Landscape of FLT3 Variations Associated with Structural and Functional Impact on Acute Myeloid Leukemia: A Computational Study.

Acute myeloid leukemia (AML) is hallmarked by the clonal proliferation of myeloid blasts. Mutations that result in the constitutive activation of the fms-like tyrosine kinase 3 (FLT3) gene, coding for a class III receptor tyrosine kinase, are significantly associated with this heterogeneous hematologic malignancy. The fms-related tyrosine kinase 3 ligand binds to the extracellular domain of the FLT3 receptor, inducing homodimer formation in the plasma membrane, leading to autophosphorylation and activation of apoptosis, proliferation, and differentiation of hematopoietic cells in bone marrow. In the present study, we evaluated the association of FLT3 as a significant biomarker for AML and tried to comprehend the effects of specific variations on the FLT3 protein's structure and function. We also examined the effects of I836 variants on binding affinity to sorafenib using molecular docking. We integrated multiple bioinformatics tools, databases, and resources such as OncoDB, UniProt, COSMIC, UALCAN, PyMOL, ProSA, Missense3D, InterProScan, SIFT, PolyPhen, and PredictSNP to annotate the structural, functional, and phenotypic impact of the known variations associated with FLT3. Twenty-nine FLT3 variants were analyzed using in silico approaches such as DynaMut, CUPSAT, AutoDock, and Discovery Studio for their impact on protein stability, flexibility, function, and binding affinity. The OncoDB and UALCAN portals confirmed the association of FLT3 gene expression and its mutational status with AML. A computational structural analysis of the deleterious variants of FLT3 revealed I863F mutants as destabilizers of the protein structure, possibly leading to functional changes. Many single-nucleotide variations in FLT3 have an impact on its structure and function. Thus, the annotation of FLT3 SNVs and the prediction of their deleterious pathogenic impact will facilitate an insight into the tumorigenesis process and guide experimental studies and clinical implications.

Structure-Based Profiling of Potential Phytomolecules with AKT1 a Key Cancer Drug Target.

Identifying cancer biomarkers is imperative, as upregulated genes offer a better microenvironment for the tumor; hence, targeted inhibition is preferred. The theme of our study is to predict molecular interactions between cancer biomarker proteins and selected natural compounds. We identified an overexpressed potential molecular target (AKT1) and computationally evaluated its inhibition by four dietary ligands (isoliquiritigenin, shogaol, tehranolide, and theophylline). The three-dimensional structures of protein and phytochemicals were retrieved from the RCSB PDB database (4EKL) and NCBI's PubChem, respectively. Rational structure-based docking studies were performed using AutoDock. Results were analyzed based primarily on the estimated free binding energy (kcal/mol), hydrogen bonds, and inhibition constant, Ki, to identify the most effective anti-cancer phytomolecule. Toxicity and drug-likeliness prediction were performed using OSIRIS and SwissADME. Amongst the four phytocompounds, tehranolide has better potential to suppress the expression of AKT1 and could be used for anti-cancer drug development, as inhibition of AKT1 is directly associated with the inhibition of growth, progression, and metastasis of the tumor. Docking analyses reveal that tehranolide has the most efficiency in inhibiting AKT1 and has the potential to be used for the therapeutic management of cancer. Natural compounds targeting cancer biomarkers offer less rejection, minimal toxicity, and fewer side effects.

Nanoparticles-based drug delivery and gene therapy for breast cancer: Recent advancements and future challenges.

Breast cancer (BC) is amongst the most lethal cancer among females and conventional treatment methods like surgery, radiotherapy and chemotherapy are not effective enough as expected and suffer concerns of low bioavailability, low cellular uptake, emerging resistance, and adverse toxicities. Gene therapy using free nucleic acids has potential to deal with key candidate genes of BC, but their effect is retarded due to poor cell uptake and instability in circulation. The rapidly evolving field of nanomedicine aiming targeted drug/gene delivery curtailing BC promises to overcome the limitations of conventional therapies. Nanoparticles can be game changer for BC gene therapy as they can be effective carrier of specific drug/gene by improving the circulation time, enhancing bioavailability, reducing the immune system based recognition chances, and delivering the gene regulator accurately. Herein, we discuss the mechanism of nanoparticles targeted drug delivery, recent advancement of therapeutic strategies of nanoparticles based carriers for small interfering RNA, and microRNA, and gene augmentation therapies in BC. We also discuss the future prospect and challenges of nanoparticle-based therapies for BC.

