We reported that T cells with anti-CD38-chimeric antigen receptors (CAR) eliminated B-cell lymphoma cells expressing CD38. To employ anti-CD38-CAR against acute myeloid leukemia (AML) blasts not expressing CD38, it is necessary to induce or increase the intensity of CD38 expression. A lactate dehydrogenase (LDH)-releasing assay and flow cytometry showed that anti-CD38-CAR T cells were cytotoxic against AML lines (THP-1 and CMK) expressing high CD38 levels (>99%), in time- and number of effector-dependent manners. In other AML lines (KG1, U937 and HL60) partially expressing CD38, CD38+ AML cells were killed by CD38-specific T cells, but CD38− AML cells remained survived. Intriguingly, 10 nM all-trans retinoic acid (ATRA) augmented CD38 expression in KG1, U937 and HL60 cells and primary leukemic cells from AML patients. Moreover, the withdrawal of ATRA from the medium decreased CD38 expression in AML cells. Killing effects of anti-CD38-CAR T cells against AML lines and AML cells were limited without ATRA, whereas CD38-specific T cells enhanced cytotoxicity on AML cells by ATRA in association with enhanced CD38 expression. These results indicate that anti-CD38-CAR T cells eliminate AML cells through CD38 expression induced by ATRA.

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Summary Using artificial receptors, it is possible to redirect the specificity of immune cells to tumour‐associated antigens, which is expected to provide a useful strategy for cancer immunotherapy. Given that B‐cell non‐Hodgkin lymphoma (B‐NHL) cells invariably express CD19 and CD38, these antigens may be suitable molecular candidates for such immunotherapy. We transduced human peripheral T cells or a T‐cell line with either anti‐CD19‐chimeric receptor (CAR) or anti‐CD38‐CAR, which contained an anti‐CD19 or anti‐CD38 antibody‐derived single‐chain variable domain respectively. Retroviral transduction led to anti‐CD19‐CAR or anti‐CD38‐CAR expression in T cells with high efficiency (>60%). The T cell line, Hut78, when transduced with anti‐CD19‐CAR or anti‐CD38‐CAR, exerted strong cytotoxicity against the B‐NHL cell lines, HT and RL, and lymphoma cells isolated from patients. Interestingly, use of both CARs had an additive cytotoxic effect on HT cells in vitro. In conjunction with rituximab, human peripheral T cells expressing either anti‐CD19‐CAR or anti‐CD38‐CAR enhanced cytotoxicity against HT‐luciferase cells in xenografted mice. Moreover, the synergistic tumour‐suppressing activity was persistent in vivo for over 2 months. These results provide a powerful rationale for clinical testing of the combination of rituximab with autologous T cells carrying either CAR on aggressive or relapsed B‐NHLs.

BMI-1 plays a critical role in regulating the activity of hematopoietic stem and progenitor cells. Patients with chronic myeloid leukemia (CML) are at a risk of developing blastic crisis (BC) even after the emergence of imatinib mesylate. In this study, to determine the relevance of BMI-1 to BC, we investigated the expression of BMI-1 in CD34(+) cells at each of the chronic phase (CP), the accelerated phase (AP), and BC by flow cytometry. Interestingly, the level of BMI-1 expression was significantly higher in CP than in controls and was further increased during the course of the disease progression (control--5.66%; CP--36.93%; AP and BC--76.41%). Curiously, mRNA levels for BMI-1 were almost consistent during the disease progression from CP to BC (control--2.21; CP--9.77; AP and BC--9.70 (BMI-1/glyceraldehyde-3-phosphate dehydrogenase ratio)). Since we further found that overexpression of BCR-ABL in human embryonic kidney-293 cells enhanced BMI-1 expression and that BMI-1 expression was increased in K562 cells, derived from patients with BC, in the presence of proteasomal inhibitors, BMI-1 was presumed to be positively regulated by BCR-ABL and further by posttranscriptional modification in the course of the disease progression. We suggest the usefulness of BMI-1 expression in CD34(+) cells as a molecular marker for monitoring patients with CML.

The expression of BMI-1 is correlated with disease progression in cancer patients. We showed that ectopic expression of BMI-1 in B-cell lymphoma cell lines, HT and RL, conferred resistance to etoposide and oxaliplatin, known to enhance sensitivity by targeting the survivin gene, but not to irinotecan, which is not relevant to the downregulation of survivin expression. The expression of survivin was not only augmented in cells transduced with BMI-1, but persisted in the presence of etoposide in cells overexpressing BMI-1. By contrast, the mock-transduced cells succumbed in the medium with anticancer drugs, with an accompanying decrease in BMI-1 and survivin expression. BMI-1 overexpression stabilized survivin post-translationally without an accompanying rise in the mRNA, suggesting survivin as a potential target for BMI-1. Knockdown of either BMI-1 or survivin restored sensitivity to etoposide in the BMI-1-overexpressing lymphoma cells. An analysis of six patients with B-cell lymphoma showed that in the drug-resistant patients, levels of BMI-1 and survivin were maintained even after drug administration. However, downregulation of both BMI-1 and survivin expression was observed in the drug-sensitive patients. Therefore, BMI-1 might facilitate drug resistance in B-cell lymphoma cells through the regulation of survivin. BMI-1 could be an important prognostic marker as well as a future therapeutic target in the treatment of drug-resistant lymphomas. (Cancer Sci 2012; 103: 34-41)