The Efficacy and Safety of Cladribine in Combination with Cytarabine ,ATRA, G-CSF and Mitoxantrone(Claag-M Regimen)in Relapsed/Refractory AML : A Phase 1/2, Prospective, Single-Arm, Multicenter Study

Background. Relapsed/refractory(R/R) AML has poor prognosis and lacks of standard treatment, while clinical trial has been recommended for these patients. It is necessary to find innovation and effective chemotherapy regimens for R/R AML. Cladribine-based regimens have been proved to be effective in R/R AML. As low-dose cytarabine (Ara-C) could reduce marrow myeloblasts and ATRA may sensitize leukemic cells to Ara-C, we performed the phase 1/2, single-arm, multicenter study of cladribine in combination with Ara-C,ATRA,G-CSF and mitoxantrone (CLAAG-M Regimen) in R/R AML (ChiCTR-OIC-16008110). Methods. R/R AML patients aged 18-70 years were eligible to enroll in this study. The objective was to assess the responses and safety of this therapy. All patients received continuous infusion cladribine 5mg/m2 on days 1-5 or 1-3 and mitoxantrone 6mg/m2 on days 1,3,5 or 1,3. ATRA was taken orally at 45mg/m2 on day 4-6 and 15mg/m2 on days 7-20. Subcutaneous injection of cytarabine 15mg/m2 every 12 hours on days 1-10 and G-CSF 300ug/d on day 0 until disease remission or absolute neutrophils count ≥ 0.5×109/L were administered. All patents received one or two courses of induction treatments. Consolidation chemotherapy consisted of cladribine, low-dose Ara-C and G-CSF with or without mitoxatrone or ATRA according to the choice of study site. Results. Between April 2014 and July 2019, a total of 52 patients from five centers were screened,data of 7 patients who did not meet the enrollment criteria and 3 patients did not follow the established treatment were excluded. Therefore, 42 patients were enrolled in this study with 22 females and 20 males (Table 1). Median follow-up duration is 202.0 days(range 12.0-1560.0) from enrollment. The median age was 46 yr (range 25-68 ) . 27 out of 42 patients were refractory AML, with 15 patients in relapsed disease.One patient did not evaluate curative effect owing to personal reason,thus among 41 estimated patients, the overall response rate (ORR= CR+PR) was 70.7%, with complete remission rate (CRR = CR + CRi) of 68.3%, while early death rate and non-remission rate were 7.3% and 22.0% respectively (Table 2). For patients achieved CR, 10 of them (24.4%) received hematopoietic stem cell transplantation (HSCT) and more patients (36.6%) were only treated with consolidation therapy. The median overall survival (OS) of all patients was 306.0 days (95%CI 150.0-462.5) and the rate of 1-year and 2-year OS were 38% and 32% (Figure A). The median disease-free survival(DFS)was 183.0 days (95%CI 16.7-349.2) with half-year survival rate of 37% and 1-year survival rate of 32% (Figure B). Hematological toxicity was the most prominent toxicity of the CLAAG-M regimens since the median duration of neutrophils<0.5×109/L and platelets<20×109/L were 22 days (range8-42) and 17 days (range5-112),respectively. WHO>2 extra-hematological toxicities were infection (21/51) and bleeding (2/51). Three patients (7.3%) died early,with one from infection and two from organ hemorrhage. In multivariate analysis, transplantation would prolong the OS (P=0.013,95%CI of HR 0.010-0.578) . Conclusion.CLAAG-M regimen is effective and tolerated in patients with R/R AML and could be as a bridging therapy regimen followed by HSCT. To confirm the clinical benefit, a randomized study will be further carried out. Disclosures No relevant conflicts of interest to declare.

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Treatment of AML Relapse After Allogeneic Hematopoietic Stem Cell Transplantation (HSCT) Using Low-Dose Azacitidine (AZA).

Blood ◽

10.1182/blood.v116.21.3445.3445 ◽

2010 ◽

Vol 116 (21) ◽

pp. 3445-3445

Author(s):

Alexandre Chiattone ◽

Rima M Saliba ◽

Borje S. Andersson ◽

Sergio Giralt ◽

Manish R Sharma ◽

...

Keyword(s):

