bluuebunny/crossref_metadata_2025_split huggingface dataset:
Application of a Standardized Flow Cytometry Panel for Defining and Monitoring the Immunophenotype of CAR-T Cells
Authors: Scarfò Irene , Gallagher Kathleen , Maus Marcela V. , Larson Rebecca , Sheehan Maegan , Brini Gabrielle , Bornheimer Scott , Hanley Mary Beth , Frigault Matthew , Leick Mark , Chekmasova Alena 
Published: November 2019

Chimeric antigen receptor T-cells (CAR-T) have emerged as an extremely promising therapy for hematological malignancies. The immunophenotype of apheresis material and the CAR-T cell product is known to be predictive of the likelihood of response to treatment of certain malignancies. Central memory and stem cell-like memory T cell phenotypes are associated with a more sustained proliferative response and long-term CAR-T persistence (Fraietta et al, Nature Medicine, 2018).
There is an unmet need for standardized methods and reagents to reliably profile the memory phenotype of CAR-Ts to better evaluate product quality, and support improvements in CAR-T manufacturing. The BD Biosciences dried memory T-cell panel contains a pre-validated mixture of 7 antibodies for the identification of naïve, stem cell memory, central memory and effector memory CD4+ and CD8+ T cell subsets. The pre-mixed dried antibody tube offers consistency in staining profiles over time and reduces the risk of operator errors. Additional drop-in antibodies can complement the panel and enable more in-depth evaluation of the T cell phenotype. Here we demonstrate the use of this panel with drop-in markers to monitor changes in expression of PD-1, TIM-3, LAG-3, HLA-DR, CD45RO, and CXCR3 on T cells transduced to express our novel anti-CD37 CAR. Cells were stained at day 0 prior to transduction, day 7, and following resting and re-stimulation, and acquired on a 12 color BD FACS Lyric.
The use of a standardized memory T-cell panel will allow us to more accurately evaluate how T-cell phenotype impacts on the efficacy and longevity of response in patients receiving CAR-T therapies.

Disclosures
Maus: INFO PENDING: Other: INFO PENDING. Bornheimer:BD Biosciences: Employment. Hanley:BD Biosciences: Employment. Frigault:Novartis: Patents & Royalties: Royalty; Arcellx, Celgene, Foundation Medicine, Kite/Gilead, Nkarta, Novartis, and Xenetic: Consultancy.
Clostridium Difficile Infection in Hematopoietic Stem Cell Transplant Patients: A National Study
Authors: Vishnuvardhan Nivetha , Naaraayan Ashutossh , Hasan Amrah , Jesmajian Stephen 
Published: November 2019

Introduction
Clostridium difficile infection (CDI) is the leading cause of infectious diarrhea among hospitalized patients. Patients undergoing hematopoietic stem-cell transplant (HSCT), either autologous stem cell transplant (Auto-SCT) or allogeneic stem cell transplant (Allo-SCT), are at high risk for CDI. We aim to describe the incidence and epidemiological trends for CDI in Auto-SCT and Allo-SCT in the United States, using the National Inpatient Sample (NIS) database provided by Healthcare Cost and Utilization Project (HCUP).
Methods
A descriptive, retrospective study was conducted on the National Inpatient Sample (NIS) Database for the years 2005-2014. Admissions with diagnosis of CDI were selected based on International Classification of Diseases-Ninth Revision (ICD-9), Clinical Modification diagnoses codes (00845). ICD-9 procedure codes were used to identify HSCT hospitalizations (autologous: 4101, 4104, 4107, and 4109; allogeneic: 4102, 4103, 4105, 4106, and 4108). Complex survey design, weights and clustering were accounted for during analysis.
Results
The mean age of patients undergoing HSCT was 45.69 + 20.35 years and 41.13% were female. There were total of 89,374 Auto-SCT patients and 66,044 Allo-SCT patients hospitalized between 2005-2014. Auto-SCTs constituted 55.5% of all the HSCTs. The incidence of Auto-SCT related hospitalizations increased from 205 per million admissions (pma) in 2005 to 291 pma in 2014 with an annual percentage change (APC) of 5.8, p<0.001. During the same period the incidence of Allo-SCT related hospitalizations increased from 151 pma to 224 pma with and APC of 6.3, p<0.001. Overall 6.67% of patients with Auto-SCT and 9.96% of Allo-SCT were diagnosed with CDI. There were statistically higher CDI rates in Allo-SCT when compared to Auto-SCT (p<0.001). The incidence of CDI in Auto-SCT patients showed a non-significant decrease from 2005-2009 and a significant increase from 2009-2014 (APC 7, p<0.001). The incidence of CDI in Allo-SCT patients increased significantly from 2005 to 2014 with an APC of 5.9, p<0.001.
Conclusion
The number of individuals being hospitalized for HSCTs in the United States is increasing and this increase is seen for both Auto-SCT and Allo-SCT procedures. One in fifteen patients undergoing Auto-SCT and one in ten patients undergoing Allo-SCT were diagnosed with CDI during hospitalization. There were significantly more CDIs in patients with Allo-SCT when compared to Auto-SCTs. The incidence of CDI in patients undergoing both Autologous and Allogeneic HSCT increased from 2005-2014 in the United States.

