bluuebunny/crossref_metadata_2025_split huggingface dataset:
Metastatic Breast Cancer in a Patient with Fanconi Anemia
Authors: Graham Jeffrey , Grenier Debjani , Ponnampalam Arjuna 
Published: October 2019

Abstract
Fanconi anemia (FA) is a rare inherited disorder characterized by progressive bone marrow failure, congenital malformations and a propensity for developing malignancies at an early age. The underlying genetic defect in FA creates a state of cellular hypersensitivity to many traditional chemotherapy agents, making the treatment of malignancies in this population particularly challenging.
We describe a 42-year-old female who presented with a solitary mass in her left breast. Core biopsy revealed an invasive ductal carcinoma that did not express estrogen (ER) or progesterone receptors (PR), but did express human epidermal growth factor receptor 2 (HER2). Staging work-up revealed diffuse skeletal metastatic disease. At her initial consultation with medical oncology, she was discovered to be pancytopenic. Further history revealed a sibling with aplastic anemia and that she had undergone chromosomal breakage testing for FA in the past, which was subsequently confirmed to be positive. She underwent a bone marrow aspirate and biopsy that showed metastatic marrow infiltration by non-hematopoietic cells. In addition there was morphological evidence of dyserythropoiesis and cytogenetic abnormalities on karyotyping, features suggestive of FA. She was initially started on trastuzumab monotherapy. Low dose radiation therapy was added due to local tumor progression. Combined HER2 directed therapy was to be implemented, but was held due to a functional decline in the patient. To date, she has not received definitive genetic testing to determine which FA subgroup she belongs to.
This case highlights two important aspects of FA. The first is the inherent increase in susceptibility to neoplasms in this group, including solid tumors such as breast cancer. The genes associated with FA are involved in deoxyribonucleic acid (DNA) repair pathways, including mutations in the breast cancer susceptibility gene, BRCA2. The second is the heightened sensitivity to the toxic effects of many standard chemotherapy and radiation treatments. This creates unique challenges in the treatment of malignancies in this population and stresses the importance of targeted therapies.

Disclosures
No relevant conflicts of interest to declare.
Calr Gene Mutation in Patients with JAK2-Negative Myeloproliferative Neoplasms
Authors: Jingyan Xu , Zhou Rong-Fu , Chen Bing , Ouyang Jian 
Published: October 2019

Abstract
Methods Genomic DNA from bone marrow and peripheral blood cells were extracted from 40 patients with MPD(male 26,female 14),Aged 13 to 76 years old, JAK2V617F mutation and CALR genetic mutations was identified by PCR- direct sequencing.
Results The number of MPD(PV15,ET25) 40 cases in patients with JAK2V617F mutation rate was 60%,among them the polycythemia vera ( PV) positive rate was 66. 7% ( 10 /15) ,and essential thrombocythemia ( ET ) positive rate was 56% ( 14 /25) . The number of JAK2V617F -negative MPD 16 cases in patients with CALR mutation rate was 12. 5% ( 2 /16),Two of the 11 patients with ET were CALR mutation positive(18.2%). Two novel mutation in CALR exon 9,c.1099_1150del ( p. chr19F12915572-12915623del) was detected in patient Tao c.1099-1151delinsT( p.chr19:12915572_12915624delinsT) was detected in patient Xu. These mutation was absent in the controls,Two novel mutation in CALR have not been reported so far.
Conclusion CALR gene mutations testing helps to JAK2V617F negative MPD diagnosis, makes the MPD early detection and treatment.

Disclosures
No relevant conflicts of interest to declare.
Inhibition of Siah2 Ubiquitin Ligase By Vitamin K3 Attenuates Hypoxia and Blocks K562-R Cells Resistance
Authors: Xu Na , Xiao Yajuan , Zhou Xuan , Li Lin , Huang Fen , Xu Bing , Xiaoli Liu 
Published: October 2019

