bluuebunny/crossref_metadata_2025 huggingface dataset:
Rapamycin (Sirolimus) and Rap-536 Increase Red Blood Cell Parameters through Distinct Mechanisms in Wild-Type and Thalassemic Mice
Authors: Acar Melih , Jupelli Madhulika , MacBeth Kyle , Schwickart Martin
Published: November 2020

Luspatercept, a fusion protein composed of a modified activin receptor type-2B (ActRIIB) and IgG1 Fc, is a novel erythroid maturation agent that has been shown to increase red blood cell (RBC) and hemoglobin (Hb) levels. Luspatercept binds to and sequesters several endogenous transforming growth factor-beta superfamily ligands, thereby diminishing Smad2/3 signaling. RAP-536, the murine analog of luspatercept, has been shown to induce erythroblast maturation and increase RBC, Hb, and hematocrit levels in wild-type (WT) mice and in murine disease models of beta-thalassemia and myelodysplastic syndromes. Previous studies have shown that rapamycin (sirolimus), an mTOR inhibitor, improved anemia in a mouse model of beta-thalassemia by activating the autophagy pathway.

In this study, rapamycin was tested in combination with RAP-536 by treating WT mice and th3/+ mice (a beta-thalassemia disease model; B6.129P2-Hbb-b1tm1Unc Hbb-b2tm1Unc/J) with vehicle, RAP-536 10 mg/kg twice weekly, rapamycin 4 mg/kg daily, or RAP-536 plus rapamycin for 2 weeks.

Single-agent dosing with either RAP-536 or rapamycin increased RBC, Hb, and hematocrit levels significantly compared with vehicle treatment in both WT and th3/+ mice (Figure). The percentage increase in RBC, Hb, and hematocrit levels upon RAP-536 or rapamycin administration was substantially higher in th3/+ mice than in WT mice. Interestingly, much higher and significant increases in RBC and Hb levels in both WT and th3/+ mice, and in hematocrit levels in th3/+ mice, were observed with RAP-536 plus rapamycin, when compared with the single agents. For example, in th3/+ mice, the increase in Hb levels compared with vehicle control was 19.2% with RAP-536 alone, 13.0% with rapamycin alone, and 44.7% with RAP-536 plus rapamycin (Figure). In th3/+ mice, RAP-536 plus rapamycin treatment also significantly reduced spleen enlargement compared with vehicle (P < 0.001). RAP-536 increased and rapamycin decreased overall reticulocyte levels and immature reticulocyte relative percentages in whole reticulocyte populations in the blood of WT mice, suggesting that RAP-536 and rapamycin have different mechanisms of action through which they increase RBC and Hb levels.

Altogether, these preclinical results provide a rationale for combining rapamycin, and potentially other mTOR inhibitors, with luspatercept to treat anemia associated with beta-thalassemia.

Disclosures

Acar: Bristol Myers Squibb: Ended employment in the past 24 months. Jupelli:Bristol Myers Squibb: Current Employment. MacBeth:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company. Schwickart:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company.
DAC+CAG Compared with the Standard 3+7 Regimen for Induction of the Medically Fit Elderly Acute Myeloid Leukemia: An Open-Label, Multicenter, Randomized, Phase III Trial
Authors: Liu Miaomiao , Zhang Yu , Zheng Weiyang , Liu Qifa
Published: November 2020

Background :Elderly subjects with acute myeloid leukemia have a very poor prognosis,only 40%-60% achieving a complete remission(CR) with intensive chemotherapy. Hypomethylating agents such as decitabine show activities in elderly AML cases. Method:This multicenter, open-label, randomized phase III trial compared the outcomes between decitabine combined with modified CAG regimen (DAC+CAG) and standard daunorubicin or idarubicin plus cytarabine (3+7) for medically fit elderly AML patients.Patients were randomly assigned to the two group in 1:1 ratio approximately. Results:A total of 110 newly diagnosed AML patients selected according to age (range 60-75), performance status (PS≤2), Charlson Comorbidity Index (CCI≤2) were analyzed.CR rate was 58.2% and 61.8% after the first cycle(P=0.963), and 70.9% and 69.1% after two cycles(p=0.780), respectively,in the DAC+CAG group and 3+7 group. There was no statistically difference of ORR after two cycles between the two groups (74.5% vs 76.4%,P=1.000).With a median follow-up of 18 months, the 1-year,3-year OS was 67.4% (95% CI:53.7-81.1%),38.9% (95% CI:16.8-61%) and 55.7%(95% CI:42-69.4%),37.6% (95% CI:20-54.8%) in the DAC+CAG and 3+7 group,respectively(p=0.383).And the 1-year,3-year LFS was 66.8% (95% CI:51.9-81.7%),34.2% (95% CI:22.2-64.2%) and 55.7% (95% CI:40-69.4%),35.8% (95% CI: 19.2-52.4%) in the two groups,respectively(p=0.255). Grade 3 or worse adverse events ,such as neutropenia (38 [69.1%] vs 46 [83.6%]), thrombocytopenia (38 [69.1%] vs 46 [83.6%]),infections(17 [30.9%] vs 25 [45.5%]) and septicemia(3[5.5%] vs 11 [20%]) are more common in 3+7 group than DAC+CAG group.The 8-week cumulative incidence of induction chemotherapy-related mortality(iTRM) were 5.5% and 12.7% (p=0.069) in DAC+CAG and 3+7 groups, respectively.Multivariate analysis revealed that allo-HSCT(p=0.004,HR=0.327,95%CI:0.152-0.703) and after first cycle achieved CR(CR1)(p=0.005,HR=0.348,95%CI:0.166-0.732) were favourable factors for OS,and the latter was also identified as a favourable factor for LFS (p=0.013,HR=0.480,95%CI:0.173-1.331). Conclusion: DAC+CAG regimen showed a similar CR rate and encouraging survival compared with the 3+7 regimen,it seems be a safe and effective regimen for medically fit older patients with AML.

