CDX2 Research Articles

Association of vitamin D receptor polymorphisms with colorectal cancer susceptibility: A systematic meta-analysis

Recent studies have reported an association between vitamin D receptor (VDR) polymorphisms and colorectal cancer (CRC) risk; however, the results are controversial. This meta-analysis was performed to investigate whether the Cdx-2, Tru9I, FokI, BsmI, TaqI, and ApaI polymorphisms were correlated with CRC susceptibility. All potential studies were retrieved by searching the PubMed, EMBASE, and Cochrane Library databases through October 2, 2021. Odds ratios (ORs) with 95% confidence intervals were used to evaluate the correlation between VDR gene Cdx-2, Tru9I, FokI, BsmI, TaqI, and ApaI polymorphisms and CRC risk. In this meta-analysis, the BsmI variant was significantly correlated with a lower risk of CRC, especially in Caucasian population (B vs b: OR 0.94, 95%CI 0.90-0.99; BB vs bb: OR 0.88; 95%CI 0.79-0.97; BB vs Bb/bb: BB vs Bb/bb: OR 0.89; 95%CI 0.81-0.98). A statistically significant result from the FokI polymorphism was observed in colon cancer rather than rectal cancer (Ff vs FF: OR 0.86, 95%CI 0.84-0.93; ff/Ff vs FF: OR 0.88, 95%CI 0.79-0.98; ff vs Ff/FF: OR 0.90, 95%CI 0.82-0.99). Similarly, Cdx-2 polymorphism was found to be associated with decreased CRC risk among Africans (C vs c: OR 0.50, 95%CI 0.33-0.75; CC vs cc: OR 0.09, 95%CI 0.01-0.77; Cc vs cc: OR 0.49, 95%CI 0.30-0.81; CC/Cc vs cc: OR 0.45, 95%CI 0.28-0.74,). Our findings indicate that VDR polymorphisms are significantly associated with CRC risk.

Role of CDX2 Marker in Patients with Colorectal Cancer

CDX2 has been proposed as a tumor suppressor in colon cancer, CDX2 gene is often amplified in colon cancer, there is a lineage survival oncogene function in some tumors, the role of CDX2 protein during CRC development remains debatable. The aim of this study investigate the effect of low CDX2 expression on overall survival (OS) for prognosis of CRC patients and estimate of therapeutic activity of CDX2 expression in predictive of chemotherapy respond. This study done in the Middle Euphrates Unit for Cancer Research, Faculty of Medicine, University of Kufa in Al-Najaf province. This study was carried out on sixty-three cases of CRC in the form of available paraffin blocks who underwent surgical resection between 2015 and 2020. Thirty_ seven blocks of normal non tumoral colorectal tissue collected randomly from archives of two private laboratories during collection of malignant tissue blocks. Nuclear low expression of CDX2 in control and patient groups were 0(0.00%) and 30 (47.6%), while high expression of this protein was 37 (100%) and 33(52.4%) in control and patient groups respectively. OS was longer in patients with low CDX2 protein expression who intake adjuvant chemotherapy (71.33%) with mean survival (24.75±2.81 month) than patients were not intake adjuvant chemotherapy (53.71%) with mean survival (15.227±1.66 month). CDX2 expression regarded as diagnostic marker for non-mucinous CRC. High of CDX2 expression can be used as an independent good prognostic biomarker to predict longer survival of patients with CRC.

Comparing the VDR Gene BsmI and CDX2 Polymorphisms in Healthy Turks and Healthy Somalians Living in Turkiye

Objectives: This study aimed to evaluate genotypic and allelic differences by comparing the Vitamin D (Vit-D) receptor (VDR) gene BsmI (rs1544410) and CDX2 (rs11568820) polymorphisms in healthy Turks and healthy Somalians living in Turkiye. Materials and Methods: The study involved 100 healthy Turkish individuals and 60 healthy Somali individuals residing in Turkiye for at least 5 years. The genotyping study was performed using the polymerase chain reaction-restriction fragment length polymorphism (PCRRFLP) and allele-specific PCR methods. Statistical analysis was performed to identify the possible differences between groups using the Chi-square and Student’s t tests for pair-wise comparisons. Results: According to the data obtained in the study with regard to the BsmI (rs1544410 A/G) and CDX2 (rs11568820 A/G) genotypes, no statistically significant difference was determined to be present regarding the frequency of carrying the mutant GG genotype in the two groups (p = 0.95 and p = 0.221, respectively). Conclusion: The study has found no significant genotypic or allelic difference to be present in terms of the BsmI (rs1544410) and CDX2 (rs11568820) gene variants between healthy Turkish individuals and Somali individuals who have spent most of their lives exposed to less sunlight while living in Turkiye for the past five years.

