pre-T Acute Lymphoblastic Leukemia Research Articles

Epigenetically dysregulated NOTCH-Delta-HES signaling cascade can serve as a subtype classifier for acute lymphoblastic leukemia.

The NOTCH-Delta-HES signaling cascade is regarded as a double-edged sword owing to its dual tumor-suppressor and oncogenic roles, in different cellular environments. In the T-cells, it supports leukemogenesis by promoting differentiation while in B-cells, it controls leukemogenesis by inhibiting early differentiation/inducing growth arrest in the lead to apoptosis. The present study was undertaken to assess if this bi-faceted behavior of NOTCH family can be exploited as a diagnostic biomarker or subtype classifier of acute lymphoblastic leukemia (ALL). In this pursuit, expression of seven NOTCH cascade genes was analyzed in bone marrow (BM) biopsy and blood plasma (BP) of pediatric ALL patients using quantitative PCR (qPCR). Further, promoter DNA methylation status of the differentially expressed genes (DEGs) was assessed by methylation-specific qMSP and validated through bisulphite amplicon sequencing. Whereas hypermethylation of JAG1, DLL1, and HES-2, HES-4, and HES-5 was observed in all patients, NOTCH3 was found hypermethylated specifically in Pre-B ALL cases while DLL4 in Pre-T ALL cases. Aberrant DNA methylation strongly correlated with downregulated gene expression, which restored at complete remission stage as observed in "follow-up/post-treatment" subjects. The subtype-specific ROC curve analysis and Kaplan-Meier survival analysis predicted a clinically applicable diagnostic and prognostic potential of the panel. Moreover, the logistic regression model (Pre-B vs Pre-T ALL) was found to be the best-fitted model (McFadden's R2 = 0.28, F1 measure = 0.99). Whether analyzed in BM-aspirates or blood plasma, the NOTCH epigenetic signatures displayed comparable results (p < 0.001), advocating the potential of NOTCH-Delta-HES cascade, as a subtype classifier, in minimally invasive diagnosis of ALL.

Acute complications observed during intensive chemotherapy in pediatric patients with acute lymphoblastic leukemia: Single-center experience.

In childhood acute lymphoblastic leukemia (ALL), very promising results were obtained thanks to the developments in treatment strategies in recent years. However, acute complications during treatment continue to be the important causes of mortality and morbidity. In this study, acute complications that develop during the treatment of ALL in childhood were evaluated. Medical records of 47 patients treated according to (ALL Intercontinental Berlin-Frankfurt-Münster) 2009 protocol between 2016 and 2021 were evaluated retrospectively. Of 47 patients, 28 (59.6%) were male and 19 (40.4%) were female. The mean age at diagnosis was 5.9±4.2 years. Forty-four patients (93.6%) were pre-B cell ALL, 3 patients (6.4%) were pre-T cell ALL. Of 47 patients, 9 (19.1%) were high risk, 32 (68.1%) were intermediate risk, and 6 (12.8%) were standard risk. Acute complications developed in 38 patients (80.8%). Among these complications, infectious complications are the most common and these were followed by gastrointestinal complications, drug-related reactions, thrombotic, neurological, and endocrine/metabolic complications, respectively. In terms of complications that may develop, the threshold of suspicion should be kept low, and patients should be treated with the same medical team in fully equipped centers with a multidisciplinary approach. Inpatient treatment strategies should be applied especially in the early stages of treatment. The importance of inpatient treatment strategy, especially in the early stages of treatment, is emphasized.

Akut Lenfoblastik Lösemide Flow Sitometri ile İmmünofenotiplemenin Prognoz Üzerine Etkileri

Background: The identification of immunophenotype subgroups is very important for the diagnosis and prognosis of acute lymphoblastic leukemia(ALL). Material and Methods: The study included 105 children with ALL(65 males, 40 females; mean age 5.9±3.8 years) who were treated TR-ALL 2000(modified) BFM treatment protocol. Results: The distributions of EGIL classification were pro-B ALL(n=1), common B ALL(n=46), pre-B ALL(n=40), pre-T ALL(n=8), cortical T ALL(n=6), and mature T ALL(n=4). Leukocyte≥100,000/mm³, lymphadenopathy≥2 cm, mediastinal involvement were commonly identified in T ALL group. T ALL had a poor response to chemotherapy according to 8th-day peripheral circulation blast counts and 15th-day bone marrow aspiration(BMA) blast counts. The recurrence, mortality, and death rate in the induction period of treatment were frequently detected in T ALL group. The variables that had prognostic potential, as indicated by univariate analyses, were leukocyte count, hepatomegaly, splenomegaly, and lymphadenopathy at the time of diagnosis, 8th-day steroid response, 15th-day BMA response, risk group, recurrence, and immunophenotyping. Multivariate Cox regression analysis demonstrated that only the leukocyte count(HR 2.51, p &amp;lt; 0.001) was a predictor of prognosis. Conclusion: Immunophenotyping may be effective in the diagnosis and prognosis of ALL, identification of risk groups, and in risk-based treatment planning. T ALL had a poor prognosis.

