Rapamycin Pretreatment Rescues the Bone Marrow AML Cell Elimination Capacity of CAR-T Cells.

Abstract

Ongoing clinical trials show limited efficacy for Chimeric antigen receptor (CAR) T treatment for acute myeloid leukemia (AML). The aim of this study was to identify potential causes of the reported limited efficacy from CAR-T therapies against AML. We generated CAR-T cells targeting Epithelial cell adhesion molecule (EpCAM) and evaluated their killing activity against AML cells. We examined the impacts of modulating mTORC1 and mTORC2 signaling in CAR-T cells in terms of CXCR4 levels. We examined the effects of a rapamycin pretreatment of EpCAM CAR-T cells (during <i>ex vivo</i> expansion) and assessed the <i>in vivo</i> antitumor efficacy of rapamycin-pretreated EpCAM CAR-T cells (including <i>CXCR4</i> knockdown cells) and CD33 CAR-T cells in leukemia xenograft mouse models. EpCAM CAR-T exhibited killing activity against AML cells but failed to eliminate AML cells in bone marrow. Subsequent investigations revealed that aberrantly activated mTORC1 signaling in CAR-T cells results in decreased bone marrow infiltration and decreased the levels of the rapamycin target CXCR4. Attenuating mTORC1 activity with the rapamycin pretreatment increased the capacity of CAR-T cells to infiltrate bone marrow and enhanced the extent of bone marrow AML cell elimination in leukemia xenograft mouse models. <i>CXCR4</i> knockdown experiments showed that CXCR4 contributes to the enhanced bone marrow infiltration capacity of EpCAM CAR-T cells and the observed reduction in bone marrow AML cells. Our study reveals a potential cause for the limited efficacy of CAR-T reported from current AML clinical trials and illustrates an easy-to-implement pretreatment strategy, which enhances the anti-AML efficacy of CAR-T cells.<i>See related commentary by Maiti and Daver, p. 5739</i>.