bluuebunny/crossref_metadata_2025 huggingface dataset:
BCR-ABL Activates IGF-1 Expression and Signaling in Chronic Myelogenous Leukemia Blast Crisis Cell Lines.
Authors: Pastural Elodie , Lakshmikuttyamma Ashakumary , Takahashi Naoto , Pearson Derek , Sheridan David , DeCoteau John F. , Ronald Geyer C.
Published: October 2019

CML blast crisis is characterized by the continued presence of the Philadelphia chromosome, which expresses the P210 BCR-ABL fusion protein, and the acquisition of additional molecular and chromosomal alterations. Evolution from CML chronic phase to blast crisis is associated with loss of heterozygosity at chromosome region 1p36, which contains the putative tumor suppressor RIZ1. We found that RIZ1 expression decreases during progression from CML chronic phase to myeloid blast crisis and that forced RIZ1 expression in CML blast crisis (CML-BC) cell lines decreases proliferation, increases apoptosis, and enhances differentiation. Furthermore, we found that RIZ1 represses IGF-1 autocrine production and blocks the activation of the IGF-1 receptor, AKT, and ERK signaling pathways in CML-BC cell lines thereby implicating RIZ1 control of the IGF-1 pathway in the regulation of these phenotypes. As BCR-ABL induces a growth factor independent phenotype due to the activation of autocrine growth factor production, we analyzed IGF-1 expression in CML-BC cell lines following exposure to imatinib. We found that imatinib treatment reduced IGF-1 mRNA and extracellular IGF-1 suggesting that RIZ1 may suppress blast crisis by counteracting the ability of BCR-ABL activate RIZ1 regulated genes. We characterized the signaling pathways used by BCR-ABL to activate IGF-1 expression and found that the HCK inhibitor PP2 and STAT5b shRNA reduced IGF-1 expression whereas the JAK2 inhibitor AG490 increased IGF-1 expression. These results suggest that BCR-ABL activates HCK, which in turn activates STAT5b and induces IGF-1 expression. To confirm the importance of BCR-ABL induced IGF-1 signaling to CML-BC cell line viability, proliferation and apoptosis, we characterized these cellular phenotypes in the presence of IGF-1 receptor blocking antibody and the IGF-1 receptor tyrosine kinase inhibitor AG1024. Blockage of autocrine IGF-1 signaling in CML-BC cell lines using an anti-IGF-1 receptor blocking antibody reduced cell viability and decreased proliferation. AG1024 treatment of CML-BC cell lines decreased proliferation and induced apoptosis. Together these results demonstrate that RIZ1 counteracts the ability of BCR-ABL to induce IGF-1 signaling in CML-BC cell lines, which influences cell proliferation, apoptosis, and differentiation. Our findings highlight RIZ-1 and IGF-1 signaling pathways as potential therapeutic targets for treating CML blast crisis.
The G Protein-Coupled Receptor cysLT1 Is Expressed in Acute Myeloid Leukemia Mediating Cell Migration and Survival.
Authors: Rether Christina , Hengstebeck Sandra , Xue Xingkui , Boehmler Andreas , Kanz Lothar , Seitz Gabriele , Mohle Robert
Published: October 2019

Previous studies have shown that the G protein-coupled receptor (GPCR) cysLT1, which recognizes inflammatory lipid mediators (cysteinyl-leukotrienes), is expressed in immature hematopoietic and tumor (eg. colon cancer) cells. In the present study, we analyzed leukemic blasts from 19 patients with newly diagnosed acute myeloid leukemia for analysis of expression and function of cysLT1. By RT-PCR, cysLT1 mRNA was found in all primary AML cells. Quantitative TaqMan PCR demonstrated high levels particularly in acute myeloblastic leukemia without maturation (FAB M1). There was also a positive correlation between the cysLT1 mRNA level and the expression of CD34 and CD-117 (c-kit), indicating that high cysLT1 expression corresponds to an AML phenotype resembling normal hematopoietic progenitor cells. CysLT1 was functionally active in AML blasts, as demonstrated by intracellular calcium fluxes and actin polymerization induced by the ligand LTD4. Similar to mRNA expression, strongest responses were seen in AML FAB M1. As LTD4 can be produced in the bone marrow by stromal cells and may contribute to bone marrow infiltration of AML, chemotaxis was analyzed. Surprisingly, already low concentrations (10nM) induced significant chemotaxis of AML blasts, while higher concentrations (up to 1 uM) were less effective in a dose-dependent manner. Incubation of myeloid leukemic cell lines (eg. KG1a) and primary AML blasts with the specific cysLT1 antagonist MK571 resulted in significantly reduced viability after 48 h in a dose-dependent manner (10nM-10uM), suggesting also an autocrine function of cysLT1 ligands. To explore signal transduction pathways involved in leukemic cell proliferation and chemotaxis, we found that in AML cell lines, LTD4 induced phosphorylation of Erk/MAP kinase, wich is related to proliferation, and Pyk2, which represents a focal adhesion kinase-like signaling molecule that links GPCRs with cell migration, while the Akt pathway was not involved. We conclude that cysLT1 is consistently expressed in acute myeloid leukemia, and mediates both chemotactic and proliferative responses. Therefore, cysLT1 antagonists, which are already used in the treatment of allergy, may improve the effectiveness of antileukemic therapy.
The Predictive Value of a “Positive” ASH Abstract in AML Therapeutics.
Authors: Estey Elihu H. , Bedikian Slyva H. , Witter Dianne C. , Pierce Sherry A. , Giles Francis J.
Published: October 2019

