bluuebunny/crossref_metadata_2025 huggingface dataset:
Pomalidomide, Bortezomib, and Dexamethasone (PVd) for Chinese Multiple Myeloma Patients at First Relapse
Authors: Qiu Lugui , An Gang
Published: November 2021

Background:

The phase 3 OPTIMISMM（NCT01734928） trial done at 133 hospitals and research centres in 21 countries but no Chinese site involved in this study. In the OPTIMISMM study ,pomalidomide, bortezomib, and dexamethasone (PVd) demonstrated excellent efﬁcacy in patients at ﬁrst relapse, including immediately after upfront lenalidomide treatment failure and other common ﬁrst-line treatments, the mPFS was 20.73 months, the ORR was 90.1%, and no new safety signals were observed. Building on these promising results, we decided to explore PVd in Chinese patients at ﬁrst relapse.

Study Design/Methods:

This is a multicenter, prospective, single-arm phase 2 study designed to evaluate the efﬁcacy and safety of PVd in Chinese patients at ﬁrst relapse.

Eligible patients were aged ≥18 years and had a diagnosis of multiple myeloma, measurable disease, and an Eastern Cooperative Oncology Group performance status of 0-2. Patients were required to have had only 1 prior antimyeloma regimen. Key exclusion criteria included creatinine clearance<30 mL/min requiring dialysis, grade ≥3 peripheral neuropathy, or grade 2 peripheral neuropathy with pain. Patients with prior exposure to bortezomib were eligible, provided they were not refractory to a bortezomib-containing regimen dosed at 1.3 mg/m 2 twice weekly.

Patient（n=62,Figure 1）will receive pomalidomide 4 mg on days 1-14 of each cycle. Bortezomib 1.3 mg/m 2 on days 1, 4, 8, and 11 of cycles 1-8 and on days 1 and 8 of cycles 9 and beyond. Dexamethasone was given on days 1, 2, 4, 5, 8, 9, 11, and 12 of cycles 1-8 and on days 1, 2, 8, and 9 of cycles 9 and beyond; patients received 20 mg of dexamethasone if aged ≤75 years and 10 mg otherwise.

The primary endpoint is ORR, the secondary endpoints are≥VGPR, MRD(-) rate, PFS, OS and safety. ORR will be assessed by the International Myeloma Working Group criteria after each cycle until PD. The Kaplan-Meier method will be used to estimate PFS and OS. Safety analysis will be conducted in the safety population, which are composed of all patients who received ≥1 dose of study medication. The trial is currently enrolling and will be open in 7 sites of China.

Disclosures: Research Sponsor: CHIATAI TIANQING PHARMACEUTICAL GROUP.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Baseline Hepcidin Does Not Predict the Response to Iron Therapy in Pregnant Women with Iron Deficiency
Authors: Hansen Rebecka , Kurtzhals Joergen , Styrishave Bjarne , Holm Charlotte
Published: November 2021

Introduction:

Hepcidin, the master regulator of iron economy, is decreased during pregnancy to facilitate adequate iron transfer across the placenta. Conversely, iron need increases substantially during pregnancy often leading to iron deficiency and subsequently anemia. The PREG-01 Study compared the efficacy and safety of intravenous (IV) ferric derisomaltose (FDI) vs. oral iron in treating persistent iron deficiency in pregnant women. The study found FDI to be efficacious and well-tolerated in pregnancy and the proportion of non-anaemic patients throughout the course of the study was significantly lower in the FDI group. In this analysis, we investigated the effect of baseline hepcidin on the response to IV and oral iron therapy.

Methods:

PREG-01 was a single-centre, open-label, randomized controlled trial. Women 14-21 weeks pregnant with persistent iron deficiency (ferritin<30 µg/L despite oral iron treatment) received a single intravenous 1000 mg dose of FDI (n=100) or 100 mg elemental oral iron daily combined with ascorbic acid (n=101). Hemoglobin (Hb), ferritin and transferrin saturation (TSAT%) levels were captured at baseline and monitored throughout the study. The effect of baseline hepcidin on achieving non-anemic status (Hb ≥ 11 g/dL) at all study visits and the effect on change in Hb, ferritin and TSAT% were investigated by estimating odds ratios from a logistic regression model with treatment as factor and interaction between treatment and baseline hepcidin. The odds ratio estimate is for an increment in baseline hepcidin of 1 ng/mL.

Results:

Mean [standard deviation (SD)] baseline Hb was 11.97 (0.93) g/dL in the FDI group and 11.75 (0.91) g/dL in the oral iron group. Baseline hepcidin was 6.42 ng/mL in the FDI and 5.32 ng/mL in the oral iron group. Baseline hepcidin was not associated with the ability to achieve non-anaemic status either in the FDI (OR 0.98; 95% CI: 0.87-1.09) or the oral iron group (OR 0.96; 95% CI: 0.88-1.05). No statistically significant associations were found between baseline hepcidin and change in Hb, ferritin or TSAT% throughout the study (Figure 1).