Investigation of the Molecular Mechanisms Underlying the Antiatherogenic Actions of Kaempferol in Human THP-1 Macrophages.

Cardiovascular disease (CVD) is causing high mortality worldwide (World Health Organization-WHO, 2015). Atherosclerosis, the hardening and narrowing of arteries caused by the accumulation of fatty acids and lipids (cholesterol plaques), is a main reason of stroke, myocardial infarction, and angina. Present therapies for cardiovascular disease basically use statins such as ß-Hydroxy ß-methylglutaryl-CoA, with <70% efficacy and multiple side effects. An in vitro investigation was conducted to evaluate the impact of kaempferol, a natural medication, in an atherosclerotic cell model. We used cytotoxicity assays, Boyden chamber invasion assays, and quantitative PCR. Affymetrix microarrays were used to profile the entire transcriptome of kaempferol-treated cell lines, and Partek Genomic Suite was used to interpret the results. Kaempferol was not cytotoxic to THP-1 macrophages. In comparison to the control, kaempferol reduced monocyte migration mediated by monocyte chemotactic protein 1 (MCP-1) by 80%. The qPCR results showed a 73.7-fold reduction in MCP-1 and a 2.5-fold reduction in intercellular adhesion molecule 1 (ICAM-1) expression in kaempferol-treated cells. In interferon gamma (IFN-γ) without kaempferol and IFN-γ with kaempferol treated cells, we found 295 and 168 differentially expressed genes (DEGs), respectively. According to DEG pathway analysis, kaempferol exhibits anti-atherosclerosis and anti-inflammatory characteristics. Kaempferol is an effective and safe therapy for atherosclerosis.

Microarray Expression Profile of Myricetin-Treated THP-1 Macrophages Exhibits Alterations in Atherosclerosis-Related Regulator Molecules and LXR/RXR Pathway.

Atherosclerosis is a chronic inflammation characterized by macrophage infiltration, lipid deposition, and arterial wall thickening. Prevention of atherosclerosis by nutraceuticals is gaining attention. Myricetin, a dietary flavonol, is claimed to possess anti-atherosclerosis properties. We studied myricetin's effect on the atherosclerosis-associated molecular mechanism. Cytotoxicity and proliferation testing to check the viability of myricetin-treated THP-1 macrophages and monocyte migration study in the presence and absence of myricetin was performed. The whole transcriptome analysis was conducted using the Affymetrix microarray platform. The Partek genomics suite for detecting differentially expressed genes (DEGs) and ingenuity pathway analysis was used to identify canonical pathways. Cytotoxicity assays exhibited no significant toxicity in THP-1 macrophages treated with different myricetin concentrations (10-200 µM). Genome-wide expression profiling revealed 58 DEGs (53 upregulated and 5 downregulated) in myricetin-treated THP-1 macrophages. Pathway analysis revealed inhibition of LXR/RXR activation and angiogenesis inhibition by thrombospondin-1 and activated phagocytosis in myricetin-treated THP-1 macrophages. The cytotoxicity assay shows myricetin as a safe phytochemical. In vitro and in silico pathway studies on THP-1 macrophages showed that they can inhibit THP-1 monocyte migration and alter the cholesterol efflux mediated via LXR/RXR signaling. Therefore, myricetin could help in the prevention of cell infiltration in atherosclerotic plaque with reduced risk of stroke or brain damage.

Advancements in CRISPR/Cas9 technology-Focusing on cancer therapeutics and beyond.

"CRISPR" is an abbreviation for Clustered Regularly Interspaced Short Palindromic Repeats, which are a characteristic of the bacterial defense system and Cas9 (or "CRISPR-associated") is a RNA-guided DNA endonuclease or molecular scissor, capable of cutting DNA strands. Both together forms the basis for CRISPR-Cas9 targeted genome editing technology and enables highly specific genomic modifications to an organism's DNA. Recent advent of high-throughput genomics has revolutionizing personalized medicine and enhanced our molecular understanding of human cancers. The development of the CRISPR/Cas9 tool has unveiled advancement of new, simplistic and efficient in vivo model systems in oncology. The usage of CRISPR/Cas9 gene editing systems for curing various cancers promises to be the next great biotechnological breakthrough in medicine. However, urgent attention is needed to assess the functional relevance of novel cancer-associated mutations and translate our molecular knowledge to therapeutics. Herein, we will review the development and applications of the exciting uses of the CRISPR/Cas9 technique for cancer research and therapy with focus on origin, progress, clinical trials, implications, and challenges ahead. Major ethical and safety concerns are perhaps unknown long term consequences of DNA manipulation and irreversibility of this procedure.