Keywords:Acute myeloid leukemia, Elderly, DAC+CAG regimen, 3+7 regimen,medically fit patients

Disclosures

No relevant conflicts of interest to declare.
Is Absolute Lymphocyte Count at Time of Diagnosis a Predictor of Overall Survival in Non-Hodgkin Lymphoma Patients?
Authors: Zatreh Mallak , Kaur Anahat , Albawaliz Anas , Bulchandani Sheetal , Khan Ghazal
Published: November 2020

Introduction:The prognostic significance of absolute lymphocyte count (ALC) has been an area of debate in non-Hodgkin-lymphoma (NHL). Several studies have reported that lymphocyte count during and after chemotherapy independently predict outcome in patients with acute myelocytic lymphoma (AML), follicular lymphoma and diffuse large B-cell lymphoma (DLBCL). Furthermore, lymphocytopenia is a poor prognostic marker in initial staging of NHL and in relapsed DLBCL. Therefore, we aimed to investigate whether ALC at time of diagnosis is an independent predictor of overall survival (OS) in NHL.

Methods:We retrospectively reviewed patients with aggressive NHL who were seen at hospitals affiliated with Saint Luke's Health System from January 2000 to December 2017 with at least 2 year longitudinal follow up after diagnosis. We retrieved 447 patients with histopathological diagnosis of DLBCL, Burkitt's Lymphoma, Follicular Lymphoma Grade IIIB, primary Effusion Lymphoma, B-cell Lymphoma unclassifiable, and high-grade B cell lymphoma. Through chart review we determined pathological and clinical characteristics of these patients, calculated the ALC at time of diagnosis, studied the treatment patterns, response to therapy and survival in our center. ALC at time of diagnosis was obtained on 358 patients based on data availability.

Results:A multivariate analysis was conducted to see if ALC at time of diagnosis is a predictor of OS by correcting for age at diagnosis, gender, race, stage at diagnosis, international prognostic index (IPI), central nervous system involvement, histopathological diagnosis and type of chemotherapy. ALC at diagnosis after controlling all variables was not a significant predictor of OS (P: 0.7126).

Conclusion:We found that ALC at time of diagnosis did not have a statistically significant effect on OS. We believe that prospective large-scale studies are needed to determine whether ALC at time of diagnosis affects OS in NHL patients.

Table

Disclosures

No relevant conflicts of interest to declare.
Plasma Exchange for COVID-19 Thrombo-Inflammatory Disease
Authors: Thomas Mari , Arulkumaran Nishkantha , Brearley David , Alwan Ferras , Singh Deepak , Lunn Michael P , Welch Anna , Clark Samuel , Raith Eamon , Reddy Ugan , Low Ryan , Leverett David , Singer Mervyn , Scully Marie
Published: November 2020

Introduction

Severe COVID-19 disease is associated with a hyperinflammatory, pro-thrombotic state and a high mortality. A thrombotic phenotype rather than a coagulopathy is suggested and we undertook plasma exchange to determine its effects on organ function and thrombo-inflammatory markers.

Methods

Plasma exchange was carried out in seven critically ill adults with severe COVID-19 respiratory failure (PaO2:FiO2 ratio <200 mmHg) requiring invasive or non-invasive ventilatory support and elevated thrombo-inflammatory markers (LDH>800 IU/L and D dimer>1000 µg/L (or doubling from baseline). Patients received a daily single volume 3 litre plasma exchange for a minimum of five days. No other immunomodulatory medications were initiated during this period. Effects on organ function, thrombo-inflammatory markers and complications were monitored. Seven patients matched for age and baseline biochemistry were a comparator group.

Results

Coagulation screening revealed no evidence of coagulopathy. However, von Willebrand Factor (VWF) activity, antigen and VWF antigen:ADAMTS13 ratio, Factor VIII and D-dimers were all elevated. Following five days of plasma exchange, plasma levels of all the above, and ferritin levels, were significantly reduced (p<0.05, Figure 1) while lymphocyte count normalized (p<0.05). The PaO2:FiO2 ratio increased from a median(IQR) of 11.6 (10.8- 19.7) kPa to 18.1 (16.0-25.9) kPa (p<0.05). Similar improvements were not observed in controls. Acute kidney injury (AKI) occurred among 5 patients in the control arm but not in patients who underwent plasma exchange.

Conclusion

Plasma exchange was associated with an improvement in oxygenation, decreased incidence of AKI, normalisation of lymphocytes and reduction in circulating thrombo-inflammatory markers including D-Dimer and VWF Ag:ADAMTS13 ratio.

Disclosures

Thomas: Ablynx: Honoraria, Other: Advisory Board; Bayer: Honoraria, Speakers Bureau; Sanofi: Honoraria, Other: Advisory Board. Scully:Takeda: Consultancy, Speakers Bureau; Alexion: Consultancy, Speakers Bureau; Sanofi: Consultancy, Speakers Bureau; Shire/Takeda: Other: Advisory Board, Research Funding, Speakers Bureau; Novartis: Other: Advisory Board, Speakers Bureau; Takeda: Speakers Bureau; Ablynx/Sanofi: Consultancy, Other: Advisory Board, Speakers Bureau.
In Hemophilia Α Plasma Treated with Emicizumab, Factor IXa in Activated Prothrombin Complex Concentrates Is the Dominant Contributor to Enhanced Thrombin Generation
Authors: Monroe Dougald , Ezban Mirella , Hoffman Maureane
Published: November 2020

