ALBERT

Description: Background: The mixed lineage leukemia (MLL) gene located on chromosome 11 band q23 normally functions as a transcription regulator of the HOX genes and is essential for normal mammalian development and hematopoiesis. Chromosomal translocations involving MLL gene represent frequent cytogenetic abnormalities found in aggressive acute leukemia, both lymphoblastic and myeloid. AL MLL rearrangements have the unique clinical, hematological and prognostic features. We aimed to study the incidence and the types of fusion genes and the clinical relevance. Results: Study samples were from 60 acute leukemia (AL) patients from Sep. 2003 to Dec. 2005. There were 28 males and 32 females. The ages were from 3 months to 54 years old. 17 patients were less than 15 years old and 43 patients were more than 15 years old. All patients were diagnosed based on FAB diagnostic criteria. The patients included 30 with ALL (10 of ALL-L1 patients, 15 of ALL-L2, 5 of ALL-L3), 28 with AML (2 of AML-M1 patients, 5 of AML-M2, 5 of AML-M4, 13 of AML-M5, 1 of AML-M6, 2 of AML-M7), 1 with mixed cell leukemia (AMLL), and 1 with NK cell leukemia. 59 patients were newly diagnosed and 1 patient was refractory. The rearrangements of MLL gene were detected by fluorescence in situ hybridization (FISH) and 6 types of common fusion genes (MLL / AF4, MLL / ENL, MLL / AF9, MLL / ELL MLL / AF6, MLL / AF10) resulting from the rearrangements of MLL gene were detected by nested RT-PCR. There arrangements of MLL gene was found in 7 out of 60 AL patients, (11.67%). Among these 7 patients, 2 were diagnosed with AML- M5 and 5 patients were diagnosed with B-ALL. The fusion genes of the 2 AML-M5 patients who had the rearrangements of MLL gene were MLL/AF9. Among 5 B-ALL patients, 2 patients were confirmed to express MLL/ENL, 1 patient was confirmed to express MLL/AF4, and the other 2 patients did not express the fusion genes. 1 of 2 AML-M5 MLL fusion gene positive patients had invasion of leukemia cell in the left leg and CR was achieved after the first course of chemotherapy. The central nervous system leukemia was got after 1 year and died. The other achieved CR with the third of chemotherapy. 1 of 5 patients died from DIC and the cerebral hemorrhage on the second day after diagnosis B-ALL of MLL fusion gene positive. 2 of 5 patients were not in remission and died from multi-organ failures and infection 3 weeks after diagnosis with positive MLL fusion gene. 1 of 5 patients B-ALL of MLL fusion gene positive got invasion of leukemia cell in thoracic and achieved CR after the second course of chemotherapy. 1 of 5 patients of B-ALL of MLL fusion gene positive achieved CR after the first course of chemotherapy and was relapse 1 year after the bone marrow transplant and then died. Conclusions: We conclude that nested RT-PCR is convenient and feasible method to detect the types of fusion genes resulting from the rearrangements of MLL gene. The clinical features of AL with MLL fusion gene include high white blood cell, invasion of multiple organs, resistant to conventional chemotherapy, easy to relapse after remission, and poor prognosis. The detection of MLL gene rearrangement is of great importance in predicting prognosis and guiding therapy in AL. Disclosures No relevant conflicts of interest to declare.

Description: Detection of BCR-ABL1 mutations that inhibit tyrosine kinase inhibitors is important for the treatment of CML patients. Sanger sequencing (SS) technique is considered the gold standard for mutation detection in a clinical laboratory. We analyzed 40 clinical samples from 22 CML patients with TKI resistance by ultra-deep sequencing (UDS) and compared the results with classical SS-based tests. UDS facilitates the detection of low levels of mutations (

Description: Tyrosine kinase inhibitors have achieved unprecedented efficacy in the treatment of chronic myeloid leukemia (CML), however, imatinib resistance has emerged as a major problem in the clinic. Because the overexpression of BCR-ABL1 critically contributes to CML pathogenesis and drug resistance, targeting the regulation of BCR-ABL1 gene expression may be a novel therapeutic strategy. In this study, we found that the transcriptional repressor MXD1 showed low expression in CML patients and was negatively correlated with BCR-ABL1. Overexpression of MXD1 markedly inhibited the proliferation of K562 cells and sensitized the imatinib-resistant K562/G01 cell line to imatinib, with decreased BCR-ABL1 mRNA and protein. Further investigations using reporter gene analysis showed that MXD1 significantly inhibited the transcriptional activity of the BCR-ABL1 gene promoter. Taken together, these data show that MXD1 functions as a negative regulator of BCR-ABL1 expression and subsequently inhibits proliferation and sensitizes CML cells to imatinib treatment. Disclosures No relevant conflicts of interest to declare.

