bluuebunny/crossref_metadata_2025 huggingface dataset:
Genomic Heterogeneity and Clonal Evolution in High Hyperdiploid Childhood Acute Lymphoblastic Leukemia
Authors: Woodward Eleanor L , Yang Minjun , Moura-Castro Larissa H , Spierings Diana CJ , van den Bos Hilda , Foijer Floris , Paulsson Kajsa
Published: November 2021

High hyperdiploid (HeH) B-cell precursor acute lymphoblastic leukemia (BCP ALL) is characterized by a very specific nonrandom gain of chromosomes; a feature distinct from other types of aneuploid tumor types that usually display very heterogeneous gains and losses of chromosomes. Gains of chromosomes X, 4, 6, 10, 14, 17, and 18 are seen in more than 75% of cases of HeH childhood ALL, and of chromosome 21 in 100% of cases. In contrast to many aneuploid malignancies, there has been little evidence of chromosomal instability (CIN) in HeH ALL and the mechanisms leading to these chromosomal gains remain unknown. The aim of this project was to determine the level of genomic heterogeneity in HeH ALL. In order to do this, we performed low-pass whole genome sequencing (WGS) of single cells isolated from diagnostic bone marrow samples from HeH ALL patients to investigate cell-to-cell heterogeneity. Single nuclei in G 1 phase from nine diagnostic childhood HeH ALL samples were isolated using fluorescence-activated cell sorting and DNA libraries were constructed for low-pass WGS. Copy number analysis for each individual cell was performed in-house using the software programs AneuFinder and Ginkgo. Homolog inheritance of chromosomes gained or lost was determined by screening for heterozygous variants and calculation of the variant allele frequencies (VAF). Sequencing 2,572 single cells showed that the nine HeH ALL patients were all relatively homogenous at the trunk of their evolution tree and that the bulk of chromosomal gains were stable and unchanging. Structural aberrations, visible as partial chromosome copy number changes, were detected at diagnosis in most cases, with a greater number of structural aberrations detected in patients with greater numbers of sub-clones (defined as three or more cells presenting with the same numerical and structural aberrations). Unique numerical chromosomal aberrations detected in two or fewer cells were relatively random and therefore indicative of nondisjunction events in recent cell divisions rather than being part of the clonal evolution of the leukemia. These results indicate very low heterogeneity in HeH ALL and suggests that the genome of these leukemias is relatively stable.

Disclosures

No relevant conflicts of interest to declare.
Highly Sensitive and Rapid Diagnosis of Mutations in Hematological Malignancies By CRISPR Technology
Authors: Liu Yin , Chen Yanling , Dang Lu , Huang Xingxu , Wang Xinjie , Zhou Fuling
Published: November 2021

With the rapid development of genomics and molecular diagnosis technologies, the treatment of hematological malignancies has entered the era of precision medicine, where individualized treatment is provided based on the unique genetic profile of each patient. Commonly-used techniques for gene mutation detection include polymerase chain reaction (PCR), sequencing approaches and real-time fluorescence quantitative PCR. However, these methods have drawbacks in sensitivity, specificity, or speed. Here, we established a mutation detection method based on CRISPR technology, which can achieve a highly sensitive, specific, and rapid diagnosis. Through a rapid release of genomic DNA, isothermal amplification, mutation enrichment, and Cas12a fluorescence reaction, this method can complete the whole diagnosis process from drawing blood to obtaining the results within one hour, without any large instruments or expensive reagents. Taking the detection of FLT3 gene D835Y/V/H/F drug-resistant mutations as an example, the sensitivity can reach 0.001%, which means 10 copies of mutation template can be detected from 999,990 copies of wild-type template. Then we used this method to detect 112 patients with acute myeloid leukemia, resulting in 11 FLT3-D835Y/V/H/F positive cases, which were verified by next-generation sequencing while missed by Sanger sequencing. We also applied this method to detect other common mutations in hematological malignancies, including NRAS gene G12D, IDH2 gene R172K, and JAK2 gene V617F. These results fully proved the high sensitivity, specificity, efficiency and universality of this method. Thus it has the potential to become a novel molecular diagnostic technology, and help point-of-care, early screening, prognosis determination and medication guidance in hematological malignancies.

Disclosures

No relevant conflicts of interest to declare.
Modeling Future Blood Requirements in Patients with Transfusion-Dependent β-Thalassemia in Greece
Authors: Souliotis Kyriakos , Golna Christina , Gountas Ilias , Tsironi Maria
Published: November 2021

Introduction : In Greece, official data confirm that blood supply is consistently declining, primarily because of decreased voluntary blood donations. At this rate, it is unclear whether there will be sufficient red blood cell (RBC) units to meet future demand for patients with β-thalassemia. We modeled future blood demand in patients with transfusion-dependent β-thalassemia (TDT) over the next 15 years to assess whether such a demand can be met.

Methods: A discrete time, stochastic, individual-based model, was developed to assess future blood needs of patients with TDT in Greece. The model adopted a cohort approach and patients enter the model shortly after birth and exit at death. The model was seeded with data from the National Registry for Hemoglobinopathies in Greece (NRHG; patient population) and data from the National Blood Centre (RBC units) (Table). To quantify the decrease in TDT incidence, a linear regression model was fitted to the incident cases as described by Voskaridou (Voskaridou E, et al. Ann Hematol 2019;98:55-66). The calculated annual reduction rate was then applied to the observed thalassemia major (TM) and thalassemia intermedia (TI) incident cases for 2012 (Voskaridou E, et al. Ann Hematol 2012;91:1451-1458). The model assumes the reduction would continue through to 2020, from which time the incidence curve would be constant.

