of endothelin activity and manipulation of different genes/enzymes/proteins related to endothelin activity will significantly enhance the extant repertoire of therapeutic options in IBD.

Promising antimalarial hits from phenotypic screens: a review of recently-described multi-stage actives and their modes of action.

Over the last two decades, global malaria cases caused by Plasmodium falciparum have declined due to the implementation of effective treatments and the use of insecticides. However, the COVID-19 pandemic caused major disruption in the timely delivery of medical goods and diverted public health resources, impairing malaria control. The emergence of resistance to all existing frontline antimalarials underpins an urgent need for new antimalarials with novel mechanisms of action. Furthermore, the need to reduce malaria transmission and/or prevent malaria infection has shifted the focus of antimalarial research towards the discovery of compounds that act beyond the symptomatic blood stage and also impact other parasite life cycle stages. Phenotypic screening has been responsible for the majority of new antimalarial lead compounds discovered over the past 10 years. This review describes recently reported novel antimalarial hits that target multiple parasite stages and were discovered by phenotypic screening during the COVID-19 pandemic. Their modes of action and targets in blood stage parasites are also discussed.

Versatile prostate cancer treatment with inducible caspase and interleukin-12.

To establish optimized conditions for immunity against prostate cancer, we compared the efficacy of multiple approaches in autochthonous and s.c. transgenic adenocarcinoma of the mouse prostate (TRAMP)-based models. Mice immunized with interleukin (IL)-12-containing apoptotic, but not necrotic TRAMP-C2 cell-based, vaccines were resistant to TRAMP-C2 tumor challenge and re-challenge, independently of the route of vaccination (s.c. or i.p.). Administration of gamma-irradiated TRAMP-C2 cells preinfected with adenovirus containing both B7-1 and IL-12 genes, unlike adenovirus containing B7-1 alone, considerably protected C57BL/6 mice from TRAMP-C2 tumor growth and extended the life span of TRAMP mice. Vaccines that included dendritic cells, instead of IL-12, were equally efficient. Whereas injections of ligand-inducible caspase-1- and IL-12-containing adenoviruses cured small s.c. TRAMP-C2 tumors, nanopump-regulated delivery of viruses led to elimination of much larger tumors. The antitumor immune responses involved CD4+-, CD8+-, and natural killer cells and were strengthened by increasing the number of vaccinations. Intraprostatic administration of inducible caspase-1- and IL-12-containing adenoviruses resulted in local cell death and improved survival of adenocarcinoma-bearing TRAMP mice. Thus, tumor cell apoptosis induced by caspase in situ and accompanied by IL-12 is efficient against prostate cancer in a preclinical model.

The Microbiota and Epigenetic Regulation of T Helper 17/Regulatory T Cells: In Search of a Balanced Immune System.

Immune cells not only affect tissue homeostasis at the site of inflammation but also exert systemic effects contributing to multiple chronic conditions. Recent evidence clearly supports an altered T helper 17/regulatory T cell (Th17/Treg) balance leading to the development and progression of inflammatory diseases that not only affect the gastrointestinal tract but also have whole-body manifestations, including insulin resistance. Epigenetic mechanisms are amenable to both environmental and circulating factors and contribute to determining the T cell landscape. The recently identified participation of the gut microbiota in the remodeling of the epigenome of immune cells has triggered a paradigm shift in our understanding of the etiology of various inflammatory diseases and opened new paths toward therapeutic strategies. In this review, we provide an overview of the contribution of the Th17/Treg balance in the development and progression of inflammatory bowel diseases and metabolic diseases. We discuss the involvement of epigenetic mechanisms in the regulation of T cell function in the particular context of dysbiosis. Finally, we examine the potential for nutritional interventions affecting the gut microbiota to reshape the T cell epigenome and address the inflammatory component of various diseases.

Modulation of interferon activity-associated soluble molecules by appendicitis and appendectomy limits colitis-identification of novel anti-colitic targets.