Description: Objective: To study the feasibility of using cyclophosphamide alone without total body irradiation (TBI) as conditioning regimen for allogeneic hematopoietic stem cell transplantation (allo-HSCT) and establish a graft-versus-host disease (GVHD) mouse model. Methods: Single cell suspension of spleen and bone marrow were prepared from donor C57/BL/6 mice (B6 mice) . The recipient B6D2F1 mice (F1 mice) were signed into 4 groups with 12 mice per each group. Groups I~III mice were conditioned with peritoneal injection of cyclophosphamide (300 mg/m 2/d) on day -5, -4 and -3 of allo-HSCT, and group IV mice were used as control with saline injection. On day 0, group I mice were injected through tail vein with a mixture of 2×107 spleen cells, 5×106 bone marrow cells and 1×104 P815 cells; group II with 5×106 bone marrow cells and 1×104 P815 cells; group III with 1×104 P815 cells and group IV with RPMI1640 culture medium. The blood leukocyte counts were analyzed, GVHD score and the pathological leukemia infiltration of skin, liver and colon were evaluated and compared on day 30 after allo-HSCT. Results: (1) The GVHD scores of group I mice were 2 to 4 on day 14 post allo-HSCT, and mice started to die on day 15, eight (8) mice survived and the GVHD scores of them were 1 to 6 with a survival rate of 66.7% on day 30. Group II mice began to die on day 6 and only one survived on day 30 with a survival rate of 8.3%. All of group III mice died on day 18. Group IV mice survived (Figure 1). (2) The leukocyte counts in group I and group II were significantly increased on day 7 post allo-HSCT(p

Description: BACKGROUND: Mantle cell lymphoma (MCL) is a type of non-Hodgkin's lymphoma(NHL), an aggressive lymphoma that does not respond well to chemotherapy. The patient has chemotherapy-refractory MCL. As the patient immunophenotype is CD19 positive, anti-CD19 Chimeric Antigen Receptor (CAR T 19) Cells are administered in order to treat his chemotherapy-refractory MCL. CASE PRESENTATION: The MCL patient, who is male and 60 years of age, has been treated at Shenzhen Second People's Hospital, The First Affiliated Hospital of Shenzhen University. Patient had been suffering from painless enlarged lymph nodes on neck, armpits and groin for two years when he was hospitalized in April 2015; he also suffered from abdominal pain since March 2015 and shortness of breath for 7 days. Physical examination: the painless enlarged lymph nodes on neck, armpits and groin from multiple bilateral, 1 × 1.5-4.5 × 5cm size. Right groin lymph node biopsy and immunohistochemistry showed NHL, B lymphocyte cell type, MCL. Bone marrow showed 7% of lymphoma cells. Immunophenotype showed positive expressions of CD5, CD19, CD20, CD23. PET/CT scan showed systemic lymph node with multiple nodules and massively increased images, with diffusion and radioactivity in the spleen having also increased. CT showed right pleural effusion a lot, hilar, chunks of retroperitoneal tumor, splenomegaly. Diagnosis was MCL IV of Group B, B lymphocyte cell type with right pleural effusion. We treated the patient with R-CHOP 4 cycles of chemotherapy from May 7, 2015. The result was poor. We infused autologous T cells transduced with a CAR T 19 cells retroviral vector with the chemotherapy-refractory MCL at doses of 6.0×10e8 CAR T 19 cells October, 2015. Patients were monitored for a response, any toxic effects, and the expansion and persistence of circulating CAR T 19 cells. After CAR T 19 cells were infused, the patient experienced chills, fever, shortness of breath, abdominal distension, abdominal pain, chest tightness, and heart palpitations for 9 days. The diagnosis was cytokine release syndrome. After the patient took an anti-IL-6R antibody, he began to recover. Enlarged lymph