Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2025 Nov 26.
doi: 10.1007/s00292-025-01507-w. Online ahead of print.

POLQ and DNA-PK inhibition in muscle-invasive bladder cancer : Enhancing radiosensitivity with novel DNA damage response inhibitors to improve radiotherapy outcomes

Julia Pannhausen  1   2 , Ahmed A Chughtai  2   3 , Michael K Melzer  4   5 , Yanchun Ma  4   5 , Julia Wirtz  1   2 , Alexander Kleger  5   6   7 , Matthias Saar  2   8 , Michael J Eble  2   3 , Danny D Jonigk  1   2   9 , Michael Rose  10   11   12 , Nadine T Gaisa  13   14   15
Affiliations

POLQ and DNA-PK inhibition in muscle-invasive bladder cancer : Enhancing radiosensitivity with novel DNA damage response inhibitors to improve radiotherapy outcomes

Julia Pannhausen et al. Pathologie (Heidelb). .

Abstract
in English, German

Background: Muscle-invasive bladder cancer (MIBC) includes histological subtypes such as urothelial carcinoma (UC) and the rarer, prognostically unfavorable, squamous cell carcinoma (SCC). Standard treatment is radical cystectomy, while alternatives like chemotherapy or radiotherapy are particularly limited in SCC. Radiosensitizers, such as DNA-dependent protein kinase (DNA-PK) and DNA polymerase theta (POLQ) inhibitors, could selectively enhance radiotherapy effects in tumor cells and represent promising approaches for clinical translation.

Objective: This study analyzed the radiosensitizing effects of DNA-PK and POLQ inhibition ex vivo in patient-derived MIBC cell lines of UC and SCC subtypes.

Materials and methods: The DNA-PK inhibitor AZD7648 (DNA-PKi) and the POLQ inhibitor ART558 (POLQi) were tested ex vivo in patient-derived SCC (p-SCC; n = 3) and UC (p-UC; n = 3) cell cultures. Effects were assessed in combination with ionizing radiation (IR) using XTT cell viability assays (IC50), clonogenic survival assays, γH2AX immunofluorescence, and comet assays.

Results: DNA-PKi strongly radiosensitized MIBC cultures ex vivo, reducing IC50-XTT values by 74-99% and survival rates by 34-64%. Under POLQi +2 Gy, IC50 decreased by 7-13%, whereas under POLQi +8 Gy it increased by 11-24%, with only ~5% reduction in survival. DNA-PKi markedly delayed DNA repair (comet tail moments 38-40%, γH2AX foci increased 11.9-13.1-fold), while POLQi showed minimal effects (comet tail moments 22-33%, γH2AX foci increased 5.4-6.0-fold).

Conclusion: DNA-PKi radiosensitized MIBC cells more effectively than POLQi, particularly SCC. DNA damage response (DDR) inhibitors thus have therapeutic potential in MIBC, depending on the target protein and tumor subtype.

Zusammenfassung: HINTERGRUND: Muskelinvasive Harnblasenkarzinome (MIBC) umfassen histologische Subtypen wie Urothelkarzinome (UC) oder seltenere, prognostisch ungünstigere Plattenepithelkarzinome (engl. squamous cell carcinoma [SCC]). Standardtherapie ist die radikale Zystektomie. Alternativen wie Chemo- oder Radiotherapie sind besonders beim SCC begrenzt wirksam. Radiosensibilisatoren wie DNA-PK- und POLQ-Inhibitoren könnten die Strahlenwirkung in Tumorzellen gezielt verstärken und bieten vielversprechende Ansätze für klinische Translation.

Ziel der arbeit: In der vorliegenden Studie wurde ex vivo die radiosensibilisierende Wirkung der Inhibition der DNA-abhängigen Proteinkinase (DNA-PK) und der DNA-Polymerase Theta (POLQ) in patientenabgeleiteten MIBC-Zelllinien der Subtypen UC und SCC analysiert.

Methoden: Der DNA-PK-Inhibitor AZD7648 (DNA-PKi) und der POLQ-Inhibitor ART558 (POLQi) wurden ex vivo in patientenabgeleiteten SCC- (p-SCC, n = 3) und UC-Zellkulturen (p-UC, n = 3) getestet. Die Effekte wurden unter Kombination mit ionisierender Strahlung (IR) mittels XTT-Zellviabilitätsassays (IC50), klonogener Überlebensassays, γH2AX-Immunfluoreszenz und Comet-Assays analysiert.

Ergebnisse: DNA-PKi führte zu einer starken Radiosensibilisierung der ex vivo MIBC-Zellkulturen, mit Reduktionen der IC50-XTT-Werte um 74–99 % und der Überlebensraten um 34–64 %. Unter POLQi +2 Gy sank der IC50-Wert um 7–13 %, unter POLQi +8 Gy stieg er um 11–24 %, und die Überlebensraten waren nur um 5 % reduziert. DNA-PKi verzögerte die DNA-Reparatur deutlich (Comet-Tail-Moments 38–40 %, Steigerung γH2AX-Foci 11,9- bis 13,1-fach), während POLQi nur minimale Effekte zeigte (Comet-Tail-Moments 22–33 %, Steigerung der γH2AX-Foci von 5,4- bis 6,0-fach).

Schlussfolgerung: Der DNA-PKi radiosensibilisierte MIBC-Zellen stärker als der POLQi, insbesondere SCC. Somit haben DNA-Damage-Response(DDR)-Inhibitoren abhängig vom Zielprotein und Subtypen im MIBC therapeutisches Potenzial.

Keywords: ART558; AZD7648; Radiosensitization; Squamous bladder cancer; Urothelial carcinoma.

PubMed Disclaimer

Conflict of interest statement

Declarations. Conflict of interest: J. Pannhausen, A.A. Chughtai, M.K. Melzer, Y. Ma, J. Wirtz, A. Kleger, M. Saar, M.J. Eble, D.D. Jonigk, M. Rose, and N.T. Gaisa declare that they have no competing interests. Patients gave written informed consent, and experiments followed the regulations of the RWTH centralized biomaterial bank and the Declaration of Helsinki. This study was approved by the Institutional Ethical Review Board of the Medical Faculty of RWTH Aachen University (EK 268/21, EK 206/09 study number 199 and 311). The supplement containing this article is not sponsored by industry.

References

    1. Barletta F, Tappero S, Panunzio A et al (2022) Differences in cancer-specific mortality after trimodal therapy for T2N0M0 bladder cancer according to histological subtype. Cancers 14(23):5766
    1. Blackford AN, Jackson SP (2017) ATM, ATR, and DNA-PK: the trinity at the heart of the DNA damage response. Mol Cell 66(6):801–817 - DOI - PubMed
    1. Ceccaldi R, Liu JC, Amunugama R et al (2015) Homologous-recombination-deficient tumours are dependent on Polθ-mediated repair. Nature 518(7538):258–262 - DOI - PubMed - PMC
    1. Choudhury A, Porta N, Hall E et al (2021) Hypofractionated radiotherapy in locally advanced bladder cancer: an individual patient data meta-analysis of the BC2001 and BCON trials. Lancet Oncol 22(2):246–255 - DOI - PubMed - PMC
    1. Chughtai AA, Pannhausen J, Dinger P et al (2022) Effective radiosensitization of bladder cancer cells by pharmacological inhibition of DNA-PK and ATR. Biomedicines 10(6):1277