FTIR SPECTROSCOPIC TRENDS AND DNA DAMAGE IN RABBIT LENS DUE TO LONG RUN OF TAMOXIFEN TREATMENT

Objective: The aim of the present work is to evaluate the molecular structure changes of the lens of rabbits and DNA damage of epithelial cells due to tamoxifen administration. Methods: Twenty four healthy New Zealand white rabbits were divided into 2 main groups. The first group is served as control (n=12) kept untreated, second one is Tamoxifen administrative group (n=12) received orally daily dose of 15 mg/kg. Rabbits were decapitated after 2, 4, 6 and 8 mo, respectively. Using fourior transform infrared (FTIR) to study the molecular structure changes due to tamoxifen and comet assay analysis for discovering DNA damage. Results: FTIR data indicated that tamoxifen affects structural components in NHOH and fingerprint region. Increases of β-turns of the protein secondary structure while, reducing the content of both α-helix after 8 mo and Turns appeared for all periods of administrative tamoxifen were observed. On the other hand tamoxifen induced a statistically significant increase in comet assay parameters as tail moment compared to control animals that indicated DNA damage due to single or double strand break. Conclusion: Tamoxifen uses for more than 6 mo may lead to changes in the molecular structure of the lens and damage of DNA cells. An ophthalmic baseline examination prior to anti-cancer treatment may help detect any pre-existing ocular condition and lead to reduction of ocular side effects when predisposed patients are screened and examined regularly during and after chemotherapeutic therapy

Optimization of the Formulation and Design of Oligonucleotide-based Pharmaceuticals for the Purpose of Gene Therapy

Oligonucleotides (ONs) are short sequences of nucleic acids which may be used in a therapeutic context to modulate gene expression. According to their target, ONs can be classified into two main classes: antisense ONs which target mRNA and antigene ONs that target chromosomal DNA. In order to be pharmaceutically efficient, both kinds of ONs have to possess enough stability against degrading enzymes and rapid clearance. They must pass the cell membrane, and in some cases the nuclear membrane, and bind with enough specificity and high affinity to their target site to successfully exert their desired effect. In fact, the use of natural nucleic acids as drugs is hindered by both their inherent instability in biological fluids and their highly charged nature, which hampers their cellular uptake. Therefore, research in the field of ON-based pharmaceuticals focuses on two main strategies: chemical modification of nucleic acids to produce analogues with better stability and binding properties, and development of delivery systems to further stabilize the ONs and enhance their cellular uptake. Splice-switching antisense ONs (SSOs) made of phosphorothioate 2’-O-methyl RNA are promising therapeutics for several disorders caused by aberrant splicing. However, as other ONs, their usefulness is hindered by the lack of efficient delivery. In the first study of this thesis, four amino acid modified versions of the well-known polycation polyethylenimine (PEI) were evaluated for the formulation and delivery of SSOs. The formulations were physically characterized via assessment of their particle size and stability and this characterization was then correlated to their splice-correction efficiency after transfection into mammalian cells. Tyrosine-modified PEI (PEIY) was identified as a successful delivery system for SSOs as shown by splice-correction efficiency of 80% measured in HeLa705; a cell-model containing a mutated –globin intron sequence found in –thalassemia splicing disorder. In the second study, a new cell penetrating peptide (PepFect 14) was developed and investigated for the formulation and delivery of SSOs using cell-models for two splicing disorders; -thalassemia and Duchenne muscular dystrophy. The feasibility of incorporating this delivery system into solid formulations via solid dispersion technique was also demonstrated. The formed solid formulations were as active as the freshly prepared nanocomplexes in solution even when stored at elevated temperatures for several weeks. In the third study, PepFect 14 was evaluated for the formulation and delivery of another kind of ONs: short interfering RNA (siRNA) in different cell lines. RNA interference effect was obtained at low siRNA doses with a unique kinetic profile. Solid formulations were then prepared and assessed for their stability in gastric conditions. PF14/siRNA solid formulations showed marked stability after incubation with simulated gastric fluid, which is extremely acidic and contains proteolytic enzymes. The fourth study of this thesis addressed design optimization of the newly developed antigene ON, Zorro-LNA (Zorro). Here, double-strand invasion was proven as the mechanism by which Zorro binds to duplex DNA. The original Zorro, targeting both strands of the DNA duplex, was made of two ONs connected via a 7-nucleotide linker. In this report, the possibility to synthesize Zorro as a bi-directional single-stranded ON was investigated, thus reducing the size, facilitating the design and improving Zorro efficiency. In conclusion, this thesis has dealt with developing formulation strategies for two different types of ON-based pharmaceuticals; SSOs and siRNA. Optimizing the design of Zorro LNA as an antigene ON has been also investigated. These findings may represent a step in the development of ON-based drug products as a new class of therapeutics

