bluuebunny/crossref_metadata_2025_split huggingface dataset:
Statins Potentiate the Cytotoxic Effect of ABT-199 in Diffuse Large B Cell Lymphoma
Authors: Fruman David A. , Lee Jong-Hoon Scott , Vo Thanh-Trang T , Bhatt Shruti , Schatz Jonathan H. , Letai Anthony 
Published: October 2019

Abstract
BCL-2 is a key pro-survival protein that is highly expressed in many leukemias and lymphomas. ABT-199 (venetoclax) is a small molecule inhibitor of BCL-2 that has demonstrated impressive responses in chronic lymphocytic leukemia (CLL) leading to FDA approval for second line treatment of patients with 17p deletion. However, other hematologic malignancies are less responsive to ABT-199 as a single agent, suggesting that combinations of targeted therapies may be required to elicit more promising responses. We have investigated the potential of combining ABT-199 with HMG-CoA reductase (HMGCR) inhibitors (statins), which have known anti-cancer potential in hematologic malignancies. Using multiple chemically distinct statin compounds, we observed profound synergistic induction of apoptosis when combined with ABT-199 in both human diffuse large B cell lymphoma (DLBCL) as well as acute myeloid leukemia (AML) cell lines. This synergy was also seen in primary murine B lymphoma cells over-expressing MYC and BCL-2. Importantly, addition of exogenous mevalonate completely rescued cells from the combination, confirming on-target efficacy of HMGCR inhibition. Using BH3 profiling, we found that simvastatin significantly primed lymphoma cells for undergoing apoptosis (termed mitochondrial priming). Notably, the degree of priming correlated with its ability to synergize with ABT-199, suggesting that BH3 profiling may be used to predict patient responses. The combination did not synergize to kill normal human peripheral blood mononuclear cells from healthy donors, suggesting that statins may selectively prime cancer cells for apoptosis. Mechanistic studies support the hypothesis that statins synergize with ABT-199 by suppressing protein prenylation, particularly protein geranylgeranylation. In support, the addition of exogenous geranylgeranyl pyrophosphate (GGPP) completely rescued cells from the effects of simvastatin. Furthermore, selective inhibition of protein geranylgeranyl transferase (GGT) increased priming and was sufficient to recapitulate the effects of simvastatin in combination with ABT-199. Statins and GGT inhibitors increased the mitochondrial abundance of a subset of BH3-only pro-apoptotic proteins. Lastly, we have identified Rap1A de-prenylation as a marker of pharmacodynamic response to statins in vivo. Thus, this project highlights a novel combination for use in aggressive lymphomas, establishes its efficacy and tolerability using preclinical models, and provides proof-of-concept to warrant investigation of its clinical potential.

Disclosures
Letai: AbbVie: Consultancy, Research Funding; Astra-Zeneca: Consultancy, Research Funding; Tetralogic: Consultancy, Research Funding.
Utilization of Mate-Pair Sequencing to Characterize Complex and Novel TCF3 Translocations
Authors: Rowsey Ross , Smoley Stephanie A. , Williamson Cynthia , Sukov William , Greipp Patricia , Ketterling Rhett P. 
Published: October 2019

