Independent Prognostic Factor For EFS Research Articles

NUP98::NSD1 and FLT3/ITD co-expression is an independent predictor of poor prognosis in pediatric AML patients

ObjectivePatients who carry NUP98::NSD1 or FLT3/ITD mutations are reported to have poor prognosis. Previous studies have confidently reported that the poor outcome in younger AML patients is owning to dual NUP98::NSD1 and FLT3/ITD positivity, with a high overlap for those two genetic lesions. In this study, we assessed the prognostic value of the presence of both NUP98::NSD1 and FLT3/ITD in pediatric AML patients.MethodsWe screened a large cohort of 885 pediatric cases from the COG-National Cancer Institute (NCI) TARGET AML cohort and found 57 AML patients with NUP98 rearrangements.ResultsThe frequency of NUP98 gene fusion was 10.8% in 529 patients. NUP98::NSD1 fusion was the most common NUP98 rearrangement, with a frequency of 59.6%(34 of 57). NUP98::NSD1 -positive patients who carried FLT3/ITD mutations had a decreased CR1 or CR2 rate than those patients carried FLT3/ITD mutation alone (P = 0.0001). Moreover, patients harboring both NUP98::NSD1 fusion and FLT3/ITD mutation exhibited inferior event-free survival (EFS, P < 0.001) and overall survival (OS, P = 0.004) than patients who were dual negative for these two genetic lesions. The presence of only NUP98::NSD1 fusion had no significant impact on EFS or OS. We also found that cases with high FLT3/ITD AR levels ( > = 0.5) with or without NUP98::NSD1 had inferior prognosis. Multivariate analysis demonstrated that the presence of both NUP98::NSD1 and FLT3/ITD was an independent prognostic factors for EFS (hazard ratio: 3.2, P = 0.001) in patients with pediatric AML. However, there was no obvious correlation with OS (hazard ratio: 1.3, P = 0.618). Stem cell transplantation did not improve the survival rate of cases with NUP98 fusion or NUP98::NSD1 AML in terms of EFS or OS.ConclusionPresence of both NUP98::NSD1 and FLT3/ITD was found to be an independent factor for dismal prognosis in pediatric AML patients. Notably, lack of FLT3/ITD mutations in NUP98::NSD1 -positive patients did not retain its prognostic value.

Fusion partner–specific mutation profiles and KRAS mutations as adverse prognostic factors in MLL-rearranged AML

AbstractMixed-lineage leukemia (MLL) gene rearrangements are among the most frequent chromosomal abnormalities in acute myeloid leukemia (AML). MLL fusion patterns are associated with the patient's prognosis; however, their relationship with driver mutations is unclear. We conducted sequence analyses of 338 genes in pediatric patients with MLL-rearranged (MLL-r) AML (n = 56; JPLSG AML-05 study) alongside data from the TARGET study's pediatric cohorts with MLL-r AML (n = 104), non–MLL-r AML (n = 581), and adult MLL-r AML (n = 81). KRAS mutations were most frequent in pediatric patients with high-risk MLL fusions (MLL-MLLLT10, MLL-MLLT4, and MLL-MLLT1). Pediatric patients with MLL-r AML (n = 160) and a KRAS mutation (KRAS-MT) had a significantly worse prognosis than those without a KRAS mutation (KRAS-WT) (5-year event-free survival [EFS]: 51.8% vs 18.3%, P < .0001; 5-year overall survival [OS]: 67.3% vs 44.3%, P = .003). The adverse prognostic impact of KRAS mutations was confirmed in adult MLL-r AML. KRAS mutations were associated with adverse prognoses in pediatric patients with both high-risk (MLLT10+MLLT4+MLLT1; n = 60) and intermediate-to-low–risk (MLLT3+ELL+others; n = 100) MLL fusions. The prognosis did not differ significantly between patients with non–MLL-r AML with KRAS-WT or KRAS-MT. Multivariate analysis showed the presence of a KRAS mutation to be an independent prognostic factor for EFS (hazard ratio [HR], 2.21; 95% confidence interval [CI], 1.35-3.59; P = .002) and OS (HR, 1.85; 95% CI, 1.01-3.31; P = .045) in MLL-r AML. The mutation is a distinct adverse prognostic factor in MLL-r AML, regardless of risk subgroup, and is potentially useful for accurate treatment stratification. This trial was registered at the UMIN (University Hospital Medical Information Network) Clinical Trials Registry (UMIN-CTR; http://www.umin.ac.jp/ctr/index.htm) as #UMIN000000511.

Expression of mitochondrial genes predicts survival in pediatric acute myeloid leukemia

Deregulated mitochondrial metabolism and biogenesis have been studied in acute myeloid leukemia (AML); yet, the relevance of mitochondrial-encoded gene expression on AML outcomes is unknown. This study was conducted to assess clinical significance of expression of mitochondrial-encoded genes, namely ND3, SDHB, Cytochrome b, Cytochrome C, and ATP6, in pediatric AML. Pediatric AML patients from July 2013 to June 2016 were enrolled in this prospective study. Relative genes expression was determined using real-time PCR, and expressed as fold change. 123 AML patients were enrolled, median age 10 (range 0.7-18years). ND3 gene expression was significantly increased in poor-risk cytogenetics (P = 0.03). In univariate analysis, high ND3 and ATP6 gene expression was significantly associated with inferior EFS (P = 0.01 and P = 0.04, respectively) and OS (P = 0.02 and P = 0.01, respectively), whereas, in multivariate analysis, ND3 gene expression emerged as the only independent prognostic factor for EFS and OS (P = 0.04 and P = 0.03). ND3 gene expression is a significant predictor of EFS and OS in pediatric AML, and should be evaluated as a potential biomarker.

