bluuebunny/crossref_metadata_2025_split huggingface dataset:
Harvesting of Peripheral Blood Stem Cells Using Plerixafor (MOZOBIL) and Granulocyte-Colony Stimulating Factor in Patients Who Had Never Undergone Stem Cell Collection.
Authors: Teoh Gerrard , Goh Keith 
Published: October 2019

Abstract
Abstract 4236

Introduction
The collection of peripheral blood stem cells (PBSC) can be tedious and expensive. Moreover, PBSC yields can be severely compromised in patients who previously received chemotherapy. The use of plerixafor (MOZOBIL) for PBSC mobilization has been shown to significantly improve CD34 stem cell yields in patients whom PBSC collections have been adequate. However, this requires the patient to undergo further PBSC harvesting sessions; adding to the patient's inconvenience, exposure to adverse events and costs. We therefore investigated whether using plerixafor at the outset of PBSC mobilization would improve patient acceptability; reduce PBSC collection days and accordingly the patient's costs.

Patients and Methods
Three adult patients who had never undergone PBSC harvesting were mobilized using subcutaneous (SC) granulocyte-colony stimulating factor (G-CSF) 10 μg/kg daily starting 4 days prior to PBSC harvesting; plus SC plerixafor 240 μg/kg 10 hours prior to PBSC harvesting. Both G-CSF and plerixafor were continued for as long as was required for PBSC collections. Two patients had multiple myeloma (MM) and were both in stringent complete remission (sCR). The third patient suffered from adult-onset spinomuscular atrophy (SMA) and stem cells were required for possible re-infusion therapy. Importantly, the bone marrows of all 3 patients were assessed to be normal prior to PBSC mobilization and collection. The target PBSC CD34 yield was 2 million CD34 cells/kg per PBSC collection.

Results
As can be seen, the CD34 yields (expressed as x million CD34 cells/kg) exceeded expectations:
All patients tolerated the procedure very well and were discharged within 24 hrs of completing the final PBSC collection. The only adverse event was grade 1 restless legs syndrome in Patient #1 following the first injection of plerixafor. It was improved with oral diazepam and did not recur. Both patients with MM reported a sense of “rejuvenation” 2-3 weeks after PBSC collection. All patients were pleased with the relatively complication-free procedure.

Conclusions
Upfront collection of PBSCs using G-CSF/plerixafor is both highly effective and safe. There is great patient acceptability as well as much fewer PBSC collection days and an accompanying reduction in procedure costs. The added advantage of using the SPECTRA Optia cell separator is worth further investigations.

Disclosures:
Off Label Use: Plerixafor (MOZOBIL) - Use in patients who had never undergone stem cell collection..
A High Resolution Comparative Genomic Hybridization Array of Adult T-Cell Leukemia-Lymphoma in Individuals of African Descent.
Authors: de Lima Rosangela Gomes , Xu Suying , Hedges Dale , Brites Carlos , Barber Glen N. , Fan Yao-Shan , Ramos Juan Carlos 
Published: October 2019

Abstract
Abstract 4241
Adult T-cell leukaemia/lymphoma (ATLL) is a highly aggressive malignancy with a poor prognosis caused by the human T-cell lymphotropic virus type-1 (HTLV-I). While no specific chromosome or genetic abnormalities have been clearly proven to contribute to the pathogenesis of ATLL, recent comparative genomic hybridization (CGH) studies have demonstrated frequent genetic lesions (gains and losses) involving specific chromosomal regions containing a number of genes of potential cancer relevance. In this study, using a high resolution Agilent platform, we performed a comprehensive CGH analysis of 60 ATLL DNA specimens obtained from individuals of African descent from the United States, the Caribbean, and Brazil. Our preliminary results demonstrate the presence of some alterations already reported by others, such as gains in chromosomes 1q22, 4p16, 7p22, 8q24, 9q33-34, 14q31-q32, 16q16, 18q23, 19p13 and deletions at 6q14, 9p21 and q21 and 14q11. Additionally, we have observed several genetic alterations not previously reported in a significant percentage of our ATLL tumors, such as specific gains in regions of chromosomes 1q, 2q, 5p, and 17p and q, and deletions in 13q and 15q. Most of these regions harbor potential cancer related genes but some contain genes of unknown function. Our preliminary results confirm there are common genetic alterations between ATLL patients of Japanese and African origin, but also distinct genetic lesions between both ethnic groups perhaps contributing to the pathogenesis of ATLL. A more comprehensive analysis including validated findings will be presented at the meeting.