Abstract
Background and objective: Hypoxia has been shown to favor the self-renewal of murine and human hematopoietic stem cells. Hypoxia as a key feature of the “stem cell niches” in vivo, and studies found that hypoxia modified proliferation and differentiation of chronic myeloid leukemia (CML) stem cells.The E3 ubiquitin ligase Siah2 is an important regulator of the hypoxic response,which has been implicated in the regulation of the hypoxia response, as well as in the control of Ras, JNK/p38/NF-κB, MAPK signaling pathways. In the present study, we identified that SIAH2 induced k562 cells resistance to imatinib by hypoxia-inducible factor (HIF)-1a activated vascular epithelial growth factor (VEGF) pathway in hypoxia micro environment. In this study we show that SIAH2/Hif-1α induced K562 cell remain in G0 stage and resistance to imatinib, and we verified that vitamin K3 (SIAH2 inhibitor) reversed K562-R drug resistance in hypoxia microenvironment.
Methods: We detected Siah2 expression levels in K562-wild type (K562-W) and K562-imatinib-resistance type (K562-R) cell lines by western blot analysis. Those two cell lines were further cultured for 24 h and 48 h under the condition of normal and hypoxia concentration of oxygen (1%, 5%), and treated K562-R with 0A5A15A30mM vitamin K3 for 72 hours in hypoxia concentration, explored cell cycle and apoptosis by flow cytometry (FCM) dyed by Annexin-V; analyzed the expression levels of Siah2, HIF-1α respectively by real-time PCR and western blot.
Results: The protein of Siah2 and HIF-1α was significantly higher in K562-R compared with K562-w cells (P<0.01). Cell cycle analysis showed a 3% increase in K562-W G0/G1 cells in 1% O2 compared with normal O2, and 7% in K562-R. Under hypoxia condition, the cellular apoptotic ratio of K562-R was 5.46%, much less than 11.08% in K562-W cells. G0 cell proportion increased significantly with the long time Hypoxia (P<0.01); After being cultured in 1% oxygen concentration for 24 hours, we confirmed Siah2,HIF-1α were all up regulated in both cell lines, moreover, it was more obvious in K562-R cells. Siah2 protein expression increased along treated with vitamin K3 concentration (0, 5, 15, 30 mM) (P<0.05), on the contrary ,HIF-1α protein expression decreased with vitamin K3 concentration (P<0.05). the proportion of G0 cell was decreased in K562-R cells treated with Vitamin K3 (15mM)for 48h Under the condition of 1% oxygen concentration compared with control group (P=0.02).
Conclusions: Hypoxia up-regulated of Siah2 and Hif-1α expression in K562-R, promoted cell apoptosis and arrested cells in G0 stage, and reduced cell sensitivity to Imatinib. Vitamin K3 could inhibit Siah2 and lowered Hif-1α in K562-R. These findings reveal an effective treatment for the identification of Siah2 inhibitors and would reverse TKI resistance for CML patients.

Disclosures
No relevant conflicts of interest to declare.
Favorable Clinical Outcome and Unique Characteristics in Association with Twist1 overexpression in De Novo Acute Myeloid Leukemia
Authors: Huang Cih-En , Chen Chih-Cheng , Gau Jyh-Pyng , You Jie-Yu , Chen Yi-Yang , Chou Hui-Ju , Lung Jrhau , Yang Muh-Hwa 
Published: October 2019

Abstract
Epithelial-mesenchymal transition (EMT) is a critical process for inducing stem-like properties of epithelial cancer cells. However, the role of EMT inducers in hematologic malignancies is unknown. Twist1, an EMT inducer necessary for cell migration, has recently been found to have transcriptionally regulatory activity on the expression of Bmi1, and these two are capable of promoting tumorigenesis in a synergized manner. Knowing that Bmi1 expression is essential for maintenance of leukemic stem cells, we speculate that Twist1 might govern the pathogenesis of acute myeloid leukemia (AML) development as well.
We used AML cell lines KG1a and THP1 cells, which had low and high expression levels of Twist1, respectively to demonstrate that upregulated Twist1 increased Bmi1 expression in AML. Bmi1 overexpression endued leukemic cells a higher proliferative potential and increased resistance to apoptosis. In primary AML samples, there was strong positive correlation between the expression levels of Twist1 and Bmi1. AML Patients whose leukemic blasts harbored overexpressed Twist1 had a more aggressive clinical phenotype, but they were more likely to have a better clinical outcome after standard therapy. In vitro studies confirmed that Twist1-overexpressing leukemic cells were more susceptible to cytarabine, but not daunorubicin, cytotoxicity.
Our findings suggest that, in a subset of AML patients, Twist1 plays a prominent role in the pathogenesis of the disease that leads to unique clinical phenotypes. The result indicates that Twist1 could represent a powerful biomarker that serves as a prognostic as well as therapeutic guide for patients with AML.