Conclusions:

In a population of pregnant women with iron deficiency, but otherwise healthy, baseline hepcidin was overall low. Although there was a trend for an association between baseline hepcidin and Hb response to oral iron only, baseline hepcidin did not predict the response to iron therapy.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.

OffLabel Disclosure:

Ferric Derisomaltose is an IV iron preparation indicated for the treatment of iron deficiency anemia in the US.
IGF2BP3 As an N6-Methyladenosine Reader Promotes Acute Myeloid Leukemia Progression By Regulating the Stability of Target RNA
Authors: Zhang Nan , Deng Jianchuan , Zhou Fuling
Published: November 2021

Background: N6-methyladenosine (m6A) is the most common post-transcriptional modification of eukaryotic mRNA. Recent evidence suggests that dysregulated m6A-associated proteins and m6A modifications play a pivotal role in the initiation and progression of diseases such as cancer. Here, we identified that IGF2BP3 is specifically overexpressed in acute myeloid leukemia (AML), which constitutes a subtype of this malignancy associated with poor prognosis and high genetic risk.

Methods: Bioinformatics analysis of public databases was performed to screen the differentially expressed m6A regulators in AML. Clinical samples were collected to detect the expression of IGF2BP3 in AML by RT-qPCR. The effects of IGF2BP3 on the proliferation, apoptosis and cycle of AML cells were detected by CCK-8 and flow cytometry. RNA-seq was used to identify target genes of IGF2BP3 by integrating analysis with RIP-Seq, iCLIP-Seq and MeRIP-Seq data sets.

Results:High expression of IGF2BP3 is closely associated with poor prognosis of AML and is higher in patients with high genetic risk group. IGF2BP3 was the lowest expressed in AML-M3 and the highest expressed in RUNX1 mutant type. IGF2BP3 is required for maintaining AML cell survival in an m6A-dependent manner, and knockdown of IGF2BP3 suppressed dramatically induces apoptosis, reduces proliferation and impaired leukemic capacity AML cells in vitro and in vivo. Mechanistically, IGF2BP3 interacts with RCC2 mRNA and stabilizes the expression of m6A-tagged RNA.

Conclusions:We provided compelling evidence to demonstrate that m6A reader IGF2BP3 contributed to tumorigenesis and poor prognosis of AML, which can serve as a target to develop therapeutics for cancer treatment.

Disclosures

No relevant conflicts of interest to declare.

Disclosures

No relevant conflicts of interest to declare.
GRK2 Regulates ADP Signaling in Platelets Via P2Y 1 and P2Y 12
Authors: Zhao Xuefei , Cooper Matthew , Yanman Yanki , Baltz Aiden , Michael James , McKenzie Steven Edward , Stalker Timothy J. , Zhu Li , Ma Peisong
Published: November 2021

Abstract

Venous thromboembolism (VTE), heart attack, and stroke are all diseases in which platelets play a role, through inappropriate platelet activation and subsequent thrombus formation. Most platelet agonists activate platelets via G protein-coupled receptors (GPCRs), which are targeted by many antiplatelet drugs. Along with thrombin and TxA 2, ADP has long been recognized for its important role in hemostasis and thrombosis. It activates platelets via GPCRs, P2Y 1 and P2Y 12. However, little is known about the negative feedback mechanisms governing P2Y receptor-mediated platelet activation and thrombus formation.

Here, we provide the first evidence that GPCR kinase 2 (GRK2) serves this regulatory role during platelet activation and thrombus formation by using a platelet-specific GRK2 deletion mouse model and a GRK2-specific inhibitor in human platelets. Deletion of GRK2 in mouse platelets causes increased platelet accumulation following laser-induced injury in cremaster muscle arterioles, particularly in the shell region of thrombi. In addition, this deletion increases ADP-induced pulmonary thromboembolism. GRK2 -/- platelets also have increased platelet aggregation in response to ADP, but not to PAR4 receptor agonist, TxA 2, or convulxin. Underlying these changes in GRK2 -/- platelets is an increase in Ca 2+ mobilization, Akt phosphorylation, and Rap1 activation in response to ADP, and an attenuated rise of cAMP levels in response to ADP in the presence of prostaglandin I 2. Furthermore, platelet aggregation can be restored in GRK2 -/- platelets in response to ADP re-stimulation, indicating that GRK2 contributes to ADP receptor desensitization. To further assess the role of GRK2 in the P2Y 12 signaling pathway in vivo, we examine laser-induced thrombus formation in WT and GRK2 -/- mice treated with the P2Y 12 antagonist, cangrelor. Cangrelor treatment eliminates the phenotypic difference in platelet accumulation between WT and GRK2 -/- mice in response to injury. Using a specific GRK2 inhibitor, pharmacologic inhibition of GRK2 activity in human platelets results in an increase in platelet activation in response to ADP. Finally, our biochemical studies show that GRK2 binds to endogenous Gβγ subunits during platelet activation.