FTIR ANALYSIS FOR RETINA ASSOCIATED WITH DIABETIC CHANGES AND TREATMENT WITH OAT

Objective: Diabetes is known to induce oxidative stress along with deranging various metabolisms. One of the most serious complications of diabetes, a disease that has seen a worldwide increase in the prevalence, is diabetic retinopathy, which is a leading cause of acquired blindness. The aim of this study is to evaluate the effect of oat on the diabetic-induced oxidative stress and if this can attenuate the development of diabetic retinopathy.Methods: Changes on retina structure were performed by using the application of Fourier transform infrared spectroscopy.Results: The results demonstrated that diabetic retinopathy was associated with changes on the retina structure which appear after received a single dose of streptozotocin (STZ) 60 mg/kg. These changes clearly appeared in the NH-OH, CH and fingerprint regions. The use of oat in case of diabetic was associated with different beneficial effects on the retina constituents, as showed by the changes toward control of the same Fourier transform infrared spectroscopy bands.Conclusion: Oat can be considered as a novel treatment modality for diabetic retinopathy and further studies is required to optimize dosing and formulations that are maximally effective.Â

Investigating the Allosteric Behavior of Malate Dehydrogenase from Escherichia coli

Regulatory mechanisms of malate dehydrogenase from E.coli (eMDH) involving NADH as an allosteric effector were investigated. The reaction was studied in both directions: malate oxidation and oxaloacetate reduction. When malate was the variable substrate, a plot of rate against substrate concentration was sigmoidal in the presence of 0.065 mM NADH, which indicates the presence of an allosteric site for NADH on the enzyme. Binding of NADH at the allosteric site causes conformational changes in the active site and, thereby, changes the catalytic activity of the enzyme. An increase in Km value, from 1.3 to 3.9 mM malate was observed, which indicates a decrease in the enzyme affinity for the substrate. When eMDH was chemically modified with 5\u27-p-flurosulfonyl-benzoyladenosine (FSBA) in the presence of 0.15 mM NADH as a protecting agent, the allosteric behavior was abolished, which suggests that FSBA is modifying specific amino acid residues in the allosteric site and, therefore, preventing NADH from binding. eMDH was inactivated by FSBA in the absence of NADH. The inactivation appears to result from covalent modification of His 177 in the active site, which is believed to be crucial for the catalytic mechanism of eMDH

Synthesis and Characterization of New phenolic Schiff bases Derivatives Based on Terephthaladehyde

A variety of new phenolic Schiff bases derivatives have been synthesized starting from Terephthaladehyde compound, all proposed structures were supported by FTIR, 1H-NMR, 13C-NMR, Elemental analysis, some derivatives evaluated by Thermal analysis (TGA)

Investigating the Allosteric Behavior of Malate Dehydrogenase from Escherichia coli

Regulatory mechanisms of malate dehydrogenase from E.coli (eMDH) involving NADH as an allosteric effector were investigated. The reaction was studied in both directions: malate oxidation and oxaloacetate reduction. When malate was the variable substrate, a plot of rate against substrate concentration was sigmoidal in the presence of 0.065 mM NADH, which indicates the presence of an allosteric site for NADH on the enzyme. Binding of NADH at the allosteric site causes conformational changes in the active site and, thereby, changes the catalytic activity of the enzyme. An increase in Km value, from 1.3 to 3.9 mM malate was observed, which indicates a decrease in the enzyme affinity for the substrate. When eMDH was chemically modified with 5\u27-p-flurosulfonyl-benzoyladenosine (FSBA) in the presence of 0.15 mM NADH as a protecting agent, the allosteric behavior was abolished, which suggests that FSBA is modifying specific amino acid residues in the allosteric site and, therefore, preventing NADH from binding. eMDH was inactivated by FSBA in the absence of NADH. The inactivation appears to result from covalent modification of His 177 in the active site, which is believed to be crucial for the catalytic mechanism of eMDH