DIAGNOSTIC UTILITY OF IMMUNOHISTOCHEMISTRY IN SUBTYPING ACUTE LYMPHOBLASTIC LEUKEMIA: A 2 YEARS’ EXPERIENCE

OBJECTIVE: To determine the diagnostic importance of immunohistochemistry (IHC) in classifying acute lymphoblastic leukemia (ALL) in a tertiary care center.METHODS: This cross-sectional descriptive study was done from January-2017 to December-2018 at Rehman Medical Institute, Peshawar, Pakistan. Out of 133 cases diagnosed as precursor lymphoid leukemia, two cases were excluded due to inadequacy of the aspirate smears and 131 cases were included in the study. The immune-phenotype was detected by IHC with immune-markers viz terminal deoxynucleotidyl transferase (TdT), CD34, CD10, CD79a, CD3, myeloperoxidase (MPO), CD117 and paired box protein 1 (PAX1). RESULTS: Out of 131 cases included 99 (75%) were males and 32 (25%) were females. Mean age of the study participants was 20±16 years (range=16-65 years). Majority of the cases presented with hepatomegaly (n=113/131, 87%), followed by pallor (n=105/131, 80.1%), splenomegaly (n=89/131, 68%) and lymphadenopathy (n=82/131, 63%). Based on IHC, 114 (87.02%) cases were successfully classified to specific subtypes and 17 (13%) cases could not be assigned into any subtype. Eighty-six cases (65.7%) were of Pre-B cell ALL, 17 (13%) cases were T-cell ALL, 8 (6.1%) cases were Pre-T cell ALL while 3 (2.3%) cases were Pro-B lineage.CONCLUSION: Study concludes that majority of the patients were male and presented with hepatomegaly and pallor. IHC is effective method to sub-classify ALL into various immune-phenotypes in low resource countries where flow cytometry is unavailable. Pre-B cell ALL is common than T-cell ALL. KEY WORDS: (MeSH); (Non-MeSH); (MeSH); (MeSH), (MeSH); (MeSH); (MeSH); (MeSH)

Very long intergenic non-coding RNA transcripts and expression profiles are associated to specific childhood acute lymphoblastic leukemia subtypes.

Very long intergenic non-coding RNAs (vlincRNAs) are a novel class of long transcripts (~50 kb to 1 Mb) with cell type- or cancer-specific expression. We report the discovery and characterization of 256 vlincRNAs from a cohort of 64 primary childhood pre-B and pre-T acute lymphoblastic leukemia (cALL) samples, of which 61% are novel and specifically expressed in cALL. Validation was performed in 35 pre-B and pre-T cALL primary samples. We show that their expression is cALL immunophenotype and molecular subtype-specific and correlated with epigenetic modifications on their promoters, much like protein-coding genes. While the biological functions of these vlincRNAs are still unknown, our results suggest they could play a role in cALL etiology or progression.

Limfocytarne zapalenie opon mózgowo-rdzeniowych w przebiegu dokanałowego leczenia liposomalną cytarabiną u chorego na ostrą białaczkę limfoblastyczną

Central nervous system (CNS) involvement at diagnosis of acute lymphoblastic leukemia (ALL) is found in approximately 5% of patients and is a poor prognosis factor. Introduction of prophylactic strategies reduced involvements rate from 30% to 10% at relapse. Prophylactic regimens include: intrathecal chemotherapy (methotrexate, cytarabine [Ara-C], dexamethasone), irradiation of the whole brain, and systemic chemotherapy penetrating to CNS. Liposomal Ara-C is a prolonged-release formulation of Ara-C dedicated to intrathecal use. Its half-time is about 2 weeks. It has been used in prophylaxis and treatment regimens. Clinical trials proved its better efficacy and patients’ quality of life than its standard formulation. However, in comparison with classic triple intrathe¬cal therapy, using liposomal Ara-C may have higher risk of neurotoxicity. In the paper we report a case of 47-years-old patient with pre-T ALL who after third injection of liposomal Ara-C developed a symptomless lymphocytic meningitis with the maximal cytosis in cerebrospinal fluid of 1600 cells/μl. The review of the literature on neurotoxicity of liposomal Ara-C is hereby presented.

Safety and Efficacy of Enoxaparin Prophylaxis in Adults Receiving Peg-Asparaginase Containing Induction Regimens for Acute Lymphoblastic Leukemia (ALL)

