Anti-Idiotypic Nanobodies Mimicking an Epitope of the Needle Protein of the Chlamydial Type III Secretion System for Targeted Immune Stimulation.

The development of new approaches and drugs for effective control of the chronic and complicated forms of urogenital chlamydia caused by Chlamydia trachomatis, which is suspected to be one of the main causes of infertility in both women and men, is an urgent task. We used the technology of single-domain antibody (nanobody) generation both for the production of targeting anti-chlamydia molecules and for the subsequent acquisition of anti-idiotypic nanobodies (ai-Nbs) mimicking the structure of a given epitope of the pathogen (the epitope of the Chlamydial Type III Secretion System Needle Protein). In a mouse model, we have shown that the obtained ai-Nbs are able to induce a narrowly specific humoral immune response in the host, leading to the generation of intrinsic anti-Chlamydia antibodies, potentially therapeutic, specifically recognizing a given antigenic epitope of Chlamydia. The immune sera derived from mice immunized with ai-Nbs are able to suppress chlamydial infection in vitro. We hypothesize that the proposed method of the creation and use of ai-Nbs, which mimic and present to the host immune system exactly the desired region of the antigen, create a fundamentally new universal approach to generating molecular structures as a part of specific vaccine for the targeted induction of immune response, especially useful in cases where it is difficult to prepare an antigen preserving the desired epitope in its native conformation.

A High-Homology Region Provides the Possibility of Detecting ß-Barrel Pore-Forming Toxins from Various Bacterial Species.

The pathogenicity of many bacteria, including Bacillus cereus and Staphylococcus aureus, depends on pore-forming toxins (PFTs), which cause the lysis of host cells by forming pores in the membranes of eukaryotic cells. Bioinformatic analysis revealed a region homologous to the Lys171-Gly250 sequence in hemolysin II (HlyII) from B. cereus in over 600 PFTs, which we designated as a "homologous peptide". Three ß-barrel PFTs were used for a detailed comparative analysis. Two of them-HlyII and cytotoxin K2 (CytK2)-are synthesized in Bacillus cereus sensu lato; the third, S. aureus α-toxin (Hla), is the most investigated representative of the family. Protein modeling showed certain amino acids of the homologous peptide to be located on the surface of the monomeric forms of these ß-barrel PFTs. We obtained monoclonal antibodies against both a cloned homologous peptide and a 14-membered synthetic peptide, DSFNTFYGNQLFMK, as part of the homologous peptide. The HlyII, CytK2, and Hla regions recognized by the obtained antibodies, as well as an antibody capable of suppressing the hemolytic activity of CytK2, were identified in the course of this work. Antibodies capable of recognizing PFTs of various origins can be useful tools for both identification and suppression of the cytolytic activity of PFTs.

Analysis of Transferrin in the Urine of Patients with Bladder Cancer Using Nanobodies.

It is known that the saturation ratio of transferrin (Tf) with iron in human blood is an important clinical parameter. Specific antibodies can be used to analyze subtle changes in the relative abundance of different forms of transferrin potentially associated with a pathological process. Recently, the authors of this study were able to obtain and characterize highly specific single-domain antibodies (nanobodies) that predominantly recognize the iron-saturated (holo-Tf) or iron-unsaturated (apo-Tf) form of transferrin. In this work, under conditions closer to physiological than in the previous experiments, we further demonstrated that these unique nanobodies have extremely high differential binding specificity for different forms of Tf in different human biological fluids. Using these nanobodies, we were able to analyze for the first time relative abundance of the transferrin forms in urine samples from the patients with bladder cancer (BC). We have shown that increase in the concentration of total Tf in the urine samples normalized for creatinine is associated with the degree of progress and growth of malignancy of BC. In the samples of healthy donors and in the early stages of BC (G1), Tf is detected in much smaller amounts (compared to the later stages) and only with additional concentration of the studied samples. For most of the studied urine samples from the BC patients, it is expected (as previously shown in the case of Tf in the blood of terminal ovarian cancer patients) that the concentration of apo-Tf is clearly higher than holo-Tf, especially in the case of the most advanced muscle-invasive BC. It was a surprise for us that approximately equal amounts of apo-Tf and holo-Tf were found in the urine samples of some patients with BC. We hypothesized that the holo-Tf fraction in this case could be largely represented by the "secondary complexes" formed by apo-Tf in combination with ions other than Fe3+, which accumulate in the urine of some cancer patients and are able to bind to apo-Tf, changing its conformation towards holo-Tf. By using inductively coupled plasma mass spectroscopy (ICP-MS), we obtained first results confirming our hypothesis. Preparation of the holo-Tf in these urine samples was found to be highly enriched in zinc and nickel. Also, relative enrichment in cadmium has been observed in this preparation, but at much lower concentrations. The obtained data indicate that the used nanobody, while recognizing predominantly the iron-saturated form of transferrin (holo-Tf), is also capable of binding transferrin in association with other metal ions that are different from iron. This ability could potentially open up new possibilities for investigation of relative abundance of various metal ions in association with transferrin in human biological fluids in normal and pathological conditions.

