HIC1 Research Articles

Computational Analysis of Single Nucleotide Polymorphisms in Human HIC1 Gene.

Background A putative tumor suppressor gene called HIC1 (hypermethylated in cancer) is situated at 17p13.3, a locus where the allelic loss occurs often in human malignancies, including breast cancer. Hypermethylated in cancer 1 protein is a protein that in humans is encoded by the HIC1 gene and it's aHomo sapiens(Human). This gene functions as a growth regulatory and tumor repressor gene. The molecular function of HIC1 gene includes DNA-binding transcription factor activity, sequence-specific DNA binding, DNA binding, histone deacetylase binding, protein binding, metal ion binding, nucleic acid binding, DNA-binding transcription repressor activity, RNA polymerase II-specific, DNA-binding transcription factor activity, RNA polymerase II-specific. The biological process of HIC1 gene includes multicellular organism development, negative regulation of Wnt signaling pathway, positive regulation of DNA damage response, signal transduction by p53 class mediator regulation of transcription, DNA-templated, negative regulation of transcription by RNA polymerase II, Wnt signaling pathway, transcription, DNA-templated, intrinsic apoptotic signaling pathway in response to DNA damage, cellular response to DNA damage stimulus. The study aimed to predict the stability and structure of the protein that will arise from single nucleotide polymorphisms (SNPs) in the human HIC1 gene. Methodology To investigate the possible negative effects associated with these SNPs, bioinformatic analysis is typically essential. The following tools were employed for forecasting harmful SNPs: scale-invariant feature transform (SIFT), Protein Analysis Through Evolutionary Relationships (PANTHER), nonsynonymous SNP by Protein Variation Effect Analyzer (PROVEAN), and nonsynonymous SNP by Single Nucleotide Polymorphism Annotation Platform (SNAP). Results The present study identified a total of 36 SNPs using the SIFT approach, which were shown to have functional significance. Twenty-six were determined to be tolerable, whereas 10 were shown to be detrimental. Out of 20 SNPs, seven (P370A, P646S, R654P, A476T, S400S, D666N, D7V) SNPs were predicted as "Possibly damaging" and seven (L9F, G468R, G490R, L482R, S12W, G489D, S12P) were identified as "probably benign", and six (R725G, G620S, A56V, E463D, D394N, L338V) were identified as "probably damaging" according to the predictions made by PANTHER tools. The majority of the pixels on the strip were red, indicating that the gene changes may have dangerous consequences. These results highlight the need for more research to fully comprehend how these mutations affect the hic1 protein's function, which is essential for the emergence of different types of cancer. Conclusion The current research has provided us with essential information about how SNPs might be used as a diagnostic marker for cancer, given that SNPs may be candidates for cellular changes caused by mutations linked to cancer.

Abstract B051: Genome-wide association studies of 26,187 women without ovarian cancer identifies 12 genetic loci of blood CA125

