Extramedullary Relapse of the AML Transformed from MDS Following Auto-HSCT: A Case Report

The treatment for AML (Acute myeloid/myelogenous leukemia) transformed from MDS (myelodysplastic syndrome) is difficult and controversial clinically, especially in elder patients. In this case report, we diagnosed a 59-year-old female patient with AML-M2a transformed form MDS which might be caused by her chemotherapy for mastocarcinoma. After achieving complete remission (CR) through combined chemotherapy, autologous peripheral blood stem cell transplantation (auto-PBSCT) was attempted. Following auto-HSCT, marrow showed continuous CR but the patient later developed extramedullary bone-infiltration relapse. Then local radiotherapy has been applied, and the patient now has prolonged survival. This is the first (or a successful) case report of auto-HSCT in an elderly patient with AML transformed from MDS.     

Cytogenetic analysis and clinical significance of chromosome 7 aberrations in acute leukaemia

The analysis was performed on bone marrow cells derived from 96 patients with acute leukaemia (AL): 76 with acute myelogenous leukaemia (AML) and 20 with acute lymphoblastic leukaemia (ALL). Aberrations of chromosome 7 were revealed in 20 (21%) of 96 analysed cases: in 14 (18%) with AML and in six (30%) with ALL. Structural aberrations, present in 13 patients (eight with AML and five with ALL), were unbalanced and led to partial monosomy (12 cases) or trisomy (four cases) of chromosome 7. Twelve (86%) out of 14 AML and all the ALL patients with chromosome 7 aberrations had complex karyotypes in their bone marrow cells. Monosomy 7 and 7q losses were frequently observed in the AML group, whereas, in the ALL group, gains in 7q and losses in the short arms constituted most chromosome 7 aberrations. The occurrence of monosomy, or of losses in 7q, results in a worse response to induction therapy in AML patients. The complete remission (CR) rate was significantly lower in this group in comparison to the group of AML patients with a normal karyotype (p = 0.01) in bone marrow cells.     

Aggressive chemotherapy in adult primary myelodysplastic syndromes. A report on 29 cases

Twenty-nine adult patients with primary myelodysplastic syndromes (MDS) and an excess of marrow blasts were treated by aggressive chemotherapy while still in MDS phase (20 cases) or after progression to ANLL (9 cases). Median age was 47.5 (range 18-68). Twenty-eight patients received a combination of Rubidazone and Ara C and 1 received High dose Ara C. Fourteen patients (48%) achieved complete remission (CR), 5 (17%) were treatment failures (F) and 10 (35%) died during therapy induced aplasia (DA). Median disease free survival was 8.5 months. Median survival of the whole population was 6 months from the onset of treatment, and 17 months in patients achieving CR. These results were significantly less favorable than those obtained at our institution in de novo ANLL with the same chemotherapy regimens. No statistically significant prognostic factors of treatment outcome emerged but patients with normal cytogenetic findings seemed to have both a higher CR rate and longer remissions than patients with abnormal karyotypes. Patients under 50 did not have higher CR rates than older patients, although they had longer remissions (with 3 out of 6 CRs exceeding 2 years). Finally, treatment outcome and survival were identical in patients treated in the MDS phase and in those treated after progression to ANLL. Combination chemotherapy is a highly toxic approach in MDS and essentially seems to benefit younger patients with a normal karyotype, in whom some long remissions can be obtained.     

Molecular dissection of valproic acid effects in acute myeloid leukemia identifies predictive networks

Histone deacetylase inhibitors (HDACIs) like valproic acid (VPA) display activity in leukemia models and induce tumor-selective cytotoxicity against acute myeloid leukemia (AML) blasts. As there are limited data on HDACIs effects, we aimed to dissect VPA effects in vitro using myeloid cell lines with the idea to integrate findings with in vivo data from AML patients treated with VPA additionally to intensive chemotherapy (n = 12). By gene expression profiling we identified an in vitro VPA response signature enriched for genes/pathways known to be implicated in cell cycle arrest, apoptosis, and DNA repair. Following VPA treatment in vivo, gene expression changes in AML patients showed concordant results with the in vitro VPA response despite concomitant intensive chemotherapy. Comparative miRNA profiling revealed VPA-associated miRNA expression changes likely contributing to a VPA-induced reversion of deregulated gene expression. In addition, we were able to define markers predicting VPA response in vivo such as CXCR4 and LBH. These could be validated in an independent cohort of VPA and intensive chemotherapy treated AML patients (n = 114) in which they were inversely correlated with relapse-free survival. In summary, our data provide new insights into the molecular mechanisms of VPA in myeloid blasts, which might be useful in further advancing HDAC inhibition based treatment approaches in AML.     