Fatty Acid Composition of Dry and Germinating Pollen of Gymnosperm and Angiosperm Plants.

A pollen grain is a unique haploid organism characterized by a special composition and structure. The pollen of angiosperms and gymnosperms germinate in fundamentally similar ways, but the latter also have important features, including slow growth rates and lower dependence on female tissues. These features are, to some extent, due to the properties of pollen lipids, which perform a number of functions during germination. Here, we compared the absolute content and the fatty acid (FA) composition of pollen lipids of two species of flowering plants and spruce using GC-MS. The FA composition of spruce pollen differed significantly, including the predominance of saturated and monoene FAs, and a high proportion of very-long-chain FAs (VLCFAs). Significant differences between FAs from integumentary lipids (pollen coat (PC)) and lipids of gametophyte cells were found for lily and tobacco, including a very low unsaturation index of the PC. The proportion of VLCFAs in the integument was several times higher than in gametophyte cells. We found that the absolute content of lipids in lily pollen is almost three times higher than in tobacco and spruce pollen. For the first time, changes in the FA composition were analyzed during pollen germination in gymnosperms and angiosperms. The stimulating effect of H2O2 on spruce germination also led to noticeable changes in the FA content and composition of growing pollen. For tobacco in control and test samples, the FA composition was stable.

Risk Effects of rs1799945 Polymorphism of the HFE Gene and Intergenic Interactions of GWAS-Significant Loci for Arterial Hypertension in the Caucasian Population of Central Russia.

The aim of this case-control replicative study was to investigate the link between GWAS-impact for arterial hypertension (AH) and/or blood pressure (BP) gene polymorphisms and AH risk in Russian subjects (Caucasian population of Central Russia). AH (n = 939) and control (n = 466) cohorts were examined for ten GWAS AH/BP risk loci. The genotypes/alleles of these SNP and their combinations (SNP-SNP interactions) were tested for their association with the AH development using a logistic regression statistical procedure. The genotype GG of the SNP rs1799945 (C/G) HFE was strongly linked with an increased AH risk (ORrecGG = 2.53; 95%CIrecGG1.03-6.23; ppermGG = 0.045). The seven SNPs such as rs1173771 (G/A) AC026703.1, rs1799945 (C/G) HFE, rs805303 (G/A) BAG6, rs932764 (A/G) PLCE1, rs4387287 (C/A) OBFC1, rs7302981 (G/A) CERS5, rs167479 (T/G) RGL3, out of ten regarded loci, were related with AH within eight SNP-SNP interaction models (<0.001 ≤ pperm-interaction ≤ 0.047). Three polymorphisms such as rs8068318 (T/C) TBX2, rs633185 (C/G) ARHGAP42, and rs2681472 (A/G) ATP2B1 were not linked with AH. The pairwise rs805303 (G/A) BAG6-rs7302981 (G/A) CERS5 combination was a priority in determining the susceptibility to AH (included in six out of eight SNP-SNP interaction models [75%] and described 0.82% AH entropy). AH-associated variants are conjecturally functional for 101 genes involved in processes related to the immune system (major histocompatibility complex protein, processing/presentation of antigens, immune system process regulation, etc.). In conclusion, the rs1799945 polymorphism of the HFE gene and intergenic interactions of BAG6, CERS5, AC026703.1, HFE, PLCE1, OBFC1, RGL3 have been linked with AH risky in the Caucasian population of Central Russia.