The therapeutic efficacy of interferons (IFNs) in ulcerative colitis is minimal. However, IFN activity-associated molecules have been inadequately investigated. Appendicitis and appendectomy (AA), when done while young, protect against colitis development later. Our novel murine AA model protects against colitis. This therapeutic target-identifying study enumerates IFN activity-associated molecules involved in this protection. Mice with 2 laparotomies were controls (sham-sham/SS). Distal colons were harvested (4 AA-group colons and 4 SS-group colons). Microarray-analysis/reverse transcriptase-polymerase chain reaction-validation was done from RNA from each (3-days/28-days-post-AA). Gene set enrichment analysis (GSEA) software was used to analyze distal colonic gene sets associated with 46 IFN activity-related genes. More AA-upregulated gene sets were associated with IFIT1, IFIT2, IFIT3, IRF7, IFI35, and IFI44 (False Discovery Rate-FDR <5% and P<0.001), although only IFIT1, IFIT2, IFIT3, and IFI44 showed individual gene upregulation (P<0.05). More AA-downregulated gene sets were associated with IRF1, IRF2, IRF4, IRF8, IRF9, IRF2BP1, IFRD1, IFRD2, and IFIH1 (FDR <5%/P<0.001); although only IRF2BP1 showed individual gene downregulation (P<0.05). There was significant upregulation (P<0.05) of IFNZ; and downregulation of IRF2BP2 and IFI30, despite no major associated GSEA differences. IFIT1, IFIT2, IFIT3, and IFI44, with profound AA-induced individual/GSEA upregulation, and their immunomodulatory/ antiproliferative activity, are the best molecules to investigate therapeutic potential. IRF4, IRF8, IRF2BP1, IFRD1, and IFRD2, owing to their profound AA-induced gene set downregulation, and because of their diverse lymphocytic activity, are good targets to competitively inhibit or to treat with exogenous products in knockout animals.

Autophagy suppression by appendicitis and appendectomy protects against colitis.

BACKGROUND: When done at a young age, appendicitis followed by appendectomy (AA) offers protection against ulcerative colitis development in later life. We developed the first ever murine AA model. Using this model, we showed earlier that previous AA ameliorated colitis. We aimed to determine whether autophagy genes contribute to the anti-colitis protection conferred by AA, and if so, to delineate the autophagy-linked genes involved in this. METHODS: Mice with 2 laparotomies each served as controls (sham-sham). Distal colons were harvested (4 AA-group colons, 4 sham-sham group colons), and RNA extracted from each. The RNA was taken through microarray analysis or reverse transcription-polymerase chain reaction validation. Gene set enrichment analysis software was used to analyze the microarray data. RESULTS: Out of 28 key autophagy-related genes investigated (VPS15, VPS34, FIP200, ATG03, ATG04A, ATG04B, ATG05, ATG07, ATG10, ATG12, ATG13b, ATG14, ATG16L1, BECN1, GABARAPL1, IRGM1, IRGM2, LAMP2, LC3A, LC3B, RAB7A, UVRAG, NOD2, XBP1, LRRK2, ULK1, ULK2, PTPN2), 7 have genetic associations with inflammatory bowel diseases (ATG16L1, IRGM1, NOD2, XBP1, LRRK2, ULK1, PTPN2). There was slight upregulation of IRGM1, FIP200, and ATG04A (P < 0.05), but no variations with the other 25 genes. In contrast, gene set enrichment analysis revealed that AA downregulated 74 gene sets (associated with 28 autophagy genes) while upregulating only 5 (false discovery rate <5%; P < 0.001) gene sets. Additionally, 22 gene sets associated with the 7 autophagy + inflammatory bowel disease-associated genes were downregulated by AA, whereas only 3 were upregulated. The genes with maximum AA-induced gene set suppression were VPS15, LAMP2, LC3A, XBP1, and ULK1. CONCLUSIONS: AA induces profound autophagy suppression in the distal colon. The AA-induced upregulation of individual genes (IRGM1, FIP200, ATG04A) could be a reflection of complex compensatory changes or the initial abnormality that led to the pronounced autophagy suppression. Autophagy suppression by AA may induce lesser antigen processing, leading to lesser cross-reactive immunity between microbes and self-antigens, and subsequent amelioration of colitis.

Sorafenib downregulates ERK/Akt and STAT3 survival pathways and induces apoptosis in a human neuroblastoma cell line.

