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Hsa-miR-181a-5p, hsa-miR-182-5p, and hsa-miR-26a-5p as potential biomarkers for BCR-ABL1 among adult chronic myeloid leukemia treated with tyrosine kinase inhibitors at the molecular response
BMC Cancer volume 22, Article number: 332 (2022)
Tyrosine kinase inhibitors (TKIs) as first-line therapy for Chronic Myeloid Leukemia (CML) show a high success rate. However, a low number of patients with long-term treatment-free remission (TFR) were observed. Molecular relapse after imatinib discontinuation occurred at 50% at 24 months, with 80% occurrence within the first 6 months. One of the reasons for relapse is untimely TKIs discontinuation caused by large errors from estimates at very low-level or undetectable disease, thus warranting new biomarkers for CML.
Next Generation Sequencing (NGS) was used to identify microRNAs (miRNAs) at the molecular response in CML adult patients receiving TKIs treatment. A total of 86 samples were collected, 30 from CML patients responsive and 28 from non-responsive to imatinib therapy, and 28 from blood donors. NGS was conducted whereby 18 miRNAs were selected and validated by real-time RT-qPCR in triplicate.
Hsa-miR-181a-5p was expressed significantly (p-value< 0.05) with 2.14 and 2.33-fold down-regulation in both patient groups, respectively meanwhile hsa-miR-182-5p and hsa-miR-26a-5p were significant only in the non-responsive group with 2.08 and 2.39 fold up-regulation. The down-regulation was consistent with decreased amounts of BCR-ABL1 in patients taking TKIs regardless of molecular responses. The up-regulation was consistent with the substantial presence of BCR-ABL1 in CML patients treated with TKIs at the molecular response.
Therefore, these miRNAs have potential as new therapeutic biomarkers for BCR-ABL1 status in adult CML patients treated with TKIs at molecular responses. These could improve current approaches and require further analysis to look for targets of these miRNAs in CML.
CML occurs mainly in adults with Philadelphia chromosome (Ph) present in most patients  as a result of a reciprocal translocation between chromosome 9 and 22 that gives rise to BCR-ABL1 transcripts, t(9;22)(q34.1;q11.2) [2, 3]. There are 3 phases of CML: chronic, accelerated, and blast crisis; CML is treated using Tyrosine Kinase Inhibitors (TKIs). Imatinib (Glivec®, Novartis), a first-generation TKI, has been widely used since the Food and Drug Administration (FDA) approval in 2001, given orally (400 mg) once daily, and patients in the chronic phase usually respond well to treatment [1, 4, 5]. It is a targeted therapy whereby imatinib competes for the BCR-ABL1 tyrosine kinase site to prevent phosphorylation and inhibit proliferation . Patients treated with imatinib are usually experiencing mild or moderate adverse effects .
Treatment progress is measured based on hematologic, cytogenetic, and molecular responses. Complete Hematologic Response (CHR) is when clinical presentations return to normal meanwhile, Complete Cytogenetic Response (CCyR) is when Ph is not detected. A molecular response is the major treatment endpoint with the optimal molecular response at BCR-ABL1 transcript level ≤ 10% by 3 months, ≤1% by 6 months is similar to CCyR and ≤ 0.1% by 12 months, known as Major Molecular Response (MMR). Beyond optimal response is known as warning or failure whereby each indicates treatment continuation, carefully considering or changing according to the patient’s conditions. Patients non-responsive or intolerant to imatinib will be administered second and third-generation (2G or 3G) TKIs as necessary, whereby nilotinib, dasatinib, and bosutinib, 2GTKIs are now choices of front-line treatment. These enable patients to not only achieve MMR faster but also much deeper molecular responses than imatinib. However, 2G and 3G TKIs showed harsher side-effects and clinical constraints without improved overall survival .
The BCR-ABL1 transcripts level quantification is by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). It is a ratio of BCR-ABL1 transcripts to ABL1 transcripts or another internationally accepted control transcript regarding the International Randomized Study of Interferon and STI571 (IRIS) trial. It is referred to as the International Scale (IS). The IRIS standardized baseline is considered as 100% and below this or 1, 0.1, 0.01% (MR4), 0.0032% (MR4.5) and 0.001% (MR5) correspond to 2, 3, 4, 4.5, and 5-log reduction from the IRIS baseline respectively. MR4, MR4.5, and MR5 are known as deep molecular responses (DMR) . A conversion factor converts laboratory ratio to an IS ratio and is determined through sample exchange with an established reference laboratory or calibrated to WHO International Genetic Reference Panel .
Patients who achieved DMR (MR4, MR4.5) have a quality of life near the general population; enable cessation studies in suitable patients. It is promising as losing MMR is uncommon after a year in treatment-free remission (TFR) otherwise indicates failure. Based on shared decision making, requirements for TKI discontinuation include first chronic phase, motivated patient, no prior treatment failure, typical e13a2 or e14a2 BCR-ABL1 transcripts, more than 5 years on TKI, and achieved DMR for more than 2 or 3 years for MR4.5 and MR4 respectively .
DMR of 4.5-log reduction or lower is preferable for discontinuation of imatinib by controlled studies due to fewer events observed and better clinical outcomes [8,9,10]. These low-level estimates are measured by using RT-qPCR which gives substantial errors, particularly from Poisson variation, with a limit of detection (LoD) of 10− 4, 10–4.5, and below [11,12,13]. These lead to disagreements in reporting as recommended by EUTOS, particularly in assigning targets to 3 for less than 3 copies [11, 12, 14]. Among suggestions for reporting of the result were to include confidence limit , average targets or as an individual result for undetectable replicate , and LoD was proposed . In most centres, the routine LoD for RT-qPCR is MR4.5 . All these have compromised the consistency of DMR which could hinder timely TKI discontinuation; therefore, a new therapeutic biomarker is necessary.
MicroRNAs (miRNAs) are small non-protein-coding, endogenous RNAs around 22 nucleotides long and function as regulators in gene expression. The expression could be down-regulated or up-regulated. Mature miRNAs are incorporated in the RNA-induced silencing complex and annealed to mRNA. Regulation of gene expression is by mRNA cleavage or translational repression . miRNAs have been studied extensively in cancer, and studies showed miRNAs have potential as a tumour suppressor or oncogene with potential use in the diagnosis, prognosis, and treatment of cancers [18,19,20,21,22,23,24,25].
NGS is suitable for detecting low-level variants as it allows deep sequencing with high sensitivity and high sequencing coverage. There are several platforms available with unique specifications . Miseq system (illumina®), an easy-to-use NGS instrument that offers short sequencing run times, long read lengths, and high data quality, is suitable for targeted resequencing. The sequencing workflow involves library preparation, cluster generation, sequencing by synthesis, and data analysis. Except for library preparation, the rest are carried out by the instrument making it a reliable platform.
In this study, we investigate miRNAs of adult CML patients, responsive and non-responsive to imatinib therapy at the molecular response. Imatinib is the front-line treatment used to treat CML, and in the event of intolerance or failure, other approved TKIs are an option as appropriate. Current treatment success is measured by the minimal presence or undetectability of BCR-ABL1 transcripts by RT-qPCR. miRNAs from the whole blood of these patients were profiled using NGS and compared with normal control. miRNAs suitable for the study were selected and verified by real-time RT-qPCR, the known gold standard for quantifying gene expression. Therefore, this study aimed to identify miRNAs that have therapeutic potential concerning TKIs at the molecular response in CML. These miRNAs could lead to discovering conditions that favour TKI discontinuation accurately, thus making long-term TFR achievable.
Patients and samples
A cross-sectional study was conducted among adults aged 18 and above in Ampang Hospital and National Blood Centre (NBC) Malaysia between 2013 and 2014. Of these respondents, 30 were CML patients responsive to imatinib therapy, and 28 were CML patients non-responsive to imatinib therapy taken from Ampang Hospital, while 28 were blood donors mostly from NBC. Peripheral blood samples were collected from all the respondents. A standardized data collection form was used to collect information from the respondents. The collection form has three domains that were sociodemographic information, treatment history with imatinib, and treatment status responsive or non-responsive. Responsive is defined as a CML patient assessed and confirmed by the clinician through clinical diagnostics and evaluations, receiving imatinib not less than 18 months, the latest BCR-ABL1IS ratio was 0.1% or less and ABL1 control gene equalled 10,000 copies or more. Non-responsive, defined as any of the above criteria that are not met and treated with imatinib, other TKIs, or other drugs as necessary. Control, defined as normal people taken from healthy donors, mostly was from the NBC who came to donate blood. The lab used a conversion factor of 0.81 to convert the BCR-ABL1 ratio to the IS ratio. Blood samples and clinical data were collected from the clinic and sent to the research laboratory.
Preparation of samples
Peripheral blood (2.5 ml) was collected from each participant using BD® Vacutainer Safety-Lok™ Blood Collection Set and drawn into PAXgene® Blood RNA tube (PreAnalytiX). Purification of total RNA including miRNA was conducted using; PAXgene® Blood miRNA Kit (PreAnalytiX) with silica-membrane technology. The quantity of total RNA was measured by NanoDrop® ND-1000 spectrophotometer. Meanwhile, the quality of total RNA and small RNA were analysed by the 2100 Bioanalyzer (Agilent) using Eukaryote Total RNA Nano assay (Agilent) for RNA Integrity Number (RIN) and Small RNA assay (Agilent) for the amount (%) of miRNA.
Next generation sequencing
Total RNA of a CML patient responsive to imatinib therapy with RIN 7.7, a CML patient non-responsive to imatinib therapy with RIN 7.4, and a normal control with RIN 8.1 were selected for NGS. TruSeq Small RNA kit (illumina®); was used to prepare libraries for subsequent cluster generation. A library was generated from each sample and labelled using a unique index sequence each. The kit contained adapters designed to directly and specifically ligate to a 5′-phosphate and a 3′-hydroxyl group of mature miRNAs. The ligated small RNA was reverse transcribed and amplified with a unique index per tube to create cDNA constructs according to the protocol (Catalogue # RS-930-1012, Part # 15004197 Rev. C, March 2011). The cycling conditions for PCR amplification were 30 s at 98 °C, 15 cycles of 10 s at 98 °C, 30 s at 60 °C, and 15 s at 72 °C, followed by 10 min at 72 °C and hold at 4 °C. Each sample was analysed using High Sensitivity DNA assay (Agilent) and obtained around 150 bp band and peak each at a 100x dilution.
The cDNA constructs (library) were purified using 6% Novex TBE PAGE Gel, 1.0 mm (Invitrogen). Libraries with unique indexes were pooled, added Novex DNA Loading Dye (Invitrogen), and loaded into 2 wells (25 μl each). The gel ran for 60 min at 145 V in the XCell SureLock™ Mini-Cell electrophoresis unit (Invitrogen™) containing 1X TBE buffer. Upon completion, the gel was removed and stained with 0.5 mg/ml ethidium bromide solution for 2 to 3 min. It was viewed under UV light, and bands between 160 and 145 bp were cut out using a razor blade. The excised gel fragment was passed through a 0.5 ml Gel Breaker tube by centrifugation, added 300 μl of Ultra-Pure water, passed through a 5 μm Filter tube by centrifugation, and concentrated by ethanol precipitation. The pellet was then resuspended in 10 μl 10 mM Tris-HCl, pH 8.5, and kept at − 150 to -25 °C. The purified sample library (different indexes) was validated using Agilent’s DNA 1000 assay (obtained a single sample peak around 150 bp) and High Sensitivity DNA assay (diluted 100x and obtained 152 bp peak, 1500 pmol/l) measured using the 2100 Bioanalyzer (Agilent).
The purified sample library was normalized to a 4 nM library using 10 mM Tris-Cl pH 8.5 with 0.1% tween 20, denatured to a 20 pM library in 1 mM NaOH using 0.2 N NaOH and pre-chilled HT1, and diluted to 13 pM using pre-chilled HT1. These were prepared according to Preparing Libraries for Sequencing on the Miseq® guide: Part # 15039740 Rev.D October 2013 (illumina®). The supplied 10 nM PhiX library was diluted to 4 nM using 10 mM Tris-Cl pH 8.5 with 0.1% tween 20, denatured to 20 pM, and further diluted to 12.5 pM. The denatured and diluted PhiX control was spiked in at least 5% to the purified sample library. The libraries were then dispensed into sample reservoir of reagent cartridge according to MiSeq® Reagent Kit v2 (50 cycles) Reagent Preparation Guide (illumina®) and ran on Miseq (illumina, USA) using Standard Flow Cell with 14 tiles imaged top and bottom for 15 M reads (approximately 5 M reads per sample). Clusters were generated and sequenced by synthesis with a single read, and then were imaged using LED and filter combinations meant for each fluorescently-labelled nucleotide. BaseSpace (illumina®) aligned, assembled, and analysed the reads. miRNAs of the responsive sample and miRNAs of the non-responsive sample were compared to miRNAs of the normal control. Eighteen miRNAs were selected mainly with regards to substantial expression level and clinical significance.
Real-time reverse transcriptase-quantitative PCR (RT-qPCR)
The selected miRNAs were ordered as ready-to-use 96-well plate Custom miScript™ miRNA PCR Array (Qiagen). The array plate consisted of the 18 miRNAs, normalisation controls (SNORD61, SNORD95, SNORD96A, RNU6–2), reverse transcription control (miRTC), and positive PCR control (PPC) in the 24 × 4 format. cDNA was prepared using miScript® II RT kit (Qiagen) for a 20 μl reaction. The reaction consisted of 4 μl 5x miScript HiSpec Buffer, 2 μl 10x miScript Nucleic Mix, 2 μl miScript Reverse Transcriptase Mix, RNase-free water, and template RNA. A starting input of 250 ng total RNA (containing miRNA) was used as recommended, which will result in 0.5 to 1 ng cDNA per array well. The reaction was incubated for 60 min at 37 °C, followed by 5 min at 95 °C, and placed on ice. The 5x miScript HiSpec buffer facilitated the conversion of mature miRNAs into cDNA while suppressing long RNAs.
miScript® SYBR® Green PCR kit (Qiagen); was used with our 96-well plate Custom miScript™ miRNA PCR Array. A reaction mixture consisted of 1375 μl 2x QuantiTect SYBR® Green PCR Master Mix, 275 μl 10x miScript Universal Primer, 1000 μl RNase-free water, and 100 μl template cDNA. Out of this, 25 μl per well was added to the verified array plate. Then the array plate was placed in Light Cycler 480 (Roche), a real-time cycler. The cycling conditions used were; initial activation for 15 min at 95 °C, 45 cycles of; denaturation for 15 s at 94 °C followed by annealing for 30 s at 55 °C, and extension for 30 s at 70 °C. PCR amplification was followed by dissociation curve analysis performed for 10 s at 95 °C, 1 min at 65 °C, and a continuous 95 °C. The miScript PCR system used; allowed sensitive and specific detection and quantification of the mature miRNAs in triplicate in all case and control samples.
A threshold value of 1.15 was used across all PCR runs and obtained threshold cycle (CT) of PPC (CTPPC) value equivalent to 19 + 2 and CTmiRTC- CTPPC value less than 7, thus allowing comparison of results. These indicated high-quality RNA samples, correct cycling program run, and the correct threshold was used, and no apparent inhibition of the reverse-transcription reaction occurred, respectively. In addition, a single melting peak was exhibited in each sample representing a specific PCR product. The Light Cycler uses a crossing point with the Fit Points method while Gene Globe Data Analysis Centre (Qiagen) uses CT, which is similar. Data was formatted in an Excel Spreadsheet using a provided template and uploaded into the Gene Globe Data Analysis Centre in a web browser (QIagen). Normalisation was conducted using the Global Threshold Cycle Mean of expressed miRNAs. Relative quantification was performed using ∆∆CT method; whereby ∆CT = CTmiRNA - CT normalisation factor, while ∆∆CT = ∆CT (experimental sample) – ∆CT (control sample). Fold change was calculated as 2-∆∆CT whereby greater than 1 was reported as fold up-regulation while less than 1 as fold down-regulation. P-values were calculated based on the Student’s t-test of the replicate 2(−∆CT) values for each miRNA. Any significant miRNA observed will be linked to TKIs received and the clinical outcomes of patients.