Abstract Background: CA125 is elevated in women with ovarian cancer and is the single best ovarian cancer screening biomarker candidate to date, although two randomized screening trials showed no clinically significant difference in ovarian cancer mortality. Consideration of genetic variants that influence blood CA125 levels in healthy women could improve its performance as an ovarian cancer screening biomarker. Furthermore, while studies have suggested CA125 is associated with ovarian carcinogenesis and progression, its biological role remains unclear. Thus, we aimed to identify genetic variants of CA125 in women without ovarian cancer. Methods: We conducted a genome-wide association analysis to identify common genetic variants associated with CA125 using data on 26,187 women without ovarian cancer, leveraging existing blood CA125 data and genetic data from Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial (PLCO), European Prospective Investigation into Cancer and Nutrition (EPIC), Nurses’ Health Studies (NHS/NHSII), and New England Case Control Study (NEC). All genotype data were inputted using TOPMed. For blood CA125 measurement, PLCO, NHS/NHSII, and NEC used the CA125II assay and EPIC used the MSD assay. We used multivariable linear regression models adjusting for age, study, and population ancestry (top 4 principal components). Analyses were conducted separately by CA125 assay, genotyping platforms, and menopausal status at blood draw. Because epidemiological predictors of CA125 differ by menopausal status, we conducted analyses stratified by menopausal status. When we meta-analyzed these results, there were no significant heterogeneity by menopausal status in majority of the genome-wide significant SNPs. Therefore, we performed a fixed-effects meta-analysis to combine summary statistics across all the results. Results: We identified 12 independent loci associated with CA125 at a genome-wide significance level (p<5 × 10−8), of which one SNP (rs73005869) had been previously linked with CA125 and is located in the MUC16 gene. In our meta-analysis, 7 SNPs were associated with lower and 5 were associated with higher levels of CA125. We discovered SNPs on chromosome 16 in the TET2 gene that were significantly associated with increased CA125 levels. We also discovered SNPs in MSLN, a gene reported to be overexpressed in ovarian tumors that contributes to metastasis in the peritoneal microenvironment, were associated with lower CA125 levels. SNPs in the HIC1 gene on chromosome 17, a tumor suppressor gene whose aberrant methylation has been observed in ovarian tumors, was associated with lower CA125 levels. Conclusion: We identified 12 independent loci associated with blood CA125 levels in women without ovarian cancer, providing novel biological insights on the role of CA125. Accounting for these genetic factors that influence blood CA125 levels could improve the performance of CA125 as ovarian cancer screening biomarker in average risk women, substantially improving survival through earlier detection of this deadly disease. Citation Format: Naoko Sasamoto, Jihye Kim, Constance Turman, Shelley S. Tworoger, Rudolf Kaaks, Renée T. Fortner, Daniel W. Cramer, Kathryn L. Terry, Nicolas Wentzensen, Peter Kraft. Genome-wide association studies of 26,187 women without ovarian cancer identifies 12 genetic loci of blood CA125 [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr B051.

The antileukemic activity of decitabine upon PML/RARA-negative AML blasts is supported by all-trans retinoic acid: in vitro and in vivo evidence for cooperation

The prognosis of AML patients with adverse genetics, such as a complex, monosomal karyotype and TP53 lesions, is still dismal even with standard chemotherapy. DNA-hypomethylating agent monotherapy induces an encouraging response rate in these patients. When combined with decitabine (DAC), all-trans retinoic acid (ATRA) resulted in an improved response rate and longer overall survival in a randomized phase II trial (DECIDER; NCT00867672). The molecular mechanisms governing this in vivo synergism are unclear. We now demonstrate cooperative antileukemic effects of DAC and ATRA on AML cell lines U937 and MOLM-13. By RNA-sequencing, derepression of >1200 commonly regulated transcripts following the dual treatment was observed. Overall chromatin accessibility (interrogated by ATAC-seq) and, in particular, at motifs of retinoic acid response elements were affected by both single-agent DAC and ATRA, and enhanced by the dual treatment. Cooperativity regarding transcriptional induction and chromatin remodeling was demonstrated by interrogating the HIC1, CYP26A1, GBP4, and LYZ genes, in vivo gene derepression by expression studies on peripheral blood blasts from AML patients receiving DAC + ATRA. The two drugs also cooperated in derepression of transposable elements, more effectively in U937 (mutated TP53) than MOLM-13 (intact TP53), resulting in a “viral mimicry” response. In conclusion, we demonstrate that in vitro and in vivo, the antileukemic and gene-derepressive epigenetic activity of DAC is enhanced by ATRA.

The promoter methylation drives down-regulation mode of HIC1 in gastric cancer, its molecular characteristics and downstream functional pathways

Gastric cancer is a common malignant tumor of the gastrointestinal tract with a high incidence and mortality rate. Previous results have suggested that the HIC1 gene might be a tumor suppressor candidate in gastric cancer. However, several critical points need to be elucidated: (1) The correlation of HIC1 promoter methylation with its specific expression level in gastric cancer; (2) The molecular characterization of HIC1 promoter methylation; (3) The possible mechanism by which HIC1 performs its inhibitory role in gastric cancer. To address these questions, we retrieved data from TCGA database to analyze HIC1 promoter methylation levels and transcript expression data, and performed targeted region bisulfite sequencing on three stable HIC1 down-regulated cell lines and normal control cell lines, and performed whole transcriptome and metabolite assays in HIC1 knockout cell lines by CRISPR-Cas9 technique. Results demonstrated that HIC1 promoter hypermethylation might be a crucial driving force leading to its down-regulation in HIC1 expression in gastric cancer. This implicated that promoter CG methylation of HIC1 might play a major role in the development of gastric carcinogenesis. Besides, HIC1 may suppress gastric cancer progression by maintaining the normal cellular metabolism, and inhibiting the mTOR signaling pathway activity.