Abstract
Rearrangements involving TCF3 (E2A) at 19p13.3 are a common cytogenetic abnormality in B-lymphoblastic leukemia/lymphoma (B-LL) across all age groups accounting for at least 5% of B-LL. The most common translocation partner, PBX1 at 1q23, is associated with an unbalanced der(19) or a balanced t(1;19) in 80% vs. 20% patients, respectively, and results in a PBX1/TCF3 fusion transcriptional activator. Several other rearrangement partners have been described, and as such TCF3 is approaching the dubious status of a "promiscuous" gene partner. Clinically, our laboratory uses a dual-color, dual fusion PBX1/TCF3 FISH probe to characterize the most common gene fusion; however, this FISH strategy also allows for the identification of atypical disruptions of the TCF3 gene region, and may suggest a gene partner based on abnormal metaphases/metaphase-FISH.
Interrogation of our pediatric leukemia database (ages 0-30) identified 219 patients with abnormal TCF3 FISH results. We selected 23 samples for mate-pair evaluation to test the methodology, characterize novel rearrangements and identify new TCF3 fusion partners. In total, 19 patient samples were successfully sequenced by Mate-Pair, including 13 samples with variants of the 1;19 translocation. Of the samples with TCF3/PBX1 fusion, 3 patients demonstrated novel insertional translocation events characterized by mate-pair. A total of 6 samples with undefined/unknown TCF3 partner genes were evaluated. The mate-pair results identified 5 samples with a t(12;19)(p13.31;p13.3), resulting in TCF3/ZNF384 fusion. Interestingly, two of the ZNF384 rearrangements were observed in the unbalanced form, suggesting a similar mechanism related to the der(19)t(1;19) regularly observed in the common PBX1-TCF3 rearrangement. From a karyotype perspective, since these rearrangements result in only a small portion of chromosome 12p translocated onto distal 19p, this rearrangement would "cryptic" by conventional karyotyping. One patient sample demonstrated a new TCF3 gene fusion partner (TEF), associated with a t(19;22)(p13.3;q13.2).
Overall, our results demonstrate the utility of mate-pair evaluation as an adjunct methodology in the identification of novel TCF3 rearrangements in B-LL. We have successfully expanded the application of mate-pair to characterize other novel/cryptic abnormalities in various hematologic and oncologic neoplasms encountered in our laboratory.

Disclosures
No relevant conflicts of interest to declare.
TULA-2 Inhibits the Platelet FcγRIIA-Syk Pathway and Heparin-Induced Thrombocytopenia in Mice
Authors: Zhou Yuhang , Abraham Shaji , Edelstein Leonard C. , Dangelmaier Carol , Tsygankov Alexander , Kunapuli Satya P. , Bray Paul F. , McKenzie Steven 
Published: October 2019

Abstract
Heparin-induced thrombocytopenia (HIT) is a life-threatening disease in which IgG antibodies against the heparin-PF4 complex activate platelets via FcγRIIA. We previously reported that TULA-2 serves as a negative regulator of FcγRIIA pathway by dephosphorylating Syk in HEL cells. To further investigate the effect of TULA-2 on the FcγRIIA pathway and HIT, we crossed TULA-2-/- with FcγRIIA+/+ mice. Ablation of TULA-2 resulted in hyperphosphorylation of Syk, LAT, and PLCγ2 in platelets after FcγRIIA activation. Integrin activation, calcium mobilization, and P-selectin exposure were also enhanced in TULA-2-/- murine platelets compared to TULA-2+/+. Further, anti-GPIX antibody-induced HIT-like thrombocytopenia and thrombin generation were also augmented in TULA-2-/- mice (Figure A). We also found that decreased TULA-2 level shortened tail-bleeding time in mice (Figure B), suggesting a role of TULA-2 in physiological hemostasis.
Additionally, TULA-2 KO and WT platelets did not show significant differences in platelet spreading and clot retraction, indicating that outside-in signaling is not affected by the absence of TULA-2. At the protein level, TULA-2 heterozygous knockout (TULA-2+/-) platelets express 50% as much protein as their wildtype counterparts. Interestingly, TULA-2+/- mice showed significantly increased platelet reactivity and more severe thrombocytopenia in vivo compared with TULA-2+/+ mice. Together the data indicate that not only the absence of TULA-2, but also the relative level of TULA-2 expression modulate FcγRIIA-mediated platelet reactivity, HIT pathogenesis, and hemostasis. Considering that TULA-2 is also a key regulator in platelet GPVI pathway, measuring TULA-2 expression may be a valuable predictor for HIT susceptibility and platelet reactivity in general.
Figure. Figure.