ASH abstracts describing outcomes of trials of new therapies (Rxs) are presumably read in anticipation that positive results, although necessarily preliminary, will herald therapeutic advances. Here we examine the predictive value of a “positive” (pos) ASH abstract. We reviewed abstracts, presented in oral or poster sessions from 1993–2001, dealing with early phase trials of new drugs, used alone or together with more standard Rx, in adult AML (APL excepted). A research nurse (RN), a medical editor (ME), and E. Estey (EE) reviewed the abstracts independently, with the latter’s decision final. There was 80–85% concordance among the 3 reviewers, with EE tending to view fewer abstracts as pos. 63/91 abstracts (69%) were judged pos from a therapeutic standpoint (based typically on conclusions that the rx was “active” or “effective”), 14 were considered “negative”, and 14 inconclusive. The 63 pos abstracts involved 37 separate drugs, i.e. several drugs were the subject of > 1 pos abstract, including gemtuzumab, and topotecan combinations (5 abstracts each) and PSC and IL-2 (without histamine) (4 abstracts each). A “pub med” search indicated that 45/63 (71%) pos abstracts, covering 27 of the 37 separate drugs, subsequently appeared in peer-reviewed journals, with half the unpublished abstracts appearing at the 2000 or 2001 ASH meetings. The pos conclusion was unaltered in 38 of the 45 and was changed to negative in 5 and to inconclusive in 2. Three of the 37 pos drugs have either migrated into clinical practice (gemtuzumab) or been pos in a randomized trial (IL-2 + histamine, cyclosporine + infusional daunorubicin + ara-C). Four of the pos drugs (PSC833 + chemotherapy. HuM195, fludarabine + ara-C, troxacitabine + ara-C) have appeared, in some pt subsets, negative in randomized trials. However, 81% (30/37) of the drugs reported pos in ASH abstracts remain, at least 5 years from date of ASH publication, either outside the scope of clinical practice or unevaluated in randomized trials. We suggest that the reasons for this phenomenon be explored.
Comorbidity Scoring Using the Hematopoetic Cell Transplantation Comorbidity Index (HCTCI) Is Predictive of Early Death Rate and Event Free Survival (EFS) in Patients over 60 Years of Age Receiving Induction Therapy for Acute Myeloid Leukemia (AML).
Authors: Giles Francis , Borthakur Gautam , Ravandi Farhad , Faderl Stefan , Verstovsek Srdan , Pierce Sherry , Jabbour Elias , Cortes Jorge , Kantarjian Hagop
Published: October 2019

The majority of patients diagnosed with AML who are 60 years of age or over are not offered specific anti-leukemia therapy. Outcomes in the minority who are offered standard induction are considerably inferior to those in younger patients. While a major factor is AML with inherently worse biology, comorbidity seen in the elderly which increases vulnerability to the adverse events of induction cytotoxic chemotherapy is also a major factor. The HCTCI is useful in predicting nonrelapse mortality and overall survival post stem cell transplantation (Sorror. Blood.2005;106:2912) As this scale represents a refinement of the Charlson Comorbidity Index, and was developed specifically in patients with hematologic malignancies, we investigated whether it might also be applicable to elderly patients receiving AML induction chemotherapy. Pre-therapy comorbidity scoring by the HCTCI was assessed in a cohort of 177 patients over 60 years of age with AML treated at MDACC with standard idarubicin and ara-C induction since 1990. Median patient age was 70 (60–89), 64% were male, 26% had PS 2 or above, 60% had an antecedent hematological malignancy, and 90% had non favorable cytogenetics (33% -5/-7). 22% of patients had HCTCI scores of 0, 30% had scores of 1–2, and 48% scores of 3 or more. Complete response rates in the 3 groups were 64%, 43%, and 42% respectively (p=0.051). Early death (defined as death within 28 days from time of commencing induction therapy) was 3%, 11%, and 29% respectively (p= <0.001). HCTCI score was also highly correlated with EFS with a score of 0 associated with a median EFS of 26 weeks in contrast to a median EFS of 6 weeks in those with a HCTCI score greater than 0 (p=0.03). Even in the probably highly selected subset of patients over 60 years of age currently deemed suitable for standard induction therapy for AML, scoring of pre-treatment comorbidity using the HCTCI is predictable of early death rates and EFS. The further investigation of HCTCI scoring in AML induction, including its value relative to other standard prognostic indicators, is warranted. Stratification of patients for current therapy or for protocol entry based on HCTCI scores seems reasonable.
Impact of Comorbidities on the Outcome of Elderly Patients with Acute Myeloid Leukemia.
Authors: Etienne Anne , Esterni Benjamin , Charbonnier Aude , Prébet Thomas , Coso Diane , Stoppa Anne-Marie , D’Incan Evelyne , Doglio Marie-Françoise , Sainty Danièle , Arnoulet Christine , Mozziconacci Marie-Joëlle , Gastaut Jean-Albert , Vey Norbert
Published: October 2019

Elderly patients with acute myeloid leukemia (AML) have a very poor prognosis, explained by the biologic characteristics of the disease but also by host-related factors including comorbidities. The objective of this study was to determine the prognostic role of comorbidities in this population. Between 1995 and 2004, 133 patients aged 70 years or older with newly diagnosed acute non promyelocytic leukemia, were treated with intensive induction chemotherapy (“3+7” regimen) at a single institution. Comorbidities at diagnosis were retrospectively evaluated using an adapted form of the Charlson Comorbidity index (CCI) (Sorror et al., Blood2004;104:961–968). Seventy-five patients achieved a complete response (CR) (56.4%) and 23 patients died during induction chemotherapy (17.3%). Comorbidity scores at diagnosis were 0 for 83 patients (68%), 1 for 16 patients (13.1%), and > 1 for 23 patients (18.7%). The median overall survival (OS) was 9 months (95% CI: 6–13) and was significantly poorly affected by high leukocyte count (≥ 30 G/l) and elevated serum creatinine level (>1.5 x upper normal limit). By multivariate analysis, we identified 4 adverse prongnostic factors for CR: unfavorable karyotype, leukocytosis ≥ 30 G/l, expression of CD34 on blast cells and a CCI > 1. Age was not found to be a significant prognosic factor for either CR or survival. When a score of 0 or 1 was affected to each prognostic variable, patients could be separated into a low-risk group (score= 0–1; 22% of patients), an intermediate-risk group (score= 2; 41% of patients) and a high-risk group (score≥3 ; 37% of patients), having 87%, 63% and 37% chance of achieving CR following conventional induction chemotherapy, an 8-week mortality rate of 9%, 22% and 34% (p=0.02), and a 2-year overall survival rate above 50%, less than 30% and 15%, respectively (p=0.01).

