ESTRO 2022 - Abstract Book

S370

Abstract book

ESTRO 2022

was confirmed by an increased HR capacity and replication fork stability after irradiation and resulted in significantly decreased yH2AX- and RPA foci after irradiation (p<0.001). The avoidance of radiation-induced replication stress resulted in a significantly lower accumulation of dsDNA in the cytoplasm and a low cGAS/STING/IRF3 activation. Indeed, the proportion of ALDH1-positive CSC correlated significantly with the amount of cytosolic dsDNA after irradiation (p<0.001). Strikingly, the CSC showed an endogenously increased expression of PD-L1 and PD-L2, which was increased ~3 fold after fractionated irradiation (5x5.2 Gy). The inhibition of ATR led to a distinct radiosensitization of the radioresistant CSC (EF=3) and significantly reduced PD-L1 expression. Furthermore, ATR-inhibition to a significant increase of nuclear IRF3 after irradiation, thus an increased activation of the intracellular immune response (p<0.001). Conclusion The results show that CSC express more PD-L1 and PD-L2 and minimize the formation of cytosolic DNA after irradiation through enhanced DSB repair and protection of replication forks by HR. Disruption of the ATR-CHK1 signaling pathway by ATR inhibition leads to radiation sensitization, reduced PD-L1 expression and increased activation of the cGas-STING pathway. Therefore, we hypothesize that inhibitors to inactivate the S-phase DNA damage response, such as ATR, may be used to further develop existing therapies in the future. I. Gorodetska 1 , A. Offermann 2 , J. Püschel 3 , V. Lukiyanchuk 1 , D. Gaete 4 , A. Kurzyukova 5 , V. Labitzky 6 , F. Schwarz 1 , T. Lange 6 , F. Knopf 7 , B. Wielockx 4 , M. Krause 8 , S. Perner 2 , A. Dubrovska 1 1 Helmholtz-Zentrum Dresden-Rossendorf - Institute of Radiooncology (OncoRay), Biomarkers for the Individualized Radiotherapy, Dresden, Germany; 2 Institute of Pathology, University Hospital Schleswig-Holstein, Pathology, Research Center Borstel, Leibniz Lung Center, Luebeck, Germany; 3 Helmholtz-Zentrum Dresden-Rossendorf - Institute of Radiooncology (OncoRay), Biomarkers for the Individualized Radiotherapy, Dresden , Germany; 4 Institute of Clinical Chemistry and Laboratory Medicine, Technische Universität Dresden, Dresden, Germany; 5 CRTD - Center for Regenerative Therapies,Technische Universität Dresden, Center for Healthy Aging, Dresden, Germany; 6 Institute of Anatomy and Experimental Morphology, Center for Experimental Medicine, University Cancer Center Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; 7 CRTD - Center for Regenerative Therapies, Technische Universität Dresden, Center for Healthy Aging, , Dresden, Germany; 8 Institute of Radiooncology (OncoRay), Helmholtz-Zentrum Dresden-Rossendorf, Dresden, Germany Purpose or Objective Aldehyde dehydrogenase (ALDH) high activity is a marker of PCa stem cells with increased therapy resistance, enhanced DNA double-strand break repair, and activation of epithelial to mesenchymal transition (EMT). We investigated the role of two ALDH genes, ALDH1A1 and ALDH1A3, in regulating prostate cancer radiation resistance, cancer stem cell (CSC) phenotype and metastases formation. Materials and Methods We conducted our study by employing in vitro radiobiological clonogenic and spherogenic assays and in vivo syngeneic mouse model, zebrafish larval and mouse xenograft tumor models. We also validated our findings on the patient’s tumor samples by immunohistochemical staining of ALDH1A1 and ALDH1A3 protein expression (N=613). Results We found that ALDH1A1 was highly upregulated, whereas ALDH1A3 was significantly downregulated in radiation-resistant (RR) cells. Radiobiological clonogenic analyses after siRNA-mediated knockdown, as well as western blot analysis of key DNA damage response proteins, demonstrated that both ALDH1A1 and ALDH1A3 contribute to the regulation of PCa cell radiosensitivity. We analyzed the association of the functional gene signatures with ALDH1A1 and ALDH1A3 gene expression in the TCGA gene expression dataset and found a high positive correlation of the ALDH1A1 expression with WNT/ β -Catenin, extracellular matrix and adhesion, EMT, angio- and osteogenesis gene sets. Contrary, the ALDH1A3 showed a negative correlation with those gene sets but a high correlation with androgen receptor (AR) signalling. We next validated our findings in knockdown experiments, as well as by using inhibitors against AR and CTNNB1 ( β -Catenin) genes. We also compared the initial prostate-specific antigen (iPSA) serum level preoperatively between patients with tumors overexpressing ALDH1A1 and ALDH1A3 and observed a significantly increased iPSA level in patients with ALDH1A3 overexpressing tumors. Mechanistically, ALDH1A1 and ALDH1A3 differentially influence the TGFB1 mRNA gene expression, a potent inducer of EMT. Our in vivo zebrafish larval model showed that PCa cells lacking ALDH1A3 exhibit higher extravasation potential than those with suppressed ALDH1A1. The syngeneic mouse model demonstrated that Aldh1a1 depleted cells had reduced bone homing potential while Aldh1a3 suppressed cells formed a higher number of tumor nodules in the bone marrow. The analysis of the patient’s cohort showed the overexpression of ALDH1A1 (N=613) in lymph node and distant metastasis, whereas the ALDH1A3 protein (N=325) was highly expressed in primary prostate tumors. Additionally, Kaplan-Meier survival analyses of PCa patients with clinical follow-up information showed that high ALDH1A1 expression predicts disease recurrence. Conclusion Overall, ALDH1A1 and ALDH1A3 have opposite effects on PCa metastatic dissemination, with ALDH1A1 associated with increased metastatic burden and biochemical failure in PCa patients. OC-0426 ALDH genes as regulators of prostate cancer radioresistance, stemness and bone metastases

OC-0427 Interplay of cancer stemness and amino acid transporters regulates radioresistance in HNSCC