The majority of the respondents were Malays (51%), followed by Chinese (34%) and Indians (15%). There were more males than females in this study. The hallmark of CML (BCR-ABL1 transcript) was present in all patients who were diagnosed with CML. In responsive patients, BCR-ABL1 ratios showed a declining pattern with imatinib therapy. Common chromosomal abnormalities; usually seen with CML were observed such as; trisomy 8, which was seen in responsive and non-responsive patients. Loss of chromosome Y, an age-related chromosomal abnormality, was seen in Ph-negative older male patients. This study showed that the number of responsive and non-responsive patients was similar in Malays. The responsiveness is markedly higher among the Chinese. However, the numbers of Indian respondents were too small for comparison (Table 1).
In responsive patients, 53% achieved MMR of 7 Malays, 6 Chinese, and 3 Indians. The remaining 47% achieved DMR of 6 Malays, 6 Chinese, and 2 Indians, in which 40% were with 4-log reduction (MR4) and 7% with 4.5-log reduction (MR4.5) from the IRIS baseline. Longer duration on imatinib of around 6 years (mean 71.64 ± 34.50 months) was observed in DMR patients compared to about 5.5 years (mean 66 ± 30.43 months) in MMR patients. Overall mean duration on imatinib was 5.7 years with 53% being on imatinib for more than 5 years (Table 2). In non-responsive patients, imatinib was changed to other TKIs (82%) or other treatment options (11%) according to mutations in the kinase domain and patients’ conditions and tolerability. BCR-ABL1IS ratio was < 0.1% in 29% of patients, and the average of the latest available BCR-ABL1IS ratio was 13.7% (Table 3).
Profiles of miRNA expression by NGS showed a similar amount of mature miRNA was present in all samples, with 40.8% in normal control, 40.9% in responsive sample, and 40.3% in the non-responsive sample. A list of miRNAs with various expression levels was generated from each patient sample and compared to normal control. Among these, hsa-miR-181-5p expressed 0.45 and 0.51 times in responsive and non-responsive samples respectively.
These were consistent with real-time RT-qPCR data whereby significant down-regulations of hsa-miR-181a-5p were observed in both responsive (Group 1) and non-responsive (Group 2) groups when compared to the control group (Figs. 1 and 2 respectively). Fold change of hsa-miR-181a-5p was 0.47 (95% CI = 0.36, 0.58, p-value = 0.000006 that was < 0.05) in responsive group (Fig. 3) and 0.43 (95% CI = 0.31, 0.55, p-value = 0.000024 that was < 0.05) in non-responsive group (Fig. 4) which corresponded to 2.14 and 2.33 fold down-regulation respectively compared to control group.
In addition, another 2 miRNAs were also significant but only in the non-responsive group. These were hsa-miR-182-5p with fold change of 2.08 (95% CI = 1.60, 2.57, p-value = 0.000012 that was < 0.05) and hsa-miR-26a-5p with fold change of 2.39 (95% CI = 1.82, 2.96, p-value = 0 that was < 0.05) as shown in Fig. 4 which corresponded to 2.08 and 2.39 fold up-regulation respectively when compared to the control group. The statistical analyses of real-time PCR data were as shown in Tables 4 and 5. Comparison of these biomarkers to BCR-ABL1 and IS levels were presented in Tables 2 and 3.
In the responsive group, a reference gene, RNU6–2, a small non-coding RNA, was also significant with fold change of 0.48 (95% CI = 0.37, 0.58, p-value = 0 that was < 0.05) corresponded to 2.10 fold down-regulation when compared to the control group (Fig. 3).
We looked at our CML patients concerning imatinib therapy, a front-line treatment for CML in our government hospitals; because of treatment discontinuation for optimal CML responders as suggested by the European LeukemiaNet (ELN) 2013 and also ELN 2020. Data showed that half of our responsive patients received imatinib therapy for more than 5 years with a mean age of 48 years old, an average of 5.5 years for patients with MMR, and 6 years for DMR. These indicated on average, MMR achieved by 18 months extended survival for at least 4 years, and DMR is achievable with a longer time on therapy in our CML patients. DMR was observed in almost half of the responsive patients, and if consistent, these patients can be considered for imatinib discontinuation studies, as minimum requirements are at least 24 months of DMR , and previously 12 months of DMR . DMR was reported suitable for imatinib discontinuation as it has survival benefits and lesser events . However, determining the right time for imatinib discontinuation is critical. Long-term TFR is still low, with relapse after discontinuation being considerably high in remission patients. The weighted mean molecular relapse rate of CML was 42%, and mean molecular remission was more than 37 months before discontinuation of imatinib. A shorter duration of imatinib therapy and a shorter time with an undetectable level of BCR-ABL1 transcripts were reasons for relapse . Therefore sufficient time on imatinib and consistency in DMR is crucial before imatinib discontinuation.
Management of CML has remarkably improved over the years with a high success rate for treatment using TKIs in chronic patients. This is assisted by new analysis techniques complementing existing ones for comprehensive detection and monitoring of CML. RT-qPCR, a sensitive technique has become a gold standard in the molecular response and is used to analyse treatment response, especially from TKIs as the first line. Since RT-qPCR has limitations at very low-levels thus new biomarkers are needed not only to assist analysis but also to understand better molecular and cellular interactions and associations to prolong TFR.
Numerous novel findings; were reported from CML studies using NGS, a new technology with relatively high sensitivity and high coverage. In CML patients treated with imatinib, NGS has enabled identification of novel BCR-ABL1 fusions gene comprising BCR intron 14 and ABL1 intron 2 breakpoints, giving partial deletion of SH3 domain . Meanwhile, with whole-genome sequencing, NGS has enabled identification of e13a2-like BCR-ABL1 fusion comprising broken BCR exon 13 and 9 ABL1 intron 1 nucleotides forming a novel chimeric exon . In our study, NGS has assisted in identifying miRNAs at the molecular response in local CML patients responsive and non-responsive to imatinib therapy. Validation by custom array real-time RT-qPCR has identified hsa-miR 181-5p, hsa-miR-182-5p, and hsa-miR-26a-5p as potential new therapeutic biomarkers in response to treatment using TKIs in CML. Hsa-miR 181-5p could be a tumour suppressor miRNA and might be involved in major or partial suppression of CML. Meanwhile, hsa-miR-182-5p and hsa-miR-26a-5p could be oncomiRs in which their expressions may indicate non-responsiveness to TKIs treatment or relapse.
Hsa-miR-181a-5p was significantly down-regulated in both the responsive and non-responsive to imatinib groups compared with the control group. The down-regulation indicated significant interactions between hsa-miR-181a-5p and TKI therapies. These showed that imatinib, nilotinib, dasatinib, and bosutinib were taken by CML patients, over time had caused inhibition to CML cell proliferation, led to apoptosis, and thus reduced the amount of BCR-ABL1 in CML patients. The down-regulation observed in the responsive to imatinib group represented 3 to 4.5-log reduction (MMR to MR4.5) of BCR-ABL1 levels from the IRIS baseline with an average of 5.7 years of intake. Similarly, down-regulation observed in the non-responsive to imatinib group represented less than 1 to 5-log reduction (approaching MMR to MR5) of BCR-ABL1 levels from the IRIS baseline. These down-regulations of hsa-miR-181a-5p indicated a positive correlation to log reduction or reduced levels of BCR-ABL1 in CML patients treated with TKIs. Therefore hsa-miR-181a-5p has the potential to be used as a therapeutic biomarker for the positive performance of TKIs shown by reduced levels of BCR-ABL1 indicating patients were responding to TKIs treatment in CML. Hsa-miR-181a-5p could be a tumour-suppressive miRNA that prevents mRNA from coding specific protein directly or associated with CML in patients treated with TKIs.
On the other hand, hsa-miR-182-5p and hsa-miR-26a-5p were significantly up-regulated in the non-responsive to imatinib group compared with the control group. The fold up-regulation was 2.08 and 2.39, respectively, and was observed only in this group. These were patients whereby most of their latest available BCR-ABL1IS ratios were more than 0.1% that was neither in MMR nor did DMR, with a substantial average, and only 7%  treated with imatinib. Fold regulation of these miRNAs in the imatinib responsive group, whereby BCR-ABL1 in these patients was minimal or absent (MMR or DMR), was not significant compared with the control group. It indicated imatinib had reduced the amount of BCR-ABL1 in these responsive patients to the extent not significantly different compared with healthy donors. Thus the significant up-regulation could indicate that hsa-miR-182-5p and hsa-miR-26a-5p have a positive correlation to the substantial presence of BCR-ABL1 in CML patients treated with TKIs. Therefore, these miRNAs could function as new therapeutic biomarkers for the substantial presence of BCR-ABL1 in CML patients treated with TKIs. Hsa-miR-182-5p and hsa-miR-26a-5p could be oncogenic miRNAs (oncomiRNAs) and might be associated with CML in patients treated with TKIs.
Studies have shown hsa-miR-181a-5p as a therapeutic biomarker, has potential use for clinical improvement in cancers. It is demonstrated in gastric cancer cells whereby hsa-miR-181a-5p has been extensively studied and reported as a potential regulator of MEG2, a tumour suppressor gene , regulated RASSF6, and in combination predicts poor prognosis in gastric cancer . It also inhibited MTMR3 expression in AGS gastric cancer cells and was identified as novel autophagy . In non-small-cell-lung cancer tissues and cell lines, hsa-miR-181a-5p was reported significantly reduced and has a potential role in tumour suppression by partially targeting Kras , a protein that regulates cell growth. In breast (BC) and colon cancers (CC), hsa-miR-181a-5p was reportedly down-regulated and inversely related to matrix metalloproteinase-14, which is elevated in tumours, in which, to prevent cancer metastasis in BC and CC, is by elevating hsa-miR-181a-5p . In hepatocellular carcinoma: miR-181 was reported to significantly turn on the MAPK/JNK pathway, the regulator of cell proliferation, and by limiting it, would suppress the pathway . Hsa-miR-181a-5p, a tumour suppressor is considered as a prognostic marker in Acute Myeloid Leukemia patients treated with intensive induction chemotherapy and autologous stem cell transplant .
Higher expression of hsa-miR-182-5p as observed in our study was also seen in other studies as in prostate cancer (PCa); lung squamous cell carcinoma tissues analysed from The Cancer Genome Atlas database, the Gene Expression Omnibus database, and real-time qPCR ; and also in colorectal cancer cells using tumorigenic variant cell line MICOL-14tum compared to MICOL-14h-tert cells . In PCa, the biphasic role of hsa-miR-182-5p was observed, with a higher expression in localized PCa and contrarily lower expression in aggressive cancers . Lower expression was also observed in renal cancer whereby hsa-miR-182-5p was down-regulated in tumour tissue compared with adjacent normal tissues, and overexpression decreased tumour growth in mice, demonstrating antitumor effect .
Hsa-miR-26a-5p expression in our study is consistent with a bigger-sample size study than ours on CML patients indicating the expression of miR-26a is influenced by the response to TKIs used . The study showed miR-26a was differentially expressed; in HL-60.BCR-ABL cells when treated with tyrosine kinase inhibitors when compared to HL-60 cells. miR-26a expression levels increased gradually with both dasatinib and nilotinib treatments. The increased expression levels with dasatinib and nilotinib were consistent with our findings of hsa-miR-26a expression was up-regulated in CML patients molecularly non-responsive to imatinib compared with controls whereby most patients were treated with nilotinib or dasatinib. Thus, with dasatinib and nilotinib treatments, up-regulation of hsa-miR-26a was observed in CML patients’ non-responsiveness to imatinib. However, HL-60.BCR-ABL cells treated with imatinib mesylate showed increased miR-26a expression at 4-h but decreased expression at 8-h. It correlates with miR-26a expression was lower in chronic phase patients (62.5% Complete Cytogenetic Response (CCyR) and 37.5% imatinib-resistant) when compared with healthy individuals. miR-26a expression was also lower in imatinib-resistant patients when compared to patients who achieved CCyR (< 1% BCR-ABL1) . Hsa-miR-26a-5p expression in our study was not significant in CML patients molecularly responsive to imatinib compared with controls as these were patients either in MMR or DMR (≤ 0.1% to undetectable BCR-ABL1). These showed that with imatinib, expressions of hsa-miR-26a were decreasing from responsiveness towards resistance. Thus these expressions indicated that imatinib, nilotinib, and dasatinib had altered BCR-ABL kinase activity differently.
The main limitation of this study was the low number of patient samples collected, which was due to only a small number of patients fulfilling all the required criteria within the 2-year duration. Hence, a longer time of sample collection is appropriate. In addition, our custom plate of 4 × 24 introduced variations between plates for replicates on different array plates. It made determining a suitable threshold challenging. Instead, using an array plate of 3 × 32 for triplicate is recommended.
These studies showed that hsa-miR-181a-5p, hsa-miR-182-5p, and hsa-miR-26a-5p are promising therapeutic biomarkers and can be manipulated to improve cancer treatment. In this study, only 18 miRNAs from NGS profiles were validated using real-time RT-qPCR; thus the significant miRNAs observed could be solely or partially involved in the decreasing and or increasing of BCR-ABL1 in CML patients. Further studies are needed to determine their targets and the effect of involvement in CML. Knowing their interactions and targets in CML will give their net effects that allow manipulation by inhibiting oncomiRs or stimulating tumour suppressor miRNAs. These will give new values for TKIs responses which could assist in determining the accurate time for TKIs discontinuation and better prediction of relapse in CML. Timely discontinuation of TKIs in CML remission patients is crucial for long-term TFR. Prolong TFR would subsequently allow CML patients to achieve a good quality of life as healthy people.
Hsa-miR-181a-5p has the potential as a therapeutic biomarker for BCR-ABL1 at below current LoD in CML adult patients treated with TKIs at the molecular response and could be a tumour-suppressive miRNA. Meanwhile, hsa-miR-182-5p and hsa-miR-26a-5p have potential as therapeutic biomarkers for the substantial presence of BCR-ABL1 in CML adults and could be oncomiRNAs.
Availability of data and materials
The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.
Tyrosine Kinase Inhibitor
Chronic Myeloid Leukemia
Treatment free remission
Next Generation Sequencing
Reverse transcriptase-quantitative PCR
International Randomized Study of Interferon and STI571
Deep Molecular Response
Complete Cytogenetic Response
Limit of Detection
RNA Integrity Number
Positive PCR Control
Reverse Transcription Control
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Ferreira AF, Moura LG, Tojal I, Ambrósio L, Pinto-Simões B, Hamerschlak N, et al. Corrigendum to “ApoptomiRs expression modulated by BCR-ABL is linked to CML progression and imatinib resistance” [Blood Cells, Mol Dis. 53, (1–2), (June–August 2014) 47–55]. Blood Cells Mol Dis. 2015;55(4):420.
We would like to thank the Director-General of Health Malaysia for his permission to publish this article, the Director of Institute for Medical Research for his technical support, respondents and assisting staff from Hospital Ampang and National Blood Centre, Malaysia; Asma Shahirah Azali and Muhammad Asyraf Noh for their comprehensive samples and clinical data collection.
This work was supported by a Research Grant from the Ministry of Health, Malaysia (NMRR 12–926-13248). The funding body had no role in the design of the study and collection, analysis, and interpretation of data and in writing the manuscript.
Ethics approval and consent to participate
All procedures performed involving human participants were approved by the Medical Research and Ethics Committee (Ministry of Health Malaysia, P12–837), in accordance with the ethical standards of the institutional and or national research committee, and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. Written informed consent was obtained from all the respondents in the study before blood collection.
Consent for publication
The authors declare that they have no competing interests.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
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Cite this article
Mohd Yacob, A., Muhamad, N.A., Chang, K.M. et al. Hsa-miR-181a-5p, hsa-miR-182-5p, and hsa-miR-26a-5p as potential biomarkers for BCR-ABL1 among adult chronic myeloid leukemia treated with tyrosine kinase inhibitors at the molecular response. BMC Cancer 22, 332 (2022). https://doi.org/10.1186/s12885-022-09396-5
- Chronic myeloid leukemia
- Tyrosine kinase inhibitors
- Next generation sequencing