Disclosures
No relevant conflicts of interest to declare.
Features of an Unusual AML with t(4;12)/Pdgra-ETV6 As the Sole Cytogenetic Abnormality Associated with Myelodysplastic Changes: A Case Report and Literature Review
Authors: De Jitakshi , Medeiros L. Jeffrey 
Published: November 2019

The t(4;12) cytogenetic abnormality is rare in AML, and is associated with a unique morphologic and immunophenotypic profile. Pseudo-lymphoid morphology of myeloid blasts (appearing as atypical lymphocytes), multilineage dysplasia, absence of eosinophilia, aberrant CD7 and lack of myeloperoxidase (MPO) expression, low frequency of FLT3 mutations, and treatment refractoriness are some of the characteristics of this AML subtype previously described in the literature.
We report a case of AML with t(4;12) in a middle-age male who presented with pancytopenia: WBC 3.34 (65% lymphocytes), Hgb 6.9, Plt 13, and no known prior history of a myeloid neoplasm. Bone marrow aspirate revealed 78% blasts that appeared large with uneven chromatin distribution, prominent nucleoli, and inconspicuous granules; and significant dysplastic features in the erythroid and megakaryocyte elements. Immunostains performed on the trephine core biopsy showed blasts expressed CD34 (subset), CD71 (subset), and were CD117+ Bcl-2+ CD43+ CD7+ MPO-. Flow cytometry identified 74% myeloblasts, positive for CD34, CD38, CD123, CD117, CD33, HLA-DR, CD13 (subset), CD56, CD7, CD71; negative for monocyte-specific markers. The morphologic and immunophenotypic features were compatible with AML with MDS-related changes.
Karyotype analysis identified t(4;12)(q12;p13) in 18/20 cells, and no additional chromosomal abnormalities. PDGFRA translocation to 12p was confirmed using PDGFRA and ETV6 FISH probes. Analysis of frequent genomic alterations in AML identified BCOR p.Leu1646Profs, TET2 p.Gln1627Ilefs, U2AF1 p.Ser34Phe mutations; and no FLT3 mutation.
Conclusions: In AML with t(4;12)/PDGFRA-ETV6, though lack of a FLT3 mutation is relatively favorable, BCOR and U2AF1 mutations and multilineage dysplasia are unfavorable prognostic factors. Synergism of the additional oncogenic mutations with the PDGFRA activating mutation can be postulated. The t(4;12)/PDGFRA-ETV6 should be recognized in the WHO classification as a recurrent balanced translocation in AML with MDS-related changes.

Disclosures
No relevant conflicts of interest to declare.
M6PR-Specific Targeting of Lysosomal Heparanase Regulates Platelet Hemostatic Function in Mice
Authors: Eaton Nathan , Drew Caleb , Dlugi Theresa A. , Hoffmeister Karin M. , Falet Hervé 
Published: November 2019