Low Dose ◽

Conflicts Of Interest ◽

Disease Free Survival ◽

Control Group ◽

Hematopoietic Stem ◽

Dismal Prognosis ◽

Baseline Characteristics ◽

And Control

Abstract Abstract 3445 Background: Relapsing AML/MDS after HSCT has a dismal prognosis, with few patients achieving long-term control of the malignancy. AZA is a hypomethylating agent that is moderately active against AML/MDS, and may have beneficial immunomodulatory effects after HSCT. We have shown that a significant minority of patients with recurrent disease respond to this drug. Here, we present long-term follow-up after salvage treatment regimens that included AZA, to treat AML/MDS that recurred after HSCT. Patients and Methods: Twenty-three patients received low-dose AZA for recurrence. Decision to use AZA was based on clinical assessment of slow progression of disease and relatively slower disease ‘tempo' and relatively small AML bulk. AZA cohort preparative regimens for 1st HSCT were myeloablative in 12 cases, and of reduced intensity in 11 cases. AZA was used prior to or without a 2nd HSCT (n=17), or after a 2nd HSCT (n=6). Outcomes were compared to controls (n=18) that relapsed ≥ 8 months after HSCT, and did not receive AZA (8 months representing the median disease free survival (DFS) for AZA-treated patients). The control group included all patients that relapsed ≥ 8 months after allogeneic HSCT using myeloablative busulfan 130 mg/m2 and fludarabine 40 mg/m2 for 4 days. AZA was studied as a time dependent variable. AZA and controls had similar baseline characteristics as described in the Table, although median DFS after the first HSCT was 8 (range: 2–51) and 17 (range: 7–59) months, favoring the control group (p=0.08). AZA was administered outpatient, with good tolerance. Fatigue and nausea were commonly observed toxicities. Doses were 8 mg/m2 (n=1), 16 mg/m2 (n=3), 24 mg/m2 (n=10), 32 mg/m2 (n=5), 40 mg/m2 (n=2), and 75 mg/m2 (n=2), administered subcutaneously for 5 days, in 28–32-day cycles. Results: Median number of cycles was 4 (range, 1–44). With a median follow-up of 18 months for AZA and control patients, median survival after relapse was 17 versus 6 months, respectively for AZA and control patients. 11 (48%) AZA patients are alive, while 2 (11%) control patients are alive. Two-year overall survival (OS) for AZA and control groups was 40% and 10%, respectively. AZA and controls had similar baseline characteristics as described in the Table. Conclusion: Low-dose AZA was a well tolerated outpatient treatment that may improve survival after AML/MDS recurrence in selected cases. Major determinants of survival in this setting, however, were remission duration after HSCT, and use of a 2nd HSCT. Disclosures: No relevant conflicts of interest to declare.

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Pre-Emptive Therapy with IFN-α-2b for Acute Leukemia Patients with High Risk of Relapsing Tendency Post Allo-HSCT

Blood ◽

10.1182/blood.v128.22.2250.2250 ◽

2016 ◽

Vol 128 (22) ◽

pp. 2250-2250

Author(s):

XiaoWen Tang ◽

Xiaoji Lin ◽

Aijing Wang ◽

Feng Chen ◽

Xiao Ma ◽

...

Keyword(s):

Survival Rate ◽

High Risk ◽

Acute Leukemia ◽

Median Time ◽

Conflicts Of Interest ◽

Disease Free Survival ◽

Hematopoietic Stem ◽

Free Survival ◽

Donor Chimerism ◽

Significant Difference

Abstract Objective: To determine the efficacy and safety of IFN-α-2b pre-emptive therapy for acute leukemia(AL) patients with relapsing tendencies after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Methods: Retrospectively analyzed 986 acute leukemia patients undergoing allo-HSCT from Jan ,2006 to Mar ,2014 in our hospital. After allo-HSCT, 986 AL patients were periodically monitored the minimal residual disease(MRD) including: bone marrow smear, leukemia-associated immunophenotype (LAIP), leukemia specific or related fusion genes, and donor chimerism through multi-parameter detection to evaluate disease status. Patients were given IFN- a -2b 3 million units / day by subcutaneous injection for preemptive treatment once a relapse tendency was detected, such as: increasing proportion of blast in bone marrowbetween 3-5%, or MRD>1.0×10-3, or leukemia specific fusion gene transfrom negative to positive, or dynamic incressing copy number of WT1 more than 200 copies/104 abl, or decreasing of donor chimerism(≤ 90%). There were 98 patients who presented increasing tendency of MRD and were enrolled in this study. Among them, 31 patients received IFN-α-2b pre-emptive therapy, and 67 patients received non-IFN-α-2b therapy such as: withdraw immunosupressant, traditional DLI or DC-CIK immunotherapy. Results: There were no significant differences in disease characteristics between two groups. For the 31 patients who received IFN-α-2b pre-emptive therapy(IFN group), the median time of IFN-αtreatment was 60 days (range: 5-720 days), Twenty five patients had responsed to the treatment without progressing to hematological relapse (response rate 80.6%). 2 patients developed to hematological relapse again after temporary response; 3 patients had no response and eventually progressed to hematological relapse. Regarding 67patients who received non-IFN-α-2b therapy(non IFN group), 22 patients responsed to the treatment (RR 32.8%), 45 patients failed to the treatment and progressed to hematological relapse at a median time of 35 (range: 6-940) days, There was significant difference of RR between two group(P=0.000) . 31 patients of IFN group tolerate well and no patient terminated therapy due to side effects. During the treatment of IFN, 18 patients(58.1%) developed GVHD: 6 patients (19.4%) with aGVHD and 14 (45.2%) with limited cGVHD . The median follow-up time was 21 (4.5-78.5) months. 22 of 31 cases of IFN group maintained disease-free survival. The 5-year overall survival rate (OS) and the leukemia-free survival rate (LFS) of IFN group were 47.0%±13.9% and 38.7%±13.1% respectively. However, the 5-yr OS and LFS of non IFN group were 14.5%±10.7% and12.5%±9.4% respectively.The difference were significantly (P=0.000,P=0.002 respectively). Patients with GVHD had significantly better response than patients without GVHD (88.9% vs 53.8%, P=0.043, P <0.05). Conclusion: IFN-α-2b pre-emptive therapy can effectively prevent high-risk patients with relapsing tendencies for disease progression post allo-HSCT. Further large-scale investigation is warranted. Disclosures No relevant conflicts of interest to declare.

