Refractory Anemia With Excess Blasts In Transformation Research Articles

Second Malignancy after Childhood AML Treatment - a Retrospective Analysis of the AML-BFM Stud

In the past decades the outcome in ped AML patients improved continuously. However, secondary malignancies seem to be a severe side-effect, affecting the quality of life in survivors. This retrospective data analysis in patients treated according to AML-BFM protocols, is focused the occurrence of secondary malignancies after AML treatment and evaluates the consequences thereof. Between 1993 - 2019 (total n=2797) 51 patients were documented with a secondary malignancy (SM) after AML treatment. 5 pat (10%) were excluded due to a lack of data concerning the SMs exact diagnosis or a clear differentiation from AMLs relapse. Patients were included if the de novo AML occurred before <18yrs. 46 pat (f=20; m=26) remained in the analysis. All pat were treated acc to the current AML-BFM protocol version at the time of their AML diagnosis and for their SM acc. to local standard of care. For AML treatment the cumulative dose of i.v.-cytarabine ranged from 41,380 to 51,380 mg/m2(median 47,380 mg/m2), i.th. cytarabine from 140 to 370 mg age and weight-depended (median 330 mg), etoposide was given constantly with 950 mg/m2 and from 2004 to 2017 anthracyclines with cumulative doses of 500 mg/m² were applied (idarubicin/mitoxantrone calculated with equivalence dosage of 1:5). Since 2004 the i.th. therapy was added with a max. dose of 70 mg prednisolone and 84 mg MTX, both depending on age and weight. The median age on initial AML was 7.3 yrs, ranged from 0.1-17.5 yrs while the median age at date of SM diagnosis was 14.4 yrs, ranged from 0.86-30.2 yrs. The median percentage of BM blasts was 61% (0 - 100%), the median WBC of 7,200/ μl (670 - 214,000/μl) and hemoglobin was 8.7 g/dl (4.6-14.2 g/dl). Acc. to the FAB classification, FAB M4/M5 (43%) was the largest group, followed by FAB M0/M1/ M2 (35%), FAB M6/M7 (6%), M3 (6%) and others (6%). In 63% of the cohort a normal karyotype was detected. Beside a single patient (2%) all others received CR during AML treatment. The median time between AML and SM was 3.6 yrs (0.6 to 19.7 years). Interestingly the median time differs between AML followed by blood cancers with 3.2 yrs (0.8- 15.2 yrs) to 8.1 yrs (0.6-19.7 yrs) for AML followed by a solid tumor. Leukemia (n=13, 28%), such as ALL (n=10) or CML (n=3), was the most frequent SM, however also secondary myelodysplastic syndromes (n=8, 17%; including RAEB and RAEB in transformation), lymphoma (n=7, 15%), head and neck tumors (n=6, 13%), skin cancer (n=3, 7%), CNS tumors (n=3, 7%) and other diagnoses (n=6, 13%) were reported. None of the patients was reported with known genetic disorders explaining the SM, but in 9 cases (20%) pre-existing diseases (malignant and non-malignant) was documented, often as multi-morbidity (e.g., heart, lung, neurological diseases). According to a median follow-up-period of 11.7 yrs (0.6-22.3 yrs) after AML diagnosis, the 5-yr-overall survival (OS) rate of the cohort was 97±2% after AML and decreases to 78±6% from the date of SM. The OS of 97±2% resulted from a single patient, which was diagnosed with the SM and died at the same day. The 5-yr-event-free-survival (EFS), defined as death by any cause, relapse (independently of AML or SM) and third malignancy, was 87±5% vs. 71±7%. Patients suffering from blood cancers as SM (n=29, 63%) had a slightly poorer outcome in comparison to solid tumor patients (n=17, 37%) with a 5-yr-OS of 79±8% vs. 88±2% (p 0.223) and 5-yr-EFS of 65±9% vs. 82±9% (p 0.097). With a cumulative incidence of 2%, SM are extremely rare after ped AML. In contrast, in the same period three times more t-AML cases were reported, therefor AML itself is the most frequent treatment-related malignancy in ped patients as previously published. The results showed a shorter period between AML followed by blood cancers than solid tumors, which suggesting either that AML and the SM are pathophysiologically related whereas solid tumors may be therapy-induced only or that hematopoiesis is more sensitive for toxic effects or shows a higher proliferation rate. Due to the very limited case numbers and lack of further genetic results to exclude e.g. predisposition syndromes, these hypotheses are speculative and cannot be proven. Based on the frequency of SM after ped AML and the outcome thereof, a further therapy intensification to improve especially the AMLs EFS is justifiable. Caused on the results and the revealed median intervals, a long-term follow up for ped AML patients into their adult life is highly recommendable to learn more about these rare events.

Refractory Anemia With Excess Blasts In Transformation (Acute Myeloid Leukemia with 20-29% Blasts) in Older Adults is Less Aggressive Than Acute Myeloid Leukemia With ≥30% Blasts

BackgroundThe current WHO Classification defines acute myeloid leukemia (AML) in most cases as a myeloid neoplasm with ≥20% bone marrow (BM) or peripheral blood (PB) blasts. However, it is controversial whether AML cases with 20-29% BM blasts, referred to as oligoblastic AML or refractory anemia with excess blasts in transformation (RAEBT), should be considered as AML or myelodysplastic syndrome (MDS) in terms of treatment approach and prognosis. MethodsWe retrospectively studied 573 patients aged ≥50 years diagnosed after the year 2004 with de novo AML, including 142 with 20-29% BM blasts (RAEBT) and 431 with ≥30% BM blasts (AML30). Patients with t(15;17), t(8;21), inv(16)/t(16;16), or history of any prior myeloid neoplasm were excluded. Clinicopathologic and genetic features, treatments, and outcome of RAEBT were compared to AML30. The clinicopathologic and genetic features of RAEBT were also compared to 152 de novo MDS patients with 10-19% BM blasts (RAEB2) diagnosed over the same time period. Fisher's exact and Wilcoxon tests were used to compare categorical and continuous variables between groups, respectively. Overall survival (OS) was estimated using the method of Kaplan and Meier and the log-rank test was used to compare OS between groups. Univariate and multivariable Cox proportional hazards regression was used to assess the impact of the two groups along with other risk factors in OS. ResultsCompared to AML30 patients, RAEBT patients were older and had lower white blood count and BM cellularity, but had higher hemoglobin, platelet count, and UKMRC karyotype risk score and greater morphologic dysplasia (Table 1).RAEBT and RAEB2 patients were similar with respect to age, peripheral counts, BM cellularity, and karyotype risk scores. In the subsets of patients in whom they were assessed, FLT3 and NPM1 mutations and monocytic features occurred more commonly in AML30 than in RAEBT (Table 1). RAEBT patients were treated more often with hypomethylating agents and less often with intensive therapy (anthracycline/cytarabine) compared to AML30 patients, whereas allogeneic stem cell transplant frequency was similar. With 55.5 months median follow-up, OS of RAEBT patients was superior to that of AML30 patients (p=0.003)(Figure 1). Multivariable analysis showed that RAEBT (p<0.0001), hemoglobin ≥10.3 g/dL (p=0.045), intermediate UKMRC karyotype score (p<0.0001), normal bone marrow karyotype (p=0.0005), intensive therapy (p<0.0001), and stem cell transplant (p<0.0001) were associated with longer OS.Table 1Comparison of RAEBT and AML30 groupsRAEBT (n=142)AML30 (n=431)p valueMedian age in years6763<0.0001Median white blood count (x 109/L)2.389.20<0.0001Median hemoglobin (g/dL)9.809.200.0007Median platelets (x 109/L)76600.01Median BM cellularity (%)6090<0.0001Morphologic dysplasia(% with >10% erythroid / myeloid / megakaryocytic dysplasia)53 / 53 / 4631 / 25 / 27<0.0001 / <0.0001 / <0.0001Monocytic or myelomonocytic differentiation23/115 (20%)137/342 (40%)<0.0001UKMRC karyotype scores(% intermediate / adverse / missing)61 / 35 / 569 / 26 / 50.049FLT3-ITD mutation3/94 (3%)59/247 (23%)<0.0001NPM1 mutation1/55 (2%)46/151 (30%)<0.0001Supportive care only11/142 (8%)38/431 (9%)0.86Hypomethylating agent as initial therapy36/142 (25%)36/431 (8%)<0.0001Intensive therapy as initial therapy73/142 (51%)310/431 (72%)<0.0001Stem cell transplant41/142 (29%)118/431 (27%)0.7412-month OS [95% CI]60% [46-74%]51% [47-55%]0.00348-month OS [95% CI]29% [22-36%]20% [17-24%]▪ ConclusionsRAEBT is more similar to RAEB2 than to AML30 in terms of its clinicopathologic and cytogenetic features. In spite of less frequent treatment with intensive chemotherapy and higher karyotype risk, RAEBT patients had superior OS compared with AML30 patients. In a prior study (Estey E et al. Blood 1997;90:2969), RAEBT and AML30 patients treated with intensive therapy had similar outcomes; the advent of hypomethylating agent therapy may in part account for the differences found in our study. Our findings favor considering RAEBT in a disease category separate from AML with higher blast counts. Disclosures:No relevant conflicts of interest to declare.