The Beneficial Effects of Static Magnetic Field and Iron Oxide Nanoparticles on the Vitrification of Mature Mice Oocytes.

This study was conducted to evaluate the effects of static magnetic field (SMF) and nanoparticles (NPs) on the vitrification of cumulus-oocyte-complex (COC). To this end, the non-vitrified (nVit) and vitrified groups (Vit) that contain NPs, with or without SMF were labeled nVit_NPs, nVit_NPs_SMF, Vit_NPs, and Vit_NPs_SMF, respectively. The non-toxic dosages of NPs were first determined to be 0.008% w/v. The survival, apoptosis, and necrosis, mitochondrial activity, fertilization rate, subsequent-derived embryo development, and gene expressions were examined. The viability rates obtained by trypan blue and Anx-PI staining were meaningfully smaller in the Vit groups, compared to the nVit groups. The JC1 red/green signal ratios were reduced considerably in the Vit group, compared to the nVit. Transmission electron microscopy (TEM) was performed to assess the entry of the NPs into the oocytes. TEM images showed that NPs were present in nVit_NPs, and Vit_NPs. Thereafter, the effects of NPs and SMF on in vitro fertilization (IVF) were examined. The difference in blastocyst rates between nVit and Vit_NPs_SMF groups was significant. Finally, Nanog, Cdx2, Oct4, and Sox2 genes were evaluated. There were substantial differences in Cdx2 gene expressions between the Vit_NPs and nVit groups. The expression of Nanog in Vit was significantly higher than those of the Vit_NPs, Vit_NPs_SMF, and nVit groups. The data presented here provide deeper insight into the application of iron oxide nanoparticles in COC vitrification. It appears that using SMF and supplemented CPA by NPs inhibits cryoinjury and promote the embryo development capacity of vitrified-warmed COCs.

Alpha lipoamide inhibits diabetic kidney fibrosis via improving mitochondrial function and regulating RXRα expression and activation.

Previous studies have shown mitochondrial dysfunction in various acute kidney injuries and chronic kidney diseases. Lipoic acid exerts potent effects on oxidant stress and modulation of mitochondrial function in damaged organ. In this study we investigated whether alpha lipoamide (ALM), a derivative of lipoic acid, exerted a renal protective effect in a type 2 diabetes mellitus mouse model. 9-week-old db/db mice were treated with ALM (50 mg·kg-1·d-1, i.g) for 8 weeks. We showed that ALM administration did not affect blood glucose levels in db/db mice, but restored renal function and significantly improved fibrosis of kidneys. We demonstrated that ALM administration significantly ameliorated mitochondrial dysfunction and tubulointerstitial fibrotic lesions, along with increased expression of CDX2 and CFTR and decreased expression of β-catenin and Snail in kidneys of db/db mice. Similar protective effects were observed in rat renal tubular epithelial cell line NRK-52E cultured in high-glucose medium following treatment with ALM (200 μM). The protective mechanisms of ALM in diabetic kidney disease (DKD) were further explored: Autodock Vina software predicted that ALM could activate RXRα protein by forming stable hydrogen bonds. PROMO Database predicted that RXRα could bind the promoter sequences of CDX2 gene. Knockdown of RXRα expression in NRK-52E cells under normal glucose condition suppressed CDX2 expression and promoted phenotypic changes in renal tubular epithelial cells. However, RXRα overexpression increased CDX2 expression which in turn inhibited high glucose-mediated renal tubular epithelial cell injury. Therefore, we reveal the protective effect of ALM on DKD and its possible potential targets: ALM ameliorates mitochondrial dysfunction and regulates the CDX2/CFTR/β-catenin signaling axis through upregulation and activation of RXRα. Schematic figure illustrating that ALM alleviates diabetic kidney disease by improving mitochondrial function and upregulation and activation of RXRα, which in turn upregulated CDX2 to exert an inhibitory effect on β-catenin activation and nuclear translocation. RTEC renal tubular epithelial cell. ROS Reactive oxygen species. RXRα Retinoid X receptor-α. Mfn1 Mitofusin 1. Drp1 dynamic-related protein 1. MDA malondialdehyde. 4-HNE 4-hydroxynonenal. T-SOD Total-superoxide dismutase. CDX2 Caudal-type homeobox transcription factor 2. CFTR Cystic fibrosis transmembrane conductance regulator. EMT epithelial mesenchymal transition. α-SMA Alpha-smooth muscle actin. ECM extracellular matrix. DKD diabetic kidney disease.Schematic figurewas drawn by Figdraw ( www.figdraw.com ).

