Publication: Tiroid kanserinde ısı şok proteinlerinin rolünün araştırılması
Abstract
Birçok kanserde olduğu gibi, ana sinyal yolaklarında görevli anahtar proteinleri kodlayan genlerde olan değişiklikler tiroid kanseri oluşumunda da önemli rol oynar. HSP90; kanser tedavisinde ilgi çeken terapötik bir hedeftir. Kanser hücrelerinde ekspresyonu artan HSP90’ın rolü sinyal yolaklarında görevli proteinlerin fonksiyonlarının devamını sağlamaktır. Çalışmamız, BRAF V600E mutasyonuna sahip olduğu bilinen papiller tiroid kanseri hücre hattı B-CPAP ile gerçekleştirildi. Birçok farklı kanser hücre hattında HSP’lerin üretiminde inhibisyon yaparak hücrelerde apoptoz gerçekleştirdiği bilinen flavonoid Quercetin’in 12.5 ile 150µM’lık konsantrasyonları papiller tiroid kanseri hücre hattına 24 ve 48 saat inkübasyonlar ile uygulandı, Hücre canlılığı WST-1 testi ile ölçüldü, canlılığın artan doza bağımlı biçimde azaldığı ve 100µM’lık konsantrasyonda canlılığın %52.5 olduğu tespit edildi. Apoptoz analizi için Quercetin’in 24 saat inkübasyonu ve 50-150µM’lık konsantrasyon aralığı seçildi ve akış sitometresinde Annexin V-FITC ve PI boyaları ile analizi gerçekleştiridi. Elde edilen bulgulara göre, 100µM’lık konsantrasyonda apoptoz oranı %35.1 idi. Apoptoz ayrıca floresan mikroskobisinde Hoechst boyaması ile gösterildi. Son olarak Western blot analizi ile Quercetin uygulanan hücreler ile kontrol grubu arasındaki HSP90 seviyeleri karşılaştırıldı. Elde edilen bulgularda, HSP90 seviyelerinin artan Quercetin konsantrasyonuna bağlı olarak azaldığı tespit edildi. Bu bulgulardan yola çıkarak ilerde yapılacak çalışmalar ile Quercetin’in B-CPAP hücreleri üzerindeki apoptik moleküler mekanizmalarının ve kombinasyon terapilerin araştırılması gerektiğini düşünmekteyiz. Papiller Tiroid Kanseri, Quercetin, HSP90
As in many other cancers, genetic alterations that occurs in genes encoding for key proteins of major signalling pathways are the driving force for the tumorigenesis of thyroid cancer. HSP90 is an emerging therapeutic target of interest for the treatment of cancer. HSP90, which is over expressed in cancer cells, maintains the function of signalling proteins. In our study, we have worked with B-CPAP; thyroid papillary cancer cell line which is known to have BRAF V600E mutation. We have administered the flavonoid, Quercetin which is known to induce apoptosis by inhibiting HSP production on various cancer cell lines, at different concentrations (between 12.5 and 150 μM) for 24 and 48 hours. We have measured cell viability using WST-1 assay. We found that the viability decreases in a dose dependent manner and at 100 μM, the viability was 52.5%. Then, we chose Quercetin concentrations between 50-150 μM for 24 hours, for the apoptosis analysis in flow cytometry with Annexin V-FITC and PI stains. We found that at 100 μM the apoptosis percentage was 35.1. Apoptosis of cells are also observed with the application of Hoechst stain in florescence microscopy. Finally we used Western blotting to compare HSP90 levels between cells treated with Quercetin and the control group. We found that HSP90 levels decreased with increasing Quercetin concentrations. In the light of the obtained data, in future studies it would be useful to investigate the apoptotic mechanisms of Quercetin and use combinational therapies on B-CPAP cells. Key words : Papillary Thyroid Cancer, Quercetin, HSP90
As in many other cancers, genetic alterations that occurs in genes encoding for key proteins of major signalling pathways are the driving force for the tumorigenesis of thyroid cancer. HSP90 is an emerging therapeutic target of interest for the treatment of cancer. HSP90, which is over expressed in cancer cells, maintains the function of signalling proteins. In our study, we have worked with B-CPAP; thyroid papillary cancer cell line which is known to have BRAF V600E mutation. We have administered the flavonoid, Quercetin which is known to induce apoptosis by inhibiting HSP production on various cancer cell lines, at different concentrations (between 12.5 and 150 μM) for 24 and 48 hours. We have measured cell viability using WST-1 assay. We found that the viability decreases in a dose dependent manner and at 100 μM, the viability was 52.5%. Then, we chose Quercetin concentrations between 50-150 μM for 24 hours, for the apoptosis analysis in flow cytometry with Annexin V-FITC and PI stains. We found that at 100 μM the apoptosis percentage was 35.1. Apoptosis of cells are also observed with the application of Hoechst stain in florescence microscopy. Finally we used Western blotting to compare HSP90 levels between cells treated with Quercetin and the control group. We found that HSP90 levels decreased with increasing Quercetin concentrations. In the light of the obtained data, in future studies it would be useful to investigate the apoptotic mechanisms of Quercetin and use combinational therapies on B-CPAP cells. Key words : Papillary Thyroid Cancer, Quercetin, HSP90