Subanalysis of Patients with Secondary Acute Myeloid Leukemia (sAML) with Refractory Anemia with Excess of Blasts in Transformation (RAEB-t) Enrolled in a Phase 3 Study of CPX-351 Versus Conventional 7+3 Cytarabine and Daunorubicin

Subanalysis of Patients with Secondary Acute Myeloid Leukemia (sAML) with Refractory Anemia with Excess of Blasts in Transformation (RAEB-t) Enrolled in a Phase 3 Study of CPX-351 Versus Conventional 7+3 Cytarabine and Daunorubicin

Anabolic Steroids in Myelodysplastic Syndromes: A Systematic Review

Background Anabolic steroids, such as danazol, have been used for years in the treatment of myelodysplastic syndromes (MDS). Their successful use has been documented in aplastic anemia, hypoplastic MDS, and immune thrombocytopenia, among other hematologic conditions. In patients with low-risk MDS, available treatment options are limited. Danazol is frequently used to improve symptoms and modify the disease course; however, its efficacy and optimal dosing schedule are unclear. Methods We conducted a systematic review of the literature to identify studies that used anabolic steroids, including danazol, in the treatment of MDS. In accordance to guidelines, our systematic review protocol was registered with the International Prospective Register of Systematic Reviews (PROSPERO) under ID CRD42019121467. We included case series with more than 15 patients, observational studies (cohorts and case control), and randomized trials where anabolic steroids were used to treat MDS. Following PRISMA guidelines, 2 independent reviewers assessed the studies obtained through the search strategy. Discrepancies were resolved by a third reviewer. Due to the heterogeneity of the findings, no meta-analysis was performed. Results A total of 1000 studies were identified through our search. After removing duplicates and undergoing Level 1 screening by 2 reviewers, 69 full text articles were assessed. Thirteen studies were included in the final review as the other studies did not meet inclusion criteria. Of these, 3 were retrospective cohorts, 8 prospective cohorts, and 2 randomized trials. A total of 366 patients with MDS were exposed to anabolic steroids throughout the studies with diverse outcomes reported. FAB classification was used by most of the studies to categorize MDS, with one exception that used the 2008 WHO classification. Diagnoses included refractory anemia, refractory anemia with ring sideroblasts, refractory anemia with excess blasts (RAEB), RAEB in transformation, and chronic myelomonocytic leukemia. All but 2 studies used danazol as their intervention, with the exceptions using other anabolic steroids as well as danazol. Dosage varied across studies with the majority aiming for a dose of danazol 600mg daily. Patient characteristics and interventions are summarized in Table 1. The intervention was sustained for at least 1 month and up to 12 months across studies. Outcomes reported are diverse with positive and negative studies identified. Unfortunately, outcomes are not comparable due to the presence of different response definitions with different hematologic improvement cutoffs. Reports of response were highly variable, with studies showing 5 - 75% response rates depending on their study response definitions. Multiple studies reported a trend in platelet count improvement. Response duration was not specified by all studies and was highly variable across the reports. Response definitions and outcomes are shown in Table 2. No significant adverse effects to danazol were reported in any of the studies reviewed. No specific analysis of benefit throughout the different MDS groups was reported in any of the studies. Conclusion Our systematic review highlights the significant lack of data and irregular results across studies. The heterogeneity of included populations, MDS classification, response criteria, and interventions, do not allow for evidence based recommendations regarding the use of danazol or anabolic steroids in MDS patients. Given the limited treatment options for low-risk MDS patients, particularly in low and middle income countries, further well designed studies are needed. Disclosures No relevant conflicts of interest to declare.

Impact of HFE gene Variants on Iron Overload, Overall Survival and Leukemia-Free Survival in Myelodysplastic Syndromes

