Publication: Kitozan/ PAI-1 sirna nanopleksılerinin hepatosellüler karsinoma hücre hatlarında anti- anjiyogenik ve anti-karsinojenik etkilerinin araştırılması
Abstract
Amaç: Anjiyogenez invaziv tümör büyümesi, metastazı ve kanser ilerlemesininkontrolünde önemli bir nokta oluşturur. Bu amaçla en yaygın çalışılan gen susturmateknolojisi RNAi (RNA interference), gen ekspresyonunu inhibe edilebilmesi(susturma), yüksek spesifisite ve hedeflenme özellikleri nedeniyle tedavidedenenmektedir. Anjiyogenez ve tümörle ilgili faktörlerin siRNA (small interferingRNA)’ lar ile hücre kültürü ortamında inhibe edilmesiyle ilgili çalışmalar mevcuttur.siRNA’ ların terapötik verimini artırmak için uygun taşıyıcı sistem olarakkullandığımız güvenli bir biyopolimer olan kitozan büyük önem taşımaktadır.Sunulan tez kapsamında hepatosellüler karsinomada (HCC) anjiyogenezi tetikleyenbir faktör olan PAI-1’ i susturarak anjiogenezin durdurulması planlanmaktadır.Gereç ve Yöntem: PAI-1‘ in aşırı ekspresyonu, HCC’ lı hastalarda tümör invazyonuve metastazı gelişimine yol açar. Anjiyogenezde etkin olan PAI-1 ekspresyonunuengellemek için ticari olarak satılan siRNA spektroskopik ve elektroforetikkontrollerinden sonra kitozan ile kompleksleşme oranı belirlenmiştir. Komplekslerinkarakterizasyonu HepG2C3a ve Hep3B hücrelerinde in vitro transfeksiyonu etkinliğiincelenmiş ve PAI-1 ekspresyonu elisa yöntemi ile tayin edilmiştir. Hazırlanankomplekslerin TEM görüntüleri ile boyut/ zeta ölçümlerinin uygun olduğu tespitedildi.Bulgular: HCC hücre hatlarında uygun komplekslern PAI-1 gen ekspresyonunuyaklaşık %50 oranında inhibe ettiği istatiksel oralara anlamlı azaldığı gözlenmiştir.Sonuç: Elde edilen sonuçlar kitozan siRNA-PAI-1 için uygun bir taşıyıcı sistemolduğu desteklenmiştir.
Objective: Angiogenesis is an important factor in the control of invasive tumorgrowth, metastasis and cancer progression. For this purpose, the most widely studiedgene silencing technology, RNAi (RNA interference), gene expression inhibition(silencing), high specificity and targeting properties are being tried in the treatment.There are studies on the inhibition of angiogenesis and tumor related factors withsiRNA (small interfering RNA) in cell culture media. Chitosan, a safe biopolymer,which we use as a suitable carrier system, is of great importance to increase thetherapeutic efficiency of siRNAs. In this thesis, it is planned to stop angiogenesis bysilencing PAI-1 (plasminogen activator inhibitor-1), a factor that triggersangiogenesis in hepatocellular carcinoma (HCC).Materials and Methods: Overexpression of PAI-1 leads to tumor invasion andmetastasis in patients with HCC. The rate of complexation with chitosan wasdetermined after spectroscopic and electrophoretic controls of commerciallyavailable siRNA to inhibit PAI-1 expression, which is effective in angiogenesis.Characterization of complexes The efficacy of in vitro transfection in HepG2C3a andHep3B cells was examined and PAI-1 expression was determined by elisa method.TEM images and zeta measurements of the complexes were found to be appropriate.Results: In the HCC cell lines, it was observed that suitable complexes inhibitedPAI-1 gene expression by approximately 50% and decreased statistically.Conclusion: The results obtained were supported to be a suitable carrier system forchitosan siRNA-PAI-1.
Objective: Angiogenesis is an important factor in the control of invasive tumorgrowth, metastasis and cancer progression. For this purpose, the most widely studiedgene silencing technology, RNAi (RNA interference), gene expression inhibition(silencing), high specificity and targeting properties are being tried in the treatment.There are studies on the inhibition of angiogenesis and tumor related factors withsiRNA (small interfering RNA) in cell culture media. Chitosan, a safe biopolymer,which we use as a suitable carrier system, is of great importance to increase thetherapeutic efficiency of siRNAs. In this thesis, it is planned to stop angiogenesis bysilencing PAI-1 (plasminogen activator inhibitor-1), a factor that triggersangiogenesis in hepatocellular carcinoma (HCC).Materials and Methods: Overexpression of PAI-1 leads to tumor invasion andmetastasis in patients with HCC. The rate of complexation with chitosan wasdetermined after spectroscopic and electrophoretic controls of commerciallyavailable siRNA to inhibit PAI-1 expression, which is effective in angiogenesis.Characterization of complexes The efficacy of in vitro transfection in HepG2C3a andHep3B cells was examined and PAI-1 expression was determined by elisa method.TEM images and zeta measurements of the complexes were found to be appropriate.Results: In the HCC cell lines, it was observed that suitable complexes inhibitedPAI-1 gene expression by approximately 50% and decreased statistically.Conclusion: The results obtained were supported to be a suitable carrier system forchitosan siRNA-PAI-1.