Prdm12 regulates inhibitory neuron differentiation in mouse embryonal carcinoma cells.

The epigenetic regulatory system significant influences the fate determination of cells during developmental processes. Prdm12 is a transcriptional regulator that modulates gene expression epigenetically. The Prdm12 gene has been shown to be expressed in neural tissues, specifically during development, but its detailed function is not fully understood. This study investigated the function of the Prdm12 gene in P19 mouse embryonic tumor cells as a model for neural differentiation. A decrease in the expression of neuron-specific genes and the alterations of dendrites and axons morphology was confirmed in Prdm12-knockout P19 cells. In addition, almost no astrocytes were generated in Prdm12-knockout P19 cells. Comprehensive gene expression analysis revealed that there was a reduction in the expression of the inhibitory neuron-specific genes Gad1/2 and Glyt2, but not the excitatory neuron-specific gene VGLUT2, in Prdm12-knockout P19 cells. Furthermore, the expression of inhibitory neuron-related factors, Ptf1a, Dbx1, and Gsx1/2, decreased in Prdm12-knockout P19 cells. Gene expression analysis also revealed that the Ptf1a, Hic1, and Foxa1 genes were candidate targets of Prdm12 during neurogenesis. These results suggest that Prdm12 regulates the differentiation of inhibitory neurons and astrocytes by controlling the expression of these genes during the neural differentiation of P19 cells.

GLAD-PCR assay of DNA methylation sites in regulatory regions of some tumor-suppressor genes in breast cancer

Hypermethylation of the RcgY sites is shown for many cancer diseases. such aberrant methylation, suppressing the gene activity, occurs at early stages of carcinogenesis. Recently, using glad-pcR assay, we have detected aberrantly methylated RcgY sites, which can be considered to be epigenetic markers of colorectal, lung, and gastric cancers. in breast cancer, methylation of the regulatory regions of ALX4, BMP2, CCND2, CDH13, CDX1, FOXA1, GALR1, GATA5, GREM1, HIC1, HMX2, HS3ST2, HOXC10, ICAM5, LAMA1, RARB, RASSF1A, RUNX3, RXRG, RYR2, SFRP2, SOX17, TERT, and ZNF613 tumor-suppressor genes is reported. in the present work, we determined aberrantly methylated RcgY sites in the regulatory regions of these genes in dNa preparations from breast cancer tissues. the study of dNa samples from 30 tumor and 22 normal mammary tissue samples demonstrates a high diagnostic potential of selected R(5mc)gY sites in regulatory regions of CCND2, BMP2, GALR1, SOX17, HMX2, and HS3ST2 genes with total index of sensitivity and specificity for R(5mc)gY detection in tumor dNa 90.0 % and 100.0 %, respectively.

DIAPH2, PTPRD and HIC1 Gene Polymorphisms and Laryngeal Cancer Risk.

AIM, DIAPH2, PTPRD and HIC1 are the cell glycoprotein, which play an important role in the occurrence and development of tumors. This study was designed to assess the association between DIAPH2, PTPRD and HIC1 SNPs and laryngeal cancer risk. PATIENTS AND METHODS: This study including 267 patients with histologically confirmed laryngeal cancer and 157 controls. The relationship between genetic variations DIAPH2 (rs6620138), PTPRD (rs3765142) and HIC1 (rs9901806) and the onset of laryngeal cancer were investigated. Statistical analysis to calculate the relationship between DIAPH2, PTPRD and HIC1 genes polymorphism and pathogenesis of laryngeal cancer. RESULTS: The results showed that rs6620138 DIAPH2 polymorphism could increase the onset risk of laryngeal cancer. Statistically significant differences in allele distribution of rs6620138 DIAPH2 and rs9901806 HIC1 in the case and control groups subgroups. CONCLUSIONS: This study results suggested that genetic variation of rs6620138 DIAPH2 polymorphism is related to the susceptibility to laryngeal cancer. Our results provide a basis to begin basic research on the role of DIAPH2 gene in the pathogenesis of laryngeal cancer.