Background:Peg-asparaginasecan cause dysregulationof pro-coagulant and anti-coagulant proteins, resulting in a significant risk of venous thromboembolism(VTE) in ALL, particularly in adults undergoing induction chemotherapy. In late 2014, we adopted a policy at our institution to use prophylactic dose ofenoxaparin 40 mg daily for up to 3 weeks following the administration of peg-asparaginase. Patients are also to undergo monitoring of complete blood count, and coagulation parameters including fibrinogen and anti-thrombin. Enoxaparin may be held if patients develop severe thrombocytopenia (platelet count <30,000/µL). Cryoprecipitate is given if fibrinogen drops below 100 mg/dl. In the present study, we aim to analyze primarily the safety, and secondarily the efficacy ofenoxaparin prophylaxis in this setting and highlight the changes in coagulation parameters following peg-asparaginase use during induction therapy.Methods:This is a single-center study of17 consecutive adult patients (>18 years old) treated for ALL with peg-asparaginasecontaining induction regimen before and after the establishment of institutional policy to useenoxaparinprophylaxis. Patients were identified from hospital research database. The diagnoses were verified. Medical records were reviewed for the occurrence of any clinically relevant bleeding and VTE within 3 months of receiving peg-asparaginase. Levels of coagulation parameters, whenever available, were collected at baseline and at weekly intervals for up to 4 weeks after receiving peg-asparaginase. The risk of bleeding and VTE among patients receivingenoxaparinwas compared to those who did not receiveenoxaparin. Fisher exact test was used for testing the association of categorical variable, and binomial exact test was used for computing confidence intervals. T-test or nonparametric test was used for comparing the continuous variables depending on whether or not the data were normally distributed.Results:The patient population consisted of 59% females. Mean age was 35 years. Histology consisted of Pre-B ALL in 76%, pre-T ALL in 18% and mixed lineage leukemia in 6%. 88% percent received pediatric-type chemoregimenand intramuscular peg-asparaginase; the most common doses of peg-asparaginaseincluded 2000 mg/m2 (41%) and 2500 mg/m2 (35%). Duration ofenoxaparin prophylaxis was 21 days in 66.7% and 7 days in 33.3%. Average changes in coagulation parameters at baseline and at weekly intervals following the use of peg-asparaginase therapy are highlighted in figures 1 and 2. The drop inantithrombin and fibrinogen levels was the highest during the first two weeks after the administration of peg-asparaginase. 41% of patients required cryoprecipitate (median 15 units, range 5-35 units over 4 weeks). No patients, including theenoxaparin group, had any clinically relevant bleeding event. The risk of VTE was 18% (95% confidence 4-43%); 2 out of 3 VTE events were cerebral venous thrombosis. No VTE occurred in theenoxaparin prophylaxis group (n=9), whereas 37% of patients (3 out of 8 patients), who did not receiveenoxaparin, developed VTE.Enoxaparin prophylaxis was associated with a trend towards a lower risk of VTE, however, the difference was not statistically significant (p=0.08).Conclusion:Within the limits of this study, in adult ALL patients receiving peg-asparaginase, the use of daily prophylactic dose ofenoxaparinmay not be associated with an increased risk of clinically relevant bleeding, and may have a potential to lower the risk of VTE. Patients receivingenoxaparin, however, should be carefully monitored for the need of platelet transfusion and cryoprecipitate supplementation. The difference in the risk of VTE was not statistically significant, which may be, in part, due to a small sample size.Antithrombindepletion may also contribute to a reduction in the efficacy ofenoxaparin. Larger prospective studies should be performed to validate the preliminary findings. [Display omitted] [Display omitted] DisclosuresArmitage:Spectrum Pharmaceuticals: Consultancy; Roche: Consultancy; Conatus - IDMC: Consultancy; GlaxoSmithKline IDMC: Consultancy; ZiopharmOncology: Consultancy; Tesaro bio Inc: Membership on an entity's Board of Directors or advisory committees. Lunning:Genentech: Consultancy; AbbVie: Consultancy; Bristol-Myer-Squibb: Consultancy; Juno: Consultancy; Spectrum: Consultancy; Pharmacyclics: Consultancy; Celgene: Consultancy; Gilead: Consultancy; TG Therapeutics: Consultancy.

Persistence of Cytogenetic Abnormalities at Complete Remission Is Not Prognostic for Relapse-Free or Overall Survival in Adult Patients with Acute Lymphoblastic Leukemia

Background: Cytogenetic abnormalities are identified at the time of diagnosis in approximately 80% of patients with acute lymphoblastic leukemia (ALL). In acute myelogenous leukemia (AML), the detection of persistent abnormal cytogenetics at complete remission (ACCR) is associated with shorter relapse-free survival (RFS) and overall survival (OS) compared to patients with normal cytogenetics at CR (NCCR). However, the incidence and prognostic significance of ACCR in adult patients with ALL is unknown.Methods: We evaluated 324 adult patients with ALL and abnormal cytogenetics at baseline who were treated on frontline induction chemotherapy protocols between 6/2001 and 3/2015 and achieved complete remission (CR) or CR without platelet recovery (CRp). Bone marrow specimens for cytogenetic and minimal residual disease (MRD) assessment were obtained at the time of CR. Cytogenetic abnormalities were classified according to standard conventions (Moorman et al., Blood 2007;109:3189-97). MRD by multi-parameter flow cytometry (MFC) was assessed with a sensitivity of 0.01% using a 15-marker, 4-color panel in the first half of the study period; subsequently a 6-color panel was used.Results: 272 patients (84%) had cytogenetic analysis performed at the time of remission and were evaluable for this analysis. Median age was 48 years (range, 16-84) and median WBC at presentation was 11.6 x109/L (range, 0.4-629.4 x109/L). A hyperCVAD backbone was used for induction in 237 patients (87%) and augmented BFM was used in 35 (13%). All patients with Philadelphia positive (Ph+) ALL were treated with a tyrosine kinase inhibitor added to the chemotherapy regimen. 245 patients (90%) had pre-B ALL, 14 (5%) had pre-T ALL and 13 (5%) had Burkitt or Burkitt-like leukemia. Cytogenetics at baseline were Ph+ in 119 (44%), hyperdiploid in 31 (11%), complex in 18 (7%), MLL rearranged in 16 (6%), hypodiploid in 9 (3%) and miscellaneous in 79 (29%). Median time to CR was 23 days (range 14 to 84 days).Among the 272 patients, ACCR was observed in 26 (9.6%). Baseline characteristics associated with ACCR were Ph+ ALL (62% of ACCR group vs. 42% of NCCR group, P=0.055) and longer mean time to CR (29.8 ± 15.6 days for ACCR group vs. 26.0 ± 9.7 days for NCCR group, P=0.07). Median RFS was 22.4 months (range, 12.3 months to not reached) for patients with ACCR vs. 47.7 months (range, 29.5 to 125 months) in those patients with NCCR (P=0.31). Median OS did not differ between the ACCR (98.7 months [range, 17.1 months to not reached]) and NCCR groups (67.3 months [range, 46.6 months to not reached], P=0.86). There was also not a significant difference in RFS or OS between the ACCR and NCCR groups when only Ph+ patients were evaluated. Among the 227 patients evaluable for MRD by MFC, MRD positivity at CR was observed in 78 patients (34%) and was highly associated with shorter RFS and OS (P<0.01 for both). The specificity of ACCR for detecting MRD (as assessed by MFC) was 43%, and there was overall poor correlation between these two methods for the detection of residual disease (P=0.47). When patients were stratified by MRD status as assessed by MFC, the presence of absence of persistent cytogenetic abnormalities did not add additional prognostic information (Fig. 1 and 2).Conclusions: There is poor association between MRD assessment by MFC and the presence or absence of cytogenetic abnormalities at CR in adult patients with ALL. Although ACCR has prognostic significance in AML, ACCR is not associated with adverse outcomes in ALL and therefore should not be used to guide prognostication or therapeutic decisions. [Display omitted] [Display omitted] DisclosuresFaderl:Celgene Corp.: Other: Advisory Board. Burger:Pharmacyclics LLC, an AbbVie Company: Research Funding. Konopleva:Novartis: Research Funding; AbbVie: Research Funding; Stemline: Research Funding; Calithera: Research Funding; Threshold: Research Funding. Cortes:Pfizer: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Teva: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; ARIAD Pharmaceuticals Inc.: Consultancy, Research Funding.