Results: The model estimates a decrease in both target population, primarily due to the implementation of the national prenatal screening program and TM related mortality, (9.2%, 18.6%, and 26.0% in 2025, 2030, and 2034, respectively) and in demand for RBC units (100,665, 89,428 and 80,565, respectively) over the next 15 years. After accounting for any blood donated, based on latest (2017) data as projected to 2020, Greece is expected to be short of 56,716 RBC units to meet needs of patients with TDT in 2020. By 2025, 2030, and 2034, cumulative required RBC units for patients with TDT are estimated to be 307,150, 501,698, and 617,074, respectively. Assuming RBC units are divided among patients ≥ 18 years and those < 18 years in line with the population mix in the NRHG, cumulative RBC units required for adult patients with TDT are estimated to be 292,837, 464,337, and 554,083 by 2025, 2030, and 2034, respectively.

Conclusions: Though blood demand for patients with TDT is expected to decrease in line with a decrease in the patient population, it will remain a challenge for the healthcare system in Greece to meet the demand, primarily due to a continuing shortage of available RBC units from voluntary donations. This annual deficit is estimated to accumulate to over 600,000 units over the next 15 years.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Incidence of Autoimmune Diseases, Molecular Characteristics and Clinical Outcomes of Chronic Myelomonocytic Leukemia in an Asian Population: A Single-Center Experience
Authors: Ponvilawan Ben , Tunsing Pattaraporn , Sukpanichnant Sanya , Owattanapanich Weerapat
Published: November 2021

Background: Chronic myelomonocytic leukemia (CMML) is the most common subtype of the rare myelodysplastic/myeloproliferative neoplasms. Some studies have described the possible association with autoimmune disease. With the potential transformation into acute myeloid leukemia, it is known to have an unfavorable prognosis. Previous studies reported TET2, SRSF2, ASXL1, and KRAS/NRAS as the most occurring somatic mutations in CMML. However, the clinical and molecular characteristics of CMML patients in Asia remained fairly unknown.

Materials and Methods: Patients with CMML who were diagnosed in Siriraj Hospital, which is the largest comprehensive cancer center in Thailand, from 2010 to 2020 were identified from the hospital database. Clinical and laboratory data, along with pathological results, were collected from the same database. The results of the 25-gene targeted DNA sequencing panel designed for myeloid neoplasms were also extracted if the results were available. Continuous variables were represented as medians and their associated ranges. Overall survival (OS) was determined using Kaplan-Meier estimator and log-rank test.

Results: A total of 67 patients with CMML were included in our study. The median age of the cohort was 67 years old with a similar number of male and female patients in the cohort. Thirteen (19.4%) patients were diagnosed with autoimmune diseases, mainly autoimmune hemolytic anemia and immune thrombocytopenic purpura. According to cytogenetic risk for CMML, 35 (64.8%), 6 (11.1%), and 13 (24.1%) patients were classified as low, intermediate, and high risk, respectively. The clinical characteristics and laboratory results of patients were shown in Table 1. For the 15 patients with available molecular results, 13 had at least 1 somatic abnormality with TET2 (46.7%), ASXL1, NRAS, SRSF2 (each 26.7%), and KRAS (20.0%) being the common genetic mutations. The median mutation number was 4 (range, 0-6). The molecular landscape was demonstrated in Figure 1. Overall survival (OS) rates at 1 and 2 years were 74% and 57%, respectively. Patients with MDS-CMML tended to have longer median OS compared to MPN-CMML [37.9 (13.6-62.2) months vs. 19.7 (10.1-29.4) months] but did not reach statistical significance (p=0.124) (Figure 2).

Conclusions: Asian patients with CMML had a considerable amount of associated autoimmune diseases. The genetic mutations contributing to CMML in our cohort also differed from previous reports.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Palbociclib and Ponatinib Suppress Acute Myeloid Leukemia in Patient-Derived Xenograft
Authors: Busa Daniel , Loja Tomas , Jeziskova Ivana , Folta Adam , Mayer Jiri , Culen Martin
Published: November 2021

Aims: Palbociclib, a breast cancer approved CDK4/6 inhibitor, and ponatinib, a BCR/ABL1 inhibitor with a multi-kinase activity approved for chronic myeloid and acute lymphoid leukemia, were previously shown to be effective in vitro against acute myeloid leukemia (AML). Here, we aimed to test this effect in a patient-derived xenograft model.

Methods: Two newly diagnosed AMLs (AML #1: myelomonocytic AML, intermediate cytogenetic risk; AML #2: AML with myelodysplasia-related changes, poor cytogenetic risk) were xenotransplanted into NOD SCID gamma mice. Treatment was initiated at detection of approximately 5-20% hCD45+ cells in mouse peripheral blood (PB). Palbociclib, ponatinib, vehicle, and venetoclax as a comparative treatment, were administered orally for 3 weeks, 5 days per week. Chemotherapy (cytarabine+doxorubicine, AraC/Dox) 5+3 regimen served as a positive control. Azacitidine served as another comparative drug and was administered subcutaneously, five days per week in 3 cycles - 1 week on, 1 week off.

