Higher Methylation Research Articles

Selective adsorption of unmethylated DNA on ZnO nanowires for separation of methylated DNA.

DNA methylation is a crucial epigenetic modification used as a biomarker for early cancer progression. However, existing methods for DNA methylation analysis are complex, time-consuming, and prone to DNA degradation. This work demonstrates selective capture of unmethylated DNAs using ZnO nanowires without chemical or biological modifications, thereby concentrating methylated DNA, particularly those with high methylation levels that can predict cancer risk. We observe varying affinities between methylated and unmethylated DNA on ZnO nanowires, which may be influenced by differences in hydrogen bonding strength, potentially related to the effects of methylation on DNA strand behavior, including self-aggregation and stretching inhibition. As a result, the nanowire-based microfluidic device effectively collects unmethylated DNA, leading to a significantly increased ratio of methylated to unmethylated DNA, particularly for collecting low-concentration methylated DNA. This simplified microfluidic device, composed of ZnO nanowires, enables direct separation of specific methylated DNA, offering a potential approach for DNA methylation mapping in clinical disease diagnostics.

Extensive methylation analysis of circulating tumor DNA in plasma of patients with gastric cancer

DNA methylation is known to be involved in tumor progression. This is the first study to perform an extensive methylation analysis of plasma circulating tumor DNA (ctDNA) using targeted bisulfite sequencing in gastric cancer (GC) patients to evaluate the usefulness of ctDNA methylation as a new biomarker. Sixteen patients who received chemotherapy for recurrent GC were included. After confirmation of the methylation status of 63 genes using the Cancer Genome Atlas (TCGA) dataset, the methylation status in paired tumor and non-tumor tissues and plasma were investigated using targeted bisulfite sequencing in these genes. Forty-four of the 63 genes were significantly hypermethylated in GC patients in the TCGA cohort. Of these 44 genes, hierarchical clustering showed that five (SPG20, FBN1, SDC2, TFPI2, SEPT9) were particularly hypermethylated in tumor compared to non-tumor tissues in our GC cohort. In plasma methylation analysis, patients with high methylation of these genes had significantly worse overall survival than those with low methylation (log-rank P = 0.009). In a patient who underwent blood sampling at multiple points, the methylation levels of these five genes varied closely with clinical tumor status. The plasma ctDNA methylation levels of these five genes could be useful as a noninvasive prognostic biomarker for GC.

Improving Process Design for Reaching Energy Efficiency, Environmentally Friendly, and Producing High Purity Methyl Chloride of Dehydrochlorination Process of Methanol and Hydrogen Chloride

Methyl chloride, also known as chloromethane, plays a vital role in producing various industrial goods. The current demand for methyl chloride in Indonesia exceeds production levels, making the design of a methyl chloride plant essential. This research focuses on improving methyl chloride production economically and operationally by exploring plant design using simulations that emphasize energy efficiency and high purity. The objective of this research is to develop a process design for producing methyl chloride from methanol and hydrogen chloride, aiming for energy efficiency, an environmentally friendly factory, and high-purity methyl chloride products. The research employed an iterative simulation method to compare the basic and modified processes for methyl chloride production. The process involved constructing a simulation model using Aspen HYSYS, analyzing the simulation results using Aspen Energy Analyzer V12, and iteratively adjusting process parameters until achieving the desired performance or results. The research findings indicate that the methyl chloride modification process exhibits a lower energy requirement compared to the methyl chloride base process. Moreover, the modification process demonstrates minimal carbon emissions, establishing it as a sustainable and environmentally friendly design. Additionally, the methyl chloride produced in the modification process achieves a higher percentage of purity. In the initial process, the methyl chloride purity stood at 98.17%, while in the modified process, it saw an elevation to 99.35%. Considering these three aspects, the modification process is conclusively more efficient than the basic process system. Copyright © 2024 by Authors, Published by Universitas Diponegoro and BCREC Publishing Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).

