ALBERT

Description: Abstract 1878 Introduction: The unbalanced translocation of der(1;7)(q10;p10) is relatively rare in myeloid neoplasms, which was reported at frequencies of 2% in de novo myelodysplastic syndrome (MDS), 0.5% in de novo acute myeloid leukemia (AML) and 3–7% in therapy-related myeloid neoplasms. Although chromosome 7 abnormality is generally considered as a poor prognostic factor in MDS and AML, it is still unclear whether or not this unbalanced translocation has a negative impact on prognosis. In addition, there appears to be some unique clinicopathological features such as male predominance, low blast counts, high hemoglobin level, presence of eosinophilia and a high rate of sole chromosomal abnormality. Methods: We retrospectively analyzed 122 der(1;7)(q10;p10) and -7/del(7q) adult patients with MDS or AML who were diagnosed at our institute between February 1995 and March 2010. According to the French-American-British (FAB) classification, 29 patients had AML (M0, n=5; M1, n=6; M2, n=10; M4, n=3; M5, n=1; M6, n=4) and 93 patients had MDS (RA, n=50; RARS, n=2; RAEB, n=35; RAEB-t, n=3; CMML, n=3). We compared the clinicopathological features and outcome of 33 der(1;7) patients with those of 89 -7/del(7q) patients. Results: The median age was 71 years for the der(1;7) patients and 67 years for -7/del(7q) patients (p=0.29). Male predominance was observed in both patients in the der(1;7) and the -7/del(7q) (p=0.52). The proportion of the sole abnormality in the der(1;7) was significantly higher than that in the -7/del(7q) (58% vs 13%, p

Description: Background: Oral tacrolimus (Tac) was first developed as a twice-daily formulation (Tac BID) and has been widely used in solid organ and allogeneic hematopoietic stem cell transplantation (allo-SCT). Lifelong immunosuppression is required to preserve graft function after solid organ transplantation, and medical nonadherence of transplant recipients has been identified as a major cause of allograft failure. It has been well documented that frequent drug dosing affects patients' adherence. In response to this potential nonadherence problem, a once-daily modified release formulation of oral tacrolimus (Tac QD) has been developed. Results of randomized prospective phase III studies have indicated that Tac QD is well tolerated with similar efficacy and safety profiles as those of Tac BID in recipients after organ transplantation. Although Tac BID has been widely used in allo-SCT recipients, there are no available data on the use of Tac QD in this population. In this study, we compared retrospectively the efficacy of Tac QD versus Tac BID administration. The objective of this study was to investigate whether Tac QD is as effective as Tac BID in the setting of allo-SCT from unrelated donors. Methods: The study cohort included 77 consecutive patients who received allogeneic bone marrow transplantation from unrelated donors for treatment of hematologic malignancies between 2000 and 2014: AML (n=35), ALL (n=15), CML (n=8), MDS (n=12), NHL (n=5), and MPN (n=2). Patients received Tac iv from day -1 with short-term methotrexate (MTX) on days 1, 3, 6, and 11; Tac iv was converted to either Tac QD (n=37) or Tac BID (n=40) when the patients were engrafted and could tolerate oral medication. Doses were modified to maintain a whole-blood trough concentration of 8-12 ng/ml. Tac BID was administrated until Oct 2008, and Tac QD was used from Nov 2008. Kaplan-Meier estimates were used for overall survival (OS), and cumulative incidence estimates were used for acute GVHD, chronic GVHD, and non-relapse mortality (NRM). Results: Median age was 53 years (range: 23-66) for Tac QD cohorts and 39 years (range: 17-56) for Tac BID cohorts. Tac QD patients were older (P