Results: Significant reduction of disease burden and prolongation of overall survival (OS) were seen with palbociclib and the reference treatment venetoclax in both AMLs, and with chemotherapy in AML#1 (Fig. 1). Ponatinib prolonged OS in AML#1 but failed to provide reduction of disease burden in PB. Interestingly, azacitidine induced the longest remission (<1% hCD45+ cells in PB), for almost 10 weeks, but only in 2/4 mice for AML#2. Treatment toxicity manifested by weight decrease was only seen with chemotherapy (AML#1: 24% weight loss, p = 0.0001; AML#2: 19% loss, p = 0.04), and was also accompanied with early mouse mortality in 3/4 mice for AML#2. None of the tested treatments lead to complete AML eradication and a gradual relapse was seen in PB. The AML infiltration was higher in bone marrow than in spleen at final analysis at relapse (AML#1, 83 vs 73% of hCD45+ cells, mean, p = 0.02; AML#2, 95 vs 65%, p = 0.0001). No changes in AML phenotype were observed between treated and vehicle mice in case of AML#1, except for azacitidine which decreased monocyte (SSCdim, CD45high; 94 vs 96%, mean, p = 0.03), CD34+ (2 vs 5%, p = 0.01) and primitive CD34+CD38- cell (0.1 vs 0.3%, p = 0.005) percentages. For AML#2, blast (SSClow, CD45dim) percentage compared to vehicle (70%) was decreased by chemotherapy (24%, p = 0.01) and increased by venetoclax (80%, p = 0.03) and azacitidine (87%, p = 0.03); monocytes (6% for vehicle) were decreased by chemotherapy (1%, p = 0.004), venetoclax (3%, p = 0.01), and azacitidine (0%, p = 0.008); CD34+ cells were increased by venetoclax (59 vs 39%, p = 0.02).

Summary: Palbociclib, and partially ponatinib, demonstrated AML suppression in vivo. This encourages further investigation of their efficacy in different AML subtypes and in combination with other drugs.

Funding: Supported by MUNI/A/1595/2020.

Figure 1 Figure 1.

Disclosures

Mayer: AOP Orphan Pharmaceuticals: Research Funding.
Haploidentical Peripheral Blood Stem Cell Transplantation (Haplo-PBSCT) with Cyclophosphamide Pos Transplant (CyPost) in High-Risk Hematologic Malignancies. Results of 5 Years in Transplant Center in Northeast of Mexico
Authors: Baltazar Severiano , Pequeño Luevano Myrna Patricia , González Villarreal Guadalupe , Sotomayor Duque Guillermo , Ramos Vazquez Raul , Cruz Contreras Diego , Guajardo Leal Martha Lilia , Sorkee Davila Helga Patricia , Carrizalez Villarreal Jose Alfredo , Cedillo Jose Luis , Ramirez López Alba Nydia , Borjon Cabada Isabel , Quirino Marquez Christian , Mecott Estudillo Arantxa , Avila Contreras Gerardo , Nava Gutiérrez Willy , Aldama Gutiérrez Dora , Garza Escobar Juan Daniel , Garcia Cerda Yarezi , Valerio Bugarin Victor , Machuca Adame Karen , Gudiño Cobos Luis Omar , De Leon Cantu Rosa Elva , Hernadez Valdez Roberto
Published: November 2021

Introduction: Haploidentical peripheral blood stem cell transplantation ( Haplo-PBSCT) is a curative option in patient with hematology malignancy. In Mexico is very difficult to find an identical HLA donor , for this reason Haplo PBSCT it is an option in our public Health institution.

Materials and methods: we aimed a retrospective study since (2017 -2021) in our Hospital Unidad Médica de alta Especialidad 25, Instituto Mexicano del Seguro Social (IMSS UMAE 25) Monterrey, Mexico. We include patients over 15 years of age who were undergo Haplo PBSCT with hematolgy malignancies . We performed Analysis of Overall and Relapse-free survival with kaplan meier curves and incidence of acute and chronic graft versus host disease (GVHD) and percentage of Transplant-Related Mortality (TRM).

Results: we analized a 38 patients, the median age was 35 years (range 15-64). Of the total , 25 were male (65%) and 13 famale (34%). The rest of baseline data is described in table 1.

The median time of neutrophil engrafment was days 17 ( 11-26 ) and platelets was 19.5 days ( 12-38 ). The Incidence of Primary Graft failure was 18 % ( seven patients). Patients with graft had 99-100% chimerism at day 100. The Incidence of aGVHD was 26 % and the cGVHD was 16%. DLI was used in four patients to treat passenger lymphocyte syndrome (2 patients) and ( 2 patients) with severe graft rejection.

The principal cause of death was sepsis in eleven patients (29%). Two patients with acute leukemia (AML and ALL) died after development of post transplant lymphoproliferative disease. The TRM was 28.2%. There were no relapse-related deaths during follow-up after Haplo-PBSCT.

The median of survival was 4.5 months ( range 1-44) with Overall Survival (OS) of 42 % at 3 years ( figure 1).

Conclusion: Our analysis shows results that were comparable with those published in first world international transplant centers . Relapse remains the major cause of transplant failure, in our population the antigenic disparity between donor and recipient can strengthen the immune response versus disease. Haplo-PBSCT is a feasible transplantation method in patients with hematological malignancies in developing countries with limited resources.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Racial Disparity in Mortality Rates Among Black and White Americans with Multiple Myeloma
Authors: Akinyemi Oluwasegun Austine , Ogunbona Oluwaseun B , Abodunrin Faith O , Oni Olanrewaju
Published: November 2021

Background: In the United States, Multiple myeloma (MM) is the second most common hematologic malignancy and the most common hematologic malignancy in blacks. It is a fatal plasma cell dyscrasia with a median overall survival of 5 to 10 years. Different studies have highlighted a racial disparity in survival outcomes between Blacks and Whites patients' population with MM. However, the reason for the observed racial disparity in survival outcomes is not clearly understood.

Aim: To explore the role of socioeconomic variables including access to insurance and, median Income on the racial disparity in outcome among patients with MM.

Method: We conducted a retrospective analysis of all hospitalizations associated with the primary diagnosis of MM in the National Inpatient Sample (NIS 2008-2014) database using the relevant International Classification of Diseases, Ninth Revision, Clinical Modification codes. The study outcome was the mortality rates between black patients and a propensity score matched control group of white patients with MM.