Expressions of SHOX2, RASSF1A and PTGER4, and the relationship between their methylation and clinicopathological characteristics in patients with lung cancer

Background: To explore the expressions of short stature homobox2 (SHOX2), Ras-association domain family 1A (RASSF1A) and prostaglandin E receptor 4 (PTGER4), and the relationship between their methylation and clinicopathological characteristics in patients with lung cancer (LC). Methods: The surgical specimens of cancer tissues and para-carcinoma tissues were collected from 50 patients with LC in the Affiliated Hospital of Hebei University of Engineering between January and November 2023. The expressions of SHOX, RASSF1A and PTGER4 proteins in cancer tissues and para-carcinoma tissues were detected by immunohistochemistry, and methylation status of SHOX, RASSF1A and PTGER4 genes in peripheral venous blood was detected by sulfite-modified real-time fluorescence quantification. The positive expression rates of SHOX2, RASSF1A and PTGER4, and positive rates of SHOX2, RASSF1A and PTGER4 genes methylation in cancer tissues and para-carcinoma tissues were compared. The relationship between SHOX2, RASSF1A, PTGER4 methylation and clinicopathological characteristics in LC patients was compared by real-time fluorescence quantitative PCR. Results: The positive expression rates of SHOX2, RASSF1A and PTGER4 in cancer tissues were 44.0%, 54.00% and 50.00%, significantly lower than those in para-carcinoma tissues (90.00%, 96.00%, 82.00%, P<0.05). The methylation positive rates of SHOX2, RASSF1A and PTGER4 genes in cancer tissues were 48.00%, 32.00% and 64.00%, significantly higher than those in para-carcinoma tissues (16.00%, 4.0%, 14.00%, P<0.05). In terms of different pathological types and TNM staging, methylation positive rates of SHOX2 and RASSF1A genes were the highest in patients with adenocarcinoma and TNM staging at stage IV. In terms of different pathological types, methylation positive rate of PTGER4 gene was the highest in patients with TNM staging at stage IV (P<0.05). Conclusion: The low expressions and high methylation of SHOX2, RASSF1A and PTGER4 are common in LC patients. The methylation positive rates of SHOX2 and RASSF1A genes are related to pathological types and TNM staging, and methylation positive rate of PTGER4 gene is only related to pathological types.

Blood methylation biomarkers are associated with diabetic kidney disease progression in type 1 diabetes.

DNA methylation differences are associated with kidney function and diabetic kidney disease (DKD), but prospective studies are scarce. Therefore, we aimed to study DNA methylation in a prospective setting in the Finnish Diabetic Nephropathy Study type 1 diabetes (T1D) cohort. We analysed baseline blood sample-derived DNA methylation (Illumina's EPIC array) of 403 individuals with normal albumin excretion rate (early progression group) and 373 individuals with severe albuminuria (late progression group) and followed-up their DKD progression defined as decrease in eGFR to <60 mL/min/1.73m 2 (early DKD progression group; median follow-up 13.1 years) or end-stage kidney disease (ESKD) (late DKD progression group; median follow-up 8.4 years). We conducted two epigenome-wide association studies (EWASs) on DKD progression and sought methylation quantitative trait loci (meQTLs) for the lead CpGs to estimate genetic contribution. Altogether, 14 methylation sites were associated with DKD progression ( P <9.4×10 -8 ). Methylation at cg01730944 near CDKN1C and at other CpGs associated with early DKD progression were not correlated with baseline eGFR, whereas late progression CpGs were strongly associated. Importantly, 13 of 14 CpGs could be linked to a gene showing differential expression in DKD or chronic kidney disease. Higher methylation at the lead CpG cg17944885, a frequent finding in eGFR EWASs, was associated with ESKD risk (HR [95% CI] = 2.15 [1.79, 2.58]). Additionally, we replicated meQTLs for cg17944885 and identified ten novel meQTL variants for other CpGs. Furthermore, survival models including the significant CpG sites showed increased predictive performance on top of clinical risk factors. Our EWAS on early DKD progression identified a podocyte-specific CDKN1C locus. EWAS on late progression proposed novel CpGs for ESKD risk and confirmed previously known sites for kidney function. Since DNA methylation signals could improve disease course prediction, a combination of blood-derived methylation sites could serve as a potential prognostic biomarker.