THE IMMUNOPHENOTYPE OF ADULT T ACUTE LYMPHOBLASTIC LEUKEMIA IN MOROCCO

There is paucity of detailed studies of adult T cell acute lymphoblastic leukemia (T-ALL) in developing countries reflecting the condition of these patients including clinical and biological features. This study was carried out to analyze the immunophenotypic characteristics of 40 Moroccan patients with T-ALL and its association with biological and clinical features. Between 2006 and 2009, 130 adult patients diagnosed with acute lymphoblastic leukemia (ALL) were immunophenotyped by 3-color flow cytometry using a panel of monoclonal antibodies. Cases presenting features of a T-lineage phenotype were subjected to detailed analysis including immunophenotypic, clinical and biological parameters. Proportion of T-ALL among ALL Moroccan patients was 31.0%. Median age of patients was 28 years. Twenty-nine patients were females and 11 were males. 45.0% of patients (18/40) had features of immature T-ALL stages (pro-T and pre-T ALL), 30.0% (12/40) of CD1a+ cortical T-ALL stage and 25.0% (10/40) had a characteristic phenotype of medullary T-ALL. The frequencies of progenitor cell markers CD10, CD34 and TdT expression were 14.0; 57.5% and 50.0% respectively. The aberrant expression of B lineage associated antigen CD79a were positive in 20.5% of the cases and the aberrant expression of myeloid antigens CD13 and/or CD33 was found in 22 (55.0%) cases. No significant association was encountered between TdT, CD34 or myeloid antigens positivity and high risk features at presentation as age, sex, and white blood cells. However, myeloid antigens (CD13 and/or CD33) was significantly associated with T-cell maturation stages (p = 0.009). To the best of our knowledge, this is the first report from North Africa of immunophenotypic study on adult T-ALL. Our findings indicate that the proportion of T-ALL among ALL in Morocco is similar to that reported in others Mediterranean countries like France and Italy and that myeloid-associated antigens expression is frequently associated with immature immunophenotype.

Akut Lenfoblastik Lösemide İzole Optik Sinir Relapsı: İki Olgu Sunumu

Notwithstanding extensive chemotherapy, intrathecal methotrexate treatments and prophylactic central nervous system radiotherapy, extramedullary relapses can still be seen in leukemia patients. Isolated optic nerve relapses, one of its symptoms being a sudden vision loss that occurs prior to a bone marrow relapse, are rarely seen in acute lymphoblastic leukemia patients. Sudden vision loss can be cured by an early diagnosis and rapid treatment. In this study, we investigate two male patients (10 and 12 years old) with optic nerve relapses, sudden vision loss complaints and pre-T acute lymphoblastic leukemia in the light of the literature.

Impactof NOTCH1, FBXW7and SETD2 Mutations On Early Treatment Response In Childhood T-Cell Acute Lymphoblastic Leukemia：A China Children's Leukemia Group Study

