Affinity of Tau antibodies for solubilized pathological Tau species but not their immunogen or insoluble Tau aggregates predicts in vivo and ex vivo efficacy

Background

A few tau immunotherapies are now in clinical trials with several more likely to be initiated in the near future. A priori, it can be anticipated that an antibody which broadly recognizes various pathological tau aggregates with high affinity would have the ideal therapeutic properties. Tau antibodies 4E6 and 6B2, raised against the same epitope region but of varying specificity and affinity, were tested for acutely improving cognition and reducing tau pathology in transgenic tauopathy mice and neuronal cultures.

Results

Surprisingly, we here show that one antibody, 4E6, which has low affinity for most forms of tau acutely improved cognition and reduced soluble phospho-tau, whereas another antibody, 6B2, which has high affinity for various tau species was ineffective. Concurrently, we confirmed and clarified these efficacy differences in an ex vivo model of tauopathy. Alzheimer’s paired helical filaments (PHF) were toxic to the neurons and increased tau levels in remaining neurons. Both toxicity and tau seeding were prevented by 4E6 but not by 6B2. Furthermore, 4E6 reduced PHF spreading between neurons. Interestingly, 4E6’s efficacy relates to its high affinity binding to solubilized PHF, whereas the ineffective 6B2 binds mainly to aggregated PHF. Blocking 4E6's uptake into neurons prevented its protective effects if the antibody was administered after PHF had been internalized. When 4E6 and PHF were administered at the same time, the antibody was protective extracellularly.

Conclusions

Overall, these findings indicate that high antibody affinity for solubilized PHF predicts efficacy, and that acute antibody-mediated improvement in cognition relates to clearance of soluble phospho-tau. Importantly, both intra- and extracellular clearance pathways are in play. Together, these results have major implications for understanding the pathogenesis of tauopathies and for development of immunotherapies.

     

Discovery of quinolinone derivatives as potent FLT3 inhibitors

Recently some fms-like tyrosine kinase 3 (FLT3) inhibitors have shown good efficacy in acute myeloid leukemia (AML) patients. In an effort to develop anti-leukemic drugs, we investigated quinolinone derivatives as novel FLT3 inhibitors. Two substituted quinolinones, KR65367 and KR65370 were subjected to FLT3 kinase activity assay and showed potent inhibition against FLT3 kinase activity in vitro, with IC₅₀ of 2.7 and 0.57 nM, respectively. As a measure of selectivity, effects on the activity of other kinases were also tested. Both compounds have negligible activity against Met, Ron, epidermal growth factor receptor, Aurora A, Janus kinase 2, and insulin receptor; with IC₅₀ greater than 10 μM. KR compounds showed strong growth inhibition in MV4;11 AML cells and increased the apoptotic cell death in flow cytometric analyses. A decrease in STAT5 phosphorylation by KR compounds was observed in MV4;11 cells. Furthermore, in vitro evaluation of compounds structurally related to KR65367 and KR65370 showed a good structure-activity relationship.     

Identification and characterization of salt responsive miRNA-SSR markers in rice (Oryza sativa)

Salinity is an important abiotic stress that affects agricultural production and productivity. It is a complex trait that is regulated by different molecular mechanisms. miRNAs are non-coding RNAs which are highly conserved and regulate gene expression. Simple sequence repeats (SSRs) are robust molecular markers for studying genetic diversity. Although several SSR markers are available now, challenge remains to identify the trait-specific SSRs which can be used for marker assisted breeding. In order to understand the genetic diversity of salt responsive-miRNA genes in rice, SSR markers were mined from 130 members of salt-responsive miRNA genes of rice and validated among the contrasting panels of tolerant as well as susceptible rice genotypes, each with 12 genotypes. Although 12 miR-SSRs were found to be polymorphic, only miR172b-SSR was able to differentiate the tolerant and susceptible genotypes in 2 different groups. It had also been found that miRNA genes were more diverse in susceptible genotypes than the tolerant one (as indicated by polymorphic index content) which might interfere to form the stem-loop structure of premature miRNA and their subsequent synthesis in susceptible genotypes. Thus, we concluded that length variations of the repeats in salt responsive miRNA genes may be responsible for a possible sensitivity to salinity adaptation. This is the first report of characterization of trait specific miRNA derived SSRs in plants.     