Gremlin-2 is a novel tumor suppressor that negatively regulates ID1 in breast cancer

BackgroundBreast cancer is one of the most common cancers in women and is closely associated with obesity. Gremlin-2 (GREM2), an antagonist for bone morphogenetic proteins (BMPs), has been considered an inhibitor of adipogenic differentiation in adipose-derived stromal/stem cells. However, the role of GREM2 in breast cancer cells remains largely unknown, and its signaling mechanism has yet to be clarified.MethodsBioinformatics analysis was conducted using public databases. Breast cancer cells overexpressing mock or GREM2 were used for in vitro and in vivo studies. Cell viability, colony formation, migration, and animal studies were performed to investigate the role of GREM2 in breast cancer cells. Screening of target genes affected by GREM2 overexpression in breast cancer cells was performed through RNA sequencing (RNA-seq) analysis.ResultsThe expression level of GREM2 mRNA was significantly reduced in both breast cancer tissues and cell lines. Kaplan-Meier analysis showed that low expression of GREM2 and high methylation of the GREM2 promoter were each associated with poor patient survival. The low mRNA expression of GREM2 in breast cancer cells was increased by the demethylating agent decitabine. Breast cancer cells overexpressing GREM2 decreased cell proliferation when compared to control cells, both in vitro and in vivo. Through comparison of RNA-seq analysis between cell lines and tissue samples, gene ontologies that were consistently upregulated or downregulated by GREM2 in breast cancer were identified. In particular, the expression of inhibitor of DNA-binding-1 (ID1) was repressed by GREM2. BMP2 is one of the upstream regulators that increases the expression of ID1, and the expression of ID1 reduced by GREM2 was restored by overexpression of BMP2. Also, the migration ability of breast cancer cells, which had been suppressed by GREM2, was restored by BMP2 or ID1.ConclusionsLow expression of GREM2 in breast cancer cells is associated with hypermethylation of the GREM2 promoter, which may ultimately contribute to poor patient survival. GREM2 participates in regulating the expression of various genes, including ID1, and is involved in suppressing the proliferation of breast cancer cells. This suggests that GREM2 has the potential to act as a novel tumor suppressor in breast cancer.

A two-site combined prediction model based on HOXA9 DNA methylation for early screening of risks of meningioma progression

To establish a recurrence risk prediction model for meningioma based on HOXA9 DNA methylation. Meningioma-related datasets were downloaded from GEO database for screening homeobox genes (HOXs) with prognostic values using differential methylation and ROC curve analysis and Cox regression analysis. The differentially methylated CpG sites with high predictive efficacy were selected to establish the risk prediction model using Lasso-Cox regression analysis, based on which the patients were divided into high- and low-risk groups by the cutoff value. The methylation levels of CpG sites were verified at the cell and tissue levels using methylation-specific PCR (MS-PCR). Clinical meningioma tissue samples were used to validate the predictive efficacy of the model. HOXA9 methylation level was significantly up-regulated in meningiomas (P< 0.001) and showed a high diagnostic efficiency (AUC=0.884) as an independent risk factor for overall survival (P< 0.01) positively correlated with the degree of malignancy and poor prognosis of meningioma (P< 0.05). Risk stratification by HOXA9 methylation was more accurate than WHO grading for predicting recurrence and patient survival time. The AUCs of the sites cg03217995 and cg21001184 were both above 0.8 for meningioma diagnosis and above 0.6 for predicting recurrence. The patients' clinical characteristics differed significantly between the high- and low-risk groups (P< 0.001), and the prediction score of the model was an independent prognostic factor for meningioma (P< 0.05). MSPCR results showed that the methylation levels of the two sites increased significantly in meningioma cells. In clinical samples, the combined model showed a high prediction efficiency (AUC=0.857), and the predicted risk of progression was highly consistent with the patients' actual condition. High HOXA9 methylation level is a predictor for poor prognosis of meningiomas, and the combined prediction model based on its CpG sites provides a new approach to early screening of meningioma patients at risk of progression.

Reduced expression of FOXE1 in differentiated thyroid cancer, the contribution of CPG methylation, and their clinical relevance.