Taken together, we have demonstrated for the first time that 1) GRK2 plays a negative regulatory role in platelet activation by attenuating ADP-dependent signaling, 2) it does this by limiting P2Y 1 and P2Y 12-mediated signaling, 3) GRK2 interacts with Gβγ and functions as a signaling hub in platelets for fine-tuning GPCR signaling, and 4) although the potential inhibition of GRK2 can be beneficial for treatment of heart diseases, maintaining GRK2 activity in platelets could be beneficial for prevention of thrombotic diseases.

Disclosures

No relevant conflicts of interest to declare.
CUX1 Deficiency Potentiates RAS Signaling to Drive Malignancy
Authors: An Ningfei , Imgruet Molly K , Khan Saira , Jueng Lia , Gurbuxani Sandeep , McNerney Megan E.
Published: November 2021

-7/del(7q) is prevalent in high-risk myeloid neoplasms and frequently co-occurs with gain-of-function mutations in the RAS pathway. Herein, we identify a genetic interaction between RAS and the 7q-encoded transcription factor, CUX1, that encompasses hematopoietic malignancies and solid-tumors. Mice with both oncogenic NrasG12D and Cux1 knockdown developed accelerated myeloid malignancies with leukemic transformation. Oncogenic RAS imparts increased self-renewal on CUX1-deficient hematopoietic stem/progenitor cells (HSPCs). Reciprocally, CUX1 knockdown amplifies RAS signaling through reduction of negative regulators of RAS/PI3K signaling. Accordingly, NrasG12D;Cux1-knockdown HSPCs have heightened growth factor-sensitivity and downstream RAS pathway activation. Double mutant HSPCs were responsive to PIK3 or MEK inhibition. Similarly, low expression of CUX1 in primary AML samples correlates with sensitivity to the same inhibitors, suggesting a viable therapy for malignancies with CUX1 inactivation. This work demonstrates an unexpected convergence of an oncogene and tumor suppressor gene on the same pathway.

Disclosures

No relevant conflicts of interest to declare.
Improving Residents' Knowledge of Peripheral Smears
Authors: Ashraf Sara , Shenouda Mina
Published: November 2021

Background: A peripheral smear is an important diagnostic tool, inexpensive, reliable and quick. It reflects the functional status of the bone marrow and can identify many blood disorders, some of which are life threatening and very time sensitive. Peripheral smears are useful to assess cytopenic states (eg, anemia, leukopenia, thrombocytopenia) or identify hematologic emergencies such as thrombocytopenic purpura/hemolytic uremic syndrome, acute myeloid leukemia, and disseminated intravascular coagulation etc. Sometimes it is sufficient to make a diagnosis solely based on them which can be life saving when time is of the essence. Based on our literature search, there are no studies to assess internal medicine residents' knowledge and understanding of peripheral smears.

Methods: We created a multiple-choice questions survey of 13 questions for residents to fill out, involving common findings on peripheral smears and those found in hematologic emergencies that every resident should be aware of. Response rate in all three program years (PGY) was 100%. 16/16 PGY-1, 10/10 PGY-2 and 9/9 PGY-3 completed the survey.

Results: Average pre-intervention score for PGY-1 was 42%, PGY-2 was 34% and PGY-3 was 30%. Questions most commonly wrong were regarding target cells, metamyelocytes, giant platelets and schistocytes. Most were correct about microcytosis and Auer rods. We then conducted the same survey weeks after our intervention, which was interactive, small group didactics sessions on peripheral smears. We compared results of the survey before and after didactics to check for improvement. Post-intervention score for PGY-1 was 98%, PGY-2 and PGY-3 were 96%.

Conclusions: Most medical residents are not aware of common interpretations of peripheral smears and this does not seem to improve with each progressive year. Improving residents' knowledge of peripheral smears is a cost effective and quick measure that can improve patient care, in addition to potentially improving internal medicine board results. In the future, we aim to make a curriculum for peripheral smear review for residents. We can compare results of the survey with hematology oncology fellows.

Disclosures

No relevant conflicts of interest to declare.
Magnetism-Induced Cell-Targeting Transplantation Improves Early Hematopoietic Reconstitution in a Murine Bone Marrow Transplantation Model
Authors: Mai Qiusui , Zhou Yongjun , Wang Zhangyuan , Chen Quanfeng , Lv Yufang , Wu Qiaolan , Huang Hao , Zhang Qiuxia , Zhang Linlin , Li Chengyao , Liu Qifa , Jiang Qianli
Published: November 2021

Background

Early hematopoietic reconstitution is essential for improving survival and reducing complications after hematopoietic stem/progenitor cell (HSC/HPC) transplantation (HSCT/HPCT). Increasing HSC/HPC homing to the bone marrow is a potential approach for promoting hematopoietic reconstitution.

