GBM remains a therapeutically challenging malignancy, partly due to its complex cytokine mediated microenvironment. This study established a novel microfluidic platform to maintain patient derived GBM tissue biopsies ex vivo for up to 12 days, enabling the dynamic profiling of cytokine responses under treatment (1μM GSK (Glycogen Synthase Kinase) 3368715 + 10μM Temozolomide). The combination of biochemical assays (LDH activity), histological analysis (H&E staining and Immunohistochemistry (IHC) for apoptotic markers), cytokine proteome profiling and ELISA assays provides evidence that GBM tissues can be maintained in a viable state within the novel microfluidic perfusion device for at least 12 days. Cytokine profiling was performed using Proteome Profiler array for 105 different cytokines, followed by ELISA validation for seven key cytokines (VEGF, MMP9, CHI3L1, IL6, IL8, Serpin E1, and Angiopoietin-2).
Multivariate analysis showed time significantly influenced cytokine profiles (p = 0.0288), with an overall downregulation over time. Treatment (temozolomide plus arginine methylation inhibitor) had no significant global effect on all cytokines expression level (p = 0.1977), but did significantly affect the seven selected cytokines (p = 0.0048). Univariate analysis confirmed VEGF (p = 0.0004) and MMP9 (p = 0.0046) were significantly downregulated with treatment, while CHI3L1 was upregulated (p = 0.0088). Additionally, gender (p = 0.0050) and age group (<60 vs. >60 years; p = 0.0021) were both significant covariates influencing cytokine expression. However, these findings should be interpreted with caution given the small sample size (n=13), particularly the underrepresentation of female patients (3 females vs. 10 males) and the age imbalance (4 under 60 vs. 9 over 60).
Three of the study cytokines, VEGF, MMP9, and CHI3L1, were identified as responsive biomarkers under drug treatment, supporting further exploration of these targets. The platform provides a valuable tool for preclinical therapeutic screening and biomarker discovery. With further optimisation, it could support personalised treatment strategies and contribute to the development of more effective, patient-tailored approaches for managing GBM.
