Pilot Study of Redirected Haploidentical Natural Killer Cell Infusions for B-Lineage Acute Lymphoblastic Leukemia

1.0. Rationale

In contrast to the well-established cytotoxicity of NK cells against AML cells, their capacity to lyse ALL cells is generally low and difficult to predict. We sought to overcome this intrinsic resistance by transducing CD56+ CD3- NK cells with chimeric receptors directed against CD19, a molecule widely expressed by malignant B cells. Expression of anti-CD19 receptors linked to CD3zeta overcame NK resistance and markedly enhanced NK cell-mediated killing of leukemic cells. This result was significantly improved by adding the 4-1BB costimulatory molecule to the chimeric anti-CD19-CD3zeta receptor: the cytotoxicity produced by NK cells expressing this construct uniformly exceeded that of NK cells whose signaling receptors lacked 4-1BB, even when natural cytotoxicity was apparent (Imai et al., Blood 2005). NK cells expressing anti-CD19 receptors also exerted vigorous anti-ALL activity in a murine model of leukemia (Shimasaki et al., Cytotherapy 2012). Our findings indicate that enforced expression of signaling receptors by NK cells might circumvent inhibitory signals, providing a novel means to enhance the effectiveness of anti-ALL NK cell therapy.

The great anti-leukemic efficacy of genetically modified NK cells shown in our preclinical studies, together with the demonstrated feasibility of infusing durable haploidentical NK cells in a non-HSCT setting and the established expertise by the NUH team in cell therapy (the only center in Asia accredited by Foundation for the Accreditation of Cellular Therapy, FACT), form a compelling rationale for the clinical testing of these NK cells.

The preparation of the key reagent (anti-CD19-BB-zeta mRNA) is finalized in the Tissue Engineering & Cell Therapy (TECT) Laboratory at NUH, where the GMP-compliant MaxCyte electroporator is located. The feasibility of large-scale expansion of NK cells has been demonstrated (Shimasaki et al. Cytotherapy 2012), and the feasibility of large-scale electroporation validated in the TECT laboratory.

We will use flow cytometric and MRD technologies, to determine the presence of persistent disease , and will include in this study only patients with a limited amount of residual disease (<1% leukemic lymphoblasts among normal bone marrow cells). We will use the same MRD methods to monitor the effects of treatment infusions. We do not expect that the conditioning regimen will have much effect by itself on leukemic cell counts, as the patients eligible for the study will have disease that is resistant to many anti-leukemic drugs. Nevertheless, because the conditioning regimen itself may have some salutary effects, it will be important to screen the peripheral blood and/or bone marrow for the presence of leukemic blast cells through all stages of the procedure, i.e., before, during, and after conditioning and after the NK cell infusion. The presence of leukemic cells in will be closely monitored by flow cytometry or PCR amplification of antigen-receptor genes (sensitivity for either method: 1 leukemic cell in 10,000) to shed some light on the relative effect of each intervention.