Result: There were 185,000 MM associated hospitalization in the NIS (2008-2014). Among the study population, 66.7% were whites, 20.8% were blacks, and 7.7% were Hispanics. The mean age was 66.5 years for blacks and 70.2years for whites. Among the Black population, 21.6% had private insurance compared to 25.1% of their White counterparts. Also, 45.6% of Black participants were in the lowest income quartile compared to only 19.6% of Whites. Only 13.5% of Blacks were in the highest income quartile compared to 28.7% of Whites. 73.4% of Blacks were on public insurance compared to 70.9% of Whites. There was no difference in the mortality rate between blacks and a control group of whites matched for insurance, median income, sex and age (49.4% vs 50.6%, p =0.42).

Conclusion: Multiple myeloma may not be more deadly in Blacks compared to Whites, and the observed disparity in outcomes in the literature are likely due to the effects of social determinants of health such as income and access to insurance.

Disclosures

No relevant conflicts of interest to declare.
New Approach to Decipher GATA2 Deficiency Syndrome: Focus on the Recurrent GATA2 R396Q Mutation
Authors: Largeaud Laetitia , Jamrog Laura , Fregona Vincent , Hamelle Camille , Dufrechou Stephanie , Prade Naïs , Hebrard Sylvie , Sellam Esmaa , Bayet Manon , Didier Christine , Gerby Bastien , Delabesse Eric , Pasquet Marlene , Broccardo Cyril
Published: November 2021

Germline GATA2 mutations are identified in a complex disorder termed GATA2 deficiency syndrome. Clinical heterogeneous manifestations are associated with a wide diversity of molecular alterations (missense, frameshift, nonsense, intronic or splicing mutations). Truncating mutations decrease GATA2 expression suggesting a haploinsufficiency mechanism while molecular consequences of missense mutations are ill-known. We focus our research on the most recurrent GATA2 mutation, GATA2 R396Q.

Our in vitro results indicate that the ectopic expression of GATA2 R396Q has a strong impact on the clonogenicity associated with an excessive granulocyte differentiation. This data motivates the elaboration of a more physiological approach through the development of a new knock-in mouse model with endogenous expression of Gata2 R396Q mutation. This model allowed us to address the question of the impact of this missense mutation on hematopoiesis at steady state and in challenging conditions.

The Gata2R396Q/+ mice showed an abnormal distribution of the immature progenitors Lin - Sca-1 + Kit + (LSK) subpopulations, mainly defines by an increase of the number of aberrant Long-Term Hematopoietic Stem cells (LT-HSC) contrasting with the decrease of LT-HSC population in Gata2+/- mouse model. Functional assays on Gata2R396Q/+ LSK cells revealed also qualitative defects. Indeed, their reconstitution capacity and their response to inflammatory or chemotherapy stimuli seemed to be largely affected. To address at the molecular level the impact of the missense mutation in these progenitors, we combined gene expression analysis with chromatin gene accessibility. These analyses revealed a strong disruption of the cells' ability to respond to stimuli, such as IFN or TNFα signaling, confirming what we observed at the cellular level in functional assays. Furthermore, specific RNA sequencing of the LT-HSC, ST-HSC and MPP3-4 subpopulations showed that the majority of these molecular perturbations are detected in every subpopulation of the LSK cells. In addition, we are able to establish a precise genetic signature in LT-HSC that may be related to their major functional defects.

Combination of in vitro and in vivo approaches leads to improve our understanding of GATA2 deficiency syndrome. Modelization of the most recurrent germline Gata2 mutation revealed a different phenotype than the haploinsufficient mouse model. Furthermore, expression of Gata2 R396Q mutation qualitatively impacts the LSK compartment mimicking functional and molecular defects observed in the GATA2 deficiency patients.

Disclosures

Delabesse: Novartis: Consultancy; Astellas: Consultancy. Pasquet: Novartis: Consultancy.
ASXL1 Mutation Cooperating with RUNX1 Mutation Further Perturbs the Differentiation of Neutrophils
Authors: He Jinping , Liu Qifa
Published: November 2021

Acute myeloid leukemia(AML) is characterized by an accumulation of myeloid progenitors. This disease can be caused by various germline mutations, including runx1 and asxl1. Recently several reports showed that mutations of ASXL1 and RUNX1 tend to coexist in AML patients. However, the impact on hematopoiesis remained uncertain. Thus far, lack of an animal model was the main limitation to study the function of the double mutations. In this work, we studied the combination role of runx1 and asxl1 by establishing a novel in vivo zebrafish model. First, we identified the characterization of 6646 patients with myeloid maliganacies in the cbioprotal database. Second, we generated a zebrafish model with double mutations by crossing the ENU-induced runx1 mutant and the CRISPR-CAS9 edited asxl1 mutant. Our results showed that truncation was the most frequent mutation type in RUNX1-mutated patients(56.3%) and ASXL1-mutated patients(87.8%). Besides, the median survival time of patients carrying double mutations was shorter than those with single RUNX1 mutation(9.61 months vs. 16.06 months ) or single ASXL1 mutation(9.61 months vs. 14.47 months). In zebrafish models, we found that the pu.1(myeloid precursor marker) positive cells increased while the gata1(proerythrocyte marker) didn't change in double mutants . Furthermore, mpo and lyz (neutrophil marker) positive cells further reduced while mpeg1 and mfap4(monocyte marker) positive cells didn't show significant difference in double mutants.Overall, we demonstrated that ASXL1 mutation cooperating with RUNX1 mutation further impeded the differentiation of the myeloid progenitors into mature neutrophils and it possibly explained the cause for a worse prognosis of patients with double mutations.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Prophylactic Administration of Antibodies Specific for the HPA-1a Epitope Prevents Alloimmunization and Platelet Destruction in a Murine Model of Fetal Neonatal Alloimmune Thrombocytopenia
Authors: Zhi Huiying , Sheridan Douglas , Newman Peter J.
Published: November 2021