Methods

We proposed the transplantation of HSCs/HPCs with a magnetism-induced cell-targeting transplantation (MagIC-TT) strategy. HSCs/HPCs were magnetized with CD45 microbeads. The biological characteristics (morphology, proliferation, viability, and ferroptosis) and target migration ability of these cells were studied in vitro. The hematopoietic reconstitution experiments were constructed in vivo in autologous and allogeneic bone marrow transplantation models with grouping showed as Table 1. The therapeutic effects were assessed by survival, donor chimerism, routine blood examination and histological analysis. We also performed transcriptomic sequencing for further mechanistic studies.

Results

The biological characteristics was found no significant difference between the MagIC-TT and non-MagIC-TT groups, while migration ability was greatly improved with MagIC-TT (Data not showed). The survival rate was higher in the MagIC-TT group and significantly different in the allogeneic model (P<0.05). Hematopoietic reconstitution of donor chimerism, WBCs, HGB, and PLTs was faster in the MagIC-TT groups (within 22 days) (Figure 1). Confocal observation showed that more donor cells (eGFP +) were retained in the injected femur of the MagIC-TT group than in the femur of the non-MagIC-TT group 7 days after transplantation (P<0.05) (Figure 2). The severity of aGVHD (Assessed by survival, body weight, back arching, fur losting, diarrhea etc.) was reduced in the MagIC-TT groups in the allogeneic model (P<0.05) (Data not showed). Transcriptome sequencing revealed differentially expressed genes (DEGs) involved in localization/locomotion and pathways, cytokine-cytokine receptor interactions and chemokine signaling pathways between the two groups (Figure 3).

Conclusion

The MagIC-TT strategy improves HSC/HPC homing, resulting in faster hematopoietic reconstitution in a murine bone marrow transplantation model.

Key words

Magnetism; Hematopoietic Stem Cell Transplantation (HSCT); Cell Homing; Hematopoietic Reconstitution; Ferroptosis

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Ruxolitinib for Steroid-Refractory Acute Graft-Versus-Host Disease: A Single Centre Retrospective Study
Authors: Suo Yuan , Liu Jiapei , Sun Yiming , Wu Qiaoyuan , Jin Hua , Liu Qifa
Published: November 2021

Background

Acute graft-versus-host disease (aGVHD) is the major cause of non-relapse morbidity and mortality after allogeneic hematopoietic stem cell transplantation(allo-HSCT). But only 40% of the patients will respond to first-line therapies Corticosteroids. Ruxolitinib, a selective Janus kinase (JAK) 1/2 inhibitor, reduces the incidence and severity of GVHD while preserving graft-versus-leukemia effects in preclinical models. The retrospective study evaluated the efficacy of ruxolitinib compared with other second-line therapies for steroid-refractory aGVHD (SR-aGVHD) in a single-center.

Methods

A total of 90 patients who developed SR-aGVHD after HSCT were evaluated in this retrospective study.They were treated with ruxolitinib (n=45) or other salvage-therapies (n=45) including MTX, basiliximab, etanercept and MSC at our institution. The primary endpoint was the overall response rate (ORR) at Day 28. Additional endpoints included overall survival (OS), cumulative incidence rates of failure-free survival (FFS), relapse, non-relapse mortality (NRM) and cGVHD. Treatment failure was defined as 1) addition of new systemic therapy for aGVHD, 2) NRM, 3) relapse, 4) progression of hematologic disease. FFS and OS were calculated from the day of starting the use of second-line treatments for aGVHD.

Results

The median age was 34 years (range 14-69). At the time of enrollment 25 patients had grade Ⅱ disease, 39 with grade Ⅲ disease, 26 with grade IV disease. The skin was involved in 54.4%, lower gastrointestinal (GI) tract in 78.9% and liver in 20.2%. With a median follow-up of 1.33 years, the ORR at day 28 was higher in ruxolitinib group than non-ruxolitinib group (62.2% [95% CI, 47.5%-77.0%] vs. 26.7% [95% CI, 13.2%-40.1%], P=0.001) (Table 1). The 1-year OS was 64.4 % and 45.5% in the two groups, respectively (P=0.0382). Ruxolitinib treatment also improved the 1-year cumulative incidence of FFS (57.8% vs. 26.6%, P=0.002), while the 1-year cumulative incidence of relapse did not differ significantly (9.6% vs. 20.0%, P=0.195). The 1-year cumulative incidence of NRM was lower in the ruxolitinib group than the non-ruxolitinib group (24.4% vs. 45.3%, P=0.023). The 1-year and 3-year cumulative incidence of cGVHD were 17.8% vs. 33.3% and 26.8% vs. 44.4% between the ruxolitinib group and non-ruxolitinib group (P=0.10 and P=0.04).