NPM1 High Mutant Allele Burden is an Adverse Prognostic Factor for AML Patients with Mutated NPM1

To investigate the clinical features, accompanying gene mutation characteristics and prognostic factors of adult patients with acute myeloid leukemia with mutated NPM1 (NPM1+AML). Seventy-three patients with newly diagnosed adult NPM1+AML were selected. The mutations of 22 genes were detected by second generation sequencing and 43 fusion genes of AML were detected by real-time fluorescent quantitative PCR. The Kaplan-Meier survival curve and Cox multivariate regression analysis were used to study the prognostic factors. A total of 74 NPM1 site mutations were detected in 73 patients with NPM1+AML. The incidence rates were 92.0% L287fs, 2.7% Q289fs and W288fs, 1.4% L258fs and Q289H, among which 1 patient had 2 NPM1 mutations; the different mutation sites had no effect on the prognosis of NPM1+AML. The median value of NPM1 variant allele frequency (VAF) was 35.4% (1.8%-56.6%). Based on the uppermost quartile of 38.4%, the patients were classified as NPM1 VAF>38.4% (NPM1highAML) and NPM1 VAF≤38.4% (NPM1lowAML). Compared with NPM1lowAML, the early mortality rate was statistically significantly higher (33.3% vs 7.3%, P<0.05), and median EFS (148 d，95%CI 58-238 d vs 372 d，95%CI 264-480 d) (P<0.01) and median OS (179 d 95%CI 6-352 d vs 444 d) (P<0.01) were significantly shorter in NPM1high AML. A total of 126 accompanying gene mutation sites were detected in 87.7% of patients with NPM1+AML. The patients with NRAS gene mutation displayed a higher rate of complete remission (100% vs 58%) (P<0.05) and longer median OS (not reached to 320 d, 95%CI 150-490 d) (P<0.05). The 43 fusion genes were examined in 65 out of 73 cases of NPM1+AML, and in all the patients the fusion gene test was negative. Multivariate analysis showed that NPM1 VAF>38.4% was an independent prognostic factor for EFS (HR=3.1, 95% CI 1.6-6.4, P<0.01) and OS (HR=3.0, 95% CI 1.4-6.2, P<0.01). The NPM1 gene mutation in AML patients often is accompanied by other gene mutations, while the coexistence of fusion genes is rare; high NPM1 mutant allele burden is an independent prognostic factor for adult AML patients with mutated NPM1.

Coexpression of CD5 and CD43 predicts worse prognosis in diffuse large B-cell lymphoma.

Both CD5 and CD43 are expressed on the surface of B lymphocytes of definite phase and associated with the adverse outcome in diffuse large B‐cell lymphoma (DLBCL). However, the relationship between CD5 and CD43 expression and the prognostic value of CD5/CD43 coexpression in DLBCL are unknown. We herein determined the correlation between CD5 and CD43 expression, as separate factors or in combination, with the clinicopathological features and survival of 200 patients with DLBCL receiving standard chemotherapy with or without rituximab. Among these DLBCL patients, CD5 expression, CD43 expression, and CD5/CD43 coexpression were detected in 18 (9%), 57 (27%), and 10 (5%) patients, respectively, and all were positively correlated with advanced age and nongerminal cell type. CD5‐positive and CD43‐positive DLBCL patients had poorer event‐free survival (EFS, P < 0.001) and overall survival (OS, P < 0.001) than CD5‐negative and CD43‐negative patients, respectively. CD5/CD43 coexpression was correlated with a significantly worse prognosis than CD5 or CD43 expression alone. Univariate analysis showed that CD5 expression, CD43 expression, and CD5/CD43 coexpression were all adverse prognostic factors for DLBCL patient survival, and CD5/CD43 coexpression was associated with a greater relative risk for recurrence and death than either CD5 or CD43 expression alone. Multivariate analysis demonstrated that CD5/CD43 coexpression was an independent prognostic factor for EFS (P < 0.001) and OS (P < 0.001) in DLBCL. In conclusion, our data indicate that DLBCL patients with CD5/CD43 coexpression represent a specific subgroup with a significantly worse prognosis than those expressing either marker alone.

ERCC1 and ERCC2 as predictive biomarkers to oxaliplatin-based chemotherapy in colorectal cancer patients from Egypt

BackgroundThe impact of Excision repair cross-complementation group 1 (ERCC1) and group 2 (ERCC2) expression levels on the efficacy of oxaliplatin-based chemotherapy is still controversial. The present study was conducted to determine the predictive value of these molecular biomarkers in stage III and IV colorectal cancer (CRC) patients receiving oxaliplatin (OX)-based chemotherapy as first-line treatment. MethodsThe study included 80 CRC patients who received first line oxaliplatin based chemotherapy The expression levels of ERCC1 and ERCC2-mRNA and proteins were determined in the primary tumors by quantitative real time reverse transcription polymerase chain reaction(RT-qPCR) and immunohistochemistry (IHC); respectively. The results of mRNA expression were correlated with patients' characteristics, response to treatment, overall- and event free survival (OS & EFS). ResultsSixty four out of the 80 patients were legible for assessment of ERCC1 and ERCC2 expression. The cut-off levels of ERCC1and ERCC2-RNA were 3.8×10−3& 4.6×10−3; respectively. Reduced ERCC1 and ERCC2 RNA expressions were detected in 50 (78.1%) and 48 (75%) cases, respectively whereas reduced proteins were detected in 48 cases (75%) for ERCC1 and ERCC2. After The median follow up period was 30.5months (range: 7–104months), Patients with low mRNAERCC1levels showed significantly longer OS (p=0.011) and EFS (p˂0.001). However, no significant relation was found between ERCC2 levels and OS or EFS. In multivariate analysis performance status (PS), stage of the disease and ERCC1-mRNA expression were independent prognostic factors for EFS whereas tumor histology and stage of the disease were independent factors for OS. ConclusionsERCC1 expression levels may help in selecting patients who benefit from oxaliplatin chemotherapy in stage III & IV CRC. Further large trials are needed to validate these data.

