Abstract Objective. Carbon is an ion species of significant radiobiological interest, particularly in view of its use in cancer radiotherapy, where its large Relative Biological Efficiency is often exploited to overcome radio resistance. A growing interest in highly pulsed carbon delivery has arisen in the context of the development of the FLASH radiotherapy approach, with recent studies carried out at dose rates of 40 Gy s⁻¹. Laser acceleration methods, producing ultrashort ion bursts, can now enable the delivery of Gy-level doses of carbon ions at ultra-high dose rates (UHDRs), exceeding 10⁹ Gy s⁻¹. While studies at such extreme dose rate have been carried out so far using low LET particles such as electrons and protons, the radiobiology of high-LET, UHDR ions has not yet been explored. Here, we report the first application of laser-accelerated carbon ions generated by focussing 10²⁰ W cm⁻² intense lasers on 10–25 nm carbon targets, to irradiate radioresistant patient-derived Glioblastoma stem like cells (GSCs). Approach. We exposed GSCs to 1 Gy of 9.5 ± 0.5 MeV/n carbon ions delivered in a single ultra-short (∼400-picosecond) pulse, at a dose rate of 2 × 10⁹ Gy s⁻¹, generated using the ASTRA GEMINI laser of the Central Laser Facility at the Rutherford Appleton Laboratory, Didcot, Oxfordshire, UK. We quantified carbon ion-induced DNA double strand break (DSB) damage using the 53BP1 foci formation assay and used 225 kVp x-rays as a reference radiation. Main Results. Laser-accelerated carbon ions induced complex DNA DSB damage, as seen through persistent 53BP1 foci (11.5 ± 0.4 foci/cell/Gy) at 24 h and significantly larger foci (1.69 ± 0.07 μm²) than x-rays induced ones (0.63 ± 0.02 μm²). The relative foci induction value for laser-driven carbon ions relative to conventional x-rays was 3.2 ± 0.3 at 24 h post-irradiation also confirming the complex nature of the induced damage. Significance. Our study demonstrates the feasibility of radiobiology investigations at unprecedented dose rates using laser-accelerated high-LET carbon ions in clinically relevant models.