Myelodysplastic syndromes (MDS) are myeloid neoplasms defined by peripheral cytopenia, dysplasia of myeloid cells and an increased risk to transform to secondary acute myeloid leukemia (AML). Although several laboratory and molecular risk factors and scoring systems have been established, little is known about genetic features contributing to disease evolution, AML development, and survival. Accumulating evidence suggests that iron metabolism and transfusion frequency are important prognostic determinants in MDS. It has also been described that patients with MDS frequently display HFE gene variants. However, little is known about the clinical impact of HFE variants. We examined the HFE status by restriction fragment length polymorphism (RFLP) analysis covering 2 common variants (H63D and C282Y) in 167 patients with MDS (observation-period: 1992-2019) and in 494 healthy Austrian controls. The median age of our MDS patients at diagnosis was 69 years (range 27-85 years) with a f:m ratio of 1:1.13. In the MDS cohort the international prognostic scoring systems, IPSS and IPSS-R, were applied, and in 94 patients a ´myeloid mutation´ screen was performed by next-generation sequencing (NGS). In all patients, serum ferritin levels were recorded at diagnosis and during follow up. Variant HFE species (H63D and/or C282Y) were detected in 63/167 patients with MDS (38.4%) and in 170/494 controls (34.4%). Median ferritin levels at diagnosis were slightly higher in HFE-mutated patients (409 mg/dL; range: 23-7415 mg/dL) compared to non-mutated patients (346.5 mg/dL; range: 10-5450 mg/dL) (p=0.62). During the first 2 years of transfusion therapy, HFE-mutated patients showed a slightly faster increase in serum ferritin levels compared to non-mutated patients. However, again, the difference was not significant, and patients with massive iron overload were found in both cohorts. When examining French-American-British (FAB) subgroups of MDS, the percentage of patients withHFE variants was higher in the group with refractory anemia, RA (22/53=41.5%) or RA with ring sideroblasts, RARS (17/39=43.9%) than in the group of RA with excess of blasts, RAEB (16/46=34.8%) or RAEB in transformation (5/17=29.4%). In CMML, 5/12 patients (41.7%) were found to carry a HFE variant. Clear differences were also observed between subgroups of MDS patients classified by World Health Organization (WHO) criteria. The prevalence of HFE mutations was 50.0% (13/26) in patients with ring sideroblasts (RS) and multilineage dysplasia (MDS-RS-MLD), 50.0% in MDS-MLD (6/12), 43% (7/18) in MDS with RS and single-lineage dysplasia (MDS-RS-SLD), 42.9% (12/28) in patients with excess of blasts-1 (MDS-EB1), 38.9% (7/18) in MDS-SLD, 21.4% (6/28) in MDS-EB2, and 20% (2/10) in MDS with isolated 5q-. When splitting patients according to the IPSS and IPSS-R, patients at highest risk were found to have a lower prevalence of HFE variants (IPSS high: 20%; IPSS-R very high: 15.4%) compared to other risk groups (IPSS low: 34.9%, int-1: 42.0%, int-2: 31.0%; and IPSS-R very low: 33.3%, low: 38.5%, int: 36.8%, high: 42.9%). Next, we examined correlations between HFE variants and somatic myeloid mutations in our MDS patients (n=94). No significant correlations were found when comparing expression of HFE variants with somatic mutations in ASXL1, ASXL1, SF3B1, SRSF2, TET2, DNMT3A, RUNX1, NRAS, TP53, IDH2 or EZH2 detected by NGS. Finally, we examined the HFE status in the context of clinical end points. In these studies, we found a clear difference in AML-free survival between HFE-mutated and non-mutated patients, although the difference did not reach statistical significance (p=0.089) (Figure 1A). No differences in overall survival (OS) were found when comparing patients with or without HFE variants (Figure 1B). In conclusion, the HFE variants H63D and C282Y are frequently detected in Austrian patients with MDS. These variants are more commonly detectable in low risk FAB patients and correlate roughly with higher ferritin levels at diagnosis and during transfusion therapy. In addition, HFE-mutated MDS patients have a slightly better AML-free survival. However, the correlation is weak. Therefore, we currently do not recommend screening for HFE gene variants at diagnosis in patients with MDS unless ferritin levels are excessively high or increase very rapidly during transfusion suggesting the possibility of a concomitant primary hemochromatosis. Disclosures Valent: Blueprint: Research Funding; Celgene: Honoraria; Deciphera: Honoraria, Research Funding; Pfizer: Honoraria; Novartis: Consultancy, Honoraria, Research Funding. Sperr:Novartis: Honoraria; Celgene: Consultancy, Honoraria.

A Phase 3 Study of CPX-351 versus Conventional 7+3 Cytarabine and Daunorubicin: Subanalysis of Patients with Secondary Acute Myeloid Leukemia (sAML) with Refractory Anemia with Excess of Blasts in Transformation (RAEB-t)

A Phase 3 Study of CPX-351 versus Conventional 7+3 Cytarabine and Daunorubicin: Subanalysis of Patients with Secondary Acute Myeloid Leukemia (sAML) with Refractory Anemia with Excess of Blasts in Transformation (RAEB-t)

Association of red cell distribution width with clinical outcomes in myelodysplastic syndrome

Studies showed red cell distribution width (RDW) can improve the detection of morphological changes in red blood cells and the understanding of their contribution to dyserythropoiesis in myelodysplastic syndrome (MDS). The purpose of the study was to evaluate dyserythropoiesis in MDS by RDW analysis and to explore the utility of RDW in clinical practice. We retrospectively analyzed laboratory and clinical data of 101 patients (59 patients was refractory anemia (RA) according to the French-American-British (FAB) classification). In patients with RA, RDW was showed weak inverse correlation with both hemoglobin concentration (Hb) (rs = −0.37, P = 0.0035) and mean corpuscular hemoglobin concentration (MCHC) (rs = −0.36, P = 0.0047). On the other hand, RDW was showed weak correlation with the number of ringed sideroblasts in bone marrow (rs = 0.31, P = 0.023). The increased RDW (≥15.0%) was associated with shorter overall survival (OS) (P = 0.0086). In patients with refractory anemia with excess blasts (RAEB) and RAEB in transformation (RAEB-t), effect of RDW on OS was less evident. These results suggested that increased RDW might reflect dyserythropoiesis, associated with deregulated hemoglobin synthesis and iron metabolism in MDS. Furthermore, increased RDW may have potential to be a prognostic significance in RA.

311 - Subanalysis of Patients with Secondary Acute Myeloid Leukemia (sAML) with Refractory Anemia with Excess of Blasts in Transformation (RAEB-T) Enrolled in a Phase 3 Study of CPX-351 Versus Conventional 7 + 3 Cytarabine and Daunorubicin

311 - Subanalysis of Patients with Secondary Acute Myeloid Leukemia (sAML) with Refractory Anemia with Excess of Blasts in Transformation (RAEB-T) Enrolled in a Phase 3 Study of CPX-351 Versus Conventional 7 + 3 Cytarabine and Daunorubicin

Aberrant Patterns of Alternative Splicing Are Frequent Events and Harbor Prognostic Significance in Patients with Myelodysplastic Syndrome

▪BackgroundAberrant patterns of alternative splicing (AS) are reported in acute myeloid leukemia and other malignances, but have not yet been investigated in myelodysplastic syndrome (MDS). In addition, mutations in spliceosome genes are especially common in MDS. Therefore, it would be highly interesting to analyze the clinical and biological significance of aberrant AS in MDS.Materials and MethodsA total of 243 newly diagnosed MDS patients, including 78 with refractory anemia (RA), 22 with RA with ring sideroblasts(RARS), 76 with RA with excess blasts (RAEB), 24 with RAEB in transformation (RAEBT), and 43 with chronic myelomonocyticleukemia (CMMoL), who had complete clinical and genetic information were enrolled. We also included 20 healthy donors of hematopoietic stem cell transplantation for comparison. We extracted RNA from mononuclear cells (MNC) isolated from the bone marrow (BM), followed by hybridization on the microarrays of AffymetrixHuman Transcriptome Array 2.0, which contained > 6 million distinct probes covering coding, non-coding transcripts, and the exon-exon junctions among 67539 genes (44710 coding genesand 22829 noncoding genes). This comprehensive coverage of exons and junctions facilitated investigation of the splicing patterns of the genes. The aberrant AS was analyzed with the splicing index (SI) method using the default setting of AffymetrixTranscriptome Analysis Console version 3.0 software. The SI of a probe in a gene was derived from the ratio between MDS patients and normal controls in terms of the probe intensity divided by the gene expression level, ie. [intensityof probe A in gene A /gene A expression level]MDS/[intensity of probe A in gene A /gene A expression level]normal control. SI had to be less than -2 or more than 2 to be defined as an aberrant AS event.ResultsTotally 52730 of 67539 genes (78.1%) on the array were expressed in BM from both MDS patients and normal donor controls. Approximately 17565 of the 52730 expressed genes (33.3%) were differentially spliced between the MDS and normal marrow MNC (P < 0.05). As a whole, 88.9% of these aberrant spliced events were mapped to coding genome and 11.1% to noncoding regions. These aberrant AS exons fell into five common patterns, including exon skipping (57.8%), alternative donor sites (18.1%), alternative acceptor sites (17.8%), intron retention (5.6%) and mutually exclusive exons (0.7%). In average, when compared with the normal marrow MNC, there were 6.4 (4.8-12.0) more aberrant AS events in each AS gene (ASG) in the marrow MNC of 243 MDS patients.More aberrant AS events per ASG implied a more complex aberrant AS pattern.The numbers of aberrant AS events per ASG were not significantly associated with MDS subtypes and frequencies of spliceosome gene mutations. However, with a median follow up of 48.4 months, the patients with more aberrant AS events per ASG had a trend of shorter overall survival (OS) (median 20.1 vs 29.6 months, P=0.074) and a trend of shorter time to acute leukemic transformation (P=0.212) than the others (Fig.1). Further subgroup analysis showed a significantly shorter OS in the patients with more aberrant AS events per ASG than those with less events among RAEB patients (median OS: 10.8 months vs 16.9 months, P=0.007, Fig.2A). In addition, RA patients with more aberrant AS events per ASG had shorter time to acute leukemic transformation than RA patients with fewer aberrant AS events per ASG (P=0.006, Fig.2B). Among the patients without detectable mutations in spliceosome genes or genes related to epigenetic modification, those with more aberrant AS events per ASG also had a shorter OS than the others (median OS: 27.9 months vs not reached, P=0.013, Fig.3). Multivariate analysis in our cohort of 243 MDS patients revealed that higher aberrant AS events per ASG was an unfavorable prognostic factor for OS (P=0.016) independent of age, IPSS-R risk score, and mutations of SF3B1, ASXL1 and TP53.ConclusionTo our knowledge, we are the first to present the prognostic impact of aberrant AS pattern in MDS patients. Large prospective cohorts are needed to confirm our observations. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Impact of Somatic Mutations on the Outcome of Children and Adolescents with Therapy-Related Myelodysplastic Syndrome