T-cell lymphoblastic leukemia (T-ALL) is an aggressive malignancy accounting for about 15% of newly diagnosed acute lymphoblastic leukemia (ALL) in children. Although the prognosis has been improved by intensified therapies, the outcome of high risk patients is still not optimistic. Activating NOTCH1 and/or FBXW7 mutations have been reported to be the leading genetic abnormality in T-ALL with good prognosis but few studies about the role of NOTCH1 and FBXW7 in T-ALL were conducted in China and their mutation distributions and clinical prognosis are still unclear. On the other hand, loss of function mutations of SETD2, a histone methyltransferase specific for trimethylation of histone H3 on Lys36 (H3K36me3), has been found in renal cell carcinoma, gliomas and early T cell precursor ALL (ETP-ALL). SETD2is a potential tumor suppressor gene and may play a deleterious role in human cancer. We attempted to investigate the role of these mutations in Chinese T-ALL children. Thirty-seven bone marrow samples from childhood patients with newly diagnosed T-ALL (26 boys and 11 girls; age＜14 years) enrolled into the China Children's Leukemia Group (CCLG) ALL-2008 trail from September 2010 to December 2012 were analyzed for SETD2, NOTCH1, FBXW7 mutations. All exons of SETD2, exons 25-28, 34 of NOTCH1 and exons 5, 7-12 of FBXW7, including intron-exon boundaries, were sequenced by Sanger method.NOTCH1 mutations were observed in 43.2% (n=16) of the T-ALL patients. Thirteen missense mutations were found in HD-N domain, HD-C domain and TAD domain of NOTCH1 while only frameshift insertions and deletions were identified in PEST domain. Besides missense mutations, five inframe insertions were also observed in HD-N domain of NOTCH1. We only found one FBXW7 synonymous mutation in our patients, which was extremely low compared with that in other studies. SETD2 somatic mutations were found in 8.1% (n=3, one female and two males) of 37 T-ALL patients. Of the three patients, two patients had truncated SETD2 protein because of nonsense mutation (c.6229 C＞T) and frameshift insertion (c.7516_7517insTTATA) respectively. Both SETD2 and NOTCH1 mutations were found in one patient. In our study, no significant relationships between NOTCH1 status and age, sex, mediastinal or Central Nervous System (CNS) involvement, immunophenotype and cytogenetics were observed. However, White Blood Cell (WBC) count at diagnosis in patients without NOTCH1 mutation were much higher (221×109/L vs 76.95×109/L, P=0.015). Patients with NOTCH1 mutations had a better early treatment response: higher prednisone good response rate (81.2% vs 47.6%, p=0.048), higher incidence of bone marrow M1 status (blast cell＜5%) on day 15 (73.3% vs 33.3%, p=0.04), and higher rate of favorable minimal residual disease (MRD) level on day 88 (100% vs 59.9%, p=0.012). In contrast, patients without NOTCH1 mutations were more likely to fall into the high risk group (71.4% vs 21.4%, p= 0.006) according to our treatment protocol. After a median follow-up of 11 months, 1 patient did not reach complete remission (CR), 2 patients died during induction and 2 patients relapsed in 6 months. Patients who suffered relapse and induction failure are all in the NOTCH1 wild type group. SETD2 mutations seemed to have no relationship to sex, age, WBC, cytogenetics, CNS and mediastinal involvement. In contrast to previous study that found SETD2 mutations only in EPT ALL patients, none of our SETD2 mutated patients were ETP ALL, two were cortical-T ALL and one was pre-T ALL. All of the patients with SETD2 mutations were alive without the disease at the last follow-up. However, all of the patients were in the high risk group. The patient who had both NOTCH1 and SETD2 mutations responded poorly to prednisone. Although the other two patients without NOTCH1mutations had good response to prednisone, favorable BM status on day 15 and low MRD on day 33, they had very high MRD levels (0.21% and 0.44% respectively) on day 88.NOTCH1 activating mutations were found in 43.2% of pediatric T-ALL patients enrolled in the CCLG ALL-2008 study with good treatment response while FBXW7 mutation rate was much lower. SETD2 mutation was a recurrent event in pediatric T ALL not only in ETP ALL. It seems that patients with SETD2 mutations have high risk of poor response to chemotherapy regardless of NOTCH1 status. Further studies are needed to find out the prognostic significance of SETD2 mutation in pediatric T ALL. Disclosures:No relevant conflicts of interest to declare.

VPREB1 Deletions Occur Independent of Lambda-Light Chain Rearrangement and Predict Worse Outcome In Pediatric Acute Lymphoblastic Leukemia (ALL)