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Ripk3 Signaling Regulates Hematopoietic Stem Cell Number and Function during Stress

Blood ◽

10.1182/blood-2019-131332 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 3714-3714

Author(s):

Lei Zhang ◽

Huacheng Luo ◽

Jing Li ◽

Hong-Min Ni ◽

Mark Sellin ◽

...

Keyword(s):

Inflammatory Cytokines ◽

Low Dose ◽

Conflicts Of Interest ◽

Lymphoblastic Leukemia ◽

High Dose ◽

Low Doses ◽

Hematopoietic Stem ◽

Increased Risk ◽

Leukemia Development ◽

Serial Transplantation

Background: Among all tissues, bone marrow (BM) is the most sensitive tissue to ionizing radiation (IR)-induced acute tissue damage (ATD) and chronic long-term residual damage (LT-RD). BM failure and a significant reduction in blood cells (pancytopenia) often occurs within days after exposure to IR due to the massive death of proliferative hematopoietic progenitor cells (HPCs). However, due to their quiescent cell cycle status and reduced fidelity of DNA repair feature, many hematopoietic stem cells (HSCs) cannot fully eliminate such damage and enter senescence; this results in LT-RD. Abnormal dysplastic hematopoiesis is the most common LT-RD in most victims of IR, followed by an increased risk of leukemia/lymphoma development. Thus IR exposure is an established cause of BM failure and leukemia. A significant increase in the production of inflammatory cytokines is induced by IR which contributes to the pathogenesis of both ATD and LT-RD. Such inflammatory cytokines induce the activation of Ripk3-Mlkl-mediated necroptotic signaling in HSCs. However, the role of Ripk3-Mlkl signaling in IR-induced damage has not studied. Experimental procedures: The self-renewal capacity of HSCs among Ripk3-/-, Mlkl-/- and WT mice were examined and compared by serial transplantation assay. The phenotypes of ATD and LT-RD induced by different dosages of IR were compared among Ripk3-/-, Mlkl-/- and WT mice. The mechanism by which Ripk3 signaling prevents IR-induced leukemia development was studied. Results: Ripk3-Mlkl signaling is not required for hematopoiesis during homeostatic condition. However, during serial transplantation, inactivation of such signaling prevents stress-induced loss of HSCs. Interestingly, Ripk3 signaling also induces an Mlkl-independent ROS-p38-p16-mediated senescence in HSCs. Thus Ripk3-/- HSCs showed better competitive hematopoietic ability compared to Mlkl-/- and WT HSCs during serial transplantation. A sub-lethal dosage of IR (6Gy) induces Ripk3-dependent NF-κB activation and pro-survival gene expression in HSCs, which is necessary for the survival of damaged HSCs. After 6Gy IR, although DNA damage is repaired in most HSCs within 2 days, a proportion of HSCs in WT and Mlkl-/- mice fail to fully repair the damage and undergo p53-p21-dependent senescence. However such cells in Ripk3-/- mice die from apoptosis. Thus the remaining HSCs in Ripk3-/- mice should be functionally normal, while a proportion of the remaining HSCs in Mlkl-/- and WT mice remain damaged but senescent, all as demonstrated by competitive hematopoietic reconstitution assay. Multiple low-doses of IR (1.75Gy once week × 4) induce HSC exhaustion in WT mice but not in Ripk3-/- and Mlkl-/- mice. Interestingly, almost all Ripk3-/- mice develop acute lymphoblastic leukemia within 200 days after such low dose IR, while 45% of WT and 60% of Mlkl-/- mice develop thymomas within 360 days (see Figure). Mechanistically, such low-dose IR stimulates chronic inflammatory cytokine production. Such cytokines induce Ripk3-Mlkl-mediated necroptosis in response to HSC exhaustion observed in WT mice. These cytokines also induce Ripk3-ROS-p38-p16-mediated senescence in response to impaired HSC functioning observed in both WT and Mlkl-/- mice. In Ripk3-/- mice, due to the lack of both necroptotic and senescent signaling, mutant HSCs accumulate and leukemia development is accelerated. Conclusion: Ripk3 signaling plays distinct roles in HSCs in response to different doses of IR. High-dose IR induces Ripk3-dependent NF-κB/survival signaling, which is required for the survival of HSCs which fail to repair the damage. Thus temporal inhibition of Ripk3-NF-κB signaling might help to remove the damaged HSCs thus preventing the occurrence of LT-RD. However multiple low-doses of IR induces Ripk3 activation in HSCs which represses leukemia development by inducing both ROS-p38-p16-mediated senescence and Ripk3-Mlkl-mediated necroptosis. Induced activation of Mlkl-necroptosis might help to repress leukemia development by removing damaged HSCs. Disclosures No relevant conflicts of interest to declare.