Disclosures
No relevant conflicts of interest to declare.
IGH/MYC Translocation in Burkitt Lymphoma Is Associated with BRCA2 Deficiency and Synthetic Lethality By PARP1 Inhibitors
Authors: Maifrede Silvia , Martin Kayla , Podszywalow-Bartnicka Paulina , Katherine Sullivan , Langer Samantha , Nejati Reza , Dasgupta Yashodhara , Gritsyuk Daniel , Nieborowska-Skorska Margaret , Lupey Lena , Piwocka Katarzyna , Wasik Mariusz A , Tempera Italo , Skorski Tomasz 
Published: October 2019

Abstract
Burkitt lymphoma/leukemia (BL) is a highly aggressive mature B-cell neoplasm characterized by chromosomal rearrangements of the c-myc oncogene resulting in the overexpression of MYC transcription factor. The most common translocation is the t(8;14)(q24;q32) (85% of all cases), which involves MYC and IGH loci to generate IGH/MYC. Deregulation of MYC, a potent proto-oncogene and transcriptional regulator contributes to lymphomagenesis through alterations in cell cycle regulation, and cell differentiation, apoptosis, adhesion, and metabolism.
BL treatment consists of high-intensity chemotherapy protocols that include cyclophosphamide, cytarabine (AraC) and doxorubicin. Current therapies have achieved a very favorable outcome resulting in complete remission in 75% to 90% of BL patients and a survival rate of 70% to 80%. However the current treatment for BL is suboptimal in elderly patients or patients with advanced-stage diseased, in the setting of HIV infections, as well as in the setting of relapsed disease. Therefore new therapeutic strategies are necessary to improve the outcomes in BL diseases in the poor prognosis patients.
It has been reported that overexpression of MYC caused accumulation of potentially lethal DNA double-strand breaks (DSBs), which can modulate the response of tumor cells to genotoxic treatment. Therefore, we examined the consequences of DSBs accumulation in IGH/MYC-positive BL cells. Here we show that untreated and cytarabine (AraC)-treated IGH/MYC-positive BL cells accumulate high number of potentially lethal DNA double-strand breaks (DSBs) and display downregulation of BRCA2 tumor suppressor protein, which is a key element of homologous recombination - mediated DSB repair. BRCA2 deficiency in IGH/MYC-positive cells was associated with hypersensitivity to PARP1 inhibitors (olaparib, talazoparib) used alone or in combination with cytarabine in vitro. Moreover, talazoparib exerted a therapeutic effect in NGS mice bearing primary BL xenografts.
In conclusion, we postulate that BRCA2 deficiency may predispose BL cells to synthetic lethality triggered by PARP1 inhibitor, such as recently FDA approved olaparib. Moreover, PARP1 inhibitor may be useful for the treatment of other malignancies associated with deregulation of MYC, including diffuse large B-cell lymphoma (DLBCL) and ALK-positive LBCL.

Disclosures
Wasik: Gilead Sciences: Equity Ownership; Seattle Genetics: Honoraria; Novartis: Research Funding; University of Pennsylvania: Patents & Royalties: NPM-ALK as an omncogene; University of Pennsylvania: Patents & Royalties: CAR T-cells; Gilead Sciences: Research Funding; Pharmacyclics: Research Funding.
The Role of UBR5 Mutations in the Pathogenesis of Mantle CELL Lymphoma
Authors: Kutovaya Olga , Hung Stacy , Christopher Hughes , Gascoyne Randy D , Gregg Morin , Steidl Christian 
Published: October 2019

