Joyeeta Bhattacharyya

T-cell immunotherapy with a chimeric receptor against CD38 is effective in eliminating myeloma cells.

T-cell immunotherapy with a chimeric receptor against CD38 is effective in eliminating myeloma cells

BMI-1 expression is enhanced through transcriptional and posttranscriptional regulation during the progression of chronic myeloid leukemia.

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.

Synergistic and persistent effect of T-cell immunotherapy with anti-CD19 or anti-CD38 chimeric receptor in conjunction with rituximab on B-cell non-Hodgkin lymphoma

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.

Overexpression of BMI‐1 correlates with drug resistance in B‐cell lymphoma cells through the stabilization of survivin expression

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)

T-cell immunotherapy with a chimeric receptor against CD38 is effective in eradicating chemotherapy-resistant B-cell lymphoma cells overexpressing survivin induced by BMI-1

The expression of BMI-1 (B lymphoma Mo-MLV insertion region 1 homolog), a member of the polycomb-group genes (PcG), is well correlated with a poor prognosis and treatment failure among patients with malignancies such as myelodysplastic syndrome, chronic myeloid leukemia, acute myeloid leukemia and lymphoma.1, 2, 3 Recently, we found that BMI-1 renders B-cell lymphoma cells refractory to several anti-cancer drugs by inducing the expression of survivin.4 There is an urgent clinical need to find therapeutics to treat patients with lymphoma cells overexpressing BMI-1 and survivin. Although in the pre-rituximab era (Rituximab, IDEC Pharmaceuticals, San Diego, CA, USA), the long-term remission rate for patients with diffuse large B-cell lymphoma (DLBCL) was 50–60%, the addition of rituximab has led to an enormous improvement in survival. As rituximab has more significantly improved the overall survival, event-free survival and progression-free survival of patients with non-germinal center B cell-type DLBCL, which have a poor prognosis, than those with germinal center B cell-type DLBCL,5, 6, 7, 8, 9 immunotherapy with an antibody such as rituximab may function by overcoming drug-resistant genes including Bcl-2, Bcl-6 and Bcl-xL.10 We previously developed T cells with a chimeric antigen receptor (CAR) against CD38 and reported that these CD38-specific T cells effectively eliminated B-cell lymphoma cells in vitro and in vivo.11, 12 This is because CD38 is widely and highly expressed in B-cell lymphoma cells (40–50% of patients with the B-cell typed),13 especially in AIDS-associated lymphoma cells14 and DLBCL cells bearing t(14;18) with aggressiveness (100% of these patients).15

All-trans retinoic acid enhances cytotoxic effect of T cells with an anti-CD38 chimeric antigen receptor in acute myeloid leukemia

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.

Elevated interleukin-18 secretion from monoclonal IgM+ B cells in a patient with Schnitzler syndrome

yellow-brown papules on the scalp, and a hemorrhagic papule on the gingival mucosa. Biopsy specimens were obtained from a molluscum-like lesion on her trunk and a new lesion on the scalp. Histopathologic examination showed findings consistent with LCH. Results from a chest radiograph, skeletal survey, and whole-body positron emission tomography scan were normal. One month later, she presented with rapid enlargement of the cutaneous lesions and right ear pain. Computed tomographic imaging of the head, neck, and chest revealed osteolytic lesions in the right and left temporal bones, a non-enhancing soft-tissue mass within the left lobe of the thyroid gland, and multiple cavitary nodules and cystic lesions within the lungs, consistent with multiorgan LCH. Because of the progressive nature of her disease and multiorgan involvement, treatment with vinblastine, prednisone, and 6-mercaptopurine was initiated. LCH often presents with skin manifestations in children. It has been estimated that approximately 80% of patients who develop multisystem disease have cutaneous findings. A review of the literature revealed only one previous case of LCH masquerading as molluscum. Dermatologists should be aware of this potentially underrecognized presentation of LCH as early detection is important to screen for systemic involvement, and potentially limit morbidity and mortality.

Successful treatment of IgM-monoclonal gammopathy of undetermined significance associated with cryoglobulinemia and cold agglutinin disease with immunochemotherapy with rituximab, fludarabine, and cyclophosphamide

Dear Editor, A 69-year-old woman with peripheral coldness as well as pain, numbness of the hands and feet, and purpura on the extremities on exposure to cold stimulation was referred to us. Physical examination found livedo reticularis on the distal extremities (Fig. 1a), Raynaud’s phenomenon, and acrocyanosis. Laboratory investigations revealed elevated levels of IgM (698 mg dl), a markedly increased titer of cold agglutinin (>×10,240), positive for the cyoglobulin test and direct Coombs test (2+(c3d), 1+(IgG)) in serum, immeasurable red blood cell (RBC) count with rouleau formation, and negative for hepatitis C virus antibody and rheumatoid factor. We detected monoclonal IgM-κ protein by immunoelectrophoretic study. Bone marrow study revealed that normal cellularity with a few atypical lymphocytes infiltrated in the morphology under light microscopy. Further analysis by flow cytometry showed a subpopulation of B cells positive for CD19, CD20, CD5, κ and IgM increased in the bone marrow mononuclear cells (Fig. 1b). Furthermore, immunoglobulin heavy chain gene (IgH) rearrangement was detected by polymerase chain reaction (PCR)-based method, suggesting that monoclonal B cells proliferate in the bone marrow (Fig. 1c). Computed tomography did not detect any significant lymphadenopathy or hepatosplenomegaly. In accordance with these results, she was diagnosed with IgM-monoclonal gammopathy of undetermined significance (MGUS) clinically complicated with cryoglobulinemia (type 1) and cold agglutinin disease. Immunochemotherapy, consisting of rituximab (375 mgm, day 1), fludarabine (25 mg m, days 2–4), and cyclophosphamide (250 mg m, days 2–4), termed FCR, was administered every 4 weeks up to three times as a first-line treatment followed by three cycles of monthly rituximab treatment. Extensive skin lesions with livedo reticularis entirely disappeared prior to initiation of the second cycle in association with the declined serum level of IgM (Fig. 1d). There is no standard therapy for IgM-MGUS patients with cryoglobulinemia and cold agglutinin disease, because it is quite rare and IgM-MGUS is generally asymptomatic. Rituximab monotherapy is recommended for patients with symptomatic IgM-MGUS [1]. Although rituximab therapy has been found to be effective in patients with type II mixed cryoglobulinemia, especially associated with hepatitis C virus [2–6], it seems to have limited usefulness for patients with cryoglogulinemia type I or III [7]. Recently, FCR J. Bhattacharyya :K. Mihara (*) :A. Kimura Department of Hematology and Oncology, Research Institute for Radiation Biology and Medicine, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8553, Japan e-mail: kmmihara@hiroshima-u.ac.jp