Background. Recently a novel bifunctional antibody (emicizumab) that binds both factor IXa and factor X has been used to treat hemophilia A. Emicizumab has proven remarkably effective as a prophylactic treatment for hemophilia A; however there are patients that still experience bleeding. An approach to treating this bleeding in hemophilia A patients with inhibitors is to give an activated prothrombin complex concentrate (APCC; FEIBA) (disfavored in NHF MASAC #255). APCC contains a number of coaguation factors including prothrombin, factor X (FX), and factor IX (FIX). APCC also contains activated factor X (FXa) and factor IX (FIXa). Previous work has shown that when APCCs are added to hemophilia A plasma containing emicizumab there is a significant increase in thrombin generation [J Thromb Haemost 2018;16:1580-1591]. The goal of this work was to study thrombin generation in hemophilia A plasma with emicizumab and to examine the role of the zymogen and activated components of an APCC in the increased thrombin generation.

Methods. In hemophilia A plasma, thrombin generation assays were done using 100 µM lipid and 420 µM Z-Gly-Gly-Arg-AMC with or without emicizumab at 55 µg/mL which is the clinical steady state level. The reactions were initiated with low (1 pM) tissue factor (TF). The components of APCC were studied at concentrations that should mimic the levels seen in the plasma of a patient given a dose of 50 U/kg: prothrombin 1800 nM; FX 130 nM; FIX 90 nM; and FIXa 0.4 nM.

Results. When initiated with low TF, hemophilia A plasma alone had essentially no thrombin generation. As expected, adding emicizumab enhanced thrombin generation. The addition of zymogen coagulation factors, prothrombin, FIX, and FX, separately or together gave a small increase in thrombin generation. However, addition of FIXa to emicizumab gave a large increase in peak thrombin. In hemophilia A plasma with emicizumab and FIXa, addition of prothrombin further increased thrombin generation and specifically increased the peak level of thrombin. Further addition of FX or FIX, separately or together, only minimally increased thrombin generation.

Discussion. The strong contribution of factor IXa to the effects of APCCs on thrombin generation in hemophilia A plasma depends on the presence of emicizumab. In the absence of emicizumab, a study of the individual components of APCC showed that a combination of FXa and prothrombin at levels found in APCCs had the major effect on thrombin generation [Haemophilia. 2016;22:615-24]. That study found that FIXa did not increase thrombin generation. However, in the presence of emicizumab, despite the weak solution phase affinity of the bifunctional antibody for FIXa, small amounts of FIXa were the most significant contributor to thrombin generation.

Disclosures

Monroe: Novo Nordisk:Research Funding.Ezban:Novo Nordisk:Current Employment.Hoffman:Novo Nordisk:Research Funding.
Igh Translocation with Undefined Partners Is Associated with Superior Outcome in Multiple Myeloma Patients
Authors: Mao Xue-Han , Xu Yan , Yan Yuting , Zhuang Junling , Fan Huishou , Sui Weiwei , Liu Jiahui , Deng Shuhui , Yi Shuhua , Qiu Lugui , An Gang
Published: November 2020

Background: In multiple myeloma (MM), impact of specific chromosomal translocations involving IgH (14q21 locus, including t(4;14), t(11;14), and t(14;16) etc.) has been explored extensively. However, over 15% MM patients harboring IgH translocation with undefined partners have long been ignored.

Methods: A prospective non-randomized cohort study with a total of 715 newly-diagnosed MM cases was conducted, 13.6% of whom were t(14;undefined) positive. The whole cohort was divided into four groups: no IgH split (47.7%); t(14;undefined) (13.6%); t(11;14) (17.6%); and t(4;14) or t(14;16) group (21.1%).

Results: Median OS for the four groups were 84.2, not reached (NR), 58.7, and 44.2 months respectively, with p values for t(14;undefined) vs. no IgH split, t(11;14), and t(4;14)/t(14;16) groups of 0.197, 0.022 and 0.001, respectively(Figure B).In bortezomib-based group, the survival advantage gained by t(14;undefined) group was much more significant compared to t(11;14) and t(4;14)/t(14;16) groups. Importantly, t(14;undefined) turned out to be an independent predictive factor for longer OS of MM patients in multivariate analysis, especially in the context of bortezomib-treatment. Similar results were also observed in the PUMCH external validation cohort (Figure C).

Conclusion: Collectively, our data confirmed and externally validated the favorable prognosis of the t(14;undefined) groups, especially in the era of novel agents.

Disclosures

No relevant conflicts of interest to declare.
Characterization of the Immunoglobulin Heavy- and Light-Chain Repertoires in a Single Reaction
Authors: Lowman Geoffrey , Toro Michelle , Chang Jayde , Pickle Loni , Yang Chenchen , Looney Timothy
Published: November 2020

Background

B cell repertoire analysis by next-generation sequencing (NGS) has shown particular utility in the field of hematological oncology research. Some advantages provided by NGS-based techniques include a lower limit-of-detection and simpler paths to standardization compared to flow-based methods, and the elimination of specifically designed primers often required for qPCR-based methods. Owing to primer-primer interactions and incompatibility of reaction conditions, current multiplex PCR assays require separate PCR reactions to survey each immunoglobulin chain (IGH, IGK, IGL), often leading to a longer time-to-answer for samples in which no marker is initially detected. We have developed an assay for receptor analysis based on Ion AmpliSeq technology to circumvent these issues, allowing the effective use of up to thousands of primers in a single reaction. The highly multiplexed, pan-clonality NGS assay provides for efficient detection of IGH, IGK, and IGL chain rearrangements in a single reaction.