Disclosures
No relevant conflicts of interest to declare.
Retrospective Analysis of 59 Cases FLT3-ITD Mutation Positive Acute Myeloid Leukemia
Authors: Sun Jie , Hu Juan , Ying Shuangwei , Cai Zhen , He Jingsong , Zheng Weiyan , Luo Yi , Wei Guoqing , Zhang Jie , Ye Xiujin , Xie Wanzhuo , Huang He 
Published: October 2019

Abstract
Objective: Between October 2010 to May 2014, 59 patients who diagnosed as FLT3-ITD positive aAML were analyzed. All the patients were signed informed consents to agree to provide their clinical information to this study. Informed consents were approved by Ethics Committee. Peripheral white cell count at diagnose; complete remission (CR) rate; the percentage of progressing to refractory leukemia and the response to allogenic stem cell transplantation (allo-SCT) were analyzed. Better therapeutic strategy was explored.
Methods: Diagnosis was made according to the French-American-British classification system or World Health Organization diagnostic classification criteria. FLT3-ITD mutation was detected by polymerase chain reaction (PCR) and gel electrophoresis. Refractory leukemia is defined as fail to achieve CR after two courses of conventional chemotherapy, or with a short (<6–12 months) CR1 duration, or patients who have relapsed at least twice.
Results: 59 patients were diagnosed as Flt3-ITD mutation positive aAML in our centre during the last 44months, including 30 males and 29 females, with an average age at 42.2y. Flt-ITD mutation can be detected in 1/59 of M0 (1.7%); 6/59 of M1 (10.1%); 20/59 of M2 (33.8%); 9/59 of M3 (15.2%); 5/59 of M4 (8.5%); 14/59 of M5 (23.7) and 1/59 of M6 (1.7%). 23.7% (14/59) patients had high white blood cell counts (＞100*10E9/L) at diagnose. After induction chemotherapy, 50% (27/54) cases achieved CR. 53.4% (23/43, 16 cases were lost follow-up) patients met the criteria of refractory leukemia. 12 patients received allo-SCT, including 8 refractory leukemia cases. The average time from diagnose to transplantation is 6.5 months (3 to 12 months). 1 patient relapsed at 3 months after allo-SCT who was diagnosed refractory leukemia before allo-SCT, all other 11 cases remained CR with the longest follow-up time at 26 months (3 to 26 months). For the left 15 refractory leukemia cases who did not received allo-SCT, the 1-year OS rate for the 12 cases is 50% (6/12), but the 1-year EFS rate is 0 (0/12) (3 cases were lost follow-up).
Conclusion: FLT3-ITD mutation was reported to be detected in around 30% of aAML. Flt3-ITD mutation is an independent high-risk factor for aAML. Based on retrospective data from 59 cases in our centre, FLT3-ITD mutation positive aAML patients can be characterized as high white blood cell count at diagnose, lower CR rate and higher refractory leukemia percent. allo-SCT can achieve persistent remission in part of patients, even in refractory leukemia patients. Our study also implied that the CR1 is the ideal stage for FLT3-ITD mutation positive aAML patient to receive allo-SCT.

Disclosures
No relevant conflicts of interest to declare.
Upregulated TCR Zeta and Improved the IFN-γ Secretion and Cytotoxicity in T Cell from Patients with AML after IL-2, IL-7 and IL-12 Stimulation
Authors: Chen Shaohua , Shi Li , Yang Lijian , Lu Yuhong , Luo Gengxin , Zhu Kanger , Li Yangqiu 
Published: October 2019

Abstract
T cells from patients with acute myeloid leukemia (AML) display antigen receptor–mediated signaling aberrations associated with defective T cell receptor (TCR) zeta chain, a subunit of the TCR/CD3 complex. Up-regulate TCR signaling activation by TCR zeta recombinant vector transfection was showed to reverse IL-2 production and increased the activation in T cells from patients with chronic myeloid leukemia. This study was undertaken to explore the possibility that increased TCR zeta gene expression stimulated with immune cytokines may upregulate T cell receptor signaling activation in T cells from patients with AML. CD3+ T cells sorted from peripheral blood of 8 cases with AML were induced with different immune cytokines, including IL-2, IL-7, IL-12, anti-CD3 and anti-CD28 antibodies in vitro. The result showed that TCR zeta chain protein was significantly upregulated after stimulation with IL-2 + IL-7 at 72 hours (p<0.05) by western-blot analysis, the expression of Zap-70 which is TCR zeta-associated protein was increased apparently after stimulation by combined IL-7 and IL-12 at 72 hours (p<0.05). A higher level of interferon (IFN)-γ secretion was showed obviously with single stimulating factor or combination with different cytokines (p<0.05), especially in IL-12 + IL-7, CD3 + CD28 + IL-2 or CD3 + CD28 + IL-7 combination. In addition, the cytotoxicity of T cell from patients with AML was achieved after stimulation with different cytokines combinations at 7 days, especially in CD3 + CD28 + IL-2 group and CD3 + CD28 + IL-2 + IL-12 group. These result showed that downregulation of the TCR zeta chain in T cell from patients with AML is a reversible event. Combined different cytokines could restore TCR zeta chain deficiency and enhance IFN-γ production and cytotoxicity in T cells from AML patients. It is possible that upregulated TCR zeta chain expression in T cell from AML patients will effectively increase or recover their activation of antileukemia cytotoxicity.