Forkhead box E1 (FOXE1) is a transcription factor with a crucial role in thyroid morphogenesis and differentiation. Promoter hypermethylation downregulates FOXE1 expression in different tumor types; nevertheless, its expression and relationship with methylation status in differentiated thyroid cancer (DTC) remain unclear. A total of 33 pairs of matched samples of PTC tumors and non-tumors were included. Tumor cell cultures were treated with either 5-Aza-2'-deoxycytidine demethylating agent or dimethyl sulfoxide (DMSO). A real-time polymerase chain reaction (RT-PCR) and Western blotting were performed to assess FOXE1 expression. The methylation status was quantified using bisulfite sequencing. A luciferase gene assay was used to determine CpG-island functionality. Gene expression and promoter methylation of FOXE1 and FOXE1-regulated genes were also analyzed with data from The Cancer Genome Atlas (TCGA) thyroid samples. After demethylating treatment, increased FOXE1 mRNA was observed concomitantly with reduced promoter methylation of CpGisland2. A negative correlation between mRNA downregulation and an increased methylation level of CpGisland2 was observed in tumors. Diminished protein expression was also detected in some DTC cell lines and in some tumor samples, suggesting the involvement of post-transcriptional regulatory mechanisms. CPGisland2 was proved to be an enhancer. TCGA data analysis showed low FOXE1 mRNA expression in tumors with a negative correlation with methylation status and a positive correlation with the expression of most of its target genes. Reduced FOXE1 expression, accompanied by a high methylation level, was associated with PTC aggressiveness (tall cell variant, advanced extra thyroid extension, T4 American Joint Committee on Cancer (AJCC) classification), age at diagnosis (over 45 years old), and presence of a BRAFV600E mutation. FOXE1 mRNA was downregulated in DTC compared with non-tumors, followed by high CpGisland methylation. A coupling of low mRNA expression and high methylation status was related to characteristics of aggressiveness in DTC tumors.

Methylation modification is a poor prognostic factor in non-small cell lung Cancer and regulates the tumor microenvironment: mRNA molecular structure and function

Non-small cell lung cancer (NSCLC) is the most common and deadly type of lung cancer, and its poor prognosis is closely related to the complex interactions of the tumor microenvironment. Through methylation analysis of tumor tissue samples from NSCLC patients, combined with high-throughput sequencing technology, the methylation status and structural characteristics of mRNA molecules were studied. Bioinformatics tools were used to analyze the regulatory effects of methylation modification on mRNA expression and genes associated with the tumor microenvironment. The results showed that the methylation level of specific mRNA was significantly correlated with the expression changes of tumor microenvironment-related factors. In addition, methylation modification affected mRNA stability and translation efficiency, further altering the metabolic activity and immune escape capacity of tumor cells. The results showed that mRNA with high methylation level was significantly associated with poor prognosis. Methylation modification profoundly affects the tumor microenvironment and prognosis of non-small cell lung cancer by altering the structure and function of mRNA molecules.

The telomere-to-telomere gap-free genome assembly of Juglans sigillata

Juglans sigillata is an economically valuable nut crop renowned for its nutritional richness, including essential nutrients, antioxidants, and healthy fats, which boost human cardial, brain and gut health. Despite its importance, the lack of a complete genome assembly has been a stumbling block in its biological breeding process. Therefore, we generated deep coverage ultralong Oxford Nanopore Technology (ONT) and PacBio HiFi reads to construct a telomere-to-telomere (T2T) genome assembly. The final assembly spans 537.27 Mb with no gaps, demonstrating a remarkable completeness of 98.1 %. We utilized a combination of transcriptome data and homologous proteins to annotate the genome, identifying 36 018 protein-coding genes. Furthermore, we profiled global cytosine DNA methylations using ONT sequencing data. Global methylome analysis revealed high methylation levels in transposable element (TE)-rich chromosomal regions juxtaposed with comparatively lower methylation in gene-rich areas. By integrating a detailed multi-omics data analysis, we obtained valuable insights into the mechanism underlying endopleura coloration. This investigation led to the identification of eight candidate genes (e.g. ANR) involved in anthocyanin biosynthesis pathways, which are crucial for the development of color in plants. The comprehensive genome assembly and the understanding of the genetic basis of important traits like endopleura coloration will open avenues for more efficient breeding programs and improved crop quality.