Methods

We developed a single primer panel targeting the framework 3 (FR3) portion of the variable gene and the joining gene region of heavy- and light-chain loci (IGH, IGK, IGL) for all alleles found within the IMGT database, enabling readout of the complementary-determining region 3 (CDR3) sequence of each immunoglobulin chain. To maximize sensitivity, we included primers to amplify IGK loci rearrangements involving Kappa deletion and C intron elements. To evaluate performance, we conducted clonality assessment and limit-of-detection testing used gDNA from a total of 45 research samples representing common B cell malignancies. We included samples derived from peripheral blood, bone marrow, and FFPE-preserved tissues at input levels ranging from 100ng to 2µg. Finally, we further characterized the samples via a separate AmpliSeq-based multiplex PCR assay targeting rearranged TCRB and TCRG chains. Sequencing and clonality analysis was performed using the Ion GeneStudio S5 System and Ion Reporter 5.16.

Results

Clonality assessments carried out using gDNA collected from both cell line and clinical research samples (CLL, B-ALL, Multiple Myeloma, Burkitt's Lymphoma, NHL, and DLBCL) show a >90% overall positive detection rate. Assessment of linearity-of-response and limit-of-detection was carried out using cell lines diluted in PBL to between 10-3 and 10-6 by mass. The multi-receptor assay performs as expected, with linear response to the cell line frequency across the range tested, including the ability to detect clones of interest at 10-6.

Conclusions

These results demonstrate the robustness of our newly developed Ion AmpliSeq-assay for B cell receptor heavy and light chains. We expect this assay to simplify the workflow for clonality assessment and rare clone detection in B cell malignancy research.

For research use only.

Disclosures

No relevant conflicts of interest to declare.
CD19 CAR-T Cells Treatment Conferred a Sustained Remission in Patients with Chemotherapy-Refractory MRD in B-ALL
Authors: Lu Wenyi , Wei Yunxiong , Cao Yaqing , Xiao Xia , Li Qing , Lyu Hairong , Jiang Yili , Zhang Huan , Wang Jia , Li Xin , Jiang Yanyu , Juanxia Meng , Ting Yuan , Zhu Haibo , He Xiaoyuan , Jin Xin , Sun Rui , Sui Tao , Liu Kaiqi , Zhao Mingfeng
Published: November 2020

Background: The persistence or recurrence of minimal residual disease (MRD) after chemotherapy predicts relapse of B-cell acute lymphoblastic leukemia (B-ALL). CD19-directed chimeric antigen receptor-modified T (CD19 CAR-T) cells have shown excitable response in B-ALL. Recently, some studies have shown that ALL patients with lower burden had higher CR rate and lower risk of CRS after CAR-T therapy (Park, et al, NEJM. 2018; 378(5):439-448. Lee, et al, Blood.128 (22):Abstract 218.). However, its role in chemotherapy-refractory MRD-positive B-ALL remains unclear. Here we aimed to assess the effectiveness and safety of CD19 CAR-T in MRD-positive B-ALL patients.

Methods: Since January 2018, a total of 14 B-ALL patients with persistent (n=8) or recurrent (n=6) MRD were enrolled in the CAR-T clinical trials (ChiCTR-ONN- 16009862 and ChiCTR1800015164). The patients were from two different clinical trials about CAR-T. If patients were treated in an MRD positive state, they would be included in this analysis. All the patients received one or more infusions of autogenous CD19 CAR-T.

Results: Median age was 37.5 (13-62) years and 7 patients were female. The median dose of infused CAR-T cells was 6.78´106cells/kg, and 5 patients received more than one infusion. After one cycle of CAR-T infusion, 12 patients achieved MRD-negative remission, leading a response rate of 85.7%. Of the subgroup of 5 Ph-positive patients who subsequently underwent transplantation, 2 patients died due to transplant-related toxic effects, whereas the other 3 patients all currently alive without leukemia. Of the subgroup of 9 Ph-negative patients, 8 patients did not undergo subsequent transplantation (Figure 1). Three patients finally suffered CD19-positive relapse and 1 patient suffered CD19-negative relapse. Importantly, 4 patients (50%) are in ongoing molecular remission without transplantation, with a duration of response averaging 22.9 months (range: 12.1-28.6 months).

The most frequent adverse events were fever and hematopoietic toxicities. Ten patients (71.4%) had grade 1 or 2 cytokine release syndrome (CRS) and no patients died of CRS. At a median follow-up time of 599.5 days (range: 172-915 days), the probability of 2-year relapse-free survival and 2-year overall survival was 61.2%±14.0% and 78.6%±11.0%, respectively.

Conclusion: In conclusion, pre-emptive CD19 CAR-T treatment is an effective and safe approach and may confer a sustained remission in B-ALL patients with chemotherapy-refractory MRD.

Disclosures

No relevant conflicts of interest to declare.
Three-Dimensional Telomere Analysis Using Teloview® Technology Identifies Smouldering Myeloma Patients with High Risk of Progression to Full Stage Multiple Myeloma in a Proof of Concept Cohort
Authors: Louis Sherif , Rangel-Pozzo Aline , Knecht Hans , Mai Sabine
Published: November 2020

