Unveiling Arformoterol as a potent LSD1 inhibitor for breast cancer treatment: A comprehensive study integrating 3D-QSAR pharmacophore modeling, molecular docking, molecular dynamics simulations and in vitro assays.

Lysine Specific Demethylase 1 (LSD1) has been identified as a chromatin-modifying enzyme implicated in various cancer pathogeneses, highlighting the potential for novel epigenetic cancer treatments through the development of effective inhibitors. We employed 3D-QSAR pharmacophore modeling, molecular docking, and molecular dynamics simulations to identify a promising drug candidate for LSD1 inhibition. RMSD, RMSF, H-bond, and DSSP analysis demonstrated that ZINC02599970 (Arformoterol) and ZINC13453966 exhibited the highest LSD1 inhibitory potential. Experimental validation using MCF-7 and MDA-MB-231 cell lines revealed that Arformoterol displayed potent antiproliferative activity with IC50 values of 12.30 ± 1.48 µM and 19.69 ± 1.15 µM respectively. In contrast, the IC50 values obtained for the control (tranylcypromine) in exposure to MCF-7 and MDA-MB-231 cells were 104.6 ± 1.69 µM and 77 ± 0.67 µM, respectively. Arformoterol demonstrated greater LSD1 inhibitory potency in MCF-7 cells compared to MDA-MB-231 cells. Also, the expression of genes involved in chromatin rearrangement (LSD1), angiogenesis (VEGF1), cell migration (RORα), signal transduction (S100A8), apoptosis, and cell cycle (p53) were investigated. Arformoterol enhanced apoptosis and induced cell cycle arrest at the G2/M phase, both in MCF-7 and MDA-MB-231 cancer cells. Based on our findings, we propose that Arformoterol represents a promising candidate for breast cancer treatment, owing to its potent LSD1 inhibitory activity.

The identification of novel inhibitors of human angiotensin-converting enzyme 2 and main protease of Sars-Cov-2: A combination of in silico methods for treatment of COVID-19.

The angiotensin-converting enzyme 2 (ACE2) and main protease (MPro), are the putative drug candidates for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this study, we performed 3D-QSAR pharmacophore modeling and screened 1,264,479 ligands gathered from Pubchem and Zinc databases. Following the calculation of the ADMET properties, molecular docking was carried out. Moreover, the de novo ligand design was performed. MD simulation was then applied to survey the behavior of the ligand-protein complexes. Furthermore, MMPBSA has utilized to re-estimate the binding affinities. Then, a free energy landscape was used to find the most stable conformation of the complexes. Finally, the hybrid QM-MM method was carried out for the precise calculation of the energies. The Hypo1 pharmacophore model was selected as the best model. Our docking results indicate that the compounds ZINC12562757 and 112,260,215 were the best potential inhibitors of the ACE2 and MPro, respectively. Furthermore, the Evo_1 compound enjoys the highest docking energy among the designed de novo ligands. Results of RMSD, RMSF, H-bond, and DSSP analyses have demonstrated that the lead compounds preserve the stability of the complexes' conformation during the MD simulation. MMPBSA data confirmed the molecular docking results. The results of QM-MM showed that Evo_1 has a stronger potential for specific inhibition of MPro, as compared to the 112,260,215 compound.

Harnessing CRISPR/Cas9 technology in cardiovascular disease.

The CRISPR/Cas9 system is a precisely targeted bacterial defense system, used to control invading viruses. This technology has many potential applications including genetic changes in somatic and germ cells and the creation of knockout animals. Compared to other genome editing techniques such as zinc-finger nucleases and transcription activator-like effector nucleases (TALENS), the CRISPR/Cas9 system is much easier and more efficient. Most importantly, the multifunctional capacity of this technology allows simultaneous editing of several genes. The CRISPR/Cas9 system also potentially has the ability to prevent and treat human diseases. The present article addresses some key points related to the use of the CRISPR/Cas9 system as a powerful tool in cardiovascular research and as a new strategy for the treatment of cardiovascular disease (CVD).

CRISPR-Cas9 in genome editing: Its function and medical applications.