Disclosures
Chen: National Natural Science Foundation of China (no. 81100353, 81270604), the Fundamental Research Funds for the Central Universities (No. 21611447, 21612116), And Medical Science Foundation of Guangdong Province(A2011325): Research Funding.
Differentiated Expression of microRNAs May be Regulated By the Vascular Endothelial Growth Factor in Acute Myeloid Leukemia
Authors: Ye Xiujin , Zhu Lixia 
Published: October 2019

Abstract
Background: Overexpression of the vascular endothelial growth factor (VEGF) is characterized in acute myeloid leukemia (AML), which stimulates endothelial cell mitogenesis and cell migration. MicroRNAs play a crucial role in the initiation and progression of malignancies. This study aimed to find the differential expression profiling of microRNAs that related with the expression level of VEGF in AML, and further explore the mechanism of VEGF in the course of AML.
Methods: The overexpression of VEGFa lentiviral vector LV-VEGFa-O or the knockdown of VEGFa lentiviral vector LV-VEGFa-shRNA or the empty lentiviral vector LV-NC was transfected into AML cells HL60 and U937. MicroRNAs was analysed by using a commercial microarray platform (miRCURY LNA™ v.18.0, Exiqon) for microRNAs. A set of selected microRNAs and VEGF mRNA were further analyzed by using quantitative real-time PCR in 45 bone marrow (BM) from primary AML patients.
Results: We have successfully constructed AML cell lines stably overexpressing VEGFa or knockdown VEGFa. Overexpression of VEGFa followed by a few changes of microRNAs profiling, that has-miR-1273f and has-miR-4679 were up-regulated, while eb-miR-BART10-3p and has-miR-3925-3p were down-regulated. However, knockdown of VEGFa significantly affected the expression profiling of microRNAs in AML cells. Thirty-three microRNAs were lower expression level than cells transfected with LV-NC (Figure 1). According to the expression level of VEGF, the AML patients were divided into two groups: VEGF-high group and VEGF-low group. We found five microRNAs (miR-20a, miR-93, miR-16-5p, miR-17 and miR-124-5p) were higher expression level in VEGF-high group than in VEGF-low group (p<0.05) (Figure 2).
Conclusion: Five microRNAs (miR-20a, miR-93, miR-16-5p, miR-17 and miR-124-5p) were positively correlated with VEGFa in AML. It is necessary to further explore the role of the selected microRNAs in the signal pathways of the regulation of VEGFa in the course of AML.
Figure 1 Figure 1.
Figure 2 Figure 2.

Disclosures
No relevant conflicts of interest to declare.
SKIP Is Underexpressed in AML Leading to Sphingosine Kinase Hypofunction
Authors: Ghazaly Essam , Smith Paul , Gnanaranjan Chathunissa , Young Bryan D , Joel Simon , Gribben John G. , Taussig David C 
Published: October 2019