Multiple Myeloma (MM) remains incurable in spite of the recent development of several advanced therapeutic modalities. Similar to other incurable diseases, the early diagnosis and treatment of MM patients may lead to better prognosis and increased survival rates. Multiple myeloma follows two asymptomatic precursor lesions, monoclonal gammopathy of undetermined significance (MGUS) and smouldering myeloma (SMM) [1]. In current clinical practice patients diagnosed with MGUS or SMM are monitored but not treated. MGUS and SMM may progress to the full stage of multiple myeloma at any time without clinically detectable signals. While 1% of MGUS patients progress to full stage MM every year, 10% of SMM patients progress to full stage MM every year [1]. In this study, we report for the first time that the 3-dimensional telomeres analysis using the TeloView® software platform is able to identify SMM patients with high risk of progression to full stage MM. The prospective study we conducted included a proof of concept cohort of total 21 SMM patients, 16 patients that remained stable for over 5 years, and 5 patients progressed to full stage multiple myeloma within 1 to 3 years from point of diagnosis. The disease progression of high risk SMM patients was confirmed clinically by MM caused morbidity. TeloView® analysis of the 2 SMM patient-groups revealed with high significance (p <0.001) distinct telomeres profiles of the stable patients versus the patients who progressed to full stage MM across 5 independent telomeric parameters measured by TeloView®. The study was conducted blindly on the diagnostic specimens suggesting the ability of TeloView® analysis to stratify SMM patients at point of diagnosis. The results we report have the potential, for the first time, to guide evidence-based decisions to treat SMM patients with high risk of progression, addressing a critical unmet clinical need in the management of MM. Follow up studies including expanded cohorts are needed to validate the results of this study, and to confirm the utility of TeloView® technology to predict the progression of SMM patients on the level of the individual patient.

References: Boutros M. et al Genomic Profiling of Smouldering Multiple Myeloma Identifies Patients at a High Risk of Disease Progression. J Clin Oncol. 2020 Jul 20;38(21):2380-2389

Disclosures

Louis: Telo Genomics Corp.: Consultancy, Current equity holder in publicly-traded company. Knecht:Telo Genomics Corp.: Consultancy, Current equity holder in publicly-traded company. Mai:Telo Genomics Corp.: Consultancy, Current equity holder in publicly-traded company.
Bortezomib and Rituximab in Newly Diagnosed Adolescent and Adult CD20-Positive Philadelphia (Ph) Negative Precursor B-Cell Acute Lymphoblastic Leukemia: A Phase II Study
Authors: Goli Vasu Babu , Jain Hasmukh , Tembhare Prashant , Shetty Dhanlaxmi Lalit , Bonda Avinash , Nayak Lingaraj , Patkar Nikhil , Subramanian Papagudi Ganesan , Sengar Manju
Published: November 2020

Background:

The expression of CD20 in precursor B-cell ALL has been associated with poor outcomes. The addition of rituximab with intensive chemotherapy in this subset of ALL has led to improvement in the event-free survival. Further increment in outcomes require novel approaches. Bortezomib is an active drug in relapsed ALL as well as has synergistic activity with rituximab in B-cell lymphomas; thus the addition of bortezomib to rituximab and chemotherapy may improve the outcomes in CD20-positive precursor B-cell ALL.

Methods:

We conducted a phase II study to test the activity of bortezomib and rituximab in combination with a paediatric inspired regimen during induction therapy in newly diagnosed adolescent and adults (>14 years of age) with CD20-positive, Philadelphia (Ph)-negative precursor B-ALL, with bone marrow measurable residual disease (MRD) negativity at the end of induction (EOI) as the primary endpoint.

Results:

From December 2017 through August 2019, a total of 35 patients were enrolled. A total of 70.99% patients achieved EOI MRD negative status. MRD negative rates improved to 88% post consolidation. There was no significant increase in toxicity with addition of bortezomib and rituximab to standard chemotherapy. The incidence of neuropathy was 26% (<grade 3). After a median follow up of 18 months, event-free survival and overall survival were 81.8% and 84.7%, respectively.

Conclusion:

The combination of bortezomib, rituximab and paediatric-inspired ALL regimen is active and well tolerated in in de-novo CD20 positive Ph-negative precursor B-ALL.

Disclosures

No relevant conflicts of interest to declare.
Supplementation of Barium to Cold-Stored Platelets Improves Platelet Quality over 7 Days of Storage
Authors: Reddoch-Cardenas Kristin M , McIntosh Colby , Calhoun Cheresa , Barrera Gema , Lizarraga Stephanny , Bynum James , Cap Andrew P.
Published: November 2020

Background:

Hemorrhage is responsible for the majority of potentially survivable deaths amongst military and civilian sectors. Early interventions to manage bleeding during hemorrhagic injury, including administration of platelets (PLTs), are crucial for decreasing mortality. Cold-stored PLTs are an attractive treatment for active bleeding as they provide greater functional preservation and a reduced risk of bacterial sepsis than conventional room temperature stored PLTs. However, cold storage causes a rise in PLT intracellular calcium, subsequent activation of signaling pathways, and often results in aggregate formation. In this study, we hypothesized that storing PLTs in the cold with barium, a divalent cation chelated by citrate anticoagulant, would improve PLT quality.

Methods:

Apheresis PLTs in 100% plasma were collected from healthy donors (n=4). Briefly, 10 mL of PLTs were sterilely aliquotted and added to 15 mL mini-bags containing 0- (Control), 3-, or 6-mM BaCl2. Individual mini-bags were used for each timepoint and condition. Bags were incubated for 15 min at room temperature prior to long-term storage in a walk-in refrigerator (1°-6°C). Samples were assayed on days 1 (the day after collection) and 7 of storage for PLT metabolism, count, clot formation, clot retraction, and activation (phosphatidylserine, mitochondrial depolarization, GPIb, and P-Selectin). Data were reported as means±SD.

Results:

Compared to Control at day 7, BaCl2-treated bags exhibited higher PLT counts (Control: 422±215 x103 PLTs/µL; 3 mM BaCl2: 468±222 x103 PLTs/µL; 6 mM BaCl2: 697±194 x103 PLTs/µL), less PS exposure (Control: 6.3±3.5%; 3 mM BaCl2: 5.8±3 %; 6 mM BaCl2: 5.6±2.8 %), and less mitochondrial membrane depolarization. Additionally, glucose consumption and lactate production were lower in barium-supplemented PLTs. GPIb expression and P-Selectin was comparable amongst all groups. Hemostatic function, as assessed via thromboelastography and clot retraction, was not negatively affected by addition of BaCl2. In most assays, cold storage of PLTs with the higher concentration of BaCl2 (6 mM) seemed to confer the greatest advantages.