Sex-Specific Features of the Correlation between GWAS-Noticeable Polymorphisms and Hypertension in Europeans of Russia.

The aim of the study was directed at studying the sex-specific features of the correlation between genome-wide association studies (GWAS)-noticeable polymorphisms and hypertension (HTN). In two groups of European subjects of Russia (n = 1405 in total), such as men (n = 821 in total: n = 564 HTN, n = 257 control) and women (n = 584 in total: n = 375 HTN, n = 209 control), the distribution of ten specially selected polymorphisms (they have confirmed associations of GWAS level with blood pressure (BP) parameters and/or HTN in Europeans) has been considered. The list of studied loci was as follows: (PLCE1) rs932764 A > G, (AC026703.1) rs1173771 G > A, (CERS5) rs7302981 G > A, (HFE) rs1799945 C > G, (OBFC1) rs4387287 C > A, (BAG6) rs805303 G > A, (RGL3) rs167479 T > G, (ARHGAP42) rs633185 C > G, (TBX2) rs8068318 T > C, and (ATP2B1) rs2681472 A > G. The contribution of individual loci and their inter-locus interactions to the HTN susceptibility with bioinformatic interpretation of associative links was evaluated separately in men's and women's cohorts. The men-women differences in involvement in the disease of the BP/HTN-associated GWAS SNPs were detected. Among women, the HTN risk has been associated with HFE rs1799945 C > G (genotype GG was risky; ORGG = 11.15 ppermGG = 0.014) and inter-locus interactions of all 10 examined SNPs as part of 26 intergenic interactions models. In men, the polymorphism BAG6 rs805303 G > A (genotype AA was protective; ORAA = 0.30 ppermAA = 0.0008) and inter-SNPs interactions of eight loci in only seven models have been founded as HTN-correlated. HTN-linked loci and strongly linked SNPs were characterized by pronounced polyvector functionality in both men and women, but at the same time, signaling pathways of HTN-linked genes/SNPs in women and men were similar and were represented mainly by immune mechanisms. As a result, the present study has demonstrated a more pronounced contribution of BP/HTN-associated GWAS SNPs to the HTN susceptibility (due to weightier intergenic interactions) in European women than in men.

Mesenchymal Stromal Cells as a Driver of Inflammaging.

Life expectancy and age-related diseases burden increased significantly over the past few decades. Age-related conditions are commonly discussed in a very limited paradigm of depleted cellular proliferation and maturation with exponential accumulation of senescent cells. However, most recent evidence showed that the majority of age-associated ailments, i.e., diabetes mellitus, cardiovascular diseases and neurodegeneration. These diseases are closely associated with tissue nonspecific inflammation triggered and controlled by mesenchymal stromal cell secretion. Mesenchymal stromal cells (MSCs) are known as the most common type of cells for therapeutic approaches in clinical practice. Side effects and complications of MSC-based treatments increased interest in the MSCs secretome as an alternative concept for validation tests in regenerative medicine. The most recent data also proposed it as an ideal tool for cell-free regenerative therapy and tissue engineering. However, senescent MSCs secretome was shown to hold the role of 'key-driver' in inflammaging. We aimed to review the immunomodulatory effects of the MSCs-secretome during cell senescence and provide eventual insight into the interpretation of its beneficial biological actions in inflammaging-associated diseases.

Thermally activated serpentine materials as soil additives for copper and nickel immobilization in highly polluted peat.