Myelodysplastic syndromes (MDS) following chemo- and/or radiotherapy for primary malignancy in childhood are clinically heterogeneous and generally resistant to therapy. Current knowledge about the clinical course and the underlying genetic background is imprecise with the exception of structural complex karyotype indicating a particular dismal prognosis. Here, we employed a next generation sequencing (NGS) of 28 genes previously established to assess the mutational landscape in primary pediatric MDS. Using Ampliseq PCR enrichment and Miseq NGS we deep sequenced DNA from bone marrow of 80 patients (pts) with therapy-related MDS (t-MDS) included in the registry of the European Working Group of MDS in Childhood (EWOG-MDS). The 41 males and 39 females had been diagnosed of t-MDS at a median age of 13 years (yrs, range 1.2-22) following treatment of hematopoietic malignancy (HM, 44), solid tumor (ST, 22) or brain tumor (BT, 14). The interval from primary malignancy to t-MDS was 3.2 yrs (0.5-18.5). Morphology was compatible with refractory anemia with excess of blast (RAEB, 34), RAEB in transformation (RAEB-t, 9), MDS related acute myeloid leukemia (MDR-AML, 4), chronic myelomonocytic leukemia (CMML, 14) or MDS without increase of blasts (19). Cytogenetic aberrations were seen in 80% of the patients, most frequently monosomy 7 (29%) and a structural complex karyotype (22%) detected in all morphological entities and subgroups of primary malignancy.Mutations were noted in 50% of pts with 1, 2, 3 or 4 genes affected in 33%, 10%, 6% and 1% respectively. Recurrent mutations were identified in RUNX1 (13.8%), TP53 (12.5%), CBL (8.8%), ASXL1 (7.5%), PTPN11 (7.5%), SETBP1 (5.0%), NRAS (5.0%), KRAS (3.8%), BCORL (2.5%), CSF3R (2.5%), HOXA9 (1.3%), EZH2 (1.3%), PTEN (1.3%), JAK3 (1.3%), STAG2 (1.3%), RAD21(1.3%). No mutations were detected in GATA2, CEBPA, GATA1, BCOR, FLT3, JAK2, cKIT, MPL, CTCF, VPS45, CALR, SH2B3. TP53 mutations arose as an isolated genetic event in the majority of cases (8/10) unlike RUNX1 mutations frequently occurring in combination with other genetic lesions (9/11). According to primary malignancy TP53 mutations were detected in all subgroups (14% HM, 63% ST, 14% BT), whereas RUNX1 mutations were absent in the group with prior solid tumor. Mutations in TP53 were strongly correlated with a structural complex karyotype (9/10). With the exception of MDS without increase of blasts characterized by the absence of RAS pathway alterations, the mutational landscape did not segregate with the morphological subtype. With a median follow-up of 5.3 yrs (0.1-13.7) for pts alive and 68/80 pts treated with hematopoietic stem cell transplantation (HSCT), the probability of overall survival (OS) at 5 yrs was 0.44. OS was worse for patients with more advanced disease (RAEB-t/MDR-AML 0.12 vs CMML 0.29 vs RAEB 0.49 vs MDS without increase of blasts 0.61, p=0.07) and t-MDS following solid tumor (ST 0.30 vs HM 0.49 vs BT 0.46, p=0.09). As expected the detection of a structural complex identified a group with a dismal outcome (OS 0.09 vs 0.37 with normal karyotype vs 0.55 with monosomy 7/ 7q- vs 0.62 with other cytogenetic aberrations, p<0.01). Assessing the impact of mutations on outcome, pts with any mutation had a worse outcome compared to pts with no mutation (OS 0.25 vs 0.60, p<0.01). Mutations in TP53 and RAS pathway were associated with a lower OS of 0.00 and 0.19, respectively, whereas RUNX1 mutated cases without TP53 or RAS pathway alteration had an excellent outcome with 5/5 pts being alive.This study demonstrates that the outcome of patients with t-MDS is variable and may currently be best defined by the genes mutated. Although the germline status was not tested in all cases with TP53 mutation, the study indicates that presence of somatic TP53 or RAS pathway mutations will help identifying patients that are at high risk of treatment failure and therefore candidates for experimental therapy. At the same token, the majority of t-MDS patients without these high risk mutations can successfully be treated with HSCT. Thus, future counseling of patients with t-MDS and their parents will require individual molecular genotyping. DisclosuresNo relevant conflicts of interest to declare.

A new complex karyotype in a unique de novo myelodysplastic syndrome case involving ten chromosomes and monoallelic loss of TP53

Myelodysplastic syndrome (MDS) is a group of clonal stem cell disorders characterized by cytopenia, dysplasia in one or more cell lineages, and ineffective hematopoiesis. MDS is associated with high risk of progression to acute myeloid leukemia. At initial diagnosis, clonal cytogenetic aberrations are present in 40–70% of patients with de novo (primary) MDS and in 65–95% of patients with therapy-associated ones (t-MDS). Most commonly observed abnormalities present in MDS are monosomy 5 and 7, trisomy 8, deletions of long arm of chromosome 20 as well as complex karyotypes. Loss or gain of chromosomal material is known to be related to disease development and progression.In a de novo adult MDS case banding cytogenetics, refined by array-proven multicolor banding (aMCB) revealed a unique complex karyotype involving ten chromosomes that include del(5q), del(7q), deletions in parts of both chromosomes 10, and a dic(7;17). Interestingly, the dic(7;17) leads to monosomy of the tumor suppressor gene TP53. The patient had an immunophenotype consistent with refractory anemia with excess blasts in transformation (RAEB-t) according to French–American–British (FAB) classification.To the best of our knowledge, a comparable adult MDS case associated with such a complex karyotype and loss of TP53 was not previously reported. As most of complex aberrations were found simultaneously in all studied malignant cells this may be a hint on an initial one step development of this complex rearrangement by chromothripsis.