Conclusion:

Supplementation of barium to PLTs provides a promising avenue for transfusion medicine since PLT count is better preserved and activation is decreased compared to normal refrigerated storage. Future studies are warranted to determine the effects of barium supplementation on other aspects of cold-stored PLT function (e.g. procoagulant activity and PLT adhesion under physiologic flow) and with respect to alternate storage media (e.g. PLT additive solutions).

Disclosures

No relevant conflicts of interest to declare.
Therapeutic Effect of a Traditional Chinese Medicine Compound Mixture Compound Kushen Injection in Treating Multiple Myeloma
Authors: Jiang Hongqiang , Tu Honglei , Jin Yanxia , Wu Xianjin , Luo Ziyi , Chen Yanling , Zhang Dongdong , Wu Balu , Wei Yongchang , Yang Yi , Zhou Fuling
Published: November 2020

Abstract

Background: Multiple myeloma (MM) is a hematology malignant disease originated from B-cell line and still incurable. Compound Kushen Injection (CKI) as a Traditional Chinese Medicines are promising agents in our previous research for treating cancer. The effect of CKI on multiple myeloma was still unknown.

Methods: In vitro experiment, flow cytometry was used to evaluate effect of CKI on multiple myeloma cells. Optofluidic chip was applied to detect effect at single-cell level. And in vivo RPMI-8226 GFP+ B-NSG mouse model was built to assess the role of CKI in multiple myeloma treatment.

Results: CKI inhibited MM cells proliferation of and increased its apoptosis rate. And the cell cycle of MM cells was also arrested by CKI treatment. In contrast, CKI has few toxic effects on mesenchymal stem cells (MSCs) and MC3T3 cells. At the single-cell level, MM cells was died in time and dose dependent manner. Transcriptome find that the expression of MYC and TERT in CKI-treated RPMI-8226 cells was significantly down-regulated and confirmed by qRT-PCR and Western blot. Overexpression of TERT can partly reverse the inhibition effect of CKI on RPMI-8226 cells. B-NSG mouse was injected with GFP+ RPMI-8226 cells through caudal vein, and the disease was partially alleviated by decreased tumor burden in the CKI-treated group. Furthermore, it is surprising that in animal models with myeloma bone disease, the bone mass was higher in CKI treatment group than control.

Conclusions: CKI inhibits MM cells through the MYC/TERT signaling pathway and improve the quality of life of MM mouse. Our findings provide preclinical evidence to show that CKI could be a promising candidate in human MM therapy.

Disclosures

No relevant conflicts of interest to declare.
Vitamin D Levels Are Associated with a Dichotomous Distribution of Hematological Parameters in a Large Cohort of Otherwise Healthy Subjects
Authors: Sevindik Omur Gokmen , Olmuşçelik Oktay
Published: November 2020

Introduction

Vitamin D deficiency is generally associated with a slowed hematopoiesis and the final outcome is varying degrees of peripheral blood cytopenia(s). The net effect of increased levels of Vitamin D is not well defined. In this study we tried to document the effect of varying levels of Vitamin D on hematological parameters.

Material and Methods

Whole patient registry of Medipol Complexes screened and a total of 12709 patients who met the pre-defined inclusion criteria enrolled in the study. Two datasets formed, first including all patients (Dataset A), second including patients with no CBC abnormality and not having any nutritional deficiency (Iron, Vitamin B12 and Folic acid).

Results

There was a statistically significant correlation between major CBC indices showing a maximum for the second pre-defined (according to quartiles 1st group included patients with a Vitamin D below 10 ng/ml, 2nd group included Vitamin D between 10-20 ng/ml, 3rd group included vitamin D between 20-30 ng/ml and the 4th group included patients who had a Vitamin d level above 30 ng/ml) Vitamin D group. This trend was statistically significant for major hematological parameters and post-hoc analysis also revealed a positive trend through Group 2 (10-20 ng/ml). Regression analysis also documented that patients who were classified under group 2 possessed a more potent hematopoiesis when compared to others even when adjusted for age, gender and baseline CRP values (Figure 1).

Conclusion

Our study reports a statistically significant correlation between major hematological parameters and Vitamin D levels in a particularly large patient population who lacks a significant confounder like chronic illnesses or conditions which can contribute and potentially serves as a bias. Even more, the group who has a maximal capacity of hematopoiesis are the ones harboring relatively normal but not high levels of Vitamin D.

Figure

Disclosures

No relevant conflicts of interest to declare.
DNA Methylation-Mediated Silencing of Mir-204 Enhances the Occurrence of T-Cell Acute Lymphoblastic Leukemia By Upregulating MMP-2 and MMP-9 through the NF-Κb Signaling Pathway
Authors: Chen Dabing , Xiao Tingting , Lin Dandan , Zhu Haojie , Xu Jingjing , Luo Xiaofeng , Ren Jinhua , Zheng Zhihong , Chen Zhizhe , Hu Jianda , Yang Ting
Published: November 2020

Background : MicroRNAs (miR) are non-coding RNAs that play a role in regulation multiple functions in different cell types. Previous studies have shown that miR-204 is downregulated in T-ALL. We previously reported that matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) gene polymorphisms may be associated with the risk of T-cell acute lymphoblastic leukemia (T-ALL). The present study aims to decipher the role of miR-204 and MMP-2/MMP-9 in T-ALL occurrence to guide the diagnosis and treatment of T-ALL in the clinics.