Heat-treated serpentine products from mining wastes have been examined to remediate highly contaminated soil with total concentration of Cu 10470 mg/kg and Ni 5300 mg/kg. The series of laboratory and field experiments (for 10 years) were conducted. The modified Tessier method was used to assess the metals geochemical mobility. The effect of hydration on the chemical stability of the components and sorption properties of thermally activated serpentine were studied. The hydration of heat-treated serpentine decreased the leaching of the main components (Mg and Si) that indicates their partial binding in a newly formed compound-magnesium silicate. Hydration of heat-treated serpentine did not lead to the changes in the phase composition and the geochemical mobility of the precipitated Ni and Cu compounds. The hydration affected the sorption value at the 1 day of the interaction but after 30 days this difference partially leveled. A laboratory experiment showed that thermally activated serpentine was effective for the Cu and Ni sorption from sulfate solutions. The substantial changes in chemical properties of soil mixtures after ten years of the field experiment were found. In the first year of the field experiment, the pH values of soil mixtures were alkaline (9.4-9.9) and were significantly higher compared to the pH 4.0 of the initial peat soil. Over 10 years, the soil pH at the experimental sites gradually decreased and reached values of 7.2-8.6. The introduction of thermoactivated serpentines led to a decrease in the share of the most mobile exchangeable fraction. The most noticeable effect of thermoactivated serpentines on metal mobility in the polluted peat soil revealed for Cu; its migration coefficient decreased from 1.8 in the peat soil to 0.7 in the mixtures with heat-treated serpentines. The sum of Cu mobile fractions in the experimental variants became lower compared with initial peat by 50-70%, while Fe was lower by 30%, and Zn-by 80%. The increase in the proportion of the most strongly bound fraction was observed for all metals in the experimental variants compared with initial soil. The coefficient of metal accumulation for Ni and Cu was significantly lower than 1, indicating protective mechanisms in plants. The high content of mobile Mg and Ca compounds seems to be the determining factor in this process. The grass communities forming in the 10-years experiment showed high productivity and stability even under constant airborne industrial pollution. The thermally activated serpentine minerals can be recommended for the in situ remediation of landscapes with completely lost vegetation during the long-term impact of industrial emissions.

Isolation of nanobodies with potential to reduce patients' IgE binding to Bet v 1.

BACKGROUND: Recent studies showed that a single injection of human monoclonal allergen-specific IgG antibodies significantly reduced allergic symptoms in birch pollen-allergic patients. Since the production of full monoclonal antibodies in sufficient amounts is laborious and expensive, we sought to investigate if smaller recombinant allergen-specific antibody fragments, that is, nanobodies, have similar protective potential. For this purpose, nanobodies specific for Bet v 1, the major birch pollen allergen, were generated to evaluate their efficacy to inhibit IgE-mediated responses. METHODS: A cDNA-VHH library was constructed from a camel immunized with Bet v 1 and screened for Bet v 1 binders encoding sequences by phage display. Selected nanobodies were expressed, purified, and analyzed in regards of epitope-specificity and affinity to Bet v 1. Furthermore, cross-reactivity to Bet v 1-homologues from alder, hazel and apple, and their usefulness to inhibit IgE binding and allergen-induced basophil activation were investigated. RESULTS: We isolated three nanobodies that recognize Bet v 1 with high affinity and cross-react with Aln g 1 (alder) and Cor a 1 (hazel). Their epitopes were mapped to the alpha-helix at the C-terminus of Bet v 1. All nanobodies inhibited allergic patients' polyclonal IgE binding to Bet v 1, Aln g 1, and Cor a 1 and partially suppressed Bet v 1-induced basophil activation. CONCLUSION: We identified high-affinity Bet v 1-specific nanobodies that recognize an important IgE epitope and reduce allergen-induced basophil activation revealing the first proof that allergen-specific nanobodies are useful tools for future treatment of pollen allergy.

Double-doped YVO4 nanoparticles as optical dual-center ratiometric thermometers.