Targeted silencing of CDX2 gene with siRNA enhanced vincristine-induced inhibition of proliferation and induction of apoptosis of leukemia K562 cells

Purpose: To determine the effect of targeted silencing of CDX2 with siRNA on vincristine-induced inhibition of proliferation and apoptosis induction in leukemia K562 cells.
 Methods: K562 cells were divided into untreated group, vincristine group, blank group, and CDX2-siRNA group. The expression of CDX2 gene after CDX2-siRNA transfection was determined by reverse-transcription-polymerase chain reaction (RT-PCR) and immunoblotting, while MTT and flow cytometric procedures were used to evaluate the effect of vincristine on proliferation and apoptotic changes in K562 cells.
 Results: Protein expression of CDX2 did not change significantly in non-treated cells, vincristine group and blank group, while the expression of CDX2 protein in cells of CDX2-siRNA group was decreased significantly (p < 0.05) while MTT assay results showed that the absorbance of CDX2-siRNA cells was significantly lower than those of the other three groups at 12, 24 and 48 h after CDX2 gene silencing. Flow cytometry showed markedly higher percentage apoptosis of CDX2-siRNA group than in other groups at 12, 24 and 48 h after CDX2 gene silencing. There was no difference in apoptosis level between vincristine group and blank group, but apoptosis was higher in these groups than in untreated group (p < 0.05).
 Conclusion: Silencing CDX2 gene via targeting with siRNA enhances vincristine-induced suppression of growth and apoptotic changes in leukemia K562 cells, thereby enhancing the anti-tumor effect of vincristine.

Effects of LPA on the development of sheep in vitro fertilized embryos and attempt to establish sheep embryonic stem cells

Lysophosphatidic acid (LPA) is a small molecule glycerophospholipid, which regulates multiple downstream signalling pathways through G-protein-coupled receptors to achieve numerous functions on oocyte maturation and embryo development. In this study, sheep in vitro fertilized embryos were applied to investigate the effects of LPA on early embryos development and embryonic stem cell establishment. At first, the maturation medium containing estrus female sheep serum and synthetic oviduct fluid (SOF) were optimized for sheep IVF, and then the effects of LPA were investigated. From 0.1 to 10 μmol L–1, LPA had no significant effect on the cleavage rate (P>0.05), but the maturation rate and blastocyst rate increased dependently with LPA concentration (P<0.05), and the blastocyst morphology was normal. When the LPA concentration was 15 μmol L–1, the maturation rate, cleavage rate and blastocyst rate decreased significantly (P<0.05), and the blastocyst exhibited abnormal morphology and could not develop into high-quality blastocyst. Besides, the exogenous LPA increases the expression of LPAR2, LPAR4, TE-related gene CDX-2 and pluripotency-related gene OCT-4 in sheep early IVF embryos with the raise of LPA concentration from 0.1 to 10 μmol L–1. The expression of LPAR2, LPAR4, CDX-2 and OCT-4 from the LPA-0.1 μmol L–1 to LPA-10 μmol L–1 groups in early embryos were extremely significant (P<0.05), while the expression of these genes significantly decreased in 15 μmol L–1 LPA-treated embryos compared with LPA-10 μmol L–1 group (P<0.05). The inner cell mass in 15 μmol L–1 LPA-treated embryos was also disturbed, and the blastocysts formation was abnormal. Secondly, the sheep IVF blastocysts were applied to establish embryonic stem cells. The results showed that LPA made the blastocyst inoculated cells grow towards TSC-like cells. They enhanced the fluorescence intensity and mRNA abundance of OCT-4 and CDX-2 as the concentration increased from 0 to 10 μmol L–1, while 15 μmol L–1 LPA decreased OCT-4 and CDX-2 expression in the derived cells. The expression of CDX-2 and OCT-4 in the blastocyst inoculated cells of LPA-1 μmol L–1 group and LPA-10 μmol L–1 group extremely significantly increased (P<0.05), but there was significant decrease in LPA-15 μmol L–1 group compared with LPA-10 μmol L–1 group (P<0.05). Meanwhile, the protein expression of LPAR2 and LPAR4 remarkably increased after treatment of LPA at 10 μmol L–1 concentration. This study references the IVF embryo production and embryonic stem cell research of domestic animals.