Assessment of genetic diversity in salt-tolerant rice and its wild relatives for ten SSR loci and one allele mining primer of salT gene located on 1st chromosome

A heterogeneous collection of rice genotypes which included seven salt-tolerant rice lines, one salt-sensitive improved line, one wild rice (Oryza rufipogon) and one salt-tolerant wild rice relative (Porteresia coarctata) was screened with ten salt-tolerance-linked simple sequence repeat markers, of which nine were from the Saltol QTL mapped on rice 1st chromosome and the rest one from 8th chromosome, having high phenotypic variance for salt tolerance. Variation in molecular weight (in the form of base pairs) of the different amplified products using RM primers was used to find out the genetic relationship among the studied rice genotypes. Genomic DNA of the studied genotypes was also amplified with a reported allele mining primer for a salt-inducible gene (salT). The amplified products were sequenced and aligned to find out the closeness among the rice lines for the studied gene. Dendrogram derived from marker profiles showed partial similarity with salT gene-derived tree. Commonly, all the salt-tolerant lines were grouped into a single cluster, including IR36 (a salt-sensitive line) to which O. rufipogon (the wild rice) and P. coarctata (the wild rice relative) joined separately. The taxonomic identity and evolutionary relationship among the three groups (rice, wild rice and wild rice relative) were bioinformatically analysed using the nucleotide sequence of the studied salT gene.     

Development of tumor lysis syndrome (TLS): A potential risk factor in cancer patients receiving anticancer therapy

Tumor lysis syndrome (TLS) is characterized by hyperuricaemia, hyperphosphatemia, hyperkalaemia, as well as hypocalcaemia due to the breakdown of tumor cells undergoing cancer therapy (chemo/radio). Therefore it is of interest to evaluate oxidative stress using selective biological markers [Malondialdehyde (MDA), Superoxide Dismutase (SOD), Glutathione (GSH) and Catalase (CAT)] in TLS. We report the marked differences (statistically significant with control) observed among a selected set of biomarkers of oxidative stress (MDA = 8.66±1.37; SOD = 0.15±0.11; GSH = 2.25±.77; CAT = 0.76±.57) in TLS patients in addition to other conventional biomarkers. Moreover, correlation was investigated among the parameters of oxidative stress and other circulating biomarkers of TLS. Data suggest the use of SOD, MDA, and GSH as potential diagnostic biomarker for TLS with other biomarkers.     

Diversity of Arsenate Reductase Genes (<Emphasis Type="Italic">arsC</Emphasis> Genes) from Arsenic-Resistant Environmental Isolates of <Emphasis Type="Italic">E. coli</Emphasis>

A polymerase chain reaction (PCR) approach was used to assess the occurrence and diversity of arsenate reductase gene (arsC gene) in arsenic-resistant environmental E. coli strains. For this purpose, two different sets of primers were designed for the specific amplification of approximately 370-bp fragments from the arsC gene. These primers were used to screen a collection of 25 environmental arsenic-resistant strains isolated from different geographical regions of India, as well as Bangladesh. The PCR results showed that 17 out of the 25 environmental isolates (68%) contained a gene related to the arsC family. Phylogenetic analysis of the protein sequences deduced from the amplicons indicated a prevalence of arsC genes in the isolated strains. A significant divergence in the DNA sequence was found in the arsC genes among As-resistant environmental E. coli strains from this study, and arsenic resistance, a genetic character, arose from a common ancestral background.     