Crystalline inorganic nanoparticles doped with rare earth ions are widely used in a variety of scientific and industry applications due to the unique spectroscopic properties. The temperature dependence of their luminescence parameters makes them promising candidates for self-referencing thermal sensing. Here we report single phase YVO4 nanoparticles doped with different pairs of rare earth ions (Nd3+/Er3+, Tm3+/Er3+ and Nd3+/Tm3+) for contactless ratiometric thermometry within a wide temperature range of 298-573 K. The presence of dual luminescence centers in the optical thermometer allows one to circumvent the fundamental limitation of sensitivity inherent to thermometers based on thermally coupled levels. Important parameters for temperature sensing, such as relative thermal sensitivity and temperature resolution, were calculated for all synthesized samples and compared with the literature data. The YVO4:Tm3+,Er3+ sample displayed a relative sensitivity of 0.28% K-1 at room temperature, and the YVO4:Nd3+,Er3+ phosphor exhibited a high sensitivity of 0.56% K-1 at 573 K, while YVO4:Nd3+,Tm3+ demonstrated sub-degree thermal resolution. These findings demonstrate the good potential of dual-center ratiometric YVO4 thermometers and open the way toward future enhancement of their thermometric performances through variation of the doping concentration.

Fluorescence Polarization Immunoassay of Human Lactoferrin in Milk Using Small Single-Domain Antibodies.

Due to its unique structure and properties, human breast milk lactoferrin (hLF) has many nutritional and health-promoting functions in infants, including protection against inflammation and bacterial infections. The lack of LF in breastmilk or formula can result in the weakening of the infant's immune system. Noncompetitive polarization fluorescence immunoassay (FPIA) is a promising method for hLF quantification in milk and dairy products, which does not require the separation of the bound and free protein and allows to avoid time-consuming sample preparation. The use of fluorescently labeled single-domain camelid antibodies (nanobodies) for protein recognition in FPIA makes it possible to quantify relatively large antigens, in particular, hLF. In this work, we used previously obtained fluorescein isothiocyanate (FITC)-conjugated anti-hLF5 and anti-hLF16 nanobodies, which selectively recognized two different human lactoferrin epitopes, but did not bind to goat lactoferrin. The kinetics of hLF interaction with the FITC-labeled nanobodies was studied. The dissociation constant (KD) for the anti-LF5 and antiLF16 nanobodies was 3.2 ± 0.3 and 4.9 ± 0.4 nM, respectively, indicating the high-affinity binding of these nanobodies to hLF. We developed the FPIA protocol and determined the concentration of FITC-labeled anti-hLF5 and anti-hLF16 nanobodies that provided the optimal fluorescence signal and stable fluorescence polarization value. We also studied the dependence of fluorescence polarization on the hLF concentration in the noncompetitive FPIA with FITC-anti-hLF5 nanobody. The detection limit for hLF was 2.1 ± 0.2 µg/ml and the linear range for determining the hLF concentration was 3-10 µg/ml. FPIA is commonly used to assay low-molecular-weight substances; however, the use of fluorescently labeled nanobodies allows quantification of high-molecular-weight proteins. Here, we demonstrated that FPIA with fluorescently labeled nanobodies can be used for hLF quantification in milk.

Polyunsaturated and Very-Long-Chain Fatty Acids Are Involved in the Adaptation of Maloideae (Rosaceae) to Combined Stress in the Mountains.

One of the mechanisms of plant adaptation to combined stress under conditions of altitudinal zonation is changing the lipid fatty acid (FA) composition. The main changes in the FA composition occurred in the outer cell layers of the pericarp, but not in the parenchyma. Adaptation was found to be species-specific. In Cydonia oblonga Mill. and Malus domestica Borkh., the ratio of polyunsaturated 18:2 and 18:3 lipid FAs changed with increasing height, while a constitutive level of the unsaturation index (UI) and low contents of very-long-chain fatty acids (VLCFAs) were maintained. Mespilus germanica L. was characterized by a higher level of VLCFAs due to the high content of 20:0. The sum of VLCFAs in medlar increased by up to 16 % with changing altitude, which was accompanied by the changes in the ultrastructure of chloroplasts and a noticeable decrease in the UI. We attribute the differences in the adaptive strategies in C. oblonga, M. domestica and M. germanica to specific structural features of the pericarp peel. Despite different adaptation mechanisms, the studied species can grow equally successfully at the altitudes from 300 to 1200 m.

Aging three-spined sticklebacks Gasterosteus aculeatus: comparison of estimates from three structures.

