Publication: Beta-katenin genini hedefleyen shRNA’yı içeren aljinat-kitozan hidrojellerinin meme kanserinde etkisi
Abstract
Amaç: Beta-katenin (CTNNB1)’in meme kanserinin kötü prognozuyla ilişkili olduğu ve yüksek oranda aktivasyonunun meme kanseri oluşumunda ve ilerlemesinde etkili olduğu belirtilmiştir. Bu çalışmada, CTNNB1 mRNA’yı hedefleyen short hairpin RNA (shRNA) ekspresyon plazmidinin meme kanseri hücrelerine ve tümör dokularına verilmesi için hazırlanan aljinat-kitozan hidrojellerin gen taşıyıcı sistem olarak in vitro ve in vivo etkinliğinin araştırılması amaçlanmıştır. Gereç ve yöntem: pH değişimi yoluyla sodyum aljinat ve kitozan kullanılarak hazırlanan hidrojellerin karakterizasyonları yapılarak formülasyonlar optimize edilmiştir. shRNA ekspresyon plazmidi aljinat-kitozan hidrojeller ile meme kanseri hücreleri MDA-MB-231 ve 4T1 hücre hatlarına verilerek gen inhibisyonuna ve hücre proliferasyonuna etkisi belirlenmiştir. En etkili bulunan formülasyonun meme tümörü oluşturulan sıçanlara uygulanması sonrasında tümör boyutları takip edilmiştir ve tümör dokularında CTNNB1 gen ekspresyonu incelenmiştir. Bulgular: Aljinat-kitozan hidrojeller ile shRNA plazmidin meme kanseri hücrelerine transfeksiyonu sonrasında CTNNB1 ekspresyonu yaklaşık %30 oranında baskılanmıştır. Meme tümörü oluşturulan sıçanlarda shRNA plazmid içeren hidrojeller ile tedavi sonrasında tümör boyutu kontrol grubuna göre anlamlı derecede küçülmüştür ve CTNNB1 ekspresyonunun önemli derecede azaldığı immünohistokimya analizi ile gösterilmiştir. Sonuç: Aljinat-kitozan hidrojellerin bölgesel gen taşıyıcı bir sistem olarak kullanılabileceği ve meme kanserinde CTNNB1 ekspresyonunun baskılanmasının hücre proliferasyonunu ve tümör gelişimini engellediği belirlenmiştir.
Objective: Beta-catenin (CTNNB1) is associated with the poor prognosis of breast cancer and its high activation is effective in breast cancer development and progression. In this study, we aimed to investigate in vitro and in vivo efficiency of alginate-chitosan hydrogels prepared for the delivery of shRNA expression plasmid targeting CTNNB1 mRNA to breast cancer cells and tumor tissues as a gene delivery system. Material and Methods: The formulations were optimized by characterizing hydrogels which were prepared via pH change using sodium alginate and chitosan. Effects of the hydrogels on gene silencing and cell proliferation were determined after shRNA expression plasmid was administered to MDA-MB-231 and 4T1 breast cancer cell lines with alginate-chitosan hydrogels. After the application of the most effective formulation to rats with mammary tumors, tumor sizes were followed and CTNNB1 gene expression was examined in tumor tissues. Results: After transfection of the shRNA plasmid into breast cancer cells with alginate-chitosan hydrogels, beta-catenin expression was suppressed by approximately 30%. After treatment with hydrogels containing shRNA plasmid in rats with mammary tumors, tumor size was significantly reduced compared to the control group, and immunohistochemistry analysis demonstrated that CTNNB1 expression was significantly reduced. Conclusion: It was determined that alginate-chitosan hydrogels can be used as a local gene delivery system and suppression of CTNNB1 expression in breast cancer prevented cell proliferation and tumor development.
Objective: Beta-catenin (CTNNB1) is associated with the poor prognosis of breast cancer and its high activation is effective in breast cancer development and progression. In this study, we aimed to investigate in vitro and in vivo efficiency of alginate-chitosan hydrogels prepared for the delivery of shRNA expression plasmid targeting CTNNB1 mRNA to breast cancer cells and tumor tissues as a gene delivery system. Material and Methods: The formulations were optimized by characterizing hydrogels which were prepared via pH change using sodium alginate and chitosan. Effects of the hydrogels on gene silencing and cell proliferation were determined after shRNA expression plasmid was administered to MDA-MB-231 and 4T1 breast cancer cell lines with alginate-chitosan hydrogels. After the application of the most effective formulation to rats with mammary tumors, tumor sizes were followed and CTNNB1 gene expression was examined in tumor tissues. Results: After transfection of the shRNA plasmid into breast cancer cells with alginate-chitosan hydrogels, beta-catenin expression was suppressed by approximately 30%. After treatment with hydrogels containing shRNA plasmid in rats with mammary tumors, tumor size was significantly reduced compared to the control group, and immunohistochemistry analysis demonstrated that CTNNB1 expression was significantly reduced. Conclusion: It was determined that alginate-chitosan hydrogels can be used as a local gene delivery system and suppression of CTNNB1 expression in breast cancer prevented cell proliferation and tumor development.