Description: A high frequency of mtDNA somatic mutation has been observed in many tumors as well as in aging tissues. In this study, we analyzed the mtDNA control region sequence variation in 3534 single normal cells and individual blasts from 18 patients with leukemia and 10 healthy donors, to address the mutation process in leukemic cells. We found significant differences in mtDNA sequence, as represented by the number of haplotypes and the mean number of cells with each nonaggregate haplotype in a population of cells, in patients compared to controls. Patients with similar clinical leukemia types, particularly acute myeloid leukemia (AML), did not show a uniform pattern of sequence variation in single blasts. Some patients at relapse presented a complex shift of major haplotypes in single cells. Four patients showed high frequencies of cells containing mutations 189, 260, 16150, and 16488, respectively, as a result of clonal expansion and could be considered as potential markers for their respective disease progression. To our knowledge, this is the first large-scale study of mtDNA variation in single malignant cells. Our results suggest that the somatic mutation process in leukemia is complex, leading to diverse levels of genetic alterations due to either intrinsic aspects of leukemia pathophysiology or chemotherapy effects.

Description: Abstract 3048 Background: In solid organ transplantation, lifelong immunosuppression is required to preserve graft function, and medication nonadherence is a major risk factor for graft failure. Tacrolimus (Tac) was first developed as an oral twice-daily formulation (Tac BID) and has been widely used in hematopoietic stem cell and solid transplantation, but long-term adherence remains a concern. In renal transplant patients, morning dosing is associated with significantly higher adherence than evening dosing. In response to this potential adherence problem, a once-daily modified release formulation (Tac QD) has been developed with a morning dosing regimen that maximizes the potential for adherence. However, several investigators have recently reported a sustained decrease in Tac exposure in kidney transplant recipients after conversion from Tac BID to Tac QD. In this study, we measured Tac exposure after switching from Tac intravenous infusion (Tac iv) to Tac QD and analyzed the pharmacokinetics (PK) of Tac QD to investigate the correlation between area-under-curve (AUC) and trough in patients who received allogeneic hematopoietic stem cell transplantation (HSCT). This is the first report of the PK study of Tac QD in patients who received allogeneic HSCT. Methods: Patients who were 15–65 years of age and received HSCT from unrelated donors were eligible. They received Tac iv 0.03mg/kg by continuous infusion beginning one day before transplantation, and administration was converted to Tac QD at a 1:4 ratio when the patients engrafted and could tolerate oral medication. Doses were modified to maintain whole-blood trough concentration of 8–12 ng/ml. To analyze PK of Tac QD, plasma samples were obtained at baseline and at 0, 1, 2, 3, 6, 12, and 24 hours after the once-daily administration. Plasma concentration of Tac was determined by an ELISA method. Results: A total of 10 patients with hematological malignancies (AML 6, ALL 1, MDS 2, NHL 1) were enrolled in the PK study. Median age was 45 (23–65) years. Five patients received myeloablative preparative regimens, and 5 patients received reduced intensity regimens, and stem cell sources were bone marrow (BM) from HLA-matched unrelated donor (n=4), BM from HLA DRB1 mismatched unrelated donor (n=4), or cord blood (n=2). After conversion from Tac iv to Tac QD, six out of 10 patients (60%) showed a sustained decrease in Tac exposure and required an increase in their Tac QD daily dose. One patient experienced a quick decrease of more than 78% in trough level of Tac QD, and he developed grade II acute GVHD after the conversion. No other patients developed grade II-IV acute GVHD. None of the patients had to discontinue the agent because of adverse effects, and none developed treatment-related mortality within 100 days after HSCT. In PK analysis, median area-under-curve (AUC) was 246 ng·h/ml. There was a strong correlation between AUC and trough level (Figure 1). Obtaining over 240 ng•h/ml of AUC required 7.5 ng/ml of whole-blood trough levels of tacrolimus. Conclusions: Despite initial reports showing the bioequivalence of Tac QD with Tac BID, we found that 60% of patients experienced a sustained decrease in Tac exposure. Therefore, the conversion from Tac iv to Tac QD should be performed under close medical supervision. Our study demonstrated that AUC and trough concentration level curves showed a strong correlation in patients who underwent allogeneic HSCT. The whole-blood trough should be maintained above 7.5ng/ml to provide an adequate level of AUC. If the trough level of Tac QD can be maintained above 7.5 ng/ml, Tac QD can be as effective as Tac iv and stable administration can be maintained for the patients with reduced stress because of the easier administration schedule. Our findings indicate that the use of Tac QD instead of Tac BID for GVHD prophylaxis is beneficial for patients undergoing allogeneic HSCT without moderate toxicities. Disclosures: No relevant conflicts of interest to declare.

