Glioma hücrelerinde temozolomid ve KC7F2 ajanlarının kombinasyonlarının moleküler biyolojik açıdan incelenmesi

Malign beyin tümörleri nöronal hastalıklar içerisinde önemli bir yer tutmakta olup Dünya Sağlık Örgütü (WHO) tarafından histolojik ve invaziflik ve genetik özellikleri baz alınarak I, II, III ve IV derece olarak sınıflandırılmıştır. Glioblastom Multiform (GBM) 2016 WHO raporunun son değerlendirilmesine göre Astrositom alt sınıfının IV dereceli en invaziv ve malignant üyesidir. Malign beyin tümörlerinin önemli bir kısmını oluşturarak, tedaviye karşı direnci ve zayıf yanıt verme özelliğinden, bunların yanısıra topografik yapısı nedeniyle kan beyin baryerinin kimyasal tedaviye karşı oluşturduğu engel, ayrıca ağır klinik ifadesinden dolayı tedavisi zorlaşmakta ve yeni tedavi yöntemleri ve yeni terapötik ajanların geliştirilmesine ihtiyaç duyulmaktadır. Konvansiyonel tedavi yöntemleri olarak cerrahi rezeksiyon, radyoterapi ve kemoterapi uygulanmakta olup hastalarda medyan sağkalım süresi 12.5 aydır. Glioblastom tedavisinde Temozolomid(TMZ) yaygın olarak kullanılmakta ve primer tedavi yöntemi sayılmaktadır. Aliklleyici ajanlar sınıfının bir üyesi olan TMZ kan beyin bariyerini kolayca geçe bilmesinden dolayı güncel olarak en etkin ilaç sayılmakta. Buna rağmen radyoterapi ile kombinasyon halinde uygulandığında sağkalım süresi 15 ayı geçmemekte. Bunun nedeni TMZ`ye karşı oluşan direnç mekanizmasıdır. Glikoz metabolizması, enerji üreten ve ATP olarak depolayarak tüm hücresel süreçler için enerji sağlayan karmaşık bir enerji üreten makinedir. Bir glükoz molekülü, tam glikolitik katabolizmadan sonra 38 molekül ATP üretir. Otto Warburg'un sayısız çalışması ve temel hipotezine göre, kanser metabolizması normal hücrelerden tamamen farklıdır. Kanser hücreleri mitokondriyal adım girmeden sadece ilk glikolitik adımı gerçekleştirerek anaerobik fenotipe eğilim gösterirler. Sonuçta bu hücreler laktik asit üretir ve salgılarlar ve mikro-ortamlarını aerobik koşullarda bile daha asidik yaparlar. Bu olgu “Warburg hipotezi” ve ya “anaerobik glikoliz” olarak tanımlanmakta. Glikoliz enzimleri ilk olarak bu fenotipik ifadenin temel aktörleri olmasına rağmen, bazı genetik ve epigenetik faktörler de istisna değildir. Bu nedenle deneyimizde kanser metabolizmasını hedef almak için KC7F2 etken maddesini kullandık. Çalışmamızda kanser metabolizmasının hedefleyerek TMZ kematerapötik ajanının etkisi ile kombinasyon halinde kanser hücre proliferasyonu, sağkalımı, apoptoz, metabolizma genlerinin ekspresyonu üzerindeki etkiyi gözlemlemek için iki farklı etken maddenin ayrı ayrı ve kombine halde etkisini inceleyerek değerlendirdik. Deneyimizin sonucunda etken maddelerin kombinatif etkisinin TMZ alkilleyici ajanının etkin dozunu düşürdüğünü ve etkisinin arttığını gözlemlemiş olduk, ayrıca etkilerinin metabolik açıdan değerlendirererek aerobik glikolizi baskıladığını da gözlemledik.Brain tumors occupy an important place in neuronal diseases and are classified as I, II, III and IV by World Health Organization (WHO) based on their histological, invasive and genetic characteristics. Glioblastom Multiforme (GBM) is the most invasive and malignant member of the IV grade of the subclass Astrocytoma according to the last assessment of the2016 WHO report. Due to the resistance to treatment and weak response, as well as the topographical structure of the blood brain barrier, which is a major part of malignant brain tumors, treatment is also difficult due to the severe clinical manifestation, and new treatment methods and new therapeutic agents are needed. Conventional treatment methods are surgical resection, radiotherapy and chemotherapy. Median survival time is 12.5 months in patients. Temozolomide(TMZ) is widely used in the treatment of glioblastoma and is considered as the primary treatment modality. TMZ, a member of the class of cognitive agents, is currently considered the most effective drug because it can easily pass through the blood brain barrier. However, when administered in combination with radiotherapy, the survival time does not exceed 15 months. The reason for this is the resistance mechanism against TMZ.Glucose metabolism is a complex energy producing machine that generates energy and stores it as ATP and provides energy for all cellular processes. A glucose molecule produces 38 molecules of ATP after full glycolytic catabolism. According to Otto Warburg's numerous studies and basic hypothesis, cancer metabolism is completely different from normal cells. Cancer cells tend to the anaerobic phenotype only by performing the first glycolytic step without entering the mitochondrial step. As a result, these cells produce lactic acid and make the secretions and micro-media even more acidic in aerobic conditions. This phenomenon is attributed to the Warburg hypothesis and either as anaerobic glucolysis. Although glycolysis enzymes are the primary actors of this phenotypic expression, some genetic and epigenetic factors are no exception. For this reason, we experimentally used KC7F2 active ingredient to target cancer metabolism. In our study, we evaluated cancer metabolism in combination with the effect of TMZ chemotherapeutic agent, examining the effect of two different agents separately and in combination to observe the effects of cancer cell proliferation, survival, apoptosis, expression of metabolism genes on expression. As a result of our experience, we observed that the combined effect of the active ingredients reduced the effective dose of the TMZ alkylating agent and that the effect was increased.We have also observed that the effect of the combined teraphy is assessed from a metabolic point of view and that it suppresses aerobic glycolysis