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Randomized Phase II Study of Two Schedules of Flavopiridol (Alvocidib, F) Given as Timed Sequential Therapy (TST) Wtih Ara-C and Mitoxantrone (FLAM) for Adults with Newly Diagnosed, Poor-Risk Acute Myelogenous Leukemia (AML)

Blood ◽

10.1182/blood.v116.21.186.186 ◽

2010 ◽

Vol 116 (21) ◽

pp. 186-186 ◽

Cited By ~ 1

Author(s):

Judith E. Karp ◽

John M. Pagel ◽

B. Douglas Smith ◽

Jacqueline M Greer ◽

D. Michelle Drye ◽

...

Keyword(s):

Phase Ii ◽

Kinase Inhibitor ◽

Conflicts Of Interest ◽

Disease Free Survival ◽

Sequential Therapy ◽

Myelogenous Leukemia ◽

Newly Diagnosed ◽

Hematopoietic Stem ◽

Cyclin Dependent Kinase Inhibitor ◽

Poor Risk

Abstract Abstract 186 Acute Myeloid Leukemia - Therapy, excluding Transplantation: Pediatric and Adult AML Therapy F is a protein bound, cytotoxic, cyclin dependent kinase inhibitor. A prior Phase II trial of TST with FLAM, with F given by one hour bolus daily × 3 for adults with newly-diagnosed AML with poor-risk features demonstrated that the complete remission (CR) rate was 30/45 (67%) with median overall survival (OS) and disease-free survival (DFS) for CR patients being 12.6 and 13.3 months, respectively. We now compare FLAM using bolus F (50 mg/m2 daily × 3; Arm A) vs. FLAM using F given by pharmacologically-derived “hybrid” schedule (30 mg/m2 bolus over 30 min followed by 40 mg/m2 in a 4 hr infusion daily × 3; Arm B) in 70 newly-diagnosed AML patients (pts) with poor-risk features. Results: Pt demographics are presented below. Age # < 50 Secondary AML Adverse Genetics MDS/MPD t-AML Single Complex Flt3 ARM A (n = 36) 59ü(24–78) 3 19 5 6 13 3 Total 24/36 = 67% Total 22/36 = 61% ARM B (n = 34) 58ü(20–73) 5 16 9 8 10 5 Total 25/34 = 74% Total 23/34 = 68% Grade > 3 tumor lysis occurred in 4/70 (6%) with 1 death (A), 1 transient hemodialysis (A), 1 transient hyperkalemia (B), and 1 discontinuation of therapy (B). Four pts (6%) died from regimen toxicity before day 60 (1 A, 3 B). Median time for ANC >500/uL was Day 33 (range 22–71), and platelets > 50,000/uL Day 30 (21-80) for both arms. CR rate in Arm A is 23/36 (64%) including 16/24 (67%) with prior MDS and 13/19 (68%) with adverse cytogenetics and 3/3 (100%) with FLT3-ITD. CR rate in Arm B is 24/34 (71%) including 16/24 (67%) with prior MDS, 12/18 (67%) with adverse cytogenetics, and 4/5 (80%) with FLT3-ITD. As of 7/1/10, 20/23 Arm A CR pts have received chemotherapy and/or allogeneic hematopoietic stem cell transplantation (HCT) in CR: 15/23 (65%) of these pts remain alive and in continuous CR for up to 14+ months, 2 relapsed 4 months post-HCT, and 2 died (1 FLAM consolidation, 1 HCT). Similarly, 20/24 Arm B CR pts have received chemotherapy and/or HCT in CR: 12/20 (60%) remain alive and in continuous CR for up to 18+ months. Of Arm B pts receiving FLAM consolidation, 1 relapsed at 2 months, 1 died at 8 months of cardiac failure, and 2 died during therapy. Three were unable to receive a second cycle and 1 refused. Overall, 51/70 (73%) of all pts and 40/47 (85%) of CR pts are alive 2+ - 19+ months after FLAM. Conclusions: TST with FLAM induces CR in >60% of newly diagnosed, poor-risk AML pts, including those with prior MDS and adverse genetics. There does not appear to be major difference in toxicity or responses between the two F schedules (bolus vs. “hybrid” bolus-infusion). Thus far, allogeneic HCT has been successfully undertaken in 21/47 (45%) of first CR patients, median age 57 (20-64), with 2 early relapses and 1 death from GVHD. Bolus F may be easier to administer than hybrid F and is therefore recommended for further study in newly diagnosed AML pts. These salutary results of FLAM in poor-risk pts will now be evaluated broadly in adults with AML and compared to traditional cytotoxic chemotherapy induction regimens. Disclosures: No relevant conflicts of interest to declare.

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A Brief Use of Imatinib Immediately Before Hematopoietic Stem Cell Transplantation (HSCT) in Children with Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia (Ph+ALL). Results of the Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG) Study Ph+ALL04

Blood ◽

10.1182/blood.v118.21.2554.2554 ◽

2011 ◽

Vol 118 (21) ◽

pp. 2554-2554

Author(s):

Atsushi Manabe ◽

Hirohide Kawasaki ◽

Motoaki Chin ◽

Atsushi Sato ◽

Kimikazu Matsumoto ◽

...