Conclusions

Our study demonstrated that ruxolitinib is effective than other second-line treatments in patients with SR-aGVHD due to the higher response rates and the improvement of prognosis. Furthermore, ruxolitinib could reduce the incidence and severity of chronic GVHD in aGVHD patients.

Keywords: Ruxolitinib; Steroid-refractory; Acute graft-versus-host disease; Haploidentical hematopoietic stem cell transplantation

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
CCR2 Expression Promotes Diffuse Large B Lymphoma Cell Survival and Invasion
Authors: Yan-Li Li , Hu Quan-Quan , Wen Zhao-Feng , Li Qian , Zhai Zhi-Min
Published: November 2021

Objective: Diffuse large B cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin lymphoma world wide. It is a phenotypically and genetically heterogeneous disease, accounting for 30-40% of all cases. 50%-70% of patients can be cured by the R-CHOP regimen, but nearly one-third of patients develop relapsed or refractory disease. CC chemokine receptor 2 (CCR2), the high affinity receptor of CC-Chemokine Ligand 2 (CCL2), which is the most representative of the CC chemokine family members, has be regarded to involve in tumor growth, angiogenesis, epithelial mesenchymal transition, metastasis and immune escape etc.. In recent years, the role and mechanism in DLBCL has not been reported yet. Our preliminary study showed that high expression of CCR2 was correlated with clinicopathological characteristics, and an adverse prognostic factor for overall survival (OS) and progression-free survival (PFS) of DLBCL patients. The purpose of this study is to investigate the role of CCR2 expression in DLBCL cells proliferation and migration by in vitro and in vivo.

Methods: CCR2 expression were analyzed in human DLBCL cell lines (SUDHL-2, SUDHL-4, SUDHL-6, OCI-Ly8 and OCI-Ly10) by Western blot (WB). SUDHL-2 and OCI-Ly8 cells were incubated with CCR2 antagonist SC-202525 (Santa Crutz Biotechnology), and control cells were left untreated. The proliferation, migration, apoptosis and signaling pathway were detected by CCK8, transwell, flow cytometry (FC) and WB, respectively in vitro. The engraftment, tumor growth, dissemination and survival time were observed in BALB/c nude mice.

Results: CCR2 were expressed in all human DLBCL cell lines (relative CCR2 expression was higher in SUDHL-2, SUDHL-4 and OCI-Ly8 than in SUDHL-6 and OCI-Ly10 cell lines). Blockade of CCR2 expression signaling with CCR2 antagonist inhibited tumor cell proliferation, migration and anti-apoptosis ability. The signaling involved in the proliferation and migration of DLBCL cells by activating PI3K/Akt signaling pathway, and induced apoptosis through activation of P38MARK signaling pathway. Expression of CCR2 was also associated with increased engraftment, tumor growth and dissemination, and decreased survival time in xenograft mice. Furthermore, administration of CCR2 antagonist decreased tumor growth and dissemination of DLBCL cells, and increased survival time in the xenograft model.

Conclusions: Our study demonstrates that CCR2 plays an important role in the development of DLBCL by stimulating cell proliferation, migration and anti-apoptosis. The inhibition of CCR2 may, therefore, be a potential target for anticancer therapy in DLBCL.

Disclosures

No relevant conflicts of interest to declare.
Enhancing Diversity in Oncology Clinical Trials: Findings of a Case Study in Improving Black Patient Representation in a Multiple Myeloma Trial
Authors: Bhandari Sudip , Lagor Charles , Mueller Judith , Whyte Warren , Heilbroner Samuel
Published: November 2021

Background:

Black patients are underrepresented in multiple myeloma (MM) clinical trials. Despite the promise of Real-World Data (RWD), little research exists on RWD's usage to address this health disparity. In collaboration with a large pharmaceutical partner, we used RWD from commercial datasets (ConcertAI's Electronic Medical Record and claims datasets) aimed at identifying sites with a large Black patient population. We recommended including these sites in a recent clinical trial of Chimeric Antigen Receptor T cell (CAR-T) therapy for MM patients.

Methods:

We used the following criteria to identify promising sites: (1) high Black patient density, (2) access to a CAR-T accredited parent organization within 100 miles, (3) a hematologist/oncologist who treats MM patients, and (4) a history of treating MM patients with a Proteasome Inhibitor (PI) and Lenalidomide (Len) in the first line of therapy. For (1), sites were ranked using the lower 95% confidence interval for the percent of Black MM patients at the site. For (4), only sites with at least five MM patients who received PI and Len were included.

Our data sources were: ConcertAI's Electronic Medical Record (EMR) and claims datasets to link each patient to a site, and Google maps API to identify the CAR-T center nearest each oncology site. The patients in our data sets were not identifiable, and our research was conducted in compliance with the Health Insurance Portability and Accountability Act.

After having identified and filtered promising sites, we curated individual candidates in the order of Black patient density. The purpose of curation was to validate a final list of sites.