miR-17 in imatinib resistance and response to tyrosine kinase inhibitors in chronic myeloid leukemia cells

In this study we examined the expression levels of miR-17 which possesses oncogenic activities through downregulation of CDKN1A, p21 and E2F1 tumor suppressor genes, in imatinib sensitive and resistant chronic myeloid leukemia (CML) cells. On the other hand, we also determined the expression levels of miR-17 in response to tyrosine kinase inhibitors imatinib, nilotinib and dasatinib used for the treatment of CML. Methods: The expression profiles of miR-17 were analysed by Stem-Loop reverse transcription (RT) polymerase chain reaction (PCR). Results: The results revealed significant increase in the expression levels of miR-17 in imatinib sensitive and resistant cells compared to peripheral blood mononuclear cells (PBMCs). On the other hand, significant decrease was observed in miR-17 levels in response to imatinib, nilotinib and dasatinib. Conclusion: These results may imply that miR-17 can be used for diagnosis and treatment of CML.Scientific and Technological Research Council of Turke

Enhanced Anti-cancer Potency Using a Combination of Oleanolic Acid and Maslinic Acid to Control Treatment Resistance in Breast Cancer

Purpose: The phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/ mTOR) pathway is a complex intracellular metabolic pathway that leads to cell growth and tumor proliferation and plays a key role in drug resistance in breast cancer. Therefore, the anti-cancer effects of oleanolic acid (OA), maslinic acid (MA), and their combination were investigated to improve the performance of the treatment strategy. Methods: We investigated the effect of OA and MA on cell viability using the WST-1 method. The synergistic effect of the combination was analyzed by isobologram analysis. In addition, the effects of the two compounds, individually and in combination, on apoptosis, autophagy, and the cell cycle were investigated in MCF7 cells. In addition, changes in the expression of PI3K/AKT/mTOR genes involved in apoptosis, cell cycle and metabolism were determined by quantitative RT-PCR. Results: MA, OA, and a combination of both caused G0/G1 arrest. Apoptosis also increased in all treated groups. The autophagosomal LC3-II formation was induced 1.74-fold in the MA -treated group and 3.25-fold in the MA-OA-treated group. The combination treatment resulted in increased expression of genes such as GSK3B, PTEN, CDKN1B and FOXO3 and decreased expression of IGF1, PRKCB and AKT3 genes. Conclusion: The results showed that the combination of these two substances showed the highest synergistic effect at the lowest dose and using MA-OA caused cancer cells to undergo apoptosis. The use of combination drugs may reduce the resistance of cancer cells to treatment

Adipose Tissue Mesenchymal Stem Cells Exposed To Oxytocin and Sunitinib are Synergistically Dystrophic