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Congress promised instructors education loan forgiveness вЂ” but hired creditors that managed to get impossible
The Department of Education acknowledged Thursday it might have done a more satisfactory job of assisting the tens and thousands of teachers along with other general public sector employees who had been guaranteed loan forgiveness under a government-funded system that wound up rejecting 99 per cent of candidates.
вЂњFor ten years you would imagine youвЂ™re with this path then you learn itвЂ™s all a tale,вЂќ said Kelly Finlaw, a 36-year-old art instructor in nyc.
In 2007, Congress began the general public Service Loan Forgiveness (PSLF) system to simply help relieve the burden of education loan incentivize and debt workers to get into general public service. The written text of this legislation had been clear: workers of qualifying general public solution companies whom received direct federal loans and had been in income-based payment plans could, after ten years of regular payments, have actually the remainder of the loans forgiven.
Nonetheless, simply one percent of these candidates were accepted, in accordance with a report that is recent the us government Accountability workplace, even with Congress streamlined the procedure in 2018 for rejected candidates.
One obstacle that is major borrowers happens to be the вЂњconfusingвЂќ process that will require them to utilize first for example system which is why theyвЂ™re ineligible and then make an application for another, the GAO report discovered.
The Department of Education acknowledged the intricacies of the application process at a Congressional hearing for the matter on Thursday.
“we are able to do a more satisfactory job of describing those requirements that are complicated borrowers,” said Jeff Appel, manager of policy liaison and execution into the DOE’s workplace of Federal scholar help.
That admission comes far too late for the lots and lots of instructors, nurses, police force officers, along with other general public sector employees for who 99 % of loans had been refused.
For many years, Finlaw made constant re payments, adding $30,000 towards her balance that is total of120,000. Her loan servicer confirmed that she ended up being in the path that is right loan forgiveness.
After ten years, a letter was got by her into the mail. It was thought by her had been likely to inform her that the remainder of her loan had been paid down, depending on this program. She had fantasies of escaping https://spot-loan.net/payday-loans-ca/ the tenant trap and buying her place that is own near college where she adored to the office.
She sat straight straight straight down with her roomie to together open it. вЂњ we was thinking ‘This is it,’вЂќ she told NBC Information. Then the letter was read by her. It told her that because certainly one of her loans wasnвЂ™t the kind that is right of, she didnвЂ™t qualify.
To create matters more serious, after interest is added, she nevertheless has $120,000 to visit spend down her loan.
вЂњIвЂ™m maybe maybe not planning to state the things I stated, as it wasn’t good,вЂќ said Finlaw.
Now this woman is one of the instructors suing Department of Education Secretary Betsy DeVos over her agencyвЂ™s administration associated with system.
Mike Giambona, a 42-year-old middle school psychologist from Ca, told a comparable story.
вЂњI called the organization servicing my loans in addition they said that my loans had been eligible, and I also simply had to continue to operate in general public service for ten years,вЂќ he told NBC Information.
Then one time he got a page when you look at the mail from their servicer, advising him that his loan ended up beingnвЂ™t the right type of repayment plan and then he didnвЂ™t qualify. He will be from the hook for the amount that is entire.
вЂњIt had been just like an out-of-body experience,вЂќ he stated. вЂњIвЂ™ve had therefore numerous conversations with more and more people over time. We did everything asked of me. If some body had explained We needed seriously to create modification, I would personally have.вЂќ
Again and again, rejected borrowers inform a type for the exact same tale: Their loan servicer, one of many businesses subcontracted by the Department of Education, told them every thing these people were doing ended up being fine, provided that they kept making regular re payments.
Detractors associated with the system state it qualified a lot of employees and might have experienced the unintended result of encouraging borrowers to battle more financial obligation than they might pay for.
” The denial that is high in PSLF are by Congressional design, perhaps perhaps not by accident or failed execution by the Department,вЂќ Education Department Press Secretary Angela Morabito stated whenever contacted by NBC Information.
NBC News reached away for remark to many regarding the loan that is major: PHEAA/FedLoan Servicing, Great Lakes, Navient, and Nelnet.
вЂњWe provide complete details regarding the different payment options, deferment/forbearance choices and loan forgiveness programs, including teacher loan forgiveness and public solution loan forgiveness, on our web site,вЂќ Paul Hartwick, a representative for Navient, told NBC Information.
FedLoan, Nelnet and Great Lakes referred requests for remark into the Department of Education.
вЂњThis will cost the authorities numerous billions of dollars,” stated Richard Fossey, a teacher of training legislation and policy during the University of Louisiana at Lafayette. “It is the equivalent that is governmental of Titanic hitting an iceberg.”
The DOE stated it agreed with all recommendations for improvement made by the watchdog GOA, and has already begun to implement other changes thursday.
Ben Popken is really a senior company reporter for NBC Information.