Synthesis and Antibacterial Evaluation for Some New Schiff-bases Derived from P-aminoacetanilide

تمتلك مشتقات قواعد شف اهمية كبيرة في مجال الكيمياء الصيدلانية ويمكن ان تستخدم في تحضير مختلف انواع المركبات ذات الفعالية الحيوية. في هذا البحث تم تحضير مشتقات جديدة من قواعد شف بخطوات متسلسلة. الحامض الجديد (I) تم تحضيره من تفاعل ثنائي كلورواسيتك اسد مع مولين من بارا- امينو استانلايد. تم تحويل الحامض (I) الى الاستر (II) بمفاعلته مع كبريتات ثنائي مثيل وبوجود كاربونات الصوديوم اللامائية مع الاسيتون الجاف كمذيب. هيدرازايد الحامض (III) تم تحضيره من اضافة 80% من الهيدرازين المائي الى الاستر باستخدام الايثانول كمذيب. الخطوة الاخيرة تضمنت تحضير قواعد شف (IV-VIII) عن طريق تفاعل هيدرزايد الحامض مع الالديهايدات الاروماتية المناسبة مع استخدام حامض الخليك الثلجي كعامل مساعد. تم تشخيص المشتقات الجديدة بواسطة طيف الاشعة تحت الحمراء والبعض منها بواسطة طيف الكتلة وطيف الرنين النووي المغناطيسي. المشتقات (IV-VIII) تم فحص فعاليتها البكتيرية ضد بكتريا القولون وبكتريا المكورات العنقودية الذهبية. جميع المركبات التي تم فحصها اظهرت فعالية ضد هذين النوعين من البكتريا.Derivatives of Schiff-bases possess a great importance in pharmaceutical chemistry. They can be used for synthesizing different types of bioactive compounds. In this paper, derivatives of new Schiff bases have been synthesized from several serial steps. The acid (I) was synthesized from the reaction of dichloroethanoic acid with 2 moles of p-aminoacetanilide. New acid (I) converted to its ester (II) via the reaction of (I) with dimethyl sulphate in the present of anhydrous of sodium carbonate and dry acetone. Acid hydrazide (III) has been synthesized by adding 80% of hydrazine hydrate  to the new ester using ethanol as a solvent. The last step included the preparation of new Schiff-bases (IV-VIII) by the reaction of acid hydrazide with appropriate aromatic aldehydes and using glacial acetic acid as a catalyst. New derivatives were diagnosed by FT-IR, and by mass and 1HNMR spectroscopy (some of them). Derivatives (IV-VIII) were screened for their antibacterial against E. Coli (G-) and staph. aureus (G+). All tested compounds were found to have activity against the two kinds of bacteria

The Yolk Sac Abnormalities, Maternal Serum Level of Cancer Antigen 125 (CA-125) and Beta Human Chorionic Gonadotropin (B-HCG) as an Early Predictors of First Trimester Pregnancy Loss in Patients with Threatened Miscarriage

Background: Pregnancy loss before 20 weeks is considered a miscarriage, as is the loss of a fetus weighing less than 500 grams before viability. A medical emergency, threatened miscarriage affects 15–25% of pregnancies. Aim and objectives: The goal of this study was to assess the predictive value of maternal blood levels of Cancer Antigen 125 (CA-125) and beta-human chorionic gonadotropin (B- HCG) in individuals at risk of miscarriage during the first trimester. Subjects and methods: This study was a prospective cohort study. This study included 120 pregnant women with threatened abortion between (6-11 weeks) and followed up till end of 14th week. Results: 36(30%) of pregnant women aborted, while 84(70%) of women continued till 14th weeks of pregnancy. At a cut-off value of 45 U/ml, the CA125 test was shown to have a sensitivity of 88.9% and a specificity of 77.5%, respectively, while also having a positive predictive value of 79.8% and a negative predictive value of 87.5%. At a cut-off value of 18.501 mlIU/ml, the B-HCG test's sensitivity and specificity were determined to be 96.3 and 88.9, respectively, with a positive predictive value of 89.7% and a negative predictive value of 96%. Conclusion: Even before fetal morphology can be investigated sonographically, abnormalities in the size of the yolk sac can be utilized as a good prognostic sign of early pregnancy loss. Pregnancy viability can be estimated from first trimester serum CA 125 and Beta HCG measurements