Disclosures:
No relevant conflicts of interest to declare.
A Novel Recurrent Chromosomal Inversion, Inversion(9)(p21q34), in Ph-Positive Leukemia Patients.
Authors: Pan Jinlan , Zhang Jun , Xue Yongquan , Wu Yafang , Shen Juan , Wang Yong , Liu Dandan , Chen Suning 
Published: October 2019

Abstract
Abstract 4242
A pericentric inversion(9)(p21q34) was identified in five Ph-positive leukemia patients including four patients with chronic myeloid leukemia(CML) in chronic phase(CP) and one patient with acute myeloid leukemia (AML) since 1998. Among them, two were males, three wrer females. The median age is 31 years (range 21-52 years). Conventional karyotypic analysis with R-banding technique showed that the inv(9)(p21q34) always occurred in the der(9)t(9;22)(q34;q11) and was accompanied by the der(22)t(9;22)(q34;q11) in all metaphases analyzed in four patients with CML-CP at diagnosis. One patient with AML presented three clones: one with normal karyotype, one with sole t(9;22)(q34;q11), one with inv(9)(p21q34) involving the der(9)t(9;22) and additional t(8;12)(q12;p11). FISH using LSI BCR/ABL dual-color, dual fusion probe, chromosome painting(CP) with the paint probes for chromosome 9p and 9q and RT-PCR using the primers of the BCR/ABL fusion genes were performed in four of them. FISH showed the coexistence of clone with sole t(9;22) and another with inv(9)(p11q34) and t(9;22) in four patients in whom, one patient also showed a deletion of partial sequence from BCR on der(9)t(9;22)inv(9)(p21q34). RT-PCR revealed a b3a2 transcript for BCR/ABL fusion gene transcript. FISH proved that the inv(9)(p21q34) disrupted the ABL/BCR fusion gene at the molecular level. However, it does not appear to have any biological significance because BCR/ABL fusion gene is thought to be involved in the pathogenesis of CML, while ABL/BCR fusion gene is merely mechanical consequence of the t(9;22) translocation. As far as we know, inv(9)(p21q34) has not been reperted previously. Thus, it should be regarded as a novel rare but recurrent secondary chromosomal anomaly. In this series, one lost to follow-up, one patient transformed into B cell acute lymphocytic leukemia, who and other two patients survivals of 28 days, 13 and 34 months, respectively. Only one remains alive. Their dismal outcome probably suggests inv(9)(p21q34) having unfavourable impact on prognosis. At present, no firm conclusion can be drawn from this study. More patients with this anomaly need to be investigated to elucidate its prognostic significance.

Disclosures:
No relevant conflicts of interest to declare.
Osteoclasts Are Involved in the Maintenance of Dormant Leukemic Cells.
Authors: Yokota Asumi , Kimura Shinya , Tanaka Ruriko , Takeuchi Miki , Yao Hisayuki , Sakai Kazuki , Nagao Rina , Ashihara Eishi , Maekawa Taira 
Published: October 2019

Abstract
Abstract 4245
We have previously reported that zoledronic acid (ZOL) had the anti-tumor activity against chronic myelogenous leukemia in the murine models and it could induce apoptosis to leukemic cells (Kuroda, et al. Blood, 2003). ZOL is a widely used agent to treat osteoporosis or hypercalcemia of malignancy because of its activity to induce apoptosis to osteoclasts (OCs), which are specialized cells to resorb the bone matrix. In this study, we hypothesized that OCs might be involved in the survival or the proliferation of leukemic cells in the bone marrow (BM) microenvironment which consists of osteoblasts (OBs), OCs, blood vessels, and so on.
In order to verify our hypothesis, we used the in vitro co-culture system. Ba/F3 wt bcr-abl cells were indirectly co-cultured with the murine OBs, OCs, and/or the bone slices derived from the murine femoral bones in the transwell chambers. OBs and OCs enhanced the proliferation of Ba/F3 wt bcr-abl cells. Interestingly, OCs significantly suppressed the proliferation of Ba/F3 wt bcr-abl cells in the presence of the bone slices. Next, we examined the cell cycle status of Ba/F3 wt bcr-abl cells co-cultured in each condition. Ki-67-negative populations were reported to be in G0 phase, thus 7-AAD and Ki-67 double staining can distinguish population in G0 phase from that in G1 phase. It was revealed that OCs with bone slices increased the Ki-67-negative dormant populations in co-cultured Ba/F3 wt bcr-abl cells. Recently, it has been reported that TGF-β1 is a candidate cytokine that maintain the hibernation state of hematopoietic stem cells in the BM niche. Furthermore, TGF-β1 is one of the major cytokines accumulated in the mineralized bone matrix. Thus we evaluate the TGF-β1 levels in the culture supernatants of OBs, OCs, and/or bone slices by ELISA. The concentrations of TGF-β1 were significantly higher in the supernatants of OBs, OCs, and bone slices, thus it might be suggested that TGF-β1 was released from the bone slices by bone-resorbing OCs (Figure). Furthermore, TGF-β-neutralizing antibody reduced Ki-67-negative dormant populations in Ba/F3 wt bcr-abl cells co-cultured with OBs, OCs, and bone slices.
In conclusion, our in vitro data demonstrate that OCs might maintain the dormant population of leukemic cells in the BM microenvironment and this effect was yielded, in part, by TGF-β1 released from the bone matrix by OCs. It was also suggested that OCs could be an effective target of a novel therapy against minimal residual disease of leukemia, which are thought to be resulted from the dormant population of leukemic cells residing in the BM microenvironment.

