Abstract 105P
Background
Rhabdomyosarcoma (RMS) is the most frequent soft tissue sarcoma in childhood. This study evaluated the efficacy of SFX-01(Evgen Pharma plc, UK), a complex of synthetic d,l-sulforaphane stabilized in alpha-cyclodextrin, in two RMS in vitro models, alveolar (ARMS) and embryonal (ERMS), as single agent and in combination with ionizing radiation (IR).
Methods
Cell cycle and apoptosis were analysed by flow cytometry, whilst the modulation of specific genes was measured by real-time PCR and Western blotting assays. Irradiation was carried out exposing RMS cells to a single dose of 4 Gy and the radiosensitivity was assessed by clonogenic and modified high density survival assays. DNA damage repair (DDR)-related markers were studied by Western blotting and Protein Simple WES technology. The effects of SFX-01 on cancer stem cells were assessed by evaluating rhabdosphere formation in ultra-low attachment plates.
Results
SFX-01 (10 μM) treatment reduced the growth of RH30 (ARMS) and RD (ERMS) cells by inducing G2 cell cycle arrest with the concomitant Cyclin B1 up-regulation and triggering early-apoptosis. Interestingly, the specific knocking down of Cyclin D1 expression by RNA interference seems to enhance the cytostatic/cytotoxic effects induced by SFX-01 on RMS cells. We also found both p62 and LC3 II over-expression after SFX-01 exposure, this suggesting a block of late-stage autophagy. SFX-01 drug synergized with IR by strongly enhancing the effects of both single-agent treatments by showing a drastic reduction of clonogenic ability, block of DDR (persistent activation of γ-H2AX) and increased G2/M cell cycle arrest. Finally, pre-treatment with SFX-01, alone and in combination with IR, significantly affected the ability to form rabdospheres in both the parental and the clinically relevant radioresistant RMS cells.
Conclusions
For the first time, we described the antitumor activity of SFX-01 in preclinical models of RMS tumours. SFX-01 and IR have strong cytotoxic effects and drastically inhibit the formation of cancer stem cell-derived tumorspheres, this indicating that the combined treatment might have therapeutic benefits, mainly in the clinical managements of patients with aggressive RMS disease.
Clinical trial identification
Editorial acknowledgement
Legal entity responsible for the study
The authors.
Funding
Sapienza University of Rome and The Italian Foundation for Cancer Research (AIRC).
Disclosure
All authors have declared no conflicts of interest.