Prognostic Impact of ABCA3 Expression in Pediatric Acute Myeloid Leukemia

Abstract Background. Despite progress in the molecular and genetic classification of pediatric acute myeloid leukemia (AML), the prognosis remains heterogeneous. The ATP-binding cassette transporter A3 (ABCA3) seems specifically involved in the resistance of pediatric AML to intensive chemotherapy. However, studies having investigated the prognostic impact of ABCA3 expression have yielded conflicting results with respect to patient outcomes while the small sample size of these studies precluded the use of multivariate analysis. Here we investigated the prognostic impact of ABCA3 expression in a representative series of homogeneously treated pediatric AML. Methods. Samples derived from 233 patients with available high-quality RNA and enrolled in the ELAM2 protocol (NCT00149162). qRTPCR amplification of 2 conserved ABCA3 mRNA sequences was performed with GUS and ABL as reference genes. Primer sets were complementary to exons 6-7 and exons 19-20 junctions. Patients were classified according to their standardized cytogenetic and molecular (NPM1 mutations, FLT3-ITD, CEBPA double mutations) risk subgroups (Rubnitz JE, Blood 2012;119:5980-5988, Creutzig U, Blood 2012;120:3187-3205). Treatment consisted of 1 induction course (AraC and mitoxantrone) and 3 consolidation courses (course 1 and 3 with high dose AraC); all children with either intermediate or high-risk disease were candidates for hematopoietic stem cell transplant (HSCT) in complete remission (CR) after 1 to 2 consolidation courses. Results. The discovery cohort included 120 patients. Median age, median WBC, CR rate, relapse rate, median follow-up, 5-years EFS, DFS, and OS were 9.4 years, 19.3 G/L, 95%, 29%, 60 months, 58±6%, 61±6%, and 71±5 months, respectively. The two primer sets yielded consistent results (R=0.9, p&lt;10-4, Spearman Rank Correlation). Lower ABCA3 expression was positively associated with CBFB-MYH11 AML (p=0.002) and thereby with favorable cytogenetics (p=0.036) and low-risk AML (p=0.027). Higher ABCA3 expression was associated with higher relapse rate (p=0.006), shorter EFS (5-years, 34±9 vs 61±6 % p=0.0005), DFS (36±9 vs 62±6% p=0.0028), and OS (49±12 vs 79.5±5% p=0.0007). Multivariate analyses identified age, WBC, risk group, and ABCA3 expression as independent prognostic factors for EFS, DFS, and OS (Table 1). The validation cohort included 113 patients in whom the proportions of AML1-ETO- and MLL-positive AML were significantly higher than in the discovery cohort: 26,5% vs 6,7% (p&lt;10-4) and 24.8 vs 14.2% (p=0.03). There was no significant difference in patients' outcome between the 2 cohorts. Using the same cutoff value in the validation cohort, higher ABCA3 expression remained significantly associated with shorter 5-years EFS: 63±7% vs 43±9% (p=0.025) with a trend for shorter DFS: 45±9 vs 53±11% (p=0.065). Multivariate analyses identified ABCA3 expression as an independent negative prognostic factor for EFS and DFS (Table 1). In the entire patients population, ABCA3 expression independently predicted EFS, DFS, and OS (not shown). In the low- (n=74) and adverse-risk (n=59) groups, higher ABCA3 expression remained associated with shorter 5-years EFS (low: 46±12 vs 75±7%, p=0.006; adverse: 12±10 vs 44±16%, p=0.018), DFS (low: 49±13 vs 75±7%; high: 12±11 vs 45±16%, p=0.016), and OS (low: 76±10 vs 94±4%; adverse: 32±14 vs 57±18%, p=0.046). Conclusion. ABCA3 expression represents an independent prognostic factor in pediatric AML. As they indicate that the level of ABCA3 expression is significantly associated with survival for currently accepted cytogenetic and molecular prognostic categories, our findings suggest that assessing ABCA3 expression will permit a better assessment of disease risk. Finally our results suggest that inhibiting ABCA3 expression, such as with indomethacin, could be beneficial in order to overcome drug resistance in pediatric AML. Disclosures No relevant conflicts of interest to declare.

Age and Gemtuzumab Ozogamicin Influence the Prognostic Impact of TET2 Expression and Splicing in Intensively Treated Patients with Acute Myeloid Leukemia