The targeted genome modification using RNA-guided nucleases is associated with several advantages such as a rapid, easy, and efficient method that not only provides the manipulation and alteration of genes and functional studies for researchers, but also increases their awareness of the molecular basis of the disease and development of new and targeted therapeutic approaches. Different techniques have been emerged so far as the molecular scissors mediating targeted genome editing including zinc finger nuclease, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9). CRISPR-Cas9 is a bacterial immune system against viruses in which the single-strand RNA-guided Cas9 nuclease is linked to the targeted complementary sequences to apply changes. The advances made in the transfer, modification, and emergence of specific solutions have led to the creation of different classes of CRISPR-Cas9. Since this robust tool is capable of direct correction of disease-causing mutations, its ability to treat genetic disorders has attracted the tremendous attention of researchers. Considering the reported cases of nonspecific targeting of Cas9 proteins, many studies focused on enhancing the Cas9 features. In this regard, significant advances have been made in choosing guide RNA, new enzymes and methods for identifying misplaced targeting. Here, we highlighted the history and various direct aspects of CRISPR-Cas9, such as precision in genomic targeting, system transfer and its control over correction events with its applications in future biological studies, and modern treatment of diseases.

Anti-inflammatory Effects of Valproic Acid in a Rat Model of Renal Ischemia/Reperfusion Injury: Alteration in Cytokine Profile.

Valporic acid (VPA) has been implicated to have anti-inflammatory and anti-oxidant activities in several ischemic/reperfusion (I/R) injury models. This study intended to evaluate whether VPA could affect the inflammatory/anti-inflammatory cytokines balance and severity of renal I/R injury in rat. I/R injury was induced in two groups of animals, vehicle normal saline and VPA-treated (IP injection, 150 mg/kg) rats, by 45 min occlusion of both left and right renal arteries followed by 3, 24 and 120 h reperfusion in separate groups. After each time point, kidneys and blood samples were collected for cytokine genes (TNF-α, IL-1ß, IL-10 and TGF-ß) expression analysis and histological examinations in the kidney tissues. Serum creatinine levels were measured for evaluation of renal function. We observed significantly downregulated mRNA expressions for IL-1ß and TNF-α in blood and tissue samples 24 and 120 h post I/R injury in VPA-treated animals compared to control groups (P < 0.0001). On the other hand, mRNA expression levels for IL-10 and TGF-ß were significantly increased in the blood samples from VPA-treated animals at two time points after I/R injury (P < 0.0001) and at 120 h in tissue samples (P < 0.001). Histopathology analysis showed downgraded ischemic changes in VPA group compared to sham control. Also, decreased serum creatinine levels were observed in VPA-treated animals particularly 120 h post I/R injury (P < 0.0001) that was correlated with less pathological changes in this group. Our results indicate that VPA can attenuate pro-inflammatory responses and augment the anti-inflammatory condition in favor of faster renal recovery from ischemic changes and improved renal function after renal I/R injury.

Cytotoxic T Lymphocyte Antigen-4 Gene Variants in Type 2 Diabetic Patients with or without Neuropathy.

Many studies have shown that cytotoxic T lymphocyte antigen-4 (CTLA-4) gene variants are associated with several autoimmune diseases particularly type 1 diabetes. Due to the lack of consistent data for this association with type 2 diabetes (T2D), this study explored the possible influence of CTLA-4 gene polymorphisms at -1722 (T/C), -318 (C/T), and +49 (G/A) positions for susceptibility to T2D in relation with neuropathy. One hundred and eleven unrelated patients with T2D [49 patients with diabetic peripheral neuropathy (DPN) and 62 patients without PDN] and 100 healthy ethnic- and gender-matched controls were included in this study. The dimorphisms at -1722 (C/T), -318 (C/T) and +49 (A/G) for CTLA-4 gene were determined using ARMS-PCR. The CTLA-4 (+49 G/G) and (+49 A/A) genotypes were found to be positively and negatively associated with T2D, respectively (p=0.03). The -318 C/T and T/T genotypes were more frequent in patients than controls and -318 C/C genotype was shown to be protective for T2D (p=0.003). ACT and GTT Haplotypes were less and more frequent in controls and patients, respectively (p=3.86×10-7 and p=2.29×10-5). Genotypes distribution among T2D patients with and without DPN compared to healthy controls showed significantly lower frequencies for -318 C/C and +49 A/A genotypes and significantly higher frequencies for -318 C/T and T/T genotypes as well. Our findings indicate that CTLA-4 (+49 A/G) and (-318 C/T) genotypes could be considered as genetic risk factors associated with susceptibility or protection for T2D.