Besides α-granules and dense granules, which play critical roles in and beyond hemostasis, circulating blood platelets and their precursor cells megakaryocytes contain lysosomes, the contents of which are also secreted during platelet activation. In their delivery to the lysosome, acid hydrolases bearing phosphomannosyl residues are trafficked from the trans-Golgi network to the acidic late-endosomal compartment via the mannose 6-phosphate receptor (M6PR) pathway. To determine the role of M6PR-specific targeting of lysosomal enzymes in platelet function, platelet parameters were investigated in M6pr-/- mice lacking the 46-kDa M6PR, the physiological role of which is unclear. M6pr-/- mice had normal platelet count but displayed an increased number of distinct proplatelet-like cells compared to control mice, as determined by immunofluorescent microscopy. Moreover, transmission electron microscopy revealed the presence of abnormal membrane tubulations, elongated and electron-dense granules, and large vacuole-like structures within resting M6pr-/- platelets. M6pr-/- platelets expressed normally major glycoproteins on their surface and von Willebrand factor and fibrinogen in their α-granules. M6pr-/- mice were hyper-thrombotic, as assessed by tail bleeding time, and M6pr-/- platelets adhered to type I collagen with a significantly greater propensity than control platelets under arterial shear in in vitro flow experiments. Heparanase, an endo-β-glucuronidase that cleaves extracellular matrix heparan sulfate proteoglycans, is the most abundant lysosomal enzyme in platelets. Thus, its contribution to the phenotype of M6pr-/- mice was investigated further. Heparanase expression was decreased in the bone marrow megakaryocytes and blood platelets of M6pr-/- mice and increased in M6pr-/- plasma, as evidenced by immunoblot and fluorescent microscopy analysis, consistent with its mistargeting in the absence of M6PR. Interestingly, pharmacological inhibition of heparanase with OGT 2115 normalized the adhesion of M6pr-/- platelets to collagen in vitro, indicating that increased plasma heparanase contributes to the thrombotic phenotype of M6pr-/- mice. Taken together, the data suggest that the M6PR-specific targeting of lysosomal heparanase plays a critical role in platelet function, thereby regulating hemostasis.

Disclosures
No relevant conflicts of interest to declare.
Simultaneous Inhibition of BCR-ABL1 Tyrosine Kinase and PAK1/2 Serine/Threonine Kinases Exerts Synergistic Effect Against Chronic Myeloid Leukemia Cells
Authors: Flis Sylwia , Bratek Ewelina , Chojnacki Tomasz , Piskorek Marlena , Skorski Tomasz 
Published: November 2019

Tyrosine kinase inhibitors (TKIs) revolutionized the treatment of BCR-ABL1 tyrosine kinase - positive chronic myeloid leukemia in chronic phase (CML-CP). However, it is unlikely that TKIs will "cure" the disease in majority of patients because CML-CP cells are elusive targets even for the most advanced therapies employing second and third generation of TKIs. Therefore, new treatment modalities are necessary to improve therapeutic outcomes.
We showed before that class I p21-activated serine/threonine kinases (PAKs) remained active in TKI-naive and TKI-treated CML-CP leukemia stem and early progenitor cells. The aim of the study was to test whether simultaneous inhibition of signaling pathways activated by BCR-ABL1 and PAK kinases may improve the treatment of CML. Special attention was focused on PAK1 and PAK2, which are expressed in hematopoietic cells and can play an important role in the promotion of CML cells proliferation and survival. PAK kinases were targeted by small molecule inhibitor IPA-3 (inhibitor of PAK1) and shRNA construct for PAK2, BCR-ABL1 kinase was inhibited by imatinib. The studies were carried out using (i) primary CML-CP stem/early progenitor cells and normal hematopoietic counterparts isolated from the bone marrow of newly diagnosed CML-CP patients and healthy donors, respectively, (ii) CML-blast phase cell lines (K562 and KCL-22), and (iii) BCR-ABL1-transformed 32Dcl3 cell line cells.
We show here that inhibition of PAK1 or/and PAK2 kinases activity enhanced the effect of IM against CML cells without affecting normal counterparts. We observed that the combined use of IM with IPA-3 increased growth inhibition and apoptosis of leukemia cells. To evaluate the type of drugs interaction median effect analysis method was used. The studies revealed that the type and strength of drug interaction depend on drug concentration. Generally, combination of IM with IPA-3 at the 50% of the cell kill level (EC50) generated synergistic effect. Altogether, we postulate that BCR-ABL1 kinase inhibitor should be combined with PAK1/2 inhibitor to facilitate eradication of CML cells.

Disclosures
No relevant conflicts of interest to declare.
Vaccine Response to a Neo Polyssacharide Antigen Typhim Vi on the Identification of Secondary Immunodeficiencies in Hematological Malignancy
Authors: Ochoa-Grullón Juliana , Benavente Cuesta Celina , Cordero Torres G , Pérez López Cristina , Peña Cortijo Ascensión , Rodríguez de la Peña Antonia , Álvarez Carmona Ana , Mateo Morales Marta , Rodríguez de Frías Edgard , Guevara-Hoyer Kissy , Fernández-Arquero Miguel , Fernández-Paredes Lidia , Perez de Diego Rebeca , Sánchez-Ramón Silvia 
Published: November 2019