Background. In AML, the frequency of TET2 mutations increases with age ranging from 1-5% in children to 6-27% in adults. These mutations have both been reported to exert unfavorable or neutral effects on patient outcomes. Besides mutations, fluctuation in TET2 expression has been evidenced in AML while we recently found that TET2 exon 2 skipping (TET2E2S) is associated with favorable outcome in adult AML (Mint-Mohamed et al., ASH 2014 and submitted). Here we investigated the effect TET2 expression and TET2E2S in homogeneously treated adult and pediatric AML patients with intensive chemotherapy (IC).Methods. Samples derived from 341 patients with available high-quality RNA and enrolled in the ELAM2 protocol (n=120) and the ALFA-0701 protocol (n=221).qRTPCR amplification of 2 conserved TET2 mRNA sequences was performed with GUS and ABL as reference genes. TET2 exon 2 skipping was quantified throughexon(E)-specific qRTPCR (qESPCR) amplification of E1E3 (spliced) and E2E3 (unspliced) TET2 isoforms. TET2E2S was quantified by calculating the ratio E1E3/E2E3. Patients were classified according to their standardized cytogenetic and molecular (NPM1 mutations, FLT3-ITD, CEBPA double mutations) risk subgroups. In children, treatment consisted in 1 induction course (AraC and mitoxantrone) and 3 consolidation courses (course 1 and 3 with high dose AraC); children with either intermediate or high-risk disease were candidates for hematopoietic stem cell transplant (HSCT) in complete remission (CR) after 1 to 2 consolidation courses. Adult patients were given a 3+7 induction course without (control group, n=110) or with fractionated intravenousgemetuzumab ozogamicin (GO) (n=111) as already described (Castaigne S, Lancet 2012; 379:1508-1516).Results. In pediatric AML, median age, median WBC, CR rate, relapse rate, median follow-up, 3-years EFS, DFS, and OS were 9.4 years, 19 G/L, 95%, 29%, 48 months, 60±5%, 63±6%, and 72±5%, respectively. Lower TET2 expression was associated with inferior EFS (3-years: 44±10%vs 69±6%, log-rank p = 0.01), DFS (51±8%vs 64±5% p=0.02), and OS (60±8%vs 83±3%, p=0,0003) while TET2E2S was associated with inferior EFS (44±8%vs 70±7%, p = 0.006) and OS (61±7%vs 81±4%, p=0.01). For TET2 expression, multivariate analysis identified WBC, age, risk group, and TET2 expression as independent prognostic factors for EFS and OS, and age, WBC, and TET2 expression for DFS (Table 1). For TET2 splicing, WBC and TET2E2S independently predicted EFS while TET2E2S alone independently predicted DFS (Table 1). In the low- (n=29) and intermediate-risk (n=62) groups, lower TET2 expression remained associated with shorter 3-years EFS. There was no significant correlation between TET2 expression and TET2E2S and TET2E2S retained its negative prognostic impact in the 85 patients with higher TET2 expression (3-years EFS 48±8vs 84±6%, p=0.004). In adult AML, median age, median WBC, CR rate, relapse rate, median follow-up, 3-years EFS, DFS, and OS were 62 years, 8 G/L, 76%, 59%, 47 months, 26±3%, 30±4%, and 44±4%, respectively. In the entire population, TET2 expression and splicing had no significant effect on outcome. In the 110 patients treated with IC alone, TET2 expression was associated with inferior OS (25±9vs 29±9%, p=0.02) while TET2E2S was associated with better EFS (17±5vs 7±6%, p=0.008), DFS (28±8vs 8±6%, p=0.02), and OS (46±8vs 29±8%, p=0.05). TET2E2S, age, and disease risk represented independent prognostic factors for EFS (Table 1). In the GO arm, TET2 expression had no significant effect on outcome whereas TET2E2S was associated with inferior EFS (27±5vs 60±12%, p=0.01), DFS (31±6vs 63±12%, p=0.05), and OS (39vs 72±11%, p=0.04).Conclusion. Inchildren, higher TET2 expression confers a favorable outcome whereas TET2E2S represents an independent unfavorable prognostic factor. In adult treated with IC alone, present results confirm the favorable prognostic impact of TET2E2S and the weak prognostic effect of TET2 expression. In contrast, in patients fulfilling the same inclusion criteria, TET2E2S loses its favorable prognostic effect and is even associated with unfavorable outcome when GO is associated to IC. While TET2 expression and splicing might represent useful biomarkers, the data highlight the strong difference in tumor biology between adult and pediatric AML and suggest distinct relationships between TET2E2S and the cytotoxic effect of GO plus IC versus IC alone. [Display omitted] DisclosuresThomas:Pfizer: Consultancy.

ABCA3 Expression Predicts Response to Gemtuzumab Ozogamicin in AML

Background. The ATP binding cassette transporter 3 (ABCA3) has been recently found to induce a significant reduction in cytotoxicity following exposure to anthracyclines, mitoxantrone, etoposide, Ara-C, vincristine, and rituximab. ABCA3 acts through the modulation of multivesicular bodies (MVB) and contributes to drug sequestration in late endosomal organelles, i.e. MVB and lysosomes. Studies having investigated the prognostic impact of ABCA3 expression in AML have yielded conflicting results as ABCA3 expression has both been reported to exert unfavorable or neutral effects on patient outcomes. In addition, the small sample size of these studies precluded the use of multivariate analyses.Methods. Our goal was to investigate the prognostic impact of ABCA3 expression in adult patients with AML treated with IC with or without gemtuzumab ozogamicin (GO). To this end we investigated the relationship between ABCA3 expression and EFS in a representative series of 221 AML homogeneously treated in the ALFA-0701 trial. qRTPCR amplification of conserved ABCA3 mRNA sequences, as identified with FasterDB database, was performed with GUS and ABL as reference genes. Primer sets were complementary to conserved ABCA3 exons 6-7 and exon 19-20 junctions. Patients were given a 3+7 induction course without (control group, n=110) or with fractionated intravenous GO (n=111) (Castaigne S, Lancet 2012; 379:1508-1516).Results. Among the 278 randomized patients, 221 had available bone-marrow diagnostic samples with high-quality RNA. The same benefits associated with GO were observed in the 221 patients from the present study as in the entire trial population. Overall, median age, CR rate, relapse rate, median follow-up, 3-years EFS were 62.1 years, 76.5%, 66%, 47.45 months, 28±3%, respectively. There was no significant difference in the level of ABCA3 expression between responders and non-responders. In the 169 responders, ABCA3 expression at diagnosis was more than 3-fold higher in the 111 remitters who subsequently relapsed than in the 58 patients who remained in persistent CR (p=0.033). The level of ABCA3 expression was significantly lower in ELN favorable group than in intermediate and adverse risk AML (p= 0.004) and negatively correlated with CD33 expression (R=-0.272, p<10-4). Through univariate analysis, higher ABCA3 expression was associated with shorter EFS (3-years: 22±3 vs 45±7 % p=0.002). Multivariate analysis identified age, treatment arm, and ELN risk group as independent prognostic factors for EFS. In the control group, there was no significant association between ABCA3 expression and CR rate, relapse rate, and EFS. In the 111 patients within the GO arm, there was no significant difference in the level of ABCA3 expression between responders and non-responder whereas in the 89 responders, ABCA3 expression at diagnosis was more than 7-fold higher in the 53 remitters who subsequently relapsed than in the 36 patients who remained in persistent CR (p=0.006). Through univariate analysis, higher ABCA3 expression was associated with shorter EFS (3-years: 22±5 vs 64±9 % p=0.0002). Multivariate analysis identified ABCA3 expression, cytogenetics, CD33 expression, and ECOG as independent prognostic factors for EFS (Figure 1).Conclusion. WhileABCB1 has been previously found to attenuate GO-induced cytotoxicity in AML cells (Walter RB, Blood 2003; 102:1466-1473), present results indicate that higher ABCA3 expression independently predicts poor outcome in AML patients treated with fractionated GO and intensive chemotherapy (IC). GO is an anti-CD33 antibody carrying a toxic calicheamicin derivative that, after hydrolytic release within lysosomal vesicles, induces DNA strand breaks, apoptosis, and cell death. Whether the clinical effect of ABCA3 expression relies on the modulation of CD33 internalization, calicheamicin release or combination thereof is under investigation. Finally our results encourage inhibiting ABCA3, such as with indomethacin, in order to overcome drug resistance in AML treated with GO-IC. [Display omitted] DisclosuresThomas:Pfizer: Consultancy.

