CD30CAR⁺ EBV-CTLs can control tumor growth in vivo while retaining their ability to migrate to EBV⁺ tumor and expand. To evaluate in vivo homing, NT EBV-CTLs or CTLs transduced with CD30CAR and sorted for transgene expression were injected intravenously in SCID mice implanted subcutaneously with autologous LCLs.³²  Both NT and CAR⁺ EBV-CTLs were labeled with the eGFP-FFLuc gene to monitor their trafficking and expansion using an in vivo imaging system (Xenogen-IVIS Imaging System). (A) Circa 50% of CD30CAR⁺ EBV-CTLs, as assessed by the goat anti–human IgG (H + L) Ab, are expressing the FFLuc transgene as GFP⁺. (B) The signal of EBV-CTLs is localized to the EBV⁺ tumors and is elevated in mice receiving either control (top panels) or CD30CAR⁺ EBV-CTLs (bottom panels). (C) The bioluminescence fold expansion of CTLs at the tumor site is shown. To evaluate the contribution of costimulation²¹  by EBV antigen, EBV-CTLs transduced with irrelevant CAR or CD30CAR were injected intraperitoneally in SCID mice bearing EBV⁻/CD30⁺ L428 tumor that was transgenic for FFLuc. EBV-CTLs were transferred 7 days after tumor implant. Tumor growth was monitored using the in vivo imaging system. (D) By 7 days after CTL infusion, tumor growth measured as maximum photon/s/cm²/sr (p/s/cm²/sr) was significantly greater in mice receiving control CTLs (top panels) compared with mice receiving CD30CAR⁺ EBV-CTLs (middle panels). Persistence of tumor control can be observed in mice receiving CD30CAR⁺ EBV-CTLs and intraperitoneal injection of irradiated EBV-infected cells, which thus provide the appropriate costimulation (bottom panels). Panel E illustrates the results of 6 mice per group implanted with the CD30⁺ L428 cell line. Bars represent average of light emission (± SD) (P < .05).