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text:
Point It Out Stamp Set
- INTRO VIDEO
Get the Bundle! (5% off!): Point it Out Stamp & Die Bundle
Coordinating die: Point It Out Die Set
When you want to draw attention to an element on your project, these stamps will help you "point it out!" The trendy arrows would look great stamped in different colors to highlight your photos, page titles, or journaling.
This is a 4” x 6” clear stamp set containing 12 stamps made in the USA from high quality photopolymer.
Love all the sayings!

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Are you looking for the best Medigap insurance in Avonmore, PA? We can help you find affordable and quality Medicare Supplemental Insurance in Pennsylvania. You can either call 888-602-5343 or enter your Zip Code above and fill out the form. We will help you find the best Avonmore, PA Medigap insurance options.Buying Medigap, or Medicare Supplemental Insurance, is the key for many people over age 65 to be able to afford to use their Medicare coverage. Medigap is a separate policy that is sold by private insurance carriers that adhere to Pennsylvania laws.
For the most part, Medigap covers co-insurance, co-payments, and even includes medical coverage if faced with an emergency when traveling outside of the United States. Older Medigap plans also covered deductibles, though those were mostly phased out in 2010. Medigap covers costs seen in Medicare Part A and Part B Plans. Medicare Advantage plans are also available though not in conjunction with Medigap benefits. Likewise, be aware that it is illegal to sell a Medigap holder Medicare Savings plans.
The reason people in Avonmore, PA buy Medigap is to cover where an individual’s money will fall short. Buying a Medigap policy will require monthly premiums in addition to Part A and Part B Plan premiums. Though, the fact is that there are still ways to save on Medigap coverage.
For one, apply for policies while in the first 6 months of your Medicare Part and Part B plan policies. During this period, guarantee issuance is extended to anyone over the age of 65 within the first six months. It means that there are no denials or increases in premiums because of pre-existing conditions, based upon past medical history, or current medical problems that spring up.
If an individual waits they may pay higher rates and fees for extending past the deadline. It is better to shop around and compare quotes and do so during the open enrollment period. The majority of Medicare Supplemental plans are standard and demarcated by letters A through N. Each plan covers benefits but have added benefits. For instance, part D covers prescriptions.
That means that while cheap Medicare Supplemental Insurance plans A through G charge higher premiums, costs that are out-of-pocket are not as expensive as for Plan K through Plan N. Those carry lower premiums and like benefits but at the trade-off of costing more at the time of service.
Employer Retiree Benefits
The next way to save money beyond following the open enrollment dates is to find out if a former or present employer has a Medigap policy of its own. In general, these will be less costly than an individual policy that you buy on your own. Though, the warning and word to the wise is to carefully consider if the company is financially stable or not.
Whenever purchasing many types of Medicare plans, you actually cancel the old policy before instating the new policy. That is not the case where Medicare A and B are concerned. The reason, recall, is that affordable Medicare Supplemental insurance coverage is meant to fill the gaps of conventional original Medicare policies.
It requires still being covered under Medicare. The other benefit is to know when shopping around that the policies are the same whether you buy with Humana or Aetna or BlueCross BlueShield, for instance.
What is worth looking at is the reputation of the health insurance company, the quality of the coverage they offer, and the credit worthiness rating of the insurance company. The reason to look at the credit worthiness of the company offering up the insurance is to ensure that they are likely to have the money to pay out benefits and claims as agreed in the policy paperwork.
In addition to looking for insurance carriers that have an A or greater rating in the creditworthiness area, it is important to find out about the customer service and the customer satisfaction ratings of the companies you are comparing as well. It pays to have the company that pays claims within a timely manner, without a heroic effort on your part or the part of your doctors.
Otherwise, you will spend time having to contact the Pennsylvania insurance department or commissioner to file official complaints to get the claims paid within the acceptable two-month window. All of the Plan C Medigap coverage will be the same as that same plan from another insurer. Look at service quality and price. Look at comparing insurance company offerings to locate a cheap medicare supplement insurance plan in Avonmore, PA .
You will also need to look for separate prescription drug coverage, which is designated by Medicare Part D. This will add in an additional premium in addition to other costs. The key is that the majority of people who qualify for cheap medicare supplemental insurance are 65 and older, and look for their Medigap during open enrollment.
In addition , the other requirements stipulate Medicare Part B and Part A participants are eligible. Once you choose a inexpensive Medicare Supplement insurance it is as if it is written in stone and can not be altered.
To be eligible for cheap Medicare Supplemental insurance plans in Avonmore, PA apply for coverage offered by insurers within your state. All policies are available in each state, though most states will have one single policy available for Medicaid plan holders.
Everyone at age 65 and older and who is also enrolled in the Medicare Parts A and B is eligible to purchase a supplemental plan. It is advisable to decide on a plan during the 6 months of open enrollment to pay the most favorable policy premiums. When comparing costs during the open enrollment, it means pre-existing conditions are not factored into the pricing for the premiums. It is the hands-down best way to save money on a Medigap Supplemental plan in Avonmore, PA .
Always find out the Pennsylvania laws prior to shopping. They can vary a bit, though it is important to recall that the policies are uniform across states. Use online free quotes searches to find all of the options available to you. Narrow it down based upon credit worthiness of each of the insurers, along with their quality of care and their speed of paying out claims.
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More Tips About Avonmore, PA Medicare Supplemental Insurance
Sometimes, the health insurance marketplace can feel like a minefield. It will not have to feel that way, however. There is loads of tips offered that can assist you get the very best offers and the best treatment out of the funds and time you place into working with health insurance policy firms. Go through on for some helpful tips.
When you have made the decision on purchasing personalized wellness insurance policy, you ought to get a copy of exactly what the strategy will seem like ahead of you make the last buy. Make certain that you study all of the fantastic print, exceptions and clauses, so that you will know what exactly you might be getting and what coverage you may conclude up being denied.
Dental insurance can actually assist cut the expense on dental repairs. Your tooth are a very crucial component of your wellness, but a expensive one particular for most men and women. Obtaining dental insurance will help to lower the whole expense of all your dental work so you can find the money for to have a healthful mouth.
If you graduated from university just lately and buying for wellness insurance coverage, then you can select from a couple of distinct choices. You can use worker-based insurance if you've got a job. Or, if you're underneath twenty-six, you can remain on the insurance prepare of your mothers and fathers.
Understand you and your people well being position when shopping for insurance policies. If you are a single healthy individual, you can take the threat and go for a lower-cost, tremendous-high, deductible strategy, as the odds are that you will not need to have to use it as frequently. If there is a historical past of ailment or if you have young children, you will want to pay out a little far more for every thirty day period for a lower deductible.
Get overall health insurance policies via a group. Whether it be an staff team, a union association, or other group that functions with particular classes of men and women, examine to see if they have well being insurance policy. A lot of teams offer you discounted health insurance policies policies if you sign up for it as a group member, so join up!
The large expense of well being insurance has triggered numerous individuals to reconsider getting coverage. But accountable people recognize they never know when sickness/ailment is likely to arise, so to protect by themselves in the function this occurs, they decide on to purchase overall health insurance.
Even if you think you have found the excellent wellness insurance coverage organization, it does not hurt to appear close to for other folks. By buying all around, you could locate that there is an even much better well being insurance policy company for you and your specific needs. You may possibly even conserve income by searching around.
If your overall health insurance coverage arrives by means of your employer, you clearly will not have a lot decision about who insures you and your loved ones. You do, nevertheless, have some choices about what possibilities you want. Be as active a buyer of your healthcare insurance, as attainable. Just take the time to understand the philosophical and real variances in between HMOs and PPOs and the attendant differences in cost composition. You need to be armed with this info, in get to make wise choices about your health care insurance policies.
You never know when ailment or injuries will strike, and the costs of remedy can be fiscally devastating. It is wise to have wellness insurance to minimize the chance that sudden healthcare bills will lead to fiscal hardship. From bare-bones policies to all-inclusive alternatives, there is a well being insurance coverage policy to go well with any circumstance.

