bluuebunny/crossref_metadata_2025_split huggingface dataset:
Assessing Proteostasis and Proteasome Stress In Light Chain Amyloidosis
Authors: Oliva Laura , Palladini Giovanni , Cerruti Fulvia , Pengo Niccolò , Cascio Paolo , Merlini Giampaolo , Sitia Roberto , Cenci Simone 
Published: October 2019

Abstract
Abstract 3992
Recently, proteasome inhibitors (PI) proved powerful against multiple myeloma (MM), the neoplastic transformation of plasma cells. The balance between proteasome expression and degradative workload (mainly contributed by protein synthesis) proved a crucial determinant of apoptotic sensitivity of MM cells to proteasome inhibition (Bianchi et al, Blood 2009). Light chain amyloidosis (AL) is a plasma cell dyscrasia caused by a bone marrow plasma cell clone synthesizing structurally unstable, misfolded, monoclonal immunoglobulin (Ig) light chains, which polymerize into amyloid fibrils. Interestingly, AL is proving even more sensitive than MM to PI in clinical trials with unprecedented response rates (>80%) rapidly achieved in previously untreated patients (Kastritis et al, J Clin Oncol 2010), raising the question as to whether, and if so why, AL cells are intrinsically more sensitive than MM to PI. We hypothesized that AL cells suffer from intense proteasome stress linked to the synthesis of the misfolded Ig light chain, thereby facing constitutive proteotoxicity.
To test this hypothesis, we set out to optimize purification of primary bone marrow plasma cells from AL patients, and determine: intrinsic sensitivity to the PI bortezomib (by FACS); proteasome activity (by fluorogenic assays); accumulation of ubiquitinated (Ub) proteins and Ig light chain (by immunofluorescence).
Our ex vivo studies demonstrated twofold higher PI sensitivity in AL plasma cells as compared to primary MM cells (EC50 in 24 hr apoptosis assays: AL, 8.3 ± 2.2 nM; MM, 15.1 ± 3.0 nM). We also found that, similar to MM cells, proteasome activity of primary AL plasma cells varies greatly among different patients (5.2 ± 3.6 nM substrate specifically cleaved by the chymotryptic β-peptidase activity per cell per min). Furthermore, accumulation of Ub proteins strongly correlates with light chain content, suggesting a crucial role for paraprotein synthesis and/or retention on proteasome stress. Interestingly, unlike MM cells, we failed to detect a clear correlation between proteasome activity and ex vivo assessed PI sensitivity, possibly due to intracellular toxicity of the misfolded light chain. The resulting hypothesis that different mutations could result in different intrinsic proteotoxicity in AL cells is currently being tested.
In conclusion, our integrated approach indicates that AL cells are intrinsically more sensitive to PI than MM cells, providing a potential explanation for the excellent clinical responses. Moreover, we established a technological platform to investigate proteostasis and proteotoxic stress in primary AL cells. This strategy may help investigate the efficacy of proteostasis regulators on plasma cell dyscrasias, including MM, and identify molecular markers of clinical use to predict disease severity and response to therapy.

Disclosures:
No relevant conflicts of interest to declare.
Myeloid-Derived Suppressive Cells Belonging to the Leukemic Clone Account for Immunosuppression In CMML
Authors: Droin Nathalie , Itzykson Raphael , Rameau Philippe , Morabito Margot , Braun Thorsten , Louache Fawzia , Zhang Yanyan , Botton Stephane De , Cordonnier Catherine , Quesnel Bruno , Fenaux Pierre , Vainchenker William , Solary Eric 
Published: October 2019