Our results show that comorbidity is an important prognostic factor for elderly patients with AML which may help the physicians’ decision making. Its capture at diagnosis should be prospectively evaluated in larger study for its future use for risk-stratification among elderly patients treated for AML.
VEGF-TRAP Exhibits Potent In Vivo Anti-Tumor Activity in Acute Myeloid Leukemia Xenograft Models.
Authors: Lal Deepika , Park Jennifer A. , Drake Allison S. , Hoelcle Brandon J. , Wang Eunice S.
Published: October 2019

Vascular endothelial growth factor (VEGF) is known to play a pivotal role in solid tumor angiogenesis, but its role in hematological cancers is less well established. Recent data have shown that subsets of acute leukemias express aberrant VEGF receptors (VEGFR) and rely on both autocrine and paracrine VEGF-VEGFR mechanisms for proliferation, migration, and extramedullary disease. We evaluated the efficacy of a soluble decoy receptor for VEGF-A (VEGF-TRAP, provided through collaboration with CTEP and Sanofi-Aventis/Regeneron Pharmaceuticals) in xenografts of human acute myeloid leukemia (AML) established in SCID mice. All three AML cell lines (ML-2, HEL, HL60/tVCR) used in these studies expressed VEGF isoforms and VEGFR by RT-PCR. The HL60/tVCR line has previously been shown to express Pgp glycoprotein conferring resistance to multiple chemotherapy agents. Mice bearing subcutaneous leukemia xenografts were treated with PBS (vehicle control) or VEGF-TRAP (5 or 25 mg/kg) twice a week for three weeks (for a total of six doses). We found that VEGF-TRAP therapy consistently reduced leukemia tumor volumes by an average of 76–93% versus controls in all three xenograft models. VEGF-TRAP treated tumors were visibly less vascular than control tumors and demonstrated large necrotic areas with only small islands of viable cells by immunohistochemistry. In vivo administration of VEGF-TRAP, which binds VEGF secreted both by tumor (human) and host (murine) cells, was significantly more effective in reducing leukemia tumor volumes (87%) than anti-human VEGF antibody (bevacizumab (Avastin)) treatment (5 mg/kg three times per week) (68% tumor reduction, p<0.05). Moreover, mice treated with VEGF-TRAP also demonstrated a marked elevation of murine VEGF levels (up to thousand fold higher than control or Avastin-treated animals), which persisted over the duration of therapy despite much smaller overall tumor volumes. These data show that the in vivo anti-VEGF effects mediated by VEGF-TRAP may result in significant feedback response loops in the host microenvironment as well as potential direct effects on tumor cells. Additional experiments investigating the effects of VEGF-TRAP in systemic leukemia mouse models are ongoing. Because VEGF-TRAP treatment alone was not curative, we further examined the effects of combination VEGF-TRAP and chemotherapy (cytarabine or doxorubicin) treatment on subcutaneous leukemia xenograft growth. Data in HL60/tVCR and ML-2 mouse models suggest a potential additive effect with concomitant VEGF-TRAP and doxorubicin (3 mg/kg 2x/week) therapy but not concomitant VEGF-TRAP and cytarabine (2.0 mg 5x/week) therapy.

CONCLUSIONS: These preclinical data demonstrate the potent in vivo tumor activity of VEGF-TRAP, a novel VEGF inhibitor, in AML. Our results support further clinical development of this agent in acute myeloid leukemia, particularly in combination with anthracycline chemotherapy.
ZM447439, a Novel Aurora Kinase Inhibitor, Induces Growth Arrest and Apoptosis of Human Leukemia Cells.
Authors: Ikezoe Takayuki , Nishioka Chie , Tasaka Taizo , Taniguchi Ayuko , Komatsu Naoki , Kuwayama Yoshio , Togitani Kazuto , Phillip Koeffler H. , Taguchi Hirokuni
Published: October 2019

The Aurora kinases play an important role in chromosome alignment, segregation, and cytokinesis during mitosis. Aberrant expression of these kinases have been shown in solid tumors and considered to associate with aneuploidy and carcinogenesis. This study found that Aurora kinase A and B were aberrantly expressed in human leukemia cell lines (n=15, e.g., PALL-1, -2, HL-60, NB4, MV4-11, etc.) as well as freshly isolated leukemia cells from individuals with acute myeloid leukemia (n=44) compared to peripheral blood mononuclear cells from healthy volunteers (n=12), as measured by real-time PCR. In addition, ZM447439, a novel selective Aurora kinase inhibitor, induced growth inhibition, accumulation of cells with 4N DNA content and apoptosis in human leukemia cells as measured by thymidine-uptake, cell cycle analysis and annexin V staining, respectively. Especially, profound growth inhibition occurred in PALL-1 and -2 cells which possess wild-type p53 gene. Of note, ZM447439 did not inhibit clonogenic growth of myeloid committed stem cells harvested from healthy volunteers. Taken together, inhibition of Aurora kinase may be a promising treatment strategy for the individuals with leukemia.
Sorafenib Potently Inhibits ETV6-PDGFRβ and FLT3 Mutants, and Shows Activity Against FLT3 Mutants That Are Resistant to Other Small Molecule Inhibitors.
Authors: Lierman Els , Lahortiga Idoya , Van Miegroet Helen , Mentens Nicole , Marynen Peter , Cools Jan
Published: October 2019