Suppressive effect of Withania somnifera root powder on experimental gouty arthritis: An in vivo and in vitro study

The effect of Withania somnifera L. Dunal root powder on paw volume and serum lysosomal enzyme activities was investigated in monosodium urate crystal-induced rats. The levels of beta-glucuronidase and lactate dehydrogenase were also measured in monosodium urate crystal incubated polymorphonuclear leucocytes (PMNL). A significant increase in the level of paw volume and serum lysosomal enzymes was observed in monosodium urate crystal-induced rats. The increased beta-glucuronidase and lactate dehydrogenase level were observed in untreated monosodium urate crystal incubated polymorphonuclear leucocytes. On treatment with the W. somnifera root powder (500/1000 mg/kg body weight), the above changes were reverted back to near normal levels. W. somnifera also showed potent analgesic and antipyretic effect with the absence of gastric damage at different dose levels in experimental rats. For comparison purpose, non-steroidal anti-inflammatory drug (NSAID) indomethacin was used as a standard. These results provide evidence for the suppressive effect of W. somnifera root powder by retarding amplification and propagation of the inflammatory response without causing any gastric damage.     

Conformation dependent monoclonal antibodies distinguish different replicating strains or conformers of prefibrillar Aβ oligomers

Background

Age-related neurodegenerative diseases share a number of important pathological features, such as accumulation of misfolded proteins as amyloid oligomers and fibrils. Recent evidence suggests that soluble amyloid oligomers and not the insoluble amyloid fibrils may represent the primary pathological species of protein aggregates.

Results

We have produced several monoclonal antibodies that specifically recognize prefibrillar oligomers and do not recognize amyloid fibrils, monomer or natively folded proteins. Like the polyclonal antisera, the individual monoclonals recognize generic epitopes that do not depend on a specific linear amino acid sequence, but they display distinct preferences for different subsets of prefibrillar oligomers. Immunological analysis of a number of different prefibrillar Aβ oligomer preparations show that structural polymorphisms exist in Aβ prefibrillar oligomers that can be distinguished on the basis of their reactivity with monoclonal antibodies. Western blot analysis demonstrates that the conformers defined by the monoclonal antibodies have distinct size distributions, indicating that oligomer structure varies with size. The different conformational types of Aβ prefibrillar oligomers can serve as they serve as templates for monomer addition, indicating that they seed the conversion of Aβ monomer into more prefibrillar oligomers of the same type.

Conclusions

These results indicate that distinct structural variants or conformers of prefibrillar Aβ oligomers exist that are capable of seeding their own replication. These conformers may be analogous to different strains of prions.

     

Anastrozole Eliminates the Improvement Effects of Nandrolone on Hippocampal Synaptic Plasticity in Adolescent Male Rats

Biology Bulletin - Background: Nandrolone Decanoate (ND) is one of the most popular androgenic anabolic steroids (AASs) compounds that is widely abused during adolescence. Despite clear evidence...     

A Convenient Method for the Synthesis of (Prop-2-Ynyloxy)Benzene Derivatives via Reaction with Propargyl Bromide,Their Optimization,Scope and Biological Evaluation

A highly convenient method has been developed for the synthesis of (prop-2-ynyloxy) benzene and its derivatives. Differently substituted phenol and aniline derivatives were allowed to react with propargyl bromide in the presence of K₂CO₃ base and acetone as solvent. The compounds were synthesized in good yields (53–85%). Low cost, high yields and easy availability of compounds helped in the synthesis. Electron withdrawing groups favor the formation of stable phenoxide ion thus in turn favors the formation of product while electron donating groups do not favor the reaction. Phenol derivatives gave good yields as compared to that of aniline. As aprotic polar solvents favor S_N2 type reactions so acetone provided best solvation for the reactions. K₂CO₃ was proved to be good for the synthesis. Antibacterial, Antiurease and NO scavenging activity of synthesized compounds were also examined. 4-bromo-2-chloro-1-(prop-2-ynyloxy)benzene 2a was found most active compound against urease enzyme with a percentage inhibition of 82.00±0.09 at 100 µg/mL with IC₅₀ value of 60.2. 2-bromo-4-methyl-1-(prop-2-ynyloxy)benzene 2d was found potent antibacterial against Bacillus subtillus showing excellent inhibitory action with percentage inhibition of 55.67±0.26 at 100 µg/ml wih IC₅₀ value of 79.9. Based on results, it can be concluded that some of the synthesized compounds may have potential antiurease and antibacterial effects against several harmful substances.