In order to assess the accuracy and reliability of age estimates from calcified structures in the three-spined stickleback Gasterosteus aculeatus, we evaluated intra and inter-reader repeatability from three structures: otoliths, gill covers and pelvic spines). Average age estimates were also compared between the structures. The overall intra-reader repeatability of age estimates were highest for otoliths (69%), lowest for gill covers (53%) and intermediate for spine cross-sections (63%). Although four of the seven readers had the highest intra-reader repeatability score for spine cross-sections, the inter-reader variance in this structure was much higher than in others. Otoliths were the easiest in terms of their pre-analysis treatment and exchange of materials (as digital images) between readers. In addition, otoliths are more well-studied compared with the other structures with respect to their development through ontogenesis; hence, age estimates based on otoliths should be the most reliable. Therefore, our recommendation is that whenever possible, analysis of otoliths should be the preferred approach for aging G. aculeatus.

Male and female sex hormones in primary headaches.

BACKGROUND: The three primary headaches, tension-type headache, migraine and cluster headache, occur in both genders, but all seem to have a sex-specific prevalence. These gender differences suggest that both male and female sex hormones could have an influence on the course of primary headaches. This review aims to summarise the most relevant and recent literature on this topic. METHODS: Two independent reviewers searched PUBMED in a systematic manner. Search strings were composed using the terms LH, FSH, progesteron*, estrogen*, DHEA*, prolactin, testosterone, androgen*, headach*, migrain*, "tension type" or cluster. A timeframe was set limiting the search to articles published in the last 20 years, after January 1st 1997. RESULTS: Migraine tends to follow a classic temporal pattern throughout a woman's life corresponding to the fluctuation of estrogen in the different reproductive stages. The estrogen withdrawal hypothesis forms the basis for most of the assumptions made on this behalf. The role of other hormones as well as the importance of sex hormones in other primary headaches is far less studied. CONCLUSION: The available literature mainly covers the role of sex hormones in migraine in women. Detailed studies especially in the elderly of both sexes and in cluster headache and tension-type headache are warranted to fully elucidate the role of these hormones in all primary headaches.

Solutions to Yang-Mills Equations on Four-Dimensional de Sitter Space.

We consider pure SU(2) Yang-Mills theory on four-dimensional de Sitter space dS_{4} and construct a smooth and spatially homogeneous magnetic solution to the Yang-Mills equations. Slicing dS_{4} as R×S^{3}, via an SU(2)-equivariant ansatz, we reduce the Yang-Mills equations to ordinary matrix differential equations and further to Newtonian dynamics in a double-well potential. Its local maximum yields a Yang-Mills solution whose color-magnetic field at time τ∈R is given by B[over ˜]_{a}=-1/2I_{a}/(R^{2}cosh^{2}τ), where I_{a} for a=1, 2, 3 are the SU(2) generators and R is the de Sitter radius. At any moment, this spatially homogeneous configuration has finite energy, but its action is also finite and of the value -1/2j(j+1)(2j+1)π^{3} in a spin-j representation. Similarly, the double-well bounce produces a family of homogeneous finite-action electric-magnetic solutions with the same energy. There is a continuum of other solutions whose energy and action extend down to zero.

Ulipristal acetate versus placebo for fibroid treatment before surgery.

