bluuebunny/crossref_metadata_2025_split huggingface dataset:
What Is The Optimal Dose Of Methotrexate In Primary Central Nervous System Lymphoma Treatment Regimens?
Authors: Dalia Samir , Price Samantha L , Forsyth Peter , Bello Celeste M. , Shah Bijal D. , Sokol Lubomir , Sotomayor Eduardo M. , Jaglal Michael V. 
Published: October 2019

Abstract

Introduction
Primary central nervous system lymphoma (PCNSL) is a rare disorder with a poor prognosis. The mainstay of treatment is single agent or combination high dose (≥3.5g/m2) methotrexate (HDMTX) based regimens. There is no consensus as to which dose of HDMTX improves outcomes in patients with PCNSL but doses of MTX greater than 3 g/m2 intravenously achieve therapeutic cerebrospinal fluid (CSF) concentrations.

Purpose
To determine if there is an optimal or total dose of HDMTX in PCNSL therapies that results in improved progression free (PFS) or overall survival (OS).

Methods
Patients at Moffitt Cancer Center with PCNSL were identified using our institutional database between January 1, 2000 and September 30, 2011. Patients with complete treatment data who were treated with HDMTX were included in this study. HDMTX was defined as MTX at a dose ≥ 3.5g/m2. Patient demographics, clinical, and treatment data were collected and analyzed. Treatment information collected included the starting dose of HDMTX, IV rituximab use, MTX toxicity and clearance, cycles of MTX, and total amount of MTX administered (g/m2).
Data were analyzed using descriptive statistics and the Kaplan-Meier (KM) method was used to estimate median PFS and OS using the log rank test. P value of <0.05 was considered significant. All data was analyzed using SPSS statistical software version 21.0.

Results
A total of 51 patients were identified (Table 1).
Median PFS was 13months (0-33) and median OS 43months (29-57). The addition of IV rituximab or other chemotherapy failed to improve PFS or OS. HDMTX dose reductions or the total dose of HDMTX administered did not significantly impact PFS or OS. Similarly, when comparing dosing of HDMTX there was no significant difference in 8g/m2 versus 3.5g/m2 (PFS p=0.56, OS p=0.68), or between patients receiving 8g/m2 versus <8g/m2 (PFS p=0.77, OS p=0.6) (Figure 1). Patients receiving 8g/m2 versus those receiving <8g/m2 of HDMTX had similar baseline characteristics except for more liver function abnormalities in the 8g/m2 group.

Conclusions
Differences in initial dosing of HDMTX or total dose of HDMTX therapy did not influence outcomes in our patients with PCNSL. Dose reductions in HDMTX, addition of other chemotherapeutic agents, or rituximab were not associated with improved PFS or OS. An intriguing plateau was observed in OS in the 8gm/m2 arm despite similar PFS, suggesting that the receipt of novel therapies in the relapsed setting may contribute to OS. Multicenter collaborative clinical trials are needed to further assess the optimal initial dose of HDMTX to administer in PCNSL.

Disclosures:
No relevant conflicts of interest to declare.
Primary Mediastinal Large B-Cell Lymphoma: Investigation On The Role Of Rituximab and PET During Treatment
Authors: Zinzani Pier Luigi , Pellegrini Cinzia , Casadei Beatrice , Derenzini Enrico , Broccoli Alessandro , Gandolfi Letizia , Quirini Federica , Argnani Lisa , Stefoni Vittorio 
Published: October 2019