Low dose cytosine arabinoside in refractory anemia with excess of blasts in transformation

Myelodysplastic syndromes (MDS) are heterogeneous diseases. Patients with blast counts of more than 20% of nucleated bone marrow cells have a high risk of short survival. We treated six patients with refractory anemia with excess of blast in transformation (RAEBiT) with low dose cytosine arabinoside (LD Ara-C). We had one partial remission (PR), surviving 16 weeks and two complete remissions (CR), surviving 22 and 55+ months. Myelosuppression was dominant in all patients, but was not as serious as with conventional remission-induction treatments for leukemias. Bone marrow aplasia occurred in all responding patients, but a differentiation effect is possible too. Maintenance therapy with LD Ara-C may be important for the two long-lasting CR.     

Molecular mechanism of the schedule-dependent synergistic interaction in EGFR-mutant non-small cell lung cancer cell lines treated with paclitaxel and gefitinib

The regimen of cytarabine, aclarubicin and G-CSF (CAG) has been widely used in China and Japan for treatment of acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). We searched literature on CAG between 1995 and 2010 and performed a meta-analysis to determine its overall efficacy using a random-effects or fixed-effects model. Thirty five trials with a total of 1029 AML (n = 814) and MDS (n = 215) patients were included for analysis. The CR rate of AML (57.9%) was significantly higher than that of MDS (45.7%) (p < 0.01). No difference in CR was noted between the new (56.7%) and relapsed/refractory AML (60.1%) (p > 0.05). The CR rate was also significantly higher in patients with favorable (64.5%) and intermediate (69.6%) karyotypes than those with unfavorable one (29.5%) (p < 0.05). Remarkably, the CR rate of CAG was significantly higher than those of non-CAG regimens (odds ratio 2.43). CAG regimen was well tolerated, with cardiotoxicity in 2.3% and early death in 5.2% of the cases. In conclusion, CAG regimen was an effective and safe regimen for the treatment of AML, and may be more effective than non-CAG regimens. Randomized controlled trials are strongly recommended to evaluate its efficacy and safety in comparison with the current standard treatment.     

Detection of minimal residual disease (MRD) after bone marrow transplantation (BMT) by multi-parameter flow cytometry (MPFC)

Multi-parameter flow cytometry (MPFC) was used to detect minimal residual disease (MRD) following bone marrow transplantation (BMT) in 21 patients. Bone marrow (BM) was analyzed pre-transplant and 3–4 months post-BMT while the patients were in clinical and morphological remission. MRD was detected by identifying cells with aberrant antigen expression and/or leukemia-associated phenotype (LAP) using MPFC. Prior to BMT, 8 out of 21 patients exhibited normal antigen expression based on normal BM samples while 13 BM aspirates had abnormal MPFC. Pre-BMT MPFC was abnormal in all 10 patients who were not in complete remission (CR) (>5% blasts in BM) as well as 3 patients acute lymphoblastic leukemia (ALL) who were in CR. In BM from ALL patients, an abnormal uniform B cell population was observed however antigen expression patterns varied greatly between patients. BM from acute myeloblastic leukemia (AML) patients showed an abnormal distribution of CD34+ cells. In addition, a correlation was observed between pre-BMT cytogenetics and MPFC. Only 2 out of 8 (25%) patients with normal MPFC pre-autologous bone marrow transplantation (ABMT) relapsed (AML), while 6 out of 13 (46%) patients with abnormal pre-BMT MPFC relapsed including 2 out of 3 patients who were transplanted in clinical CR. Pre-BMT MPFC may thus be an effective tool for detection of MRD by detection of a pre-transplant MPFC abnormality.     