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Huda Lutfi is both a visual artist and cultural historian living and working in Cairo. Lutfi works like an urban archaeologist, constantly digging up found objects and images as loaded fragments of history. Her training as a cultural historian has influenced her work as a visual artist, juxtaposing historical past and present, as well as traversing and fusing cultural boundaries. In her works she references Egypt’s social, cultural and popular artistic traditions, trying to integrate these into contemporary art practices. In her attempts to do so, she works with mixed media, using painting, text, collage, photographs, found objects, ready-mades, assemblage, video and installation. Presently Lutfi is working on a solo exhibition in Cairo, in which she references the rich archive of images and texts she collected since January 2011. She has exhibited both locally and internationally: in Cairo, Alexandria, Paris, London, Liege, Thessalonica, Marseille, Frankfurt, Bonn, Bamako, Dakar, Virginia (USA), Bahrain, Dubai and Tunisia.
On Air: Huda LutfiThe ikono On Air Festival is showing "Cairo Resonances", a new film by Huda Lutfi (Soundtrack: Jacob Kirkegaard / Camera and Editing: Karim El-Sharnouby)
Links- Ahram online: Huda Lutfi - The artist and the historical moment - Huda Lutfi at Culture Base - Huda Lutfi at re-title.com - Huda Lutfi: The Artist and the Historical Moment - Huda Lutfi on Sultan Gallery's Blog
VideosHuda Lutfi: The Artist and the Historical Moment: A Testimony Summer Academy: Aesthetics and Politics lecture entitled "The Artist and the Historical Moment: A Testimony" presented by Huda Lutfi at the American University in Cairo. Image: Huda Lutfi, "Stripping off the Garments". Photo: Courtesy The Townhouse Gallery.
Curators' SpecialPart of the curatorial selection of Karin Adrian von Roques

domain: artistic


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Hailing from Kansas City, Missouri, rapper JL B. Hood, born Jason Varnes, got his start as one-third of B.
Hood, a rap group sometimes known as Brotherhood, in which he was joined by his two brothers. JL released his first mixtape, Just Landed, in 2011, immediately catching the attention of both fans in his local hip-hop scene and more established rap artists. He was featured on Tech N9ne's 2011 single "Worldwide Choppers," still new into his career but trading scattershot verses with luminaries like Busta Rhymes, Twista, Yelawolf, and others. He continued to gain steam, releasing his Adderall mixtape and proper debut solo LP, Brain Scatter, in 2013 and touring in support of huge names as well as headlining his own shows. More collaborations and mixtape releases followed, including Brain Scatter 2 in 2014 and Symptoms in 2015. He eventually signed a deal with Strange Music, and in 2017 he released Tech N9ne Presents Dibkis on the label, a massively produced album with cameo appearances from Church Boi, Joey Cool, Tech N9ne, and many others. ~ Fred Thomas, Rovi