Abstract 480: Aberrant promoter hypermethylation of HIC1 and SOX1 in circulating cell free DNA as a potential biomarker for noninvasive early detection of ovarian cancer

Abstract Background: Aberrant CpG island hypermethylation is an early event during carcinogenesis and a significant mechanism of tumor suppressor gene silencing. Assessment of tumor specific methylation changes in tissue or body fluids might provide insights into disease mechanisms and discovery of biomarkers. Ovarian cancer remains the most lethal gynaecological malignancy. Non-specific symptoms at early stage, presentation of disease at advanced stage and lack of sufficiently sensitive screening modalities have rendered significant challenge to its prompt diagnosis. To date, no single biomarker is able to accurately detect early OC in either tissue or body fluid. Aberrant promoter hypermethylation in cell free tumor-specific DNA can be indicative of highly specific cancer signals. Methods: Herein, we concurrently evaluated the promoter methylation of HIC1 and SOX1 gene in 120 ovarian tissue samples and validated in 70 matched serum cell free DNA of cancerous and non cancerous samples by multiplex MethyLight assay (a high throughput quantitative, TaqMan based real time PCR technique). Additionally, promoter methylation pattern of these genes were validated by clonal bisulfite sequencing. The ROC (Receiver-operator characteristic) curves were constructed to evaluate the diagnostic performances of both individual and combined gene panel. Results: In singleplex assay, high methylation sensitivity and specificity was evident for SOX1 (78.8% and 80.0%, respectively; AUC= 0.88) and HIC1 (80.0% and 77.14%, respectively; AUC=0.85) in tissue samples. In the multiplex assay, when either or both gene promoter showed methylation, the highly sensitivity and specific discrimination of EOC patients from normal controls was observed for SOX1 + HIC1 (83.53% and 88.57%, respectively; AUC=0.89). We identified DNA methylation of HIC1 and SOX1 as best discriminator between cancer and non-neoplastic tissue. Methylated SOX1 and HIC1 occurred in 53.3% and 71.1% of OC serum samples when their promoter methylation was analyzed in singleplex assay (AUC=0.81 and 0.88, respectively). In cell free DNA, this novel panel achieved a sensitivity of 80.0% and a specificity of 96.0% with an area under the ROC curve of 0.93 for discriminating OC samples from normal control. The results of MethyLight were highly concordant with those of clonal bisulfite sequencing. Methylated Individual gene as well as in combined panel exhibited better discriminatory efficiencies to identify ovarian cancer at various stages of disease when analyzed in tissue and serum cell free DNA. Conclusion: We report a QPCR based non-invasive epigenetic biomarker assay with high sensitivity and specificity for OC screening. Our findings also reveal the potential utility of methylation based detection of circulating cell free tumor DNA in clinical management of ovarian cancer. Citation Format: Alka Singh, Manisha Sachan. Aberrant promoter hypermethylation of HIC1 and SOX1 in circulating cell free DNA as a potential biomarker for noninvasive early detection of ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 480.

Aberrant DNA Methylation in Bladder Cancer among Saudi Arabia Population

AbstractTumor biomarkers developed based on the aberrant deoxyribonucleic acid (DNA) methylation patterns in bladder cancer (BC) hold great promise due to their stability, specificity, and known associations with the disease. No study has investigated DNA methylation patterns in BC patients from Saudi population. We analyzed DNA methylation levels of 48 tumor suppressor genes loci in 24 bladder tissues (19 BC and 5 control samples) using Human Tumour Suppressor Genes EpiTect Methyl II Complete PCR Array (Qiagen, Hilden, Germany). We identified significant difference in DNA hypermethylation levels at E2F1, ERBB2, HIC1, OPCML, SFN, SFRP1, SFRP2, SPARC, and TERT gene loci between controls and cancerous samples. SCGB3A1 was differentially methylated in nonmuscle invasive versus muscle invasive BC samples. Results suggest that these aberrant DNA methylation patterns in BC are disease and population specific and can be developed as distinct DNA methylation-based biomarkers for BC detection.

Association between Gene Promoter Methylation and Cervical Cancer Development: Global Distribution and A Meta-analysis.