Abstract
Abstract 3997
We have shown previously that cells identified as monocytes in the peripheral blood of patients with chronic myelomonocytic leukemia (CMLL) included a variable proportion of CD14-negative, CD24-positive immature granulocytes. These cells synthesize and secrete alpha-defensin 1–3 that inhibit M-CSF-driven monocyte differentiation into macrophages through interaction with the P2Y6 purinergic receptor (Droin N et al, Blood 2010).
In the present study, we show that these CD14-,CD24+ immature granulocytes also inhibit the proliferation of autologous lymphocytes activated with anti-CD3 and anti-CD28 antibodies through cell-cell contact. This functional property suggested that these cells could be “myeloid-derived suppressive cells” (MDSC), which was supported by their phenotype that included expression of CD15 marker at their surface and survivin, S100A8 and S100A9, Cyclin D2 and Cyclin D3 at the mRNA level. STAT3 and STAT6 were found constitutively phosphorylated in these immature granulocytes that responded to Toll-like receptor agonists such as LPS or Pam-6. CD14-positive monocytes of the leukemic clone activated these MDSC through production of IL-13 and induction of arginase 1 mRNA, which could be reproduced by recombinant IL-13. On the other hand, activation of these MDSC did not require induction of the nitric oxide synthase mRNA, in agreement with their granulocytic origin.
We were able to generate immature myeloid cells expressing CD24 with morphology similar to that of peripheral blood MDSC by in vitro culture of various subpopulations of bone marrow CD34-positive cells obtained from CMML patients, including the most immature CD34+/CD38-/CD90+ cells. Furthermore, generation of these cells could be recapitulated in vivo by xenotransplantation of CMML CD34+ cells in NOG mice, albeit with lower efficacy than CD14+ cells. In patients with high grade CMML included in a phase II clinical trial, decitabine was observed to decrease both CD14+,CD24- monocytes and CD14-,CD24+ immature granulocytes.
Altogether, these data suggest that CMML initiating cells generate CD14-positive monocytes and, in most patients, an additional population of CD14-negative immature granulocytes with suppressive properties towards innate and acquired immune response. Generation of these cells may account for the high sensitivity of CMML patients to autoimmune and infectious diseases.

Disclosures:
Fenaux: Celgene: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Janssen Cilag: Honoraria, Research Funding; ROCHE: Honoraria, Research Funding; AMGEN: Honoraria, Research Funding; GSK: Honoraria, Research Funding; Merck: Honoraria, Research Funding; Cephalon: Honoraria, Research Funding.
Efficacy of Azacitidine In the Treatment of Chronic Myelomonocytic Leukemia
Authors: Teichman Melissa L. , Wetzstein Gene A. , Ho Viet Q. , Lancet Jeffrey E. , List Alan F. , Komrokji Rami S. 
Published: October 2019

Abstract
Abstract 4017

Background:
Chronic myelomonocytic leukemia (CMML) is a heterogeneous disease sharing features of myelodysplastic syndromes (MDS) and myeloproliferative neoplasms (MPN). FDA approved indications for azacitidine and decitabine include CMML as subset of MDS. Fewer than 10 patients with CMML, however, were treated in each of the original studies. In this study we report our institutional experience of azacitidine treatment of CMML patients.

Methods:
This was a retrospective review of CMML patients who received azacitidine at Moffitt Cancer Center. The primary endpoint was determining response rate to azacitidine utilizing International Working Group 2006 criteria (IWG 2006). Secondary objectives were to assess treatment tolerance and overall survival. Descriptive statistics were used for baseline characteristics and response rates. Kaplan-Meier estimates were used for evaluation of overall survival.

Results:
Between July 2004 and December 2009, 35 CMML patients treated with azacitidine were identified. Table-1 summarizes baseline characteristics of those patients. Based on Dusseldorf CMML risk criteria one patient (2.9%) was low risk, 17 (48.7%), intermediate, 7 (20%) high risk and 10 (28.6%) were unknown. According to MD Anderson CMML risk model, 11 (31.4%) were low risk, 12 (34.3%) int-1, 2 (5.7%) int-2, 1 (2.9%) high risk and 9(25.7%) unknown. The median number of azacitidine cycles was 6.0 (1-34)
The best response rates by IWG 2006 criteria were complete response (CR) 5 (14.3%), marrow CR 4 (11.4%), partial response (PR) 1 (2.9%), and hematological improvement (HI) 7 (20%). The overall response rate was 48.6%. The median OS was 25 month (95%CI 13.8–36.1 mo).

Conclusions:
In this retrospective analysis, response to azacitidine in CMML was similar to response rates reported in other MDS patients on azacitidine studies. The median overall survival is comparable to AZA-001 randomized clinical study.