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The Samoa National Provident Fund, (SNPF) members can expect their annual dividends and bonus at nothing less than 8 percent of their total contributions starting this year.
The new minimum threshold was announced this week by the SNPF Executive noting that it reaffirms the Fund’s priority to maximise benefits for its members and contributors.
Under the SNPF law, the Fund is mandated to pay an annual bonus to contributors of not less than four percent at the end of the fiscal year.
But in the last ten years, the minimum required by law was surpassed with annual dividends paid out averaging 7percent to 8 percent.
However, in 2018, the SNPF set a new record with its ten percent bonus paid to contributors for a total of over $53 million (US$20.3 million) injected into the local economy.
Meanwhile, the Samoa National Provident Fund have announced concessions for businesses to allow a smooth transition while they adjust to the mandatory increase for SNPF members’ contributions to the Fund.
The SNPF will waive the penalty fee for late payments of contributions and will allow partial payments, so the business community can adjust to the increase, which comes into effect in July.
SNPF Chief Executive Pauli Prince Suhren says that Government, through the Fund’s Board and Management, are not blind to the crucial role of and contribution by the Business Sector to the development of the local economy.
Pauli and the Samoa Chamber of Commerce met this week to discuss the increase, amongst other issues.
SOURCE: SAMOA GOVT/PACNEWS
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The Metropolitan Police are demanding £7,000 from a woman they paid damages to after she discovered that her long-term partner was a police spy. The claim is part of their ongoing campaign for secrecy around political undercover police units who have committed human rights abuses.
Helen Steel has been a lifelong social justice campaigner. In the 1990s she was one of the defendants in the McLibel trial, which arose after McDonald’s sued campaigners for libel over a leaflet produced by London Greenpeace. McDonald’s spent millions on the case, but a public support campaign meant the trial was dubbed ‘the greatest corporate PR disaster in history’.
POLICE SPIES AND CORPORATE SPIES
At the trial it was revealed that London Greenpeace had been infiltrated by several corporate spies hired by McDonald’s. But it was only years later that it emerged the group was also infiltrated by undercover police officers from the now-disgraced Special Demonstration Squad. One of them, Bob Lambert, co-wrote the What’s Wrong With McDonald’s leaflet that caused the trial, though this fact was kept from the court. Another officer in the group, John Dines, deceived Steel into a two-year relationship. They lived together, discussed starting a family and planned to spend the rest of their lives with one another. Then Dines feigned a breakdown and disappeared from Steel’s life back into his police career.
In 2011, Steel was one of a group of eight similarly deceived women who brought a legal case against the Metropolitan Police for abuse by five undercover officers. The Met spent many years and huge sums of public money obstructing the case.
NEITHER CONFIRM NOR DENY
After almost three years of the Met claiming they could ‘neither confirm nor deny’ (NCND) that anyone was an undercover officer – a tactic Steel forensically dismantled at a later public inquiry hearing – in 2014 the courts forced their hand.
The women challenged the police use of NCND. In July 2014 they won an important victory when the High Court ruled that there was no legitimate public interest in the Met Police asserting NCND in respect of the allegations that officers had engaged in long term intimate sexual relationships while undercover.
The Court also ruled that as Bob Lambert and Jim Boyling had already been publicly confirmed as undercover officers, the police could no longer maintain NCND in respect of their identities.
MET POLICE CONTINUE TO HIDE THE TRUTH
However, regarding Mark Jenner and John Dines, the Judge said that although the evidence amassed by the women was overwhelming, and it was surely only a matter of time until they were confirmed, he could not force the naming of people who hadn’t outed themselves.
As Steel said at the time
‘It is very disappointing that despite the overwhelming evidence our former partners John Dines and Mark Jenner were also undercover SDS officers, the Judge has allowed the Met to continue to hide the truth about them.’
Steel put in an appeal against this decision. These men were not private individuals, they had been acting as public servants, so the public had a right to know.
A few months after this appeal was lodged the Met held talks with the women to seek a settlement for their civil claims. Just before Christmas 2014 the Met agreed to apologise to the women, though it wasn’t finalised and published for another 11 months.
On 31st December 2014, the Appeal Court agreed that NCND was an important issue and Steel’s argument was well-grounded. They granted her leave to appeal.
COURT GRANTS APPEAL, MET CLAIMS DECEPTION
The Met, with their tactic of trying anything to undermine those they have victimised, attempted to get the appeal struck out. They claimed Steel had misled the appeal court by not informing them that a settlement had been agreed. This was an underhanded trick, given that the settlement hadn’t been finalised, she was unrepresented for the appeal, and the events happened over the Christmas period when people are generally not focussed on legal proceedings.
Steel argued that it was in the public interest to name those responsible for the abuses. A hearing for the Met’s strike-out application took place in July 2015. Steel was unrepresented, and mentally exhausted from the long battle for the truth. She reluctantly acceded to the court’s twofold advice.
Firstly, if she lost the appeal she would be liable to pay the Met’s legal costs, which could wipe out her entire damages in the main claim. Secondly, the forthcoming public inquiry would provide a safer route to argue about the use of NCND and the release of spycops’ names as there were no costs risks.
STEEL DROPS APPEAL, MET CLAIMS £10,000 COSTS
Despite the hearing lasting only about an hour, the police then claimed over £10,000 costs. Although later reduced to £7,000, the ludicrous amounts charged act as a deterrent, intimidating members of the public seeking accountability for wrongdoing committed by police officers. The threat of such an award can be used by the police as a means to intimidate people out of seeking redress.
The police’s whole argument – that a settlement was agreed – rested on them issuing an apology admitting these men were Met officers who inexcusably abused women. The Met concede they were wrong, and that the women who were deceived into relationships were blameless. Why should officers who have abused members of the public be allowed to hide behind a wall of secrecy?
The apology came with damages for the harm caused by the extreme deception. The Met are now trying to claw money back from a woman they victimised because she tried to get them to do something that they should have done anyway.
PARTIAL CONFIRMATION, MORE DENIAL
The Undercover Policing Inquiry eventually confirmed that Dines was a police officer in December 2016 – a grudging and minimal admission that Steel excoriated. To this day, the police won’t admit Mark Jenner was the undercover officer Mark Cassidy, even though he’s been publicly identified since January 2011.
Women deceived into intimate relationships by undercover police officers want to ensure that these human rights abuses never happen to anyone else. This requires the Met to stop protecting the identities of the abusers. It also requires a legal system that allows funding to enable those who have been abused to challenge their abusers without the risk of becoming bankrupt or losing their homes.
For the Met to have abused these women is horrific enough. For them to inflict the second injustice of legal tricks and obstructions compounds their cruelty. To then to go after Helen Steel for money is an utterly outrageous further leap into the shameless bullying and corruption that has driven their response to the spycops scandal from the start.

domain: law


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Dylan Wang is back on screen again after Love Between Fairy and Devil (苍兰决)！
Did you watch the famous Love Between Fairy and Devil (苍兰决) drama? If you did, you have probably felled in love with Dylan Wang like how we did. He is set to return in another iQIYI costume period original, Unchained Love (浮图缘), a revenge flick that explores the palace politics set in ancient China.
Together with Dylan Wang (王鹤棣), Yukee Chen (陈钰琪), Peter Ho (何润东) and Zeng Li (曾黎) will also be appearing in Unchained Love (浮图缘).
In Unchained Love, Wang follows his role as the overlord Dongfang Qingcang and continues to show off his penchant for charismatic and “boss-like” characters. This time as the overbearing and arrogant Xiao Duo (肖铎), an imperial eunuch who wields the supreme power in court, only answering to the emperor himself. Six years ago, Xiao Duo’s brother was murdered in the capital’s streets. Since that day, Xiao Duo has vowed revenge on those responsible for the killings.
And Peter Ho (何润东) is finally back on screen after a year’s hiatus! Peter Ho is known throughout the industry in many identities. While he is usually known for his macho and manly character, this time, he plays a new challenging role as a timid and simple-minded royal prince who is looked down upon by others in court.
Zeng Li(曾黎), who boasts over 20 years of acting experience plays Empress Rong An (荣安皇后). She holds the crown prince hostage and tries to solidify her place as Empress Dowager in the new reign. Also, she holds a candle for Dylan Wang’s handsome eunuch Xiao Duo. She often tries to seduce him. Gradually, forming a grudge as she gets rebuffed and outsmarted by him time and again.
Xiao Duo collaborates with Mu Rong Gao Gong to plan a change in the powerhouse within the palace. By aiding the ascend of Mu Rong Gao Gong as emperor. Bu Yin Lou, a lady-in-waiting, was supposed to be sacrificed upon the emperor’s death but was saved by Xiao Duo. Placed under his care of him. It was through living in close proximity that feelings started developing between the two but as it was a forbidden relationship, both swept their feelings under the mat.
For the better sake of both, Bu Yin Lou decided to sacrifice herself and sent herself into the deeply complicated palace only for Xiao Duo to misunderstand her intention of doing so as betraying him. Will the two overcome all odds and ends on a good note?
Unchained Love is exclusively streaming on iQIYI & apps only.