AbstractAbstract 273VPREB1 deletions occur in 1/3 of pediatric ALL (pALL) patients, but have been considered a normal consequence of λ light chain rearrangement due to VPREB1‘s location among λ chain variable region genes. Recently, VPREB1 deletions were reported in high risk pALL samples with BCR-ABL1-like gene expression (Den Boer 2009) and VPREB1 under-expression was found in a 38-gene signature associated with relapse (Kang 2010). We characterized VPREB1 deletions to determine if they are part of normal λ light chain rearrangement or represent a bona fide copy number alteration (CNA) that predicts worse outcome in pALL. We first used the genome-wide Molecular Inversion Probe (MIP) 330K Cancer Panel (Affymetrix) to identify VPREB1 deletions in a cohort of 52 pALL patients from the University of Utah (n=47 Precursor B-cell (Pre-B), 4 Precursor T-cell (Pre-T), 1 relapse Pre-B ALL). We found focal VPREB1 deletions in 25% (n=12/48) of Pre-B ALL and no (n=0/4) Pre-T ALL patients. We also identified a distinct and often homozygous deletion (14 kb in length) located ≂f 78 kb upstream of VPREB1 that could represent a potential enhancer region; this possible enhancer deletion was seen in 16.7% (n=8/48) of Pre-B and none of the Pre-T ALL patients. 50% of Pre-B ALL samples had at least one deletion of B-cell developmental gene (SPI1, IKZF1, BCL11A, TCF3, EBF1, PAX5, FOXP1, LEF1, VPREB1, BLNK), and of these, half (n=12/24) had deletion only of VPREB1 or its possible enhancer. We next sorted blood leukocytes from 10 healthy volunteers for quantitative PCR (qPCR) analysis on κ V-J junction (deletion=attempted κ rearrangement), λ V-J junction (deletion=attempted λ rearrangement), LOC96610 and PRAME (distal to VPREB1, among λ-light chain variable genes), VPREB1, and VPREB1 enhancer (proximal to VPREB1, among λ-light chain variable genes). As predicted for normal light chain rearrangement, we observed no deletions in monocytes; 100% of κ-sorted B-cells had κ V-J deletions without lambda deletions; and 100% of λ-sorted B-cells had both κ V-J and λ V-J deletions. We also analyzed 10 mature B-cell (Burkitt) lymphoma samples and observed VPREB1 deletions only in lambda-expressing samples with both κ and λ V-J deletions, and deletions always extended contiguously downstream to λ chain V-J region. We re-analyzed 30 Pre-B ALL samples (Utah cohort) and 11 Pre-B ALL cell lines by qPCR. Results validated MIP CNA patterns and differed from normal sorted B-cell and mature B-cell (Burkitt) results, displaying several types of rearrangements: 1) No light chain rearrangements with no VPREB1/enhancer deletion (n=5/30 [17%] clinical samples and 2/11 [18%] Pre-B cell lines), 2) Normal light chain (κ or κ/λ) rearrangement with no VPREB1/enhancer deletion (n=5/30 [17%] samples and 6/11 [55%] cell lines), 3) Normal light chain (κ or κ/λ) rearrangements with contiguous VPREB1/enhancer deletion (n=1/30 [3%] samples and no cell lines), 4) λ-light chain rearrangement with non-contiguous VPREB1/enhancer deletion (n=13/30 [43%] samples and 2/11 [18%] cell lines), and 5) No λ light chain rearrangement and focal VPREB1/enhancer deletion (n=6/30 [20%] samples and 1/11 [9%] cell lines). In the Utah Pre-B ALL cohort (n=47), VPREB1/enhancer deletions were significantly associated (Fisher's Exact Test, 2-tailed) with Hispanic ethnicity (p= 0.013), relapse (p= 0.027), death (p=0.026), and Day 14 M2 Marrow (5-25% blasts; p=0.038). Finally, we examined the publicly available St. Jude Children's Research Hospital ALL dataset (NEJM 351:533) and found VPREB1 under-expression associated with higher LC50 (resistance) for DNR (p=0.04) and VCR (p=0.04), but not PRED or ASP. In summary, we have shown VPREB1 deletions occur outside the context of normal light chain rearrangement. Unlike physiologic λ rearrangements, VPREB1 deletions do not always extend to the λ V-J junction in pALL. We also identified a focal, homozygous deletion just proximal to VPREB1 that may affect a possible enhancer region for this B-cell developmental gene. VPREB1/enhancer deletions were the only B-cell developmental gene lost in 25% of our cohort, perhaps defining a new Pre-B ALL subtype. Finally, deletion of VPREB1 (and its possible enhancer) predict worse clinical outcome and VPREB1 down-regulation correlates with in vitro drug resistance. Further studies of VPREB1‘s function in pediatric ALL may improve our understanding of leukemogenesis and could further refine clinical risk stratification. Disclosures:No relevant conflicts of interest to declare.

A case of chronic myeloproliferative syndrome followed by precursor T-cell acute lymphoblastic leukemia

Interlineage switch from myeloid to lymphatic malignancies is a rare phenomenon. Progression from a BCR-ABL negative chronic myeloproliferative disorder (CMPD) to acute lymphoblastic leukemia (ALL) has been reported in very few cases. We describe the case of a 62-year-old man who developed precursor T-cell (pre-T) ALL 18 months after the diagnosis of an unclassifiable chronic myeloproliferative syndrome (CMPD, U), which had been treated with hydroxyurea (HU) over 12 months. The transformation from CMPD to pre-T-ALL was accompanied by clonal evolution from normal to a high hyperdiploid karyotype with an i(17q): 52,XY,+X,+2,+13,+14,+15,i(17)(q10),+19. Diverse pathways should be considered in this transformation process: although therapeutic induction of ALL is extremely rare and no MLL/11q23 rearrangement was detected by chromosome banding analyses and interphase fluorescence in situ hybridization (FISH), T-lineage ALL might have been caused by HU therapy for the CMPD. Chance coincidence of both disorders seems improbable, given the short interval from the diagnosis of the CMPD to the development of the pre-T-ALL, but nonetheless must be considered. A third explanation might be provided by a spontaneous interlineage switch, which would give further support to the theory that the CMPDs are disorders of a pluripotent stem cell. Interlineage switch might result from an aberrant differentiation of the malignant clone or from selection within a mixed population.