Lack of association between rs1800795 (-174 G/C) polymorphism in the promoter region of interleukin-6 gene and susceptibility to type 2 diabetes in Isfahan population.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is an inflammatory autoimmune disease that mostly affects older adults. The etiology of T2DM includes both genetic and environmental factors. rs1800795 (-174 G/C) single nucleotide polymorphism (SNP) linked with autoimmune disorders predispositions, identified by Genome-Wide Association Study among genes, which immunologically related is considerably over signified. The goal of this study was to evaluate the association between rs1800795 (-174 G/C) polymorphisms in the promoter of interleukin-6 (IL-6) gene with susceptibility to T2DM in a subset of the Iranian population. MATERIALS AND METHODS: In this case-control study, 120 healthy subjects and 120 patients with T2DM were included. Genomic DNA obtained from whole blood samples and the polymerase chain reaction was used to amplify the fragment of interest contain rs1800795 SNP, restriction fragment length polymorphism method was applied for genotyping of the DNA samples with NlaIII as a restriction enzyme. SPSS for Windows software (version 18.0, SPSS, Chicago, IL, USA) was performed for statistical analysis. RESULTS: No significant differences were found between healthy controls and T2DM patients with respect to the frequency distribution of the cytokine gene polymorphism investigated. Odds ratio, adjusted for sex, age, and smoking status has displayed similar outcomes. CONCLUSION: These results indicated that the rs1800795 SNP is not a susceptibility gene variant for the development of T2DM in the Isfahan population. Further studies using new data on complex transcriptional interactions between IL-6 polymorphic sites are necessary to determine IL-6 haplotype influence on susceptibility to T2DM.

HLA class II alleles and risk for peripheral neuropathy in type 2 diabetes patients.

The potential impact of human leukocyte antigen (HLA) genotype variations on development of diabetic peripheral neuropathy (DPN) is not well determined. This study aimed to identify the association of HLA class II alleles with DPN in type 2 diabetes (T2D) patients. Totally 106 T2D patients, 49 with DPN and 57 without DPN, and 100 ethnic-matched healthy controls were analyzed. Both groups of the patients were matched based on sex, age, body mass index (BMI) and duration of T2D. Polyneuropathy was diagnosed using electrodiagnostic methods. HLA-DRB1 and DQB1 genotyping was performed in all subjects by the polymerase chain reaction with sequence-specific primers (PCR-SSP) method. T2D patients with DPN showed higher frequencies of HLA-DRB1*10 and DRB1*12 alleles compared to control group (P = 0.04). HLA-DQB1*02 allele and HLA-DRB1*07-DQB1*02 haplotype were associated with a decreased risk for developing DPN in T2D patients (P = 0.02 and P = 0.05 respectively). Also, patients with severe neuropathy showed higher frequencies of DRB1*07 (P = 0.003) and DQB1*02 (P = 0.02) alleles than those with mild-to-moderate form of neuropathy. The distribution of DRB1 and DQB1 alleles and haplotypes were not statistically different between all patients and healthy controls. Our findings implicate a possible protective role of HLA-DQB1*02 allele and HLA-DRB1*07-DQB1*02 haplotype against development of peripheral neuropathy in T2D patients. Therefore, variations in HLA genotypes might be used as genetic markers for prediction and potentially management of neuropathy in T2D patients.

Association between the apelin rs2235306 gene polymorphism and metabolic syndrome.

AIM: The rs2235306 of apelin polymorphism has been shown to be associated with fasting plasma glucose levels and hypertension. The present study aimed to investigate the impact of the apelin rs2235306 gene polymorphism on the risk of metabolic syndrome (MeS) in a sample of the Iranian population. MATERIALS AND METHODS: This population-based cross-sectional study was performed on 151 subjects with MeS and 149 without MeS, as defined by ATPIII criteria. Apelin rs2235306 polymorphism detection was done using the tetra amplification refractory mutation system-polymerase chain reaction. Because the apelin gene is located on the X chromosome, statistical analyses were conducted in a sex-specific manner. RESULTS: Our findings proposed that the apelin rs2235306 polymorphism was not associated with MeS susceptibility in the codominant, dominant, and recessive inheritance models tested (OR = 0.93, 95% CI = 0.51-1.71 for TC vs. TT; OR = 2.39, 95% CI = 0.70-8.16 CC vs. TT; OR = 1.09, 95% CI = 0.62-1.93 for TC+CC vs. TT; and OR = 2.45, 95% CI = 0.73-8.21 for CC vs. TT+TC). We found that the apelin TC+CC genotypes were associated with lower HDL-cholesterol in women without MeS. CONCLUSION: Our findings indicated no association between the apelin rs2235306 polymorphism and MeS. However, the results suggest that healthy females carrying apelin TC+CC genotypes have lower HDL-cholesterol in comparison with those carrying TT, which remains to be confirmed.