Interactions and relevance of blast percentage and treatment strategy among younger and older patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS).

Acute myeloid leukemia (AML) is defined as ≥20% myeloblasts, representing a change from original guidelines where ≤30% blasts were considered as myelodysplastic syndromes (MDS), and 20-29% blasts classified as refractory anemia with excess blasts in transformation (RAEB-T). Whether the diagnostic bone marrow blast percentage has current value with regards to patient prognostication or identification of optimal treatment strategies is unclear. We retrospectively studied 1652 treatment-naïve adults with MDS or AML and ≥10% blasts from January 2000 to April 2014. Patients with 20-29% blasts were more similar to MDS patients in terms of advanced age, increased frequency of poor-risk cytogenetics, lower WBC count, and less frequent NPM1 and FLT3-ITD mutations. Median overall survival of MDS and RAEB-T were similar, 16.0 and 16.0 months, compared to 13.5 months for AML with ≥30% blasts (P = 0.045). Multivariate analysis showed inferior survival with increased age (HR 1.81 age 60-69, HR 2.68 age ≥70, P < 0.0005); poor-risk cytogenetics (HR 2.25, P < 0.0005); therapy-related disease (HR 1.44, P < 0.0005); and markers of proliferative disease including WBC ≥25 × 10(9) /L (HR 1.35, P = 0.0003), elevated LDH count (HR 1.24, P = 0.0015), and peripheral blasts (HR 1.25, P = 0.004). Among younger patients (≤60 years), intensive AML-type therapy resulted in similar outcomes regardless of blast percentage, suggesting this to be optimal therapy in this context. Among older patients (≥70 years), patients with 20-29% blasts had similar outcomes to patients with <20% blasts, and better than those with ≥30% blasts. In addition, among older patients, epigenetic therapy provided at least equivalent outcome to intensive chemotherapy.

Clinical Significance and Pathopysiological Function of the Tim-3/Galection-9 Pathway in Myelodysplastic Syndromes

Abstract Background: Galectin-9 (Gal-9), a member of the galectin family, plays a crucial role in inflammation and tumorigenesis involving angiogenesis, cell adhesion, immune escape, and cancer cell survival. In acute myeloid leukemia (AML), Gal-9 is highly expressed in some AML cell lines and binds with T-cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) on T cells and leukemic stem cells. The Tim-3/Gal-9 pathway may be associated with disease progression by inducing T-cell dysregulation and mediating autocrine proliferation of leukemic cells. However, the role of the pathway in myelodysplastic syndromes (MDS) remains unclear. In the current study, we investigated the prognostic impact of plasma Gal-9 levels and the pathophysiological roles of Gal-9 and Tim-3 in MDS. Methods and results: 1) To investigate the prognostic impact of Gal-9, plasma Gal-9 levels were measured by ELISA in 92 MDS patients: 30 with refractory anemia (RA); 3 with RA with ringed sideroblasts (RARS); 18 with RA with excess blasts (RAEB); 14 with RAEB in transformation (RAEB-t); and 29 with acute leukemia transformed from MDS (AL-MDS). The plasma Gal-9 level increased with disease progression (mean ± SD: control 3.4 ± 0.9; RA, 9.3 ± 5.7; RARS, 10.2 ± 6.8; RAEB, 9.7 ± 6.5; RAEB-t, 17.2 ± 9.6; AML-MDS, 20.1 ± 14.1 ng/ml). Furthermore, patients with AL-MDS in International Prognostic Scoring System (IPSS) high-risk categories (Intermediate-2/High) had significantly higher Gal-9 levels compared with IPSS low-risk (Low/Intermediate-1) patients (Low/Intermediate-1 vs Intermediate-2/High, p = 0.002; Low/Intermediate-1 vs AML-MDS, p = 0.004). The overall survival of MDS patients with plasma Gal-9 higher than the normal level (10 ng/mL) was significantly shorter than that of other patients (log-rank p &lt; 0.001) even in lower-risk (RA, RARS) cases (log-rank p = 0.0369). Moreover, multivariate analysis showed that a high level of Gal-9 was an independent predictor of shorter survival in MDS patients as well as IPSS. 2) Bone marrow mononuclear cells were isolated from some MDS patients, and Tim-3 expression on granulocytes, monocytes, lymphocytes, and blasts was analyzed using flow cytometry (FCM). Tim-3 expression on blasts tended to be higher in AL-MDS patients than in MDS patients. 3) To investigate whether MDS cells can produce Gal-9, we analyzed Gal-9 levels in supernatants of cell cultures of 3 human AL-MDS cell lines, F-36P, SKM-1, and MDS-L, and MDS blasts obtained from an AL-MDS patient by ELISA. Gal-9 was detected in all cell culture supernatants and its concentration was higher when cultured in medium containing 10% fetal bovine serum (FBS) than in 2.5% FBS medium in F-36P and SKM-1 cells. The concentration of Gal-9 produced from MDS blasts was increased time dependently in 2-6 day culture. Furthermore, recombinant human Gal-9 enhanced the cell proliferation of F-36P cells. 4) After analyzing Tim-3 expression in MDS cell lines using real-time quantitative PCR and FCM, all cell lines expressed relatively low levels of Tim-3 mRNA. Extracellular expression of Tim-3 was detected and induced by adding cell culture supernatants of the human stromal cell line HS-5 in F-36P cells. To elucidate the signaling pathway inducing Tim-3 expression, we investigated Tim-3 mRNA expression in F-36P cells treated with HS-5 supernatant with several signal transduction inhibitors (STAT3 inhibitor, U0126; MAPK/ERK; AG490; JAK2, LY294002; PI3K, PDTC; NF-κB inhibitor). Upregulation of Tim-3 gene expression in F-36P cells treated with HS-5 supernatant was inhibited by the MEK inhibitor U0126. These results suggest that the MAPK/ERK pathway is involved in Tim-3 expression induced by HS-5 supernatant. Conclusions: In MDS, the elevation of plasma Gal-9 levels is associated with disease progression, including leukemic transformation, and with shorter survival. Furthermore, its receptor Tim-3 is upregulated on blasts in AL-MDS patients. Our data suggest that the Gal-9/Tim-3 pathway plays a key role in MDS disease progression. Although further study is required to clarify the detailed functions of the interaction between Gal-9 and Tim-3, the current study could lead to the development of a new immunotherapeutic strategy via the blockade of this pathway in MDS. Disclosures No relevant conflicts of interest to declare.