Alterations of transcriptome expression, cell cycle, and mitochondrial superoxide reveal foetal endothelial dysfunction in Saudi women with gestational diabetes mellitus.

Gestational diabetes mellitus (GDM) affects one in four Saudi women and is associated with high risks of cardiovascular diseases in both the mother and foetus. It is believed that endothelial cells (ECs) dysfunction initiates these diabetic complications. In this study, differences in the transcriptome profiles, cell cycle distribution, and mitochondrial superoxide (MTS) between human umbilical vein endothelial cells (HUVECs) from GDM patients and those from healthy (control) subjects were analysed. Transcriptome profiles were generated using high-density expression microarray. The selected four altered genes were validated using qRT-PCR. MTS and cell cycle were analysed by flow cytometry. A total of 84 altered genes were identified, comprising 52 upregulated and 32 downregulated genes in GDM.HUVECs. Our selection of the four interested altered genes (TGFB2, KITLG, NEK7, and IGFBP5) was based on the functional network analysis, which revealed that these altered genes are belonging to the highest enrichment score associated with cellular function and proliferation; all of which may contribute to ECs dysfunction. The cell cycle revealed an increased percentage of cells in the G2/M phase in GDM.HUVECs, indicating cell cycle arrest. In addition, we found that GDM.HUVECs had increased MTS generation. In conclusion, GDM induces persistent impairment of the biological functions of foetal ECs, as evidenced by analyses of transcriptome profiles, cell cycle, and MTS even after ECs culture in vitro for several passages under normal glucose conditions.

A novel homozygous nonsense mutation in CCDC88A gene cause PEHO-like syndrome in consanguineous Saudi family.

Progressive encephalopathy, edema, hypsarrhythmia, and optic atrophy (PEHO) syndrome is an unusual Mendelian phenotype of unidentified origin that causes profound intellectual disability, optic nerve/cerebellar atrophy, epileptic seizures, developmental progress, pedal edema, and early death. Uncharacteristic affected individuals are often classified as having PEHO-like syndrome, although they may be misdiagnosed as having epileptic encephalopathy, a potential result of early birth. In this study, we report a consanguineous Saudi family with a novel homozygous nonsense mutation of the CCDC88A gene causing PEHO-like syndrome. The children were suffering from developmental delay, epilepsy, mental disability, optic nerve/cerebellar atrophy, and pedal edema. Whole exome sequencing was conducted for the members of the family who have the disorder to study the novel mutation. Whole exome sequencing data analysis, confirmed by subsequent Sanger sequencing validation, identified a novel homozygous nonsense mutation c. 1292G > A, which was caused by p.Trp431* stop gain. This mutation was ruled out in 100 unrelated healthy controls. The nonsense homozygous mutation detected in this study has not yet been reported as pathogenic in the literature or various databases. In conclusion, a complete loss of protein function due to premature stop gain was caused by a mutation in exon 12 of CCDC88A. This loss may lead to PEHO phenotype. CCDC88A gene may therefore play an important and critical role for multiple aspects of normal human neurodevelopment.

Role of Overexpressed Transcription Factor FOXO1 in Fatal Cardiovascular Septal Defects in Patau Syndrome: Molecular and Therapeutic Strategies.

Patau Syndrome (PS), characterized as a lethal disease, allows less than 15% survival over the first year of life. Most deaths owe to brain and heart disorders, more so due to septal defects because of altered gene regulations. We ascertained the cytogenetic basis of PS first, followed by molecular analysis and docking studies. Thirty-seven PS cases were referred from the Department of Pediatrics, King Abdulaziz University Hospital to the Center of Excellence in Genomic Medicine Research, Jeddah during 2008 to 2018. Cytogenetic analyses were performed by standard G-band method and trisomy13 were found in all the PS cases. Studies have suggested that genes of chromosome 13 and other chromosomes are associated with PS. We, therefore, did molecular pathway analysis, gene interaction, and ontology studies to identify their associations. Genomic analysis revealed important chr13 genes such as FOXO1, Col4A1, HMGBB1, FLT1, EFNB2, EDNRB, GAS6, TNFSF1, STARD13, TRPC4, TUBA3C, and TUBA3D, and their regulatory partners on other chromosomes associated with cardiovascular disorders, atrial and ventricular septal defects. There is strong indication of involving FOXO1 (Forkhead Box O1) gene-a strong transcription factor present on chr13, interacting with many septal defects link genes. The study was extended using molecular docking to find a potential drug lead for overexpressed FOXO1 inhibition. The phenothiazine and trifluoperazine showed efficiency to inhibit overexpressed FOXO1 protein, and could be potential drugs for PS/trisomy13 after validation.

