Feasibility of Clinical Production of Autologous NKG2D CAR-T

Although there are often differences in the cells harvested from patients by way of venipuncture or apheresis, investigators say that despite this variation in starting materials, they were able to manufacture commercial-grade engineered cells from 100% of the samples from both healthy donors and patients with hematologic malignancies. 

This means that the required parameters of T cell purity and expansion, transduction, and functional activity against NKG2D — as well as release criteria — were all met for the samples prepared for all of the study participants, the authors wrote in an article published in Cytotherapy.¹

The researchers made these natural killer (NK) cell-based chimeric antigen receptors (CAR) by fusing the human gene NKG2D to the cytoplasmic signaling domain of CD3ζ. Thus, the engineered cells gained activity against NKG2D ligands, which, according to researchers, are expressed in more than 70% of human cancers.

When the investigators compared the products they could produce from healthy donors with those they made using malignant donor cells, they observed both products were comprised of mostly CD3⁺T cells. However, they also noticed the product from healthy cells had more CD8+ cells than the product produced from diseased cells, which the researchers could not explain. They suggested the significant difference in CD4/8 ratio between the groups (P= .003) was because “exposure to prior agents such as lenalidomide during treatment for malignancy may affect both apheresis and final product composition from donors with malignancies.”¹

Their method of manufacture allowed them to produce enough modified T cells (dosages from 1 × 10⁶to 3 × 10⁷viable CD3⁺cells) of investigational drugs for first-in-human phase 1 studies featuring a NKG2D CAR-T for the treatment of multiple myeloma (MM) and acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS). Preclinical studies in animal models have shown that this type of construct promotes antitumor responses.

The T-cell purity in the final products was a median of 98%, even in samples where the starting materials had as few as 3% CD3⁺cells. Sterility and stability measures of final products met all requirements for release; the researchers successfully manufactured product from all 38 samples examined across the study, with vector-specific NKG2D CAR surface expression of 66% and 93% of CD8⁺and CD4⁺, respectively.

Within 9 days of apheresis, investigators released and infused the fresh NKG2D CAR-T. They hypothesized that product composition and expansion could be even further improved by employing T-cell selection strategies.

“Methods used to produce CAR T cells, in addition to genetic construct and clinical trial design, are essential determinants of in vivo efficacy,” the authors wrote. “Stepwise efforts to improve scalability, cost, and reproducibility are necessary and may well enable the broad commercialization of these products.”¹

Disclosures: Multiple authors declare affiliations with the pharmaceutical industry. Please refer to the original paper for a complete list of disclosures.

Reference

1. Murad JM, Baumeister SH, Werner L, et al. Manufacturing development and clinical production of NKG2D chimeric antigen receptor–expressing T cells for autologous adoptive cell therapy. Cytotherapy. 2018;20(7):952-963.