Stem cell transplantation for leukemias following myelodysplastic syndromes or secondary to cytotoxic therapy

Two main forms of therapy-related myelodysplastic syndrome and acute myeloid leukemia (t-MDS/AML) have been recognized. The most frequent type, occurring after treatment with alkylating agents, is characterized by abnormalities of chromosomes 5 and/or 7 and t-MDS/AML following treatment with topoisomerase II inhibitors and is associated with molecular aberrations of MLL (11q23) and AML-1 (21q22). Individuals with certain polymorphisms associated with impaired detoxification of cytotoxic agents have an increased risk of developing MDS or AML after treatment of unrelated cancers. Multidrug chemotherapy is less effective for patients with MDS, or AML following MDS, or t-MDS/AML when compared with primary AML, and results in lower complete remission (CR) rates and lower long-term survival. Patients with good risk cytogenetic features, such as t(15; 17), t(8; 21) and inversion 16 are an exception as their treatment outcome is comparable with primary AML patients. Patients who attain a polyclonal and/or a cytogenetic CR may be candidates for autologous stem cell transplantation. For the remaining patients, the only curative option is allogeneic stem cell transplantation with stem cells from a histocompatible sibling or an alternative donor. Reduced intensity conditioning regimens may be considered for patients older than 50 years or patients with comorbidities. The advice is to treat patients early after diagnosis and preferably before progression as these patients have the highest chance of a favorable outcome.     

Stem cell transplantation in pediatric leukemia and myelodysplasia: state of the art and current challenges

The role of stem cell transplantation in the treatment of leukemia and myelodysplasia (MDS) in children has changed over the past decade. In pediatric acute lymphoblastic leukemia (ALL), the overall cure-rate is high with conventional chemotherapy. However, selected patients with a high-risk of relapse are often treated with allogeneic hematopoietic stem cell transplantation (allo-HSCT) in first remission (CR1). Patients with a bone-marrow relapse who attain a second remission frequently receive HSCT. High minimal residual disease (MRD) levels directly prior to HSCT determines the relapse risk. Therefore, MRD positive patients are eligible for more experimental approaches such as intensified or experimental chemotherapy pre-HSCT, as well as immune modulation post-HSCT. In pediatric acute myeloid leukemia (AML) the role of allo-HSCT in CR1 is declining, due to better outcome with modern multi-agent chemotherapy. In relapsed AML patients, allo-HSCT still seems indispensable. Targeted therapy may change the role of HSCT, in particular in chronic myeloid leukemia, where the role of allografting is changing in the imatinib era. In MDS, patients are usually transplanted immediately without prior cytoreduction. New developments in HSCT, such as the role of alternative conditioning regimens, and innovative stem cell sources such as peripheral blood and cord blood, will also be addressed.     

Transplantation of allogeneic CD34+-selected cells followed by early T-cell add-backs: favorable results in acute and chronic myeloid leukemia

BACKGROUND: The aim of this study was to investigate preservation of anti-leukemic activity and protection from opportunistic infections after transplantation of allogeneic + cells in patients with hematologic malignancies and bad prognosis. Methods Thirty-three patients [median age 42 years, range 23-55 years, diagnosis AML/myelodysplastic syndrome (MDS) 14, ALL nine, CML seven and multiple myeloma (MM) three] received myeloablative conditioning followed by infusion of selected CD34+ cells from matched unrelated donors (31) or HLA-identical siblings (two). Early donor lymphocyte infusions (DLI; 0.5 and 1.0 x 10(6) CD3+ cells/kg) were given while patients were on immunosuppressive therapy. RESULTS: Ninety-seven per cent of patients engrafted and 24 of 29 patients surviving more than 30 days received at least one pre-emptive DLI. Three patients (10%) developed acute (a)GvHD (two grade I-II, one grade III-IV) spontaneously, and 16 patients (67%) developed aGvHD after DLI (12 grade I-II, four grade III-IV). Eight of 24 evaluable patients developed chronic (c)GvHD (33%, six limited, two extensive). After a median follow-up of 590 days (range 138-1610 days) 18 patients were alive (55%), 16 in complete remission (CR), one in hematologic and one in molecular relapse. Seven patients died after relapse (21%) and eight died from transplantation-related causes (24%). Patients with myeloid malignancies had a significantly better survival than patients with ALL or MM (74%+/-10 vs. 30%+/-13, P<0.05). DISCUSSION: Early pre-emptive low-dose DLI following transplantation of selected CD34+ cells from unrelated donors after myeloablative conditioning is feasible and effective without undue toxicity, especially in patients with myeloid malignancies.     

