Background: Glioblastoma (GBM), accounting for 55% of primary malignant brain tumors, is characterized by its highly aggressive nature, poor prognosis, and high recurrence rate. Surgical resection is the primary treatment, yet recurrence is common, and survival rates remain dismal. Recurrent GBM lacks established interventions and standardized therapies. YSCH-01 is an oncolytic adenovirus, developed based on cancer-targeting gene-viro-therapy strategy (1), that has been incorporated with an interferon-like immune anticancer gene (L-IFN). Our preclinical studies have demonstrated significant tumor inhibition by YSCH-01 in both cell-line derived xenograft and patient-derived xenograft models (2). This investigator-initiated trial aims to evaluate the safety and efficacy of YSCH-01 in subjects with recurrent GBM. Methods: Six subjects with recurrent GBM are planned to be enrolled, receiving intracranial injections of YSCH-01 via Ommaya reservoir at a dosage of 5.0×1010 VP, once every 3 weeks for a total of 5 administrations, with a dose-limiting toxicity (DLT) assessment period of 3 weeks. Ommaya reservoir implantation will precede drug administration by one day. Inclusion criteria specify pathology-confirmed recurrent GBM, age 18-75 years, expected survival ≥ 12 weeks, and Karnofsky Performance Status (KPS) score ≥ 50 points, with at least one enhanced lesion of ≥ 1 cm diameter. Safety will be evaluated based on DLT and adverse events (AEs) assessed using CTCAE 5.0. Efficacy will be assessed every 5 weeks using Response Assessment in Neuro-Oncology (RANO) criteria, with overall survival follow-up at 12 weeks for two years. Two of the planned six cases have been enrolled, completing the DLT period without DLTs. The trial is registered with Clinical Trial Registry under NCT05914935. Clinical trial information: NCT05914935.