Abstract
Intro: Mantle cell lymphoma (MCL) accounts for 6% of non-Hodgkin lymphomas and represents a particularly challenging disease with patient outcomes inferior to most other lymphoma subtypes. Using targeted capture sequencing of MCL biopsy samples, we recently reported frequent mutations (18%) in UBR5, a gene encoding an E3 ubiquitin-protein ligase that has not been previously implicated in lymphomagenesis. All mutations were clustered within 100bp in or around exon 58 of UBR5 and truncate the reading frame or change a key lysine residue. These mutations are predicted to result in the loss of the conserved cysteine residue in the HECT-domain, which is responsible for binding the ubiquitin co-factor. The recurrence and clustering of UBR5 mutations suggest their critical pathogenic involvement in a subgroup of MCL that might be therapeutically targetable. The aim of this study is to determine UBR5 mutation-associated proteome changes and altered cell signaling.
Methods: As seen in MCL patients, mutations in exon 58 of UBR5 were introduced to three MCL cell lines (Granta-519, Jeko-1, and Mino) using the CRISPR-Cas9 genome engineering tool. First, mass spectrometry-based immunoprecipitation proteomics (IP-MS) was employed to identify differences in UBR5 interacting partners between UBR5 mutant and wildtype (WT) cells. Candidate UBR5 interacting proteins were validated by flow cytometry, western blotting, co-immunoprecipitation, and immunofluorescence. Next, global proteomes of UBR5 mutants and WT were analyzed by Tandem Mass Tag (TMT)-based mass spectrometry quantification to identify proteins with differential expression due to the UBR5 mutations.
Results: The IP-MS analysis of WT vs UBR5 mutants revealed histone and cell cycle control proteins as candidate differential UBR5 interacting proteins (p<0.05). Particularly, histones H1, H4, and H2AFX, as well as the cell cycle genes CDC5L, BUB3, MAP4, RAD50 and CDK11B were identified as candidate UBR5 interacting partners. The global proteome analysis identified a set of differentially expressed genes (mutant vs wt; p<0.05) that are common among the MCL cell lines with the same direction of change. Gene ontology analysis of this set revealed DNA damage response, chromosome organization, and cell cycle response pathways as the predominant pathways affected. Moreover, our preliminary functional studies indicate constitutive phosphorylation of H2AFX in UBR5 mutants vs WT in line with the role of UBR5 in DNA damage response.
Conclusions: Our results are consistent with UBR5 functioning as a key regulator of cell signalling and strongly suggest UBR5 as a novel regulator of histone modifications and DNA damage response. These findings provide an experimentally valid platform for further functional investigation and testing of target therapies for MCL harbouring UBR5 mutations.

Disclosures
No relevant conflicts of interest to declare.
Identification of Imatinib-Sensitizing Genes in Chronic Myeloid Leukemia with a Genome-Scale Crispr Knock-out Screen
Authors: Lewis Matthieu , Prouzet-Mauleon Valerie , Richard Elodie , Turcq Beatrice , Iggo Richard , Mahon Francois-xavier 
Published: October 2019

Abstract
Background: Resistance to tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia (CML) can either originate from mutations in the BCR-ABL1 gene, which are mostly well characterized, or emerge from unknown alternative mutations elsewhere in the genome. Small hairpin (sh)RNA screens have been used to discover such genes but are becoming limited due to sup-optimal protein depletion and non-reliable off-target effects. More efficient screening techniques in human cells are now available as a result of the increasing understanding of the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) /Cas9 system.
Aims: Our goal is to uncover imatinib (IM)-sensitizing genes that cause IM resistance when knocked-out. Characterizing these genes may help understand the mechanisms of IM uptake, metabolism, degradation and/or activity in CML cells. Additionally, we also expect to unveil alternative, BCR-ABL1 independent, oncogenic pathways in CML cells.
Methods: In order to find other genes involved in IM resistance, we performed a genome scale CRISPR knock-out (GeCKO) screen, which contains 121,413 sgRNAs that target 20,914 protein coding genes and miRNAs. We transduced one sgRNA per cell and challenged the K562-GeCKO cell pool to IM selection. We compared the abundance of sgRNAs between pre/post-IM treatment by next generation sequencing (NGS).
Results: After IM selection, the sgRNAs from surviving cells were identified by NGS and unveiled potential IM-sensitizing genes. The most enriched sgRNAs (FDR < 0.01) targeted genes involved in transcriptional (KLF1, MED24) and translational (EIF2AK1, UBE2M) regulation, apoptosis (BAX, BCL2L11) and cell cycle regulation (BAP1, SPRED2). Subsequent screens on LAMA84 cells are currently underway in order to validate our findings. Additionally, the establishment of individual gene knock-out cell lines are in progress in order to fully understand the role of each gene in IM resistance.
Summary/Conclusion: Using a CRISPR knock-out screen, we produced a list of 19 genes (FDR < 0.05) that may play a role in IM resistance. Encouragingly, a subset of these genes (BAX, BAP1, BCL2L11 and SPRED2) have already been correlated to CML progression and/or TKI resistance in the past. We aim to bolster our findings by establishing individual gene KO cell lines and study resistance in LAMA84 cells. The utilization of CRISPR libraries may not only help understand TKI resistance in CML, but also help identify numerous novel genes involved in drug resistances for a myriad of different diseases.

