Rational Design of Novel Anticancer Small-Molecule RNA m6A Demethylase ALKBH5 Inhibitors

The RNA 6-N-methyladenosine (m6A) demethylase ALKBH5 has been shown to be oncogenic in several cancer types, including leukemia and glioblastoma. We present here the target-tailored development and first evaluation of the antiproliferative effects of new ALKBH5 inhibitors. Two compounds, 2-[(1-hydroxy-2-oxo-2-phenylethyl)sulfanyl]acetic acid (3) and 4-{[(furan-2-yl)methyl]amino}-1,2-diazinane-3,6-dione (6), with IC50 values of 0.84 mu M and 1.79 mu M, respectively, were identified in high-throughput virtual screening of the library of 144 000 preselected compounds and subsequent verification of hits in an m6A antibody-based enzyme-linked immunosorbent assay (ELISA) enzyme inhibition assay. The effect of these compounds on the proliferation of selected target cancer cell lines was then measured. In the case of three leukemia cell lines (HL-60, CCRF-CEM, and K562) the cell proliferation was suppressed at low micromolar concentrations of inhibitors, with IC50 ranging from 1.38 to 16.5 mu M. However, the effect was low or negligible in the case of another leukemia cell line, Jurkat, and the glioblastoma cell line A-172. These results demonstrate the potential of ALKBH5 inhibition as a cancer-cell-type-selective antiproliferative strategy.Peer reviewe

An Adult`s Vitiligo in Estonia: Study of 155 Patients

Background: Vitiligo is a common depigmentary disorder characterized by white patches of the skin, hair and mucous membranes due to selective destruction of melanocytes. Objective: The objective of this study was to analyze the clinical characteristics, coexisting diseases, presence of autoantibodies and autoimmune polyglandular syndrome (APS) in Estonian adult vitiligo patients. Methods:Adult patients with vitiligo were called to participate in the study at the Dermatology Department of Tartu University from January 2005 to July 2008. One hundred fifty five subjects were examined in 141 of those the level of thyroid peroxidase antibodies (TPO-Ab), gastric parietal cell antibodies (PCA), antinuclear antibodies (ANA), antiadrenal cortex antibodies (AAA) and rheumatoid factor (RF) in blood were measured. Results: Study group (mean age 44.9 years, mean age of vitiligo onset 28.5 years, mean duration of vitiligo 16.9 years) consisted of 44 males and 111 females. Vitiligo vulgaris was the most common clinical type (81.3%), followed by acrofacial, focal, segmental and universal vitiligo. Two-thirds of subjects reported a coexisting disease and 36.7% had one or more disease of autoimmune origin. The presence of autoantibodies was established in 49.6%. TPO-Ab was found in 36.9%, PCA in 14.2%, ANA and AAA both in 2.8% and positive RF in 7.8% cases. 17 subjects had APS 3, 35 had subclinical APS 3 and two subjects had APS 4. Conclusions: Vitiligo vulgaris was the most frequent clinical type. Vitiligo was associated with other autoimmune diseases, the presence of autoantibodies in the blood was frequent (especially TPO-Ab) and many subjects had APS

The PRO2268 Gene as a Novel Susceptibility Locus for Vitiligo

Letter to the edito

Polymorphisms in Toll-like receptor genes are associated with vitiligo

Background: The members of Toll-like receptor (TLR) family are responsible for recognizing various molecular patterns associated with pathogens. Their expression is not confined to immune cells and have been detected in skin cells such as keratinocytes and melanocytes. As part of a generated response to pathogens, TLRs are involved in inducing inflammatory mediators to combat these threats. It is therefore not surprising that TLRs have been implicated in inflammatory skin diseases, including atopic dermatitis and psoriasis. Likewise, as key players in autoimmunity, they have been associated with a number of autoimmune diseases. Based on this, the role of TLRs in vitiligo could be suspected, but is yet to be clearly established. Methods: In order to conduct a genetic association analysis, 30 SNPs were selected from TLR1-TLR8 and TLR10 regions to be genotyped in Estonian case-control cohort consisting of 139 vitiligo patients and 307 healthy control individuals. The patients were further analyzed in subgroups based on sex, age of onset, occurrence of vitiligo among relatives, extent of depigmented areas, vitiligo progression activity, appearance of Köbner's phenomenon, existence of halo naevi, and incidence of spontaneous repigmentation. Results: The most notable finding came with SNP rs179020 situated in TLR7 gene, that was associated in entire vitiligo (Padj = 0.0065) and also several subgroup analyses. Other single marker and haplotype analyses pointed to TLR3, TLR4, and TLR10 genes. Conclusions: This study investigated the genetic regions of nine TLR genes in relation to vitiligo susceptibility. The main results were the associations of TLR7 SNPs with vitiligo, while several other associations were obtained from the remaining TLR gene regions. This suggests that in addition to other inflammatory skin diseases, TLRs affect the development of vitiligo, thus making them interesting targets for future research