Neuroblastoma is a common solid tumor in children and its tumorigenicity is enhanced by the expression of survival pathways such as Akt and signal transducer and activator of transcription 3 (STAT3). Sorafenib is a multikinase inhibitor that also inhibits STAT3 signaling and induces apoptosis. In this study, we will examine the efficacy of sorafenib on a human neuroblastoma cell line (SK-N-AS) and also investigate its possible mechanisms. After cells reached 50-60% confluence, they were treated with various concentrations of sorafenib (0, 0.1, 1, 5, 10 and 20 microM) for different periods of time. The cell viability and apoptosis were determined by MTS colorimetric assay and TUNEL, respectively. Phosphorylation of Akt1/2/3 (p-Akt1/2/3), extracellular signal-regulated kinase 1/2 (p-ERK1/2), STAT3 (p-STAT3), and AMP-activated protein kinase alpha subunit (p-AMPKalpha) were determined with Western blot. The results indicate that as early as 2 hours post-treatment, cell viability was significantly decreased at 10 microM concentration. In 24 hours or longer treatment groups, sorafenib at 5 microM and above significantly decreased cell viability. TUNEL assay showed a significant increased of apoptosis in 5 and 20 microM treatment groups 24 hours after treatment. Western blots showed a decrease of p-ERK1/2, p-Akt1/2/3, p-STAT3, and p-AMPKalpha expression levels in various sorafenib treatment groups. Our results indicate that sorafenib significantly decreased cell viability and increased apoptosis in human neuroblastoma cell line in association with down-regulation of p-ERK1/2, p-Akt, p-STAT3 survival pathways. These data suggested potential clinical application of sorafenib in the treatment of neuroblastoma.

Molecular mechanisms of vacuum therapy in penile rehabilitation: a novel animal study.

BACKGROUND: Penile rehabilitation (PR) is widely applied after radical prostatectomy. Vacuum erectile device (VED) therapy is the one of three PR methods used in the clinical setting that improve erectile function (EF) and is the only PR method which may preserve penile length. However, its unknown mechanism hampered doctors' recommendations and patients' compliance. OBJECTIVES: To assess the effects of VED therapy on erectile dysfunction (ED) in a rat model of bilateral cavernous nerve crush (BCNC) and to investigate the molecular mechanism of VED in postprostatectomy ED. DESIGN, SETTING, AND PARTICIPANTS: This was an experimental study using Sprague-Dawley rats in three groups: sham, BCNC, and BCNC plus VED. INTERVENTION: Intervention included BCNC, electrical stimulation of the cavernous nerve (CNS), and VED therapy. MEASUREMENTS: At the end of a 4-wk period, CNS was used to assess EF by maximum intracavernosal pressure (ICP)/mean arterial pressure (MAP) ratio and duration (area under the curve [AUC]). For the structural analyses, whole rat penis was harvested. Terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling assay was used for the assessment of apoptotic indices (AI). Immunohistochemistry was performed for endothelial nitric oxide synthase (eNOS), α-smooth muscle actin (ASMA), transforming growth factor beta 1 (TGF-ß1), and hypoxia inducible factor-1α (HIF-1α). Staining for Masson's trichrome was utilized to calculate the smooth muscle/collagen ratios. RESULTS AND LIMITATIONS: EF was improved with VED therapy measured by ICP/MAP ratios and AUC. VED therapy reduced HIF-1α expression and AI significantly compared with control. Animals exposed to VED therapy had decreased TGF-ß1 expression, increased smooth muscle/collagen ratios, and preserved ASMA and eNOS expression. CONCLUSIONS: To our knowledge, this is the first scientific study to suggest that VED therapy in the BCNC rat model preserves EF through antihypoxic, antiapoptotic, and antifibrotic mechanisms.

Poloxamer 188 inhibition of ischemia/reperfusion injury: evidence for a novel anti-adhesive mechanism.

Poloxamer 188 (P188) is a nonionic block copolymer surfactant that has rheologic, anti-thrombotic, anti-inflammatory, and cytoprotective activities. Several mechanisms have previously been proposed, but none explain all of the observed effects. In this study, superior mesenteric artery occlusion (SMAO) was employed as a clinically relevant model of ischemia/reperfusion. The superior mesenteric artery of rats was clamped for one hr and and followed by reperfusion with P188 or saline for five hr, after which tissues were harvested for expression microarray, histologic, enzymatic, and western blot analyses. The results demonstrated that P188 significantly inhibits the entire spectrum of inflammatory, coagulation, and apoptotic responses produced by SMAO. This supports the existence of a novel mechanism that recognizes two types of adhesive interactions. The first, specific receptor-ligand adhesion, governs interactions between cells and molecules and is unaffected by P188. P188 affects only the second type, hydrophobic adhesion, which is responsible for the integrity of membranes and conformation of proteins. Hydrophobic interactions are non-specific because they derive from repulsion of water rather than from affinity of molecules for one another. Ischemic membranes develop defects that expose underlying hydrophobic structures and trigger multiple deleterious responses. Fat emboli and hydrophobic proteins such as fibrin produced by the injury further compromise the microcirculation. The unique structure of P188 facilitates its rapid, but gentle, binding to any exposed hydrophobic domain, restoring normal non-adhesive surfaces and thereby preventing activation of the entire spectrum of deleterious reactions.