Purpose: Recent clinical successes in the use of chimeric antigen receptor (CAR) T cell therapy has revolutionized cancer therapy. However, only 20% to 30% of patients achieve long term survival benefits, and distinguishing responders from non-responders early remains a challenge with conventional imaging techniques. Thus, there is an urgent need to develop new biomarkers to distinguish responsive and resistant patients, both to improve standard of care and to assess the antitumor activity of experimental immunotherapies. This study aims to determine the efficacy of a novel granzyme activated imaging probe to image treatment response to CAR T cell therapy in a mouse model.Methods: Immunodeficient mice were obtained and inoculated subcutaneously with Raji tumors. CD8+ T cells were obtained from donor blood and transduced to express anti-CD19 receptors. CAR T cells were then expanded in vitro with IL-2. After tumors were palpable, mice were treated with empty or anti-CD19 CAR T cells intravenously. Mice were then injected with 64Cu-GRIP B at 24 hours post CAR T cell administration. PET/CT studies were performed on a dedicated Inveon small animal scanner. Post mortem radiotracer uptake was quantified as %injected dose/g (%ID/g) values for tumor and normal tissue. Tumors from one representative animal per group was prepared for autoradiography. Results: ROI analysis of static PET/CT images indicated that 64Cu-GRIP B uptake in treated tumors rose from 0.5 to 2 hours post injection. Tumoral uptake of the probe was higher in anti-CD19 CAR T versus vehicle treated arm at 2 hours post injection. The mean tumoral standard uptake values for anti-CD19 CAR T and vehicle arms were 1.5%ID/cc and 0.4%ID/cc, respectively. Biodistribution data demonstrated similar uptake of the 64Cu-GRIP B probe between treatment arms as mean %ID/g for both arms was roughly 0.75. Digital autoradiography suggested substantially higher and localized uptake of the probe in the treatment arm compared to the vehicle arm in several mice. Conclusions: Using 64Cu-GRIP B, a peptide-based chemosensor whose biodistribution was engineered to be controlled by the proteolytic activity of secreted GZMB allows for imaging granzyme B mobilization by cytotoxic T cells in CAR T cell therapy. Further studies with larger samples sizes and standardized batches of CAR T cells will provide more conclusive results.
The diagnosis and management of the phenotypically heterogeneous and progressive disorders tuberous sclerosis complex (TSC) and lymphangioleiomyomatosis (LAM) is a significant clinical challenge owing to a lack of disease specific biomarkers. Because the genetic mutations in TSC1 and/or TSC2 characteristic of TSC and LAM activate mTORC1 signaling, we hypothesized that disease burden could be measured with PET using 89Zr-transferrin (Tf). Toward this goal, we show that spontaneous renal cystadenomas arising in a genetically engineered mouse model heterozygous for Tsc2 were readily detectable with 89Zr-Tf PET. Moreover, subcutaneous implants of TSC and LAM cell line models consistently harbored high avidity for 89Zr-Tf in vivo. Deeper mechanistic studies showed that transferrin receptor expression and Tf biology were mTORC1 regulated in TSC and LAM models. Finally, the early treatment effects of clinically approved and experimental systemic therapies for TSC and LAM were interpretable using 89Zr-Tf PET. In summary, these data advance a translatable molecular imaging strategy that may be capable of detecting and longitudinally monitoring whole body TSC and LAM disease burden.
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