Reductive dehalogenase of Dehalococcoides mccartyi strain CBDB1 reduces cobalt- containing metal complexes enabling anodic respiration.

Microorganisms capable of direct or mediated extracellular electron transfer (EET) have garnered significant attention for their various biotechnological applications, such as bioremediation, metal recovery, wastewater treatment, energy generation in microbial fuel cells, and microbial or enzymatic electrosynthesis. One microorganism of particular interest is the organohalide-respiring bacterium Dehalococcoides mccartyi strain CBDB1, known for its ability to reductively dehalogenate toxic and persistent halogenated organic compounds through organohalide respiration (OHR), using halogenated organics as terminal electron acceptors. A membrane-bound OHR protein complex couples electron transfer to proton translocation across the membrane, generating a proton motive force, which enables metabolism and proliferation. In this study we show that the halogenated compounds can be replaced with redox mediators that can putatively shuttle electrons between the OHR complex and the anode, coupling D. mccartyi cells to an electrode via mediated EET. We identified cobalt-containing metal complexes, referred to as cobalt chelates, as promising mediators using a photometric high throughput methyl viologen-based enzyme activity assay. Through various biochemical approaches, we show that cobalt chelates are specifically reduced by CBDB1 cells, putatively by the reductive dehalogenase subunit (RdhA) of the OHR complex. Using cyclic voltammetry, we also demonstrate that cobalt chelates exchange electrons with a gold electrode, making them promising candidates for bioelectrochemical cultivation. Furthermore, using the AlphaFold 2-calculated RdhA structure and molecular docking, we found that one of the identified cobalt chelates exhibits favorable binding to RdhA, with a binding energy of approximately -28 kJ mol-1. Taken together, our results indicate that bioelectrochemical cultivation of D. mccartyi with cobalt chelates as anode mediators, instead of toxic halogenated compounds, is feasible, which opens new perspectives for bioremediation and other biotechnological applications of strain CBDB1.

Hypermethylation of the sodium channel beta subunit gene promoter is associated with colorectal cancer

AimsTo better understand the role of sodium channel beta subunit (SCNN1B) in the initiation and progression of colorectal cancer (CRC) and to identify potential biomarkers for the early detection and prognosis of CRC.MethodsA total of 74 pairs of CRC tissues and their adjacent normal tissues were collected between October 2016 and November 2017. The methylation levels of the SCNN1B promoter region in CRC tissues and their adjacent normal tissues were investigated by pyrosequencing. The expression of both SCNN1B mRNA and protein were detected by RT‒qPCR and immunohistochemistry, respectively.ResultsThe results showed that the methylation levels of the SCNN1B promoter region were significantly higher in CRC tissues than in adjacent normal tissues. The expression levels of SCNN1B mRNA and protein were significantly lower in the CRC tissues than in their adjacent normal tissues. Moreover, Pearson’s correlation analysis showed that the methylation levels of the SCNN1B promoter were negatively correlated with the SCNN1B mRNA levels in CRC tissues. In addition, the high methylation levels and low mRNA expression of SCNN1B showed a significant association with advanced tumour stage, increased risk of lymph node metastasis and poor prognosis of CRC patients.ConclusionThis study suggested that the decreased expression of SCNN1B due to its promoter hypermethylation may play an important role in the progression and prognosis of CRC, and the methylation levels of the SCNN1B promoter may serve as an effective molecular marker for predicting the progression and prognosis of CRC.