Vitamin D Receptor Genetic Polymorphisms Associate With a Decreased Susceptibility to Extremity Osteomyelitis Partly by Inhibiting Macrophage Apoptosis Through Inhibition of Excessive ROS Production via VDR-Bmi1 Signaling.

Background: Previous studies had reported that vitamin D receptor (VDR) gene polymorphisms were related to the development of several inflammatory disorders. However, potential links between such variations and the risk of developing a bone infection and underlying mechanisms remain unclear. This study aimed to analyze potential associations between VDR genetic variations and susceptibility to extremity osteomyelitis (OM) in a Chinese Han population and investigate potential mechanisms. Methods: Between January 2016 and August 2020, altogether 398 OM patients and 368 healthy controls were genotyped for six VDR gene polymorphisms, including ApaI (rs7975232), BsmI (rs1544410), FokI (rs2228570), TaqI (rs731236), GATA (rs4516035), and Cdx-2 (rs11568820) by the SNaPshot genotyping method. Then, male C57BL/6 mice were randomly divided into vitamin D–standard, –excess, –deficient, and –rescued groups. One week after making the model surgery, OM occurrence and severity were assessed using the bacterial count and histopathological staining. In vitro, phagocytosis, apoptosis, and bactericidal ability of macrophages were evaluated by overexpression or knockdown of VDR protein. Results: Significant associations were found among rs7975232, rs1544410, and OM development by the recessive model (AA vs. AC + CC, p = 0.037, OR = 0.594), homozygous model (AA vs. CC, p = 0.033, OR = 0.575), and heterozygous model (CT vs. CC, p = 0.049, OR = 0.610), respectively. Patients with the AA genotype of rs7975232 had a relatively higher mean level of vitamin D than those with AC and CC genotypes (22.5 vs. 20.7 vs. 19.0 ng/ml). Similarly, patients with CT genotype of rs1544410 had a relatively higher mean vitamin D level than those with CC genotype (20.94 vs. 19.89 ng/ml). Outcomes of in vivo experiments showed that the femoral bacterial load of vitamin D–deficient mice was highest among different vitamin D dose groups, with the most severe histopathological features of infection, and vitamin D supplementation partly reversed the changes. While in vitro experiment results revealed that active vitamin D promoted phagocytosis and sterilization of macrophages and inhibited apoptosis during infection. Reactive oxygen species (ROS) inhibitor inhibited apoptosis of macrophages induced by bacterial infection. Active vitamin D inhibited excessive ROS production in macrophages via the VDR-Bmi1 signaling pathway. Conclusion: In this Chinese cohort, ApaI and BsmI are associated with a decreased risk of OM development by influencing serological vitamin D level, the latter of which reduced macrophage apoptosis with inhibition of excessive ROS production via the VDR-Bmi1 signaling pathway.

Cryoprotectant role of exopolysaccharide ID1 in the vitrification/in-straw warming of in vitro-produced bovine embryos.

The cold-adapted bacterium Pseudomonas sp. ID1 produces the extracellular exopolysaccharide ID1 (EPS ID1) with cryoprotective activity. This study was designed to optimize the vitrification/in-straw warming protocol of in vitro-produced (IVP) blastocysts by adding EPS ID1 to the vitrification media. Day 7-expanded blastocysts were vitrified/warmed using the VitTrans device after the addition of 0 or 100 μg/mL EPS ID1 to the vitrification media. Blastocysts vitrified by the Cryotop method and fresh non-vitrified blastocysts served as controls. Outcomes were assessed in the warmed embryos in terms of survival rates and mRNA relative abundances of BAX, BCL2, GPX1, and CDX2 genes. No differences in survival rates were observed at 3h post-warming between vitrification treatments. At 24 h post-warming, the addition of EPS prior to vitrification with the VitTrans device produced similar survival rates to Cryotop-vitrified embryos and similar hatching rates to fresh non-vitrified or Cryotop-vitrified embryos. No differences emerged in BCL2 gene expression. Lower BAX (p < .05) and higher GPX1 (p < .05) and CDX2 (p < .1) gene expression were observed in expanded and/or hatched blastocysts derived from VitTrans-EPS-vitrified embryos when compared to those from the non-supplemented group. In conclusion, addition of EPS not only promoted blastocyst survival and hatching after VitTrans vitrification/warming but also modified the expression of genes associated with better embryo quality.

