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Background Epigenetic mechanisms play an important role in the chemoresistance of acute myeloid leukemia (AML). The clinical response to epigenetic modifier-based chemotherapy in patients with relapsed/refractory AML (r/r AML) is unclear. This multicenter clinical trial evaluated the safety and efficacy of epigenetic modifiers (chidamide and decitabine) in combination with aclarubicin, cytarabine, and granulocyte colony-stimulating factor (G-CSF) in patients with r/r AML. Results Adult patients with r/r AML were treated with chidamide, decitabine, cytarabine, aclarubicin, and G-CSF (CDCAG). The primary measures were overall response (OR), overall survival (OS), and safety. Next-generation sequencing was performed to analyze the correlation between gene mutations and response. A total of 93 patients with r/r AML were enrolled. Overall, 24 patients had a complete remission (CR) and 19 patients achieved CR with incomplete blood count recovery (CRi). The overall response rate (ORR) was 46.2%. The overall survival of these 43 patients who achieved CR/CRi was significantly longer than that of patients who failed to achieve remission (563 vs 152 days, P < 0.0001). Of the patients with mutations in epigenetic and transcription factor-related genes, but without internal tandem duplications in FMS-like tyrosine kinase3 (FLT3-ITDs), 55.6% achieved CR/CRi, whereas the ORR was 28.2% for patients with mutations in other genes. Conclusions The CDCAG regimen was well tolerated and effective in r/r AML. Patients with epigenetic and transcription factor-related gene mutations, but without FLT3-ITD mutations, may benefit from this regimen. Trial registration Clinical Trials, NCT02886559. Registered 01 September 2016

Unmanipulated haploidentical peripheral blood stem cell transplantation (haplo-PBSCT) has been an established treatment to cure high-risk leukemia/lymphoma. Relapse is the main cause of treatment failure for patients with relapsed/refractory disease or with very high-risk gene mutations such as TP53, TET2, and DNMT3a. In this study, we aimed to establish the tolerance and efficacy of prophylactic donor lymphocyte infusion (DLI) with G-CSF-primed peripheral blood progenitors for prevention of relapse in these very high-risk patients after haplo-PBSCT. The prophylactic DLI was given at a median of 77 days after transplantation in 31 of 45 consecutive patients with very high-risk leukemia/lymphoma. The median dose of CD3+ cells for infusion was 1.8 × 107/kg. The 100-day incidences of acute graft-versus-host disease (GVHD) grades 2–4 and 3–4 after DLI were 55.3% and 10.2%. The 2-year incidences of chronic GVHD and severe chronic GVHD were 52.0% and 18.2%. The 2-year incidences of non-relapse mortality and relapse were 33.1% and 32.5%. The 2-year probabilities of overall survival and relapse-free survival were 40.1% and 31.9%. Poor-risk gene mutations (p = 0.029), disease in non-remission status prior to transplantation (p = 0.005), and donors older than 40 years of age (p = 0.043) were associated with relapse after DLI. In multivariate analysis, disease in non-remission status prior to transplantation was an independent risk factor of relapse (hazard ratio = 4.079; p = 0.035). These data showed the feasibility of the prophylactic DLI in the haplo-PBSCT setting and the anti-leukemic efficacy in very high-risk leukemia/lymphoma. Electronic supplementary material The online version of this article (10.1007/s00277-018-3482-7) contains supplementary material, which is available to authorized users.