DNA methylation is the main epigenetic event for gene silencing and is associated with carcinogenesis. In this meta-analysis, we evaluated the association between the methylation of the promoter regions of APC, CADM1, CCNA1, CDH1, DAPK, FHIT, HIC1, MAL, MGMT, hMLH1, P16, PAX1, RAR-β, and RASSF1 genes and the risk of cervical cancer development and progression. Overall, 194 eligible studies were identified assessing the associations of promoter methylation status of aforementioned genes with low- and high-grade squamous intraepithelial lesions (LSIL and HSIL) and cervical cancer development. The majority of studies were conducted on Caucasian and Asian populations, whereas rare studies were available on the African population. Promoter methylation frequencies were shown to be significantly higher in LSIL and HSIL cervical cancer cases as compared to control specimens for CADM1, CCNA1, CDH1, DAPK1, FHIT, MAL, P16, PAX1, RAR-β, and RASSF1 genes. A moderate association was found between HIC promoter methylation, whereas APC, MGMT, and hMLH1 promoter methylation was not correlated with cervical cancer development. Promoter methylation could be considered as a noninvasive biomarker for early cervical lesions, making them highly promising targets for a personalized therapeutic approach.

ZBTB gene expression in HIV patients: a possible new molecular mechanism of viral control.

HIV infects its target cell and integrates into its genome as an essential step in its replication cycle. Proviral DNA is also subjected to the same transcriptional regulation as the host cell genome by its own transcriptional factors, with activating or repressive activity. There is a clear interaction between the presence of transcriptional repressors and a decrease in the rate of HIV replication, promoting gene silencing in infected cells, which serve as viral reservoirs. This represents a major obstacle for HIV eradication. The ZBTB gene family comprises 49 genes that encode transcription factors that have a repressor function in differentiation and development of cells of the lymphopoietic lineage, including the main target cells of HIV, CD4+ T cells. In this cross-sectional study, we evaluated the expression profile of ZBTB genes in CD4+ T cells of HIV-positive individuals with different levels of infection control. We found upregulation of gene expression of ZBTB4 (p < 0.01), ZBTB7B (p < 0.001), and ZBTB38 (p < 0.05) and downregulation of ZBTB16 (p < 0.01) in HIV-positive patients compared to HIV-negative individuals. Interestingly, in a deeper analysis, we observed that elite controllers had the highest levels of expression of the ZBTB38, ZBTB2, HIC1, ZBTB7A, ZBTB7B (ThPOK) and ZBTB4 genes, showing 2.56- to 7.60-fold upregulation compare to the ART-naïve group. These results suggest a possible contribution of these ZBTB transcriptional repressors in HIV-positive patients and a possible new molecular mechanism of viral control.

Expression and Clinical Significance of Computer-aided HIC-1 in Colon Cancer

Objective: To investigate the expression and clinical significance of HIC-1 in colon cancer. Methods: RT-PCR, immunohistochemistry, and computer image analysis were used to detect the expression levels of HIC-1 gene and protein in 200 patients with colon cancer and the corresponding paracancerous tissues to analyze the relationship between the expressions and clinicopathological characteristics as well as their correlation. Results: the expression of HIC-1 mRNA was up-regulated in the colon cancerous tissues of 141 patients (70.5%) and in the paracancerous tissues of 21 patients (10.5%) with colon cancer, and the difference was statistically significant (P = 0.001). The expression of HIC-1 mRNA was correlated with Dukes stage, lymph node metastasis, and differentiation degree. The up-regulated expression of HIC-1 mRNA in colon cancer and the corresponding paracancerous tissues were 104 (52.0%) and 121 (60.5%), respectively. The expression of HIC-1 mRNA was not correlated with any clinicopathological characteristics of colon cancer. The immunohistochemical staining intensity of HIC-1 in cancerous tissues was 1.79 ± 0.11, higher than that in paracancerous tissues (P = 0.002). The expression level of HIC-1 was correlated with Dukes stage and lymph node metastasis. The expression of HIC-1 in colon cancerous and paracancerous tissues was weaker than that of HIC-1, which was not correlated with clinicopathological characteristics. There was a negative correlation between the expression of computer-aided and HIC-1 in colon cancerous tissues (r = -0.63, P < 0.01). Conclusions: The expression of HIC-1 can promote the progression of colon cancer, and it may have an antagonistic effect.