Disclosures:
Lancet: Celgene: Research Funding. List:Celgene: Research Funding. Komrokji:Celgene: Research Funding, Speakers Bureau.
Thrombocytopenia Predicts for Poor Survival In Patients with Lower Risk Myelodysplastic Syndromes (MDS)
Authors: Cruz Raymond , Galili Naomi , Khan Ghulam Sajjad , Raza Azra 
Published: October 2019

Abstract
Abstract 4021
The incidence of thrombocytopenia in MDS is reported to be as high as 40–60% in various reports and is more commonly seen in patients presenting with higher risk disease (∼80%) by International Prognostic Scoring System (IPSS) compared to Int-1 (∼50%) or low risk MDS (12%). Thrombocytopenic MDS patients are at an increased risk of bleeding and hemorrhagic complications have been reported in 3– 50% patients with fatal hemorrhage occurring in 14–20% cases. The prognostic significance of thrombocytopenia in lower risk MDS patients (low+Int-1 risk) has not been clearly defined. To determine whether thrombocytopenia in lower risk MDS patients affects survival, we performed a retrospective study of 1,402 patients diagnosed with MDS between 1987 and 2010. Of these, 687 patients were classified by IPSS as follows: 22%(151/687) were identified as having low risk, 44% (301/687) had Int-1 risk, 4% 26/387) had Int-2, and 30% (209/687) had high risk disease. The overall incidence of thrombocytopenia (platelets <100,000/ul) across all groups was 54% (373/687) with the highest incidence being in the higher risk groups (46%), followed by Int-1 (43%) and low risk (7.5%). Of the 452 lower risk patients, 41% were thrombocytopenic (187/452). Overall, 256 lower risk patients had platelets of <150,000 and these were further divided into three subgroups based on platelet counts ranging between: 1) 100–149,000/ul; 2) 50–99,000/ul; 3) less than 50,000/ul. Median overall survival (OS) was obtained for each group. Statistical significance was calculated using Kruskal-Wallis rank sum test of equality of population. Non-thrombocytopenic lower risk patients showed a statistically significant higher median OS compared to thrombocytopenic patients (3.2 vs. 1.3 yrs p < 0.0005). Median OS progressively decreased with declining platelet range (2.4, 1.6 and 1.2 years) below 150,000/ul. Patients whose platelet counts were less than 50,000/ul had survival rates that were comparable to Int -2 patients (1.2 yrs). We conclude that the presence of thrombocytopenia predicts for poor survival in lower risk MDS patients and this information may be helpful in making therapeutic decisions.

Disclosures:
No relevant conflicts of interest to declare.
Prognostic Relevance of Genomic Aberrations In Light Chain Only Multiple Myeloma
Authors: Jiang Nan , Qi Connie , Trieu Young , Reece Donna E. , Chang Hong 
Published: October 2019

Abstract
Abstract 4036

Background:
Multiple myeloma (MM) is characterized by an expansion of clonal plasma cells with production of monoclonal immunoglobulin. The majority of MM patients produce an intact immunoglobulin, but in a subset of patients (≂f15%), the tumor produces monoclonal light chains only (LCO). Although specific genomic aberrations have emerged as a major prognostic factor in MM, the genomic changes and their prognostic impact on LCO myeloma patients are not clear.

Methods:
A total of 86 patients with LCO MM identified by urine and serum immunoelectrophoresis were included in this study. They were all uniformly treated with high dose chemotherapy followed by autologous stem cell transplant (ASCT). The genomic risk factors including del(13q), del(17p), t(4;14), 1q21 gain and 1p loss– were evaluated by cytoplasmic fluorescence in situ hybridization (cIg-FISH) in clonal plasma cells and correlated with patients clinical outcomes.

Results:
cIg-FISH detected del(13q) in 41%, t(4;14) in 12%, del(17p) in18%, 1q21 gain in 25%, and 1p loss in 19% of the evaluable cases. In our entire cohort, the median post-transplant follow-up was 36.5 months with a median progression free survival (PFS) of 24.8 months [95% confidence interval (CI): 18.4–31.3] and overall survival (OS) of 68.8 months (95% CI: 50.2–87.5). Patients with del(13q) and 1q21 gains had a significantly shorter PFS (median 15.8 vs. 33.4 months, p=0.002; median 19.1 vs. 33.4 months, p=0.011, respectively), and shorter OS (median 56.2 vs. 80.4 months, p=0.021; median 26.9 vs. 77.9 months, p=0.006, respectively), than those without such genetic abnormalities. In addition, 1p loss was associated with a significantly shorter PFS (median 18.2 vs. 37.9 months, p=0.001). However, there was no significant difference in PFS or OS in patients with or without the high-risk genetic factors t(4;14) or del(17p). On multivariate analysis adjusting for all 5 genetic risk factors, del(13q) was an independent prognostic factor for PFS (p=0.011) and OS (p=0.045).