Methods: Expression of miR-204 was determined in the bone marrow and peripheral blood samples from 70 T-ALL patients and 70 healthy volunteers by real-time quantitative PCR (RT-qPCR). Bisulfite sequencing PCR (BSP) was used to detect the DNA methylation levels of the miR-204 promoter region in T-ALL patients and T-ALL cell lines.The effect of miR-204 on cell proliferation was evaluated with the cell counting kit-8 solution (CCK-8) assay and by Hoechst and PI double staining. The binding site of miR-204 on IRAK1 was predicted by the Primer Premier 5.0 and the defined binding sequences were used to construct luciferase-tag plasmids. The regulation of IRAK1 expression by miR-204 was evaluated by RT-qPCR and Western blot analysis. With the purpose to confirm the role of MMP-2 and MMP-9 in the occurrence of T-ALL, we investigated the effect of related proteins on T-ALL cells using Western blot. To determine that miR-204 affects the occurrence of T-ALL disease by regulating the NF-KB signaling pathway, RT-qPCR and Western Blot were used for verification.

Results: DNA methylation directly affects the miR-204 expression in the promoter region when T-ALL developed. Moreover, overexpression of miR-204 inhibited the proliferation and enhanced the apoptosis of T-ALL cells. Notably, overexpression of miR-204 inhibited IRAK1, which in turn inhibited the proliferation and enhanced the apoptosis of T-ALL cells. Furthermore, IRAK1 enhanced the expression of MMP-2 and MMP-9 through phosphorylation of of p65 NF-κB, and miR-204 modulated MMP-2 and MMP-9 expression through the IRAK1/NF-κB signaling pathway.

Conclusion s : Our results demonstrate that in T-ALL cells, DNA methylation-mediated silencing of miR-204 regulates the expression of MMP-2 and MMP-9 through increased transcription of IRAK1, and activation of the NF-κB signaling pathway. These data provide a potential mechanism for the role of MMP-2 and MMP-9 in the occurrence of T-ALL. Further studies will be needed to demonstrate whether demethylation of miR-204 may be a promising treatment for T-ALL.

Disclosures

No relevant conflicts of interest to declare.
Impact of the Availability of Novel Drugs on Overall Survival of Multiple Myeloma Patients: A Real-World Study
Authors: Arcuri Leonardo Javier , Silva Cinthya Correa , Ovigli Danielle , Helman Ricardo , Kerbauy Mariana , Hamerschlak Nelson
Published: November 2020

Introduction: Multiple myeloma is an incurable hematologic malignancy. However, many novel drugs (lenalidomide, carfilzomib, daratumumab, elotuzumab and ixazomib) have recently been approved in Brazil for this disease. Proving that the availability of these drugs increases survival is not straightforward, since pivotal trials with these drugs were designed to show benefit in progression-free survival and not in overall survival.

Hypothesis: We have hypothesized that the availability of novel drugs would reduce the risk of death.

Methods: This is a real-world, unicenter analysis that included patients with multiple myeloma treated between 1993 and 2017. The availability of novel drugs (lenalidomide, carfilzomib, daratumumab, elotuzumab and ixazomib) was treated as a time-dependent covariate and analyzed by Cox model. The analysis was controlled for the International Staging System (ISS) and for age.

Results: With a median follow-up of 6 years, we included 151 patients with multiple myeloma. Fourty percent had ISS-I, 34% ISS-II and 26% ISS-III. Mean age was 60.5 y/o (SD: 12.2). There has been 51 deaths, and 5-y overall survival was 90% for ISS-I, 80% for ISS-II and 54% for ISS-III (p=0.0004). In multivariable analysis, age (HR=1.06, p<0.0001) and ISS-III (HR=4.51, p<0.0001) were risk factors for death, while the availability of two or more novel drugs was protective (HR=0.31, p=0.006). The availability of a single novel drug was also protective (HR=0.58), although it did not reach statistical significance (p=0.13). Adjusted Simon-Makuch survival plot is in figure 1 (adjusted for ISS III and 60 y/o).

Discussion:

Our results show that the availability of two or more novel drugs increases overall survival in multiple myeloma. This is an important finding, since the pivotal trials were designed to show advantages in progression-free survival and, once achieved, the experimental drug was offered to the control group, hampering overall survival analyses. Overall survival benefit, therefore, are more likely to be seen in real-world studies. We included the availability of novel drugs as a time-dependent, and the interpretation of our results is that patients alive when two novel drugs became available had benefit in overall survival. Our study has two weaknesses. Two or more novel drugs were more likely to be available for patients from a more recent time. This difference could be due to changes in supportive care, but there has been no major change in supportive care for multiple myeloma patients. Another weakness is that patient alive in the more recent years may have not received novel drug-based treatment. Considering that multiple myeloma is an incurable disease, this is unlikely for the majority of patients. In conclusion, the availability of two or more novel drugs for multiple myeloma improves survival.

Figure

Disclosures

No relevant conflicts of interest to declare.
Phase Ib Trial with Dose Expansion of the Bruton's Tyrosine Kinase (BTK) Inhibitor, Ibrutinib, in Combination with Rituximab and Lenalidomide in Patients with Refractory/Recurrent Primary Central Nervous System Lymphoma (PCNSL) and Refractory/Recurrent Secondary Central Nervous System Lymphoma (SCNSL)
Authors: Grommes Christian , Piotrowski Anna , Pentsova Elena , Nolan Craig , Francis Jasmine , DeAngelis Lisa , Schaff Lauren , Mellinghoff Ingo K
Published: November 2020

BACKGROUND: Primary CNS Lymphoma (PCNSL) is an aggressive primary brain tumor. Outcome and treatment options for patients with recurrent/refractory (r/r) disease are poor. We have observed promising efficacy of single-agent ibrutinib in r/r PCNSL and secondary CNS lymphoma (SCNSL). We hypothesize that combing ibrutinib with the immunomodulatory imide lenalidomide and theanti-CD20 antibody rituximab would be adding clinical efficacy. In this phase 1b trial, we investigate the toxicity of ibrutinib in combination with rituximab and lenalidomide in r/r PCNSL/SCNSL.