Predicting toxicity through computers: a changing world

The computational approaches used to predict toxicity are evolving rapidly, a process hastened on by the emergence of new ways of describing chemical information. Although this trend offers many opportunities, new regulations, such as the European Community's 'Registration, Evaluation, Authorisation and Restriction of Chemicals' (REACH), demand that models be ever more robust

Discovery of small molecules that activate RNA-methylation through cooperative binding to the METTL3/14/WTAP complex active site

Chemical modifications of RNA provide an additional, epitranscriptomic, level of control over cellular functions. N-6-methylated adenosines (m6As) are found in several types of RNA, and their amounts are regulated by methyltransferases and demethylases. One of the most important enzymes catalyzing generation of m6A on mRNA is the trimer N-6-methyltransferase METTL3-14-WTAP complex. Its activity has been linked to such critical biological processes as cell differentiation, proliferation, and death. We used in silico-based discovery to identify small-molecule ligands that bind to METTL3-14-WTAP and determined experimentally their binding affinity and kinetics, as well as their effect on enzymatic function. We show that these ligands serve as activators of the METTL3-14-WTAP complex

Glial cell line-derived neurotrophic factor receptor REarranged during transfection agonist supports dopamine neurons in Vitro and enhances dopamine release In Vivo

Background Motor symptoms of Parkinson's disease (PD) are caused by degeneration and progressive loss of nigrostriatal dopamine neurons. Currently, no cure for this disease is available. Existing drugs alleviate PD symptoms but fail to halt neurodegeneration. Glial cell line-derived neurotrophic factor (GDNF) is able to protect and repair dopamine neurons in vitro and in animal models of PD, but the clinical use of GDNF is complicated by its pharmacokinetic properties. The present study aimed to evaluate the neuronal effects of a blood-brain-barrier penetrating small molecule GDNF receptor Rearranged in Transfection agonist, BT13, in the dopamine system. Methods We characterized the ability of BT13 to activate RET in immortalized cells, to support the survival of cultured dopamine neurons, to protect cultured dopamine neurons against neurotoxin-induced cell death, to activate intracellular signaling pathways both in vitro and in vivo, and to regulate dopamine release in the mouse striatum as well as BT13's distribution in the brain. Results BT13 potently activates RET and downstream signaling cascades such as Extracellular Signal Regulated Kinase and AKT in immortalized cells. It supports the survival of cultured dopamine neurons from wild-type but not from RET-knockout mice. BT13 protects cultured dopamine neurons from 6-Hydroxydopamine (6-OHDA) and 1-methyl-4-phenylpyridinium (MPP+)-induced cell death only if they express RET. In addition, BT13 is absorbed in the brain, activates intracellular signaling cascades in dopamine neurons both in vitro and in vivo, and also stimulates the release of dopamine in the mouse striatum. Conclusion The GDNF receptor RET agonist BT13 demonstrates the potential for further development of novel disease-modifying treatments against PD. (c) 2019 International Parkinson and Movement Disorder SocietyPeer reviewe

Three Thousand Years of Continuity in the Maternal Lineages of Ancient Sheep (Ovis aries) in Estonia

lthough sheep (Ovis aries) have been one of the most exploited domestic animals in Estonia since the Late Bronze Age, relatively little is known about their genetic history. Here, we explore temporal changes in Estonian sheep populations and their mitochondrial genetic diversity over the last 3000 years. We target a 558 base pair fragment of the mitochondrial hypervariable region in 115 ancient sheep from 71 sites in Estonia (c. 1200 BC – AD 1900s), 19 ancient samples from Latvia, Russia, Poland and Greece (6800 BC – AD 1700), as well as 44 samples of modern Kihnu native sheep breed. Our analyses revealed: (1) 49 mitochondrial haplotypes, associated with sheep haplogroups A and B; (2) high haplotype diversity in Estonian ancient sheep; (3) continuity in mtDNA haplotypes through time; (4) possible population expansion during the first centuries of the Middle Ages (associated with the establishment of the new power regime related to 13th century crusades); (5) significant difference in genetic diversity between ancient populations and modern native sheep, in agreement with the beginning of large-scale breeding in the 19th century and population decline in local sheep. Overall, our results suggest that in spite of the observed fluctuations in ancient sheep populations, and changes in the natural and historical conditions, the utilisation of local sheep has been constant in the territory of Estonia, displaying matrilineal continuity from the Middle Bronze Age through the Modern Period, and into modern native sheep