Targeting T-cell malignancies using anti-CD4 CAR NK-92 cells.

Peripheral T-cell lymphomas (PTCLs) are a group of very aggressive non-Hodgkin's lymphomas (NHLs) with poor prognoses and account for a majority of T-cell malignancies. Overall, the standard of care for patients with T-cell malignancies is poorly established, and there is an urgent clinical need for a new approach. As demonstrated in B-cell malignancies, chimeric antigen receptor (CAR) immunotherapy provides great hope as a curative treatment regimen. Because PTCLs develop from mature T-cells, these NHLs are commonly CD4+, and CD4 is highly and uniformly expressed. Therefore, CD4 is an ideal target for PTCL CAR immunotherapy. To that effect, we created a robust third-generation anti-CD4 CAR construct (CD4CAR) and introduced it into clonal NK cells (NK-92). CD4CAR NK-92 cells specifically and robustly eliminated diverse CD4+ human T-cell leukemia and lymphoma cell lines (KARPAS-299, CCRF-CEM, and HL60) and patient samples ex vivo. Furthermore, CD4CAR NK-92 cells effectively targeted KARPAS-299 cells in vivo that modeled difficult-to-access lymphoma nodules, significantly prolonging survival. In our study, we present novel targeting of CD4 using CAR-modified NK cells, and demonstrate efficacy. Combined, our data support CD4CAR NK cell immunotherapy as a potential new avenue for the treatment of PTCLs and CD4+ T-cell malignancies.

Novel anti-CD3 chimeric antigen receptor targeting of aggressive T cell malignancies.

Peripheral T-cell lymphomas (PTCLS) comprise a diverse group of difficult to treat, very aggressive non-Hodgkin's lymphomas (NHLS) with poor prognoses and dismal patient outlook. Despite the fact that PTCLs comprise the majority of T-cell malignancies, the standard of care is poorly established. Chimeric antigen receptor (CAR) immunotherapy has shown in B-cell malignancies to be an effective curative option and this extends promise into treating T-cell malignancies. Because PTCLS frequently develop from mature T-cells, CD3 is similarly strongly and uniformly expressed in many PTCL malignancies, with expression specific to the hematological compartment thus making it an attractive target for CAR design. We engineered a robust 3rd generation anti-CD3 CAR construct (CD3CAR) into an NK cell line (NK-92). We found that CD3CAR NK-92 cells specifically and potently lysed diverse CD3+ human PTCL primary samples as well as T-cell leukemia cells lines ex vivo. Furthermore, CD3CAR NK-92 cells effectively controlled and suppressed Jurkat tumor cell growth in vivo and significantly prolonged survival. In this study, we present the CAR directed targeting of a novel target - CD3 using CAR modified NK-92 cells with an emphasis on efficacy, specificity, and potential for new therapeutic approaches that could improve the current standard of care for PTCLs.

Refractory Adult Primary Autoimmune Neutropenia that Responded to Alemtuzumab.

Primary autoimmune neutropenia (P-AIN) is an extremely rare disease. The most effective treatment for primary P-AIN is a granulocyte colony-stimulating factor; however, no curative treatment has been reported. We herein report a case of an adult P-AIN patient with a relatively mild medical history (irrespective of the severe neutropenia) who showed a sustained hematological response over seventeen months after the initiation of treatment with subcutaneous Alemtuzumab.

Suppression of graft-versus-host disease by naturally occurring regulatory T cells.

Studies of graft-versus-host disease after allogeneic bone marrow transplantation have shown that there are subsets of freshly isolated donor T cells that induce the disease and subsets that suppress the disease. The balance of subsets in the graft determines disease severity. The authors' work on the nature of the regulatory-suppressor T cells and their mechanisms of action is summarized in this article.

Host conditioning with total lymphoid irradiation and antithymocyte globulin prevents graft-versus-host disease: the role of CD1-reactive natural killer T cells.

Our previous studies in mice showed that the nonmyeloablative conditioning regimen of fractionated irradiation of the lymphoid tissues (total lymphoid irradiation; TLI) and depletive anti-T-cell antibodies (anti-thymocyte serum) markedly increased the percentage of regulatory DX5+ and natural killer 1.1+ T cells in the mouse spleen, and prevented acute lethal graft-versus-host disease (GVHD) in BALB/c mice (H-2(d)) following the transplantation of bone marrow (BM) and peripheral blood mononuclear cells (PBMC) from C57BL/6 (H-2(b)) donors. The object of the current study was to determine whether the TLI and anti-thymocyte serum regimen protected natural killer T-cell deficient CD1(-/-) BALB/c mice against GVHD after BM and PBMC transplantation from C57BL/6 donors, and whether a similar conditioning regimen of TLI and anti-thymocyte globulin (ATG) can prevent GVHD in Lewis rat (RT1(l)) hosts after BM and PBMC transplantation from ACI rat (RT1(a)) donors. The experimental results in mice showed that, although wild-type BALB/c hosts are protected in association with a marked increase in CD1- reactive T cells expressing the invariant TCR identified with a CD1 tetramer reagent; CD1(-/-) BALB/c hosts are not. Studies of chimeric donor cells in mice protected from GVHD showed donor T-cell polarization to a Th2 cytokine pattern. Results in rats showed that approximately 1000 fold more donor PBMC cells were required to induce a similar incidence of lethal GVHD in TLI and ATG conditioned hosts as compared with hosts conditioned with single-dose total-body irradiation or total-body irradiation and ATG. Surviving TLI and ATG conditioned rat hosts were complete chimeras. In conclusion, the TLI and ATG/anti-thymocyte serum conditioning regimen protects against GVHD in rats and mice, and regulatory natural killer T cells are required for protection.

Unique patterns of surface receptors, cytokine secretion, and immune functions distinguish T cells in the bone marrow from those in the periphery: impact on allogeneic bone marrow transplantation.

The "conventional" NK1.1(-) T cells from mouse blood and marrow were compared with regard to surface receptors, cytokine secretion, and function. Most blood NK1.1(-) CD4(+) and CD8(+) T cells expressed the naive CD44(int/lo)CD62L(hi)CD45RB(hi) T-cell phenotype typical of those in the peripheral lymphoid tissues. In contrast, most marrow NK1.1(-) CD4(+) and CD8(+) T cells expressed an unusual CD44(hi)CD62L(hi)CD45RB(hi) phenotype. The blood NK1.1(-) CD4(+) T cells had a naive T-helper cytokine profile and a potent capacity to induce lethal graft versus host (GVH) disease in a C57BL/6 donor to a BALB/c host bone marrow transplantation model. In contrast, the marrow NK1.1(-) CD4(+) T cells had a Th0 cytokine profile and failed to induce lethal GVH disease, even at 20-fold higher numbers than those from the blood. NK1.1(-) CD8(+) T cells from the blood but not the marrow induced lethal GVH disease. Nevertheless, the marrow NK1.1(-) CD8(+) T cells induced potent antitumor activity that was augmented by marrow NK1.1(-) CD4(+) T cells and facilitated hematopoietic progenitor engraftment. The inability of marrow CD4(+) and CD8(+) T cells to induce GVH disease was associated with their inability to expand in the blood and gut of allogeneic recipients. Because neither the purified marrow CD4(+) or CD8(+) T cells induced GVH disease, their unique features are desirable for inclusion in allogeneic bone marrow or hematopoietic progenitor transplants.