Description: Purpose: Acute lymphoblastic leukemia (ALL) remains a difficult disease to treat in adults and less than 40% of ALL patients could be alive at 5 years post-diagnosis. We investigated whether intensive conditioning regimens improve outcome of patients with ALL. Methods: Retrospectively, we analyzed 24 patients (median age; 29 years) with ALL who underwent allogeneic transplantation in our institute between February 1991 and October 2003. The median follow-up time was 7.7 years. Sixteen patients were in first complete remission (CR1) and five patients were in second CR (CR2) and three patients did not attain CR (non-CR) at the time of transplantation. Five patients with Philadelphia (Ph) chromosome-positive ALL were included. Preparative regimens were cyclophosphamide (CY), total body irradiation (TBI) and etoposide based regimen (n=23) or CY, TBI, and cytosine arabinoside (n=1) followed by marrow (n=21) or peripheral blood (n=3) transplant from unrelated (n=7) or related donors (n=17). Results: The 8-year overall survival (OS) rate and the 8-year disease-free survival rate for all cases were 48% and 39%, respectively. The 8-year OS of Ph negative and positive patients were 56% and 20%. The 8-year OS of CR1, CR2 and non-CR patients were 50%, 60% and 0%, respectively. Overall probability of the 8-year relapse rate was 47%. Treatment related mortality was 25%. Conclusion: These results suggest that CY/TBI based intensive regimens are feasible for ALL patients and may be effective for Ph negative ALL patients in not only CR1 but also CR2.

Description: Purpose: High-intensity conditioning regimens before allogeneic stem cell transplantation (all-SCT) could reduce relapse but may increase nonrelapse mortality. We have investigated the efficacy and feasibility of an intensive preparative regimen of busulfan (BU), cyclophosphamide (CY), and total body irradiation (TBI) for patients with myeloid malignancies. Patients and methods: From November 1990 to February 2005, 80 patients (acute myeloid leukemia (AML) 33, chronic myeloid leukemia (CML) 29, myelodysplastic syndrome (MDS) 18) treated with BU (8 mg/kg), CY (120 mg/kg), and TBI (10 Gy) followed by related (n=46) or unrelated (n=34) allo-SCT. Thirty-eight consecutive patients with AML in 1st remission, CML in chronic phase, and MDS (refractory anemia) defined as a standard risk group, and 42 patients with advanced AML, CML, and MDS (more than 5% marrow blasts) defined as a poor risk group. RESULTS: With a median follow-up was 6.9 years, the 7-year actuarial overall survival (OS) rate was 58% (standard risk 70% vs poor risk 47%, p=0.0076), and the probability of 7-year relapse rate was 18% (standard risk 17% vs poor risk 18%, p=0.4777). Nonrelapse mortality was 29% (standard risk 26%, poor risk 31%). Multivariate analysis indicated that age older than 40 years (p=0.018) and received allo-SCT before 1998 (p=0.006) had independent predictive value for nonrelapse mortality. CONCLUSION: The BU- CY- TBI regimen resulted in decreased relapse, especially in poor risk patients. The results of this study suggested that the BU- CY- TBI is an effective and feasible preparative regimen for allo-SCT in patients with myeloid malignancies.

