Critical Roles of Chemoresistant Effector and Regulatory T Cells in Antitumor Immunity after Lymphodepleting Chemotherapy.

Antitumor immunity is augmented by cytotoxic lymphodepletion therapies. Adoptively transferred naive and effector T cells proliferate extensively and show enhanced antitumor effects in lymphopenic recipients. Although the impact of lymphodepletion on transferred donor T cells has been well evaluated, its influence on recipient T cells is largely unknown. The current study demonstrates that both regulatory T cells (Tregs) and effector CD8(+) T cells from lymphopenic recipients play critical roles in the development of antitumor immunity after lymphodepletion. Cyclophosphamide (CPA) treatment depleted lymphocytes more efficiently than other cytotoxic agents; however, the percentage of CD4(+)CD25(+) Foxp3(+) Tregs was significantly increased in CPA-treated lymphopenic mice. Depletion of these chemoresistant Tregs following CPA treatment and transfer of naive CD4(+) T cells augmented the antitumor immunity and significantly suppressed tumor progression. Further analyses revealed that recipient CD8(+) T cells were responsible for this augmentation. Using Rag2(-/-) mice or depletion of recipient CD8(+) T cells after CPA treatment abrogated the augmentation of antitumor effects in CPA-treated reconstituted mice. The transfer of donor CD4(+) T cells enhanced the proliferation of CD8(+) T cells and the priming of tumor-specific CD8(+) T cells originating from the lymphopenic recipients. These results highlight the importance of the recipient cells surviving cytotoxic regimens in cancer immunotherapies.

DDX3X induces primary EGFR-TKI resistance based on intratumor heterogeneity in lung cancer cells harboring EGFR-activating mutations.

The specific mechanisms how lung cancer cells harboring epidermal growth factor receptor (EGFR) activating mutations can survive treatment with EGFR-tyrosine kinase inhibitors (TKIs) until they eventually acquire treatment-resistance genetic mutations are unclear. The phenotypic diversity of cancer cells caused by genetic or epigenetic alterations (intratumor heterogeneity) confers treatment failure and may foster tumor evolution through Darwinian selection. Recently, we found DDX3X as the protein that was preferentially expressed in murine melanoma with cancer stem cell (CSC)-like phenotypes by proteome analysis. In this study, we transfected PC9, human lung cancer cells harboring EGFR exon19 deletion, with cDNA encoding DDX3X and found that DDX3X, an ATP-dependent RNA helicase, induced CSC-like phenotypes and the epithelial-mesenchymal transition (EMT) accompanied with loss of sensitivity to EGFR-TKI. DDX3X expression was associated with upregulation of Sox2 and increase of cancer cells exhibiting CSC-like phenotypes, such as anchorage-independent proliferation, strong expression of CD44, and aldehyde dehydrogenase (ALDH). The EMT with switching from E-cadherin to N-cadherin was also facilitated by DDX3X. Either ligand-independent or ligand-induced EGFR phosphorylation was inhibited in lung cancer cells that strongly expressed DDX3X. Lack of EGFR signal addiction resulted in resistance to EGFR-TKI. Moreover, we found a small nonadherent subpopulation that strongly expressed DDX3X accompanied by the same stem cell-like properties and the EMT in parental PC9 cells. The unique subpopulation lacked EGFR signaling and was highly resistant to EGFR-TKI. In conclusion, our data indicate that DDX3X may play a critical role for inducing phenotypic diversity, and that treatment targeting DDX3X may overcome primary resistance to EGFR-TKI resulting from intratumor heterogeneity.

[A case of mediastinal growing teratoma syndrome with acute megakaryoblastic leukemia].

We report a case of a 38-year-old man who was diagnosed with a mediastinal germ cell tumor. After induction chemotherapy, the tumor marker levels normalized, but the tumor itself continued to grow. Surgical resection was performed successfully, but the patient developed acute megakaryoblastic leukemia 6 months later, and induction and consolidation therapies failed to achieve remission. Leukemia cells invaded the central nervous system following hematopoietic stem cell transplantation, and the patient died 5 months after being diagnosed with leukemia. This very rare case of a mediastinal germ cell tumor met the criteria for "growing teratoma syndrome", against a background of acute megakaryoblastic leukemia.

DEAD/H (Asp-Glu-Ala-Asp/His) box polypeptide 3, X-linked is an immunogenic target of cancer stem cells.