流式细胞术动态监测微小残留病灶在急性髓系白血病中的意义*

目的:探讨急性髓系白血病(AML)在治疗过程中应用多参数流式细胞术(MFC)动态微小残留病灶(MRD)的意义。方法:选择2015年1月至2017年2月在我院血液科收治的60例AML患者,在诱导治疗第8天,第21天,每次巩固治疗前1天,结束化疗随访过程中,检测骨髓形态学变化和应用MFC监测患者骨髓MRD,并动态随访。定义未发现异常表型细胞(MRD10~(-4))为阴性,其余为阳性。结果:平均随访时间为11个月(3到16个月),早期MRD(诱导化疗第8天)阴性的患者占58.33%,MRD阳性的患者占41.67%。MRD阴性的患者在诱导治疗接受第21天100%达到CR,MRD阳性的患者80%达到CR。第一次巩固治疗结束后,MRD阴性的患者预后明显较MRD阳性的患者好。结论:动态监测MRD对预测AML患者对治疗的反应和预测复发有重要意义。     

Day-6 bone marrow aspirate for the prediction of response to remission induction therapy for acute myelogenous leukaemia

Seventy-two adults were treated for acute myelogenous leukaemia (AML). Forty-two had previously untreated AML and 30 had AML after a preleukaemic phase, refractory AML or relapsed AML. The previously untreated patients received a 7-day course of cytosine arabinoside (100 or 200 mg/m2 daily), daunorubicin and vincristine while the remaining patients received a 7-day course of cytosine-arabinoside (1 g/m2 q 12h for 6 days) and amsacrine (on day 7). The percentage of malignant cells and the reduction in the percentage of malignant cells were determined by means of bone marrow aspirates taken on day 6 of the chemotherapy course and at the time of diagnosis. Both variables correlated significantly with the ultimate treatment outcome; the reduction in the percentage of malignant cells correlated even more significantly than the absolute percentage malignant cells in the day-6 bone marrow. By means of multiple regression analysis it became possible to calculate the probability of achieving complete remission for the individual patient; this is given by the equation: probability = 1.9-0.009X (% malignant cell reduction). In addition, the mean percentage of malignant cells in the day-6 bone marrow was significantly higher for patients who failed to achieve than those who entered complete remission. Eighty-six per cent of the patients with less than 20% malignant cells on day 6 entered remission, while 75% of the patients with more than 21% malignant cells failed to achieve complete remission (p less than 0.001). Although all of these calculations support the predictive value of the day-6 bone marrow aspirate, the 95% confidence intervals are too large to allow reliable and safe predictions; therefore more patients must be studied to demonstrate the reliability of this test.     

Identification of B-cell translocation gene 1 as a biomarker for monitoring the remission of acute myeloid leukemia

Acute myeloid leukemia (AML) is a biologically heterogeneous disease of the hematopoietic system characterized by a clonal accumulation of immature blast cells in bone marrow. We used a proteomic approach based on two-dimensional electrophoresis and mass spectrometry to search for biomarkers related to the complete remission (CR) state of AML patients. We detected one AML-related protein, which was identified as the B-cell translocation gene 1 (BTG1) protein that belongs to anti-proliferative protein family. In the CR state of AML-M2 and M3 patients (by French-American-British subtype classification), the BTG1 protein was upregulated in bone marrow mononuclear cells. It was also expressed robustly in normal bone marrow mononuclear cells. In addition, the BTG1 levels in AML-M2 patients in a non-remission state after therapy did not increase as they did before therapy. Overexpression of BTG1 mRNA was also observed in the CR state of all-trans-retinoic acid (ATRA)-treated AML-M3 patients and ATRA-treated HL-60 cells. Taken together, these results suggest that BTG1 may play a role in the differentiation process of myeloid cells and can therefore be used as a potential treatment-related biomarker for monitoring the remission status of AML-M2 and M3 patients.     