Disclosures:
No relevant conflicts of interest to declare.
A Novel Mechanism of Nilotinib-Induced Apoptosis; Sphingolipids.
Authors: Baran Yusuf , Gencer Emel Basak , Camgoz Aylin , Avcu Ferit , Ural Ali Ugur 
Published: October 2019

Abstract
Abstract 4246
Chronic myeloid leukemia (CML) is a hematological malignancy resulting from the reciprocal translocation of chromosomes 9 and 22 that generates BCR/ABL oncogene. Nilotinib is a rationally designed, specific BCR/ABL tyrosine kinase inhibitor. Ceramide is a novel regulator of cell growth and proliferation, differentiation, senescence, cell cycle and also acts a strong apoptotic molecule while its conversion to antiapoptotic glucosyle ceramide (GC) and sphingosine-1-phosphate (S1P) by glucosyle ceramide synthase (GCS) and sphingosine kinase-1 (SK-1) enzymes result in more aggressive and resistant cancers. In this study, we studied the roles of ceramide metabolising genes in nilotinib induced apoptosis and possibility of increasing the sensitivity of BCR/ABL positive K562 and Meg-01 cells to nilotinib through targeting ceramide metabolism.
The cytotoxicity analyses of nilotinib, C8:ceramide to induce de novo generation of ceramides, 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP) to inhibit GCS and SK1 inhibitor were conducted by XTT cell proliferation assay. The changes in caspase-3 enzyme activity and mitochondrial membrane potential (MMP) were measured by caspase-3 colorimetric assay and JC-1 MMP detection kit, respectively. Expression analyses of ceramide synthase (LASS) genes, SK-1 and GCS genes were performed by RT-PCR.
We have shown that nilotinib induces apoptosis and inhibits cell-cycle progression in K562 and Meg-01 cells in a dose dependent manner. We have shown significant synergistic apoptotic effects of nilotinib in combination with C8:ceramide or PDMP or SK-1 inhibitor by XTT cell proliferation assay in addition to the changes in caspase-3 enzyme activity and changes in mitochondrial membrane potential, as compared to any agent alone. These results revealed that increasing de novo generation of ceramides or inhibiting conversion of ceramides to antiapoptotic GC or S1P increased sensitivity of BCR/ABL CML cells to nilotinib. More importantly, RT-PCR results revealed that there were significant decreases in expression levels of SK1 in response to increasing concentrations of nilotinib. On the other hand increases in expression levels of LASS2, -4, -5, and -6 ceramide synthase genes were determined in a dose dependent manner as compared to untreated controls.
It was shown for the first time by this study that targeting ceramide metabolism in addition to inhibition of BCR/ABL by nilotinib induces apoptosis synergistically in BCR/ABL positive K562 and Meg-01 CML cells.
This study was supported by The Scientific and Technological Council of Turkey

Disclosures:
No relevant conflicts of interest to declare.
Detection of Enzymatic Activity of Free Circulating Tyrosine Kinase in Patients with Chronic Myelogenous Leukemia (CML) and the Demonstration of Lower Levels in Resistant Patients.
Authors: Yeh Chen-Hsiung , Abdool Adam , Bruey Jean-Marie , Albitar Maher 
Published: October 2019

