Background: High grade gliomas (HGG) including glioblastoma (GBM) are among the most aggressive brain cancers, with patients exhibiting highly variable treatment responses in both newly diagnosed (ND) and recurrent disease. Temozolomide (TMZ) + radiation therapy is the guideline directed standard of care (SOC) in the ND setting; it has remained relatively unchanged for > 15 years, despite variable patient responses. Current available biomarkers do not inform personalized therapy. Functional drug response testing using patient specific tumor cells may have the potential to inform effective therapy selection, thus advancing functional precision oncology. Progression free survival (PFS) is a meaningful surrogate to overall survival (OS) in GBM and therefore, represents a measure of clinical benefit. Methods: The 3D-PREDICT clinical study (NCT03561207) allows enrollment of HGG patients with ECOG ≤ 3, perhaps representing a more accurate real-world population compared to most clinical studies in the same patient cohort. Tumor tissue was prospectively collected during SOC biopsy/resection and analyzed in an ex vivo cell culture test using a panel of agents prescribed as HGG therapeutics. Results: Data pertain to 56 3D-PREDICT patients who had > 6 months of follow up as of December 31, 2021 or experienced progression/death < 6 months post tissue collection. There were 42 3D-PREDICT patients who had IDH wild type ND GBM and received SOC. PFS analysis of these patients showed 3D ex vivo testing was able to prospectively predict TMZ clinical responders vs. non-responders (Kaplan-Meier, p = 0.039; HR 0.516, 95% CI 0.234,1.137). Test predicted TMZ responders had a relative PFS advantage of 3.7 months. Of the 42 patients, 38 had known MGMT methylation status with 23 patients (60%) being unmethylated. Test predicted responders (9) included unmethylated patients; test predicted non-responders (33) included methylated patients. The data suggest ex vivo testing of patient specific tumor tissue may identify ND HGG patients a priori who respond to TMZ, irrespective of MGMT methylation status. Beyond TMZ response prediction, the test assesses tissue response to 11 additional known HGG therapies and therefore may provide a tool to inform potential alternative treatments to TMZ for ND patients. Also evaluated were 14 3D-PREDICT recurrent HGG patients who received test directed salvage therapy; mean PFS after tissue collection from re-resection was 9.0 months (range 2.3 – 24.7 mo) for 9 patients at first recurrence, and 5.7 months (range 1.8 – 11.4 mo) for 5 patients with 2 to 6 recurrences. These examples demonstrate initial test utilization informing salvage therapy selection, correlated with PFS improvements in recurrent HGG. Conclusions: This functional 3D ex vivo cell culture platform provided survival benefit in analyzed ND and recurrent cohorts. Clinical trial information: NCT03561207.