Results:

We identified 17 promising clinical trial sites affiliated with 16 healthcare systems in the mid-west, mid-Atlantic, southeastern, and southwestern regions of the United States (table 1). Our RWD captured an average of 141 MM patients (range: 6-791) who were treated at the 17 sites from 2015-2020. Thirty-nine percent of the patients were Black (range: 13-67%). This percentage was three times the recruitment rate of black patients in MM trials in the US (13%). On average, the sites were 44 miles driving distance (range: 0.8-96 miles) from the closest CAR-T center, had eight hematology/medical oncology specialists on staff (range: 1-17), and had previous interventional trial experience (13 sites had experience with MM trials). All of the identified sites were community-based sites, and none of the sites were previously identified by our pharmaceutical partner.

Conclusions:

We demonstrated that RWD can be leveraged to identify clinical trial sites with a high potential for Black patient recruitment, thereby addressing a known health disparity problem within multiple myeloma (MM) clinical trials.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Magnetic Resonance Imaging Reveals Significantly Increased Cerebral Blood Flow in the Hemoglobin E Mouse Model
Authors: Cui Min-Hui , Branch Craig A , Hirsch Rhoda Elison
Published: November 2021

Hemoglobin E (HbE) is the most common hemoglobin mutation world-wide. EE individuals exhibit a mild anemia as do those with β-thalassemia trait. A paradox arises with the combination of two relatively benign genes in HbE/β-thalassemia (HbE/β-thal) giving rise to highly morbid symptoms, anemia, growth retardation, developmental retardation, thalassemic bone type development, chronic leg ulcers (for a review, S Fucharoen and DJ Weatherall 2012) and often early mortality arising from cardiac failure (N Olivieri, Z Pakbaz et al. 2011). One approach to understanding the characteristics of HbE and this paradox was to generate a transgenic mouse model, expressing solely human HbE. This HbE mouse model exhibits a mild oxidative stress that parallels that observed in human EE individuals (QY Chen, EE Bouhassira et al. 2004, QY Chen, ME Fabry et al. 2012). These transgenic mice also exhibit mild cardiac dysfunction with depressed left ventricular contraction. We present here the first evidence of a significant increase in cerebral blood flow in the full knockout HbE (HbEKO) transgenic mouse model expressing solely human HbE compared to both the HbE+HbF (γ, gamma)-globin transgenic mouse, and the normal background C57 mouse. Cerebral blood flow is assessed non-invasively by MRI imaging. All protocols were approved by the Albert Einstein Institutional Animal Care and Use Committee.

Comparing the HbEKO mouse (without human γ globin) to the C57 normal background mouse, a significant increase of over 20% in thalamus cerebral blood flow at baseline is observed (p=0.008). There is also an approximate 18% reduction in thalamus cerebral blood flow comparing HbE low γ mice to the full HbEKO (no γ) (p=0.011). Concomitant with these observations, no significant difference is observed comparing these low gamma HbE mice to the normal C57 background mouse (p=0.384). The goodness of the data is also seen in the relatively small variation in cerebral blood flow amongst the individual mice in each subset. These findings are of particular relevance to reports of neurologic symptoms, intracerebral hemorrhage, and brain infarct in HbE/β-thal patients (V Wong, YL Yu et al. 1990, S Das, S Dubey et al. 2019).

In conclusion, these results suggest a direct role of HbE RBC initiating altered cerebral blood flow that when further complexed with β-thal could lead to intracerebral hemorrhage and other cerebral pathophysiology. The finding that the HbEKO mice with high HbF are not significantly different in cerebral blood flow from C57 mice may lend further support to therapeutic approaches enhancing the production of HbF in severe hemoglobinopathies, such as in sickle cell anemia, HbE/β-thal, and β-thal individuals.

Disclosures

No relevant conflicts of interest to declare.
Inherited Thrombophilias Are Associated with an Increased Risk of COVID-19 Associated Venous Thromboembolism: A Prospective Population-Based Cohort Study
Authors: Stevens Hannah P , Canovas Rodrigo , Peter Karlheinz , Tran Huyen , Kaplan Zane , McFadyen James David
Published: November 2021

Background: COVID-19 is associated with high rates of venous thromboembolism (VTE). The impact of common inherited thrombophilias on the development of COVID-19-associated VTE (COVID-19 VTE) is not well understood.

Objective: To determine if the presence of inherited thrombophilias modifies the risk of COVID-19 VTE or COVID-19 mortality.

Methods: Prospective population-based cohort study evaluating adult participants of the UK Biobank diagnosed with COVID-19 between November 2019 and May 2021. Individuals were of European descent and aged between 45 and 69 at recruitment to UK Biobank. We evaluated six single nucleotide polymorphisms including rs6025 (Factor V Leiden mutation) and rs1799963 (Prothrombin mutation) in addition to two polygenic risk scores (PRS-VTE and PRS-ABO). A genome-wide association study was performed for associations with COVID-19 VTE. COVID-19 VTE was defined using International Classification of Diseases and Related Health Problems, Tenth Revision (ICD-10) codes for VTE following COVID-19 diagnosis. COVID-19 mortality was defined using ICD-10 codes for COVID-19 on the death certificate.