Disclosures
Mahon: ARIAD: Honoraria; PFIZER: Honoraria; BMS: Consultancy, Honoraria; NOVARTIS PHARMA: Consultancy, Honoraria, Research Funding.
Dysregulation of ANKRD11 Influenced Hematopoisis By Histone Acetylation-Mediated Gene Expression in Myelodysplastic Syndrome
Authors: Zhao Youshan , Xu Feng , Guo Juan , Zhao Sida , Chang Chunkang , Li Xiao 
Published: October 2019

Abstract
Background and Object In addition to histone deacetylation, the importance of histone over-acetylation induced oncogene transcription in initiation and progression of myelodysplastic syndrome (MDS) has been proposed recently. Our previous whole-exome sequencing identified a new somatic mutation, ANKRD11, an important factor in histone acetylation regulation. Its roles in MDS pathophysiology need to be clarified.
Methods The next generation target sequencing (Including ANKRD11) was carried out in 320 patients with MDS using the MiSeq Benchtop Sequencer. ANKRD11 mRNA expression in bone marrow of MDS was measured by real-time PCR. Loss and gain of function assay were carried out in myeloid cell lines K562, MEG-01£¬or SKM-1 to observe the influence on cell proliferation and differentiation . The levels of histone acetylation at H3 and H4 were detected by Western blot.
Results Target sequencing in a cohort of 320 MDS patients identified 14 of ANKRD11 mutations (4.38%, Fig.1), which were confirmed by Sanger sequencing. Meanwhile, no ANKRD11 mutations in 100 normal controls were defined. ANKRD11 mutations occurred frequently in exons 10 and 9. The mRNA expression levels of ANKRD11 were significantly decreased in MDS patients, especially in ANKRD11mutant patients (Fig.2). ANKRD11 knockdown in K562 and MEG-1 resulted in growth inhibition, cell cycle arrest and erythroid/megakaryocytic differentiation retardant. In MDS cell line SKM-1, the arrested differentiation was rescued by over-expression of ANKRD11. Consistent with a role for ANKRD11 in histone acetylation, ANKRD11 KD increased acetylation of histones H3 and H4 at H3K14 and H4K5 and resulted in the upregulation of genes involved in differentiation inhibilation (SOX6, P21, et al). Finally, the ANKRD11 KD-mediated influence on cell proliferation and differentiation were reversed by inhibiting histone acetyltransferase activity.
Conclusion Our assay defined that ANKRD11 was a crucial chromatin regulator that suppress histone acetylation and then decrease gene expression during myeloid differentiation, providing a likely explanation for its role in MDS pathogenesis. This study further support histone acetylase inhibitor as a potential treatment in MDS.
Figure ANKRD11mutation distribution (a) and coexist with other mutations (b). Figure. ANKRD11mutation distribution (a) and coexist with other mutations (b). Figure The mRNA expression levels of ANKRD11in our MDS (A, C) subset and GEO data (B). Figure. The mRNA expression levels of ANKRD11in our MDS (A, C) subset and GEO data (B). Changes of histone acetylation in ANKRD11-KD cell line (MEG-01). ANKRD11 KD significantly increased acetylation of histones H3 and H4 at H3K14 and H4K5. Changes of histone acetylation in ANKRD11-KD cell line (MEG-01). ANKRD11 KD significantly increased acetylation of histones H3 and H4 at H3K14 and H4K5.