METHODS: Eligible patients had r/r PCNSL/SCNSL or newly diagnosed SCNSL, age≥18, ECOG≤2, normal end-organ function, and with any number and type of prior therapies. Enrollment followed the 3+3 design. Ibrutinib was dosed at 560mg daily (dose level 1) and 840mg (level 2, 3, 4); lenalidomide was dosed at 10mg daily (day 1-21) (level 1, 2), 15mg daily (day 1-21) (level 3) and 20mg daily (day 1-21) (level 4); rituximab was dosed at 500mg/m2 (all dose levels). Rituximab was given for 6 cycles, lenalidomide for 12 cycles, and ibrutinib ongoing.

RESULTS: Fifteen patients have been enrolled; 3 at dose level 1-3 and 6 at level 4. Median age was 69 years (range 56-82); 4 were women. Median ECOG was 1 (0: 5, 1: 7, 2: 3). Eleven had r/r PCNSL. Eleven had recurrent parenchymal disease; two had additional cerebrospinal fluid (CSF) involvement; two had both. Nine patients had recurrent and 6 refractory disease. No DLT occurred. One grade 4 lymphopenia was observed. The following grade 3 events occurred: rash (in 3 patients); lymphopenia (in 2); AST, ALT, lung infection, and elevated aPTT (in 1). The most common adverse events were thrombocytopenia, lymphopenia, and rash. No Aspergillosis infection was observed. After a median follow-up of 6.9 months, 14/15 were evaluated for response with 13/15 (73%) showing a response: 4 CR, 7 PR, and 2 SD, 1 PD. Median PFS was 3.03 months; not yet reached for Dose level 4 (only 1/6 has developed progression with a median follow up of 5 months).

CONCLUSION: Patients with CNS lymphoma tolerate the combination of rituximab, lenalidomide and ibrutinib well. No dose limiting toxicities were observed. No Aspergillosis infection occurred with this combination regimen. Continued enrollment is ongoing.

Disclosures

Grommes: BTG: Consultancy; Ono: Consultancy; Kite: Consultancy. Schaff:Debiopharm: Consultancy.
Loss of FXR Signaling Promotes Chronic Liver Injury in Sickle Cell Disease
Authors: Vats Ravi , Novelli Enrico M , Gladwin Mark T , Sundd Prithu , Pradhan Tirthadipa
Published: November 2020

Hepatic crisis is an emergent complication affecting sickle cell disease (SCD) patients, however, the molecular mechanism of sickle cell hepatobiliary crisis remains poorly understood. We have used intravital imaging to show that sinusoidal ischemia occurs in the liver of transgenic-humanized SCD mice under basal condition. SCD mice manifested progressive hepatomegaly, liver injury and hyperbilirubinemia. RNA-sequence analysis identified dysregulation of genes encoding proteins responsible for fibrosis, bile acid synthesis, and bile transport in the SCD mice liver. Live injury was the result of NF-κB-dependent inhibition of FXR signaling and its downstream targets in hepatocytes, leading to loss of canalicular bile transport and bile metabolism. Blocking NF-κB activation rescued FXR-signaling, ameliorated liver injury and resolved sinusoidal ischemia in SCD mice. Moreover, administration of FXR agonist (GW4064) also ameliorated liver injury seen in SCD mice. These findings are the first to identify that FXR-dependent impaired bile secretion promotes intrahepatic bile accumulation, which contributes to hepatobiliary injury of SCD. Improved understanding of these pathways could potentially benefit the development of new therapies to treat sickle cell hepatic crisis.

Disclosures

No relevant conflicts of interest to declare.
Towards a better understanding of the low recall of insertion variants with short-read based variant callers
Authors: Delage Wesley J. , Thevenon Julien , Lemaitre Claire
Published: November 2020

Background

Since 2009, numerous tools have been developed to detect structural variants using short read technologies. Insertions >50 bp are one of the hardest type to discover and are drastically underrepresented in gold standard variant callsets. The advent of long read technologies has completely changed the situation. In 2019, two independent cross technologies studies have published the most complete variant callsets with sequence resolved insertions in human individuals. Among the reported insertions, only 17 to 28% could be discovered with short-read based tools.

Results

In this work, we performed an in-depth analysis of these unprecedented insertion callsets in order to investigate the causes of such failures. We have first established a precise classification of insertion variants according to four layers of characterization: the nature and size of the inserted sequence, the genomic context of the insertion site and the breakpoint junction complexity. Because these levels are intertwined, we then used simulations to characterize the impact of each complexity factor on the recall of several structural variant callers. We showed that most reported insertions exhibited characteristics that may interfere with their discovery: 63% were tandem repeat expansions, 38% contained homology larger than 10 bp within their breakpoint junctions and 70% were located in simple repeats. Consequently, the recall of short-read based variant callers was significantly lower for such insertions (6% for tandem repeats vs 56% for mobile element insertions). Simulations showed that the most impacting factor was the insertion type rather than the genomic context, with various difficulties being handled differently among the tested structural variant callers, and they highlighted the lack of sequence resolution for most insertion calls.

Conclusions

Our results explain the low recall by pointing out several difficulty factors among the observed insertion features and provide avenues for improving SV caller algorithms and their combinations.