Abstract 2183: Interrogating the molecular profile of colorectal cancer: detection of clinically actionable alterations in Hispanics

Abstract Background: Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths among men and women and the third cause of cancer deaths in the US. Tumor profiling techniques have led to the identification of actionable alterations to guide treatment and improve survival in cancer patients. However, comprehensive genetic profiling of colorectal cancer tumors from Hispanics living in Puerto Rico (PRH) has not been performed. Our study aimed at understanding the common CRC carcinogenic molecular pathways that affect our Hispanic population by evaluating colorectal tumors from molecularly characterized ethnically diverse cohorts. Methods: We retrospectively evaluated the mutational profile of CRC tumors from 218 PRH that underwent NGS testing from 2015 to 2020 (provided by CARIS Life Sciences and the Precision Oncology Alliance). We estimated the prevalence of somatic mutations of PRH CRC tumors and compared them with those reported in The Cancer Genome Atlas (TCGA) Pan-Cancer Atlas Clinical Data and the AACR Project Genomics Evidence Neoplasia Information Exchange (GENIE), both available through the cBioPortal for Cancer Genomics. Descriptive statistics were performed to characterize the database. Results: Among the top mutated genes for CRC tumors in PRH were APC (81.9%, n=182), TP53 (75.3%, n=178), KRAS (56.2%, n=185), PIK3CA (18.0%, n=178) and SAMD4 (12.6%, n=175). For PRH, the most frequent gene amplifications were: CDX2 (20.1%, n=184), CDKN1B (6.6%, n=183), and HNRNPA2B1 (5.4%, n=184). Significant differences in the mutational frequencies of CRC driver genes, such as APC, TP53, PIK3CA, FBXW7, CDX2, CDKN1B, and HNRNPA2B1, and actionable genes, such as KRAS/NRAS, BRAF and for ERBB2 amplifications, among PRH were observed when compared to the TCGA and GENIE datasets. The prevalence of MSI tumors for PRH was 2.0%. Conclusion: This is the first study to report the mutational profile of CRC tumors from PRH and to describe differences in mutational frequencies when compared other non-Hispanic and U.S. mainland Hispanic populations using data from TCGA and GENIE. Differences in prevalence of immunotherapy and targeted associated biomarkers may contribute to the observed disparities in survival and response to therapies. Understanding the most common carcinogenic molecular pathways that affect Hispanics with CRC is crucial to guide research efforts in the development of new therapeutic modalities incorporating genomically diverse populations. Citation Format: Ingrid M. Montes-Rodriguez, Hilmaris Centeno-Girona, Camila Rivera-Lynch, Noridza Rivera, Marcia Cruz-Correa. Interrogating the molecular profile of colorectal cancer: detection of clinically actionable alterations in Hispanics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2183.

A Case of Metastatic Breast Cancer that Responded to Anti-HER2 Therapy with Amplification of the ERBB2 Gene in the FoundationOne®CDx Gene Panel Test after Completion of Standard Therapy

A Case of Metastatic Breast Cancer that Responded to Anti-HER2 Therapy with Amplification of the ERBB2 Gene in the FoundationOne®CDx Gene Panel Test after Completion of Standard Therapy

Epigenetic reprogramming in cloned mouse embryos following treatment with DNA methyltransferase and histone deacetylase inhibitors

We examined the effects of DNA methyltransferase inhibitor – RG108, and histone deacetylase inhibitor – SAHA, on the reprogramming parameters of cloned mouse embryos produced by somatic cell nuclear transfer into oocytes. The programming parameters studied included dynamics of histone reacetylation, developmental rate, DNA methylation, and transcript levels of genes, all of which are pivotal to lineage specification and blastocyst formation. At the pronuclear stage, somatic nucleus-transplanted oocytes treated with 5 µM SAHA presented higher histone acetylation at H3K9, H3K14, H4K16 and H4K12, compared to untreated clones (p < 0.05). At the morula stage, cloned embryos treated with 5 μM RG108 or 5 μM SAHA presented lower DNA methylation intensity compared to untreated clones (p < 0.05), resembling the intensity levels of fertilized embryos. However, these effects were not observed when RG108 and SAHA were used in combination. The rate of morula formation was significantly higher in cloned embryos treated with 5 µM SAHA than in untreated clones, whereas treatment with RG108 resulted in no obvious effects on morula formation rates. On the other hand, the combined treatment with RG108 and SAHA resulted in inferior rates of cloned morula formation, compared to untreated clones. At the blastocyst stage, the aberrant expression levels of key developmental genes Oct4 and Cdx2, but not Nanog, were corrected in cloned embryos by the treatment with RG108. This is similar to the intensity levels seen in fertilized embryos. The expression of Rpl7l1 gene was significantly higher in embryos treated with both RG108 and SAHA than in untreated and in control groups. In summary, the present study showed that SAHA and RG108, when applied separately, improve the rate and quality of cloned mouse embryos.