BACKGROUND: The efficacy and safety of oral ulipristal acetate for the treatment of symptomatic uterine fibroids before surgery are uncertain. METHODS: We randomly assigned women with symptomatic fibroids, excessive uterine bleeding (a score of >100 on the pictorial blood-loss assessment chart [PBAC, an objective assessment of blood loss, in which monthly scores range from 0 to >500, with higher numbers indicating more bleeding]) and anemia (hemoglobin level of ≤10.2 g per deciliter) to receive treatment for up to 13 weeks with oral ulipristal acetate at a dose of 5 mg per day (96 women) or 10 mg per day (98 women) or to receive placebo (48 women). All patients received iron supplementation. The coprimary efficacy end points were control of uterine bleeding (PBAC score of <75) and reduction of fibroid volume at week 13, after which patients could undergo surgery. RESULTS: At 13 weeks, uterine bleeding was controlled in 91% of the women receiving 5 mg of ulipristal acetate, 92% of those receiving 10 mg of ulipristal acetate, and 19% of those receiving placebo (P<0.001 for the comparison of each dose of ulipristal acetate with placebo). The rates of amenorrhea were 73%, 82%, and 6%, respectively, with amenorrhea occurring within 10 days in the majority of patients receiving ulipristal acetate. The median changes in total fibroid volume were -21%, -12%, and +3% (P=0.002 for the comparison of 5 mg of ulipristal acetate with placebo, and P=0.006 for the comparison of 10 mg of ulipristal acetate with placebo). Ulipristal acetate induced benign histologic endometrial changes that had resolved by 6 months after the end of therapy. Serious adverse events occurred in one patient during treatment with 10 mg of ulipristal acetate (uterine hemorrhage) and in one patient during receipt of placebo (fibroid protruding through the cervix). Headache and breast tenderness were the most common adverse events associated with ulipristal acetate but did not occur significantly more frequently than with placebo. CONCLUSIONS: Treatment with ulipristal acetate for 13 weeks effectively controlled excessive bleeding due to uterine fibroids and reduced the size of the fibroids. (Funded by PregLem; ClinicalTrials.gov number, NCT00755755.).

Identification of molecular subtypes of gastric cancer with different responses to PI3-kinase inhibitors and 5-fluorouracil.

BACKGROUND & AIMS: Almost all gastric cancers are adenocarcinomas, which have considerable heterogeneity among patients. We sought to identify subtypes of gastric adenocarcinomas with particular biological properties and responses to chemotherapy and targeted agents. METHODS: We compared gene expression patterns among 248 gastric tumors; using a robust method of unsupervised clustering, consensus hierarchical clustering with iterative feature selection, we identified 3 major subtypes. We developed a classifier for these subtypes and validated it in 70 tumors from a different population. We identified distinct genomic and epigenomic properties of the subtypes. We determined drug sensitivities of the subtypes in primary tumors using clinical survival data, and in cell lines through high-throughput drug screening. RESULTS: We identified 3 subtypes of gastric adenocarcinoma: proliferative, metabolic, and mesenchymal. Tumors of the proliferative subtype had high levels of genomic instability, TP53 mutations, and DNA hypomethylation. Cancer cells of the metabolic subtype were more sensitive to 5-fluorouracil than the other subtypes. Furthermore, in 2 independent groups of patients, those with tumors of the metabolic subtype appeared to have greater benefits with 5-fluorouracil treatment. Tumors of the mesenchymal subtype contain cells with features of cancer stem cells, and cell lines of this subtype are particularly sensitive to phosphatidylinositol 3-kinase-AKT-mTOR inhibitors in vitro. CONCLUSIONS: Based on gene expression patterns, we classified gastric cancers into 3 subtypes, and validated these in an independent set of tumors. The subgroups have differences in molecular and genetic features and response to therapy; this information might be used to select specific treatment approaches for patients with gastric cancer.

Integrated epigenomics identifies BMP4 as a modulator of cisplatin sensitivity in gastric cancer.

OBJECTIVE: Cisplatin is a widely used gastric cancer (GC) chemotherapy; however, genetic factors regulating GC responses to cisplatin remain obscure. Identifying genes regulating cisplatin resistance could aid clinicians in tailoring treatments, by distinguishing cisplatin sensitive patients from those who might benefit from alternative platinum therapies, and highlight novel targeted strategies for overcoming cisplatin resistance. Here integrated epigenomics is applied to identify genes associated with GC cisplatin resistance. DESIGN: 20 GC cell lines were subjected to gene expression profiling, DNA methylation profiling and drug response assays. The molecular data were integrated to identify genes highly expressed and unmethylated specifically in cisplatin-resistant lines. Candidate genes were functionally tested by several in vitro and in vivo assays. Clinical impact of candidate genes was also assessed in a cohort of 197 GC patients. RESULTS: Epigenomic analysis identified bone morphogenetic protein 4 (BMP4) as an epigenetically regulated gene highly expressed in cisplatin-resistant lines. Functional assays confirmed that BMP4 is necessary and sufficient for the expression of several prooncogenic traits, likely mediated through stimulation of the epithelial-mesenchymal transition. In primary tumours, BMP4 promoter methylation levels were inversely correlated with BMP4 expression, and patients with high BMP4-expressing tumours exhibited significantly worse prognosis. Therapeutically, targeted genetic inhibition of BMP4 caused significant sensitisation of GC cells to cisplatin. Notably, BMP4-expressing GCs also did not exhibit cross resistance to oxaliplatin. CONCLUSIONS: BMP4 epigenetic and expression status may represent promising biomarkers for GC cisplatin resistance. Targeting BMP4 may sensitise GC cells to cisplatin. Oxaliplatin, a clinically acceptable cisplatin alternative, may represent a potential therapeutic option for BMP4-positive GCs.