Adult T-cell acute lymphoblastic leukemia: biologic profile at presentation and correlation with response to induction treatment in patients enrolled in the GIMEMA LAL 0496 protocol

AbstractBetween 1996 and 2000, 90 newly diagnosed adult patients with T-acute lymphoblastic leukemia (T-ALL) were registered in the Gruppo Italiano Malattie Ematologiche dell'Adulto (GIMEMA) Leucemia Acuta Limfoide (LAL) 0496 protocol. Cases were centrally processed for morphology, immunophenotype, cytogenetics, molecular biology, and multidrug resistance (MDR). Twenty-two patients were females and 68 were males. Four percent of cases were pro-T, 47% pre-T, 39% cortical T, and 10% mature T-ALL. Fifty-six percent of patients with pro-T + pre–T-ALL achieved complete remission (CR) compared with 91% for cortical + mature cases (P = .002). CD34 expression was associated with a significantly lower CR rate: 54% versus 84% (P = .009). Thirty-one (36.5%) of 85 patients had an abnormal karyotype, the most common abnormality (15%) being a partial del(6q). The cytogenetic profile did not impact on CR achievement. MDR1 function, present in 26% of cases, correlated significantly with CR achievement (P = .004). A highly significant (P = .001) difference in CR rate was observed between patients who did not express the CD13/CD33/CD34 antigens and were MDR functionally negative (96%) compared with patients positive for at least one of these markers (57%). Multivariate analysis showed an impact on CR achievement for CD33 expression and MDR1 function. An extensive biologic workup of adult T-ALL cases at presentation is recommended in order to design tailored therapeutic strategies aimed at improving CR rates.

Significado pronóstico de los subtipos inmunológicos de la leucemia aguda linfoblástica T del adulto. Estudio de 81 pacientes

T-cell acute lymphoblastic leukemia (ALL) includes 4 immunological subtypes: pro-T, pre-T, thymic or cortical and mature. In some studies, pro-T and mature subtypes have a poor prognosis. The objective of this study was to describe the clinical characteristics, the result of treatment and the prognosis of the immunological subtypes of T-cell ALL in 81 adult patients included in 2 protocols of the Spanish PETHEMA group (ALL-96 and ALL-93). Between 1993 and 2003, 81 adult patients from 22 Spanish hospitals were included in two PETHEMA protocols: ALL-96 for standard-risk patients, and ALL-93 for high- risk patients. The main clinical and biological parameters as well as the rate of response to treatment, the frequency of complete remission , disease free survival and overall survival were compared in each T-cell ALL subtype. Of the 64 evaluable patients the distribution of the immunological subtypes was: 3 pro-T, 17 pre-T, 22 thymic or cortical and 22 mature. Patients with mature T-ALL had higher frequency of central nervous system involvement and myeloid antigen expression than those of the remaining subgroups. Patients with mature T-cell ALL had a slow rate of response to treatment in comparison with patients wit pre-T and mature T-cell ALL but this did not translate to significant differences in frequency of complete remission (77% vs 94%), disease free survival (42% vs 46%) and overall survival (29% vs 47%). Although patients with mature T-cell ALL had a slow rate of response to treatment and their survival tended to be shorter, in the present study there were no statistically significant differences in the prognosis of the different subtypes of T-cell ALL.

Growth suppression of pre-T acute lymphoblastic leukemia cells by inhibition of notch signaling.

Constitutive NOTCH signaling in lymphoid progenitors promotes the development of immature T-cell lymphoblastic neoplasms (T-ALLs). Although it is clear that Notch signaling can initiate leukemogenesis, it has not previously been established whether continued NOTCH signaling is required to maintain T-ALL growth. We demonstrate here that the blockade of Notch signaling at two independent steps suppresses the growth and survival of NOTCH1-transformed T-ALL cells. First, inhibitors of presenilin specifically induce growth suppression and apoptosis of a murine T-ALL cell line that requires presenilin-dependent proteolysis of the Notch receptor in order for its intracellular domain to translocate to the nucleus. Second, a 62-amino-acid peptide derived from a NOTCH coactivator, Mastermind-like-1 (MAML1), forms a transcriptionally inert nuclear complex with NOTCH1 and CSL and specifically inhibits the growth of both murine and human NOTCH1-transformed T-ALLs. These studies show that continued growth and survival of NOTCH1-transformed lymphoid cell lines require nuclear access and transcriptional coactivator recruitment by NOTCH1 and identify at least two steps in the Notch signaling pathway as potential targets for chemotherapeutic intervention.

Cellular characteristics of acute leukemia cells simultaneously expressing CD13/CD33, CD7 and CD19.

Of 832 acute leukemia patients, including 580 acute myeloblastic leukemia (AML), 197 pre-B acute lymphoblastic leukemia (ALL) and 55 pre-T ALL, 26 cases (3.1%) of CD13/CD33+CD7+CD19+ acute leukemia were found. A total of 20 patients were diagnosed as AML, two as pre-B ALL and four as pre-T ALL. Based on the relative intensity of expression of CD7 and CD19, CD13/CD33+CD7+CD19+ acute leukemia patients were subclassified into three categories. Type I (CD7 > CD19) included ten AML and four pre-T ALL, having cellular characteristics similar to CD7+ AML and CD13/CD33+CD7+ ALL. Type II (CD7 < CD19) consisted of four AML with t(8;21) and two pre-B ALL. Type III (CD7 = CD19) included six AML. CD13/CD33+CD7+CD19+ acute leukemia frequently expressed stem cell associated molecules, such as CD34 (88.5%), HLA-DR (96.2%) and mRNA for MDR1 (72.2%), GATA-2 (87.5%) and SCL (25.0%). Simultaneous expression of cytoplasmic CD3 and myeloperoxidase in some leukemia cells implies that CD13/CD33+CD7+CD19+ acute leukemia cells have the potential to differentiate into various lineages. These data suggest that a small population of acute leukemia patients with distinct phenotype, CD13/CD33+CD7+CD19+ acute leukemia, may originate from hematopoietic stem cells.