An increasing healthcare challenge in the management of hematological malignancy (HM) is secondary immunodeficiency (SID), either caused by the underlying malignancy or by treatments, including B-cell-targeting therapies. From January 2019, the EMA (European Medicines Agency) included the evaluation of specific antibody (Ab) responses to better select patients for immunoglobulin replacement therapy (IgRT). Antibody response to polysaccharide immunizations is primordial in the immunological evaluation of the patients and it is thought to be the first Ab response to be lost in the rising of an immunodeficiency. Clinically, an impaired polysaccharide response means that the patient may not be protected to new encounters with encapsulated bacteria, having therefore higher risk of severe or recurrent infections. We evaluated Ab responses to pneumococcal (PPV) and Salmonella typhi (TV) pure polysaccharide immunization in a cohort of 42 HM patients and 24 healthy controls. Pre-post specific Ab serum concentrations were measured by ELISA (Binding Site, UK), before immunizations and at 4 weeks. Globally, the prevalence of TV titers pre-immunization was lower (9%) compared to PPV pre-immunization titers (76%) (p&lt;0.001). There were more HM patients' non-responders for TV (88%, unable to mount a &gt;3-fold increase in TV titers) than PPV non-responders (62%) (p&lt;0.0001), and no-response to TV correlated better to infectious history. By ROC analysis, pre-post 5-fold TV increase was the best cut-off to discriminate HM with recurrent infections and controls (sensitivity 91%, specificity 100%). TV Ab responses were superior to PPV to identify SID and defective primary responses, notably in patients with high PPV baseline titers in the conjugate vaccine era, therefore overcoming the limitations in identifying patients with impaired polysaccharide responses after pneumococcal conjugate vaccine immunizations. Our results suggest that specific Ab response to S typhi polysaccharide is a useful complementary assay in the diagnosis and management decision of SID to HM, that may help to predict patients more prone to infections and guide a more risk-adapted infection-prophylaxis.

Disclosures
No relevant conflicts of interest to declare.
Ruxolutinib in Steroid Refractory Graft Versus Host Disease (SR-GVHD): Systemic Literature Review
Authors: Mohammed Saleh Mostafa F. , Hashmi Shahrukh K. 
Published: November 2019

Background: Graft versus host disease (GVHD) is a main cause of morbidity and mortality in patients having undergone allogeneic hematopoietic stem cell transplantation (HSCT). About 30-40% of patients have steroid‐refractory GVHD (SR‐GVHD) after the first‐line use of high doses of corticosteroids with a poor prognosis .Ruxolitinib is a promising treatment for SR-GVHD. However, data regarding optimum dosing, response rates and associated adverse events are scarce. Herein, we provide the first systemic review of literature for the use ruxolutinib in GVHD.
Methods: A Medline (PubMed), google scholar, OVID and Cochrane Database of Systematic Reviews search using key words "Ruxolutinib and GVHD", "Ruxolutinib and SR-GVHD" was undertaken in June 2019. Only peer reviewed databases were searched and search was restricted to human studies of acute and chronic GVHD only.
Results: 16 publications, as listed in Table 1. Only one was a prospective trial, all others were retrospective studies, case series (5), and case reports (2). Overall response, ranged 45% - 100%, complete response was noted in 5.2% -80% patients. Time to response was variable from 1-12 weeks. Cytopenias and infectious complications were frequently reported with dose reduction or interruptions needed in most studies. Maintained responses were reported in a small proportion after ruxolutinib discontinuation.
Conclusion Ruxolutinib has promising efficacy in SR-GVHD , however cytopenias and infectious complications reported frequently mandate close monitoring. Results of ongoing prospective trials could provide answers for optimum dosing and response assessment, and management of related adverse events.
Table

Disclosures
No relevant conflicts of interest to declare.
An External Validation Study for Current Risk Scoring Models in Elderly Patients with Newly Diagnosed Acute Myeloid Leukemia
Authors: Liu Chia-Jen , Ying-Chung Hong , Yeh Chiu-Mei , Tsai Chun-Kuang , Hsiao Liang-Tsai , Wang Hao-Yuan , LIU Yao-Chung , Gau Jyh-Pyng , Ko Po-Shen , Liu Jin-Hwang , Chen Po-Min 
Published: November 2019