Description: Abstract 4342 (Purpose) We retrospectively analyzed the data regarding patients who received fractionated administration (FR) of gemtuzumab ozogamicin (GO) and standard administration (ST) of GO for the treatment of their refractory and relapsed acute myeloid leukemia in order to review the efficacy and safety of GO in each type of administration. (Method) The patients with relapsed and refractory acute myeloid leukemia (excluding patients with acute promyelocytic leukemia) who were treated with the ST of GO received 2 doses of monotherapy with 9 mg/m2 GO as a 2-hour intravenous (iv) infusion with a 14–28-day interval. The fractionated GO group received 2 doses of GO monotherapy administered as a 2-hour iv infusion of 3 mg/(m2·day) on days 1, 3, and 5 with a 14–28-day interval. (Result) Eleven patients received ST and 9 received FR. The median age of all patients was 66.1 (55–77) years. Overall response rates (CR+CRp) in the ST and FR groups were 27% and 33%, respectively. The adverse events (grade 3–4) in the ST and FR groups were neutropenia (100% vs. 100%), thrombocytopenia (90.9% vs. 88.9%), and hypertransaminasis (18.2% vs. 0%). Grade 3–4 hypertransaminasis was seen only in the ST group. Infusion reactions occurred in 5 patients: 3 in the ST group and 2 in the FR group. The median CD33 positivity of blast cells of patients who received CR or CRp was 90.9% (78.2–99.5%). (Discussion) The response rates of the ST and FR groups were similar. However, the incidence of grade 3–4 hypertransaminasis tended to be higher in the ST group. These data suggest that FR of GO is less toxic than ST. Thus, FR of GO is safer than ST even for Japanese patients with acute myeloid leukemia. Disclosures: No relevant conflicts of interest to declare.

Description: Introduction Reactivation of HHV-6 and HHV-7 (HHV-6/7) are associated with encephalopathy, skin rush, institutional pneumonia, and cytopenia in patients with hematological malignancies. Our colleges in the department of virology reported that reactivation of saliva HHV-6/7 was measurable in healthy person and non-immunosuppressive patients although reactivation of serum HHV-6/7 was not detected. Hence determination of HHV-6/7 from salvia sample must be more sensitive than from serum sample. Clinical significance of reactivation of HHV-6/7 has not been well-analyzed in multiple myeloma (MM) patients treated with novel agents. Patients and Methods We prospectively analyzed saliva sample from MM patients treated with bortezomib (BOR), lenalidomide (LEN) or thalidomide from May 2013 to January 2016. Saliva sample was harvested on day 1 of each cycles. DNA copy numbers of HHV-6/7 in saliva samples were also measured using real-time PCR. We analyzed factors associated with reactivation of HHV-6/7 in univariate analysis by Fisher's exact test. We evaluated the correlation between HHV-6/7 ratio and M-protein ratio, which was calculated using DNA copy number of HHV-6/7 and M-protein level at each time points, by Pearson's product moment correlation coefficient. The cutoff value of reactivation of HHV-6 and HHV-7 were defined as 1 x103 and 1 x105 copy/mL. High and low corticosteroid (CS) groups were defined depending on daily estimated dose of predonisolone; the cutoff value was 100mg/body. Results One hundred and ninety-one saliva samples were collected from 20 patients. Median age was 69 years. Patients with newly diagnosed and relapsed or refractory MM were 14 and 6. Seventeen patients were treated with bortezomib (BOR) and 7 patients were treated with lenalidomide (LEN). All patients received either dexamethasone or predonisolone. The cumulative incidence of reactivation of HHV-6 and HHV-7 were 78.2% and 65.5%. High CS was related with reactivation of HHV-6 and HHV-7. LEN was related with only reactivation of HHV-7 while BOR did not induce reactivation of HHV-6/7. There was significant reverse correlation between HHV-7 ratio and M-protein ratio (r=-0.34, P