Abstract
Background: Sphingosine kinase interacting protein (SKIP) has been shown to be mostly silenced by hypermethylation in AML [1]. SKIP interacts with and regulates the function of sphingosine kinase (SK) enzyme. SK activity results in phosphorylation of sphingosine (SPH) to form sphingosine 1 phosphate (S1P), which promotes cell survival and resistance to apoptosis. On the other hand, S1P precursors ceramide (CER) and SPH mediate antiproliferative and apoptotic responses. SKIP has been reported to negatively regulate SK1 activity in fibroblasts. Therefore, we investigated the consequences of SKIP silencing in primary AML cells. In addition, we studied the effects of SKIP re-expression in leukemic cell lines.
Methods: CTS and K562 cells were transfected with SKIP gene using standard techniques. SKIP is normally silenced in both cell lines. Intracellular and extracellular S1P, SPH and CER were measured by UPLC-MS/MS. In addition, intracellular SK activity was determined based on C17 S1P production from C17 SPH substrate. Chemosensitivity to doxorubicin, Imatinib and Ara-C in transfected cells was also studied.
Results: In Primary AML cells, intracellular S1P and CER concentrations were reduced compared to G-CSF-mobilized peripheral blood mononuclear cells. In addition, SK activity was found to be downregulated in AML primary cells. When we transfected leukemic cell lines with SKIP gene, S1P and CER showed at least 2 fold increase in intracellular and extracellular basal levels compared to vector alone control (Figure 1). Further studies confirmed a significant increase in intracellular SK activity in SKIP transfected compared to vector alone cells, based on C17 S1P production (8.8 ± 2.6 vs 1.4 ± 0.4 ng/106 cells respectively after 24 hrs, p< 0.05). This increase in S1P and CER was associated with increasing apoptotic signals as evidenced by Annexin V staining and cleaved PARP expression. Moreover, chemosensitivity to Imatinib and AraC was significantly increased in SKIP transfected cell lines. These experiments confirm the regulation of SK1 function by SKIP.
Conclusion: These data indicate that SKIP is downregulated in AML leading to reduced SK activity, which ultimately inhibits the apoptosis response.
Figure 1. Effect of SKIP transfection on intracellular concentrations of S1P. Figure 1. Effect of SKIP transfection on intracellular concentrations of S1P.
1. Saied, M.H., et al., Genome wide analysis of acute myeloid leukemia reveal leukemia specific methylome and subtype specific hypomethylation of repeats. PLoS One, 2012. 7(3): p. e33213.
EAG and PS contributed equally
JG and DT contributed equally

Disclosures
No relevant conflicts of interest to declare.
Genomic Analisys of Notch Mutations in a Case of Alagille Syndrome with Acute Lymphoblastic Leukemia
Authors: Bernardi Simona , Tononi Paola , Marin Vargas Sergio , Bassi Giulio , Takam Kamga Paul , Perbellini Omar , Ambrosetti Achille , Meneghini Vittorio , Scaffidi Luigi , Martinelli Giovanni , Iacobucci Ilaria , Pizzolo Giovanni , Delledonne Massimo , Krampera Mauro 
Published: October 2019

Abstract
Alagille syndrome (ALGS), or arteriohepatic dysplasia, is a congenital multisystem disease due to Notch signalling pathway mutations, most commonly affecting JAGGED-1 (ALGS type 1), and more rarely NOTCH-2 (ALGS type 2), leading to hepatic, lung, renal and ocular dysfunction (chronic cholestasis, peripheral pulmonary artery stenosis, dysplastic kidneys pigmentary retinopathy), and skeletal abnormalies (minor vertebral segmentation, characteristic facies, posterior embryotoxon/anterior segment defects). ALGS is an autosomal dominant disease, but it is characterized also by variable penetrance and clinical expression and somatic/germline mosaicism.
A 20-year-old man with ALGS was admitted to the University Hospital of Verona because of pancytopenia. Following analyses led to the diagnosis of Philadelphia chromosome/bcr-abl-negative, CD10-positive, B-lineage acute lymphoblastic leukemia (common B-ALL).
In order to identify the genetic components involved in this complex phenotype, we sequenced the exome of a bone marrow sample collected from the patient. By genome interpretation with Knome pipeline applied to the reference genome UCSC hg19, we found missense variants both in NOTCH-2 (E38K) and JAGGED-1 (P871R) genes that are mainly involved in the syndrome, although their effect on protein function was predicted not to be deleterious. However, we detected putative damaging mutations in genes such as PAX5 (R38H) and NOTCH-1 (K1821N), which might be strongly related to the observed disease. In fact, PAX5 is a member of PAX protein family of transcription factors implicated into regulation of early development that binds NOTCH-2 and likely altering its functionality. On the other hand, NOTCH-1 is involved in cell growth and proliferation and thus the predicted alteration of function of the corresponding protein may have an important role in neoplastic transformation.

Disclosures
Martinelli: Novartis: Consultancy, Speakers Bureau; BMS: Consultancy, Speakers Bureau; Pfizer: Consultancy; ARIAD: Consultancy.