nodes got shrunken after being infused with CAR T19 cells for 7 days. The peripheral blood CD19 B lymphocytes cell is 0. The right pleural effusion gradually decreased. The lymphadenopathy was cured after 4 months of infusion, with pleural effusion disappearing after 5 months. Q-PCR was used to detect the amount of CAR T 19 cells, which stood at 1.68x10e6 copies/L after 6 months of infusion. After treatment with CAR T 19 cell therapy for 10 months, the patient's state is good. CONCLUSIONS: Cancer immunotherapy as a method of cancer treatment is the most effective after conventional treatments such as radiotherapy, chemotherapy, and surgery. For CD19+ lymphoma, CD19 is a very favorable target, because the expression of CD19 is limited to B cells and not expressed in other tissues or cells. Currently, the efficacy of this treatment in treating MCL remains to be seen. Effects of chemotherapy on the patient's MCL is negligible, due to the fact that his lymph nodes and lymph tissue have become malignant. As a result we chose the CAR T19 cell therapy genetic engineering technique as a method of treatment, to which the patient has responded well. Disclosures No relevant conflicts of interest to declare.

Description: PURPOSE: Arsenic trioxide (ATO) is a very effective drug for acute promyelocytic leukemia (APL). However, little has been reported concerning its use as front-line therapy for complete remission of newly diagnosed APL with t(15;17) chromosomal abnormality. METHODS: The adult patients newly diagnosed with APL with t(15;17) chromosomal abnormality (which would invariably lead to a positive PML/RAR alpha fusion gene) were received 4-5 weeks of ATO administered intravenous drip at a daily dose of 10mg(concentration 2%)/day as induced chemotherapy. When a complete remission was achieved by the ATO induction, a three weeks interval would be observed, after which 5 + 3 days of maintenance (with cytarabine and anthracycline, respectively) would be carried out. A further interval of 2 weeks would then take place, after which all-trans retinoic (ATRA) would be administered oral for another 2 weeks. A final interval of 2 weeks after the administration of ATRA would take place before a recurrence of the chemotherapy regimen, which would restart again with ATO induction. After PML/RAR alpha fusion gene Q-PCR tested negative, the regimen would continue recurring, ensuring negativity of the gene for 2 years. After 2 years the regimen would end, and PML/RAR alpha fusion gene of the patients were to be observed/monitored once every 3-6 months for 10 years. Central nervous system leukemia (CNSL), which might be caused by APL, was to be prevented by lumbar punctures chemotherapy once every 6-8 weeks after complete remission of the patients. Six times were taken. RESULTS: Out of 496 for newly diagnosed APL from 2005 to 2015, we identified 113 patients as newly diagnosed APL. All patients achieved complete response after treatment with ATO induced chemotherapy. Sixty-two out of 65 (95.38%) patients with APL were cured with ATO induction and subsequent 5 + 3 maintenance (cytarabine and anthracycline) therapy from 2005 to 2010. The other three patients were relapsed or had refractory disease and then died. Forty-six out of 48 patients (95.83%) achieved complete cytogenetic remission from 2011 to 2015. The other two patients were relapsed or had refractory disease and then died. All patients had no serious adverse events. Five patients had dyspnea, fever, weight gain, peripheral edema and were successfully treated with dexamethason and furosemide. CONCLUSIONS: Arsenic trioxide is a very effective drug as the front-line therapy for newly diagnosed acute promyelocytic leukemia. Disclosures No relevant conflicts of interest to declare.