Acute respiratory failure and pulmonary thrombosis in leukemic children

Acute respiratory failure (ARF) in an 11-year-old child with pre-T acute lymphoblastic leukemia (ALL) at the beginning of induction therapy was observed, connected with a pulmonary thrombosis and not with an infective origin. A systematic search for this pathology identified six other children with the same pulmonary complication, five of whom where in the early phase of acute nonlymphoblastic leukemia (ANLL) and one in induction therapy for ALL in marrow relapse. At the beginning of the symptomatology, all children presented severe hypoxia and hypercapnia, with no or minimal chest radiograph abnormalities and no clear hemodynamic involvement. In all patients the arteriography and nuclear imaging studies confirmed the diagnosis. The causes of the thrombi could be connected with neoplastic emboli after cell lysis and/or with the vascular damage resulting from antiblastic therapy. Intravenous urokinase treatment and respiratory assistance had been successfully carried out in six of seven children.

Prethymic phenotype and genotype of pre-T (CD7+/ER-)-cell leukemia and its clinical significance within adult acute lymphoblastic leukemia [see comments

Pretreatment blast cells from 739 adults with acute lymphoblastic leukemia (ALL) were immunophenotyped as part of a prospective treatment protocol study. Among 192 patients (26%) with T lineage ALL, 47 (6%; 24% of T lineage ALL) had lymphoblasts without sheep erythrocyte rosette formation, but with pan-T antigen CD7 on the membrane and intracellular CD3 proteins mostly in perinuclear accumulation. The T- cell surface antigens CD5 and/or CD2 and focal acid phosphatase were additional markers of this subgroup traditionally called pre-T ALL, whereas thymocyte antigen CD1 as well as CD4 and CD8 antigens were not expressed. Hematopoietic progenitor cell markers, namely terminal deoxynucleotidyl transferase (TdT), and in part common ALL antigen (CD10), HLA-DR antigens, and/or My-10 (CD34), a unique antigen of marrow cells absent in thymus cells, further characterized this immature T-ALL form of putative prothymocytic phenotype (CD7+/intracellular CD3+/TdT+/My-10+/-/HLA-DR+/-/CD10+/-). The prethymic T cell character was supported by germ-line T-cell receptor beta genes found in 21 of 36 patients analyzed. In five cases only T gamma-chain genes were rearranged. Fifteen patients, however, had rearrangements of both T beta and T gamma genes. Immunoglobulin heavy chain genes were rearranged only in two cases. Pre-T ALL differed significantly from E-rosette+ T-ALL in some presenting clinical features, namely mediastinal mass, lymphoadenopathy, and platelet count, and independently of clinical factors in prognosis (P = .02, median remission duration: 15.7 v 33.5 months, and P = .02, median survival time: 24.6 v 50.7 months). We conclude that ALL classification based solely on T- or B-cell lineage affiliation is not sufficient but needs further subdivision according to relevant maturation stages as exemplified here within the T-cell axis. The putative prethymic T cell progenitor phenotype described might help elucidate the sequence of genetic events that commit normal hematopoietic cells to the T-cell lineage.

Prethymic Phenotype and Genotype of Pre-T (CD7+/ER-)-Cell Leukemia and Its Clinical Significance Within Adult Acute Lymphoblastic Leukemia

Pretreatment blast cells from 739 adults with acute lymphoblastic leukemia (ALL) were immunophenotyped as part of a prospective treatment protocol study. Among 192 patients (26%) with T lineage ALL, 47 (6%; 24% of T lineage ALL) had lymphoblasts without sheep erythrocyte rosette formation, but with pan-T antigen CD7 on the membrane and intracellular CD3 proteins mostly in perinuclear accumulation. The T-cell surface antigens CD5 and/or CD2 and focal acid phosphatase were additional markers of this subgroup traditionally called pre-T ALL, whereas thymocyte antigen CD1 as well as CD4 and CD8 antigens were not expressed. Hematopoietic progenitor cell markers, namely terminal deoxynucleotidyl transferase (TdT), and in part common ALL antigen (CD10), HLA-DR antigens, and/or My-10 (CD34), a unique antigen of marrow cells absent in thymus cells, further characterized this immature T-ALL form of putative prothymocytic phenotype (CD7+/intracellular CD3+/TdT+/My-10+/-/HLA-DR+/-/CD10+/-). The prethymic T cell character was supported by germ-line T-cell receptor beta genes found in 21 of 36 patients analyzed. In five cases only T gamma-chain genes were rearranged. Fifteen patients, however, had rearrangements of both T beta and T gamma genes. Immunoglobulin heavy chain genes were rearranged only in two cases. Pre-T ALL differed significantly from E-rosette+ T-ALL in some presenting clinical features, namely mediastinal mass, lymphoadenopathy, and platelet count, and independently of clinical factors in prognosis (P = .02, median remission duration: 15.7 v 33.5 months, and P = .02, median survival time: 24.6 v 50.7 months). We conclude that ALL classification based solely on T- or B-cell lineage affiliation is not sufficient but needs further subdivision according to relevant maturation stages as exemplified here within the T-cell axis. The putative prethymic T cell progenitor phenotype described might help elucidate the sequence of genetic events that commit normal hematopoietic cells to the T-cell lineage.