Abstract
Abstract 4247
Tyrosine kinases (TKs) regulate a wide variety of cellular signaling events, and aberrant activation of TKs is directly associated with tumorigenesis and cancer progression. Targeted therapy using TK inhibitors (TKIs) is commonly explored for treating various types of cancers. Measuring TK activities for diagnosis and monitoring of therapeutic efficacy may be particularly important for patients treated with TKIs. In this study, we examined the clinical utility of measurement of free circulating TK (cTK) activity directly from plasma in CML patients. The study included 46 patients with newly-diagnosed CML, 24 drug (imatinib, dasatinib, or nilotinib)-resistant CML patients, and 38 apparently healthy control subjects. Plasma levels of cTK activity were measured using a chromogenic universal TK assay kit in which samples and standards were added to a 96-well microtiter plate immobilized with synthesized poly (Glu-Tyr) peptide substrates. Specific TK activity was determined by spectrophotometry. Measured cTK activity at baseline was ∼1000-fold higher in plasma than in paired serum specimens. Overall, plasma cTK activity was significantly higher in newly diagnosed CML patients (median=802 U/mL, range=18-3932 U/mL) than in healthy donors (median=83 U/mL, range=0.6-853 U/mL) or drug-resistant CML patients (median=202 U/mL, range=17-1215 U/mL) (P<0.001). In drug–resistant CML patients, plasma cTK activity was closely correlated with cellular BCR-ABL1 kinase activation, as indicated by downstream phospho-CRKL (P=0.0005) and phospho-STAT-5 (P=0.003) levels. TKI treatment of these patients also resulted in a corresponding decrease in cTK activity over time reflecting response to therapy. Nevertheless, cTK activity was not associated with BCR-ABL1 expression level and was independent of BCR-ABL1 mutation type. In conclusion, measuring circulating TK activity in plasma/serum may reflect the leukemic cells in CML patients and can be used to evaluate response to therapy. In addition, our plasma results support previous data showing that resistant cells in CML patients demonstrate lower TK activity as compared with naïve leukemic cells.

Disclosures:
No relevant conflicts of interest to declare.
Cartridge-Based Automated BCR-ABL mRNA Quantification: Towards Simplified Standardisation.
Authors: Cayuela Jean-Michel , Macintyre Elizabeth , Darlington Meryl , Abdelali Raouf Ben , Fund Xavier , Villarèse Patrick , Piterboth William , Tulliez Michel , Raffoux Emmanuel , Sigaux Francois , Rea Delphine , Bories Dominique , Seror Valerie 
Published: October 2019

Abstract
Abstract 4261
Molecular monitoring of chronic myeloid leukaemia patients treated with tyrosine kinase inhibitors (TKI) is essential for therapeutic stratification. In this context inter-laboratory reproducibility is mandatory. This requires standardization and strict alignment to international scale (IS), as established by the IRIS laboratories. We compared economic and analytical performances, including the possibility of reporting results on the international scale, of an automated cartridge-based assay (Xpert BCR-ABL MonitorTM, Cepheid) to classical non-automated analysis on 181 blood samples from TKI treated CML patients in up to three experienced laboratories. The automated Xpert BCR-ABL MonitorTM assay greatly improved reproducibility between non standardized laboratories. Reproducible sensitivity covered 5 logs, on condition that pre-analytical delays were less than 6 hours. Results were convertible to the international reporting scale. Assessment of average costs showed that this automated assay could be economically relevant for annual activity levels below 300, although cost stability was achieved at 600, compared to 1200 annual tests for non-automated testing. The Xpert BCR-ABL MonitorTM assay could therefore be used in local laboratories for routine quantification of e13/e14-a2 transcripts and might contribute to simplified standardization. However, its prognostic impact relative to non-automated quantification remains to be tested prospectively within appropriate clinical trials.

Disclosures:
No relevant conflicts of interest to declare.
Imatinib as Front-Line Treatment in Chronic Myelogenous Leukemia: How Important is the Achievement of Complete Cytogenetic Response after 3 Months?
Authors: Latagliata Roberto , Breccia Massimo , Carmosino Ida , Vozella Federico , Volpicelli Paola , Stefanizzi Caterina , Santopietro Michelina , Cuia Rosa De , Nanni Mauro , Mancini Marco , Alimena Giuliana 
Published: October 2019