Unexplained high and persistent methyl bromide emissions in China

Methyl bromide (CH3Br) is an important ozone-depleting substance whose use is regulated under the Montreal Protocol. Quantifying emissions on the national scale is required to assess compliance with the Montreal Protocol and thereby ensure the timely recovery of the ozone layer. However, the spatial-temporal patterns of China’s national CH3Br emissions remain unclear. Here we estimate the national emissions of CH3Br in China during 2011−2020 using atmospheric observations at 10 sites across China combined with an inversion technique (top-down) and compare those with an updated inventory of identified emission sources (bottom-up). Measured CH3Br mole fractions are enhanced well above the background mole fractions, especially at sites in eastern China. Top-down emission estimates exceed bottom-up estimates by 5.5 ± 1.4 gigagrams per year, with the largest fraction (60%) of observationally derived CH3Br emissions arising from underestimated or unidentified emissions sources. This study shows the potential impacts of the unaccounted emissions on stratospheric ozone depletion, with implications for the Montreal Protocol.

In-situ water separation enhanced methyl glycolate oxidation to methyl glyoxylate by catalytic membrane reactor

The selective oxidation of methyl glycolate by oxygen is a green route for the synthesis of methyl glyoxylate, which however faces great challenges like the consecutive hydrolysis reaction between methyl glyoxylate and by-product water. Herein, we fabricated an in-situ water separation catalytic membrane reactor (CMR) to suppress the hydrolysis of methyl glyoxylate, which can promptly and preferentially remove water from the reaction system by a hydrophilic NaA membrane. The MoO3@NaA CMR achieved a high methyl glyoxylate selectivity of 84.0 % at 260 °C and 2 bar, being 32.4 % higher than that of the conventional fixed-bed reactor (FBR), while the methyl glycolate conversion was almost the same for both modes. Moreover, the deactivation of the MoO3 catalyst caused by water in CMR was also greatly inhibited, exhibiting excellent stability as compared to FBR. The in-situ water separation strategy in this work contributed to a new concept to improve the selectivity of easily hydrolyzable products.

DNA methylation-driven gene FAM3D promotes colorectal cancer growth via the ATF4-SESN2-mTORC1 pathway.

Globally, colorectal cancer (CRC) is the malignant tumor with the highest mortality rate after lung cancer. Abnormal DNA methylation drives dysregulated gene expression, thereby promoting CRC progression and leading to poor prognosis. We identified a 3-CpG methylation signature that is independently associated with CRC prognosis. The model consists of three methylation-driven genes: FAM3 Metabolism Regulating Signaling Molecule D (FAM3D), DAPP1, and PIGR. However, the prognostic significance, biological function, and related mechanisms of the individual methylation-driven gene FAM3D in CRC have not been studied. Here, we discovered that FAM3D expression was reduced in CRC tissues and cells, and that high methylation and low expression of FAM3D were independent prognostic risk factors for CRC. In addition, FAM3D promoted the growth and movement of CRC cells in vitro and the proliferation in nude mice, mainly by inhibiting ATF4 transcription and downregulating SESN2 expression, and ultimately activating mTORC1. Furthermore, FAM3D resulted in reduced sensitivity of CRC cells to oxaliplatin, cisplatin, and 5-fluorouracil. Our study showed that FAM3D activates the mTORC1 pathway through the ATF4-SESN2 axis and promotes the malignant progression of CRC, which contributes to predict CRC prognosis and guide individualized treatment.

8815 ANPEP regulates one-carbon metabolism in prostate cancer

Abstract Disclosure: A. El-Kenawi: None. R. Putney: None. A. Berglund: None. K. Yamoah: None. African American men (AAM) experience disproportionately higher rates of both prostate cancer (PCa) incidence and mortality compared to their European American counterparts (EAM). To investigate some of the biological factors which contribute to these disparities, we carried out an unbiased genomic expression analysis of PCa by ancestry using the matched GRID and VANDAAM cohorts. In this analysis, we discovered aminopeptidase N (ANPEP, APN, CD13), to be the most differentially expressed functional gene in both self-identified and ancestry derived AAM compared with EAM. Further computational analyses revealed that ANPEP correlates with signatures of cholesterol transport, estrogen and androgen receptor (AR) signaling. The functional role of ANPEP in regulating cholesterol metabolism and AR signaling remains unclear. Further computational analyses demonstrate that expression of ANPEP predominantly correlates with various amino acid transporters. In addition, comprehensive metabolomic characterization revealed that ANPEP-overexpressing cells exhibit altered level of one-carbon metabolism metabolites. The one-carbon metabolic pathway regulates amino acids homeostasis and epigenetic mechanisms. Metabolomics-based approach to assess the consumption of over 15 amino acids in patient-derived explants validate enrichment of one-carbon metabolism circuits in AAM. Thus, we further investigated whether ANPEP impact high methylation capacity by performing genome-wide methylation profiling in cells. We unraveled that ANPEP overexpression is associated with elevated methylation of DNA. Our future studies will assess whether ANPEP regulation of methylation capacity have an impact on AR binding to DNA. Presentation: 6/3/2024

