SIVI BİYOPSIDE SİRKÜLE TÜMÖR HÜCRESİ YAKALAMAK İÇİN DNA APTAMERİ-İMMOBİLİZE EDİLMİŞ BLOK KOPOLİMER YÜZEY

Yıl 2025, Cilt: 15 Sayı: 3, 345 - 353, 15.09.2025

Sadık Kağa , Elif Kağa

https://doi.org/10.16919/bozoktip.1700824

Öz

Amaç: Dolaşımdaki sıvı biyopsi bileşenlerini tespit etmek, erken kanser tanısı ve hastalık ilerlemesinin izlenmesi için önemlidir. Mevcut biyosensörler, dolaşımdaki kanser hücrelerini hastalığın ileri evrelerinde tespit edebilmektedir, bu da daha etkili tekniklerin araştırılmasına yol açmaktadır. Bu çalışmada, dolaşımdaki kanser hücrelerinin hassas tespiti için AS1411 aptameriyle fonksiyonelleştirilmiş ve PGMA-b-PPDSMA blok kopolimeriyle modifiye edilmiş yeni bir yüzey geliştirilmiştir.
Gereç ve Yöntemler: PGMA-b-PPDSMA blok kopolimeri, NMR spektroskopisi ve GPC ile sentezlenmiş ve karakterize edilmiştir. Cam lamlar, APTES ile modifiye edilerek blok kopolimerin PGMA epoksi grupları aracılığıyla kovalan bağlanması sağlanmıştır. PPDS'nin piridil disülfid grupları, tiol-modifiye edilmiş anti-nükleolin DNA aptameri (AS1411)'nin bölgesel immobilizasyonunu mümkün kılmıştır. Immobilizasyon verimliliği ve aptamer spesifitesi, MCF-7 meme kanseri hücreleri ile floresan mikroskopi kullanılarak doğrulanmıştır.
Bulgular: NMR ve GPC sonuçları, 4.8 kDa PGMA ve 1.2 kDa PPDSMA bloklarından oluşan 6 kDa moleküler ağırlığa sahip PGMA-b-PPDS blok kopolimerinin sentezlendiğini ve saflaştırıldığını göstermiştir. Cam yüzeylerdeki APTES modifikasyonu, FTIR spektrumundaki 1500-1650 cm-¹ aralığında NH2 grubuna ait emilim sinyalleriyle doğrulanmıştır. Tiol-modifiye AS1411 aptamerinin PGMA-b-PPDS blok kopolimerinin PPDS grubuna bağlanma verimliliği, reaksiyonla oluşan piridin-2-tiyon yan ürününün 250-400 nm arasında emilim değişimi ile zaman içinde izlenmiştir.
Sonuç: DAPI boyama ve yıkama işlemlerinden sonra elde edilen floresan mikroskopi görüntüleri, AS1411 ile modifiye edilmiş yüzeylere MCF-7 hücrelerinin seçici bağlandığını göstermiştir. Bu sonuçlar, yüzeyin erken evre sıvı biyopsi uygulamalarında dolaşımdaki kanser hücrelerinin hassas bir şekilde yakalanması için umut verici bir platform sunduğunu göstermektedir.

Anahtar Kelimeler

Meme kanseri , Polimerler , DNA Aptamerleri , Biyoalgılayıcılar , Likit Biyopsi

Proje Numarası

22.TEMATİK.012

DNA APTAMER-IMMOBILIZED BLOCK COPOLYMER SURFACE FOR CIRCULATING TUMOR CELL CAPTURE IN LIQUID BIOPSY

Öz

Objective: Detecting circulating liquid biopsy components is essential for early cancer diagnosis and monitoring disease progression. Current biosensors detect circulating tumor cells in later stages, prompting a search for more effective techniques. In this study, we developed a novel surface modified with PGMA-b-PPDSMA block copolymer and functionalized with AS1411 aptamer for sensitive detection of circulating tumor cells.
Materials and Methods: The PGMA-b-PPDSMA block copolymer was synthesized and characterized by NMR spectroscopy and GPC. Glass slides were modified with APTES, allowing covalent attachment of the block copolymer via PGMA epoxy groups. The pyridyl disulfide groups of PPDSMA enabled site-specific immobilization of thiol-modified anti-nucleolin DNA aptamer (AS1411). Immobilization efficiency and aptamer specificity were validated using fluorescence microscopy with MCF-7 breast cancer cells.
Results: NMR and GPC results showed that PGMA-b-PPDSMA block copolymer with a molecular weight of 6 kDa, consisting of 4.8 kDa PGMA and 1.2 kDa PPDSMA block, was synthesized and purified. APTES modification on glass surfaces was confirmed by absorbance signals in the range of 1500-1650 cm-¹ belonging to the NH2 group in the FTIR spectrum. The binding efficiency of thiol-modified AS1411 aptamer to the PPDS group of PGMA-b-PPDSMA block copolymer was followed by absorbance change between 250-400 nm of pyridin-2-thione by-product formed during the reaction with time.
Conclusion: After DAPI staining and washing, Fluorescence microscopy images showed selective binding of MCF-7 cells to the AS1411-modified surfaces. These results suggest the surface provides a promising platform for sensitive capture of circulating cancer cells in early-stage liquid biopsy applications.

Anahtar Kelimeler

Liquid biopsy , Breast cancer , Polymer , DNA Aptamers , Biosensors , Breast cancer

Etik Beyan

Ethical approval was not required for this study, as only commercially available or previously established cell lines were used.

Destekleyen Kurum

This study was supported by the Afyonkarahisar Health Sciences University, Scientific Research Project Unit grant 22.TEMATİK.012.

Proje Numarası

22.TEMATİK.012

Kaynakça

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