Comprehensive molecular biomarker identification in breast cancer brain metastases.

BACKGROUND: Breast cancer brain metastases (BCBM) develop in about 20-30% of breast cancer (BC) patients. BCBM are associated with dismal prognosis not at least due to lack of valuable molecular therapeutic targets. The aim of the study was to identify new molecular biomarkers and targets in BCBM by using complementary state-of-the-art techniques. METHODS: We compared array expression profiles of three BCBM with 16 non-brain metastatic BC and 16 primary brain tumors (prBT) using a false discovery rate (FDR) p < 0.05 and fold change (FC) > 2. Biofunctional analysis was conducted on the differentially expressed probe sets. High-density arrays were employed to detect copy number variations (CNVs) and whole exome sequencing (WES) with paired-end reads of 150 bp was utilized to detect gene mutations in the three BCBM. RESULTS: The top 370 probe sets that were differentially expressed between BCBM and both BC and prBT were in the majority comparably overexpressed in BCBM and included, e.g. the coding genes BCL3, BNIP3, BNIP3P1, BRIP1, CASP14, CDC25A, DMBT1, IDH2, E2F1, MYCN, RAD51, RAD54L, and VDR. A number of small nucleolar RNAs (snoRNAs) were comparably overexpressed in BCBM and included SNORA1, SNORA2A, SNORA9, SNORA10, SNORA22, SNORA24, SNORA30, SNORA37, SNORA38, SNORA52, SNORA71A, SNORA71B, SNORA71C, SNORD13P2, SNORD15A, SNORD34, SNORD35A, SNORD41, SNORD53, and SCARNA22. The top canonical pathway was entitled, role of BRCA1 in DNA damage response. Network analysis revealed key nodes as Akt, ERK1/2, NFkB, and Ras in a predicted activation stage. Downregulated genes in a data set that was shared between BCBM and prBT comprised, e.g. BC cell line invasion markers JUN, MMP3, TFF1, and HAS2. Important cancer genes affected by CNVs included TP53, BRCA1, BRCA2, ERBB2, IDH1, and IDH2. WES detected numerous mutations, some of which affecting BC associated genes as CDH1, HEPACAM, and LOXHD1. CONCLUSIONS: Using complementary molecular genetic techniques, this study identified shared and unshared molecular events in three highly aberrant BCBM emphasizing the challenge to detect new molecular biomarkers and targets with translational implications. Among new findings with the capacity to gain clinical relevance is the detection of overexpressed snoRNAs known to regulate some critical cellular functions as ribosome biogenesis.

e-GRASP: an integrated evolutionary and GRASP resource for exploring disease associations.

BACKGROUND: Genome-wide association studies (GWAS) have become a mainstay of biological research concerned with discovering genetic variation linked to phenotypic traits and diseases. Both discrete and continuous traits can be analyzed in GWAS to discover associations between single nucleotide polymorphisms (SNPs) and traits of interest. Associations are typically determined by estimating the significance of the statistical relationship between genetic loci and the given trait. However, the prioritization of bona fide, reproducible genetic associations from GWAS results remains a central challenge in identifying genomic loci underlying common complex diseases. Evolutionary-aware meta-analysis of the growing GWAS literature is one way to address this challenge and to advance from association to causation in the discovery of genotype-phenotype relationships. DESCRIPTION: We have created an evolutionary GWAS resource to enable in-depth query and exploration of published GWAS results. This resource uses the publically available GWAS results annotated in the GRASP2 database. The GRASP2 database includes results from 2082 studies, 177 broad phenotype categories, and ~8.87 million SNP-phenotype associations. For each SNP in e-GRASP, we present information from the GRASP2 database for convenience as well as evolutionary information (e.g., rate and timespan). Users can, therefore, identify not only SNPs with highly significant phenotype-association P-values, but also SNPs that are highly replicated and/or occur at evolutionarily conserved sites that are likely to be functionally important. Additionally, we provide an evolutionary-adjusted SNP association ranking (E-rank) that uses cross-species evolutionary conservation scores and population allele frequencies to transform P-values in an effort to enhance the discovery of SNPs with a greater probability of biologically meaningful disease associations. CONCLUSION: By adding an evolutionary dimension to the GWAS results available in the GRASP2 database, our e-GRASP resource will enable a more effective exploration of SNPs not only by the statistical significance of trait associations, but also by the number of studies in which associations have been replicated, and the evolutionary context of the associated mutations. Therefore, e-GRASP will be a valuable resource for aiding researchers in the identification of bona fide, reproducible genetic associations from GWAS results. This resource is freely available at http://www.mypeg.info/egrasp .