Molecular mechanisms underlying eicosapentaenoic acid inhibition of HDAC1 and DNMT expression and activity in carcinoma cells

DNA methylation and histone acetylation, the most studied epigenetic changes, drive and maintain cancer phenotypes. DNA methyltransferase (DNMT) dysregulation promoted localized hypermethylation in CpG rich regions while upregulated histone deacetylases (HDAC) deacetylated histone tails. Both changes led to close chromatin conformation, suppressing transcription and silencing tumor suppressor genes. Consequently, HDAC and DNMT inhibitors appeared to reprogram the transcriptional circuit and potentiate anti-tumoral activity. Here, we report that eicosapentaenoic acid (EPA), a fatty acid with anti-cancer properties, inhibited HDAC1 and DNMT expression and activity, thus promoting tumor suppressor gene expression. In hepatocarcinoma cells (HCC) EPA bound and activated PPARγ thus downregulating HDAC1 which sequentially reduced expression of DNMT1, 3A and 3B. At the same time, activated PPARγ physically interacted with DNMT1 and HDAC1 in a CpG island on the Hic-1 gene to assemble PPARγ/DNMT1 and PPARγ/HDAC1 protein complexes, which exited from DNA. When EPA and PPARγ were no longer bound, the protein complexes separated into individual proteins. Consequently, DNMT1 and HDAC1 down-regulation and release from DNA inhibited their activities. Overall, EPA-bound PPARγ induced re-expression of the tumor suppressor gene Hic-1. In the present study PPARγ emerged as a master regulator acting synergistically through diverse targets and ways to reveal the epigenetic action of EPA as an HDAC1 and DNMT1 inhibitor.

Chronic prostatitis alters the prostatic microenvironment and accelerates preneoplastic lesions in C57BL/6 mice

BackgroundChronic prostatitis has been supposed to be associated with preneoplastic lesions and cancer development. The objective of this study was to examine how chronic inflammation results in a prostatic microenvironment and gene mutation in C57BL/6 mice.MethodsImmune and bacterial prostatitis mouse models were created through abdominal subcutaneous injection of rat prostate extract protein immunization (EAP group) or transurethral instillation of uropathogenic E. coli 1677 (E. coli group). Prostate histology, serum cytokine level, and genome-wide exome (GWE) sequences were examined 1, 3, and 6 months after immunization or injection.ResultIn the EAP and E. coli groups, immune cell infiltrations were observed in the first and last months of the entire experiment. After 3 months, obvious proliferative inflammatory atrophy (PIA) and prostatic intraepithelial neoplasia (PIN) were observed accompanied with fibrosis hyperplasia in stroma. The decrease in basal cells (Cytokeratin (CK) 5+/p63+) and the accumulation of luminal epithelial cells (CK8+) in the PIA or PIN area indicated that the basal cells were damaged or transformed into different luminal cells. Hic1, Zfp148, and Mfge8 gene mutations were detected in chronic prostatitis somatic cells.ConclusionChronic prostatitis induced by prostate extract protein immunization or E. coli infection caused a reactive prostatic inflammation microenvironment and resulted in tissue damage, aberrant atrophy, hyperplasia, and somatic genome mutation.

Abstract LB-237: Circadian dysregulation via exposure to light at night alters DNA methylation of cancer-associated genes in murine mammary tissue