Immunohistochemical Expression of CDX2 in Gastric Carcinoma.

Background & Objective:Gastric cancer (GC) persists to be a major health issue globally, and the need to investigate new molecular markers for improving the survival of patients continues. CDX2 is a homeobox caudal protein family member encoded by the CDX2 gene and is probably playing a role in intestinal epithelial differentiation and proliferation. This study aimed to assess the expression of this protein in gastric cancer cells in addition to its correlation with multiple clinicopathological parameters. Methods:This observational retrospective study was carried out on 80 gastric cancer cases in Erbil, Iraq. CDX2 protein immunoexpression in tumor cells, as well as its correlation with several clinicopathological criteria, were investigated.Results:CDX2 was detected in 38.75% of GC patients. We found a significant correlation between CDX2 expression and the age of patients (P=0.02). Even though the protein was more expressed in tumors with negative lymphovascular invasion and intestinal GC, there was no significant correlation between the expression of this protein and invasion. In addition, CDX2 expression was not significantly correlated with patient gender, tumor grade, nodal status, and tumor stage. Conclusion:CDX2 expression was observed to be downregulated in younger patients. It could be due to the higher frequency of diffuse GC, in which CDX2 is expressed less than the intestinal type, in younger individuals.

New Wenshen Shengjing Decoction Improves Early Embryonic Development by Maintaining Low Levels of H3K4me3 in Sperm.

Background New Wenshen Shengjing Decoction (NWSSJD), a traditional Chinese compound medicine, has significant effect on spermatogenesis disorder and can significantly improve sperm quality. Many components in NWSSJD can induce epigenetic modifications of different types of cells. It is not yet known whether they can cause epigenetic modifications in sperm or early embryos. Objective This study investigated the effect of NWSSJD on mouse early embryonic development and its regulation of H3K4me3 in mouse sperm and early embryos. Methods Spermatogenesis disorder was induced in male mice with CPA (cyclophosphamide). NWSSJD was administrated for 30 days. Then, the male mice were mated with the female mice with superovulation, and the embryo degeneration rate of each stage was calculated. Immunofluorescence staining was used to detect the expression of H3K4me3 in sperm and embryos at various stages. Western blotting was performed to detect methyltransferase SETD1B expression. The expressions of development-related genes (OCT-4, NANOG, and CDX2) and apoptosis-related genes (BCL-2 and p53) were measured with qRT-PCR. Results Compared with the CPA group, NWSSJD significantly reduced the H3K4me3 level in sperms, significantly increased the number of normal early embryos (2-cell embryos, 3-4-cell embryos, 8-16-cell embryos, and blastocysts) per mouse, and reduced the degeneration rate of the embryos. The expression levels of H3K4me3 and methyltransferase SETD1B in early embryos were significantly elevated by NWSSJD. Additionally, NWSSJD significantly promoted BCL-2 expression, while reducing p53 expression, thus inhibiting embryonic cell apoptosis. Moreover, the expressions of development-related genes OCT-4 and CDX2 were significantly increased by NWSSJD, but NANOG expression had no significant difference. Conclusion NWSSJD may promote early embryonic development possibly by maintaining low H3K4me3 levels in sperms and normal H3K4me3 modification in early embryos and by inhibiting embryonic cell apoptosis.

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.

The effects of embryo splitting on Cdx2, Sox2, Oct4, and Nanog gene expression in mouse blastocysts.

Embryo splitting is utilized in reproduction biotechnology. The blastomeres resulting from the splitting of an embryo in two-, four- or eight-cell stages can develop into separate embryos that are genetically similar to the other blastomeres. The present work studied the effects of splitting on embryo pluripotent gene expression (Cdx2, Sox2, Oct4, and Nanog) in mice. Two-cell embryos were isolated from stimulated mice. The embryos were grouped into "split" and "non-split" groups. The zona pellucida was removed from the split group and the blastomeres were distributed before being co-cultured with mouse embryo fibroblasts to the blastocyst stage. Normal (non-split) blastocysts were co-cultured in the same way. The 3.5-day-old blastomeres were collected as the control group. For molecular evaluation, real-time PCR was conducted to analyze changes in Cdx2, Sox2, Oct4, and Nanog gene expression. Moreover, the blastocyst formation rate, overall blastocyst rate, and the number of newborns were statistically analyzed. The findings showed that embryo splitting increased blastocyst formation, overall blastocysts, developmental potential embryos, and the number of infants. Furthermore, the split and non-split (control) groups showed equal expression of pluripotent genes (Cdx2, Sox2, Oct4, and Nanog) in the molecular analysis. It can be concluded that the growth and developmental potency of sister blastocysts derived from split two-cell stage mouse embryos are the same as those of normal blastocysts. So, there are no significant differences in gene expression between the split and non-split groups.