Description: CASE PRESENTATION: A 68-year-old male was diagnosed with CLL/SLL in November 2007. Bone marrow asp/bx: 36.5% lymphocytes, 78% CD19, 65% ATM (11q22 deleted) positive cells, 13.5% D13S25 (13q14.3 deleted). On December 10, 2009, the patient took FCR scheme for five cycles, followed by FR scheme for one cycle, and then a month of Chlorambucil. On September 5, 2013, the patient took BR scheme for four cycles with no effect. From March 2015 to Feb 2016, 420 mg of Ibrutinib was administered daily. On January 15, 2016, the patient developed swollen lymph nodes in his right neck with intermittent lumps, fever and nausea. He was admitted into the hospital at Feb 2, 2016. Test results: multiple swollen superficial lymph nodes over the body, with the biggest measuring 60×30mm on the right neck, with no tenderness. Supplementary tests: peripheral white blood cells (WBC) 11.94×10E9/L, lymphocyte 7.5×10E9/L, CD19 cells 6.73×10E9/L, bone marrow lymphocyte 62%, peripheral blood lymphocyte 52%. Immunophenotype: CD5, CD19, CD20dim, CD23, CD11b dim, HLA-DR expression, visible CD5+CD19+ cell clusters, and visible immunoglobulin cKappa with restricted expression. On March 10, 2016, peripheral blood platelet 60 × 10E9/L, CD19 cells 1.94×10E9/L, lactate dehydrogenase 460U/L, FER 115.6ng/ml, hepatitis B virus carrier. Diagnosis: CLL/SLL IV stage, ATM (11q22) deletion, D13S25 (13q14. 3) positive, CD19 positive. Relapse of CLL/SLL occurred again after four months and at this stage the patient was considered for therapy in a clinical trial of CD19-specific chimeric antigen receptor (CAR-) T cell therapy. Ethical approval and informed consent were obtained for anti-CD19 CAR T Cell treatment of ibrutinib resistance in relapsed/refractory CLL/SLL. We infused autologous T cells transduced with a CAR T 19 retroviral vector with CLL/SLL at doses of 3.3 × 10E8 CART19 cells on Mar. 16 2016. Patients were monitored for responses, toxic effects, and the expansion and persistence of circulating CART19 cells. After CART19 cells were infused, the patient experienced chills, fever, headache, weak, anorexia, nausea, shortness of breath, chest tightness, heart palpitation, hypotension and shock for 9 days. The serum levels of IFN-Υ were at their highest at day 7 after CAR T cells infusion. Serum interleukin 6 (IL-6) was at 680pg/ml and CD3+ cells were 97.5%, CD8+ cells 72.8% (18.7-32.8%), FER was 1529.5ng/ml (Normal No. 22-322ng/ml) 14 days after CAR-T cell infusion. The serum levels of IL-6 were at their highest at day14. The patient was diagnosed as having cytokine release syndrome. After the patient took the anti-IL-6R antibody and anti-TNF antibody, he began to recover gradually. Enlarge lymph nodes shrunk after being infused with CART19 cells for 7 days. The peripheral blood CD19 B lymphocytes were 0 on day 14 after infused with CAR T19 cells. Q-PCR was used to detect the amount of the peripheral blood CART19 cells, which stood at 5485 copies/μl, 924 copies/μl, 191 copies/μl respectively 2 weeks, 6 weeks and 3 months after infusing with CART19 cells. The peripheral blood CART 19 cells were not detectable 4 months after infusing with CART19 cells until present. The lymphadenopathy was decreased gradually after 14 days of infusion. The MRI test showed that lymphadenopathy reduced markedly or disappeared after 6 months of infusion. ATM (11q22 deleted) negative, D13S25 (13q14.3 deleted) negative. After treatment with CAR T 19 cell therapy for 53 months, the patient remained disease-free, the patient's lymph nodes, lymphocytes and I mmunoglobulins were normal. CONCLUSIONS : Cancer immunotherapy as a method of cancer treatment is the most effective after conventional treatments such as radiotherapy, chemotherapy, and surgery. For BTK Inhibitor resistance in relapsed and refractory CD19+ CLL/SLL, CD19 is a favorable target, because the expression of CD19 is limited to B cells and not present in other tissues or cells. Currently, the efficacy of this treatment in treating CLL/SLL remains to be seen. The effects of chemotherapy on the patient's B cell lymphoma are negligible, due to the fact that his CLL/SLL have become relapsed and refractory. As a result we chose the CAR T19 cell therapy genetic engineering technique as a method of treatment, to which the patient has responded well. Therefor, CAR T cell technology overcome the limitations of existing cancer therapies and has great potential for development and application. Disclosures No relevant conflicts of interest to declare.