A Transcriptional- and Posttranscriptional-Based Prognostic Model in Pediatric Acute Myeloid Leukemia

Background. We have recently investigated transcriptional and posttranscriptional changes in AML cells (Oncotarget, 2016, 7, 2889-909) and identified ABCA3 expression, TET2 expression and TET2 exon 2 skipping (TET2E2S) as prognostic factors in pediatric AML (See companion abstracts from Ceraulo et al.). The present study was conducted in order to test whether the combination of these prognostic factors might help stratifying patients according to their relapse risk.Methods. Samples derived from 120 patients with available high-quality RNA and enrolled in the French ELAM2 protocol. Patients were classified according to their standardized cytogenetic and molecular (NPM1 mutations, FLT3-ITD, CEBPA double mutations) risk subgroups. Treatment consisted of 1 induction course (AraC and mitoxantrone) and 3 consolidation courses (course 1 and 3 with high dose AraC); all children with either intermediate or high-risk disease were candidates for hematopoietic stem cell transplant in complete remission (CR) after 1 to 2 consolidation courses. qRTPCR amplification of 2 conserved ABCA3 (exons 6-7 and exon 19-20) and TET2 (exon 4 and exon 2-3) mRNA sequences were performed with GUS and ABL as reference genes. TET2E2S was quantified as already described (Oncotarget, 2016, 7, 2889-909). The concordance index (c-index) was used to compare the relative statistical power of the new model compared with the standardized cytogenetic and molecular-based scoring system.Results. ABCA3 expression [HR 2.6 (95% CI 1.3-5), p=0.006], TET2 expression (HR 0.5 (0.2-0.9) p=0.04, and TET2E2S (HR 2.5 (1.2-5.1), p=0.01) represented independent prognostic factors for EFS. To design a convenient prognostic score, continuous variables were dichotomized using cutoff points that were based using the Maximally Selected Rank Statistics (maxstat R-statistics package, from the R-Project). One point was attributed for lower TET2 expression (n=85), higher ABCA3 expression (n=44), and TET2E2S (n=61). Three groups were formed: lower-risk patients (0 point, n=33), intermediate-risk patients (1 point, n=41), and higher-risk patients (either 2 or 3 points, n=46). The present score identified subgroups of patients with significantly different outcome (p<10-4) (Figure 1). Patients with a low TPS (Transcriptional-Posttranscriptional Score) had significantly better EFS than either patients with a high (p<10-4) or an intermediate TPS (p=0.03) while patients with intermediate TPS had significantly better EFS than patients with high TPS (p=0.02). For OS (p=0.0004), patients with a low TPS had significantly better OS than either patients with a high (p=0.0004) or an intermediate TPS (p=0.047) while patients with intermediate TPS had significantly better OS than patients with high TPS (p=0.024). For DFS (p=0.003), patients with a low TPS had significantly better DFS than either patients with a high (p=0.0012) or an intermediate TPS (p=0.036) whereas no significant difference was noticed between patients with intermediate versus high TPS (p=0.2). Figure 1 shows that FLT3-ITD was significantly associated with intermediate TPS, EVI1 expression with high TPS and NPM1 mutations and CBFB-MyH11 leukemias with low TPS. The distribution of CR rate and relapse rate was significantly distinct among the 3 TPS categories (Figure 1). The added value of the TPS within AML genetic risk groups was evaluated through univariate analysis. The TPS permitted to identify patients with different outcome in lower and intermediate-risk AML (EFS: Log-rank, p=0.011 and 0.005, respectively). Ten of the 20 lower-risk AML corresponded to intermediate (n=7) and high (n=3) TPS. Their CR rate was 100%, none of these patients was allografted and 4 patients relapsed as compared to 0/10 in the remaining lower-risk AML cases with low TPS (p=0.02). In intermediate risk AML, the CR and relapse rates of patients with low, intermediate and high TPS were 94 and 17%, 100 and 45%, and 73 and 50 %, (CR, p=0.001; relapse, p=0.036). Overall, the present TPS model had a better c-index (0.73) compared with the standardized cytogenetic and molecular-based scoring system (0.61).Conclusion. We propose a new prognostic model that includes transcriptional and posttranscriptional data and can predict survival in pediatric AML. A validation in a larger, independent, multicenter, data set of patients is important to facilitate the translation of this model into clinical practice. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Expression of CD30 in Diffuse Large B Cell Lymphoma and Its Clinical Significance

To evaluate the expression and clinical significance of tumor necrosis factor receptor (TNFR) superfamily protein CD30 in diffuse large B cell lymhoma (DLBCL). The CD30 expression, clinical characteristics and prognosis of 63 patients with DLBCL, NOS out of 149 patients with DLBCL admitted in our hospital between January 2008 and December 2012 were analyzed retrospectively. no significant relationship existed between CD30 expression and clinical features, such as age, sex, B symptoms, staging, ECOG PS, LDH level, extranodal site involvement, IPI, GCB or non GCB type, bone marrow involvement. By univariate analysis, the clinical factors associated with general OS and EFS, included CD30, ECOG PS, B symptoms, extranodal site involvement, LDH level, IPI, bone marrow involvement and rituximab. Univariate analysis in GCB DLBCL indicated that CD30 had no significant effect on OS and EFS. However, univariate analysis in non-GCB DLBCL indicated CD30 was associated with longer OS (P=0.037) and showed a tendency of better EFS (P=0.067). In multivariate analysis, IPI and CD30 were independent prognostic factors for OS (IPI: P=0.000, 95%CI 0.042-0.374, CD30: P=0.044, 95%CI 1.055-60.613), and IPI also was independent prognostic factors for EFS (P=0.000, 95%CI 0.040-0.360). CD30+ and DLBCL have a tendency of better EFS (P=0.050, 95%CI 0.996-56.501). CD30 expression level correlates with the prognosis of DLBCL and has a certain clinical value, which may be a new prognostic index of DLBCL.