Associations of recurrent miscarriages with chromosomal abnormalities, thrombophilia allelic polymorphisms and/or consanguinity in Saudi Arabia.

BACKGROUND: Recurrent pregnancy loss (RPL) or recurrent spontaneous abortion is an obstetric complication that affects couples at reproductive age. Previous reports documented a clear relationship between parents with chromosomal abnormalities and both recurrent miscarriages and infertility. However, limited data is available from the Arabian Peninsula which is known by higher rates of consanguineous marriages. The main goal of this study was to determine the prevalence of chromosomal abnormalities and thrombophilic polymorphisms, and to correlate them with RPL and consanguinity in Saudi Arabia. METHODS: Cytogenetic analysis of 171 consent patients with RPL was performed by the standard method of 72-h lymphocyte culture and GTG banding. Allelic polymorphisms of three thrombophilic genes (Factor V Leiden, Prothrombin A20210G, MTHFR C677T) were performed using PCR-RFLP (restriction fragment length polymorphism) and gel electrophoresis. RESULTS: Data analysis revealed that 7.6 % of patients were carrier of numerical or structural chromosomal abnormalities. A high rate of translocations (46 %) was associated to increased incidence of RPL. A significant correlation between consanguineous RPL patients and chromosomal abnormalities (P < 0.05) was found. Both Factor V Leiden and Prothrombin A20210G allelic polymorphisms were significantly associated with a higher prevalence of RPL. CONCLUSIONS: This study demonstrated a strong association between RPL and the prevalence of chromosomal abnormalities and inherited thrombophilia. Given the high rate of consanguineous marriages in the Saudi population, these results underline the importance of systematic cytogenetic investigation and genetic counseling preferably at the premarital stage or at least during early pregnancy phase through preimplantation genetic diagnosis (PGD).

Metformin improves the angiogenic potential of human CD34⁺ cells co-incident with downregulating CXCL10 and TIMP1 gene expression and increasing VEGFA under hyperglycemia and hypoxia within a therapeutic window for myocardial infarction.

BACKGROUND: Cardiovascular disease (CVD) is the leading cause of morbidity and mortality in patients with diabetes mellitus (DM). To identify the most effective treatment for CVD, it is paramount to understand the mechanism behind cardioprotective therapies. Although metformin has been shown to reduce CVD in Type-2 DM clinical trials, the underlying mechanism remains unexplored. CD34(+) cell-based therapies offer a new treatment approach to CVD. The aim of this study was to investigate the effect of metformin on the angiogenic properties of CD34(+) cells under conditions mimicking acute myocardial infarction in diabetes. METHODS: CD34(+) cells were cultured in 5.5 or 16.5 mmol/L glucose ± 0.01 mmol/L metformin and then additionally ± 4 % hypoxia. The paracrine function of CD34(+) cell-derived conditioned medium was assessed by measuring pro-inflammatory cytokines, vascular endothelial growth factor A (VEGFA), and using an in vitro tube formation assay for angiogenesis. Also, mRNA of CD34(+) cells was assayed by microarray and genes of interest were validated by qRT-PCR. RESULTS: Metformin increased in vitro angiogenesis under hyperglycemia-hypoxia and augmented the expression of VEGFA. It also reduced the angiogenic-inhibitors, chemokine (C-X-C motif) ligand 10 (CXCL10) and tissue inhibitor of metalloproteinase 1 (TIMP1) mRNAs, which were upregulated under hyperglycemia-hypoxia. In addition metformin, increased expression of STEAP family member 4 (STEAP4) under euglycemia, indicating an anti-inflammatory effect. CONCLUSIONS: Metformin has a dual effect by simultaneously increasing VEGFA and reducing CXCL10 and TIMP1 in CD34(+) cells in a model of the diabetic state combined with hypoxia. Therefore, these angiogenic inhibitors are promising therapeutic targets for CVD in diabetic patients. Moreover, our data are commensurate with a vascular protective effect of metformin and add to the understanding of underlying mechanisms.

Cyclin D1 as a therapeutic target of renal cell carcinoma- a combined transcriptomics, tissue microarray and molecular docking study from the Kingdom of Saudi Arabia.