Keyword(s):

Survival Rate ◽

Medical Information ◽

Conflicts Of Interest ◽

Lymphoblastic Leukemia ◽

Cord Blood Transplantation ◽

High Dose ◽

Induction Phase ◽

Study Group ◽

Pediatric Leukemia ◽

Hematopoietic Stem

Abstract Abstract 2554 Aims: Children with Ph+ALL generally have a poor prognosis when treated with chemotherapy alone. The timing and duration of the use of imatinib has not been determined. We investigated a role of imatinib immediately before HSCT. Methods: All the patients with ALL were screened for diagnosis of Ph+ALL using RT-PCR. Children with Ph+ALL were enrolled on JPLSG Ph+ALL04 Study within 1 week of initiation of treatment for ALL. Treatment regimen consisted of 5 therapeutic phases: Induction phase (5-drug induction), Intensification phase (high-dose cytarabine and BFM Ib), Re-induction phase (4-drug re-induction), 2 weeks of Imatinib monotherapy phase (23 weeks after diagnosis), and HSCT phase (Etoposide+CY+TBI conditioning). Before and after each phase, minimal residual disease (MRD), the amount of BCR-ABL transcripts, was measured with the real-time PCR method (cut-off 50 copies/microgram RNA). The study was registered in UMIN-CTR (Medical Information, University hospital Medical Information Network - Clinical Trials Registry): UMIN ID C000000290. Results: During the period 2004–08, 42 patients were registered in the Ph+ALL04 study. Out of 42 patients, 37 patients (88%) achieved CR and 7 of 37 patients also achieved MRD-negative after induction phase. There were 13 patients who had no MRD at the beginning of imatinib monotherapy phase, and 14 patients were MRD-negative after imatinib phase, consequently, 14 patients were MRD-negative at the time of HSCT. Six patients relapsed before HSCT. In total, 31 patients received HSCT in 1st CR. All the patients had engraftment and no patients died because of complications of HSCT. Five patients relapsed after HSCT and 4 of the 5 patients were MRD-negative before HSCT and the other patient had detectable MRD although it was less than 50 copies. Twenty-six patients continue to be in 1st CR and MRD-negative for median of 3 years after diagnosis. The 3-year event-free survival rate and over-all survival rate for all the patients was 57% and 80%, respectively (figure 1). Five patients did not achieve CR after induction phase and they were treated with imatinib-contained chemotherapy. Four of the 5 patients achieved CR. All of the 4 patients received cord blood transplantation and remains in continued CR. Interpretation: The chemotherapy we employed was based on the previous high-risk regimen of TCCSG (Tokyo Children's Cancer Study Group) L-99-15 Study. The chemotherapy was intensive enough to induce MRD-negative in 13 at the time of imatinib phase and 31 of 42 patients were in CR at the time of HSCT (around 25–28 weeks after diagnosis). We planned to assess the efficacy of imatinib immediately before HSCT but it was not possible because of the low amount of MRD in most patients at the beginning of imatinib phase. Conclusion: Although EFS and OS was excellent in this study, 88% of induction rate appeared unsatisfactory and relapse occurred before HSCT in 6 out of 37 patients who achieved CR after induction phase. Earlier and longer use of imatinib may improve EFS in children with Ph+ALL and HSCT may be omitted in a subset of patients who achieve an early and deep remission status. Disclosures: No relevant conflicts of interest to declare.

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Inhibition of HDAC8 Reactivates p53 and Abrogates Leukemia Stem Cell Activity in CBFβ-SMMHC Associated Acute Myeloid Leukemia

Blood ◽

10.1182/blood.v124.21.363.363 ◽

2014 ◽

Vol 124 (21) ◽

pp. 363-363

Author(s):

Jing Qi ◽

Qi Cai ◽

Sandeep Singh ◽

Ling Li ◽

Hongjun Liu ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Bone Marrow Cells ◽