Abstract
Due to limited prospective studies, the optimal treatment of primary mediastinal large B-cell lymphoma (PMLBCL) is still a matter of debate. Third-generation MACOP-B (adriamycin, cyclophosphamide, vincristine, bleomycin, methotrexate and prednisone) regimen in combination with mediastinal radiotherapy (RT) seems to improve disease free survival of patients. In addition, the impact of additional treatment with rituximab and the role of PET are still under investigation due to controversial reported results.
As per institutional guidelines, MACOP-B plus RT was recommended in all PMLBCL patients until 2002. Aim of this report was evaluate the outcome of PMBCL patients diagnosed and treated with MACOP-B plus rituximab and consolidative mediastinal RT (30-36 Gy) after 2002. PET role was also investigated. Seventy-four patients were deemed eligible for this study (follow up of at least 2 years). Fifty patients had stage II and 24 stage IIE-IV, bulky disease was documented in 93% of patients. Median age was 34 years (range, 17-62) and 59.5% were females. All patients were evaluated by both CT and PET scan. After the final PET evaluation, PET-negative patients were observed while PET-positive patients underwent mediastinal RT.
At the end of treatment, 61 (82.4%) patients achieved a complete response (CR); 51 (68.9%) presented a positive final PET and were treated with local RT, while the other 23 (31.1%) had a negative PET. Five patients relapsed within 12 months. At 10 years, estimated overall survival was 82%, progression-free survival was 87.6% and disease-free survival (DFS) for the 61 CR patients was 90.5% (median follow-up 4 years). Regarding the DFS curve (figure 1), no statistically significant differences were observed between patients who underwent also RT (PET-positive, group 1) and patients who remained under observation (PET-negative, group 2): 90.7% (4/51 relapses) vs 90% (1/23 relapse) (p= 0.85), respectively. Comparing these results with our institutional historical series when the front-line for PMLBCL patients included only MACOP-B plus RT without any decision related to PET results (before 2002), the 10-year DFS resulted lower, i.e. 82.8%.
Although with the limitations of an observational retrospective study, the present report underlines that the additional treatment with rituximab does not change the final results in terms of CRs and DFS utilizing third-generation regimens. Moreover, the introduction of the PET-guided RT approach after MACOP-B plus rituximab allows a patient tailored strategy which reduces the use of RT and preserves clinical outcomes.
Figure 1 Figure 1.

Disclosures:
No relevant conflicts of interest to declare.
GPRC5D Is a Cell Surface Plasma Cell Marker Whose Expression Is High In Myeloma Cells and Reduced Following Coculture With Osteoclasts
Authors: venkateshaiah Sathisha Upparahalli , Bam Rakesh , Li Xin , Khan Sharmin , Ling Wen , Randal S Shelton , Yaccoby Shmuel 
Published: October 2019

Abstract
Identification of unique cell surface markers on myeloma cells is important for development of targeted therapies and detection of residual disease. GPRC5D is an orphan receptor reportedly expressed at high level in bone marrow (BM) aspirates or biopsies from myeloma patients. Other studies detected GPRC5D in skin and brain but not in other tissues. The aims of the study were to assess GPRC5D gene expression and cell surface localization in myeloma cells, the changes in expression of GPRC5D and other typical myeloma cell markers following coculture of primary myeloma cells with osteoclasts and the consequences of GPRC5D gene silencing on myeloma cell growth. Global gene expression profile (GEP), qRT-PCR and immunohistochemistry revealed exclusive high expression of GPRC5D in normal and myeloma plasma cells but not in B cells, MSCs, osteoclasts or BM mononucleated cells. GPRC5D expression was higher in myeloma plasma cells from newly diagnosed patients (n=698) compared to normal plasma cells (n=26, p<0.0001)). Among molecularly classified groups GPRC5D expression is higher in MS, MF and LB subgroups and lowest in CD2 and MM cell lines. Flow cytometry analysis and immunohistochemistry detected cell surface expression of GPRC5D in myeloma plasma cells while nuclear localization was also detected in certain myeloma cell lines (e.g. H929). Western blots analysis confirmed GPRC5D expression in whole cell lysate and nuclear fraction. We and other demonstrated phonotypical plasticity of myeloma plasma cells capable of altered expression of recognizable plasma cell markers (e.g. CD138, CD45) following coculture with stromal cells (Dezorella et al., 2009) or osteoclasts (Yaccoby, 2005). In coculture of primary CD138-selected myeloma cells with osteoclasts (n=8), CD138 (p<0,004), CD38 (p<0.001) and GPRC5D (p<0.007) were commonly and significantly downregulated, CD45 (p<0.02) was upregulated, and IRF4 expression was unaffected in cocultured myeloma cells compared to the control freshly obtained uncultured myeloma cells assessed by GEP and qRT-PCR. We also observed reduced expression of GPRC5D in MM cells purified from focal lesion compared to interstitial marrow in paired clinical samples (n=176, p<1.21E-09), suggesting that high activity of osteoclasts in osteolytic lesions mediates phenotypical alteration in myeloma cells. Stable knockdown of GPRC5D by 70% in CAG myeloma line using lentiviral particles containing shRNA had no effect on short-term growth of these cells assessed by MTT assay. These data indicate that GPRC5D is a novel cell surface marker for myeloma plasma cells and that its expression is reduced in dedifferentiated myeloma cells.