A 45-bp insertion/deletion polymorphism of UCP2 gene is associated with metabolic syndrome.

BACKGROUND: Metabolic syndrome (MeS) is being recognized as a risk factor for insulin resistance and cardiovascular disease. The present study was aimed to find out the possible association between 45-bp I/D polymorphism of uncoupling protein 2 (UCP2) and MeS. METHODS: DNA was extracted from peripheral blood of 151 subjects with and 149 subjects without MeS. 45-bp I/D variant of UCP2 was detected using polymerase chain reaction (PCR). RESULTS: Our finding showed that 45-bp I/D polymorphism was associated with protection against MeS (OR = 0.56, 95% CI = 0.34-0.92, p = 0.020 D/I vs DD and OR = 0.54, 95% CI = 0.34-0.86, p = 0.009; D/I + I/I vs D/D). The I allele decreased the risk of MeS (OR = 0.62, 95% CI = 0.44-0.90, p = 0.011) in comparison with D allele. CONCLUSION: In conclusion, our result suggests that 45-bp I/D polymorphism is associated with the risk of MeS, which remains to be cleared.

Association of promoter methylation and 32-bp deletion of the PTEN gene with susceptibility to metabolic syndrome.

Metabolic syndrome (MeS), a cluster of several metabolic disorders, is increasingly being recognized as a risk factor for type II diabetes (T2D) and cardiovascular disease. Genetic and epigenetic alteration of the phosphatase and tensin homolog deleted on chromosome ten (PTEN) has been associated with components of MeS. The aim of the present study was to investigate the possible association of a 32-bp deletion polymorphism and promoter methylation of the PTEN gene with MeS. DNA was extracted from the peripheral blood of 151 subjects with and 149 subjects without MeS. The 32-bp deletion variant of PTEN was detected by polymerase chain reaction (PCR) and PTEN promoter methylation was defined by a nested methylation­specific PCR (MSP) method. No significant differences were found in the allelic and genotypic frequencies of the 32-bp deletion variant of PTEN between the groups [odds ratio (OR), 0.77; 95% confidence interval (CI), 0.41-1.45; P=0.431]. However, patients with MeS were identified to have lower levels of PTEN promoter hypermethylation than subjects without MeS. Promoter methylation may be a protective factor against susceptibility to MeS (OR, 0.52; 95% CI, 0.29-0.92; P=0.029). Our findings suggest that PTEN promoter methylation may be a mechanism for PTEN downregulation or silencing in MeS, which remains to be fully clarified.

Association between polymorphisms of glutathione S-transferase genes (GSTM1, GSTP1 and GSTT1) and breast cancer risk in a sample Iranian population.

AIM: Genetic and environmental factors are risk factors for breast cancer. Our aim was to investigate the associations between genetic polymorphism of GST genes (GSTM1, GSTT1 and GSTP1) and susceptibility to breast cancer in an Iranian population. MATERIALS & METHODS: This case-control study was carried out on 134 patients with breast cancer and 152 healthy, cancer-free women. GSTP1 polymorphism was determined using tetra-primer amplification refractory mutation system PCR assay and GSTM1 and GSTT1 were genotyped by a multiplex PCR. RESULTS: We found that the GSTM1 null genotype is a risk factor for predisposition to breast cancer (odds ratio [OR] = 2.01; 95% CI = 1.78-3.45; p = 0.010). No significant difference was found between the groups regarding GSTT1 null genotype (p > 0.05). The GSTP1 Ile/Val and Val/Val genotypes were associated with breast cancer risk (OR = 3.29; 95% CI = 1.84-5.91; p < 0.0001 and OR = 20.68; 95% CI = 5.66-75.60; p < 0.0001, respectively). CONCLUSION: In summary, GSTM1 and GSTP1, but not GSTT1 genetic polymorphisms are associated with increased risk of breast cancer in our population.

Evaluation of UDP-glucuronosyltransferase 2B17 (UGT2B17) and dihydrofolate reductase (DHFR) genes deletion and the expression level of NGX6 mRNA in breast cancer.