Activated tyrosine kinases are implicated in the pathogenesis of chronic and acute leukemia, and represent attractive targets for therapy. Sorafenib (BAY43-9006, Nexavar®) is a small molecule B-RAF inhibitor that is used for the treatment of renal cell carcinoma. Sorafenib also has activity against receptor tyrosine kinases from the PDGFR and VEGFR families. Here we show that sorafenib is a potent inhibitor of the ETV6-PDGFRβ tyrosine kinase that is associated with chronic myelomonocytic leukemia. Oncogenic FLT3 receptor tyrosine kinase mutants implicated in the pathogenesis of acute myeloid leukemia are also potently inhibited by sorafenib, including some of the resistant mutants that confer resistance to PKC412 and other FLT3 inhibitors. Sorafenib induced a cell cycle block and apoptosis in the AML cell lines MV4-11 and MOLM-13, both expressing FLT3 with an internal tandem duplication. In contrast, the imatinib resistant KIT(D816V) mutant that is associated with systemic mastocytosis was found to be resistant to sorafenib. These results suggest that sorafenib could be used for the treatment of myeloid leukemias expressing activated forms of PDGFRβ, FLT3, and some PKC412 resistant mutants, but not for leukemias expressing imatinib resistant KIT(D816V).
Induction of Bim Facilitates Apoptosis in Leukemia Cells Treated with HDAC Inhibitors.
Authors: Stubbs Matthew C. , Kim Teresa , Krivtsov Andrei , Atadja Peter , Armstrong Scott A. , Kung Andrew L.
Published: October 2019

Lymphoblastic leukemias containing chromosomal translocations involving the Mixed Lineage Leukemia (MLL, HRX, ALL-1) gene, as well as most acute myeloid leukemias (AMLs) have relatively poor clinical prognoses due in part to intrinsic drug resistance. Therefore, new avenues are being explored for treatment of MLL-rearranged ALL and AMLs. One possible new therapeutic class currently being investigated is the histone deacetylase (HDAC) inhibitors. We utilized the histone deacetylase inhibitor NVP-LAQ824 (Novartis, Basel, Switzerland) and analyzed its effects on MLL rearranged and other myeloid leukemias. We also made use of an MLL-AF9 expressing myeloid leukemia cell line (AKLG) derived from purified murine leukemia stem cells to perform gene expression analysis on NVP-LAQ824 treated cells in order to further understand the mechanism of action of HDAC inhibitors, and to potentially identify cooperating therapeutics. NVP-LAQ824 inhibits cell growth at similar concentrations for all cell lines and primary patient samples tested (~25–50nM) as determined by MTT assay 48 hours after treatment. NVP-LAQ824 does not appear to induce apoptosis solely through inhibition of the HSP90/FLT3-ITD complex as cell lines possessing FLT3-ITD (a HSP90-chaperoned protein) and cells without this mutation have similar drug sensitivities. In fact, in cells overexpressing FLT3-ITD that are treated with NVP-LAQ824, phospho-FLT3-ITD levels do not diminish. Microarray data indicated that NVP-LAQ824 induces the BH3-only family member bim. This finding was verified by Western blotting in all cell lines and patient samples tested. Further, shRNA-mediated knockdown of Bim induced relative resistance of cells to NVP-LAQ824. The expression profile also showed similarities to gene expression patterns of dexamethasone treated cells, namely, increased bim levels and decreased expression of c-myc, raising the possibility of synergy between the two drugs. Using MTT assays, we discovered that NVP-LAQ824 in low doses (25nM) induces sensitivity to dexamethasone in glucocorticoid resistant cell lines in a glucocorticoid receptor (GR) dependent manner. Therefore, our data indicate that NVP-LAQ824 may reverse glucocorticoid resistance and may provide insight into glucocorticoid resistance in MLL rearranged leukemias. The biochemistry behind HDAC inhibitors merits further study.
A Randomized Study with or without Intensified Maintenance Chemotherapy in Patients with Acute Promyelocytic Leukemia Who Had Become Negative for PML-RARα Transcript after Consolidation Therapy: The Japan Adult Leukemia Study Group (JALSG) APL97 Study.
Authors: Asou Norio , Kishimoto Yuji , Kiyoi Hitoshi , Okada Masaya , Kawai Yasukazu , Tsuzuki Motohiro , Horikawa Kentaro , Matsuda Mitsuhiro , Shinagawa Katsuji , Kobayashi Tohru , Ohtake Shigeki , Nishimura Miki , Takahashi Masatomo , Yagasaki Fumiharu , Takeshita Akihiro , Kimura Yukihiko , Iwanaga Masako , Naoe Tomoki , Ohno Ryuzo
Published: October 2019

The use of all-trans retinoic acid (ATRA) has markedly improved therapeutic outcome in patients with acute promyelocytic leukemia (APL). Non-cross resistance between ATRA and chemotherapeutic drugs has contributed to not only a high complete remission (CR) rate but also a decrease in the relapse rate, leading to the significant improvement in disease-free survival (DFS) and overall survival (OS) rates. However, it is not clear whether maintenance chemotherapy actually prevents relapse in APL patients treated with ATRA and chemotherapy. If short-term therapy without maintenance shows identical DFS rates as compared to long-term therapy with maintenance, it would be beneficial for quality of life of patients as well as to medical costs. In these aspects, to determine an efficacy of maintenance/intensification chemotherapy, this study was designed to compare the DFS and OS rates in previously untreated adult patients with APL who showed absence of PML-RARα fusion transcript at the end of consolidation therapy and were randomly allocated to either maintenance therapy or observation. Of 302 registered, 283 patients were assessable and 267 (94%) achieved a CR. Predicted 6-year OS and DFS rates were 83.7% and 69.2%, respectively. The PML-RARα fusion gene was amplified using bone marrow samples at the diagnosis and after consolidation therapy by the reverse transcriptase-polymerase chain reaction analysis. The detection limit of PML-RARα fusion transcript in this assay was 10-4. Among 235 patients who completed 3 courses of consolidation chemotherapy, five (2.1%) were positive for the PML-RARα fusion transcript. Three of them subsequently relapsed and another one patient received allogeneic hematopoietic stem cell transplantation. On the other hand, 230 patients (97.9%) showed no PML-RARα transcript in the bone marrow cells at the end of consolidation. Of these, 175 patients were randomly assigned to receive moderately intensive and intermittent maintenance chemotherapy (n=89) or to observation (n=86). Predicted 6-year DFS was 63.2% for the chemotherapy group and 81.8% for the observation group, showing no statistically significant difference (p=0.102). Predicted 6-year OS in patients assigned to observation was 98.7% and was significantly higher than 85.9% in those allocated to maintenance therapy (p=0.013). These results indicate no benefit from adding maintenance chemotherapy in APL patients who are negative for PML-RARα fusion transcript after 3 courses of intensive consolidation.
Prognostic Analysis of Aberrant Somatic Hypermutation of RhoH in Diffuse Large B Cell Lymphoma.
Authors: Katsumi Akira , Hiraga Junji , Iwasaki Toshihiro , Abe Akihiro , Kiyoi Hitoshi , Matsushita Tadashi , Kinoshita Tomohiro , Naoe Tomoki
Published: October 2019