BACKGROUND: Renal cell carcinoma (RCC) is a seventh ranked malignancy with poor prognosis. RCC is lethal at metastatic stage as it does not respond to conventional systemic treatments, and there is an urgent need to find out promising novel biomarkers for effective treatment. The goal of this study was to evaluate the biomarkers that can be potential therapeutic target and predict effective inhibitors to treat the metastatic stage of RCC. METHODS: We conducted transcriptomic profiling to identify differentially expressed genes associated with RCC. Molecular pathway analysis was done to identify the canonical pathways and their role in RCC. Tissue microarrays (TMA) based immunohistochemical stains were used to validate the protein expression of cyclinD1 (CCND1) and were scored semi-quantitatively from 0 to 3+ on the basis of absence or presence of staining intensity in the tumor cell. Statistical analysis determined the association of CCND1 expression with RCC. Molecular docking analyses were performed to check the potential of two natural inhibitors, rutin and curcumin to bind CCND1. RESULTS: We detected 1490 significantly expressed genes (1034, upregulated and 456, downregulated) in RCC using cutoff fold change 2 and p value < 0.05. Hes-related family bHLH transcription factor with YRPW motif 1 (HEY1), neuropilin 2 (NRP2), lymphoid enhancer-binding factor 1 (LEF1), and histone cluster 1 H3h (HIST1H3H) were most upregulated while aldolase B, fructose-bisphosphate (ALDOB), solute carrier family 12 (SLC12A1), calbindin 1 (CALB1) were the most down regulated genes in our dataset. Functional analysis revealed Wnt/ß-catenin signaling as the significantly activated canonical pathway (z score = 2.53) involving cyclin D1 (CCND1). CCND1 was overexpressed in transcriptomic studies (FC = 2.26, p value = 0.0047) and TMA results also showed the positive expression of CCND1 in 53 % (73/139) of RCC cases. The ligands - rutin and curcumin bounded with CCND1 with good affinity. CONCLUSION: CCND1 was one of the important upregulated gene identified in microarray and validated by TMA. Docking study showed that CCND1 may act as a potential therapeutic target and its inhibition could focus on the migratory, invasive, and metastatic potential of RCC. Further in vivo and in vitro molecular studies are needed to investigate the therapeutic target potential of CCND1 for RCC treatment.

Assessment of Radiation Induced Therapeutic Effect and Cytotoxicity in Cancer Patients Based on Transcriptomic Profiling.

Toxicity induced by radiation therapy is a curse for cancer patients undergoing treatment. It is imperative to understand and define an ideal condition where the positive effects notably outweigh the negative. We used a microarray meta-analysis approach to measure global gene-expression before and after radiation exposure. Bioinformatic tools were used for pathways, network, gene ontology and toxicity related studies. We found 429 differentially expressed genes at fold change >2 and p-value <0.05. The most significantly upregulated genes were synuclein alpha (SNCA), carbonic anhydrase I (CA1), X-linked Kx blood group (XK), glycophorin A and B (GYPA and GYPB), and hemogen (HEMGN), while downregulated ones were membrane-spanning 4-domains, subfamily A member 1 (MS4A1), immunoglobulin heavy constant mu (IGHM), chemokine (C-C motif) receptor 7 (CCR7), BTB and CNC homology 1 transcription factor 2 (BACH2), and B-cell CLL/lymphoma 11B (BCL11B). Pathway analysis revealed calcium-induced T lymphocyte apoptosis and the role of nuclear factor of activated T-cells (NFAT) in regulation of the immune response as the most inhibited pathways, while apoptosis signaling was significantly activated. Most of the normal biofunctions were significantly decreased while cell death and survival process were activated. Gene ontology enrichment analysis revealed the immune system process as the most overrepresented group under the biological process category. Toxicity function analysis identified liver, kidney and heart to be the most affected organs during and after radiation therapy. The identified biomarkers and alterations in molecular pathways induced by radiation therapy should be further investigated to reduce the cytotoxicity and development of fatigue.

Effect of BRAF mutational status on expression profiles in conventional papillary thyroid carcinomas.

