Immunotherapy with chimeric antigen receptor (CAR) T cells has recently been shown to be successful in the treatment of B cell malignancies¹. Since epidermal growth factor receptor variant III (EGFRvIII), a glioma-specific antigen, is expressed in 24% to 67% of glioblastomas (GBM),^{1, 2} CAR-T cells have also been targeted to EGFRvIII for treating recurrent GBMs. EGFRvIII expression has been shown to promote oncogenesis and is associated with poor prognosis. Anti-EGFRvIII CAR-T cell therapy works by eliminating tumor cells without damaging normal tissue due to the tumor specificity of its target antigen³. Phase 1 clinical trials are currently ongoing to evaluate the safety and efficacy of this novel treatment paradigm. Since an acute inflammatory response has been observed in earlier similar trials⁴, it is important to evaluate treatment response using advanced MRI and MRS methods rather than relying on tumor volumetric changes. This hypothesis formed the basis of our current study in this exciting clinical trial.Eight recurrent GBM patients (3M/5F, mean age 62.4 ) with EGFRvIII expression underwent serial MRI scans after 1 and 2 months of CAR-T infusion. MRI scans included DTI, DSC and three-dimensional echo planar spectroscopic imaging (3D-EPSI). As the trial is ongoing, not all patients have undergone all the subsequent MRI scans. MRI was performed on a 3T scanner with a 12-channel phased-array head coil. DTI data were acquired using a single shot spin echo EPI sequence with parallel imaging using GRAPPA (acceleration factor = 2); TR/TE = 5000/86 ms, NEX = 3, FOV = 22 × 22 cm², b = 1000 s/mm², number of diffusion weighting directions = 30, in-plane resolution = 1.72 × 1.72 × 3 mm³. DSC T2* weighted gradient-echo echo planar images were obtained using the following parameters: TR/TE = 2000/45 ms, FOV = 22 × 22 cm², in-plane resolution = 1.72 × 1.72 × 3 mm³, and 20 slices covering the brain. EPSI was acquired using the following parameters: TR/TE = 1510/17.6 ms, spatial points = 50×50×18, FOV= 280×280×180mm³, voxel size = 5.6×5.6×10mm³, excitation angle = 73°, 512 complex points, spectral BW = 2500Hz with radiofrequency excitation pulse centered at water resonance, NEX = 1. Water suppression using frequency-selective saturation pulses and inversion-recovery nulling of lipid signal was performed with TI of 198ms. MD and FA maps were computed using in house software. Leakage corrected CBV maps were generated using Nordic ICE (Nordic Imaging Lab). NAA, Cr and Cho were computed using the metabolic imaging and data analysis system (MIDAS) package. Contrast-enhanced T1 weighted images, FLAIR, CBV and DTI maps were co-registered and a semi-automated segmentation routine was used to segment the contrast-enhancing ROI. The median MD, FA, rCBV and Cho/Cr values from this ROI were used to analyze the data and the 90th percentile rCBV values were measured to compute maximum rCBV (rCBV_max)⁵. The percent changes between the baseline and the subsequent scan (N) were calculated as (N – baseline)/baseline × 100.Representative MD, FA, CBV and Cho/Cr images from the baseline and follow-up scans in a patient are shown in Fig.1. After 1 month, an increase in MD with a concomitant decrease in FA was noted in this patient (Fig. 2). The Cho/Cr ratio decreased by 20% of the initial value, whereas the rCBV_max decreased to about 60% of the initial values after 2 months of CAR-T cell therapy in this patient (Fig. 2). Similar trends were observed in other patients.The efficacy of CAR-T cell therapy against EGFRvIII expressing GBMs has been reported in a murine model, which showed migration of CAR-T cells to the tumor with significant growth delay as well as prolonged survival³. The decrease in rCBV and Cho/Cr is suggestive of growth arrest and treatment induced response and inhibition of EGFRvIII expression since EGFRvIII is highly correlated with CBV⁵. Similarly a decrease in Cho has been shown to correlate with reduced cell proliferation, indicating cell death. A slight decrease in FA along with increase in MD also suggests cell death and increased extracellular volume fraction. However, the changes in FA and MD were not significant when measured in all patients. This observation may indicate the limited utility of DTI in monitoring response in this treatment paradigm. Our preliminary findings need to be evaluated in a larger patient cohort and correlated with clinical endpoints of progression free survival and overall survival to establish the role of CAR-T cell therapy in this subgroup of GBM patients. Advanced MRI and MRS metrics can serve as potential imaging biomarkers to assess for early treatment response in these patients.