Skipping of ATP-Binding Cassette Transporter A3 Exon 19 in AML Cells Is an Independent Prognostic Factor in Patients with Normal Cytogenetics

ATP binding cassette (ABC) transporters are a superfamily of highly conserved membrane proteins that transport a wide variety of substrates across cell membranes and confer drug resistance against a wide range of chemotherapeutic agents. We recently found that WT1, which is regularly overexpressed in AML and interact with the splicing machinery, modifies the splicing of ABC transporters A2, A3, A5, and C2. For ABCA3, WT1 knock-down in three AML cell line coupled with Affymetrix HTA2 exon arrays analysis confirmed by exon-specific PCR revealed that WT1 influences the skipping of exon 19. ABCA3 belongs in the ABC subclass and induces a significant reduction in cytotoxicity observed following exposure to DNR, mitoxantrone, etoposide, Ara-C and vincristine. The ABCA3 domain encoded by exon 19 (amino acid 805-847) is localized at the junction of the first nucleotide-binding domain and the second transmembrane domain, and is involved in ATP hydrolysis. In silico, skipping of exon 19 deletes a sequence of 32 amino acids rich in positively charged residues and is thereby assumed to increase drug efflux through increased ATP hydrolysis. The effects of the skipping of exon 19 on chemoresistance and DNR efflux are currently investigated while for the present study, we hypothesized that skipping of exon 19 of ABCA3 might negatively influence outcome in AML patients.Analyzing 132 bone marrow AML samples harvested at diagnostic confirmed the statistically significant correlation between WT1 expression and ABCA3 splicing in vivo (p<10-4, Spearman Rank Correlation). This correlation was specific because it was not observed in 37 control samples deriving from bone marrow donors or in the AML cases with 2 TET2 alternative exon usages found WT1-independent ex vivo. In the 106 patients treated with intensive chemotherapy (IC), skipping of ABCA3 exon 19 was associated with a significantly lower response rate: 39/55 (70.9%) vs 44/51 (86.3%, p = 0.045, Pearson’s chi-squared test). In complete remitters, skipping of ABCA3 exon 19 was associated with a significantly higher relapse rate: 77.1% vs 52.6% (p=0.026, Pearson’s chi-squared test). Median follow-up was 25 months. The 25 allografted patients were censored at the time of allograft. By univariate analysis ABC-A3 exon skipping of exon 19 significantly affected OS (HR=4.50 (95% confidence interval: 2.10-9.63), p<10-4), DFS (HR=3.76 (95% confidence interval: 1.87-7.42), p<10-4), and EFS (HR 3.73 (95% confidence interval: 1.38-5.13), p=0.004); higher the level of exon 19 skipping, poorer the outcome. In multivariate analysis, age, cytogenetic, and ABC A3 exon 19 skipping were identified to be independent prognostic factors for OS, EFS and DFS. Age and ABC-A3 exon exclusion were identified to be independent prognostic factor for OS in the 49 patients with normal karyotype.In order to confirm these results we investigated the prognostic impact of ABC A3 exon 19 skipping in a validation cohort of 108 additional AML case with normal cytogenetics. FLT3 internal tandem duplication (FLT3-ITD) and nucleophosmin (NPM1) exon-12 gene mutation were identified in 37 (34.3%) and 66 cases (61.1%). In the 86 patients treated with IC, the CR rate was 89,5% without significant difference between patients with or without exon 19 skipping. The relapse rate was higher in cases with exon 19 skipping (47,1% vs 23.5%) but the difference was not statistically significant. The 29 allografted patients were censored at the time of allograft. Median follow-up was 12 months. By univariate analysis ABC-A3 exon skipping of exon 19 significantly affected DFS (HR=2.03 (95% confidence interval: 1.22-3.39), p=0.007, and EFS (HR 1.62 (95% confidence interval: 1.06-2.48), p=0.027); higher the level of exon 19 skipping, poorer the outcome. In multivariate analysis, age, FLT3-ITD, ABC A3 exon 19 skipping but not NPM1 mutation were identified to be independent prognostic factors for EFS while FLT3-ITD and ABC A3 exon 19 skipping were independent prognostic factors for DFS.In conclusion ABC-A3 missplicing possesses a strong prognostic impact in AML indicating that besides whole gene transcription, quantitative analysis of alternative splicing might represent a promising tool for assessing AML aggressiveness at the time of diagnostic in patients with normal or abnormal karyotype. DisclosuresNicolini:Novartis: Consultancy.

Dose-Intensity Impacts On Survival of Adolescents and Young Adults with Acute Lymphoblastic Leukemia Treated in Adult Departments by a Pediatric Protocol (FRALLE 2000BT)