BACKGROUND: Whereas 40 % to 70 % of papillary thyroid carcinomas (PTCs) are characterized by a BRAF mutation (BRAFmut), unified biomarkers for the genetically heterogeneous group of BRAF wild type (BRAFwt) PTCs are not established yet. Using state-of-the-art technology we compared RNA expression profiles between conventional BRAFwt and BRAFmut PTCs. METHODS: Microarrays covering 36,079 reference sequences were used to generate whole transcript expression profiles in 11 BRAFwt PTCs including five micro PTCs, 14 BRAFmut PTCs, and 7 normal thyroid specimens. A p-value with a false discovery rate (FDR) < 0.05 and a fold change > 2 were used as a threshold of significance for differential expression. Network and pathway utilities were employed to interpret significance of expression data. BRAF mutational status was established by direct sequencing the hotspot region of exon 15. RESULTS: We identified 237 annotated genes that were significantly differentially expressed between BRAFwt and BRAFmut PTCs. Of these, 110 genes were down- and 127 were upregulated in BRAFwt compared to BRAFmut PTCs. A number of molecules involved in thyroid hormone metabolism including thyroid peroxidase (TPO) were differentially expressed between both groups. Among cancer-associated molecules were ERBB3 that was downregulated and ERBB4 that was upregulated in BRAFwt PTCs. Two microRNAs were significantly differentially expressed of which miR492 bears predicted functions relevant to thyroid-specific molecules. The protein kinase A (PKA) and the G protein-coupled receptor pathways were identified as significantly related signaling cascades to the gene set of 237 genes. Furthermore, a network of interacting molecules was predicted on basis of the differentially expressed gene set. CONCLUSIONS: The expression study focusing on affected genes that are differentially expressed between BRAFwt and BRAFmut conventional PTCs identified a number of molecules which are connected in a network and affect important canonical pathways. The identified gene set adds to our understanding of the tumor biology of BRAFwt and BRAFmut PTCs and contains genes/biomarkers of interest.

Comparison of microarray expression profiles between follicular variant of papillary thyroid carcinomas and follicular adenomas of the thyroid.

BACKGROUND: Follicular variant of papillary thyroid carcinoma (FVPTC) and follicular adenoma (FA) are histologically closely related tumors and differential diagnosis remains challenging. RNA expression profiling is an established method to unravel molecular mechanisms underlying the histopathology of diseases. METHODS: BRAF mutational status was established by direct sequencing the hotspot region of exon 15 in six FVPTCs and seven FAs. Whole-transcript arrays were employed to generate expression profiles in six FVPTCs, seven FAs and seven normal thyroid tissue samples. The threshold of significance for differential expression on the gene and exon level was a p-value with a false discovery rate (FDR) < 0.05 and a fold change cutoff > 2. Two dimensional average linkage hierarchical clustering was generated using differentially expressed genes. Network, pathway, and alternative splicing utilities were employed to interpret significance of expression data on the gene and exon level. RESULTS: Expression profiling in FVPTCs and FAs, all of which were negative for a BRAF mutation, revealed 55 transcripts that were significantly differentially expressed, 40 of which were upregulated and 15 downregulated in FVPTCs vs. FAs. Amongst the most significantly upregulated genes in FVPTCs were GABA B receptor, 2 (GABBR2), neuronal cell adhesion molecule (NRCAM), extracellular matrix protein 1 (ECM1), heparan sulfate 6-O-sulfotransferase 2 (HS6ST2), and retinoid X receptor, gamma (RXRG). The most significantly downregulated genes in FVPTCs included interaction protein for cytohesin exchange factors 1 (IPCEF1), G protein-coupled receptor 155 (GPR155), Purkinje cell protein 4 (PCP4), chondroitin sulfate N-acetylgalactosaminyltransferase 1 (CSGALNACT1), and glutamate receptor interacting protein 1 (GRIP1). Alternative splicing analysis detected 87 genes, 52 of which were also included in the list of 55 differentially expressed genes. Network analysis demonstrated multiple interactions for a number of differentially expressed molecules including vitamin D (1,25- dihydroxyvitamin D3) receptor (VDR), SMAD family member 9 (SMAD9), v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT), and RXRG. CONCLUSIONS: This is one of the first studies using whole-transcript expression arrays to compare expression profiles between FVPTCs and FAs. A set of differentially expressed genes has been identified that contains valuable candidate genes to differentiate both histopathologically related tumor types on the molecular level.

Transcriptomics profiling study of breast cancer from Kingdom of Saudi Arabia revealed altered expression of Adiponectin and Fatty Acid Binding Protein4: Is lipid metabolism associated with breast cancer?