The Rho family of small GTPases including Rho, Rac, Cdc42 has been well characterized as molecular switches to transduce signals from plasma membrane to the downstream effectors. RhoH, a member of the Rho family, is specifically expressed in hematopoietic cells, especially in T and B lymphocytes, and has been reported to downregulate Rac and LFA-1. As RhoH is GTPase deficient and constitutively active, GTP-bound form, the activity of RhoH is directly related to the level of expression. Aberrant somatic hypermutation (SHM) of RhoH gene has been reported in 43% of Diffuse Large B cell Lymphoma (DLBCL) patients (Pasqualucci L et.al., Nature.2001;412:341–346). Although several studies demonstrated the SHM of DLBCL and other lymphoid malignancy, few of the previous reports investigated the clinical significance of SHM. To definitively resolve this issue, we have screened for SHM of RhoH gene in the lymph node samples from 100 previously untreated DLBCL cases. These patients were diagnosed and treated at Nagoya University hospital since 1987 to 2002. Informed consent was obtained from all patients. Samples from 12 cases of reactive lymphadenopathy were also screened for negative control. Intron 1 of RhoH gene was amplified and screened by a denaturing HPLC technique (Transgenomic WAVE_). Mutant samples were confirmed by direct sequencing. We found 78 mutations (76 point mutations, 2 insertions) unique to individual tumor DNAs in 36 of 100 DLBCL cases (36%) in RhoH. Overall, 73 out of 78 were novel SHM, and five were same mutations as previously reported (Pasqualucci L et.al., Nature.2001;412:341–346). The number of mutation in one patient ranged from one to six, and 20 patients had more than one mutations. These mutations were distributed in a region throughout the first 1.6 kb of the RhoH gene. The presence of aberrant SHM in RhoH gene impacted neither on overall survival nor disease free survival (p=0.36 and 0.69, respectively). The absence of aberrant SHM correlated with advanced stage (stage III and IV, p<0.05). Taken together, our study demonstrated that although the SHM might contribute the lymphomagenesis through regulation of RhoH gene expression, it has less impacts on survival of DLBCL patients.
Association between Drug-Metabolizing Enzymes Polymorphisms and Diffuse Large B-Cell Lymphoma Risk in the Middle Eastern Population.
Authors: Siraj Abdul K. , Bu Rong , Al-Rasheed Maha , Ibrahim Muna , Bavi Prashant , Abubaker Jehad , Al-Jomah Naif , Mourad Walid , Al-Dayel Fouad , Ezzat Adnan , El-Solh Hassan , Uddin Shahab , Al-Kuraya Khawla
Published: October 2019

The last four decades have seen significant increase in the incidence of non-Hodgkin’s lymphoma (NHL) as a possible result of increasing environmental carcinogens exposure. Based on the increasing evidence for the association between carcinogens exposure related cancer risk and xenobiotic gene polymorphisms. We have undertaken a case control study on xenobiotic gene polymorphisms in Saudi individuals with a diagnosis of diffuse large B-cell lymphoma (DLBCL).

Polymorphisms in five genes (CYP1A1, GSTT1, GSTP1, GSTM1 and NQO1) were characterized in 187 individuals with DLBCL and 513 normal controls using polymerase chain reaction (PCR) based method. We chose the Saudi population as our study population because of its high consanguinity and its relative genetic homogeneity.

The CYP1A1*2C, GSTT1 null and GSTP1 TT genotype were all found to be significant predictors of DLBCL risk (odds ratio 6.62, 11.94 and 3.42 respectively). None of the other alleles tested for proved to be significant indicators of DLBCL risk. These results suggest that the risk of DLBCL may indeed be associated with xenobiotics - metabolism and thus with environmental exposures.