Valproic acid enhances fludarabine-induced apoptosis mediated by ROS and involving decreased AKT and ATM activation in B-cell-lymphoid neoplastic cells

Histone deacetylase (HDAC) inhibitors have been shown synergize with a number of cytotoxic drugs in leukemic cells. In chronic lymphocytic leukemia (CLL), the first line therapy is based on the combination of fludarabine, a nucleoside analogue, and rituximab, an anti-CD20 monoclonal antibody, and there are presently no HDAC inhibitors are used to manage CLL. In the present study, we found that the addition of valproic acid (VPA), a HDAC inhibitor, increases cell death in B-cell-neoplasm-derived cell lines, BJAB, NALM-6 and I-83. This increased apoptosis caused release of mitochondrial cytochrome c, activation of caspases, and increased reactive oxygen species (ROS). The addition of a ROS scavenger inhibited cell death induced by the VPA–fludarabine combination. In contrast, blocking the death receptor pathway failed to inhibit VPA increased fludarabine induced apoptosis. Combination of VPA and fludarabine treatment decreased both total and phosphorylated levels of AKT, an important anti-apoptotic protein, and ATM, a pivotal protein in DNA damage response. Chemical inhibition of AKT or ATM was sufficient to enhance fludarabine-induced apoptosis. We next examined patient samples from a local clinical trial where relapsed CLL patients were treated with VPA and examined the effects of VPA on AKT and ATM in vivo. After 30 days, there was a reduction in ATM levels in three out of the four patients treated, while AKT phosphorylation was reduced only in one patient. Taken together, VPA reduces ATM levels, thereby increasing ROS-dependent cell death via the mitochondrial apoptotic pathway when combined with fludarabine.     

Using peripheral blood circulating DNAs to detect CpG global methylation status and genetic mutations in patients with myelodysplastic syndrome

Myelodysplastic syndrome (MDS) is a hematopoietic stem cell disorder. Several genetic/epigenetic abnormalities are deeply associated with the pathogenesis of MDS. Although bone marrow (BM) aspiration is a common strategy to obtain MDS cells for evaluating their genetic/epigenetic abnormalities, BM aspiration is difficult to perform repeatedly to obtain serial samples because of pain and safety concerns. Here, we report that circulating cell-free DNAs from plasma and serum of patients with MDS can be used to detect genetic/epigenetic abnormalities. The plasma DNA concentration was found to be relatively high in patients with higher blast cell counts in BM, and accumulation of DNA fragments from mono-/di-nucleosomes was confirmed. Using serial peripheral blood (PB) samples from patients treated with hypomethylating agents, global methylation analysis using bisulfite pyrosequencing was performed at the specific CpG sites of the LINE-1 promoter. The results confirmed a decrease of the methylation percentage after treatment with azacitidine (days 3-9) using DNAs from plasma, serum, and PB mono-nuclear cells (PBMNC). Plasma DNA tends to show more rapid change at days 3 and 6 compared with serum DNA and PBMNC. Furthermore, the TET2 gene mutation in DNAs from plasma, serum, and BM cells was quantitated by pyrosequencing analysis. The existence ratio of mutated genes in plasma and serum DNA showed almost equivalent level with that in the CD34+/38- stem cell population in BM. These data suggest that genetic/epigenetic analyses using PB circulating DNA can be a safer and painless alternative to using BM cells.     

Phase II cooperative study with a new alkylating drug, PTT-119, in the treatment of non-Hodgkin's lymphomas

In a phase II cooperative study involving eleven Italian haematological units, the efficacy and toxicity of a new alkylating compound, PTT-119, was evaluated in 53 patients with non-Hodgkin's lymphoma (NHL). Forty-five of the patients had been previously treated with various regimens of chemotherapy, the remaining eight were at the onset of the disease. PTT-119 was scheduled at 3.0 mg/kg every three weeks for a minimum of three administrations. Seven patients achieved a complete remission (CR), 19 a partial remission (PR); the overall response rate was 49%. The median duration of response was 6 months. Most frequent adverse effects were alopecia, nausea and vomiting and phlebitis due to the drug infusion. Myelosuppression was severe only in patients with bone marrow involvement or who were heavily pretreated. No liver, cardiac or renal toxicity was recorded. These data indicate that PTT-119 is an effective drug in the treatment of NHL; the matter of its non-cross-resistance with other alkylating compounds warrants further studies.