Disclosures:
No relevant conflicts of interest to declare.
Utility Of 9-Color, 11-Parameter Flow Cytometry For Detection Of Plasma Cell Neoplasms: A Comparison With Bone Marrow Morphologic Findings and Concurrent M-Protein Studies In Serum and Urine
Authors: Behdad Amir , Ross Charles Warren , Jacques Joshua , Kota Usha , Brown Noah A. , Keren David , Stoolman Lloyd 
Published: October 2019

Abstract

Background
Multiparameter flow cytometry (MFC) is a widely available laboratory platform used for primary diagnosis of mature B-cell and plasma cell (PC) neoplasms. Ongoing advances in myeloma therapeutics have drawn attention to the need for advanced laboratory methods for monitoring disease response and defining disease eradication. Recent literature indicates that achievement of remission based on sensitive MFC criteria is prognostically superior to remissions defined by older criteria based on monitoring of paraproteins in blood and urine. As such, the International Myeloma Working Group (IMWG) recommends MFC for minimal residual disease (MRD) testing during clinical trials for multiple myeloma (MM). This study validates a sensitive 9-color, 11-parameter, two-tube MFC assay suitable for both the initial diagnosis of patients with paraproteinemias and MRD monitoring in MM.

Method
We established and validated a 9-color, 11-parameter MFC assay in our laboratory. The 9-color tube contained cytoplasmic κ/cytoplasmic λ/CD45/CD38/CD56/CD138/CD19/CD117/CD20. Results of the MFC were compared to bone marrow microscopic examinations, immunohistochemical (IHC) studies and serum/urine M-protein measurements ordered on 363 samples from patients with documented or suspected PC neoplasms and IgM paraproteinemias.

Results
Total preparation time is approximately 90 minutes (“hands-on” time under 30 minutes) for up to 3 concurrent specimens. The assay is performed in two tubes (control and PC panel) with analysis of up to 1.8 x 106 total bone marrow cells/tube. The mean instrument analysis time was ∼5 minutes/tube (range 1-9 minutes). MFC detected clonal PCs in 19% and 23% of cases in which IHC or morphologic evaluation, respectively, failed to show a clonal plasma cell population. The MFC assay consistently detected clonal PC in bone marrow aspirates when serum M-protein levels were above 1g/dl. The frequency of clonal PC detection by MFC fell in concert with M-protein levels. However, in 11% of patients, MFC detected clonal PC after serum and urine immunofixation studies turned negative.

Conclusion
We conclude that the assay described herein is suitable when immunologic studies are indicated for either primary diagnosis or MRD detection in PC neoplasms. It equals or exceeds sensitivities reported in the literature and it is readily integrated into a high volume, multipurpose clinical flow cytometry laboratory.

Disclosures:
Keren: Sebia, Inc (Norcross, GA). : Consultancy.
Role Of Cytogenetic Abnormalities At Baseline and During 5-Year Follow-Up In Multiple Myeloma Patients Treated On The Total Therapy 3 Protocol
Authors: Waheed Sarah , Sawyer Jeffrey , Rosenthal Adam , van Rhee Frits , Petty Nathan M , Usmani Saad Z , Bailey Clyde , Heuck Christoph , Hoering Antje , Barlogie Bart 
Published: October 2019

Abstract
The presence of metaphase cytogenetic abnormalities (CA) has long been considered an adverse prognosticator in multiple myeloma (MM). An update is provided of overall survival (OS) and progression-free survival (PFS) of patients treated with Total Therapy 3 (TT3). The prognostic role of CA was assessed relative to its presence at baseline (BL) and during 2 landmarks (LM) of 2 (LM2) and 5 years of follow-up (LM5). For both LM, the following CA combinations were considered: no CA at BL and up to LM; CA at BL and till LM; CA at BL, no CA till LM; no CA at BL, CA toward LM. Patients considered for evaluation had at least one cytogenetic evaluation prior to the start of therapy and at least one cytogenetic evaluation during follow-up. In Cox modeling, only patients with at least one cytogenetic evaluation prior to baseline were considered. The time-dependent variable indicating whether patients lived to reach a certain LM, and had a CA during the time between enrollment and that LM, was considered along with several typical MM predictor variables. Results showed that, for the LM2, patients with BL-no CA/LM-no CA fared best followed by the BL-CA/LM-no CA combination, with worst outcomes seen in the remainder sharing LM-CA (Figure 1a). In case of the LM5, LM-no CA was superior regardless of BL-CA status, while LM-CA conferred equally poor OS regardless of BL-CA status (Figure 1b). On multivariate analysis, LM5-CA retained the highest HR along with GEP-70-defined high-risk MM, BL-CA and creatinine elevation. We conclude that serial CA monitoring provides a simple cost-effective tool contributing to early detection of disease relapse often without clinical progression (data not shown) that also affords the early detection of MDS-associated CA, an increasing complication of MM therapy. Data will be presented on the detection at BL of GEP features forecasting CA development during 2 and 5 years of follow up.