Fetal/neonatal alloimmune thrombocytopenia (FNAIT) is a life-threatening bleeding disorder caused by maternal alloantibodies directed against paternally inherited antigens present on the surface of fetal and neonatal platelets. The human platelet alloantigen HPA-1a (formerly known as the Pl A1 alloantigen), is the most frequently implicated HPA for causing FNAIT in Caucasians. A single Leu33Pro amino acid polymorphism near the amino terminus of the integrin b3 subunit (known as GPIIIa in the platelet literature) serves as the central target for alloantibody binding, leading to clearance of both fetal and neonatal platelets, thrombocytopenia, and in the most severe cases, spontaneous- or trauma-induced intracranial hemorrhage. Unlike hemolytic disease of the newborn, which occurs in pregnancies subsequent to parturition-induced alloimmunization, an estimated 25% to 50% of FNAIT cases occur without warning during gestation of the first pregnancy. Though long proposed, there are currently no approved therapies for the prevention of FNAIT. We recently described the development of transgenic mice expressing the human HPA-1a allogeneic epitope on a murine GPIIIa backbone. Transfusion of such platelets into wild-type female mice induced the generation of high-titer anti-HPA-1a alloantibodies that can cross the placenta and recapitulate many of the relevant clinical features of FNAIT. To test the hypothesis that rapid elimination of fetal HPA-1a positive platelets from the circulation of a mother who is HPA-1a negative might prevent maternal alloimmunization and the development of FNAIT, we administered either a hyperimmune plasma-derived polyclonal anti-HPA-1a antibody derived from females having previous cases of FNAIT (termed RLYB211), or a novel human monoclonal antibody directed against the HPA-1a epitope (termed RLYB212), to wild-type female mice prior to challenging them with HPA-1a-positive murine platelets. RLYB211 and RLYB212 were each able to effect the rapid removal of HPA-1a-positive platelets from murine circulation and prevent the development of anti-HPA-1a alloantibodies. Importantly, wild-type female mice pretreated with RLYB211 prior to exposure to HPA-1a-positive platelets, and then impregnated by HPA-1a-positive males, gave birth to HPA-1a-positive pups with significantly improved platelet counts and no bleeding symptoms. These preclinical data establish the potential for prophylactic polyclonal and monoclonal anti-HPA-1a antibody therapy for the prevention of FNAIT in humans.

Disclosures

Sheridan: Rallybio: Current Employment, Current equity holder in publicly-traded company. Newman: Rallybio: Consultancy, Research Funding.
Notch Ligand Jagged1 Is a Fetal Liver Niche Factor for the Function of Embryonic Hematopoietic Stem Cells
Authors: Pajcini Kostandin V. , Shao Lijian , Paik Na Yoon , Sottoriva Kilian
Published: November 2021

Embryonic hematopoietic stem cells (HSC) expand rapidly during development in the fetal liver. Notch1 is required for emergence of the definitive hematopoietic stem cells (HSCs) from the hemogenic endothelium, and is essential for survival and function of HSCs in the fetal liver. The identity of the ligand and the ligand-presenting cell during hematopoietic development would provide valuable information of the Notch signaling mechanism in HSCs as well as the identity of key niche cells that drive the expansion and cell fate decisions of embryonic HSCs. In the present study, we have taken a comprehensive approach to determine the ligands and cells that initiate Notch signaling in the mouse fetal liver. To this end, we have performed single-cell analysis for all Notch signaling proteins and many known targets in E14.5 fetal HSCs and adult bone marrow HSCs as well as fetal liver endothelial cells. We determined that Jagged1 (Jag1) is highly expressed in both endothelial cells as well as in fetal HSCs but not in adult HSCs. We have performed conditional loss-of-function analysis of Jag1 in fetal endothelial cells as well as in fetal hematopoietic lineages, where both myeloid and megakaryocytic progenitors are shown to express high levels of Jag1. Our results indicate that while loss of endothelial Jag1 has severe effects in embryonic vascular development, loss of hematopoietic Jag1 allows for normal fetal morphology, yet severely impedes the functional ability of fetal liver HSCs to expand and differentiate. RNA-Sequencing analysis of long-term fetal HSCs in Jag1-mutant embryos (VavCreJag f/f) revealed reduced expression of Gata2, Mllt3, Hoxa7, Angpt1 and IL-12a genes in fetal HSCs, which are well-known regulators of self-renewal and expansion. Our findings indicate that Jag1 is an essential niche factor for development of HSCs in the fetal liver and for functional potential of fetal HSCs once in the bone marrow microenvironment.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Herpesvirus Infections of Intestinal Tract in the Patients with Intestinal Graft-Versus-Host Diseases: Laboratory Diagnosis Based on DNA Detection By Real-Time Quantitative PCR in Feces Samples
Authors: Lin Ren , Liu Wenjun , Huang Fen , Luo Tingyue , Mo Wenjian , Gu Yingying , Xu Na , Chang Yuan , Fan Zhiping , Liang Xinquan , Wu Yuanbin , Wang Shunqing , Du Jing , Hui Liu , Wang Caixia , Sun Jing , Liu Qifa
Published: November 2021

Background: Intestinal herpesvirus disease remains one of the major causes of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (allo-HSCT). There is lack of useful methods for etiological diagnosis of intestinal herpesvirus diseases. Here, we evaluated the efficiency of detecting herpesvirus in feces samples via real-time quantitative PCR (RQ-PCR) for diagnosis of intestinal herpesvirus diseases after allo-HSCT.