The present study was aimed to investigate the possible association between 19-base pair (bp) deletion polymorphism of the DHFR gene (rs70991108), null genotype of UDP-glucuronosyltransferase 2B17 (UGT2B17) as well as the expression level of nasopharyngeal carcinoma-associated gene 6 (NGX6) with the risk of breast cancer. This case-control study was done on 236 patients with breast cancer and 203 cancer free women. Detection of 19-bp del of DHFR was done using bi-directional PCR allele-specific amplification and UGT2B17 genotyping was performed using multiplex PCR assay. NGX6 mRNA expression level was determined by quantitative reverse transcriptase PCR in 62 breast cancerous and 62 adjacent non-cancerous tissues. Our finding showed an association between null genotype of UGT2B17 and risk of breast cancer and the null genotype increased susceptibility to breast cancer (OR: 2.99; 95 % CI: 1.94-4.60; p < 0.0001). However, no statistically significant difference was found between breast cancer patients and cancer free normal women regarding 19-bp ins/del of DHFR (χ(2) = 0.91, p = 0.63). Real-time PCR data showed that the relative expression level of NGX6 mRNA was significantly lower in cancerous than that in non-cancerous breast tissue specimens (0.936 ± 0.042 and 1.042 ± 0.039, respectively). However, NGX6 mRNA expression was not correlated with tumors grade (p > 0.05). In conclusion, the null genotype of UGT2B17 revealed to be a risk factor for breast cancer in a sample of Iranian population. Furthermore, down-regulation of NGX6 mRNA expression in breast carcinoma confirms the growing proof regarding the tumor suppressor role of NGX6.

Association between chemerin rs17173608 and vaspin rs2236242 gene polymorphisms and the metabolic syndrome, a preliminary report.

Metabolic syndrome (MeS) is associated with increased risk for type 2 diabetes mellitus (T2DM) and cardiovascular disease. There is some evidence indicating that adipokines play a role in the development of MeS. The present study was aimed to investigate the impact of chemerin rs17173608 and vaspin rs2236242 gene polymorphisms with the risk of MeS in a sample of Iranian population. This case-control study was done on 151 subjects with MeS and 149 without MeS, as defined by NCEP-ATPIII. Tetra amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR) was designed to detect the polymorphisms. Our finding showed that there are significant differences in genotype frequencies between the groups regarding vaspin rs2236242 polymorphism (χ(2)=18.74, p<0.0001). A significant protection against MeS was found for vaspin rs2236242 in allele and genotypes (Odd Ratio [OR]=0.52; 95% confidence interval [CI]=0.37-0.72; p=0.0001, T vs A; OR=0.49; 95%CI=0.29-0.82; p=0.007, TT vs TA and OR=0.17; 95%CI=0.07-0.40; p<0.0001, TT vs AA). Our finding showed positive association between chemerin rs17173608 polymorphism and risk of MeS (χ(2)=7.70, p=0.021). The G allele increased the risk of MeS (OR=1.78; 95% CI=1.14-2.78; p=0.012) as compared to the T allele. In conclusion, our data suggest for the first time a significant association between vaspin rs2236242 and chemerin rs17173608 polymorphisms and the MeS in Zahedan, southeast Iran. Further studies with large sample size and different ethnicities are required to confirm our findings.

Bi-directional PCR allele-specific amplification (bi-PASA) for detection of caspase-8 -652 6N ins/del promoter polymorphism (rs3834129) in breast cancer.

Caspase-8 (CASP8) plays a critical role in regulating apoptosis, and its functional polymorphisms may modify cancer risk. We investigated the possible association between CASP8 -652 6N ins/del (rs3834129) and the risk of breast cancer in a sample of Iranian population. This case-control study was done on 236 breast cancer patients and 203 cancer free healthy female. We designed a rapid and simple bi-directional PCR allele-specific amplification (bi-PASA) for detection of CASP8 -652 6N ins/del polymorphism. The results showed that the CASP8 -652 6N del/dl genotype was inversely associated with breast cancer risk (OR=0.33, 95% CI=0.17-0.65, p=0.001). The frequencies of the del allele in cases and controls were 29.1% and 38.6%, respectively. An inverse association between CASP8 6N del variant and the risk of breast cancer (OR=0.66, 95% CI=0.66-0.87, p=0.002) was found. In conclusion, the result suggests that the CASP8 -652 6N del polymorphism plays a protective role in susceptibility to breast cancer in our population. Further studies in other populations with larger samples are needed to confirm these findings.