Analyses of serum 25-hydroxyvitamin D levels and single nucleotide polymorphisms in the vitamin D receptor gene for acne vulgaris

Background Acne vulgaris (AV) is one of the diseases that are widespread in Egypt. It is a disease that mainly affects the pilosebaceous units of the face, neck, and others. Aim To analyze and assess the role of 25-hydroxyvitamin D levels in serum as well as different vitamin D receptor (VDR) genes (Taq-1, Apa-1, Fok-1, and Cdx-2 polymorphisms) among Egyptian patients having AV. Patients and methods A total of 300 patients with acne and another 300 healthy controls were included in the study. Patients were subdivided into four different groups: group 1 received cholecalciferol 8000 IU/day for 3 months; group 2 applied 1–2 g of topical vitamin D analog for 3 months, group 3 had no treatment, and lastly, group 4 included healthy controls. For all patients, skin manifestations were assessed. Serum 25(OH)3 D was measured in all participants before and after treatment. Different VDR gene polymorphisms were measured. Results Patients with acne showed a statistically significant decrease in serum 25(OH) D3 concentration than controls, with P value of 0.001. Regarding VDR polymorphisms (rs731236 for Taq-1, rs7975232 for Apa-1, rs2228570 for Fok-1, and rs11568820 Cdx-2 polymorphisms), the results revealed that there was a statistically significant difference between patients with acne and controls regarding Taq-1 and Apa-1 genotypes, with P values of 0.001 and 0.026, respectively, whereas regarding Fok-1 and Cdx-2 genotypes, the results revealed that there were no statistically significant differences between the studied patients and the controls (P=0.690 and 0.113, respectively). Conclusion Vitamin D and VDR polymorphisms (Taq-1, Apa-1, Fok-1, and Cdx-2) play important roles in AV.

385 MIR-24-3P REGULATES CDX2 DURING THE PROCESS OF INTESTINALIZATION OF THE CARDIAC-TYPE EPITHELIUM IN A HUMAN MODEL OF BARRETT’S ESOPHAGUS

Abstract Cardiac-type epithelium has been proposed as an intermediate stage between normal squamous epithelium and intestinal metaplasia in the development of Barrett’s esophagus. Deregulation of certain miRNAs and their effects on CDX2 expression might contribute to the intestinalization process of cardiac-type epithelium. The aim of this study was to identify miRNAs differentially expressed between CDX2 positive and negative glands of Barrett’s esophagus and to examine the function of specific miRNAs on the regulation of CDX2. Methods miRNA expression profiling using OpenArrayTM analysis in microdissected cardiac-type glands with and without fully CDX2 expression was performed in biopsies from patients who developed cardiac-type epithelium in the remnant esophagus after esophagectomy. Data were validated using real-time PCR in esophageal adenocarcinoma cell lines and in situ and real-time PCR miRNA/CDX2/MUC2 co-expression analysis in cardiac-type mucosa samples. The effect of miR-24-3p precursor transfection on CDX2 expression was assessed in the esophageal adenocarcinoma cell lines FLO-1 and KYAE-1. Results CDX2 positive glands were characterized by an unique miRNA profile with a significant downregulation of miR-24-3p, miR-520e-3p, miR-548a-1, miR-597-5p, miR-133a-3p, miR-30a-5p, miR-638, miR-625-3p, miR-1255b-1, miR-1260a and upregulation of miR-590 (Figure 1A). miRNA-24-3p was identified as potential regulator of CDX2 gene expression in three bioinformatics algorithms, and this was confirmed in esophageal adenocarcinoma cell lines (Figure 1C). Furthermore, miR-24-3p expression negatively correlates with CDX2 in cardiac-type mucosa samples with different stages of mucosal intestinalization (Figure 1B). Conclusion These results imply that miRNA-24-3p directly targets CDX2, and downregulation of miRNA-24-3p is associated with the acquisition of an intestinal phenotype in cardiac-type epithelium.

