Effects of CRISPR-Cas9 silencing of VPS28 gene on cisplatin response in chemotherapy-resistant ovarian cancer cells

Öz

Aim: Today, ovarian cancer is still common in women and is the leading cause of deaths from gynecologic cancer. Although cisplatin, a chemotherapeutic agent, is frequently used in the treatment, resistance to cisplatin develops in patients over time and genetic factors are known to be responsible for the development of resistance. In this study, it was aimed to determine the possible role of the ESCRT-I complex component VPS28 gene, which is active in multivesicular body biogenesis, in ovarian cancer cisplatin resistance. Method: After silencing the VPS28 gene in cisplatin-resistant A2780 ovarian cancer cells using the CRISPR-Cas9 method, the effects on proliferation, cisplatin response, apoptosis and cell cycle were investigated. Results: Functional analyses revealed that VPS28 gene silencing in chemotherapy-resistant ovarian cancer increased cisplatin sensitivity, induced apoptosis, and caused mitotic catastrophe by arresting the cell cycle in the S phase. Conclusion: The VPS28 gene has been shown to be involved in acquired cisplatin resistance in A2780 ovarian cancer cells, and its silencing has a synergistic effect with the cisplatin used in treatment.

Anahtar Kelimeler

• Ovarian cancer
• cisplatin resistance
• VPS28
• CRISPR-Cas9

Kemoterapi dirençli over kanseri hücrelerinde VPS28 geninin CRISPR-Cas9 yöntemi ile susturulmasının cisplatin yanıtı üzerine etkileri

Öz

Amaç: Günümüzde yumurtalık (over) kanseri kadınlarda halen yaygın olarak görülmekte ve jinekolojik kansere bağlı ölüm nedenlerinin başında gelmektedir. Tedavisinde sıklıkla kemoterapötik bir ajan olan cisplatin kullanılmakla birlikte, zamanla hastalarda cisplatine karşı direnç gelişmekte ve direnç gelişiminde genetik faktörlerin sorumlu olduğu bilinmektedir. Bu çalışmada multiveziküler cisim biyogenezinde etkin ESCRT-I kompleks bileşeni VPS28 geninin, over kanseri cisplatin direncindeki olası rolünün belirlenmesi amaçlanmıştır. Yöntem: Cisplatin dirençli A2780 over kanseri hücre serisinde VPS28 geni CRISPR-Cas9 yöntemiyle susturulduktan sonra proliferasyon, cisplatin yanıtı, apoptoz ve hücre döngüsü üzerine etkileri araştırılmıştır. Bulgular: Yapılan fonksiyonel analizler sonucunda, cisplatin dirençli over kanseri hücrelerinde VPS28 geninin susturulması ile cisplatin duyarlılığının arttığı, apoptozun indüklendiği ve hücre döngüsü S evresinde mitotik katastrofa neden olduğu belirlenmiştir. Sonuç: VPS28 geninin A2780 over kanseri hücrelerindeki kazanılmış cisplatin direncinde etkili olduğu ve susturulmasının, tedavisinde kullanılan cisplatin ile birlikte sinerjistik etki yarattığı gösterilmiştir.

Anahtar Kelimeler

• Over kanseri
• cisplatin direnci
• VPS28
• CRISPR-Cas9

Destekleyen Kurum

Bu çalışma, Türkiye Bilimsel ve Teknolojik Araştırma Kurumu (TÜBİTAK) tarafından 123Z193 Numaralı proje ile desteklenmiştir.

Etik Beyan

İn vitro çalışmalar yapıldığı için etik kurul onayına ihtiyaç duyulmamıştır.

Teşekkür

Projeye verdiği destekten ötürü TÜBİTAK’a teşekkürlerimizi sunarız. Ayrıca laboratuvar çalışmalarında bize yardımcı olan ve her türlü imkanı sağlayan Harran Üniversitesi Tıp Fakültesi Tıbbi Biyoloji Anabilim Dalı Öğretim Üyesi Arş. Gör. Dr. Neslihan Pınar Özateş’e, Harran Üniversitesi Tıp Fakültesi Fizyoloji Anabilim Dalı Öğretim Üyesi Dr. Öğr. Üyesi Seyhan Taşkın’a, Mehmet Akif Ersoy Üniversitesi Fen Edebiyat Fakültesi, Moleküler Biyoloji ve Genetik Bölümü Öğretim Üyesi Dr. Muzaffer Dükel’e teşekkür ederiz.

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