Conflicts Of Interest ◽

Cell Activity ◽

Disease Free Survival ◽

Hematopoietic Stem ◽

The Impact ◽

Acute Myeloid

Abstract The inv(16)-created CBFβ-SMMHC fusion protein inhibits differentiation of hematopoietic stem and progenitor cells (HSPCs) and creates pre-leukemic populations predisposed to acute myeloid leukemia (AML) transformation. However, the molecular mechanism underlying the leukemogenic function of CBFβ-SMMHC has been elusive. Given the low TP53 mutation rate in AML, alternative mechanisms disrupting p53 function are expected. We showed thatCBFβ-SMMHC impairs p53 acetylation and p53 target gene activation through formation of an aberrant protein complex with p53 and HDAC8 (Blood, 120: A772; 122(21): 224). We now show that CBFβ-SMMHC binds to p53 and HDAC8 independently through distinct regions and that HDAC8 mediates the deacetylation of p53 associated with CBFβ-SMMHC. In addition, we generated mice carrying a floxed Hdac8 (Hdac8f) allele and crossed with Cbfb56M/+/Mx1-Cre (Kuo YH et al, Cancer Cell 2006). Deletion of Hdac8 signifiacntly (p<0.0001) reduced the incidence of AML and prolonged disease-free survival. Pharmacologic inhibition of HDAC8 activity with HDAC8-selective inhibitors (HDAC8i) reactivates p53 and selectively induces apoptosis of inv(16)+ AML CD34+ cells while sparing normal HSPCs. To test the effect of HDAC8i on LSC engraftment and leukemia-initiating capacity, we generated Cbfb56M/+/Mx1-Cre mice with a Cre-reporter line expressing tdTomato fluorescence protein following Cre-mediated recombination. AML cells (dTomato+/cKit+) treated with HDAC8i (22d) ex vivo showed reduced engraftment (p=0.025) and enhanced survival (p=0.025) in transplanted mice. To examine whether HDAC8i 22d treatment affects the engraftment capacity on surviving cells, we transplanted equal number (2 x 106) of AML cells treated with either 22d or vehicle in another cohort of mice (n=4). We show that HDAC8i 22d treatment reduced the engraftment of dTomato+/cKit+ AML cells and enhanced survival, suggesting that the engraftment capacity is altered in addition to reducing AML cell survival. We next performed preclinical studies to determine the efficacy of in vivo administration of HDAC8i 22d. AML transplanted mice were randomized into two groups, one group treated with vehicle and the other treated with HDAC8i 22d for 2 weeks. Flow cytometry analysis revealed significantly reduced frequency (p=0.0097) and number (p=0.0101) of dTomato+/cKit+ AML cells in the bone marrow and spleen of 22d treated mice compared to vehicle treated group. To further assess the impact on LSC activity, we transplanted bone marrow cells from these treated mice into secondary recipients and analyzed for AML engraftment. Significant reduction in the frequency (p<0.0001) and the number (p=0.0006) of dTomato+/cKit+ AML cells was observed in the bone marrow and spleen. Furthermore, HDAC8i 22d treated transplants showed no signs of leukemia while vehicle treated transplants are moribund with aggressive AML. These results indicate that HDAC8 inhibition by 22d treatment effectively eliminates engraftment and leukemia-initiating capacity of AML LSCs. In conclusion, our studies identify a novel post-translational p53-inactivating mechanism and demonstrate selective HDAC8 inhibition as a promising approach to target inv(16)+ AML LSCs. Disclosures No relevant conflicts of interest to declare.

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Sorafenib Is Contributing to Salvage Chemotherapy and Is Effective Given Alone for Maintenance in AML FLT3 ITD Patients Relapsing Post Allo Hematopoietic Stem Cell Transplantation

Blood ◽

10.1182/blood.v124.21.5265.5265 ◽

2014 ◽

Vol 124 (21) ◽

pp. 5265-5265 ◽

Cited By ~ 1

Author(s):

Emilia Jaskula ◽

Janusz Lange ◽

Mariola Sedzimirska ◽

Andrzej Lange

Keyword(s):

Maintenance Therapy ◽

Low Dose ◽

Liver Toxicity ◽

Conflicts Of Interest ◽

Normal Karyotype ◽

Salvage Chemotherapy ◽

Remission Induction ◽

Comparative Genomic ◽

High Dose ◽

Hematopoietic Stem

Abstract Normal karyotype AML patients with FLT3 ITD have a higher risk of relapse than those lacking this mutation (Estey EH, Am J Hematol. 2013). Our comparative genomic hybridization study (77 patients) showed that AML cases with as compared to those without FLT3 ITD had significantly lower number of amplifications or deletions (number of total CNV aberrations: 18±4 vs 51±8, p=0.003). The latter observation in concert with that of others (Thiede, Blood 2002) suggest that FLT3 ITD mutation if present in normal karyotype patients is a key player in the pathogenesis of leukaemia and targeting this mutation may be used successfully in FLT3 ITD positive patients relapsing post HSCT. This was also the case in the two patients presented in this study. Patient UPN 952, male, 53 years old, AML M4 (FAB), normal karyotype, received alloHSCT in 2 CR after completion of two lines of remission induction therapies (first: DA3+7 (Daunorubicin, Ara-C), HAM, high dose Ara-C; second: ICE (IDA, Ara-C, VP-16)) and then promptly transplanted. He relapsed 56 days after transplantation. Patient UPN 938, female, 50 years old, AML, normal karyotype, received as an induction DA 3+7, and for consolidation HAM and high dose Ara-C and completed only one course of the maintenance therapy and transplanted in CR. Both cases were transplanted from unrelated donors (10/10 HLA A, B, C, DR and DQ alleles matched), they received myeloablative conditioning (i.v. Busulfan and Cyclophosphamide) and Cyclosporin A as GvHD prophylaxis. At relapse they received salvage chemotherapy tailored to their biological performance (UPN 938 having Age Adjusted Charlson Comorbidity Index 3 received FLAG and UPN 952 with the index 9, ECOG 3, DA2+5). Both cases received in addition Sorafenib (two times 400 mg per day). The response was prompt and the marrow was free from blasts beginning from 11 day post chemotherapy. UPN 952 received as a maintenance only one course of 6-TG with low dose of Ara-C. Due to the substantial comorbidity and liver toxicity WHO3 further chemotherapy treatment was terminated. The patient was left only on Sorafenib (2 times 200 mg per day). UPN 938 was receiving the maintenance therapy (AML protocol) in 6 – 8 weeks intervals based on low dose Ara-C with 6TG or DNR and also Sorafenib (2 times 200 mg per day). In both cases FLT3 signalling pathway (FLT3 Pathway Mutation PCR Array, SABiosciences, Qiagen) revealed a lack of any additional mutation at the check points in FLT3, KRAS, HRAS, NRAS, MEK1, PIK3CA, BRAF and PTEN genes. UPN 938 had in addition to FLT3 ITD mutation c.1807_1808insATGAATATGATCTCAAAT (p.K602_W603insYEYDLK) insertion which relevance was not so far describe. Sorafenib resistance mutations were not found. The patients were on Sorafenib for 8 (UPN 938) and 9 months (UPN 952). The course of UPN938 was uncomplicated. The second case showed mild aGvHD symptoms evolving into cGvHD (skin lesions and dry eye) slowed down with rapamicine. Up to now the patients are in CR and free from FLT3 ITD. The main observation points: - Sorafenib with chemotherapy tailored to the biologic performance of patients contributes to the efficient salvage in patients with relapsing FLT3 ITD positive AML post HSCT and is also effective given alone as a maintenance. - Side effects were seen in one out of two patients likely associated with the blocking of VEGFR signalling transmission (hand foot skin rash, von Willebrandt factor activity – 388%). Conclusion: The use of multikinase inhibitor (Sorafenib) contributes effectively to salvage therapy in FLT3+ AML patients relapsing post alloHSCTSorafenib given alone is able to maintain long-lasting remissionSide effects are individually dependent Supported by NBiR CellsTherpy grant and Byer Health Care Disclosures No relevant conflicts of interest to declare.