Table 1 Distribution of polymorphisms in healthy population and lymphoma patients. Polymorphism Genotype Control group Lymphoma patients p OR CYP1A1 −/− 384(76.5%) 104(78.8%) *2A −/2A 105(20.9%) 24(18.18%) 0.543 0.844 2A/2A 13(2.6%) 3(2.27%) 1.000 0.852 2A allele 13% 11.36% 0.659 0.839 CYP1A1 −/− 443(88.2%) 121(91.66%) *2B −/2B 50(10%) 10(7.58%) 0.505 0.732 2B/2B 9(1.8%) 1(0.76%) 0.697 0.407 2B allele 6.8% 4.55% 0.424 0.646 CYP1A1 −/− 497(99%) 125(94.7%) *2C −/2C 5(1%) 4(3.03%) 0.090 3.181 2C/2C 0 3(2.27%) 0.008 ND 2C allele 0.5% 3.8% 0.011 6.627 NQO1 C609T CC 295 (58.5%) 94 (62.7%) CT 177 (35.1%) 37 (24.7%) 0.051 0.656 TT 32 (6.4%) 19 (8.7%) 0.059 1.863 CT+TT 209 (41.5%) 56 (37.3%) 0.395 0.841 GSTP1 2293 CC 389 (76.3%) 113 (77.9%) CT 113 (22.2%) 24 (16.6%) 0.240 0.731 TT 8 (1.5%) 8 (5.5%) 0.017 3.422 CT+TT 121 (23.7%) 32 (22%) 0.739 0.910 GSTP1 A1578G AA 170 (33.5%) 56 (35%) AG 271 (53.5%) 96 (60%) 0.772 1.075 GG 66 (13%) 8 (5%) 0.013 0.368 AG+GG 337 (66.5%) 104 (65%) 0.774 0.937 GSTT1 P 385 (75%) 36 (20.1%) D 128 (25%) 143 (79.9%) <0.001 11.948 GSTM1 P 233 (45.4%) 91 (50%) D 280 (54.6%) 91 (50%) 0.300 0.832 Table 2 Distribution of combined GSTT1 and GSTM1 polymorphisms in case and control group. Genotype Control Case p OR Null: Complete deletion of GSTT1 and GSTM1 allele Present 423 (82.8%) 109 (60.9%) Double Null 88 (17.2%) 70 (39.1%) <0.001 3.087
A Small Subset of CD5-Negative Diffuse Large B-Cell Lymphomas Express Nuclear Cyclin D1 with Evidence of Amplified Cyclin D1 Signals or the t(11;14) Translocation.
Authors: Ehinger Mats , Linderoth Johan , Christensson Birger , Sander Birgitta , Ståhl Eva
Published: October 2019

The expression of cyclin D1 in malignant lymphomas is largely restricted to mantle cell lymphoma, plasma cell myeloma and hairy cell leukemia. In virtually all cases of mantle cell lymphoma and some cases of plasma cell myeloma, the t(11;14) translocation between the immunoglobulin heavy chain and the CYCLIN D1 genes can be demonstrated using fluoroscence in situ hybridization. There are also anecdotal reports of the expression of cyclin D1 in other lymphoma subtypes including small lymphocytic lymphoma, Hodgkin lymphoma and anaplastic large cell lymphoma. In our routine practice, we have encountered a few cases of diffuse large B-cell lymphomas with immunohistochemical evidence of cyclin D1-expression. Therefore, an unselected large number of diffuse large B-cell lymphomas (n=231) from archived material collected at three university hospitals in Sweden were stained for cyclin D1 and microscopically evaluated with regard to cyclin D1-expression. The lymphomas studied included 11 of anaplastic subtype, 4 of immunoblastic subtype, 6 of T-cell/histiocyte rich subtype and the remainder of centroblastic subtype. 10 (4%) of the 231 cases showed nuclear positivity for cyclin D1. 9 of these were of centroblastic subtype and 1 of anaplastic subtype. All of the cyclin D1 positive cases were negative for CD5. The cyclin D1-positive cases were further subjected to fluoroscence in situ hybridization to determine the presence of the t(11;14) translocation. One of these cases (of centroblastic subtype) showed a hybridization pattern consistent with the t(11;14) rearrangement. Two other cases (one of centroblastic subtype and one of anaplastic subtype) displayed a fluoroscence pattern with amplified cyclin D1 signals in the absence of evidence of the t(11;14) translocation, possibly related to CYCLIN D1 gene amplification. Thus, contrary to the current view, there appears to exist a certain number of cyclin D1-positive and CD5-negative diffuse large B-cell lymphomas some of which display amplified cyclin D1 signals or the t(11;14) translocation. The results emphasize the importance of CD5 and cyclin D1 immunostaining in routine diagnostics of diffuse large B-cell lymphomas. Whether CD5-negative cyclin D1-positive lymphomas with the t(11;14) translocation and classic centroblastic morphology should be classified as mantle cell lymphomas or diffuse large B-cell lymphomas remains debatable.
International T-Cell Lymphoma Classification Project: Angioimmunoblastic T-Cell Lymphoma.
Authors: Ruediger Thomas , Coiffier Bertrand , Weisenburger Dennis D. , Federico Massimo
Published: October 2019

Peripheral T-cell lymphomas are rare diseases: therefore, the International T-cell Lymphoma Project was undertaken to compare lymphomas at different sites throughout North America, Asia and Europe. Within this project, 243 angioimmunoblastic T-cell lymphomas (AILTs) were diagnosed which made up 21% of all peripheral T-cell lymphoma (PTCL). At presentation, generalized lymphadenopathy was noted in 76% of the patients. Interestingly, three patients presented with extranodal disease only. Among the skin symptoms, erythroderma was the most frequent (21% of patients). Hemolytic anemia was seen in 13% and dysproteinemia occurred in 50%, and among these monoclonal serum immunoglobulin was seen in 8% of the patients. Anemia, hypergammaglobulinemia and elevated LDH were significantly more frequent in AILT than in PTCL-unspecified. Similarly, patients with AILT had a significantly higher frequency of high stage disease (89% of the patients were stage 3 or 4), as well as worse prognostic indices. Despite this, their 5-year overall (33%) and failure-free survivals (18%) were similar to patients with PTCL-unspecified. Treatment was usually administered in combination with anthracycline. A few factors at presentation were prognostic for outcome, including the PIT (prognostic index for T-cell lymphoma; Gallamini et al.: Blood. 2004 Apr 1;103(7):2474–9), age, B-symptoms and performance status. The IPI, however, was not prognostic. Controlling for the PIT, a platelet count <150.000 μl was prognostic for overall survival whereas B-symptoms were prognostic for failure-free survival. In conclusion, AILT is an aggressive disease for which the optimum treatment has not yet been developed.
Adult T-Cell Leukemia/Lymphoma: A Clinicopathologic Study of 126 Cases from the International T-Cell Lymphoma Project.
Authors: Suzumiya Junji , Ohshima Koichi , Tamura Kazuo , Karube Kennnosuke , Uike Naokuni , Armitage James Olen O.
Published: October 2019

To assess the clinicopathological applicability and reproducibility of the new WHO classification against peripheral T/NK-cell lymphomas, International T-cell Lymphoma Study Group was organized. Our task is to show the clinicopathologic and therapeutic results of ATLL from International T-cell Lymphoma Project. In the study, 1159 cases of T/NK-cell lymphomas, excluding mycosis fungoides and Sezary syndrome, were collected from the 21 centers in 13 countries of North America, Europe and Asia. All cases were reviewed by expert hematopathologists.