Background
Acute myeloid leukemia (AML) is a common hematologic neoplasm in the elderly. The high mortality in elderly AML patients is reported to be associated with old age, poor performance status, and several disease characteristics. Several risk stratification models have been reported. Our aims were to explore risk factors for early mortality in older AML patients and compare the discrimination ability of existing prognostic models.
Methods
We enrolled newly diagnosed AML patients age 60 and above at Taipei Veterans General Hospital, a national medical center in Taiwan, between July 1, 2008 and May 31, 2017. Our primary endpoint was early mortality, defined as death within two months after initial AML diagnosis. Performance of several existing scoring systems were compared by using the Akaike information criteria (AIC) and Bayesian information criterion (BIC) calculations. Model discrimination ability was also estimated by Harrell's C statistics.
Results
A total of 478 AML patients were diagnosed during the eight-year follow-up period. After excluding young patients (age < 60) and those without a histopathologic diagnosis, the final cohort included 277 patients. The median age was 74 (range 60-96), and 171 (61.7%) of them were male. One hundred sixteen patients (41.9%) had Eastern Cooperative Oncology Group performance (ECOG) ≥ 2 and 33.9% patients had poor/adverse cytogenetics or molecular abnormalities. The two-month mortality rate was 29.9% (95% confidence interval [CI] 24.8%-35.9%). Age ≥ 80 (adjusted HR 1.95, 95% CI 1.12-3.42), having an antecedent hematologic disorder (adjusted HR 1.86, 95% CI 1.01-3.43), ECOG ≥ 2 (adjusted HR 2.06, 95% CI 1.20-3.54), complex karyotype (adjusted HR 3.13, 95% CI 1.76-5.55), BM blasts ≥ 70% (adjusted HR 1.79, 95% CI 1.02-3.13), WBC ≥ 100 ×109/L (adjusted HR 3.27, 95% CI 1.58-6.75), and creatinine > 1.3 mg/dL (adjusted HR 2.04, 95% CI 1.23-3.39) were identified as independent predictors for early mortality in the multivariate analysis. Furthermore, we systematically reviewed existing prognostic models for elderly AML. We found five scoring models that don't require additional specific examinations beyond clinical practice and later applied them to our elderly AML cohort. The performance of the five models is shown in Table. Kantarjian's prognostic model (Kantarjian H, et. al. Blood 2010) had the highest Harrell's C statistic and the ALMA score (Ramos F, et. al.Leukemia Research 2015) had the lowest AIC and BIC compared with the other prognostic models.
Conclusion
We identified seven risk factors for early mortality and compared the performance of five prognostic models for elderly AML patients. The finding may help clinicians to stratify patients and initiate appropriate management.
Table

Disclosures
No relevant conflicts of interest to declare.
Good Tolerance and Durable Remission for Anti-CD19 Chimeric Antigen Receptor T-Cell Therapy in Refractory/Relapsed Mantle Cell Lymphoma
Authors: Ye Shiguang , Zhou Lili , Li Shaoguang , Li Ping , Liang Aibin 
Published: November 2019

Mantle cell lymphoma (MCL) is one subtype of the B cell non-Hodgkin lymphomas (NHL) and has a distinguash course. Despite all efforts, MCL is nearly incurable that refractory or relapse(r/r) course will occur in almost all patients. Due to the rapid development of chemotherapy resistance, it is difficult to treat r/r MCL. A number of treatment regimens have been evaluated with various degrees of success. Cellular immunotherapy has the potential to lead significant and sustained clinical remissions in lymphoma patients, in which modification of T cells with chimeric antigen receptors (CARs) is a powerful regimen. T cells containing CARs with costimulatory domains show better activity against tumor cells. We conducted a clinical trial testing a "second-generation" CD19-specific CAR with 4-1BB costimulatory domains in patients with r/r MCL.
Six patients with r/r MCL were enrolled and all were received T-cell infusions. The age of patients ranged from 52 to 81 years old. Five patients were male and one patient was female. Five patients had stage IV MCL with bone marrow involvement and one patient was stage II. Four patients had received more than 3 lines regimens and two patients had received first-line chemotherapy regimen. Four patients received FC regimen before CAR-T infusion and two patients were not received fludarabine due to age ≥80 years old.
Treatment was well tolerated, two patients developed transient infusion reaction. No patients have grade 3-4 Cytokine Release Syndrome (CRS). Three patients have grade 2 CRS and two patients have grade 1 CRS. Objective response rate is 66.67%. One patient achieves complete remission(CR) and three patients have partial remission(PR). The response of the other two patients is stable disease(SD). The patient who has CR response has remained progression-free for 36 months and is still being followed up. Two patients who have PR response have progressed in 3 months and the other one has durable response for 7 months. In the patients who has response, the peak of fever appears at approximately day 10 after CAR-T infusion, moreover interleukin(IL)6,IL2αand interferon(IFN)-γ almost reaches the peak on the day 14.
In conclusion, adoptive immunotherapy with CD19-specific T cells was well tolerated and was associated with antitumor activity. Further large-sample and multicenter studies are needed to fully evaluate this strategy for such patients.

Disclosures
No relevant conflicts of interest to declare.