Effect of doxorubicin on Caspase-1/GSDMD-mediated pyroptosis in MCF-7 breast cancer spheroids

Işıl Aydemir; Emre Çetindağ

doi:10.19161/etd.1684512

TR EN

Doksorubisinin MCF-7 meme kanseri sferoidlerinde Caspase-1/GSDMD aracılı piroptoz üzerine etkisi

Öz

Amaç: Doksorubisin, yaygın olarak kullanılan bir antrasiklin antibiyotiği olup antitümör etkilerini DNA interkalasyonu, topoizomeraz II inhibisyonu ve oksidatif stresin indüklenmesi gibi çeşitli mekanizmalar aracılığıyla gösterir. Bu çalışma, doksorubisinin Kaspaz-1 ve Gasdermin D (GSDMD) aracılığıyla inflamatuar hücre ölümü yolu olan piroptozu aktive etme potansiyelini ve üç boyutlu (3B) hücre kültürü modelinde sferoid morfolojisi üzerindeki etkisini değerlendirmeyi amaçlamaktadır. Gereç ve Yöntem: Sferoidler 3B hücre kültürü teknikleriyle oluşturulmuş ve farklı dozlardaki doksorubisin (0,1–100 µM) ile 24 ve 48 saat süreyle tedavi edilmiştir. Sferoid sayısı ve çapı ImageJ v1.47 yazılımı ile analiz edilmiştir. Kaspaz-1 ve GSDMD protein düzeyleri, piroptotik yolların aktivasyonunu değerlendirmek için ELISA yöntemiyle ölçülmüştür. Bulgular: Doksorubisin uygulaması, sferoid sayısı ve çapında doz bağımlı azalmaya yol açmıştır. 100 µM’de, sferoid oluşumu tamamen inhibe edilmiş ve dağılmış hücre kümeleri gözlenmiştir. Kaspaz-1 ve GSDMD protein düzeyleri, yüksek doksorubisin konsantrasyonları ile anlamlı şekilde artmış, maksimum ekspresyon 25 µM’de belirlenmiştir; bu da inflamatuar hücre ölüm yollarının aktive olduğunu göstermektedir. 24 ve 48 saatlik tedavi süreleri arasında anlamlı bir fark saptanmamıştır. Sonuç: Doksorubisin, 3B sferoid yapısını bozarak ve piroptozla ilişkili proteinlerin ekspresyonunu doz bağımlı şekilde artırmaktadır. Bu bulgular, doksorubisinin tümör yapısını bozan ve inflamatuar hücre ölüm mekanizmalarını aktive eden iki yönlü etkisini vurgulayarak, solid tümörlerdeki terapötik potansiyeline dair önemli ipuçları sunmaktadır.

Anahtar Kelimeler

Effect of doxorubicin on Caspase-1/GSDMD-mediated pyroptosis in MCF-7 breast cancer spheroids

Abstract

Aim: Doxorubicin, a widely used anthracycline antibiotic, exerts its antitumor effects through multiple mechanisms, including DNA intercalation, inhibition of topoisomerase II, and the induction of oxidative stress. This study aimed to evaluate its potential to activate the inflammatory cell death pathway, pyroptosis via Caspase-1 and Gasdermin D (GSDMD), as well as its impact on spheroid morphology in a three-dimensional (3D) cell culture model. Materials and Methods: Spheroids were formed using 3D cell culture techniques and treated with increasing concentrations of doxorubicin (0.1–100 µM) for 24 and 48 hours. Spheroid number and diameter were analyzed using ImageJ v1.47. Caspase-1 and GSDMD protein levels were quantified by ELISA to assess the activation of pyroptotic pathways. Results: Doxorubicin treatment led to a dose-dependent reduction in both spheroid number and size. At 100 µM, spheroid formation was completely inhibited and replaced by disorganized cell clusters. Caspase-1 and GSDMD protein levels increased significantly with higher doxorubicin concentrations, with peak expression observed at 25 µM; suggesting activation of inflammatory cell death pathways. No significant differences were found between 24- and 48-hour treatments. Conclusion: Doxorubicin disrupts 3D spheroid structure and induces the expression of pyroptosis-related proteins in a concentration-dependent manner. These findings highlight its dual role in both impairing tumor architecture and activating inflammatory cell death mechanisms, providing insights into its therapeutic potential in solid tumors.

Keywords

Ethical Statement

Dear Editor, In our study titled "Effect of doxorubicin on Caspase-1/GSDMD-mediated pyroptosis in MCF-7 breast cancer spheroids", we used a commercially obtained cell line, the name of the company from which the cell line was obtained and the registration number are provided in the ‘’Materials and Methods’’ section of the manuscript. Cells isolated directly from humans were not used; therefore, ethical committee approval is not required.

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Details

Primary Language

English

Subjects

Cancer Cell Biology , Histology and Embryology

Journal Section

Research Article

Authors

Işıl Aydemir ^*
0000-0002-4143-7319
Türkiye

Emre Çetindağ
0000-0001-8063-8060
Türkiye

Publication Date

December 8, 2025

Submission Date

April 26, 2025

Acceptance Date

June 19, 2025

Published in Issue

Year 2025 Volume: 64 Number: 4

DOI

https://doi.org/10.19161/etd.1684512

IZ

https://izlik.org/JA66ES67PE

Cite

RIS / Bibtex

Vancouver

1.Işıl Aydemir, Emre Çetindağ. Effect of doxorubicin on Caspase-1/GSDMD-mediated pyroptosis in MCF-7 breast cancer spheroids. EJM. 2025 Dec. 1;64(4):598-605. doi:10.19161/etd.1684512