Conclusion:
Although LCO MM had a similar incidence of genetic abnormalities to common MM, only del(13q) and chromosome 1 abnormalities appear to adversely affect the survival in our cohort. Further larger, prospective studies are warranted to verify the role of these genomic aberrations in the genetic risk stratification of LCO MM.

Disclosures:
Reece: Celgene: Honoraria, Research Funding.
Molecular Profiling of Extramedullary and Medullary Plasmacytomas Compared to Multiple Myeloma
Authors: Mahindra Anuj , Amin Samir B. , Sohani Aliyah R. , Motyckova Gabriela , Patel Kishan , Vallet Sonia , Pozzi Samantha , Santo Loredana , Cirstea Diana , Toomey Christiana E. , Michaelson James S. , Hochberg Ephraim P. , Cin Paola Dal , Rodig Scott J , Hideshima Teru , Anderson Kenneth C. , Munshi Nikhil C. , Raje Noopur 
Published: October 2019

Abstract
Abstract 4042
Plasmacytomas are rare clonal proliferations of plasma cells that though cytologically identical to plasma cell myeloma, present with osseous or extraosseous growth pattern. Understanding their molecular characteristics can provide crucial insights into their pathogenesis and risk of progression to multiple myeloma (MM). To investigate the differences between extramedullary (EMP) and medullary plasmacytomas (MP) and MM without plasmacytomas, we sought to molecularly profile these tumors by tissue microarrays, gene expression, microRNA, and FISH.
We identified 85 patients from our data base with a pathological diagnosis of plasmacytoma. Of the 85 patients, 13 patients presented with EMP, and 72 had MP. Among the patients with EMP (n=13), 2 patients presented with multiple lesions. Three of 13 (23%) patients progressed to develop MM at a median of 12 months. 72 patients presented with MP, of which 21 had solitary lesions and 27 (37%) progressed to MM at a median of 20.5months. There was a male preponderance (67% vs 33%) and the median age at diagnosis was 60.5 years (range 27.7–87.6). The mean overall survival for patients with EMP was 121 months (95% confidence interval[CI] 97–144 months) and for patients with MP was 102 months (95% CI 93–128 months) {p=0.025}.
MicroRNA (miRNAs) profiling was performed on MP (n=19), EMP (n=7) and MM samples (n=66). Data was normalized using U6 endogenous control. Gene expression profiling was performed and correlated with the miRNA data to identify genes and transcripts of interest. miRNA 127, which regulates SET D8, was upregulated four fold in both MP and EMP compared to MM. miRNA 493, which regulates cadherin 11 and PTCH 1, both of which have been associated with metastatic potential in solid tumors, was similarly downregulated four fold in both MP and EMP compared to MM. A tissue microarray was created on 52 patients (8: EMP, 44: MP,) in whom paraffin-embedded tissue was available. Additional evaluation using SET 8, cadherin 11 antibodies and validation of additional functional targets is ongoing and will be reported.
Differential expression patterns of factors involved in proliferation, survival, adhesion, and stroma-tumor cell interactions may help explain plasmacytoma biology and identify factors responsible for progression to MM. These insights may help identify new therapeutic approaches and targets in the treatment of these plasma cell disorders.

Disclosures:
No relevant conflicts of interest to declare.
Targeting Syk (spleen tyrosine kinase) Inhibits the Proliferation, Migration and Viability of Multiple Myeloma Cells
Authors: Körber Ruth-Miriam , Held Stefanie AE , Daecke Solveig , von Lilienfeld-Toal Marie , Bringmann Anita , Brossart Peter 
Published: October 2019

Abstract
Abstract 4071

Introduction:
Multiple myeloma (MM) is a malignant plasma cell disorder characterized by clonal expansion of single plasma cells in the bone marrow. Despite high-dose melphalan therapy with autologous stem cell transplantation (ASCT) and the introduction of active drugs like bortezomib or lenalidomide that have been associated with improved survival MM is still incurable. Thus, the identification of novel molecular targets and additional treatment options are warranted. In B-cell malignancies such as chronic lymphocytic leukaemia (CLL) or diffuse large B cell lymphoma, the inhibition of the tyrosine kinase Syk gave promising preclinical and first clinical results.In our study, we analyzed the potential of Syk as a target in MM.