Methods: This was a multicenter, prospective study. Patients with refractory diarrhea after intestinal graft-versus-host diseases (GVHD) were enrolled in this study. Laboratory tests which consisted of morphologic examination, immunohistochemistry, in situ hybridization, and RQ-PCR of tissue homogenate were used to detect viral pathogens including cytomegalovirus (CMV), epstein-Barr virus (EBV), herpes simplex virus (HSV)-I, HSV-II, varicella zoster virus (VZV), adenovirus (ADV) and human herpes virus (HHV)-6, HHV-7. These viruses aforementioned were also detected in feces and blood samples.

Results: One hundred and seven patients with refractory diarrhea after intestinal GVHD were enrolled between January 2016 and December 2020. Based on the detection of viruses in biopsy specimens, 75 patients were diagnosed as intestinal infectious diseases including 64 accompanying with intestinal GVHD. CMV was the most frequent pathogen of intestinal infectious diseases (53.8%), followed by EBV (36.5%), bacteria (3.4%) and others (6.3%). For diagnosis of intestinal CMV diseases, the sensitivity and specificity of RQ-PCR in feces samples were better than those of blood (sensitivity: 96.9% v.s. 72.5%, p=0.004; specificity: 93.6% v.s. 75.8%, p=0.035). Similarly, the sensitivity of RQ-PCR in feces and blood samples were 88.2% and 21.9% (p<0.001) and the specificity were 98.5% and 86.3% (p=0.032) for diagnosis of intestinal EBV diseases.

Conclusion: Intestinal infectious diseases were one of the main causes of refractory diarrhea after intestinal GVHD. Herpesviruses, especially CMV and EBV, were the most common pathogens. Herpesvirus-DNA detection by RQ-PCR in feces samples was a useful diagnostic method for intestinal herpesviruses diseases.

Disclosures

No relevant conflicts of interest to declare.
Venetoclax 400mg Combined with Bortezomib and Dexamethasone Is Efficacious and Feasible for Plasma Cell Dyscrasia
Authors: Lu Jin
Published: November 2021

【Abstract】 Objective: To investigate the efficacy and safety of BCL-2 inhibitor Venetoclax (Ven) 400mg combined with bortezomib and dexamethasone in plasma cell dyscrasia with t(11,14). Methods: The data of twenty patients with plasma cell dyscrasia with t (11;14) were retrospectively analyzed.The majority of patients received Venetoclax 400mg combined with bortezomib and dexamethasone. The baseline clinical information, treatment efficacy and adverse effects after Ven-based regimen. Results: Seventeen multiple myeloma patients, two light-chain amyloidosis and one primary plasma cell leukemia (pPCL) patients were studied. Among seventeen MM patients, the median follow-up was 9.0 (0.5-27) months and the median treatment cycles was 4 (1-12);fourteen patients have been evaluated for response rate. The overall response rate for evaluable patients (ORR) was 85.7%, among that one patient (7.1%) achieved CR, four patients (28.6%) achieved VGPR and 7 (50%) achieved PR. The median progression-free survival (PFS) was11.0 months (95%CI 2.1-19.9); the median Overall survival (OS) was 14.0 months (95%CI 6.9-21.1). Two amyloidosis patients achieved VGPR and PR respectively, while one pPCL patient achieved CR. The most common grade 3 or worse treatment-emergent adverse events were thrombocytopenia (15%), infection (10%), neutropenia (5%), diarrhea (5%) and peripheral neuropathy (5%). There was no treatment-related death. Conclusion Ven (400mg) combined with bortezomib and dexamethasone is feasible, tolerated and efficacious in plasma cell dyscrasia.

【Key words】Plasma cell dyscrasia; Venetoclax; Efficacy; Safety

Disclosures

No relevant conflicts of interest to declare.
Post-Transplant Blockade of CXCR4 Improves Complete Remission Rates and Donor Stem Cell Engraftment without Aggravating Gvhd
Authors: Su Long , Fang Ming Hui , Zou Jun , Gao Su Jun , Gu Xiao Yi , Meng Xian Di , Wang Xue , Hu Zheng , Yang Yong Guang
Published: November 2021

Allogeneic hematopoietic cell transplantation (allo-HCT) is a promising therapeutic option for hematological malignancies, but relapse resulted predominantly from residual disease in bone marrow (BM) remains the major cause of treatment failure. Using immunodeficient mice grafted with laboratory-generated human B-ALL, our previous study suggested that leukemia cells within the BM were resistant to graft-versus-leukemia (GVL) effects, and mobilization with CXCR4 antagonist may dislodge leukemia cells from BM, rendering them destroyed by GVL effects. In this study, we extended this approach to patient-derived xenograft (PDX) and murine T-ALL and AML models for determining its clinical relevance and effects on GVHD and donor hematopoietic engraftment. We found that post-transplant treatment with CXCR4 antagonist AMD3100 significantly improved eradication of leukemia cells in the BM in PDX mice grafted with B-ALL cells from multiple patients. AMD3100 also significantly improved disease-free remission rates in murine T-ALL and AML models, and promoted donor hematopoietic engraftment in mice following non-myeloablative allo-HCT. Furthermore, post-transplant treatment with AMD3100 had no detectable deleterious effect on acute or chronic GVHD. These findings provide important preclinical data supporting the initiation of clinical trials exploring combination therapy with CXCR4 antagonist and allo-HCT.