Accumulating evidence suggests that most solid malignancies consist of heterogeneous tumor cells and that a relatively small subpopulation, which shares biological features with stem cells, survives through potentially lethal stresses such as chemotherapy and radiation treatment. Since the survival of this subpopulation of cancer stem cells (CSC) plays a critical role in recurrence, it must be eradicated in order to cure cancer. We previously reported that vaccination with CD133(+) murine melanoma cells exhibiting biological CSC features induced CSC-specific effector T cells. These were capable of eradicating CD133(+) tumor cells in vivo, thereby curing the parental tumor. In the current study, we indicated that DEAD/H (Asp-Glu-Ala-Asp/His) box polypeptide 3, X-linked (DDX3X) is an immunogenic protein preferentially expressed in CD133(+) tumor cells. Vaccination with DDX3X primed specific T cells, resulting in protective and therapeutic antitumor immunity. The DDX3X-primed CD4(+) T cells produced CD133(+) tumor-specific IFNγ and IL-17 and mediated potent antitumor therapeutic efficacy. DDX3X is expressed in various human cancer cells, including lung, colon, and breast cancer cells. These results suggest that anti-DDX3X immunotherapy is a promising treatment option in efforts to eradicate CSC in the clinical setting.

Thrombocytopenia with reticulin fibrosis accompanied by fever, anasarca and hepatosplenomegaly : a clinical report of five cases.

We report five cases that presented with high fever, anasarca, hepatosplenomegaly and severe thrombocytopenia with reticulin fibrosis of the bone marrow. The constellation of symptoms is not compatible with any known disease, and we had difficulty in diagnosis and treatment. The age distribution was from 47 to 56 years, and two men and three women were affected. Two patients needed hemodialysis because of renal dysfunction and oliguria with massive pleural effusion. Laboratory examinations showed normal immunoglobulin levels and no monoclonal protein. None of them showed diagnostic autoantibodies for any autoimmune diseases. Histological examination of the liver in three patients and spleen in two showed non-specific findings. Lymphadenopathy was tiny and lymph node biopsy was carried out in only one case. Histologically, paracortical hyperplasia with vascular proliferation and atrophic germinal centers resembling hyaline-vascular-type Castleman's disease or POEMS syndrome were detected. Without a definitive diagnosis, treatment was started with cyclophosphamide, hydroxydaunorubicin, vincristine and prednisolone (CHOP) regimen in one patient, semi-pulse therapy with methyl-predonisolone in three and cyclosporin A in three. Two patients achieved complete remission, two were steroid-dependent and the remaining one died of multiple organ failure. These findings suggest that this disease may be a novel clinical entity belonging to systemic inflammatory disorder with a background of immunological abnormality or a unique variant of multicentric Castleman's disease. [J Clin Exp Hematop 53(1): 63-68, 2013].

Bortezomib and dexamethasone for Japanese patients with relapsed and refractory multiple myeloma: a single center experience.

Bortezomib is a novel proteasome inhibitor, which has shown high antimyeloma activity. APEX trial, phase III randomized study for relapsed or refractory myeloma established efficacy and feasibility of bortezomib. In our study, we retrospectively investigated 60 Japanese patients with relapsed or refractory multiple myeloma (MM) who underwent bortezomib and dexamethasone (BD) therapy in our institution. Overall response rate was 75%, including 7 cases (11.7%) of complete response and 13 cases (21.7%) of very good partial response. Stable disease and progressive disease were observed in 15 patients (25%). Major ≥ grade 3 adverse events were hematological toxicities and grade 3 non-hematological toxicities included appetite loss, diarrhea and peripheral neuropathy. BD therapy was well tolerated, and produced significant response in relapsed or refractory MM patients. Recently, many worldwide trials including bortezomib or other new agents are ongoing to evaluate its efficacy not only as a therapy for relapsed or refractory disease but also as a frontline therapy. Further investigations are required to define how to use new antimyeloma agents for Japanese MM patients.

Donor cell-derived acute lymphocytic leukemia after allogeneic stem cell transplantation for multiple myeloma.

Donor cell leukemia (DCL) is a rare, but well-known, complication after allogeneic hematopoietic cell transplantation. We report a case of donor cell-derived acute lymphocytic leukemia (ALL) occurring in a 55-year-old man after allogeneic bone marrow transplantation (allo-BMT) from an HLA-matched unrelated donor for refractory multiple myeloma (MM). Molecular analysis using short tandem repeat sequences proved the ALL to be of donor origin. He underwent combination chemotherapy and second allo-BMT from an alternative donor. After second allo-BMT, extramedullary myeloma relapsed as tumor, but was successfully treated with proteasome inhibitor, bortezomib. However, he died from severe graft-versus-host disease four months after the second transplantation. Although more than 50 cases of DCL have been reported, there have been only two reports of DCL developed in MM patients including our case. This rare complication may give some insights into leukemogenesis.