Results: Demographic and clinical characteristics are shown in Table 1. Of the 13 712 COVID-19 positive individuals included in the analysis, the median age was 54 years and 52.5% were female. There were 197 (1.4%) cases of COVID-19 VTE and 890 (6.5%) died due to COVID-19. The rs6025 variant, synonymous with FVL, was associated with a 1.8-fold risk of COVID-19 VTE (95% CI 1.040-2.931) (Table 2). The risk of COVID-VTE was also increased with rs2066865 (OR 1.345; 95% CI 1.074-1.675) and the PRS-VTE (OR 1.262; 95% CI 1.081-1.468) (Table 2). COVID-19 VTE was associated with increased COVID-19 mortality (OR 2.731; 95% CI 1.885-3.901) but this study found no association between the studied inherited thrombophilias and COVID-19 mortality (Table 2). On genome-wide analysis, two novel SNPs, rs4975019 and rs2875853, located on chromosomes 4 and 16 respectively, were associated with an increased occurrence of COVID-19 VTE.

Conclusions: These data demonstrate that several inherited thrombophilias increase the risk of COVID-19 VTE and suggest that two novel SNPs are associated with COVID-19 VTE. These results suggest that certain inherited thrombophilias may assist in characterising a subgroup of COVID-19 patients at higher risk of thrombotic events who require individualised antithrombotic therapy. Future prospective studies are required to evaluate inherited thrombophilias in this patient cohort.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Manipulating MEF2C: Discovering Novel Drugs to Target ETP-ALL
Authors: Meijer Mariska T. , Cordo' Valentina , Hagelaar Rico , Smits Willem K. , Meijerink Jules PP
Published: November 2021

Ectopic MEF2C expression due to rearrangements have been recurrently found in patients with human early thymocyte progenitor acute lymphoblastic leukemia (ETP-ALL). ETP-ALL is the most immature subtype of T-ALL and is characterized by the expression of myeloid lineage markers and an ETP-like gene expression profile. In normal haematopoiesis, MEF2C expression occurs at immature stages and in myeloid- and B cell lineages but is absent in early T cell progenitors.

Here, we investigate MEF2C as an oncogene for ETP-ALL. First, we validated our initial observations in an independent T-ALL cohort that ETP-ALL patients highly express MEF2C along with BCL2, HHEX, LMO2 and LYL1 in contrast to other T-ALL subtypes. In addition, we have demonstrated that enforced MEF2C expression in the T-lineage drives a bi-phenotypic CD3 +CD19 + leukemia in mice by inhibiting a T-cell developmental program in favor a B-cell transcriptional program and that resembles human ETP-ALL. This confirms MEF2C as an oncogenic driver of ETP-ALL.

We developed a model system that enabled screening of various drug libraries for compounds that overcome developmental arrest induced by MEF2C-expression, as evidenced by the upregulation of CD3 and TCRγδ. The ETP-ALL-like LOUCY cells express MEF2C as consequence of a del5(q14-qter). Further induction of MEF2C levels (i.e. LOUCY-iMEF2C cells) blocked differentiation of LOUCY cells towards a CD3 +/TCRγδ + phenotype on OP9-DL1 or DLL4-coated plates in contrast to parental LOUCY cells. This model enabled high-throughput screening of drug libraries using a flow cytometry-based assay for drugs that further promote differentiation of LOUCY-MEF2C cells. We identified various epigenetic inhibitors that promoted differentiation of LOUCY-iMEF2C cells. Unlike AML patients, where HDAC4 was shown to play an important role in the activation of MEF2C and the propagation of the leukemia, we found that different HDACs are involved that can block or promote MEF2C function. This suggests that the role of different HDACs in MEF2C regulation is cell-type or disease dependent.

In addition, over-expression of MEF2C raised cellular resistance towards prednisolone treatment. Conversely, inhibition of MEF2C sensitized cells to prednisolone cytotoxicity, even in the presence of MEF2C-overexpression. Together, these data show that MEF2C is an oncogenic driver of ETP-ALL, and ectopic expression of MEF2C negatively affects steroid sensitivity. Specific targeting of only those HDACs that promote MEF2C activity may provide new therapeutic options in ETP-ALL by inhibiting MEF2C function and enhancing steroid sensitivity.