domain: entertainment


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Multi-Day Virtual Event Also Featured Guidance from the Council and Payment Security Experts on Challenges Faced in 2020
WAKEFIELD, Mass., 9 October 2020 — More than 1,800 stakeholders attended the North America Community Meeting, which provides a forum for leaders in the payment security industry to share knowledge, collaborate and promote understanding and adoption of PCI Security Standards and Programs. The multi-day virtual event, moderated by BBC America Anchor Katty Kay, featured insights into the development of the PCI Data Security Standard (PCI DSS) v4.0, addressed payment security challenges stemming from the COVID-19 pandemic, and the growing importance of software security. Experts included keynote speakers Dr. Jessica Barker and F.C., co-founders of Cygenta, who spoke about the increasing cyber threats during the COVID-19 pandemic.
“2020 has been a year of global challenges that have impacted every stakeholder in the payment security landscape. The Council listened to the industry and has provided guidance and adapted programs to help the industry address those challenges,” says Executive Director Lance J. Johnson. “While many of the challenges we experienced have been new and different, the Council’s mission remains the same – to support the needs of the global payments industry and enhance global payment account data security.”
The event featured a four-part series dedicated to sharing insights about how industry feedback has helped shape the current draft PCI DSS v4.0. The flagship payment data security standard is currently open for stakeholders to provide feedback during a request for comments period. Emma Sutcliffe, SVP, Standards Officer notes: “Stakeholder feedback plays an integral role to ensure that the PCI DSS continues to meet the needs of the global payment card industry. The standard is being updated to address this feedback and to support a range of environments and technologies. The draft also includes a new customized approach that will provide organizations more flexibility for meeting the security objectives of PCI DSS requirements.” For more information read PCI Perspectives Blog: Request for Comments: PCI DSS Version 4.0 Draft Standard
Other key topics discussed at the meeting included:
- PCI SSC Response to COVID-19 Challenges
PCI SSC collaborated with industry leaders and listened to stakeholder feedback related to the impacts of the pandemic to the global payments industry. In turn, the Council took action by providing supporting resources on such topics as how to perform remote assessments, extending expiry dates and deadlines and moving all in-person training to an online format. Resources can be found on the PCI Perspectives Blog, and on the newly-created hub of resources the Council created to keep the industry informed of these updates. View the PCI SSC COVID-19 Resource Page.
- PCI Software Security Framework
When the Payment Application Data Security Standard (PA-DSS) program ends on 28 October 2022, it will be replaced by the Software Security Framework. This framework standardizes and consolidates software security principles and practices for payment software and software development entities under a single requirements architecture with supporting validation and listing programs. The Council discussed the growing importance of software security and how the two programs compare, shared key timelines, and offered suggestions on how organizations can prepare for the transition.
- Upcoming Request for Comments
In addition to the PCI DSS v4.0 Request for Comment (RFC), which is currently open, the Council has two additional RFCs planned for the November/December 2020 timeframe: Card Production v3 Draft Standard and PTS HSM v4 Draft Standard. For more information visit the RFC page: Request for Comments.
- Updates to Standards
To help address the accelerating changes to payment security, the Council published updates to the PCI PIN Transaction Security (PTS) Point-of-Interaction (POI) Modular Security Requirements v6.0 and the PCI Software-based PIN Entry on COTS (SPoC) Standard. Before the end of the year, the Council will publish version 3.1 to the PIN Standard which is being updated based on feedback provided by the industry via an RFC period earlier this year.
- Board of Advisor Nomination Period
The 2021-2022 Board of Advisors nomination period runs until 26 October 2020, followed by the election period from 9-20 November 2020. The Board of Advisors represents PCI Security Standards Council (PCI SSC) Participating Organizations worldwide to ensure global industry involvement in the development of PCI Security Standards. For more information visit the PCI Perspectives Blog: Make a Difference: Serve on the 2021-2022 PCI SSC Board of Advisors
Visit the PCI SSC website for more information on all of the PCI Council’s efforts and activities including how your organization can attend the next PCI SSC Community Meeting and join the global cross-industry effort to increase payment security.
Save the date for the next two virtual PCI SSC Community Meetings:
- Europe Community Meeting: 20 – 23 October
- Asia-Pacific Community Meeting: 4 – 6 November
About the PCI Security Standards Council
The PCI Security Standards Council (PCI SSC) leads a global, cross-industry effort to increase payment security by providing industry-driven, flexible and effective data security standards and programs that help businesses detect, mitigate and prevent cyberattacks and breaches. Connect with the PCI SSC on LinkedIn. Join the conversation on Twitter @PCISSC. Subscribe to the PCI Perspectives Blog.


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Home > Tours > Andretti Indoor Karting & Game Play Card
Andretti Indoor Karting & Game Play Card
Experience the adrenaline rush of our electric go kart races with instant acceleration as you put the pedal to the metal around hairpin turns and long straightaways on our climate controlled tracks and enjoy the 120 games in the arcade with your game card!
2 to 3 Hours
Mobile & printed vouchers accepted
Fully refundable when canceled up to 24 hours prior to departure.
Enjoy fast acceleration and the rush of a high-speed electric Superkart on the world's longest multi-level track
Feel the adrenaline course through your veins around hairpin turns!
Play on over 120 Arcade Games!
Avoid the rain and Florida weather on a climate controlled track!
Adult Indoor Karting (Min Height 54'') - Seven Minute Heats of an exhilarating ride while driving a Sodi Electric Kart on a banked multi-level track AND a $10 Game Card
Child Indoor Karting (Min Height: 48") - Six Minute Heat take place on Track One. Guests will enjoy a fun ride while driving a Sodi Electric Kart on our Andretti Junior Track AND a One Hour Game Card
Additional activities at the complex
Food and Beverages
Each Driver will watch a brief video detailing the proper use of the equipment, basic kart operation and important rules and procedures, including the meaning of each flag
If you are under the age of 18, your parent or legal guardian must complete a waiver, but does not have to be present during your entire visit of the facility.
Closed toe shoes are required to race.
Complex is closed on Thanksgiving Day and Christmas Day
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At our Orlando facility, we feature the best model of electric karts in the industry, the SODI RTX. These karts are powerful, ergonomic, safe and environmentally friendly plus pack enough torque to take you all the way to victory lane. For our younger drivers, we feature the Sodi LRX electric karts that take the sweeping turns of the ‘pear-shaped’ junior track on a dime and provide all the heart-pounding thrills your little one craves!
Track 1 is a road-course style single level track for Junior drivers.
Track 2 is a A multi-level track with 25 feet of elevation changes and a scenic ‘bumped-out’ turn with a view of Universal Blvd.
Track 3 is a fast road course style track with elevation changes, six banked turns and long straightaways.
Come experience the adrenaline of being behind the wheel!
Don't skip the arcade! We take our fun pretty seriously. That means state-of-the-art, 3D, multi-player arcade games, life-sized games of your favorite apps, and your favorite timeless classics. Test your skills, compete with friends, and engage your senses. It’s Game On!
This does not include transportation to the activity. Guests must make their own way to Andretti Indoor Karting & Games Orlando located at 9299 Universal Blvd, Orlando, FL 32819


Where there’s smoke, there’s fire. High school students enrolled in Del Mar College’s 2012 Public Safety Academy offered by the Department of Public Safety Education learned the mechanics of saving a life. Using Cardiopulmonary Resuscitation (CPR) after a fire rescue, campers learned how to count chest compressions and administer oxygen to mannequins while a CPR instructor gave instructions.
On Thursday, July 26, the campers put what they’ve learned over the past three weeks to the test during a simulated rescue.
The fire rescue and CPR exercise was just one of several different activities designed to challenge this year’s academy participants. The summer program consisted of afternoon sessions that began July 9.
“The Summer Public Safety Academy educates and gives campers insight into the public safety sector,” says Patrick McBride, DMC Emergency Medical Technology Instructor and Public Safety Academy Director. “They learn how important public safety is to our community. Campers learn how important it is to make good life choices, along with engaging in physical fitness and team leadership while forming special bonds with other participants.”
The 49 students represent John Paul, West Oso, Flour Bluff, Corpus Christi College Prep, Collegiate and Corpus Christi high schools as well as South Park Middle School, Richard Milburn Academy, Skidmore-Tynan High School, London ISD and Bishop High School.
During the camp, participants were also introduced to basic first aid, including CPR, and were briefed about fire behavior, rappelling and the utilization of a self-contained breathing apparatus. Other mini courses encompassed Crime Scene Investigation exercises, better known as “CSI,” and students were introduced to the College’s weapons simulator.
IN THE PICTURE: Public Safety Academy campers perform CPR after rescuing a fire victim during a simulation performed in the Public Safety Education Department’s burn tower.