AbstractAbstract ▪3561▪This icon denotes a clinically relevant abstract Background:Adolescents (15–19y) with acute lymphoblastic leukemia (ALL) markedly benefit from pediatric rather than adult chemotherapy regimens, as highlighted by many retrospective studies (Boissel et al., JCO 2001). In young adults with ALL, there are now strong evidences that pediatric or pediatric-inspired protocols may also improve long-term outcome (Huguet et al., JCO 2009). However, whether care environment (adult vs pediatric unit) may impact this outcome remains an open issue. To address this question, we explored the outcome of adolescents and young adults (AYA, 15–29y) treated in adult hematology departments by the pediatric FRALLE 2000-BT protocol. Methods:From February 2001 to June 2011, 89 AYAs with Ph-negative ALL including 60 adolescents (15–19y) and 29 YAs (20–29y) were treated according to the pediatric FRALLE 2000-BT protocol in 13 French and Belgian adult hematology units (median follow-up, 5 years). After a prednisone prephase and a four-drug induction (prednisone, daunorubicin, vincristine and L-asparaginase), patients achieving CR received four 8-week phases of treatment: consolidation, 1st delayed intensification (DI), interphase, 2nd DI, and maintenance. According to FRALLE protocol recommendations, each phase should begin after hematological recovery of previous phase (ANC>1 G/L, Platelets>100 G/L). Twenty-five patients (18 adolescents and 7 YAs) underwent allogeneic transplantation after consolidation phase or 1stDI. Results:Apart age (median age, 18 vs 23y; p<.0001), adolescent and YAs cohorts had similar characteristics, including WBC, B/T lineage distribution, and high-risk karyotype (t(4;11)/MLL-AF4, hypodiploidy)), except for initial CNS involvement (1/60 vs 4/29 respectively, p=.02). Initial peripheral response to prednisone (PDN) at D7 and bone marrow response to chemotherapy at D21 were better in the adolescent subgroup, without reaching significance (slow PDN-responders: 30% vs 38%, p=.42; slow chemo-responders: 13% vs 21%, p=.37). Overall CR rate was 99% and did not differ between both cohorts (98% in adolescents, 100% in YAs). 5y-OS and 5y-EFS were estimated respectively at 66% and 61%. Despite similar CR rates, 5-year EFS was significantly shorter in adolescents than in YAs (47% vs 79%, p=.02). A non significant shorter 5y-OS was also observed in adolescents (58% vs 81%, p=.11). To explain this difference, we looked at dose-intensity disparities between both subgroups. During induction phase, no differences in drug cumulative doses were observed. To further explore potent difficulties to respect protocol schedules in younger patients, we compared intervals from induction D1 to start of different trial phases between both subgroups. In comparison to YAs, intervals from induction D1 to consolidation, 1st DI, 2nd DI and maintenance phases were progressively delayed in adolescents: 4 days for consolidation (p=.24), 6 days for 1st DI (p=.19), 9 days for 2nd DI (p=.005), and finally 25 days for maintenance phase (p=.0002). In univariate analysis, among age (adolescents vs YAs), B/T lineage, WBC, cytogenetics, and time to 1st DI, only age and time to 1st DI significantly affected EFS (p=.05 and p=.001 respectively). In bivariate analysis, age and time to 1st DI were identified as independent prognostic factors for EFS (p=.03 and p=.004, respectively). The same observation was done with interval time to 2ndDI. Conclusion:The outcome of AYAs treated in adult units with the pediatric FRALLE 2000 protocol is encouraging, particularly in YAs. The poorer outcome observed in adolescents may be related to increasing intervals from initial induction to the different phases of treatment. Prospective trials are required to register individual reasons that lead physicians and patients to progressively delay therapy. This study suggests that trial itself is not sufficient to improve the outcome in adolescents with cancer and that AYA programs are needed to explore the difficulties encountered by patients and care teams to adhere to therapy. Disclosures:No relevant conflicts of interest to declare.

PROGNOSTIC SIGNIFICANCE of CD68 EXPRESSION for Korean PATIENTS with HODGKIN'S LYMPHOMA

AbstractAbstract 3888 Background:A lower incidence of Hodgkin's lymphoma (HL) in Asians has been recognized and the incidence of HL in Korea was reported to be 5.3%. This low incidence of HL in Asian population has hindered the evaluation of pathogenesis and prognostic factors of the disease. Although international prognostic score (IPS) has been largely utilized as prognostic stratification tool, there has been unmet need to further clarify those with poor prognosis until an increased number of tumor-associated macrophages was identified as a predictor of poor clinical outcome. Hence, we evaluated the prognostic significance of CD68, a marker of macrophages, in Korean HL patients. Methods:We performed immunohistochemical analysis of CD68 in 144 classic HL patients treated with ABVD (n=113, 78.5%), MOPP (n=10, 6.9%), ABVD/MOPP hybrid (n=15, 10.4%) or BEACOPP (n=6, 4.2%) chemotherapy with or without radiotherapy between November 1990 and December 2009 in the Asan Medical Center. The relative percentage of CD68+ cells in relation to overall cellularity was calculated and the results were correlated with clinical outcome. Results:We examined various cutoff points of CD68 expression from 10 to 90 percentile with a rising gradient constructed using 5% steps (5%, 10%, 15%, 20%, 25%, 30%, 35% and 40%). The most significant statistical difference in disease-specific survival (DSS) was observed at a cutoff value of 20%, employing the log-rank test. The high (>=20%, n=78) CD68 group included more patients with older patients (Age 45 yr, 45.5% vs. 28.2%, p=0.032) and higher IPS (>=4, 37.9% vs. 21.8%, p=0.034) compared with the low (<20%, n=66) CD68 group. In total, 18 patients in the low CD68 group and 20 patients in the high CD68 group experienced relapse or progression. Nine patients in the low CD68 group (6 patients of progressive disease [PD], 1 of treatment-related infection during salvage treatment, 1 of moyamoya disease and 1 of unknown cause) and 15 patients in the high CD68 group (8 of PD, 5 of treatment-related infection, 1 of intraventricular hemorrhage during primary therapy, 1 of unknown cause) died by the time of data cutoff. Treatment-related mortality (TRM) was significantly higher in the high CD68 group (n=1 vs. n=6, p=0.048). The 5-year EFS rates were 74.7% and 49.5% in the low and high CD68 expression groups, respectively (P=0.009) with a median follow-up period of 5.4 years (range, 0.6–19.0 years) in surviving patients. The 5-year DSS rates were 95.7% in the low CD68 expression group and 76.5% in the high CD68 expression group (P=0.011). In the multivariate analysis with clinical variables significantly correlating with EFS/DSS in univariate analyses including IPS (>=4), age (>=45 years) and presence of B symptoms, CD68 expression was found to be an independent prognostic factor for EFS (Hazard ratio [HR] =1.846; 95% confidence interval [CI] 1.106–3.354; P=0.044) and DSS (HR = 2.955; 95% CI, 1.148–7.607; p=0.025). However, the prognostic significance of CD68 seems to be more prominent in patients with localized disease (n=48) in a subgroup analysis. While 5-year EFS (85.8% vs. 25.7%, p=0.001) and DSS (95.7% vs. 78.8%, p=0.007) for patients with localized disease were significantly higher in the low CD68 group, both EFS (66.5% vs. 56.5%, p=0.144) and DSS (92.7% vs. 75.7%, p=0.144) were not significantly different between the high and low CD68 groups. Conclusion:The number of CD68+ macrophages is a significant prognostic factor in Korean HL patients. [Display omitted] [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Autologous Stem Cell Transplantation (ASCT) in CLL. Results of a Phase III Randomized Multicenter Trial.