ATLL was diagnosed in 126 (10.9%) patients. It was rare in North America (2.0%), and Europe (1.5%), but frequent in Asia (24.8%). There were clinical subtypes of ATLL from smoldering, chronic, lymphoma to acute type, and 87% of the patients fell into lymphoma type. The median age was 60 (range; 15–92) and male to female ratio was 1. B symptoms, PS 2–4(ECOG) and two or more extranodal sites were found in 39 (31%), 29 (23%) and 43 (34%) patients, respectively. Bone marrow involvement was present in 28% of the patients. Stage I/II was seen in 10% of the patients, stage III in 18% and stage IV in 73%. There were 23 (19%) patients in score 0/1, 41 (33%) in score 2, 40(32%) in score 3 and 20(16%) in score 4/5 of International Prognostic Index (IPI). There were 20 (17%) patients with less than 150x103/ml of platelet counts, 107 (85%) with over 20% transformed T cells in absolute lymphocyte counts, and 113 (94%) with over 20% Ki-67 positive cells. The 5- year-overall survival (OS) and failure free survival (FFS) were 14.4% and 12.1%, respectively. The significant prognostic factors for OS were B symptoms (p=0.018), PS (p=0.0022), stage (p=0.019), extranodal sites (p=0.045) and IPI (p=0.019). By using the multivariate analysis, the important prognostic factors were found to be B symptoms, platelet counts (<150x103/ml), transformed T-cell counts (>20%), and Ki67+ cells (>20%). However, the Cox model analysis showed that no significant prognostic factors were found after adjusting by IPI. For treatment, the combination chemotherapy using anthracycline as an initial therapy was given in 109 of 120 (91%) treated patients. The overall and complete response rates were 69% and 33%, respectively. It is unfortunate that 71 (63%) patients had relapsed. Many drugs were used including prednisone 105 (88%), vincristine 102 (85%), cyclophosphamide 92 (77%), doxorubicin 77 (64%), etoposide 53 (44%), mitoxantrone 14 (12%), cisplatin 11 (9%), methotrexate 8 (7%) , bleomycin 6 (5%) and cytarabine 5 (4%). There was no difference in OS and FFS according to the treatment arms includ.
DA-EPOCH-R Is Highly Effective in Both BCL-6+ and BCL-6− Untreated De Novo Diffuse Large B-Cell Lymphoma (DLBCL): Study Update and Analysis of Survival Outcomes for Multiple Biomarkers.
Authors: Wilson Wyndham H. , Dunleavy Kieron , Pittaluga Stefania , Grant Nicole , Shovlin Margaret , Steinberg Seth , Raffeld Mark , Staudt Louis , Jaffe Elaine S. , Janik John
Published: October 2019

Gene expression profiling has yielded a new molecular taxonomy of DLBCL in which 3 subtypes are distinguished; germinal center B-cell (GCB) subtype, derived from a germinal center B-cell; activated B-cell (ABC) subtype, derived from a post-germinal center B-cell; and primary mediastinal B-cell (PMBL) subtype, likely derived from a “thymic” B-cell. To define the effect of pathobiology on treatment outcome, immunophenotype has been used as an “approximate surrogate” biomarker for molecular taxonomy. Recent studies suggest that rituximab (R) benefit is primarily in BCL-6− (Winter et al Blood107:4207, 2006) and BCL-2+ (Mounier et al Blood101:4279, 2003) DLBCL, both more frequent in post-germinal center subtypes. Conversely, R-CHOP appears equivalent to CHOP in BCL-6+ DLBCL, which comprise up to 75% of all cases. We analyzed the outcome of DA-EPOCH-R using the immunophenotypic biomarkers BCL-6, BCL-2, MIB-1, CD10, MUM-1, and GCB vs ABC subtypes by the method of (Hans et al Blood 103:275, 2003). Patients had untreated de novo DLBCL excluding PMBL, stage II-IV, HIV-, and adequate organ function. No patients received radiation. Characteristics of 72 patients include median (range) age 50 (19–85); stage III/IV 69%; and High Intermediate/High International Prognostic Index (3–5) (IPI) 40%. Of 71 evaluable patients, response is CR/CRu 94%. At the median potential follow-up of 43 months, PFS is 82% and OS is 79% (Fig 1). PFS by Low/Low Intermediate (0–2) and High Intermediate/High (3–5) IPI is 93% and 64%, respectively, at 43 months. PFS by BCL-6 (Fig 1), and BCL-2 and GCB vs. ABC (Fig 2) is shown. PFS of DLBCL with MIB-1 < and > 80% was 92% and 81%, respectively. There was no significant difference in survival outcome by CD10 or MUM-1 expression (data not shown). Distribution by immunophenotype is shown below for patients with available tissue.

Biomarker Distribution Biomarker Number Percent BCL-6 Neg 14 24 BCL-6 Pos 44 76 BCL-2 Neg 24 41 BCL-2 Pos 35 59 GCB 34 67 ABC 17 33 MIB-1 < 80% 13 33 MIB-1 > 80% 39 67 CD10 Neg 36 63 CD10 Pos 21 37 Mum-1 Neg 30 64 Mum-1 Pos 17 36

In conclusion, DA-EPOCH-R appears equally effective among all biomarker subgroups. Of note, DA-EPOCH-R is highly effective in BCL-6+ DLBCL, in which R does not appear to be very useful, suggesting the DA-EPOCH regimen itself may be highly effective in this group. The CALGB has initiated a Phase III randomized study of R-CHOP vr. DA-EPOCH-R to determine if DA-EPOCH-R represents a treatment advance. Gene expression profiling will be performed to assess the effect of the new molecular taxonomy and tumor biology on outcome.