Objective: Mesenchymal stem cells (MSCs) are also promising in immunosuppressed patients after organ and tissue transplantation, in addition to their current wide range of uses and research areas. Sunitinib is a receptor tyrosine kinase with immunosuppressive properties and its cytotoxic activity in different types of cells is known. Our study aimed to elucidate the effect of oxytocin on sunitinib-treated MSCs. Methods: For this purpose, commercially available rat adipose tissue-derived MSC (ADMSCs) was used. The individual or combinational effect of the active substances on viability was evaluated with WST-1, the effect on apoptosis Annexin V, the effect on oxidative stress markers MDA, CAT, GPX, and SOD ELISA tests. Results: The IC50 value of sunitinib was determined as 44.57 ?M at the 48th hour, and it was determined that oxytocin had no cytotoxic effect in doses up to 100 ?M. Treatment of the two agents in combination increased the cytotoxic effect of sunitinib. Oxytocin attenuated the effect of sunitinib on apoptosis and lipid peroxidation. Conclusion: It is important to investigate the efficacy of these two substances individually and in combination with ADMSCs with further experiments to evaluate the potential use of oxytocin in organ and tissue transplantations

Ege Üniversitesi organ nakli merkezinde karaciğer transplantasyonu alıcı ve donörlerindeki faktör V leiden gen mutasyonunun prevalansı

Liver transplantation has become a routine procedure for the treatment of end stage of chronic liver disease and acute liver failure. Factor V which is one of the coagulation factors is synthesized by the liver. Defects in coagulation factor genes may lead to the production of abnormal or deficient proteins that constellate risk factors for the development of deep-vein thrombosis. Acquired Active protein C resistance is the most encountered coagulation abnormality and in 90-95 % of cases and it is caused by a point mutation in the factor V gene. The factor V Leiden mutation is genotyped for a single base-pair change (G1691A). Patients who develop venous thrombosis after liver transplantation should be screened for thrombophilic abnormalities. Deep-vein thrombosis occurs approximately 2.7% of patients after liver transplantation. For this reason, FV Leiden mutation has determined in organ donors and recipients. In our study; recipient (191 individuals),-donors (693 individuals) totally 884 cases appealed to our department. Genomic DNA of the cases isolated and FV Leiden mutation was determined by Real Time Online PCR. Totally 884 individual, included to our study. The prevalence of the heterozygous mutation was 80 and homozygous mutation was 3. Number of normal genotype was determined 801 individuals. Routine screening for factor V Leiden mutation by polymerase chain reaction, and appropriate pre-operative and post-operative anticoagulation after transplantation might be valuable strategy to prevent tromboembolic complications in transplant recipients.Karaciğer transplantasyonu, son evredeki karaciğer hastalıklarında ve akut karaciğer yetmezliğinde rutin bir tedavi haline gelmiştir. Koagülasyon faktörlerinden biri olan Faktör V karaciğer tarafından sentezlenmektedir. Koagülasyon faktör genlerindeki defektler, anormal veya derin ven tromboz gelişimi için risk faktörü oluşturabilecek yetersiz proteinlerin oluşumuna sebep olmaktadır. Aktif protein C ye karşı kazanılmış olan direnç (APCr) en sık karşılaşılan koagülasyon anomalisidir ve % 90 - 95 oranında faktör V Leiden mutasyonu sonucu gelişmektedir. Bu mutasyon, G1691A genotiplemesi şeklinde karakterize edilmektedir. Trombofilik anomalilerde canlı donörlerin genotiplemesi mutlaka gerekmektedir. Karaciğer transplantasyonu sonrası olguların yaklaşık % 2.7' sinde derin ven trombozu gözlenmektedir. Bu amaçla, organ alıcı ve donörlerinde FV Leiden gen mutasyonu analizi yapılmalıdır. Çalışmamızda alıcı (191 kişi) ve donör (693 kişi) olarak Anabilim Dalı' miza yollanan toplam 884 olgunun kan örneklerinden genomik DNA izole edilmiş ve Faktör V Leiden mutasyonları Real-time Online PCR ile saptanmıştır. Toplam 884 olgunun dahil edildiği çalışmamızda FV Leiden gen mutasyonu için 80 olgu heterozigot, üç olgu homozigot, 801 olgu normal genotipte olduğu belirlenmiştir. Karaciğer transplantasyonu sonrasında, derin-ven trombozisi geliştiren hastaların, donör karaciğerinde homozigot Faktör V Leiden gen mutasyonu sonucu ortaya çıkan sonradan kazanılmış aktive olmuş Protein C (APC) rezistansı temeline dayandığı belirtilmiştir. FV Leiden gen mutasyonunun rutin taraması pre ve post operatif koagülasyonların kullanımı transplantasyon olgularında tromboembolik komplikasyonların gelişimini önlemede uygun bir stratejidir. Sonuç olarak bu genotipleme, hem alıcıyı hem de donörün trombozdan korunmasını sağlamaktadır