Abstract Breast cancer is a leading cause of cancer deaths among women, with an estimated 12.4% of American women diagnosed with this tumor type. Evidence suggests that exogenous disruptors can exert epigenetic changes leading to aberrant gene expression, putatively shifting the balance toward oncogenesis. While chemical pollutant exposure is well-studied, light exposure may exert diverse effects that are often overlooked. Mounting epidemiological evidence suggests that chronic circadian dysregulation is associated with increased breast cancer risk. Exposure to light at night (LAN) via ambient nighttime lighting, digital technology, shiftwork, and trans-meridian travel can alter temporal organization of cellular processes such as cell cycle regulation and tissue proliferation. Previous studies demonstrated that animals exposed to altered light cycles exhibited marked DNA methylation pattern changes, and aberrant DNA methylation is noted across multiple loci in the cancer genome. LAN exposure may thus contribute to the etiology of breast cancer via alteration of the epigenome and subsequent temporal disruption of typical gene expression patterns. We previously found (unpublished data) that mPer2::luc females exposed to 21 days of LAN (18:6 LD) exhibited profound circadian disruption particularly in mammary chain, in comparison to control-exposed (12:12 LD) mice, and that LAN exposure significantly decreased mammary ERα and ERβ expression. We are continuing to explore LAN-induced molecular alterations in the mammary via whole-genome bisulfite sequencing, to determine if inappropriate light exposure is an etiological risk factor of hormone-dependent mammary cancer. Post-pubertal female mice were exposed to LAN or control light cycles for 21d prior to mammary tissue harvest as described above for mPer2::luc knock-in mice. Genomic DNA samples were bisulfite-converted and sequenced, aligned to a bisulfite-converted genome, and analyzed to identify differential DNA methylation, using probes for regulatory regions (-500 to +2000bp), exons, introns, and CpG islands. LAN exposure induced changes in mean methylation of probes across 6.4% of regulatory regions (2044 of 32025), 5.5% of exons (1752 of 32025), 0.03% of introns (577 of 190791), and 1.9% of CpG islands (269 of 13840) individually analyzed and annotated with surrounding or downstream genes (adjusted p&amp;lt;0.05, abs. min. difference of means &amp;gt;5). Significant methylation changes resulting from LAN were noted in multiple genetic loci implicated in cancer, including directional methylation changes intermediate between control and 4T1 breast cancer models across various Hox genes, Hic1, Cdkn1c, and other genes associated with proliferation and oncogenesis, with commensurate changes in gene expression also observed. We are currently working to confirm relative expression of commensurate genes via qRT-PCR. Together, these results point to a potential mechanism by which LAN exposure may initiate cellular dysregulation within mammary tissue. Citation Format: Rebecca E. Veitch, Shay Bracha, Patrick Chappell. Circadian dysregulation via exposure to light at night alters DNA methylation of cancer-associated genes in murine mammary tissue [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-237.

Tandem repeat variation near the HIC1 (hypermethylated in cancer 1) promoter predicts outcome of oxaliplatin-based chemotherapy in patients with metastatic colorectal cancer.

The hypermethylated in cancer 1/sirtuin 1 (HIC1/SIRT1) axis plays an important role in regulating the nucleotide excision repair pathway, which is the main oxaliplatin-induced damage-repair system. On the basis of prior evidence that the variable number of tandem repeat (VNTR) sequence located near the promoter lesion of HIC1 is associated with HIC1 gene expression, the authors tested the hypothesis that this VNTR is associated with clinical outcome in patients with metastatic colorectal cancer who receive oxaliplatin-based chemotherapy. Four independent cohorts were tested. Patients who received oxaliplatin-based chemotherapy served as the training cohort (n = 218), and those who received treatment without oxaliplatin served as the control cohort (n = 215). Two cohorts of patients who received oxaliplatin-based chemotherapy were used for validation studies (n = 176 and n = 73). The VNTR sequence near HIC1 was analyzed by polymerase chain reaction analysis and gel electrophoresis and was tested for associations with the response rate, progression-free survival, and overall survival. In the training cohort, patients who harbored at least 5 tandem repeats (TRs) in both alleles had a significantly shorter PFS compared with those who had fewer than 4 TRs in at least 1 allele (9.5 vs 11.6 months; hazard ratio, 1.93; P = .012), and these findings remained statistically significant after multivariate analysis (hazard ratio, 2.00; 95% confidence interval, 1.13-3.54; P = .018). This preliminary association was confirmed in the validation cohort, and patients who had at least 5 TRs in both alleles had a worse PFS compared with the other cohort (7.9 vs 9.8 months; hazard ratio, 1.85; P = .044). The current findings suggest that the VNTR sequence near HIC1 could be a predictive marker for oxaliplatin-based chemotherapy in patients with metastatic colorectal cancer. Cancer 2017;123:4506-14. © 2017 American Cancer Society.