Methods:
The MM cell lines AMO-1, U266, MMS-1 and RPMI8226 and primary MM cells were treated with the Syk-inhibitors BAY61-3606 or piceatannol and proliferation, migration and apoptosis induction were analyzed. Effects on involved intracellular signaling cascades were determined by Western blotting.

Results:
Incubation of MM cell lines with Syk-inhibitors resulted in a reduced proliferation and and SDF-1 induced migration, that was accompanied by a concentration dependent inhibition of the MAP kinase signaling characterized by reduced phosphorylation of ERK an p38 molecules. Furthermore, the nuclear localized expression of the NF-kB members RelA and RelB was inhibited in treated cells. In addition, Syk inhibition induced apoptosis in MM cell lines and primary MM cells in a dose-dependent manner as demonstrated by flow cytometry, caspase-3 activity and PARP-1 cleavage. While there was no effect on the expression of xIAP, survivin or MCL-1, Syk inhibition reduced the expression of pro-apoptotic Bcl-2 and Bcl-xl molecules and increased the release of cytochrome c, indicating that the apoptotic cell death is mediated via the internal mitochondrial pathway. Combination of piceatannol with lenalidomide and orally bioavailable dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor NVP-BEZ235but not with bortezomib or dexamethasone enhanced the cytotoxic effects of the compound. In line with the results from previous experiments the addition of MAPK inhibitors PD98059, SP600125, U0126, SB202190 and SB203580 to piceatannol further increased the efficacy of Syk inhibition.

Conclusions:
Our results show that Syk inhibition might represent a promissing new treatment option in MM with an increased efficacy when combined with lenalidomide or MAP kinase inhibitors.

Disclosures:
No relevant conflicts of interest to declare.
Functional Role of BAALC In Leukemogenesis
Authors: Berg Tobias , Heuser Michael , Kuchenbauer Florian , Park Gyeongsin , Fung Stephen , Lin Grace , Leung Malina , Lai Courteney , Krauter Jürgen , Ganser Arnold , Humphries R. Keith 
Published: October 2019

Abstract
Abstract 4194
Cytogenetically normal acute myeloid leukemia (CN-AML) patients with high BAALC or MN1 expression have a poor prognosis. Whereas the oncogenic function of MN1 is well established, the functional role of BAALC in hematopoiesis is not known.
We therefore compared the expression of BAALC and MN1 in 140 CN-AML patients by quantitative PCR. To further assess the impact of BAALC on leukemogenesis we used retroviral gene transfer into primary murine bone marrow cells and cells immortalized with NUP98-HOXD13 (ND13) and HOXA9. Transduced cells were assessed in vitro by colony forming assays and for their sensitivity to treatment with all-trans retinoic acid (ATRA). They were also evaluated by in vivo transplantation into lethally-irradiated mice.
In the 140 CN-AML patients analyzed, the expression of BAALC and MN1 was highly correlated (R=0.71). Retroviral overexpression of MN1 or BAALC in the Hox gene-immortalized bone marrow cells did not cause upregulation of the other gene, suggesting that these genes do not regulate each other. In murine bone marrow cells BAALC did not immortalize the cells in vitro as assessed by serial replating of transduced cells in methylcellulose assays. Transplantation of transduced cells resulted in negligible engraftment of approximately 1 percent at 4 weeks after transplantation. However, co-transduction of BAALC into NUP98-HOXD13 cells (which are very sensitive to the treatment with all-trans retinoic acid) increased the 50 percent inhibitory concentration (IC50) of ATRA by 4.3-fold, suggesting a negative impact of BAALC on myeloid differentiation. We next evaluated whether the differentiation inhibiting effects of BAALC may cooperate with the self renewal-promoting effects of HOXA9 to induce leukemia in mice. Mice receiving transplants of murine bone marrow cells transduced with BAALC and HOXA9 developed myeloid leukemias with a median latency of 139.5 days that were characterized by leukocytosis, massively enlarged spleens (up to 1.02 g), anemia and thrombocytopenia. Infiltrations of myeloid cells were also found in liver, spleen, and kidney. The disease was transplantable into secondary animals. By Southern blot analysis we found one to two BAALC viral integrations per mouse, suggesting that clonal disease had developed from BAALC-transduced cells.
We demonstrate for the first time that BAALC blocks myeloid differentiation and promotes leukemogenesis when combined with the self-renewal promoting oncogene HOXA9. Due to its prognostic and functional effects BAALC may become a valuable therapeutic target in leukemia patients.

Disclosures:
No relevant conflicts of interest to declare.