Author ORCID Identifier

0000-0003-1423-6658

Date of Graduation

5-2024

Document Type

Thesis (MS)

Program Affiliation

Biomedical Sciences

Degree Name

Masters of Science (MS)

Advisor/Committee Chair

Jin S. Im, MD, PhD

Committee Member

Jeffrey J. Molldrem, MD

Committee Member

Leonid S. Metelitsa, MD, PhD

Committee Member

Alexandre Reuben, PhD

Committee Member

Koichi Takahashi, MD, PhD

Committee Member

Gheath Al-Atrash, DO, PhD

Abstract

Chimeric antigen receptor (CAR) T cells have revolutionized the treatment of hematopoietic malignancies achieving >50% complete response rates in numerous refractory/relapsed B cell malignancies. However, there are challenges that hinder CAR-T efficacy and bar the broader use of this therapy in patients. One approach to address these challenges is to create a safe allogeneic CAR cell product by using cells that do not cause graft versus host disease (GvHD). Invariant natural killer T (iNKT) cells are an ideal candidate as they are restricted to the monomorphic CD1d protein in contrast to HLA restricted αβ-T cells and therefore are safe in the allogeneic setting. Furthermore, iNKT cells possess inherent antitumor mechanisms including the expression of NK activating receptors and the capability to lyse M2 tumor associated macrophages via aberrant glycolipid presentation on CD1d. Previous work from our lab demonstrated iNKT cells derived from cord blood units (CB-iNKT) are enriched in naïve and central memory phenotype compared to iNKT cells derived from adult peripheral blood (AB-iNKT). Patients that receive CAR-T cells with higher proportions of naïve/central memory phenotype cells receive greater therapeutic benefit. Therefore, CB-CAR-iNKT cells may serve as an effective platform for allogeneic adoptive cell therapy. Here we evaluate the preclinical therapeutic potential of CB-CAR-iNKT cells by equipping them with 8F4CAR that is restricted to the acute myeloid leukemia (AML) associated antigen PR1 presented in the context of HLA/A2*01. We show CB-8F4CAR-iNKT cells expand in greater numbers compared to AB-8F4CAR-iNKT cells with equivalent 8F4CAR expression and iNKT cell purity. CB-8F4CAR-iNKT cells exhibit significantly higher CD62L expression indicating an enriched naïve and central memory phenotype compared to AB-8F4CAR-iNKT cells. While both CB-8F4CAR-iNKT cells and AB-8F4CAR-iNKT cells display potent AML cytotoxicity via 8F4CAR and iNKTCR in vitro, CB-8F4CAR-iNKT cells trend in an increase in proliferation after repeated leukemia challenge. The increased expression of CD62L and the increased proliferative potential of CB-8F4CAR-iNKT cells suggests an improvement in anti-leukemic activity over AB-8F4CAR-iNKT cells in vivo. We go on to show that CB-8F4CAR-iNKT cells are polarized to Th2 cytokine production after 8F4CAR mediated leukemia cytolysis, which is consistent with iNKTCR mediated stimulation of parental CB-iNKT cells. Importantly, CB-8F4CAR-iNKT cells display anti-leukemic activity in vivo. This thesis supports the idea that CB-iNKT cells can be an effective allogeneic CAR therapy and supports the use of CB-iNKT cells as a platform to target hematopoietic malignancies.

Keywords

natural killer T cell, iNKT, chimeric antigen receptor, CAR-iNKT, cord blood, acute myeloid leukemia

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