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FoxM1 Haploinsufficiency Drives Clonal Hematopoiesis

Blood ◽

10.1182/blood-2019-128429 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 726-726

Author(s):

Chunjie Yu ◽

Yue Sheng ◽

Zhijian Qian

Keyword(s):

Cell Cycle ◽

Progenitor Cells ◽

Low Dose ◽

Conflicts Of Interest ◽

Clonal Expansion ◽

Brdu Incorporation ◽

Quiescent State ◽

Dapi Staining ◽

Hematopoietic Stem ◽

Increased Risk

Hematopoiesis is an orchestrated process in which hematopoietic stem cells (HSCs) can self-renew and produce all lineages of blood cells. Majority of HSCs are in a quiescent state with a low growth rate. However, some genetic mutations that occur in HSCs impel HSCs to exit the quiescent state and to proliferate excessively, which enables mutant HSCs to outcompete normal HSCs and leads to clonal expansion of mutant HSCs. Myelodysplastic syndromes (MDSs) as a clonal disease, arise from the expansion of mutant HSCs and are characterized by morphologic dysplasia, ineffective hematopoiesis and an increased risk of transformation to acute myeloid leukemia. FoxM1 is one of transcription factors in the family of Fox ('Forkhead box') proteins. Analysis of public database revealed that the expression level of FOXM1 was decreased significantly in CD34 + cells from a subset of patients with MDS as compared to healthy individuals. Thus, we sought to determine whether haploinsufficiency of FOXM1 contributes to the development of MDS in mice. Our study showed that haploinsufficiency of Foxm1 led to an expansion of hematopoietic stem/progenitor cells in mice. Since FoxM1 has previously been implicated in regulation of cell cycle, we determined the cell cycle status of Foxm1 heterozygous HSCs. By BrdU incorporation assay, we showed that Foxm1 heterozygous HSCs have an increased S phase and G2/M phase as compared to control HSCs from wildtype mice. Additional analysis with Hochest33342/Pyronin-Y staining and Ki67/DAPI staining showed a significant decrease in the number of quiescent (G0) cells in Foxm1 heterozygous HSCs as compared to control HSCs. These results suggest that FoxM1 haploinsufficiency promotes HSCs to exit quiescence and to enter cell cycle, thereby leading to exhaustion of HSCs. To further assess the function of Foxm1 heterozygous HSCs in vivo, we performed competitive repopulation assay. We found that Foxm1 haploinsufficiency HSCs exhibited competitive repopulation advantage in the first and secondary recipient mice, but displayed significantly decreased capacity of repopulation in tertiary recipient mice as compared to control HSCs, suggesting that Foxm1 haploinsufficiency promoted clonal expansion of HSCs, which leads to an exhaustion of HSCs eventually. HSC proliferation can be induced by some specific immune effectors such as Toll-like receptor 4 (TLR4). Lipopolysaccharide (LPS) stimulates HSC proliferation by activating TLR4 signaling pathway. Low dose of LPS treatment over time accelerated the development of MDS in mice. Interestingly, low dose of LPS injection chronically induced defects in hematopoiesis in Foxm1 haploinsufficient mice but not the control wildtype mice. Recipient mice transplanted with Foxm1 heterozygous BM cells but not the control BM cells developed MDS-like disease with cytopenia and a decreased number of hematopoietic stem/progenitor cells after LPS stimulation. Moreover, we found that nearly half of aged Foxm1 haploinsufficient mice (20 months) developed splenomegaly. Analysis of histologic sections in Foxm1 haploinsufficient mice showed that the mice developed hematopoietic dysplasia including dysplastic megakaryocytes with bizarre-shaped nuclei in bone marrow and extramedullary hematopoiesis with accumulation of myeloid cells in spleen. RNA-seq analysis indicated that haploinsufficiency of Foxm1 perturbed multiple stem cell-maintenance mechanisms especially in metabolic processes. Taken together, our studies suggest that Foxm1 haploinsufficiency in mice causes clonal expansion of HSCs and promotes MDS-like disease, which underscores the significant role of FOXM1 downregulation in the initiation and development of human MDS. Disclosures No relevant conflicts of interest to declare.