Impact of Unbalanced Translocation Der(1;7)(q10;p10) and -7/Del(7q) on the Prognostic Value after Allogeneic Hematopoietic Stem Cell Transplantation in Myelodysplastic Syndromes: A Nationwide Retrospective Study

Background:Poor-risk karyotypic abnormalities in patients with myelodysplastic syndromes (MDS) are known to have a negative impact on outcomes after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Abnormalities in chromosome 7 have classically been categorized as a poor-risk karyotype, but comprise various patterns, in which the unbalanced translocation der(1;7)(q10;p10) and -7/del(7q) are frequently observed. To accurately estimate outcomes after allo-HSCT in MDS patients with der(1;7)(q10;p10) and -7/del(7q), we herein conducted a retrospective study among adults with de novo MDS.Methods:The clinical data of patients who had either der(1;7)(q10;p10) or -7/del(7q) as the sole karyotypic abnormality and underwent allo-HSCT between January 1999 and December 2012 were collected from the Transplant Registry Unified Management Program of the Japan Society for Hematopoietic Cell Transplantation. Patients with a normal karyotype (NK) were included as a reference group. We used the French-American-British classification to define the disease status at HSCT, as described previously (Saber W, et al. Blood 2013.); refractory anemia with excess blasts (RAEB) or RAEB in transformation at any time between diagnosis and HSCT was considered to be an advanced disease status, while others were regarded as an early disease status. Causes of death were categorized into disease-associated mortality (DAM) or transplant-related mortality (TRM); the former was defined as death after the relapse or progression of MDS, while the latter was defined as any death other than that due to MDS. Cox proportional hazards regression models were used to evaluate variables potentially affecting overall survival (OS). Fine and Gray proportional hazards models were used to evaluate variables potentially affecting DAM and TRM. DAM and TRM were estimated using cumulative incidence curves to accommodate competing events.Results:Twenty-three MDS patients with der(1;7)(q10;p10), 29 with -7/del(7q), and 347 with NK were identified among those who underwent HSCT with an early disease status. The estimated 5-year OS, DAM, and TRM by cytogenetic subcategories were as follows; 47.3%, 9.9% and 42.8% for patients with der(1;7)(q10;p10); 63.5%, 11.1%, and 25.4% for patients with -7/del(7q); and 69.5%, 5.8%, and 24.7% for patients with NK, respectively. The univariate and multivariate analyses on OS, DAM, and TRM did not identify any significant differences among the cytogenetic subcategories tested.Sixty-nine patients with der(1;7)(q10;p10), 75 with -7/del(7q), and 511 with NK were identified among those who underwent HSCT with an advanced disease status. The estimated 5-year OS, DAM, and TRM by cytogenetic subcategories were as follows; 47.2%, 18.6% and 34.2% for patients with der(1;7)(q10;p10); 33.4%, 36.8%, and 29.8% for patients with -7/del(7q); and 48.0%, 21.8%, and 30.2% for patients with NK, respectively. The univariate analysis revealed that the presence of -7/del(7q) was associated with lower OS (P =.012) and higher DAM (P =.001), whereas that of der(1;7)(q10;p10) showed similar OS (P =.781) and DAM (P =.858) to those with NK. In the multivariate analysis, the presence of -7/del(7q) was significantly associated with lower OS (Hazard ratio (HR) [95% confidential interval], 1.40 [1.01-1.92]; P =.041) and higher DAM (HR, 2.02 [1.24-3.28]; P =.005) than those with NK. No significant differences were observed in TRM among the 3 cytogenetic subcategories.Conclusion:Patients with der(1;7)(q10;p10) may benefit from allo-HSCT, achieving similar outcomes to those of patients with NK. However, the presence of -7/del(7q) adversely affected OS and DAM with significance in patients with an advanced disease status. DisclosuresMiyazaki:Shin-bio: Honoraria; Chugai: Honoraria, Research Funding; Sumitomo Dainippon: Honoraria; Celgene Japan: Honoraria; Kyowa-Kirin: Honoraria, Research Funding.

Allogeneic Transplantation in Elderly Patients with AML and MDS Comparing Reduced-Intensity Conditioning with Flamsa-Busulfan Versus Fludarabine/BCNU/Melphalan

▪By reducing treatment intensity allogeneic hematopoietic stem cell transplantation (allo-HSCT) has become feasible for elderly patients. Different reduced-intensity conditioning (RIC) regimens are available, but there is little consensus about the optimal preparative regimen to use, in particular with regard to the outcomes counterbalancing the aim of feasibility and tolerability with higher rates of relapse. Here, we retrospectively evaluate the outcome of sequential therapy employing RIC with fludarabine 30 mg/m2, cytarabine 2g/m2 and amsacrine 100 mg/m2 for 4 days (FLAMSA; Schmid C et al. JCO 2005) followed by busulfan 10 x 0.8 mg/kg (FLAMSA-Bu) compared to RIC utilizing fludarabine 5 x 30 mg/m2, carmustine (BCNU) 2 x 150 mg/m2 and melphalan 110 mg/m2 (FBM; Marks R et al. Blood 2008) in elderly patients treated consecutively at our institution between July 2005 and October 2012.We analyzed the course of 114 patients (pts) with acute myeloid leukemia (AML; n=99) or myelodysplasia (MDS; n=15) aged ≥ 59 years with 59 pts aged ≥ 66 years who were treated with either FLAMSA-Bu (n=66; n=24 ≥ 66 years) or FBM (n=48; n=35 ≥ 66 years). All patients received sero-therapy with anti-thymocyteglobuline (ATG). Median patient age was 66 years for the entire cohort (68 years FBM; 64 years FLAMSA-Bu). 36 patients (75%) of the FBM and 42 patients (63 %) of the FLAMSA-Bu group suffered from high risk disease defined as relapsed or refractory AML or refractory anemia with excess of blasts in transformation (RAEB-T). The hematopoietic cell transplantation comorbidity index (HCT-CI) was higher for the patients of the FBM group than for the FLAMSA-Bu group with 26 (54 %) versus (vs) 24 patients (36 %) scoring ≥ 2 (p 0.085). Graft source after conditioning with FBM/FLAMSA-Bu was bone marrow (1/2), G-CSF mobilized peripheral blood stem cells (40/62) and double-umbilical cord-blood (7/1). In 23 pts (20 %) HLA-matched related and in 91 pts (80 %) HLA-matched unrelated donor transplantation was performed. Engraftment failure was observed in 1 patient after FLAMSA-Bu, while engraftment was achieved in all evaluable patients of the FBM group in a median of 23 days vs 18 days after FLAMSA-Bu (p 0.003), while 7 pts with double-umbilical cord-blood transplantation where included in the FBM group vs 1 pt in the FLAMSA-Bu group. Non-hematological treatment-related acute toxicity ≥ CTC III (gastrointestinal, hepatic, cardiovascular, renal, centralnervous system) occurred in 12/48 pts (25 %) after FBM and in 18/66 pts (27 %) after FLAMSA-Bu. Incidence of severe acute (III-IV) and chronic GvHD was 22.9 %/16.6 % for FBM vs 18.2 %/19.7 % for FLAMSA-Bu, respectively. After conditioning with FBM 2/48 pts vs 9/66 pts after FLAMSA-Bu were diagnosed with a secondary malignancy (p 0.08). Non-relapse mortality (NRM) after 12 months was 26.8 % for FBM versus 25.2 % for the FLAMSA-Bu group. Incidence of relapse after FBM vs FLAMSA-Bu conditioning was 22.9 % vs 15.2 % after 1 year and 31.3 % vs 16.7 % after 2 years. Occurrence of relapse was significantly related to an incomplete or mixed chimerism (donor cells ≤ 95 % in peripheral blood and/or bone marrow) at day +30 (p 0.001).After a median follow up of 31.4 months (range 4.4-97.5) estimated overall survival (OS) and relapse-free survival (RFS) after 2 years was 55.4 % and 51.4 % for the FBM vs 58 % and 56.7 % for the FLAMSA-Bu group, respectively. Analyzing different subgroups, FBM conditioning might be favorable for pts aged ≥ 66 years when suffering from high risk AML (n=26): Within this group 1-year OS after FBM vs FLAMSA-Bu was 71.4 % vs 66.7 % (p 0.58) and 1-year RFS was 71.4 % vs 58.3 % (p 0.59), respectively. Notably, for pts at highest risk (aged ≥ 66 years and suffering from secondary or therapy-related AML; n=24) the benefit of FBM conditioning becomes more pronounced: 1-year OS after FBM vs FLAMSA-Bu was 62.5 % vs 37.5 % (p 0.26) and 1-year RFS 54.2 % vs 37.5 % (p 0.17).Both conditioning regimens are feasible, and provide similar rates of acute toxicity, NRM and GvHD. There might be evidence for a benefit of conditioning with FBM for the subgroup of “the oldest patients at highest risk”. Taking into account that there is an increasing group of ‘medically fit’ elderly patients in the field of allogeneic transplantation, prospective clinical trials are needed to investigate different conditioning regimens considering their special requirements. DisclosuresNo relevant conflicts of interest to declare.