Patient: Male, 49 Final Diagnosis: T-lymphoid/myeloid bilineal blastic transformation of CML Symptoms: Rapidly enlarging mass in left neck Medication: — Clinical Procedure: Biopsy of the left submandibular lymph nodes Specialty: Hematology Objective: Rare co-existance of disease or pathology Background: Chronic myeloid leukemia (CML) is a clonal myeloproliferative disorder characterized by the Philadelphia chromosome generated by the reciprocal translocation t(9: 22)(q34;q11). CML is usually diagnosed in the chronic phase. Blast crisis represents an advanced phase of CML. Extramedullary blast crisis as the initial presentation of CML with bone marrow remaining in chronic phase is an unusual event. Further, extramedullary blast crisis with T lymphoid/myeloid bilineal phenotype as an initial presentation for CML is extremely unusual. Case Report: Here, we report the case of a 49-year-old male with rapidly enlarged submandibular lymph nodes. Biopsy specimen from the nodes revealed a characteristic appearance with morphologically and immunohistochemically distinct myeloblasts and T lymphoblasts co-localized in 2 adjacent regions, accompanied by chronic phase of the disease in bone marrow. The presence of the BCR/ABL1 fusion gene within both cellular populations in this case confirmed the extramedullary disease represented a localized T lymphoid/myeloid bilineal blastic transformation of CML. After 3 courses of combined chemotherapy plus tyrosine kinase inhibitor treatment, the mass was completely regressed with a 3-log decrease in BCR/ABL1 transcript from baseline. Five months after the diagnosis, the patient showed diminished vision, hand tremors, and weakness of lower extremities. Flow cytometric immunophenotyping of cerebrospinal fluid revealed the presence of myeloid blasts. An isolated central nervous system relapse of leukemia was identified. Following high-dose systemic and intrathecal chemotherapy, the patient continued to do well. Conclusions: The possibility of extramedullary blast crisis as an initial presentation in patients with CML should be considered. Further, an isolated central nervous system blast crisis should be considered if neurological symptoms evolve in patients who have shown a good response to therapy.

Intermediate-risk acute myeloid leukemia (IR-AML), which accounts for a substantial number of AML cases, is highly heterogeneous. Although several mutations have been identified, the heterogeneity of AML is uncertain because novel mutations have yet to be discovered. Here we applied next generation sequencing (NGS) platform to screen mutational hotspots in 410 genes relevant to hematological malignancy. IR-AML samples (N=95) were sequenced by Illumina Hiseq and mutations in 101 genes were identified. Only seven genes (CEBPA, NPM1, DNMT3A, FLT3-ITD, NRAS, IDH2 and WT1) were mutated in more than 10% of patients. Genetic interaction analysis identified several cooperative and exclusive patterns of overlapping mutations. Mutational analysis indicated some correlation between genotype and phenotype. FLT3-ITD mutations were identified as independent factors of poor prognosis, while CEBPA mutations were independent favorable factors. Co-occurrence of FLT3-ITD, NPM1 and DNMT3A mutations was identified with associated with specific clinical AML features and poor outcomes. Furthermore, by integrating multiple mutations in the survival analysis, 95 IR-AML patients could be stratified into three distinct risk groups allowing reductions in IR-AML by one-third. Our study offers deep insights into the molecular pathogenesis and biology of AML and indicated that the prognosis of IR-AML could be further stratified by different mutation combinations which may direct future treatment intervention.

Liping Dou,1,* Jingxin Li,1,* Dehua Zheng,2,* Yonghui Li,1 Xiaoning Gao,1 Chengwang Xu,1 Li Gao,1 Lili Wang,1 Li Yu1 1Department of Hematology, Chinese PLA General Hospital, Beijing, People's Republic of China; 2Department of Hepatobiliary Surgery, Organ Transplant Center, Chinese PLA 309th Hospital, Beijing, People's Republic of China*These authors contributed equally to this workAbstract: Myeloid/lymphoid or mixed-lineage AF4 acute lymphoblastic leukemia (MLL-AF4 ALL) is a pediatric leukemia that occurs rarely in adults. MLL-AF4 ALL is typically characterized by the presence of chromosomal translocation (t(4;11)(q21;q23)), leading to expression of MLL-AF4 fusion protein. Although MLL-AF4 fusion protein triggers a molecular pathogenesis and hematological presentations that are unique to leukemias, the precise role of this oncogene in leukemogenesis remains unclear. Previous studies have indicated that microRNAs (miRs) might modulate the expression of MLL-AF4 ALL fusion protein, thereby suggesting the involvement of miR in progression or suppression of MLL-AF4 ALL. We have previously demonstrated that miR-205 negatively regulates transcription of an MLL-AF4 luciferase reporter. Here, we report that exogenous expression of miR-205 in MLL-AF4 human cell lines (RS4;11 and MV4-11) inversely regulates the expression of MLL-AF4 at both messenger RNA (mRNA) and protein level. Furthermore, miR-205 significantly induced apoptosis in MLL-AF4 cells as evidenced by Annexin V staining using fluorescence-activated cell sorting (FACS) analysis. The proliferative capacity of leukemic cells was suppressed by miR-205. The addition of an miR-205 inhibitor was able to restore the observed effects. In conclusion, these findings demonstrate that miR-205 may have potential value as a novel therapeutic agent in the treatment of MLL-AF4 ALL.Keywords: miR-205, MLL-AF4, leukemia, microRNA, oncogene expression, untranslated regions, proliferation