Abstract A060: Breast cancer-associated leukocyte DNA methylation profile in obese African-American women

Abstract Background: DNA methylation (DNAme) is a stable epigenetic mark leading to reduced gene expression. However, genome-wide DNAme profile is susceptible to disease. Therefore, specific genetic DNAme profiles could potentially serve as biomarkers associated with breast cancer (BC) and obesity as its comorbidity. Incidentally, BC-related mortality is higher in African-American women, causing a significant health disparity. In this study, we profiled obesity and breast cancer-related leukocyte DNAme in African-American women to explore methylation- associated biomarkers in these women. Methods: We used 10 African-American women with confirmed breast cancer diagnosis and 10 otherwise healthy African-American women with known Body Mass Index (BMI) &amp;gt; 30 (Obese). DNA was extracted from the buffy coat of peripheral blood using Qiagen DNA extraction kits. We used the next-generation sequencing technology to perform reduced presentation bisulfite sequencing (RRBS) using the Tecan Ovation RRBS kit. We sequenced the libraries on a NextSeq 550 at our Precision Medicine Unit. Single-end 75bp reads were generated and processed using Bismark and mapped against the GRCh38 human genome, and differentially methylated regions (DMR) were derived using MethylKit using a 1 kb window. Gene Ontology and Disease association studies were conducted using DAVID and relapsed-free survival in Kmplot. Additionally, we compared the gene sets with obese and normal DMRs that had previously been obtained similarly. Results: We found that 83% of the DMRs were in the promoter/upstream regions of the nearest genes, and 96% of DMRs were part of CpG islands, indicating potential regulatory functions of these regions. Gene ontology analysis revealed that the DMR-associated genes were also overrepresented in breast carcinoma, Type 2 Diabetes, Obesity, and Tobacco usage, according to the DisGeNET and GAD disease associations databases. Genes with DMRs in their upstream or promoter regions - include PRDM16, PTBP1, RAB1A (lower methylation in cancer with obesity), and Multiple ribosomal genes along with RRAF, XKR4 (higher methylation in cancer with obesity). We observed lower methylation profiles in multiple genes in samples with cancer, which were higher in obese samples without cancer.Conclusion: Our study suggests that a distinct peripheral DNA methylation profile is associated with minority women suffering from both cancer and obesity. Our data show that showed that breast cancer modulates leucocyte DNA methylation profile in obese individuals towards that of a more normal BMI, causing a partial reversal. This potential reversal is primarily in the upstream regions of the genes detected for methylation. The DMR genes in cancer with obesity are significantly associated with overall survival in breast cancer, thus indicating a potential link in understanding the crosstalk between obesity and cancer. Citation Format: Pranabananda Dutta, Yanyuan Wu, Jaydutt V. Duta. Breast cancer-associated leukocyte DNA methylation profile in obese African-American women [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr A060.

Selective upcycling of brominated epoxy resin by subcritical water ammonia process with waste copper-based catalyst: Production of high purity methyl pyrimidine/phenols and copper recovery