Abstract
Abstract 4271
The achievement of Complete Cytogenetic Response (CCyR) (Ph+ cells 0%) with Imatinib treatment still remains the most important objective in Chronic Myelogenous Leukemia (CML) patients. According to The European Leukemia NET guidelines, CCyR should be achieved within the 12th month of treatment while at 3 months of treatment the goal should be complete haematological response. To address the prognostic role of the early achievement of CCyR, we revised 108 chronic phase CML patients [M/F 57/51, median age 54.9 years, interquartile range (IR) 40.8 – 68.1] treated with front-line Imatinib at our Institution from June 2002 to June 2008 who had evaluable karyotype after 3 months of treatment. At onset, median WBC and PLT counts were 78.4 × 109/l (IR 34.0 – 135.9) and 399 × 109/l (IR 282 – 585), respectively. Sokal risk score was low in 52 patients (48.1%), intermediate in 49 (45.4%) and high in 7 (6.5%); a short pre-treatment phase (< 3 months) with Hydroxyurea was administered to 94/108 patients (87%). After 3 months of Imatinib treatment, 84 patients (77.7%) achieved CCyR while 24 patients (22.3%) still presented Ph+ metaphases (median value 40%, IR 20 - 80) at cytogenetic analysis. At univariate analysis, factors with a negative prognostic impact on achievement of CCyR at 3 months were palpable spleen enlargement (p=0.002), WBC count at onset > 100.0 × 109/l (p=0.01) and pre-treatment with Hydroxyurea (p=0.032); on the contrary, sex, age, Sokal risk score and PLT value did not appear to affect early CCyR achievement. Among the 84 patients in CCyR after 3 months, there were 10 failures during follow-up (6 cytogenetic relapses, 2 molecular relapses and 2 evolution to blastic phase); among the 24 patients who did not achieve early CCyR, there were 12 failures during follow-up (9 primary resistances and 3 cytogenetic relapses). The difference in the incidence of failures during follow-up in the 2 groups was highly significant (p<0.001). In conclusion, the achievement of CCyR at 3 months seems unrelated to traditional prognostic factors (Sokal); furthermore, in our experience it appears to have an important prognostic role, as patients not achieving it showed a significantly higher rate of failures during the follow-up.

Disclosures:
No relevant conflicts of interest to declare.
Long-Term Outcome of Imatinib Treatment in Chronic Myeloid Leukemia (CML) Patients in Shanghai: An International Patient Assistance Programs (GIPAP) Analysis.
Authors: Wang Ai-hua , Zhou Li , Zheng Yu , Li Jun-min , Shen Zhi-Xiang , Hu Jiong 
Published: October 2019

Abstract
Abstract 4275
Imatinib as an oral Bcr-Abl tyrosine kinase inhibitor is approved as first-line therapy for patients with CML chronic phase (CP), accelerated phase (AP) and blast crisis (BC). As the appointed clinical center for the International Patient Assistance Programs (GIPAP) program, a total of 339 CML patients with CML-CP (n=274), AP (n=30) and BC (n=35) were registered and received imatinib treatment in our center. The starting dose of Imatinib treatment was 400mg daily and dose escalation to 600mg was allowed in patients of CML-AP or BC and in patients who failed to obtained optimal response. All patients were evaluated regularly according to the GIPAP protocol. In this study, we analysed the outcome of Imatinib treatment in these patients in terms of cytogenetic response, overall and progression-free survival and adverse events. The median age was 42 (13-81) years old. The median time from diagnosis to imatinib therapy was 4.4 (0-273) months. The median follow-up was 29 (3-103) months. The overall survival for patients received Imatinib treatment were 92.8±3.1%, 57.3+11.3% and 12.3±9.7% for CML-CP, AP and BC respectively. Among patients with CML-CP, the cumulative complete cytogenetic response (CCyR) were 71.9% at 12 months and 79.9% at the censored follow-up timepoint. In prognosis analysis, there were two major impact factors associated with progression-free survival (PFS) for the CML-CP patients: achievement of CcyR and Imatinib as initial therapy. The estimated 60 month PFS was 87.1±3.7% for patients who achieved a CCyR irrespective of time and only 46.7±1.37% for patients who failed to obtain a CCyR (P<0.0001). For those patients who received imatinib as initial therapy (95.6±1.85%) had a significantly superior PFS than those patients received imatinib as secondary treatment after initial interferon therapy either due to intolerance or failed to obtain optimal response (65.9±6.4%, P<0.0001). Overall Imatinib was well tolerated and no patient switched to other treatment due to toxicity of Imatinib treatment. Overall, Imatinib as initial treatment for CML-CP was effective and safe in the first analysis of GIPAP program in Shanghai.

Disclosures:
No relevant conflicts of interest to declare.