This is an interventional, non-randomized, single site study. Brain tumor samples will be collected from patients for organoids generation and subject to panel drugs screening and QPOP analysis to derive the optimal drug combinations for treatment at the time of first high grade astrocytic glioma recurrence. The investigators hypothesize that patient-derived organoids (PDOs) mimic the biological characteristics of high grade astrocytic gliomas and serve as an ideal platform for the evaluation of drug sensitivities, accurately reflecting the patient's therapeutic response to the drugs.

This study aims to test the feasibility of Quadratic Phenotypic Optimization Platform (QPOP) application in high grade astrocytic glioma, by first establishing clinically relevant high grade astrocytic glioma organoid models, followed by the utility of QPOP-derived drug combinations in high grade astrocytic glioma.

The primary purpose of this study is not hypothesis testing, but to assess the feasibility of QPOP-guided therapy for recurrent high grade astrocytic glioma to be used in a larger scale study. Therefore, this study does not have a formal sample size, but rather, a set benchmarks to determine feasibility.

Specific Aim 1: To establish primary and corresponding temozolomide/radiotherapy (TMZ/RT)-resistant high grade astrocytic glioma patient-derived organoids.

Specific Aim 2: To determine the utility of QPOP (Quadratic Phenotypic Optimization Platform)-derived drug combinations in treating recurrent high grade astrocytic glioma.

Specific Aim 3: To evaluate the use of non-invasive imaging tools (Ga68-NEB PET/MRI and DCE-MRI) to track BBB permeability over time in high grade astrocytic glioma patients, and correlate this with pathological biomarkers, therapeutic response determined by follow up standard MRI and clinical outcomes.

Male and female subjects aged 21 years and above with suspected high grade astrocytic glioma planned for surgery/ biopsy followed by adjuvant chemoradiotherapy will be invited to participate in the pre-screening study. Subjects will only be enrolled in the main study if they had pathologically confirmed high grade astrocytic glioma, and received adjuvant treatment comprising standard-of-care therapy with surgery/biopsy followed by temozolomide and radiotherapy.

Pre-screening phase:

In pre-screening phase, patients will be approached for pre-screening consent at the time of first suspected diagnosis of a high grade astrocytic glioma and tumor will be harvested at the time of initial surgery/ biopsy for the generation of PDOs. Once the histological diagnosis of high grade astrocytic glioma is confirmed and the patient is planned for adjuvant temozolomide and radiotherapy, they will undergo baseline standard MRI plus Ga68-NEB PET/MRI and DCE-MRI imaging within one week of the planned radiotherapy simulation date. Patients will then undergo standard-of-care treatment for high grade astrocytic glioma with adjuvant concurrent temozolomide and radiotherapy (TMZ/RT)1, and will be evaluated by the primary oncologist routinely with clinical examination, laboratory tests and regular neuro-imaging. High grade astrocytic glioma organoids will be generated from resected tumour samples and QPOP analyses will be performed to identify specific drug combinations to guide clinical management at the time of first relapse.

Main study:

At the time of documented tumor recurrence, eligible patients will be reassessed for suitability to participate in the main study and approached for their informed consent to enter this phase of the study. During the main study, patients will receive QPOP-guided systemic therapy for the treatment of their relapsed high grade glioma and will be assessed regularly for safety and efficacy of this therapy. In addition, patients will undergo standard MRI plus investigational Ga68-NEB PET/MRI and DCE-MRI imaging prior to and 8 weeks (+/- 1 week) after QPOP-guided systemic therapy. Subsequent to this, radiological assessment of their disease will revert to standard clinical protocols with routine standard MRI imaging.

Inclusion Criteria:

Pre-screening:

1. Patients 21 years of age or older, with ECOG performance status 0 to 2, and with a life expectancy of more than 3 months with suspected high grade astrocytic glioma, fit for treatment comprising standard-of-care therapy with adjuvant temozolomide and radiotherapy if the diagnosis of high grade astrocytoma is pathologically confirmed.

2. Signed informed consent obtained before any study specific procedure. Subjects must be able to understand and be willing to sign the written informed consent.

   • Patients will be enrolled at the time of initial surgery but study imaging and further PDO generation will not take place if the patient is subsequently found not to meet the histological criteria or will not be receiving standard adjuvant temozolomide/ radiotherapy.

All subsequent criteria apply to the main study only:

1. Patients 21 years of age or older, with ECOG performance status 0 to 2, and life expectancy of more than 3 months with pathologically confirmed high grade astrocytic glioma, having undergone first-line standard-of-care therapy with surgery/biopsy followed by temozolomide and radiotherapy. Subjects with truncated adjuvant chemoradiotherapy may be enrolled at the Principal Investigator's discretion.
2. Documented tumor progression based on standard clinical, radiological or histological criteria, and deemed suitable for second line systemic therapy.
3. Sufficient tumor tissue available for PDO generation at baseline and at least one available or pending QPOP result.
4. Adequate organ function as defined by:

1. Bone marrow function i. Haemoglobin ≥ 9g/dl ii. Absolute neutrophil count (ANC) ≥ 1.5 x 109/L iii. Platelet count ≥ 100 x 109/L. 2. Liver function i. Bilirubin < 2.5x upper limit of normal (ULN) ii. Alanine transaminase (ALT) and aspartate transaminase (AST) < 2.5x ULN or < 5x ULN if liver metastases are present iii. Prothrombin time (PT) within the normal range for the institution. 3. Renal function i. Plasma creatinine <1.5x institutional ULN 5) Capable of swallowing tablets. 6) Recovery from any previous drug- or procedure-related toxicity to National Cancer Institute Common Terminology Criteria for Adverse Events (NCI-CTCAE) version 5.0 Grade 0 or 1 (except alopecia), or to baseline preceding the prior treatment.

Exclusion Criteria (both pre-screening/ main study)

1. Chemotherapy, radiotherapy, surgery, immunotherapy or other therapy within 2 weeks of study entry.
2. Pregnancy or breastfeeding at the point where systemic anti-cancer therapy is initiated. Women of childbearing potential must have a negative pregnancy test at the point where systemic anti-cancer therapy is initiated. Women of childbearing potential and men, must agree to use adequate contraception (barrier method of birth control) while on anti-cancer treatment and until at least 3 months after the last study drug administration.
3. Concurrent cancer which is distinct in primary site or histology from the cancer being evaluated in this study EXCEPT cervical carcinoma in situ, treated basal cell carcinoma, superficial bladder tumours (Ta, Tis & T1) or any cancer curatively treated less than 5 years prior to study entry.
4. Patients with leptomeningeal dissemination of disease and/or pure spinal high grade gliomas will be excluded.
5. Kidney disease which would clinically disqualify the subject from serial MRI scans with gadolinium contrast.