Disclosures:
Usmani: Celgene: Consultancy, Research Funding, Speakers Bureau; Onyx: Research Funding, Speakers Bureau.
Neutrophil Akt2 Plays a Critical Role In Heterotypic Neutrophil-Platelet Interactions During Vascular Inflammation
Authors: Li Jing , Kim Kyungho , Hahm Eunsil , Gordeuk Victor R. , Hay Nissim , Du Xiaoping , Cho Jaehyung 
Published: October 2019

Abstract
Platelet-leukocyte interactions on activated endothelial cells play important roles in mediating pathological thrombosis and inflammation. Heterotypic platelet-leukocyte aggregation is mediated by the interaction of two crucial receptors and counter receptors; P-selectin-P-selectin glycoprotein ligand-1 and glycoprotein Ibalpha-alphaMbeta2 integrin. In spite of extensive understanding of receptor-counter receptor interactions, it remains unclear how heterotypic cell-cell interactions are regulated under thrombo-inflammatory conditions. Using real-time fluorescence intravital microscopic analysis of Akt isoform-specific knockout (KO) mice, we have demonstrated that Akt2, but not Akt1 or Akt3, plays an important role in neutrophil adhesion to the site of TNF-alpha-induced vascular inflammation. Further, heterotypic platelet-neutrophil interactions on the activated endothelium were markedly reduced in Akt2 but not Akt1 or Akt3 KO mice. Studies with chimeric mice generated from bone marrow transplants on wild-type and Akt2 KO mice revealed that hematopoietic but not endothelial cell Akt2 regulates neutrophil recruitment and platelet-neutrophil interactions during vascular inflammation. Using in vitro reconstituted systems in which platelets and neutrophils were treated with an Akt2 specific inhibitor or cells were isolated from WT and Akt KO mice, we observed that both platelet and neutrophil Akt2 play an important role in platelet-neutrophil aggregation under shear conditions. In particular, neutrophil Akt2 was critical for membrane translocation, activation, and adhesive function of alphaMbeta2 integrin. We found that the basal phosphorylation levels of Akt isoforms are significantly increased in neutrophils and platelets of patients with sickle cell disease (SCD), which is an inherited hematological disorder with vascular inflammation and occlusion. Also, SCD patients’ neutrophils show increased alphaMbeta2 integrin activation in the absence of an agonist, in comparison with healthy donors’ cells. Inhibition of Akt2 dose-dependently reduced heterotypic aggregation of patients’ neutrophils and platelets in vitro and inhibited neutrophil adhesion and neutrophil-platelet aggregation in SCD mice, thereby improving blood flow. Our results provide important genetic and pharmacologic evidence that neutrophil Akt2 regulates alphaMbeta2 integrin function and thus plays a critical role during neutrophil recruitment and neutrophil-platelet interactions under thrombo-inflammatory conditions such as SCD.

Disclosures:
No relevant conflicts of interest to declare.
STAT3 Inhibition Promotes Potent Th1 Responses By Down Regulating Pdl-1 Expression On Tumor Cells
Authors: Vasir Baldev , Rajabi Hasan , Mills Heidi , Rosenblatt Jacalyn , Stroopinsky Dina , Luptakova Katarina , Coll Maxwell D , Kristen Palmer , Kufe Donald , Avigan David 
Published: October 2019