Description: Background Blastic plasmacytoid dendritic cell neoplasm (BPDCN) accounts for 0.44% of hematologic malignancies and is a rare aggressive myeloid neoplasm which shows high rates of cutaneous lesions, bone marrow involvement and lymphadenectasis. Central nervous system (CNS) recurrence can occur when disease progress, preventative intrathecal treatment was essential, whereas optimal treatment of relapsed or refractory BPDCN is still poorly defined. Since a transmembrane glycoprotein, CD38, was defined as a cell receptor which is express not only on lymphoid tissues but also on myeloid cells, Daratumumab (Dara) , the first-in-class human-specific anti-CD38 IgG1 mAb approved for the treatment of MM, had been currently reported to use in treating other hematological malignancies such as Acute Promyelocytic Leukemia (APL), T-acute lymphoblastic leukemia and BPDCN. Case presentation A 66-year-old male patient was admitted to hospital due to fatigue and palpitation, no cutaneous lesion and B symptoms were presented. Physical examination showed painless enlarged lymph nodes on neck, armpits and groin from multiple bilateral. Blood routine: WBC 2.18×109/L, HGB 83g/L, PLT 57×109/L. Bone marrow aspiration showed 79.5% of unclassified cells with irregular nuclei and immature chromatin pattern. Flow cytometry showed 82.26% abnormal cells were positive for CD56,CD123,CD38,CD303,CD304,TCL-1,CD5,CD4. Fluorescence in situ hybridization(FISH) showed P53/CEP17 1.2% positive on amplification, while chromosome showed normal karyotype. Next Generation Sequencing(NGS): TET2/JAK1/KMT2D/PHF6/TP53 mutation. BPDCN of stage Ⅳ/group A was diagnosed. Induction regimen VDCAP was suspended because the patient suffered from infection and grade IV cytopenia on the 3rd day of chemotherapy. Then 7 cycles of CDOP with 5 months of oral BCL-2 inhibitor Venetoclax were applied, autologous stem cell cryopreservation was conducted in the 4th to 5th cycle chemotherapy interval. Bone marrow aspiration after 8 cycles indicated that the patient received bone marrow complete remission, however minimal residual disease test harbored 0.65% abnormal dendritic cells. The patient suffered from headache and sight lost due to disease central nervous system (CNS) recurrence during preparation of autologous stem cell transplantation. The CNS relapse was confirmed by lumbar puncture, the cerebrospinal fluid White blood cell count is 278 x10e6 copies/L and flow cytometry testing for these abnormal white cells was in accord to BPDCN phenotype, meanwhile BM morphology showed 48% of abnormal dendritic cells of typical BPDCN immunophenotype with CD38 expression by flow cytometry either. After treatment with Two cycles of Daratuzumab-VRd regimen and one cycle of HD-MTX(3g/m2) in addition to Temolozomide 140mg/d for 7 days, the disease re-assessment was done, bone marrow aspiration showed 10% residual tumor cells, the amount of WBC in CSF reduced to 14 x10e6 copies/L. Therapeutic response reached Partial Remission (PR) and the patient's state is good. Conclusion BPDCN is rare, therefore no standard treatment was the first-in-class recommendation. Regimens for ALL, AML and NHL had been used for the induction therapy. Although remission rate of ALL regimens are as high as 90%, the duration of response was short, with a median survival of only 12-14 months. Targeted therapy like BCL-2 inhibitor Venetoclax, CD38 monoclonal antibody Daratuzumab and CD123-direct cytotoxin Targraxofusp had been reported to be sensitive to disease. This is the secondly reported case that treated with Daratuzumab. All two cases seemed to be effective. Currently, there is no standardized treatment for BPDCN, novel targeted therapies may improve the prognosis of BPDCN in the near future. This case also indicating that CD38 CART therapy also might be rational for the treatment on myeloid neoplastic. Disclosures No relevant conflicts of interest to declare.