Description: Abnormalities of mitochondrial DNA (mtDNA) are responsible for a variety of inherited syndromes and have been broadly implicated in aging, cancer, and autoimmunity diseases. Mutations in mtDNA have been reported in myelodysplasia and leukemia, although their pathogenic mechanism remains uncertain. We have described age-dependent accumulation of mtDNA mutations, leading to a high degree of mtDNA sequence heterogeneity among normal marrow and blood CD34 cells as well as in granulocytes (Shin M et al, Blood101:3118 [2003], 103:553 [2004], 103:4466 [2004]). In order to examine mtDNA heterogeneity in detail, we developed a method for analysis of the mtDNA control region from single cells that were sorted by flow cytometry. Highly purified populations of CD34 cells, T cells, B cells, and granulocytes were obtained from five healthy adult donors. The sequence of the individual cells’ mtDNA was compared to the aggregate mtDNA for the respective cell type and differences were expressed as a measure of mtDNA heterogeneity among cells. Overall, heterogeneity was high: for circulating CD34 cells, 38±3.4%; for T cells, 37±14%; B cells, 36±10.8%; and for granulocytes, 48±7.2% (the value for granulocytes statistically differed from CD34 cells; p = 0.03). Most intercellular heterogeneity was due to polyC tract length variability; however, mtDNA base substitution mutations were also prevalent: 15±5.5% in CD34 cells; 15±9.0% in T cells, 15±6.7% in B cells; and 33±2.4% for granulocytes (granulocytes were significantly higher than other cells; p 〈 0.01). The higher rate of base substitution in granulocytes may reflect their greater exposure to reactive oxygen species. Surprisingly, for both polyC tract length differences and point mutations, the specific mtDNA abnormalities and the proportion of circulating cells characterized by these changes were similar among different cell lineages and relatively constant over time (~2 years) in the same donors. One inference from these results is that mtDNA heterogeneity during development is fixed in the primitive lymphohematopoietic stem cell compartment. In contrast to normal adults, circulating CD34 cells from patients obtained even years after successful allogeneic stem cell transplantation showed a remarkable level of mtDNA homogeneity, similar to the uniformity we have previously observed in cord blood CD34 cells and consistent with limited numbers of stem cells active in these individuals. Leukemic blast cells (from patients with AML-M2, AML evolving from CMML, and T-PLL) also showed a high degree of homogeneity. We propose that mtDNA sequence of single cells may be utilized as a natural genetic marker of hematopoietic progenitors and stem cells; to detect minimal residual disease in leukemia; and as a measure of the accumulation of mutagenic events in mammalian cells in vivo and in vitro.

Description: [Background] In JALSG Ph + ALL protocol(IDEAMOP) study, combination therapy with conventional intensive chemotherapy with imatinib (600mg/day) provided high quality of hematologic and cytogenetic (94%) and molecular (77%) complete remission (CR) for the patients with newly diagnosed Ph + ALL(Blood2004;104:748a #2736). However, a few of patients attained molecular CR relapsed subsequently, and allogeneic hematopoietic stem cell transplantation (allo-HSCT) should be considered for younger patients as possible as they can. In order to investigate a role of allo-HSCT post IDEAMOP therapy, three patients treated in our institute were analyzed efficacy and toxicity of allo-HSCT. [Patients and Methods] Those three newly diagnosed Ph + ALL patients (M/F:2/1 Age:30,31,38 y.o.) were treated with IDEAMOP between Jan 28,2003 and Feb 13,2004. All patients attained CR after single course of induction therapy and median time of CR was 21 (19–27) days. During consolidation therapies, two of them found HLA-matched unrelated donors and one patient chose to receive cord blood transplantation. As conditioning regimen for allo-HSCT, all of them received total body irradiation, high-dose cyclophosphamide and high-dose etoposide before allo-SCT. [Results] Allo-HSCT was undergoing during their first CR(CR1). Regimen related toxicity and complications of the transplantation were well tolerated. The reconstitutions of bone marrow function were similar to other patients with Ph-negative ALL treated by allo-HSCT. Although two patients presented with either limited or extensive chronic GvHD, they were clinically controlled with or without steroid. Minimal residual disease (MRD), measured by RQ-PCR assay, was detected in two of three patients before allo-HSCT. However, MRD was not detected in any of the patients after allo-HSCT. All of three patients remain in CR at a median follow-up time of 25 (18–31) months. Allo-HSCT post JALSG-Ph+ALL (IDEAMOP)protocol Patient Status at HSCT MRD pre HSCT Cell Source MRD postHSCT cGvHD Outcome OS(M) 30Y M CR1 yes Cord Blood no limited alive 31+ 31Y M CR1 no MUD no extensive alive 25+ 38Y F CR1 yes MUD no extensive alive 18+ [Conclusion] Although the number of patients are small, imatinib (600mg/day) with intensive chemotherapy as remission induction for newly diagnosed Ph + ALL may have benefit for rapid control of disease, and may provide better outcome of allo-HSCT without additional severe toxicities.