Abstract
STAT3 signaling is upregulated in cancer cells and is thought to be a critical mediator of tumor associated immune suppression. We examined the relationship of STAT3 and PDL-1, an important negative regulator of T cell immune responses via interactions with PD-1. The 5'upstream, putative promoter region of PDL-1 gene contains four potential STAT3 binding sites close to transcription start site, suggesting a possible role of STAT3 in transcriptional regulation of PDL-1. We investigated for a possible relationship between STAT3 and PDL-1 in multiple myeloma (MM-RPMI) cell line. Using chromatin immunoprecipitation assay (ChIP assay), we observed direct binding of STAT3 to the PDL-1 promoter region in MM-RPMI cells. Moreover, an enhanced occupancy of STAT3 on the PDL-1 promoter region was also observed when the cells were treated with IL-6 (20 ng/ml). Concurrent with the above observation, IL-6 treated MM-RPMI cells showed an enhanced expression of PDL-1 protein. Treatment of RPMI-MM cells with STAT3 inhibitors AG490 and CDDO-Me for 72 hrs resulted in PDL-1 expression levels of 24.5% and 15.1% respectively, compared to 97.5% for untreated control cells as determined by FACS analysis. Based on these observations, we evaluated the effect of STAT3 inhibition by exposure to small molecule STAT3 inhibitor on response to a tumor vaccine consisting of myeloma cells fused with dendritic cells (DC). DC/myeloma fusion cells were co-cultured with autologous T cells in the presence or absence of small molecule STAT3 inhibitor (CDDO-Me). Treatment of the cocultures with CDDO-Me resulted in an increased in the percentage of CD4/IFNγ (11.57±SEM 1.16; n=3) and CD8/IFNγ (23.37±SEM 2.25; n=3) expressing cells as compared to untreated cocultures (3.1± SEM 0.72; n=3) and (4.43±SEM 0.81; n=3) respectively. Moreover, we also observed a concomitant decrease in both CD4+ and CD8+ T cells expressing IL-10 or IL-4 compared to untreated cocultures. We also noted a decrease in the percentage of immunosuppressive regulatory T cells (CD4+/CD25+/FOXP3+) in treated cocultures (8.33±SEM 0.59; n=3) as compared to untreated cocultures (28.50±SEM 2.0; n=3). These findings indicate a pivotal role of inflammatory cytokine IL-6 and its downstream activator STAT3 in regulating PDL-1 expression. Thus, strategic inhibition of STAT3/IL-6 pathway and therefore down regulation of PDL-1 could enhance the potency of tumor/DC fusion cell based cancer vaccines for immunotherapeutic treatments. Furthermore, the data demonstrate that the availability of novel adjuvants offers the potential for combination therapy that may have profound effect on the potency and durability of responses to cellular immunotherapy.

Disclosures:
No relevant conflicts of interest to declare.
Factors Influencing Peripheral Blood Progenitor Cell Yield After 2nd Mobilization With G-CSF In Allogeneic Donors
Authors: Hölig Kristina , Kramer Michael , Zimmer Kristin , Blechschmidt Matthias , Poppe-Thiede Kirsten , Bornhäuser Martin , Ehninger Gerhard 
Published: October 2019

Abstract

Purpose
2nd collection of Peripheral Blood Progenitor Cells (PBPC) from the same donor is increasingly required for the treatment of complications after allogeneic transplantation, but information regarding the efficacy of this procedure is still limited.

Methods
Within our program 212 allogeneic donors underwent a 2nd cycle of G-CSF-application and PBPC leukapheresis between 1996 and 2012. Demographical data and details of donation are shown in table 1. G-CSF (lenograstim) was administered at a median daily dose of 7.5µg/kg for 4.5d; leukapheresis was performed on day 5 and 6, if necessary. Peripheral CD 34+ counts and leukapheresis yields after both mobilization cycles were compared. The influence of age, sex, BMI, G-CSF schedule and the interval between the donations on the efficacy of the 2nd PBPC mobilization were analyzed, too.

Results
Median CD 34+ counts in peripheral blood were 53.2/µl at day 5 of 1st mobilization compared to 49.5/µl at day 5 of 2nd mobilization. Between 1st and 2nd donation, the median yield of CD 34+ cells/kg recipient b.w. decreased from 7.3 x 106 to 6.6 x 106 (p = 0.026, see table 2). The number of aphereses necessary in both donation episodes did not differ in the majority of donors (n=175; 82.5%). A strong correlation was observed between the amount of CD 34+ cells per recipient b.w. after the 1st and 2nd PBPC donation (r=0.89). The majority (65.4%, n=17) of donors with poor mobilization (<3 x 106 CD 34+ cells/kg recipient b.w.) after the 1st leukapheresis reached similar low CD 34+ counts after the 2nd mobilization. The interval between both mobilization cycles did not affect the results of the 2nd donation. According to an adjusted model, the other variables (age, sex, BMI and G-CSF-schedule) had no additional influence on mobilization efficacy in the 2nd cycle of G-CSF application.

Conclusions
In the largest group of healthy donors reported so far, we could demonstrate a slight reduction (about 10%) of PBPC yield after the 2nd mobilization that might not be clinically significant. Demographic variables affected the results of both mobilization episodes in the same way and the interval between them was of no significance. Our data support the assumption, that stem cell mobilization is probably determined by genetic factors and the outcome of a 2nd donation can easily be predicted by the results of the 1st one.

Disclosures:
No relevant conflicts of interest to declare.