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The US Dollar has trimmed its rise against a basket of its peers since President Donald Trump began his speech to Congress, which has so far focused heavily on border protections and more constrained immigration policies.
The dollar index had risen to 101.5 in the run up to Mr Trump’s speech but has since fallen back to 101.27.
Mr Trump’s speech focused on immigration restrictions early on, also hinting at higher import taxes. There were strong protectionist overtones, focused on “making it easier for companies to do business in the US and much harder for companies to leave our country.”
S&P 500 e-mini futures rose before Mr Trump began speaking and have eased off slightly to 2,369. Benchmark 10-year US Treasury yields have fallen from their highs leading up to the speech, now at 2.4 per cent.
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Mercedes GT / GT S Akrapovic Titanium Evolution Line Exhaust System
|Price:||CALL FOR PRICING|
Perfectly formed using specially developed ultra-lightweight titanium alloys and finished with hand-crafted matte carbon-fibre tailpipes. The system replaces the performance exhausts fitted to the Mercedes-AMG GT and GT S. Because it is ECE type approved with no need to remap the ECU or add any other parts, it is just a simple plug-and-play installation. The exquisite Akrapovic Evolution Line (Titanium) exhaust system adds performance and style, along with an amazing soundtrack.This stunning system includes parts cast in the company’s in-house foundry, featuring an active X connection and an additional pair of exhaust valves behind the rear mufflers, designed to provide optimal control of the exhaust sound for the best aural enjoyment.
Because of packaging requirements, the EVO system has been split into two codes. Please be aware that both part numbers (MTP-ME/T/4H and E-ME/T/4) must be ordered.
We accept no compromises! The company’s vision was clear from the very beginning, and we have not faltered. What started as an innovative,
driven group of six, is now a modern prominent company of more than 13,000 m2 of production and commercial space and with 450 employees, all of us aware that we produce exhaust systems for customers who simply do not accept mistakes – the most demanding racing teams and riders from around the world.
Only the best is good enough for us. But the best is not just the exclusive privilege of the factory teams. Now it is available to everyone!
An exhaust system looks simple at first glance. But a closer look shows that it is a complex system that has a significant effect on the performance of a racing or a regular sports car.
The experience in racing that we acquired over many years, taught us how to combine high performance, a long-life cycle and the lightest possible weight. Our numerous world championship titles and countless international and national titles are a testament to our continuous ability to fulfil our clients’ highest expectations.
The titanium we use for our exhaust systems is produced exclusively for us. It is three times stronger and is more heat resistant than any other titanium product on the market and 40% lighter than stainless steel. The high strength allows us to use thinner insulation while still achieving the highest quality standards. In fact, this brings our clients the decisive advantage on the race track.
- Performance and quality
Our goal is to make the perfect torque and the perfect power curve. Our research and development department is provided with the latest equipment, so that they are able to combine their collective know-how obtained at many race tracks and even Formula 1, and apply it in practice.
Please note that certain aftermarket exhaust systems may not comply with applicable California laws and regulations, and may therefore be prohibited for use on California highways or roads, or on roads or vehicles otherwise subject to emissions control requirements.
Akrapovi? exhaust systems for automobiles and motorcycles mounted downstream of the catalytic converter (also known as “cat-back systems”) are considered “replacement parts” in California by the California Air Resources Board (CARB), and do not require an exemption or executive order from CARB to be sold in California. However, California prohibits the use of any aftermarket exhaust system that modifies, removes or replaces original equipment catalysts, unless CARB has issued an Executive Order as to such part or system.
Further, Akrapovi? parts or exhaust systems used or intended for use on “racing vehicles” (i.e. a competition vehicle used exclusively for competition on closed-course circuits) do not require an exemption or Executive Order from CARB to be sold in California. However, such parts are prohibited from use on California public highways or roads, even if occasionally used “off-road.”??
|Wheels And Tires||+Service & Maintenance||Suspension Upgrades||Drivetrain Upgrades||Brake Upgrades||Interior Upgrades|
|Exhaust Upgrades||Engine Upgrades||Exterior Upgrades||Electronic Upgrades||Lifestyle||Installations Overview|


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Calcium supplements, taken with or without vitamin D, may increase the risk of polyps growing in the colon, suggest researchers following a large US trial.
As a result, they suggested that patients with a history of pre-cancerous serrated polyps, especially women and smokers, may wish to avoid vitamin D and calcium supplementation.
They noted, however, that further studies were recommended to confirm their results and any possible risks must be weighed against the benefits of supplementation.
They highlighted that some previous studies have suggested that calcium and vitamin D may protect against colon polyps, but results have been mixed.
To investigate further, the researchers set out to determine whether taking daily calcium and vitamin D supplements reduce the risk of serrated polyps.
They noted that serrated polyps were less common than conventional “adenomatous” polyps, but probably had the same risk of developing into cancer.
They analysed findings from a large US trial involving over 2,000 patients aged between 45 and 75 who had a history of polyps and were due to have a follow-up colonoscopsy in three to five years.
Patients were excluded if they had a family history of bowel cancer, inflammatory bowel disease, or other serious health conditions, and several factors were taken into account at the start of the study.
The remaining patients were randomly split into groups to receive either daily calcium supplements, daily vitamin D supplements, both or neither for three or five years until their colonoscopsy.
During the three to five year treatment phase of the trial, there was no effect of either calcium or vitamin D on cases of serrated polyps, according to the researchers.
However, six to 10 years after treatment began, they found increased risks of serrated polyps among patients taking calcium alone and among those taking a combination of calcium and vitamin D.
There was evidence that women and smokers were at higher risk when exposed to calcium supplements, but no association was found between vitamin D alone and the risk of serrated polyps.
Extra vitamin D for babies and pregnant women in Scotland
The results also suggest an association with calcium supplements only, not dietary calcium, said the researchers from the University of North Carolina School of Medicine.
Further research was recommended to confirm their results, which they described as “surprising”, said the study authors in the journal Gut.
But they concluded that there would be “important public health implications” if calcium and its combination with vitamin D were truly associated with an increased risk of serrated polyps.
They highlighted that calcium supplements were taken by millions of people around the world and the findings may have important implications for bowel cancer screening and prevention.
In the meantime, they suggested that patients with a history of pre-cancerous serrated polyps, especially women and smokers, may wish to avoid vitamin D and calcium supplementation.


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Flow City and Broadsign today announced an AdTECH integration that makes premium global digital-out-of-home screens available for purchase programmatically on Flow City’s DOOH-specialty demand-side-platform.
The collaboration allows media buying agencies, small businesses, and startups using Flow City to plan, purchase, and execute global DOOH campaigns with optimal visibility and exposure across new inventory located in shopping centers, supermarkets, and other high-traffic areas in the UK, France, and the US.
This includes premium inventory from Clear Channel available on more than 4,000 screens in the UK alone through their LaunchPAD programmatic offering.
Dagmara Lacka, CEO, Flow City told us “Broadsign is a DOOH leader and understands the intricacies involved in the delivery of compelling programmatic DOOH ads. Naturally, we’re thrilled to team up with them to bring our clients the unique screens and real-time reporting the Broadsign supply-side-platform (SSP) offers. The collaboration unlocks a massive pool of highly coveted DOOH inventory previously unavailable to our clients, including a large volume of desirable screens in the UK, Europe, and the US that will open up new commercial opportunities for our business to grow by nearly thirty per cent.”
We understand that Flow City clients have already started leveraging the integration to deliver programmatic DOOH campaigns promoting commercial office space, consumer goods, and luxury offerings from private jets to perfumes, cosmetics, high-end couture, private culinary services, and beyond.
Dom Dunne, Clear Channel Europe Programmatic Commercial Lead told us “Clear Channel LaunchPAD is an easy way for big brands and startups alike to engage with consumers through premium DOOH,” says Providing convenience, flexibility and scale, we know this enhancement to campaign planning and execution is a welcome addition to new and existing clients on the Flow City platform across the globe.”
Flow City helps brands take their product to the real world using digital billboards. Brands use the platform to plan and execute their growth – both on hyperlocal, national and international level. Unique sets of algorithms built especially for tactical and hyperlocal planning offer great budget efficiency.

domain: geography


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KoFIU investigation revealed that no major crypto exchanges have unlawfully issued native tokens.
Korea Financial Intelligence Unit (KoFIU), a South Korean governmental agency aiming to prevent money laundering and illegal fund flow, including terrorist financing, has rolled out an investigation into crypto exchanges and the issuance of their in-house native tokens.
According to the report written by the local news portal Korea JoongAng Daily, the investigation was started in light of the FTX collapse. On November 11th, FTX filed for Chapter 11 bankruptcy due to a price crash of its native token, FTT.
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The Act on Reporting and Using Specified Financial Transaction Information in South Korea notes that crypto exchanges are not allowed to issue native tokens, and sell, exchange, or mediate “coins listed by people related to the exchange.”
Despite the law, Korea Financial Intelligence Unit seeks to explore all possibilities.
The news report highlighted that the first round of investigations revealed that crypto exchanges are obeying the law. However, the Financial Services Commission (FSC) does not rule out the possibility of extending the probe as “there are still some doubts related” to the issuance of native tokens.
One of the companies on the KoFIU radar is a Daegu-based crypto exchange, Flata Exchange. Local news portal Yonhap reported that KoFIU aims to look into the issuance of Flat, which was first listed in January 2020, and identify whether it is a native token or not.
It is worth noting that KoFIU emphasized that major South Korean crypto exchanges, like Upbit and Bithumb, haven’t issued native tokens. However, the authority highlighted that smaller crypto exchanges operating in the country are yet to be investigated.
South Korean authorities are worried about the impact FTX collapse may have on domestic investors. Local news reports revealed that around 6,000 Korean investors are holding FTT. The total amount of FTT in Korean investors’ wallets is approximately 110,000 units.
Moreover, a separate report revealed that every month, FTX welcomed around 297,229 unique South Korean users. Therefore, it is among the three countries most impacted by FTX collapse.

domain: economics


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My last wedding of the year was the most darling and intimate event. It was in a small town surrounded by farmland and charming people. Aaron + Kendall are so absolutely adorable together, my heart could barely handle it. And being a photographer himself, Aaron knew the importance of bride and groom portraits which made my job so much easier. I'm pretty sure their pictures are some of my favorites.
Congrats you two and thanks for dodging cow pies to get those perfect shots. You guys are the best!