Figure Figure
Predictive Value of Clonal Peripheral Blood γTCR Rearrangement in Patients with Peripheral T-Cell Lymphoma.
Authors: Schuetzinger Christian , Esterbauer Harald , Skrabs Cathrin , Uffmann Martin , Raderer Markus , Hauswirth Alexander , Gaiger Alexander , Mannhalter Christine , Dieckmann Karin , Wagner Oswald , Jaeger Ulrich
Published: October 2019

Peripheral T-cell lymphoma is a rare heterogeneous disease with generally poor outcome. Known risk factors include the International Prognostic Index (IPI) and b2 microglobulin. However, there is little information on molecular risk factors. In this single center analysis we have prospectively studied the prognostic value of a clonal T-cell receptor rearrangement (TCR) determined by conventional PCR for the γTCR (sensitivity 1 in 102) from DNA from peripheral blood MNC. Thirty nine consecutive patients diagnosed between 1987 and 2006 were assessed for clonality at diagnosis.

Patient characteristics: Median age 53 years (range 28–89). M:F = 29 vs. 10. Histological diagnosis included: PTCL-NOS 7 (18%); Large cell 1 (2.5%); Large + medium-sized 5 (13%); Pleomorphic 7 (18%); Small cell 3 (8%); Small+medium mixed cell 1 (2.5%); Lennert’s type 4 (10%); Hepatosplenic 3 (8%); Intestinal 4 (10%); Kimura’s disease: 1 (2.5%);

AILD: 3 (8%). Assessment of γTCR showed monoclonality in 24 (62%) and polyclonality in 15 (38%) patients. Patients with a monoclonal TCR had higher clinical stages, higher LDH levels, higher IPI scores, and higher ß2 microglobulin levels.

Clinical stage: CS I–III (clonal 42%; polyclonal 58%) vs. CS IV (clonal 93%; polyclonal 7%). LDH above 240 U/L (clonal 78%; polyclonal 22%). IPI 0,1 (clonal 44%; polyclonal 56%), IPI 2 (clonal 75%; polyclonal 25%), IPI 3 (clonal 87%; polyclonal 13%), IPI 4,5 (clonal 60%; 40%). ß2 microglobulin above 2 mg/L (clonal 68%; polyclonal 32%). Patients with a clonal TCR tended to be older (57 vs. 49 yrs.). No major differences were found for age and sex. Induction treatment consisted of polychemotherapy (CHOP-like=34), prednisone (1), radiation (1). 3 patients received no therapy. Nine patients received subsequent involved field radiation and 6 pts. had an autologous stem cell transplant. Significant differences were observed in terms of disease outcome: The presence of a clonal γTCR (n=24) was associated with low clinical remission rates: CR 38%, PR 21%, SD 8%, PD 33%, relapse rate 38%. Remission rates in patients with polyclonal rearrangements (n=15) were: CR 66%, PR 20%, SD 7%, PD 1%, relapse rate 47%. The estimated 3-year overall survival for the monoclonal group was 26% vs. 69% for the polyclonal group (P=0.01). Importantly, patients with CS I–III and a monoclonal γTCR rearrangement (n=10) had a similarly poor survival when compared to the monoclonal group with CS IV (n=14) (18 months vs. 12 months; P=0.23). This indicates that γTCR positivity is of particular predictive value in patients with CS I–III. The data from this unique cohort suggest that γTCR-PCR is a useful prognostic tool in patients with peripheral T-cell lymphoma.

Figure Figure
Loss of Heterozygosity in Acute Myeloid Leukemia with Normal Karyotype.
Authors: Traikov Sofia , Steudel Christine , Oelschlaegel Uta , Schaich Markus , Ehninger Gerhard , Thiede Christian
Published: October 2019

Loss of heterozygosity (LOH) is detectable in many forms of cancer, including leukaemia. LOH contributes to tumorigenesis through the loss of one of the two alleles of one tumor suppressor gene at a given locus, caused by deletion or uniparental disomy (UPD). In this study, we analysed 146 patients with primary AML, using high-density single nucleotide polymorphisms (SNPs) arrays. Leukemic cells and healthy T-cells from each patient were obtained using FACS-Vantage cell sorting. The genome wide SNP analysis was carried out using Human Mapping 10K Arrays (Affymetrix). So far LOH was identified in 27 cases with normal karyotype (18.5%). Potential LOH regions were confirmed by analysis of short tandem repeat (STR) markers using DNA samples from T-cells and blasts. In 21 of these cases, STR-analysis of T-cells representing the corresponding tumor-free material confirmed the regions of UPD. This aberration affected different chromosomes, but most frequently chromosome 2, 6, 11, 13, 21, and covered between 11.5 and 88 Mb. In the remaining 6 LOH cases, long stretches of homozygosity occurring at identical positions in the T-cells and in the leukemic blasts were found. These cases were detected and separated from those with UPD.

So far, correlation of clinical variables and LOH results in this cohort did not indicate significant differences based on age, WBC-counts, sex and FAB-subtype. Additionally, we did not detect any proven correlation between FLT3-ITD, MLL-PTD, NPM1 mutations and UPD. Interestingly, 80% of the patients with UPD that had achieved CR after chemotherapy relapsed within 6 months. In contrast, the rate of relapse in patients without UPD was only 54% . Furthermore, we were able to confirm the relationship between UPD and gene aberrations. We demonstrated that the UPD cases found in chromosomes 21, 19, and 11 were correlated with point mutations in the RUNX1, CEBPA and WT1 genes, respectively. Furthermore, AML cases with and without UPD showed different gene expression profiles, revealing different expression levels for some genes involved in double strand brake repair.

In conclusion, the use of the SNP array technology allowed the identification and mapping of LOH in 18.5% of AML patients with normal karyotype. Our data also underline the necessity of analysing tumour-free material in order to confirm the somatic origin of the alteration. Furthermore, our results indicate a higher relapse rate in patients with UPD.