Incidence and Outcomes of Pediatric Myelodysplastic Syndrome in the US

Background There is lack of epidemiological data on pediatric myelodysplastic syndrome (p-MDS) in the literature. MDS became reportable to the Surveillance, Epidemiology and End Results (SEER) Program in 2001, providing an opportunity to estimate the incidence and survival of pediatric patients with MDS in the United States. Methods We utilized data from the National Cancer Institute SEER-18 to determine the incidence and long term overall survival (OS) of pediatric patients (ages 0 to 20 years) diagnosed with de novo MDS or therapy-related MDS. Inclusion criteria was diagnosis of MDS (International Classification of Diseases-Oncology, Third Edition, ICD-O-4 codes 9980/3, 9991/3, 9992/3, 9982/3, 9985/3, 9983/3, 9986/3, 9986/3, 9989/3, 9985/3, 9975/3, and 9987/3) between 2001 and 2011. Follow up was updated through the end of 2011 (November 2013 submission). Overall survival was estimated using the method of Kaplan-Meier. A Cox proportional hazard model was used to compare the effects of age, race, gender, histological subtype, and etiology (de novovs. therapy-related) on survival. Results The incidence of p-MDS was 1.16 cases/1 million population*year. A greater incidence occurred in children younger than 1 year of age possibly reflecting congenital bone marrow failure syndromes (Figure 1). A total of 314 p-MDS cases were included in the analysis with median follow up of 31 months (range 0-131). Median age of patients was 9 years; 167 (53.3%) had MDS unclassifiable (NOS), 40 (12.7%) had therapy-related MDS (t-MDS), 44 (14%) had refractory anemia with excess blasts (RAEB), 32 (10.3%) had refractory anemia (RA), 17 (5.4%) had refractory cytopenia with multilineage dysplasia (RCMD), 6 (1.9%) had refractory anemia with ring sideroblasts (RARS), 5 (1.6%) had refractory anemia with excess blasts in transformation (RAEBT), and 3 (0.9%) had MDS associated with isolated del(5q). Male patients comprised 154 (49%) of cases. Racial groups included white (218, 69.4%), 52 (15.7%) black, 37 (11.8%) of other races, and 7 (2.3%) the race was unknown. The 5 year-OS for the entire cohort was 68% (95% C.I.=62.3-73.7). Patients with t-MDS had significantly worse 5 year-OS (41.2%; 95%C.I.=23.8-58.6) compared to those with de novo MDS (71.3%; 95%C.I.=65.3-77.2; P=0.004, Figure 2). In multivariate analysis of age, race, gender, histological subtype, and etiology (de novovs. therapy-related) utilizing Cox regression model, only t-MDS was associated with higher risk of death (HR=2.07, 95% C.I.=1.25-3.42, P=0.005). Conclusions Pediatric MDS is a rare disorder, with higher incidence among children younger than 1 year of age. Over two thirds of p-MDS patients will become long-term survivors, although significantly inferior outcome is seen in t-MDS. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

Allogeneic hematopoietic stem cell transplantation in pediatric myelodysplastic syndromes: a multicenter experience from Argentina.

Allogeneic hematopoietic stem cell transplantation (AHSCT) represents the only curative treatment for the majority of pediatric patients with Myelodysplastic Syndrome (MDS). We aimed to evaluate overall survival (OS), disease-free survival (DFS), non-relapse mortality (NRM) and relapse incidence in children who underwent AHSCT for MDS in six institutions from Argentina. A retrospective analysis of 54 AHSCT was carried out in 52 patients (mean age: 9 years; range: 2-19; 35 males). MDS subtypes were refractory cytopenia of childhood (RCC) (n: 26, 50%), refractory anemia with excess blasts (RAEB) (n: 9, 18%), RAEB in transformation (RAEB-T) (n: 8, 15%) and juvenile myelomonocytic leukemia (JMML) (n: 9, 17%). At time of transplant, seven (13%) patients transformed to acute myeloid leukemia (AML) and two patients with RCC to RAEB. Donors were related in 32 cases (59%) and the stem cells source was: bone marrow (63%), peripheral blood (26%), and umbilical cord blood (11%). Five-year DFS and OS were 50% and 55% respectively; and for patients with JMML, 57% and 67% respectively. Cumulative incidence of NRM and relapse were 27% and 21% respectively. In the multivariate analysis, umbilical cord blood (HR 4.07; P = 0.025) and age ≥ 9 years at transplantation (HR 3.28; P = 0.017) were associated with lower OS; age and graft-versus-host disease (GVHD) had a higher NRM. In our series, more than half of the patients achieved long term OS with AHSCT. Less toxic conditioning regimens or more intensive GVHD prophylaxis could lead to better results in some children.

Cytogenetics and clinical features of pediatric myelodysplastic syndrome in Japan.

We analyzed the cytogenetics and clinical features of pediatric myelodysplastic syndrome (MDS) in Japan. Data on patients (<16 years) diagnosed with MDS from 1990 to 2000 were retrospectively collected from pediatric hematologists in 234 institutions. Chromosome analysis was successfully performed in 255 of 277 MDS patients. The numbers of patients with refractory anemia, refractory anemia with ringed sideroblasts, refractory anemia with excess of blasts (RAEB), refractory anemia with excess of blasts in transformation (RAEBt), chronic myelomonocytic leukemia, and juvenile myelomonocytic leukemia were 67 (24%), 51 (18%), 51 (18%), 20 (7%), and 65 (23%), respectively. The other 23 patients (8%) could not be classified specifically. The distribution of childhood MDS in Japan according to the French-American-British subclassification was similar to that in other countries. However, we identified a higher incidence of therapy-related cases. As for relationship between cytogenetics and prognoses, abnormal karyotypes were related to poorer prognoses than normal karyotype (P < 0.01). However, patients with trisomy 8 had prognoses comparable to those with normal karyotypes. Complex karyotypes were associated with poorer prognoses among RAEB and RAEBt patients. In conclusion, prognosis of pediatric MDS is related to cytogenetics. A more precise diagnosis and classification system is needed for childhood MDS.