Vitamin D Receptor Gene Polymorphisms in Childhood Brucellosis in Turkish Children

Abstract Objective The vitamin D receptor gene (VDR) polymorphisms and the risk of various infections have been studied. An association with brucellosis and vitamin D levels has been investigated but not yet with VDR gene polymorphisms. We aimed to examine the association between VDR gene polymorphisms and susceptibility to childhood brucellosis. Methods This case–control study included patients with brucellosis and healthy controls. After extracting genomic DNA using a Qiagen blood DNA isolation kit, five VDR single nucleotide polymorphisms (SNPs), including Cdx-2, FokI, BsmI, ApaI, and TaqI gene, were amplified. Genetic distribution of these SNPs of VDR gene in patient and control groups were compared. Results A total of 38 patients with brucellosis and 89 healthy controls were evaluated. The genotype distribution of Cdx2, FokI, BsmI, and ApaI polymorphisms were similar between patients and healthy controls. However, the CC homozygous genotype for VDR gene TaqI was significantly overexpressed in patients compared with controls (23.7 vs. 7.9%; p = 0.042). The frequency of the C allele of the TaqI genotype was significantly different between patients and controls (p = 0.018). On the other hand, presence of the A allele in the BsmI was associated considerably with an increased risk of brucellosis (p = 0.037). VDR polymorphism distribution was similar according to age, presence of complicated disease, and presence of bacteremia. The heterozygote TaqI polymorphism was more common in patients presented as subacute and chronic symptoms (p = 0.036). Conclusion Our results indicated the possible role in TaqI polymorphism of the VDR gene for the risk of brucellosis at the time of exposure to infection.

Abstract 612: Interrogating the molecular profile of colorectal cancer tumors in Puerto Rican Hispanics

Abstract Background: Colorectal cancer (CRC) is the second and third most common diagnosed cancer in males and females, respectively, and is the second leading cause of cancer-related deaths in Puerto Rico (PR). This differs from USA, were CRC is the third most commonly diagnosed cancer and the third leading cause of cancer-related deaths for men and woman, respectively. CRC is a heterogeneous disease and the genetic characteristics of the tumors are considered to evaluate prognostic outcome and selection of targeted therapies. Currently, techniques such as next-generation sequencing (NGS) are being used for tumor profiling, in order to classify and identify driver genetic mutations in CRC patients. Even though there have been numerous studies that have use genetic profiling to classify and to identify driver genetic mutations for CRC, comprehensive genetic profiling of cancer patients in the Hispanic population of PR has not been performed. Methods: We retrospectively evaluated the mutational profile of CRC tumors from 218 Puerto Rican Hispanics (PRH) that underwent NGS testing from 2015 to 2020. The data was provided by CARIS Life Sciences that uses a 592 gene panel in order to detect mutations, indels and copy number variants (CNV). We estimated the prevalence of somatic mutations of PRH CRC tumors and compared them with the mutational profiles reported for Colorectal Adenocarcinoma from the TCGA Pan-Cancer Clinical Data available in the cBioPortal for Cancer Genomics. Descriptive statistics were performed to characterize the database. Results: For our population, 7.8% were diagnosed with CRC before 50 years of age (early-onset CRC) and 92.2% were diagnosed after 50 years of age. These analyses showed that the most commonly mutated genes for CRC tumors in PRH are APC (82%), TP53 (76%), KRAS (56%), PIK3CA (18%), SMAD4 (13%), AMER1 (10%), FBXW7 (9%), BRAF (5%), RNF43 (4%), and MUTYH (4%). Several genes are shared among the top ten most mutated genes when comparing the CRC tumor mutational profiles from PRH with that reported in the CRC tumors from TCGA study, including APC, TP53, KRAS, and PIK3CA, having mutational frequencies of 73%, 59%, 41%, and 28%, respectively. The most common CNVs for PRH CRC tumors were CDX2 (20%), CDKN1B (7%), and HNRNPA2B1 (5%) genes. These CNVs have also been reported in the CRC TCGA Pan-Cancer study with frequencies of 6.4%, 0.3% and 0.3 % for CDX2, CDKN1B, and HNRNPA2B1 genes, respectively. Conclusion: This study is the first to reports the mutational profile of CRC tumors from Puerto Rican Hispanics. CRC tumors from PRH and the TCGA study, which has only 0.8% of Hispanic/Latino patients, share mutated genes and CNV, although with different mutational frequencies. Understanding the most common carcinogenic molecular pathways that affect PRH with CRC is crucial to guide research efforts in the discovery of new therapeutic modalities. Citation Format: Ingrid M. Montes-Rodriguez, Hilmaris Centeno-Girona, Camila Rivera-Lynch, Marievelisse Soto-Salgado, Noridza Rivera, Marcia Cruz-Correa. Interrogating the molecular profile of colorectal cancer tumors in Puerto Rican Hispanics [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 612.