BACKGROUND: Breast cancer incidence rates are increasing at an alarming rate among Saudi Arabian females. Most molecular genetic discoveries on breast cancer and other cancers have arisen from studies examining European and American patients. However, possibility of specific changes in molecular signature among cancer patients of diverse ethnic groups remains largely unexplored. We performed transcriptomic profiling of surgically-resected breast tumors from 45 patients based in the Western region of Saudi Arabia using Affymetrix Gene 1.0 ST chip. Pathway and biological function-based clustering was apparent across the tissue samples. RESULTS: Pathway analysis revealed canonical pathways that had not been previously implicated in breast cancer. Biological network analysis of differentially regulated genes revealed that Fatty acid binding protein 4, adipocyte (FABP4), adiponectin (ADIPOQ), and retinol binding protein 4 (RBP4) were most down regulated genes, sharing strong connection with the other molecules of lipid metabolism pathway. The marked biological difference in the signatures uncovered between the USA and Saudi samples underpins the importance of this study. Connectivity Map identified compounds that could reverse an observed gene expression signature CONCLUSIONS: This study describes, to our knowledge, the first genome-wide profiling of breast cancer from Saudi ethnic females. We demonstrate the involvement of the lipid metabolism pathway in the pathogenesis of breast cancer from this region. This finding also highlights the need for strategies to curb the increasing rates of incidence of this disease by educating the public about life-style risk factors such as unhealthy diet and obesity.

KDM3A knockdown regulates COMP, LOX, COL8A1 and ACOT1 genes in myocardial fibrosis.

Cardiovascular disease (CVD) is one of the main causes of death in Saudi Arabia. Cardiac remodeling plays a critical role in the pathophysiology of heart failure. Major focus of our study was to identify crucial genes involved in the pathological remodeling of the heart caused by pressure overload. We utilized various in-silico tools to analyze and interpret microarray data obtained from the Gene Expression Omnibus (GEO) database (GSE120739), including GEO2R analysis, Metascape analysis, WebGestalt analysis, and IPA (Ingenuity pathway analysis). Our findings indicate that certain genes, including Cartilage Oligomeric Matrix Protein (COMP), collagen type VIII alpha 1 chain (COL8A1) and Lysyl Oxidase (LOX) under the influence caused by knockdown of KDM3A, were down regulated by the extracellular matrix pathway. Moreover, genes, such as Acyl-CoA Thioesterase 1 (ACOT1) were up regulated by the fatty acid metabolism pathway. Overexpression of lysine-specific demethylase 3A (KDM3A) leads to the up regulation of fibrosis-related genes COMP, COL8A1, and LOX and the down regulation of ACOT1, result in enhanced fibrosis and heart failure. Our results suggest that COMP, COL8A1, LOX, and ACOT1 warrant further investigation in the development of cardiac fibrosis and as potential biomarkers for causing heart failure.

Uncovering hidden genetic risk factors for breast and ovarian cancers in BRCA-negative women: a machine learning approach in the Saudi population.

Breast and ovarian cancers are prevalent worldwide, with genetic factors such as BRCA1 and BRCA2 mutations playing a significant role. However, not all patients carry these mutations, making it challenging to identify risk factors. Researchers have turned to whole exome sequencing (WES) as a tool to identify genetic risk factors in BRCA-negative women. WES allows the sequencing of all protein-coding regions of an individual's genome, providing a comprehensive analysis that surpasses traditional gene-by-gene sequencing methods. This technology offers efficiency, cost-effectiveness and the potential to identify new genetic variants contributing to the susceptibility to the diseases. Interpreting WES data for disease-causing variants is challenging due to its complex nature. Machine learning techniques can uncover hidden genetic-variant patterns associated with cancer susceptibility. In this study, we used the extreme gradient boosting (XGBoost) and random forest (RF) algorithms to identify BRCA-related cancer high-risk genes specifically in the Saudi population. The experimental results exposed that the RF method scored superior performance with an accuracy of 88.16% and an area under the receiver-operator characteristic curve of 0.95. Using bioinformatics analysis tools, we explored the top features of the high-accuracy machine learning model that we built to enhance our knowledge of genetic interactions and find complex genetic patterns connected to the development of BRCA-related cancers. We were able to identify the significance of HLA gene variations in these WES datasets for BRCA-related patients. We find that immune response mechanisms play a major role in the development of BRCA-related cancer. It specifically highlights genes associated with antigen processing and presentation, such as HLA-B, HLA-A and HLA-DRB1 and their possible effects on tumour progression and immune evasion. In summary, by utilizing machine learning approaches, we have the potential to aid in the development of precision medicine approaches for early detection and personalized treatment strategies.