Disclosures

No relevant conflicts of interest to declare.
Prophylactic Treatment of Severe Haemophilia Á Patients with Inhibitors to FVIII with Peglip-FVIII
Authors: Timofeeva Margarita , Zozulya Nadezhda I. , Pospelova Tatiana , Kosinova Marina V , Kurtov Igor , Tuddenham Edward , Ling Gavin
Published: November 2021

Background: FVIII replacement therapy is ineffective for severe haemophilia A (HA) patients who develop inhibitors to FVIII. Patients with intractable inhibitors currently require FVIII mimetics and/or bypassing agents to prevent bleeding. PEGylated liposomes (PEGLip) have been shown to protect FVIII from anti-FVIII antibodies in ex-vivo human studies and in combination with FVIII may present an option for the prophylactic treatment of inhibitor patients.

Aims: To (a) demonstrate that PEGLip-FVIII administered intravenously (IV) to severe HA patients with history of inhibitors to FVIII enhances their clotting activity, (b) compare the number of bleeding episodes before and after PEG-Lip treatment, and (c) demonstrate that PEGLip-FVIII is well tolerated with no increase in inhibitor titres.

Methods: Stage A: Four patients with a history of inhibitors were given single IV injections of PEGLip-FVIII (simoctocog alfa) at a dose of 22mg/kg PEGLip + 35 IU/kg FVIII and assessed for clotting activity at 0 hours (pre-injection) and at 20min, 1, 2, 4, 8, 24 hours, and daily thereafter up to 7 days using Rotational Thromboelastometry. Stage B: Patients received IV injections of PEGLip-FVIII for 6-weeks at a frequency determined by the investigator based on results obtained during Stage A. Inhibitor titres were monitored throughout.

Results: Results are shown below.

Treatment with PEGLip-FVIII was highly tolerated with no clinically significant changes in inhibitor titres. No Adverse Drug Reactions were reported. The mean frequency of administration of PEGLip-FVIII was every 5.7±1.4 days. The mean number of bleeding episodes reported during Stage B was 0.5±0.9 per month (due to 1 patient) compared with 0.9±0.4 per month recorded during the 24 weeks prior to enrollment.

Conclusion: PEGLip-FVIII in inhibitor patients demonstrated efficacy in preventing spontaneous bleeds without increasing inhibitor titres, indicating a novel FVIII-based treatment for this cohort. Planned studies in a larger cohort may confirm our findings.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Liposome-Encapsulated Cytarabine and Daunorubicin (CPX-351) Induces Remission in Newly Diagnosed Pediatric Secondary Myeloid Malignancies
Authors: Caldwell Kenneth J , Hu Yixin , Onciu Mihaela , Federico Sara M , Salek Marta , Lewis Sara , Lei Shaohua , Zhang Jinghui , Nichols Kim E. , Takemoto Clifford M , Triplett Brandon M , Farrar Jason E , Rubnitz Jeffrey E. , Ribeiro Raul C , Wlodarski Marcin W.
Published: November 2021

Secondary myelodysplastic syndromes and acute myeloid leukemia (sMDS/AML) are rare myeloid neoplasms in children/adolescents and have a dismal prognosis. The mainstay therapy is hematopoietic cell transplantation (HCT) but there has been little therapeutic innovation for decades and outcomes remain poor. CPX-351, a fixed 5:1 molar ratio of liposomal cytarabine/daunorubicin, has shown favorable safety and efficacy in elderly individuals with sAML and children with relapsed de novo AML, which led the FDA to recently expand the label of CPX-351 to include pediatric patients with secondary AML, however, no data has been reported in this patient group. We report the outcomes of seven young patients with newly diagnosed sMDS/AML uniformly treated with CPX-351. Five patients had previously received chemotherapy for osteosarcoma, Ewing sarcoma, neuroblastoma, or T-ALL; one had predisposing genomic instability disorder (Cornelia de Lange syndrome); and one presented with MDS-related AML and multi-organ failure. The median age at diagnosis of myeloid malignancy was 17 (13-23) years. We identified somatic mutations and copy-number changes across 16 leukemia driver genes in six cases (including TP53 in two), abnormal karyotypes in six cases and rearrangements involving MECOM or NIM1K-TERT in two patients. Additional genomic studies identified pathogenic germline mutations in CHEK2 and SMC3 each in a single patient . Patients received 1-3 cycles of CPX-351 (100 units/m 2 on days 1, 3, and 5) resulting in complete morphologic remission without overt toxicity or treatment-related mortality. This approach allowed for adding FLT3 inhibitor as individualized therapy in one patient. Six patients were alive and leukemia-free at 0.51-3.25 years after HCT. One patient died from disease progression before HCT. Concluding, CPX-351 is an effective and well-tolerated regimen for cytoreduction in pediatric secondary myeloid malignancies warranting further investigation

Figure 1 Figure 1.

Disclosures

Triplett: Miltenyi: Other: Travel, meeting registration.
A Clinical Study of 15 Acute Leukemia Patients with Plasmacytoid Dendritic Cells Expansion
Authors: Zhou Ya-Lan , Wei Yan , Liu Xiaohong , Wang Jing , Jia Jinsong , Shi Hong-Xia , Chang Ying-Jun , Zhao Xiao-Su , Liu Yan-Rong , Ruan Guo-Rui , Huang Xiao-Jun , Jiang Hao
Published: November 2021

