ESTRO 2023 - Abstract Book

S2010

Digital Posters

ESTRO 2023

Murine CT26 colorectal cancer cells and 4T1 breast cancer cells were exposed to H2O2 for 1 hour. The toxicity and radio- modulatory effects of H2O2 were determined under static (0.1% oxygen in a hypoxic chamber) and metabolic hypoxia (micropellets) by colony formation assay. Reactive oxygen species (ROS) levels (DCFDA), DNA damage ( γ H2AX), apoptosis (Annexin-V/7-AAD) and ferroptosis (C11BODIPY) were examined by flow cytometry. Oxygen consumption rate (OCR) and mitochondrial electron transport chain (ETC) complex activity was measured by Seahorse analyzer. The oxygen pressure after H2O2 injection was analyzed in real-time under hypoxic conditions (tissue mimetic culture system (TMCS)) using an oxygen monitor. Results Non-toxic concentrations of H2O2 were determined per cell line and used subsequently. Treatment with H2O2 radiosensitized CT26 and 4T1 cells under both static and metabolic hypoxic conditions. Enhancement ratios achieved were respectively 2.38 and 1.67 under static hypoxic conditions and 1.44 and 2.21 under metabolic hypoxic conditions. ROS levels did not increase following 1 hour treatment with H2O2 whereas increased levels of ferroptosis, apoptosis and DNA damage were observed with H2O2 alone, yet did not result in an enhanced augmentation in combination with radiation. A dose- dependent decrease in OCR was observed in both cell lines after H2O2 treatment correlating to an inhibition of the activity of ETC complex I, and to a lesser extent of complex II. Injection of H2O2 in our TMCS resulted in an immediate and significant surge in oxygen levels. Conclusion The radiosensitizing effect of H2O2 has been primarily attributed to the induction of apoptosis. However, our preliminary results suggest that the main radiosensitizing mechanism of H2O2 under hypoxic conditions can be ascribed to reoxygenation of cells after the conversion of H2O2 to water and oxygen along with inhibition of cellular respiration through a complex I blockade. Both the radiosensitizing and oxygen-releasing effects of packaged H2O2 in vivo are currently under investigation. Gaining knowledge on the underlying working mechanisms of H2O2 as a radiosensitizer is vital in designing future combination therapies for cancer patients. M. Spa ł ek 1,2 , K. Bobak 3 , A. Tysarowski 4 , A. Szumera-Cie ć kiewicz 5,6 , K. Sokó ł 5 , M. W ą grodzki 7 , H. Kose ł a-Paterczyk 3 , P. Rutkowski 1 , A.M. Czarnecka 3,8 1 Maria Sklodowska-Curie National Research Institute of Oncology, Department of Soft Tissue/Bone Sarcoma and Melanoma, Warsaw, Poland; 2 Maria Sklodowska-Curie National Research Institute of Oncology, Department of Radiotherapy I, Warsaw, Poland; 3 Maria Sklodowska-Curie National Research Institute of Oncology, Department of Soft Tissue/Bone Sarcoma and Melanoma , Warsaw, Poland; 4 Maria Sklodowska-Curie National Research Institute of Oncology, Cancer Molecular and Genetic Diagnostics Laboratory, Warsaw, Poland; 5 Maria Sklodowska-Curie National Research Institute of Oncology, Department of Pathology and Laboratory Diagnostics, Warsaw, Poland; 6 Institute of Hematology and Transfusion Medicine, Diagnostic Hematology Department, Warsaw, Poland; 7 Maria Sklodowska-Curie National Research Institute of Oncology, Department of Pathology and Laboratory Diagnostics , Warsaw, Poland; 8 Mossakowski Medical Research Institute, Polish Academy of Sciences, Department of Experimental Pharmacology, Warsaw, Poland Purpose or Objective The locally advanced marginally resectable primary soft tissue sarcomas (MRSTS) require neoadjuvant treatment before the attempt of surgery. The study aimed to assess genetic changes in MRSTS that may be predictive factors in MRSTS treatment. Materials and Methods We analysed preoperative core biopsy samples of 20 patients with MRSTS who received 5 × 5 Gy radiotherapy combined with three cycles of doxorubicin-ifosfamide chemotherapy in a phase II clinical trial (NCTXXXXXXXX). Enrolled patients included eleven patients with undifferentiated pleomorphic sarcoma (UPS), one patient with leiomyosarcoma (LMS), five patients with myxofibrosarcoma (MFS), one patient with pleomorphic liposarcoma (PLPS), one patient with dedifferentiated liposarcoma (DDLPS), one patient with malignant peripheral nerve sheath tumor (MPNST). The pathological response to neoadjuvant therapy was assessed using the European Organization for Research and Treatment of Cancer-Soft Tissue and Bone Sarcoma Group (EORTC-STBSG) recommendations for pathological examination and reporting. Next-Generation Sequencing (NGS) Illumina NGS TruSight Oncology 500 kit was used to assess over 500 genes. Results The analyzed group presented a low tumor mutation burden and no microsatellite instability. The identified mutations were presented in Table 1. There were five patients with good response to therapy (A = 1, B = 1, C = 3) and 15 patients with poor response to therapy (D = 10, E = 5). All but one mutation with strong pathogenic prediction were found in poor responders to neoadjuvant radiochemotherapy (Table 1). An intronic mutation of TP53 was found in one patient with UPS who presented an excellent response to therapy. PO-2234 Genetic abnormalities as a predictive factor for neoadjuvant treatment in soft tissue sarcomas

Table 1. Results of molecular analysis Gene Type of mutation

Mutation variant in codon

Mutation variant in protein

EORTC-STBSG response grade

Subtype of sarcoma

- -

- - -

E D

PRUNE2/NTRK2 Gene fusion PRUNE2/NTRK2 Gene fusion

Leiomyosarcoma Myxofibrosarcoma Myxofibrosarcoma

D, E

MDM2

Amplification -

Malignant peripheral nerve sheath tumor

-

D

MDM2

Amplification -