As one of the best known cancer testis antigens, PRAME is overexpressed exclusively in germ line tissues such as the testis as well as in a variety of solid and hematological malignant cells including acute myeloid leukemia. Therefore, PRAME has been recognized as a promising target for both active and adoptive anti-leukemia immunotherapy. However, in most patients with PRAME-expressing acute myeloid leukemia, PRAME antigen-specific CD8(+) CTL response are either undetectable or too weak to exert immune surveillance presumably due to the inadequate PRAME antigen expression and PRAME-specific antigen presentation by leukemia cells. In this study, we observed remarkably increased PRAME mRNA expression in human acute myeloid leukemia cell lines and primary acute myeloid leukemia cells after treatment with a novel subtype-selective histone deacetylase inhibitor chidamide in vitro. PRAME expression was further enhanced in acute myeloid leukemia cell lines after combined treatment with chidamide and DNA demethylating agent decitabine. Pre-treatment of an HLA-A0201(+) acute myeloid leukemia cell line THP-1 with chidamide and/or decitabine increased sensitivity to purified CTLs that recognize PRAME(100-108) or PRAME(300-309) peptide presented by HLA-A0201. Chidamide-induced epigenetic upregulation of CD86 also contributed to increased cytotoxicity of PRAME antigen-specific CTLs. Our data thus provide a new line of evidence that epigenetic upregulation of cancer testis antigens by a subtype-selective HDAC inhibitor or in combination with hypomethylating agent increases CTL cytotoxicity and may represent a new opportunity in future design of treatment strategy targeting specifically PRAME-expressing acute myeloid leukemia.

AIM: To evaluate the role of leptin in the internal disorders during hepatic ischemia/reperfusion injury. METHODS: A rat model of 70% hepatic ischemia/reperfusion injury was established, with groups of sham-operation (Sham), 60 min ischemia/60 min reperfusion (I60’R60’), I60’R150’, I60’R240’ and I60’R360’. Serum leptin was detected by a self-produced radioimmunoassay; serum glucose, total anti-oxidation capacity, myeloperoxidase, alanine transaminase and diamine oxidase were determined by relevant kits, while histological alterations and protein levels of leptin in the lung, liver and duodenum were examined by hematoxylin-eosin staining and immunohistochemistry. Spearman’s rank correlation between leptin and other variables or grading of tissue impairment were analyzed simultaneously. RESULTS: Serum leptin in I60’R360’ was significantly higher than in Sham and I60’R240’ groups (both P < 0.05), serum glucose in I60’R360’ was higher than in Sham and I60’R150’ (both P < 0.05), and serum total anti-oxidation capacity in I60’R240’ and I60’R360’ were higher than in Sham (both P < 0.05) and I60’R150’groups (both P < 0.01). Serum myeloperoxidase in groups of I60’R240’ and I60’R360’ were lower than in I60’R150’group (both P < 0.05), serum alanine transaminase in the four reperfusion groups were higher than in the Sham group (all P < 0.05), while serum DAO in I60’R360’ was lower than in I60’R60’ (P < 0.05). Histological impairment in the lung, liver and duodenum at the early phase of this injury was more serious, but the impairment at the later phase was lessened gradually. Protein levels of leptin in the lung in the four reperfusion groups were significantly lower than in the Sham group (all P < 0.01), decreasing in the order of I60’R150’, I60’R60’, I60’R360’ and I60’R240’; the levels in the liver in I60’R60’ and I60’R240’ were higher than in the Sham group (both P < 0.01), while the levels in I60’R240’ and I60’R360’ were lower than in I60’R60’ (both P < 0.01); the levels in duodenum in I60’R240’ and I60’R360’ were higher than in Sham, I60’R60’ and I60’R150’ (all P < 0.01), while the level in I60’R150’ was lower than in I60’R60’ (P < 0.05). There was a significantly positive correlation between serum leptin and alanine transaminase (ρ = 0.344, P = 0.021), a significantly negative correlation between the protein level of leptin in the lung and its damage scores (ρ = -0.313, P = 0.036), and a significantly positive correlation between the protein level of leptin in the liver and its damage scores (ρ = 0.297, P = 0.047). CONCLUSION: Endogenous leptin fluctuates in hepatic ischemia/reperfusion injury, exerts a potency to rehabilitate the internal disorders and represents a potential target for supportive therapy.