Objective: To improve the understanding of acute leukemia with plasmacytoid dendritic cells (pDCs) expansion. Method: Bone marrow smears and flow cytometry immunophenotyping results of patients with acute leukemia and pDCs expansion at diagnosis in our clinical center from August 2018 to January 2021 were retrospectively analyzed. The clinical and biological characteristics were analyzed. Results: Among the 1039 patients with acute leukemia, 15 (1.4%) had pDCs expansion at diagnosis, with a median age of 55 (28-99) years old. The ratio of men to women was 3 to 2. The median proportion of pDCs accounting for all the nucleated cells in bone marrow smear and flow cytometry immunophenotyping were 12.0% (6.0%-50.0%) and 16.6% (4%-28%), respectively. There were 12 cases of acute myeloid leukemia (AML), 11 of them had mutations in RUNX1 and 10 were morphologically classified as AML-M4 or M5. Notably, CD56 was mostly negative (only 1 of the 15 patients expressed CD56 antigen partially) and one of these 15 patients had definite skin lesions, which were significantly different from Blastic plasmacytoid dendritic cell neoplasms. 3 cases were diagnosed as acute lymphoblastic leukemia (ALL), of which 2 cases had mutations in NOTCH1 and RAS signaling pathways. 9 out of the 15 cases had consecutive prognostic information and the median follow-up time was 14.9 (2.1-31.9) months, of which 6 received allogeneic hematopoietic stem cell transplantation(allo-HSCT) and all were in continuous remission. However, among the 3 non-transplanted patients 2 cases relapsed. Conclusion: Acute leukemia with pDCs expansion is a group of patients with characteristic mutation profiles and relatively poor prognosis, more common in AML with abnormal expansion of the mononuclear system. Allo-HSCT may improve the clinical outcomes of these patients.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
CARD19, a Novel Negative Regulator of B-Cell Tolerance
Authors: Zheng Yongwei , Yu Mei , Chen Yuhong , Xue Liquan , Zhu Wen , Fu Guoping , Burns Robert , Morris Stephan , Wen Renren , Wang Demin
Published: November 2021

The CARD11-Bcl10-Malt1 (CBM) signalosome controls TAK1 activation to regulate B-cell receptor (BCR)-induced NF-κB activation and B cell biology. The biological function of caspase recruitment domain family member 19 (CARD19), originally identified as a BCL10-interacting CARD protein (BinCARD), is not known. Here we found CARD19 strongly interacted with TAK1 but not BCL10 or other CBM components and prevented TAK1's association with TAB2, thereby inhibiting TAB2-mediated TAK1 ubiquitination and activation and subsequent NF-κB activation. CARD19 was ubiquitously expressed in hematopoietic lineages but its deficiency in mice had no effect on hematopoiesis, including B cell development and humoral immune response. CARD19 deficiency enhanced clonal deletion, receptor editing and anergy of self-reactive B cells, thus reducing autoantibody production in vivo. Mechanistically, CARD19 deficiency led to an increase of BCR/TAK1-mediated NF-κB activation. Activation of NF-κB, such as c-Rel, was responsible for the up-regulation of BCR-induced expression of the transcription factor early growth response genes 2 and 3 (Egr2, Egr3) and the E3 ubiquitin ligases, c-Cbl and Cbl-b, the important inducers of B-cell tolerance in B cells. Further, high-throughput RNA sequencing analysis revealed that CARD19 deficiency did not affect the overall antigen-induced gene expression in naïve B cells but suppressed BCR signaling to increase hyporesponsiveness of self-reactive B cells. Consequently, CARD19 deficiency prevented Bm12-induced experimental systemic lupus erythematosus (SLE) and autoimmunity in a B cell-intrinsic manner. Taken together, CARD19 negatively regulates BCR-induced NF-κB activation via blocking TAK1/TAB2 interaction and its deficiency leads to NF-κB-induced expression of Egr2/3 and c-Cbl/Cbl-b in self-reactive B cells, which enhances B-cell tolerance and thus prevents autoimmunity.

Disclosures

No relevant conflicts of interest to declare.
Clinicopathological Characteristics of Subcutaneous Panniculitis-like T-Cell Lymphoma in Asian Population: A Retrospective Cohort of 48 Patients
Authors: Ponvilawan BEN , Likasitwatanakul Pornlada , Sitthinamsuwan Panitta , Siritanaratkul Noppadol , Khuhapinant Archrob , Chairatchaneeboon Manasmon , Samutpradit Danai , Phikulsod Ployploen
Published: November 2021

Background: Subcutaneous panniculitis-like T-cell lymphoma (SPTCL) is a rare subtype of cutaneous T cell lymphoma. Despite its higher prevalence in Asia than other regions, the clinicopathological data of SPTCL in Asian population is scarce.

Materials and Methods: Patients who were diagnosed with SPTCL in Siriraj Hospital, Thailand, from 2010 to 2021 were identified. Demographic, clinical, and laboratory data were retrospectively collected from the hospital database. All statistics were performed in IBM SPSS Statistics Version 23. All continuous variables were reported as medians and their associated ranges.

Results: A total of 48 patients, comprising of 13 males and 35 females, were included in this study. The median age of the cohort was 30 years old. Seven (14.6%) patients had previous or concurrent autoimmune diseases. Seven (14.6%) patients presented with concomitant hemophagocytic lymphohistiocytosis (HLH) at diagnosis. Out of 14 patients who were tested for T-cell receptor (TCR) gene rearrangement, 9 were positive for at least one of monoclonal TCR-β, -γ, or -δ gene rearrangement. The summary of characteristics, presentations, and laboratory results of patients were shown in Table 1. 34 and 14 patients received immunosuppressant therapy (IMT) and chemotherapy (CMT) as their first line of treatment, respectively. The response rates were comparable in both groups.

Conclusions: Clinical presentations of SPTCL in patient's cohort in Asian population are similar to those reported in Western population except a higher rate of autoimmune features. IMT and CMT resulted in similar short-term outcome. Further studies on long-term outcome and the predictive markers for treatment response will be beneficial for this rare subtype of T-cell lymphoma.

Figure 1 Figure 1.

Disclosures

Khuhapinant: Astellas Pharma, Inc.: Research Funding.