Disclosures
No relevant conflicts of interest to declare.
A Method for the Measurement of Red Cell Dimensions and Calculation of Mean Corpuscular Volume and Surface Area
Authors: HOUCHIN DONALD N. , MUNN JOHN I. , PARNELL BENJAMIN L. 
Published: October 2019

Abstract
1. A photomicrographic method was devised for the measurement of the dimensions of red cells in rouleaux in plasma. With normal blood valid conclusions were drawn from 80 cell measurements with a high degree of confidence.
2. The mean corpuscular volume and the surface area were computed from these dimensions by formulae which treated the cell as a spheroid.
3. The dimensions, mean corpuscular volume and surface area of normal human red cells were as follows: 8.28 µ diameter, 1.71 µ thickness, 82 µ3 volume and 134 µ2 surface area.
4. The computed mean corpuscular volume was in agreement with the refractometric determination.
Plasma Lactic Dehydrogenase and Phosphohexose Isomerase in Leukemia
Authors: BLANCHAER M. C. , GREEN P. T. , MACLEAN J. P. , HOLLENBERG M. J. 
Published: October 2019

Abstract
Two enzymes, lactic dehydrogenase (LD) and phosphohexose isomerase (PHI), were measured in the plasma of 30 patients with leukemia and compared with the findings in 66 control subjects. Abnormally elevated PHI levels were found in both acute and chronic myelocytic leukemia, but not in lymphocytic leukemia. The plasma LD was increased above normal in acute and chronic myelocytic leukemia, in acute lymphocytic, but not in chronic lymphocytic leukemia. Both enzymes were normal or only slightly raised in three patients with the aleukemic type of the disease. Hemolytic anemia in seven leukemic patients was associated with high plasma LD values in the presence of relatively low PHI levels.
Results of serial enzyme studies from the time of diagnosis until death indicated that both plasma enzymes, but especially the LD, usually reflected changes in the course of the disease-falling during remissions and rising during relapses. In most cases this enzyme paralleled the leukocyte level but occasionally indicated the onset of a relapse or remission before the white cell count had begun to change.
Critique of the Concept of the "Pre-L.E." Cell
Authors: PASCHER FRANCES , BOROTA ALEXANDER 
Published: October 2019

Abstract
1. Cells with morphologic features apparently identical with the so-called pre-L.E. cell were demonstrated in L.E. preparations in healthy subjects and in individuals suffering from diverse cutaneous and visceral disorders, as well as in patients with systemic lupus erythematosus.
2. These cells appear to be decomposition forms without diagnostic significance, resulting from nonspecific autolytic process that sets in and progresses in the course of setting up the L.E. preparation.
Inhibition of Leukocyte Agglutination by Serum from Patients with Systemic Lupus Erythematosus: A Manifestation of the L.E. Cell Phenomenon
Authors: LEE STANLEY L. , Schultz Florence 
Published: October 2019

Abstract
Dying leukocytes suspended in normal serum tend to agglutinate spontaneously into a firm "clot." This phenomenon is associated with cell lysis and the formation of long fibrous strands of DNA-protein. It seems likely that nuclear DNA-protein actually combines in some way with a fraction of the serum proteins (possibly mucoprotein).
Such leukocyte agglutination is inhibited by serum containing the L.E. cell factor. LAI is associated with nuclear changes which are characteristic of the L.E. cell phenomenon, and it is most probable that it is an expression of the L.E. cell phenomenon.
An understanding of the chemistry of LAI may lead to useful information as to the fundamental mechanism of the L.E. cell phenomenon. Without such understanding, LAI has been used to obtain collateral information concerning the L.E. cell phenomenon: namely, to rule out any important role for DNA-ase, and to confirm the importance of blood platelets. It has also been used to prepare "pure cultures" of L.E. bodies for cytochemical study.
On a clinical level, LAI may prove a useful adjunct to the standard L.E. cell tests. However, its lower level of sensitivity and occasional "false positive" reactions make it inadequate as a primary test in systemic lupus.