Abstract
Ephrin type-B receptor 4 (EPHB4), expressed in tumors including rhabdomyosarcoma, is a suitable target for chimeric antigen receptor (CAR)-T cells. Ligand-independent activation of EPHB4 causes cell proliferation and malignant transformation in rhabdomyosarcoma, whereas ligand-dependent stimulation of EPHB4 induces apoptosis in rhabdomyosarcoma. Therefore, we hypothesized that ligand-based, EPHB4-specific CAR-T cells may kill rhabdomyosarcoma cells without stimulating downstream cell proliferation mechanisms. We developed novel CAR-T cells by targeting EPHB4 via EPHRIN B2, a natural ligand of EPHB4. The generation of EPHB4-CAR-T cells via piggyBac (PB) transposon-based gene transfer resulted in sufficient T cell expansion and CAR positivity (78.5% ± 5.9%). PB-EPHB4-CAR-T cells displayed a dominant stem cell memory fraction (59.4% ± 7.2%) as well as low PD-1 expression (0.60% ± 0.21%) after 14 days of expansion. The PB-EPHB4-CAR-T cells inhibited EPHB4-positive tumor cells without activating cell proliferation downstream of EPHB4, even after multiple tumor re-challenges and suppressed tumor growth in xenograft-bearing mice. Therefore, PB-EPHB4-CAR-T cells possess a memory-rich fraction without early T cell exhaustion and show potential as promising therapeutic agents for treating rhabdomyosarcoma and other EPHB4-positive tumors.
Highlights
There might be various factors underlying the low efficacy of chimeric antigen receptor (CAR)-T cell-based therapy against solid tumors, such as the following: (1) low levels of unique tumor-associated antigens (TAAs) that are exclusively expressed in tumor cells; (2) insufficient expansion of chimeric antigen receptor T (CAR-T) cells in vivo; (3) insufficient tumor penetration by CAR-T cells; (4) escape from immune surveillance owing to reduced tumor antigen expression; and (5) immunosuppression owing to a unique tumor microenvironment.[9,10,11]
Ephrin type-B receptor 4 (EPHB4) expression in human tumor cells To determine whether EPHB4 is a suitable target for EPHB4-CAR-T cells, we investigated the expression of EPHB4 in tumor cells
We modified the extracellular domain of the EPHRIN B2 molecule as an antigen-recognition site in order to develop novel CAR-T cells that re-direct to EPHB4
Summary
The potential of using chimeric antigen receptor T (CAR-T) cells to treat blood cancers, including chemo-refractory or relapsed leukemias, has been demonstrated.[1,2,3,4] efforts to target solid tumors using CAR-T cells have been unsuccessful.[5,6,7,8] There might be various factors underlying the low efficacy of CAR-T cell-based therapy against solid tumors, such as the following: (1) low levels of unique tumor-associated antigens (TAAs) that are exclusively expressed in tumor cells; (2) insufficient expansion of CAR-T cells in vivo; (3) insufficient tumor penetration by CAR-T cells; (4) escape from immune surveillance owing to reduced tumor antigen expression; and (5) immunosuppression owing to a unique tumor microenvironment.[9,10,11] An artificial single-chain fragment variable (scFv) or a specific binding domain, such as a specific ligand of the target molecule, was utilized, and the CAR molecule was variously modified to recognize TAAs.Ephrin type-B receptor 4 (EPHB4), a member of the largest family of receptor tyrosine kinases (RTKs), is highly expressed in various tumors, such as lung cancer,[12] colorectal cancer,[13] breast cancer,[14] esophageal cancer,[15] melanoma,[16] and malignant soft tissue sarcoma, including rhabdomyosarcoma (RMS).[17]. Ligand-dependent activation of EPHB4 induces apoptosis in some tumors.[23] EPHRIN B2-dependent stimulation of EPHB4 activates Crkl downstream of EPHB4 signaling and induces apoptosis, especially in RMS.[24] Based on these observations, we hypothesized that EPHB4 may be utilized as a potent target for the recognition of RMS cells. We developed piggyBac (PB)-mediated CAR-T cells by redirecting EPHB4 (EPHB4-CAR-T cells) and thereby demonstrated the properties and antitumor efficacy of these cells against EPHB4-positive solid tumors in vitro and in vivo
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