Abstract 878 Introduction:We investigated, in a prospective randomized study, the place of ASCT in the frontline treatment of CLL. Patients and methods:From March 2001 to December 2007, 241 patients were included in the trial. Eligibility criteria were previously untreated stage B and C CLL patients under 66 years, characterized by Matutes score 4-5, absence of cyclin D1 expression, baseline flow assessment of ZAP 70 and CD38 expression, karyotype and FISH analysis, IgHv mutational status (centralized). Preceding randomization, initial chemotherapy consisted of 3 monthly courses of mini CHOP regimen as previously described, followed by 3 monthly courses of fludarabine, IV or oral. Patients achieving CR (NCI 1996 criteria plus normal CT scan) were randomized between observation and ASCT. Non CR patients were offered cisplatin/cytosine-arabinoside/dexamethasone (DHAP) rescue and randomized whatever the response between ASCT or 3 subsequent monthly IV courses of Fludarabine-Cyclophosphamide (FC). Conditioning regimen for ASCT consisted of cyclophosphamide IV (60 mg/sqm d-5-4) and fractionated total body irradiation (10 Gy). The primary end-point of the study was event-free survival at 3 years. Responses after initial treatment (i.e.before randomization) and after completion of therapy, overall survival, side effects, prognostic significance of clinical and biological characteristics at baseline were other endpoints. Results:Baseline characteristics were: gender (M/F: 3), age (median 56.4 years, range 33.3-66), stage B (185 patients) or C (56 patients). All enrolled cases but five (236 patients) started the treatment. Among them, 206 completed the six planned courses of initial chemotherapy. For the 236 patients, CR rate was 43.6%, and overall response was 89.8%. Forty two patients were not randomized because of treatment failure (19) or patient/physician decisions (23). After an observed median follow-up of 40.2 months (Q1-Q3, 17.9-47.9) at the reference date (1/1/2009), the overall survival for the 241 patients was 87.8% (95% CI, 83.3-92.6) at 3 years and 75.4% (95% CI, 66.2-88.6) at 5 years. For the 199 randomized patients, the overall survival was 90.9% (95% CI, 86.7-95.3) at 3 years. Among them 105 patients were in CR after initial treament and were allocated to ASCT (53 patients) or observation (52 patients) with a 3 years EFS of respectively 78.7% (95% CI, 67.7-91.4) and 31.3% (95% CI, 20.1-48.8) (p<0.00001). The median EFS from the start of the studied treatment was 26.2 months in the observation arm (not reached in the ASCT arm). Randomization (p=0.0001), mutational status (p<0.00001) and 11q deletion (p=0.001) remained independent prognostic factors for EFS in a multivariate analysis. There was no difference between ASCT and observation arms in terms of overall survival with 98% (95% CI, 94.3-100) and 97.5% (95% CI, 92.2-100)respectively. For the 94 patients not achieving CR and rescued with DHAP, the EFS at 3 years was 46.5% (95% CI, 33.6-66.3) in the ASCT arm (46 patients) and 43.2% (95% CI, 29.8-62.5) in the FC arm (48 patients). There was not statistical difference between these 2 arms but for the 34 patients actually autografted the EFS was 57.8% (95% CI, 41.7-80.2). The overall survival was not different for ASCT and FC arms: 81,2% (95% CI, 70.3-93.9) and 85,4% (95% CI, 75.1-97) respectively. In multivariate analysis, the only prognostic factor influencing the EFS (p=0.0001) and the overall survival (p=0.04) was the 17p deletion. The mutational status (non mutated) and the 17p deletion remained adverse prognostic factors in patients actually autografted. For the whole study, 28 patients allocated to ASCT did not receive this treatment mainly because of mobilisation failures (14 pts). Four MDS were recorded within the follow-up frame. Conclusions:ASCT is a safe procedure which significantly improves response duration in patients attaining CR after a first line treatment. For patients not in CR, ASCT or consolidation with 3 FC courses provide similar results on response duration but ASCT could be considered in patients without 17p deletion or with mutated IGHV gene. Disclosures:Van Den Neste:Roche: Research Funding.

Prognostic scoring model based on multi-drug resistance status and cytogenetics in adult patients with acute myeloid leukemia

Clinical heterogeneicity exists within an acute myeloid leukemia (AML) patient group with the same cytogenetic risk. Multi-drug resistance (MDR) is also regarded as one of the potential prognostic factors for AML. Accordingly, the prognostic scoring model can be generated based on both consideration of cytogenetic risk and the MDR status for AML. The CR rate, event-free (EFS) and overall survival (OS) were analysed according to cytogenetic risk, MDR status and clinical factors. Prognostic score was calculated by the sum of MDR status (0 for negative, 1 for positive) and dichotomized scoring for cytogenetic risk (0 for favorable/intermediate and 1 for unfavorable cytogenetics). MDR expression was noted in 36.6% of the patients and associated with a lower CR rate (p = 0.037). MDR, cytogenetics and the use of SCT were identified as independent prognostic factors for EFS and OS. The CR rate of the group scored with 0, 1 and 2 was 81.4, 66.7, and 44.4%, respectively (p = 0.050). The prognostic scoring model depicted a discriminating role in terms of EFS (p < 0.0001) and OS (p = 0.0001). The prognostic scoring model based on cytogenetic risk and MDR provided an improved method for evaluating the prognosis in AML and helped to stratify the risk of patients with the same cytogenetic risk.