Tetrabromobisphenol A epoxy resin (TBBPAER) is the main non-metallic component of electronic waste. The scale of electronic waste production and the accompanying TBBPAER waste disposal problems represent a great opportunity for chemical upcycling. But the chemical upcycling of TBBPAER faces great challenges due to its high bromine content and thermally stability. In this study, a subcritical water ammonia (SWA) process combined with waste copper-based catalyst (WCC) selectively converted TBBPAER in high yields (63.46 % at 300°C for 15 min and 73.66 % for 60 min) to low molecular-weight liquid products including high value-added methyl pyrimidine and phenols. The electron transfer among multivalent copper species contained in the WCC promoted the production of free radicals OH, O2−, NH2, and :NH, which resulted in the efficient conversion of TBBPAER and a 99.29 % of debromination ratio. The molecular chain of TBBPAER was snipped by OH and NH2 to produce tetrabromobisphenol A groups (TAG) and ternary carbon groups (TCG). The further degradation of TAG resulted in the producing of phenol chemicals with a purity of 91.8 % (GC peak area%). The further cyclization of TCG induced by :NH produced methyl pyrimidine with a purity of 91.9 % (GC peak area%). The formation of copper ammonia complex led to the leaching/recovery of 86.6 % of copper from the WCC. The SWA-WCC approach demonstrated how TBBPAER waste could be a viable feedstock for the producing of high value-added methyl pyrimidine and phenol chemicals. This study provided a novel sustainable strategy for synchronous treatment/upcycling of the two different wastes of TBBPAER and WCC. The leaching toxicity test of Cu and Zn for the solid residue after the co-treatment showed that their leaching concentrations were much lower than the hazardous waste standard.

Novel epigenetic biomarkers for hematopoietic cancer found in twins.

This article aims to identify epigenetic markers and detect early development of hematopoietic malignancies through an epigenome wide association study of DNA methylation data. This register-based study includes 1,085 Danish twins with 31 hematopoietic malignancies and methylation levels from 450,154 5'-C-phospate-G-3' (CpG) sites. Associations between methylation levels and incidence of hematopoietic malignancy is studied through time-to-event regression. The matched case-cotwin design, where one twin has a malignancy and the cotwin does not, is applied to enhance control for unmeasured shared confounding and false discoveries. Predictive performance is validated in the independent Older Finnish Twin Cohort. We identified 67 epigenetic markers for hematopoietic malignancies of which 12 are linked to genes associated with hematologic malignancies. For some markers, we discovered a 2-3-fold relative risk difference for high versus low methylation. The identification of these 67 sites enabled the formation of a predictor demonstrating a cross-validated time-varying area under the curve (AUC) of 92% 3 years after individual blood sampling and persistent performance above 70% up to 6 years after blood sampling. This predictive performance was to a large extent recovered in the validation sample showing an overall Harrell's C of 73%. In conclusion, from a large population representative twin study on hematopoietic cancers, novel epigenetic markers were identified that may prove useful for early diagnosis.

LTBR acts as a novel immune checkpoint of tumor-associated macrophages for cancer immunotherapy.

Tumor-associated macrophages (TAMs) greatly contribute to immune checkpoint inhibitor (ICI) resistance of cancer. However, its underlying mechanisms and whether TAMs can be promising targets to overcome ICI resistance remain to be unveiled. Through integrative analysis of immune multiomics data and single-cell RNA-seq data (iMOS) in lung adenocarcinoma (LUAD), lymphotoxin β receptor (LTBR) is identified as a potential immune checkpoint of TAMs, whose high expression, duplication, and low methylation are correlated with unfavorable prognosis. Immunofluorescence staining shows that the infiltration of LTBR+ TAMs is associated with LUAD stages, immunotherapy failure, and poor prognosis. Mechanistically, LTΒR maintains immunosuppressive activity and M2 phenotype of TAMs by noncanonical nuclear factor kappa B and Wnt/β-catenin signaling pathways. Macrophage-specific knockout of LTBR hinders tumor growth and prolongs survival time via blocking TAMimmunosuppressive activity and M2 phenotype. Moreover, TAM-targeted delivery of LTΒR small interfering RNA improves the therapeutic effect of ICI via reversing TAM-mediated immunosuppression, such as boosting cytotoxic CD8+ T cells and inhibiting granulocytic myeloid-derived suppressor cells infiltration. Taken together, we bring forth an immune checkpoint discovery pipeline iMOS, identify LTBR as a novel immune checkpoint of TAMs, and propose a new immunotherapy strategy by targeting LTBR+ TAMs.