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A Retrospective Observation of Treatment Outcomes Using Decitabine-Combined Standard Conditioning Regimens Before Transplantation in Patients With Relapsed or Refractory Acute Myeloid Leukemia

Frontiers in Oncology ◽

10.3389/fonc.2021.702239 ◽

2021 ◽

Vol 11 ◽

Author(s):

Yuhang Li ◽

Longcan Cheng ◽

Chen Xu ◽

Jianlin Chen ◽

Jiangwei Hu ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Low Dose ◽

High Dose ◽

Hematopoietic Stem ◽

Hematopoietic Reconstitution ◽

Conditioning Regimens ◽

Refractory Acute Myeloid Leukemia ◽

Acute Myeloid ◽

Refractory Aml

Hypomethylating agents, decitabine (DAC) and azacitidine, can act as prophylactic and pre-emptive approaches after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and a non-intensive bridging approach before allo-HSCT. However, they are rarely used as a part of conditioning regimens in patients with relapsed or refractory acute myeloid leukemia (AML). This retrospectively study included a total of 65 patients (median, 37; range, 13–63) with relapsed or refractory AML who were treated by allo-HSCT after myeloablative conditioning regimens without or with DAC (high-dose DAC schedule, 75 mg/m2 on day −9 and 50 mg/m2 on day −8; low-dose DAC schedule, 25 mg/m2/day on day −10 to −8). DAC exerted no impact on hematopoietic reconstitution. However, patients who were treated with the high-dose DAC schedule had significantly higher incidence of overall survival (OS, 50.0%) and leukemia-free survival (LFS, 35.0%), and lower incidence of relapse (41.1%) and grade II–IV acute graft versus host disease (aGVHD, 10.0%) at 3 years, when compared with those treated with standard conditioning regimens or with the low-dose DAC schedule. In conclusion, high-dose DAC combined with standard conditioning regimens before allo-HSCT is feasible and efficient and might improve outcomes of patients with relapsed or refractory AML, which provides a potential approach to treat these patients.

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Clinical Analysis of Adult Acute Leukemia with Rearrangements of the 11q23/MLL: Multicenter Co-Operative Study.

Blood ◽

10.1182/blood.v108.11.2354.2354 ◽

2006 ◽

Vol 108 (11) ◽

pp. 2354-2354

Author(s):

Hayato Tamai ◽

Hiroki Yamaguchi ◽

Hiroyuki Hamaguchi ◽

Humiharu Yagasaki ◽

Masami Bassyo ◽

...

Keyword(s):

Remission Rate ◽

Lymphoblastic Leukemia ◽

Treatment Strategies ◽

Disease Free Survival ◽

Clinical Analysis ◽

Adult Patients ◽

Japanese Adult ◽

Acute Leukemias ◽

Hematopoietic Stem ◽

11Q23 Abnormalities

Abstract Background AL(acute leukemias) with 11q23 abnormalities involving the MLL (mixed lineage leukemia) gene comprise one category of recurring genetic abnormalities in the WHO classification. However, the prognosis of adult AL with 11q23/MLL between fusion partners are controversial especially in Asia. Methods We analyzed data on 61 Japanese adult patients diagnosed as AL with 11q23/MLL abnormalities, including the translocations t(4;11), t(9;11), t(6;11), t(11;19) between 1990 and 2006. Results Median age of the 61 patients was 48 years (range, 17–79 years). 54 patients were AML (acute myeloid leukemia) and 7 patients were ALL (acute lymphoblastic leukemia).The incidence by fusion partners in AML were as follows; t(9;11) in 29.6%, t(11;19) in 24.0%, t(6;11) in 18.5%. The incidence of patients with t(11;19) was higher than that in Caucasian and t(4;11) was lower than that in childhood. As for the FAB (French-American-British) subtype, 68.5% of the 11q23/MLL patients were M4/M5. Organ involvement was observed commonly (46.2%) in AML with 11q23/MLL patients except for t(11;19) (7.6%).The CR (complete remission) rate of AML with 11q23/MLL was not low (78.7%), but the CR duration was very short (10 months).The DFS (disease free survival) at 1year and the OS (overall survival) at 2 years were 20% and 30% in AML with t(9;11), 10% and 19% in AML with t(11;19), 10% and 30% in AML with t(6;11). In total, the DFS at 1year and the OS at 2 years were 26% and 25% in AML patients, respectively, while 50% and 50% in ALL patients, respectively. The DFS and the OS at 2 years were 44% and 53% in AML patients with 11q23/MLL who were treated with HSCT(hematopoietic stem cell transplantations), while 14% and 21% in those without HSCT respectively. Conclusions Although there were reports that the prognosis of AML with t(9;11) was not bad in Caucasian, the present study showed poor prognosis of AML with 11q23/MLL in Japanese adult patients regardless of the fusion partners. Our study revealed the short CR duration of the AML patients with 11q23/MLL in the face of the high CR rate. AML patients with 11q23/MLL who were treated with HSCT had superior outcomes to those without HSCT (P=.04 for DFS and P=.04 for OS) .This findings suggest that treatment strategies including HSCT should be considered for AML with 11q23/MLL during the first CR.

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