CD7 Expression On Blasts Of Myelodysplastic Syndromes Is Associated With Apoptosis Resistance With Decreased Expression Of The Proapoptotic Protein Bad and An Independent Unfavorable Prognostic Factor Together With The Revised IPSS Score In Patients

IntroductionCD7 is expressed on human T and NK cells and progenitors of myeloid cells as well as blasts in some patients with myelodysplastic syndromes (MDS). We showed previously that CD7 positivity of MDS blasts was associated with aggressive characteristics and poor prognosis. However, the mechanisms by which CD7-expressing MDS blasts are associated with disease progression remain unknown. In this study, we first investigated the biology of CD7-expressing MDS blasts. Second, to validate the prognostic impact of CD7 on MDS blasts, prognostic variables including the CD7 positivity of MDS blasts and a new prognostic system, the revised International Prognostic Scoring System (IPSS-R) were evaluated in Japanese MDS patients. Methods & Results1) To investigate the mechanisms regulating CD7 expression, we used MDS cell lines HNT-34 and F-36P. CD7 expression on these cells was partially down-regulated by inhibition of NFκB. 2) To investigate the survival potential in CD7+ MDS blasts, we analyzed cell cycles and spontaneous apoptosis by flow cytometry. CD7+ blasts had a cell cycle advantage compared with CD7– blasts in F-36P but not in HTN-34 cells. Compared with CD7– HNT-34 cells, CD7+ HTN-34 cells were more resistant to spontaneous and serum deprivation-induced apoptosis. 3) We then compared gene and protein expression levels of apoptosis-related proteins including Bad, Bax, Bcl-2-L1, Bcl-2, caspase-3, caspase-8, caspase-9, FADD, Fas, and FasL between CD7+ and CD7– blasts of these cell lines using real-time PCR and flow cytometry, respectively. CD7– blasts had markedly higher expression levels of the Bad gene and protein compared with CD7+ blasts in both cell lines. The expression levels of Fas and FasL were suppressed in CD7+ blasts compared with CD7– blasts in F36P cells. These results support the association of CD7 expression on MDS blasts with disease progression. 4) To reevaluate the prognostic impact of CD7 expression as well as R-IPSS, prognostic variables were analyzed in 81 MDS patients [refractory anemia (RA) 55, RA with ringed sideroblasts 7, RA with excess blasts (RAEB) 18, and RAEB in transformation 1], comprising 50 men/31 women with a median age of 67 (range 27–88) years. Immunophenotyping was performed by 3-color flow cytometry, in which blast cells were gated with a CD45-gating method, and 9 parameters including CD7 expression on MDS blasts, i.e., CD34+ myeloblast-related and B-progenitor-related cluster size, myeloblast CD45 expression, and aberrant expression of CD7, CD10, CD11, CD15, CD56, and B7-H1 on myeloblasts, were analyzed. Using the Cox proportional hazard regression model, we identified five prognostic variables: IPSS-R score; percentage of blasts in peripheral blood; CD7 expression; gender; and white blood cell count. The chi-square test showed that the IPSS-R score and CD7 expression were strong prognostic factors (P = 0.0114 and 0.006, respectively). Patients whose MDS blasts expressed high levels of CD7 (17% or more MDS blasts were CD7+) had significantly shorter survival than other patients. ConclusionOur study revealed that CD7+ MDS blasts had apoptosis resistance with decreased expression of apoptosis-related genes, especially Bad. Signaling via CD7 on MDS blasts might inhibit Bad expression and then confer apoptosis resistance. Further studies are in progress to clarify CD7 signaling in cell lines as well as in MDS blasts from patients. Furthermore, we demonstrated for the first time that IPSS-R as well as CD7 expression on blasts had a strong impact on MDS patient prognosis. Disclosures:Kurokawa:Novartis: Consultancy, Research Funding; Bristol-Myers Squibb: Research Funding; Celgene: Consultancy, Research Funding. Shibayama:celgene: Honoraria, Research Funding; Janssen: Honoraria. Naoe:Otsuka Pharmaceutical Co., Ltd, Kyowa Hakko Kirin Co., Ltd., Wyeth, and Chugai Pharmaceutical Co., Ltd.: Research Funding.

Outcome Of Patients With Myelodysplastic Syndrome (MDS) With Bone Marrow Blasts Between 10-30% Treated With Hypomethylating Agents Versus Intensive Chemotherapy

BackgroundIn 2001, the World Health Organization modified the French-American-British (FAB) classification for myelodysplastic syndrome (MDS) by folding the refractory anemia with excess blasts in transformation (RAEB-t) category into acute myeloid leukemia (AML). Whether this group of patients (pts) should be treated with AML versus MDS therapy remains controversial. A subset analysis of the AZA-001 trial showed that azacitidine prolongs overall survival (OS) in elderly pts with low blasts AML (bone marrow blasts [BM] 20-30%). AimTo compare the clinical outcome and OS of patients with MDS or AML with BM blasts between 10-30%, treated with hypomethylating agents (HMA) vs intensive chemotherapy (IC). Patients and MethodsWe conducted a retrospective analysis of newly diagnosed pts with MDS (or AML by WHO) and BM blasts between 10-30% treated with either HMA (alone or in combination with investigational therapies) or IC on clinical trials. Eligibility was based on pt characteristics and specific protocol inclusion criteria. A univariate Cox proportional hazards regression model was used to evaluate the overall effects of treatments and outcome (remission duration (RD), and OS). Then a regression model with the interactions between treatments and baseline covariates were used for subgroup analysis. The final model was obtained by a stepwise selection using 0.05 as a cut off of significant values. Results330 patients were included in the final analysis, with 93 (28%) HMA-treated pts and 237 (72%) pts treated with IC. Clinical characteristics at diagnosis are summarized in Table1. The overall response rate (ORR= CR+CR p) was 42% for the pts who treated with HMA and 60% for pts treated with IC (P = 0.01). The median RD was similar between the two groups (14.7 mos (m) vs. 14.7 mos, respectively, P = 0.74). Early induction mortality was also similar among the two groups (4-week mortality was 5% vs. 7%, respectively, and the 8-week mortality was 10% vs. 13 %, respectively). With median follow up of 37 mos (range, 1-94 mos), the median OS was 18.8 mos for pts who treated with HMA vs. 14.6 mos for pts treated with IC (P = 0.32). Moreover, the BM blasts percentage did not impact the overall outcome. In multivariate analysis, treatment with IC was associated with worse OS compared to HMA (HR 2.09, 95% CI 2.07-3.17, P = 0.003) but not for RD (HR 0.43, 95% CI 0.89-2.68, P = 0.13).Table 1Patients' characteristics and responsesCharacteristicsAll patients No.%HMA No.%IC No.%P- valueNumber330932823772Age, yearsMedian646662<0.001Range13-9013-9017-84Age, years0.03< 6518757444714360> 651434349539440SexMale194595963135570.28Female13641343710243History of prior Cx and /or Rx0.26Yes752325275021No25577687318779IPSSHigh181604148140650.02Intermediate-21073638456932Intermediate-11246763Not applicable30NA8NA22NRCytogenetic analysisDiploid/-Y13342343999430.825-/7-1113532377934Other862327245923Complex cytogeneticsYes11234333579330.76No732220225322Bone marrow blasts %10- 2016751646910343<0.000121-3016349293113457Median WBC x109/L Range2.7(0.4 123.4)2.5(0.4-25.6)2.9(0.9-123.4)0.01Median hemoglobin g/dl Range9.4 (4.2-13.5)9.7(6.8-13.5)9.1(4.2-13.5)0.02Median platelets x103/mL Range57 (3-746)62(11-588)55(3-746)0.23Median bone marrow blasts %Range19 (10-30)15(10-30)20(10-30)<0.0001 ConclusionAlthough patients with MDS or AML with BM blasts between 10-30% initially achieve a higher ORR when treated with IC compared to HMA-based therapy, the OS was better for pts treated with HMA after accounting for all other covariates. Interestingly, BM blast percentages within this cohort did not impact overall outcome suggesting that pts with BM blasts 20-30% may achieve better outcome with MDS therapy. Disclosures:No relevant conflicts of interest to declare.