Epigenetics of Human Parathyroid Tumors

Parathyroid tumors are common endocrine neoplasia associated with primary hyperparathyroidism, a metabolic disorder sustained by parathormone hypersecretion. The epigenetic scenario in parathyroid tumors is beginning to be decoded. Here, main findings are reviewed: hypermethylation of specific DNA CpG islands has been described, despite global DNA promoter hypomethylation was not detectable; embryonic-related miRNAs, belonging to the C19MC and miR‐371-373 clusters, and miR‐296, are deregulated; expression of histone H1.2 and H2B is increased; expression of histone methyltransferase EZH2, BMI1 and RIZ1 is impaired; the tumor suppressor HIC1, MEN1 and CDC73 gene products, key molecules in parathyroid tumorigenesis, may be involved in epigenetic aberrant changes. Epigenetic changes are more frequent and more consistent in parathyroid malignancies, and positively correlated with severity of primary hyperparathyroidism.

124P - Tandem repeat variation in HIC1 gene predicts outcome for oxaliplatin-based chemotherapy in patients with metastatic colorectal cancer

124P - Tandem repeat variation in HIC1 gene predicts outcome for oxaliplatin-based chemotherapy in patients with metastatic colorectal cancer

KRAS, BRAF oncogene mutations and tissue specific promoter hypermethylation of tumor suppressor SFRP2, DAPK1, MGMT, HIC1 and p16 genes in colorectal cancer patients.

Colorectal cancer is a serious disease that causes significant morbidity and mortality in developed countries. Genetic changes, such as mutations in proto-oncogenes and DNA repair genes, and loss of function in the tumor suppressor genes cause colorectal cancer development. Abnormal DNA methylation is also known to play a crucial role in colorectal carcinogenesis. In this study, frequencies of KRAS and BRAF mutations, promoter hypermethylation profiles of SFRP2, DAPK1, MGMT, HIC1 and p16 genes, and possible associations between hypermethylation of these genes and KRAS and BRAF mutations were aimed to find out. Ninety three colorectal cancer tissues and 14 normal colon mucosas were included in the study. Common twelve KRAS gene mutation were investigated with using reverse-hybridization strip assay method. BRAF V600E mutations were investigated with RFLP method. Hypermethylation status of five tumor suppressor genes were detected by using reverse-hybridization strip assay method after bisulfite modification of DNA. KRAS and BRAF mutation frequencies were determined as 54.84% and 12.9%, respectively. Promoter hypermethylation frequencies of tumor suppressor genes SFRP2, DAPK1, MGMT, HIC1 and p16 were determined as 66.7%, 45.2%, 40.9%, 40.9% and 15.1%, respectively. Statistically significant associations were found between BRAF mutation and SFRP2 and p16 tumor suppressor genes hypermethylation (SFRP2; p= 0.005, p16; p= 0.016). Compared to rectum, SFRP2 (p= 0.017) and MGMT (p= 0.013) genes have statistically significantly higher promoter hypermethylation in colon. Results of the current study have confirmed that KRAS mutations and SFRP2 hypermethylation can be used as genetic markers in colorectal cancer.

Post-mortem cytogenomic investigations in patients with congenital malformations

Congenital anomalies are the second highest cause of infant deaths, and, in most cases, diagnosis is a challenge. In this study, we characterize patterns of DNA copy number aberrations in different samples of post-mortem tissues from patients with congenital malformations. Twenty-eight patients undergoing autopsy were cytogenomically evaluated using several methods, specifically, Multiplex Ligation-dependent Probe Amplification (MLPA), microsatellite marker analysis with a MiniFiler kit, FISH, a cytogenomic array technique and bidirectional Sanger sequencing, which were performed on samples of different tissues (brain, heart, liver, skin and diaphragm) preserved in RNAlater, in formaldehyde or by paraffin-embedding. The results identified 13 patients with pathogenic copy number variations (CNVs). Of these, eight presented aneuploidies involving chromosomes 13, 18, 21, X and Y (two presented inter- and intra-tissue mosaicism). In addition, other abnormalities were found, including duplication of the TYMS gene (18p11.32); deletion of the CHL1 gene (3p26.3); deletion of the HIC1 gene (17p13.3); and deletion of the TOM1L2 gene (17p11.2). One patient had a pathogenic missense mutation of g.8535C>G (c.746C>G) in exon 7 of the FGFR3 gene consistent with Thanatophoric Dysplasia type I. Cytogenomic techniques were reliable for the analysis of autopsy material and allowed the identification of inter- and intra-tissue mosaicism and a better understanding of the pathogenesis of congenital malformations.