Comprehensive genomic meta-analysis identifies intra-tumoural stroma as a predictor of survival in patients with gastric cancer.

OBJECTIVE: Gastric adenocarcinoma (gastric cancer, GC) is a major cause of global cancer mortality. Identifying molecular programmes contributing to GC patient survival may improve our understanding of GC pathogenesis, highlight new prognostic factors and reveal novel therapeutic targets. The authors aimed to produce a comprehensive inventory of gene expression programmes expressed in primary GCs, and to identify those expression programmes significantly associated with patient survival. DESIGN: Using a network-modelling approach, the authors performed a large-scale meta-analysis of GC transcriptome data integrating 940 gastric transcriptomes from multiple independent patient cohorts. The authors analysed a training set of 428 GCs and 163 non-malignant gastric samples, and a validation set of 288 GCs and 61 non-malignant gastric samples. RESULTS: The authors identified 178 gene expression programmes ('modules') expressed in primary GCs, which were associated with distinct biological processes, chromosomal location patterns, cis-regulatory motifs and clinicopathological parameters. Expression of a transforming growth factor ß (TGF-ß) signalling associated 'super-module' of stroma-related genes consistently predicted patient survival in multiple GC validation cohorts. The proportion of intra-tumoural stroma, quantified by morphometry in tissue sections from gastrectomy specimens, was also significantly associated with stromal super-module expression and GC patient survival. CONCLUSION: Stromal gene expression predicts GC patient survival in multiple independent cohorts, and may be closely related to the intra-tumoural stroma proportion, a specific morphological GC phenotype. These findings suggest that therapeutic approaches targeting the GC stroma may merit evaluation.

Facile-Solution-Processed Silicon Nanofibers Formed on Recycled Cotton Nonwovens as Multifunctional Porous Sustainable Materials.

Limited efficiency, lower durability, moisture absorbance, and pest/fungal/bacterial interaction/growth are the major issues relating to porous nonwovens used for acoustic and thermal insulation in buildings. This research investigated porous nonwoven textiles composed of recycled cotton waste (CW) fibers, with a specific emphasis on the above-mentioned problems using the treatment of silicon coating and formation of nanofibers via facile-solution processing. The findings revealed that the use of an economic and eco-friendly superhydrophobic (contact angle higher than 150°) modification of porous nonwovens with silicon nanofibers significantly enhanced their intrinsic characteristics. Notable improvements in their compactness/density and a substantial change in micro porosity were observed after a nanofiber network was formed on the nonwoven material. This optimized sample exhibited a superior performance in terms of stiffness, surpassing the untreated samples by 25-60%. Additionally, an significant enhancement in tear strength was observed, surpassing the untreated samples with an impressive margin of 70-90%. Moreover, the nanofibrous network of silicon fibers on cotton waste (CW) showed significant augmentation in heat resistance ranging from 7% to 24% and remarkable sound absorption capabilities. In terms of sound absorption, the samples exhibited a performance comparable to the commercial standard material and outperformed the untreated samples by 20% to 35%. Enhancing the micro-roughness of fabric via silicon nanofibers induced an efficient resistance to water absorption and led to the development of inherent self-cleaning characteristics. The antibacterial capabilities observed in the optimized sample were due to its superhydrophobic nature. These characteristics suggest that the proposed nano fiber-treated nonwoven fabric is ideal for multifunctional applications, having features like enhanced moisture resistance, pest resistance, thermal insulation, and sound absorption which are essential for wall covers in housing.