title: High-affinity FRβ-specific CAR T cells eradicate AML and normal myeloid lineage without HSC toxicity.
abstract: Acute myeloid leukemia (AML) is an aggressive malignancy, and development of new treatments to prolong remissions is warranted. Chimeric antigen receptor (CAR) T-cell therapies appear promising but on-target, off-tumor recognition of antigen in healthy tissues remains a concern. Here we isolated a high-affinity (HA) folate receptor beta (FRβ)-specific single-chain variable fragment (2.48 nm KD) for optimization of FRβ-redirected CAR T-cell therapy for AML. T cells stably expressing the HA-FRβ CAR exhibited greatly enhanced antitumor activity against FRβ(+) AML in vitro and in vivo compared with a low-affinity FRβ CAR (54.3 nm KD). Using the HA-FRβ immunoglobulin G, FRβ expression was detectable in myeloid-lineage hematopoietic cells; however, expression in CD34(+) hematopoietic stem cells (HSCs) was nearly undetectable. Accordingly, HA-FRβ CAR T cells lysed mature CD14(+) monocytes, while HSC colony formation was unaffected. Because of the potential for elimination of mature myeloid lineage, mRNA CAR electroporation for transient CAR expression was evaluated. mRNA-electroporated HA-FRβ CAR T cells retained effective antitumor activity in vitro and in vivo. Together, our results highlight the importance of antibody affinity in target protein detection and CAR development and suggest that transient delivery of potent HA-FRβ CAR T cells is highly effective against AML and reduces the risk for long-term myeloid toxicity.
authors.affiliation: Ovarian Cancer Research Center, Department of Obstetrics and Gynecology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: R C
authors.initials: RC
authors.lastname: Lynn
authors.affiliation: Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, Frederick, MD, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Feng
authors.affiliation: Ovarian Cancer Research Center, Department of Obstetrics and Gynecology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: K
authors.initials: K
authors.lastname: Schutsky
authors.affiliation: Ovarian Cancer Research Center, Department of Obstetrics and Gynecology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: M
authors.initials: M
authors.lastname: Poussin
authors.affiliation: Division of Hematology and Oncology, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: A
authors.initials: A
authors.lastname: Kalota
authors.affiliation: Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, Frederick, MD, USA.
authors.forename: D S
authors.initials: DS
authors.lastname: Dimitrov
authors.affiliation: Ovarian Cancer Research Center, Department of Obstetrics and Gynecology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.|Department of Pathology and Laboratory Medicine, Abramson Cancer Center, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: D J
authors.initials: DJ
authors.lastname: Powell
mesh_terms: D000818:Animals; D002460:Cell Line; D019070:Cell Lineage; D058976:Folate Receptor 2 / Q000276:immunology*; D006412:Hematopoietic Stem Cells; D006801:Humans; D015470:Leukemia, Myeloid, Acute; D051379:Mice; D008822:Mice, Transgenic; D009000:Monocytes; D022423:Myeloid Cells; D011948:Receptors, Antigen, T-Cell / Q000276:immunology*; D057127:Single-Chain Antibodies; D013601:T-Lymphocytes / Q000276:immunology
chemical_list: C000610095:FOLR2 protein, human; D058976:Folate Receptor 2; D011948:Receptors, Antigen, T-Cell; D057127:Single-Chain Antibodies