Disclosures

No relevant conflicts of interest to declare.
Prognostic Significance of PD1, PD-L1 Expression , Pathological Subtypes and Metabolic Activity on 18F-FDG PET/CT in Refractory /Relapsing Pediatric Hodgkin Lymphoma
Authors: Khedr Reham , Khorshed Eman , Hassanein Omneya , Abdelrahman Hany , Elwakil Madeeha , Zaghloul Mohamed , Hamoda Asmaa Mohammed
Published: November 2021

Background:

Hodgkin Lymphoma (HL) is a unique disease entity both in its pathology and the young patient population that it primarily affects. Several meta-analyses have demonstrated that high PD-L1 expression levels are correlated with adverse clinical and pathologic features. Objectives: The aim of this study is to evaluate the correlation between the expression of PD-L1 and clinicopathological features, as well as the prognostic significance of PD-L1 expression with regard to interim PET response in relapsing / refractory pediatric HL. Methods: We measured the expression of PD-1/PD-L1 in the baseline diagnostic samples of children with relapsing/ refractory classical HL. The results were correlated with the pathological subtypes as well as the clinical outcome. Results: Of the 88 included patients, 77% had advanced stage HL. PD-1 expression was detected in 50% of cases, whereas PD-L1 (membranous) was expressed by tumor cells in 60% of the cases, and strongly expressed in 16% of cases. Notably, PD-L1 (cytoplasmic) was detected in 55% of the cases. There was a significant differences in the expression levels of PDL-1 between the different pathological subtypes (p = 0.006). OS of patients with PD-L1expression (Cytoplasmic) was 83% vs 91% in patients with absent expression (P=0.001). There was no prognostic significance of PD-L1 expression with regard to PET response (p=0.31). Conclusion: Although PD-L1 expressions did not show statistically significance with well-established prognostic factors, our preliminary data indicate that pathological subtypes and cytoplasmic expression of PD-L1 may have a prognostic implication on survival in pediatric HL.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Research on the Mechanism of SIRT1/AMPK Inducing Resistance to Imatinib in K562 Cells By Inhibiting Srebp to Promote Lipid Metabolism
Authors: Luo Jie , Luo Dongmei , Liang Hanying , Tan Yaxian , Liu Xiaoli , Liu Qifa , Xu Na
Published: November 2021

Background Leukemia stem cells (LSC) have the characteristics of independent of BCR-ABL kinase activity and metabolic reprogramming to support self-renewal. They are the root cause of the occurrence of chronic myeloid leukemia (CML), mediating TKI resistance and leading to clonal evolution. LSC self-renewal depends on the activity of SIRT1, the enhanced of SIRT1 induces LSC mitochondrial metabolic reprogramming. Studies have shown that SIRT1 and abnormally activated lipid metabolism are important reasons for inducing CML resistance to imatinib.

Objective Study the mechanism of SIRT1 regulating lipid metabolism and intervention strategies under imatinib therapy.

Method Liquid chromatography coupled with mass spectrometry (LC-MS) were used to analysis the lipid metabolism in imatinib-resistant CML cell line K562-R under low-glucose conditions. RT-qPCR and Western blot were used to detect the key enzymes related to lipid metabolism, and the SIRT1 inhibitor TV-6 was given to study the regulation mechanism of SIRT1 on lipid metabolism.

Result Firstly, we found that the glycolysis/gluconeogenesis pathway and fatty acid metabolism were abnormal in K562-R cells by metabolomics, compared with normal culture conditions, the expression of SIRT1 and AMPK were significantly increased, the expression of fatty acid synthesis-related genes (SREBP, FASN, ACC, SCD1) was significantly reduced, and the fatty acid β oxidation rate-limiting enzyme CPT-1 is significantly increased inK562-R cells under low-glucose conditions. Therefore, in the low-glucose environment, the expression of the above genes was reversed after the application of the SIRT1 inhibitor TV-6.

Conclusion K562-R imatinib-resistant cells can down-regulate the expression of SREBP through up-regulated SIRT1/AMPK pathway under low-glucose conditions, thereby inhibiting the expression of transcription factors involved in lipogenesis, reducing fatty acid synthesis, and providing the imatinib-resistant cells with the energy which required for proliferation.

Keyword Chronic myeloid leukemia; Metabolomics; Lipid metabolism; SIRT1/AMPK

Disclosures

No relevant conflicts of interest to declare.
The MYC/MIZ1 Complexes Are Required for Germinal Center B Cell Lymphomagenesis
Authors: Gunawan Arief Suryono , Toboso-Navasa Amparo , Chakravarty Probir , Clear Andrew James , Calaminici Maria , Calado Dinis Pedro
Published: November 2021

We previously showed that the formation of protein complexes between MYC and its partner MIZ1 (MYC-interacting zinc finger 1) is critically required for germinal center (GC) B cell expansion (Toboso-Navasa et. al., JEM 2020). MYC and MIZ1 are transcriptional activators; however, they can form a transcriptional repressor complex that represses MIZ1 target genes. High expression of MYC is commonly found in aggressive B cell lymphoma, most notably in Burkitt Lymphoma (BL) and in a fraction of Diffuse B cell lymphomas (DLBCLs). In DLBCL, MYC positivity is associated with poorer prognosis, especially when co-expressed with BCL2, and increased proliferative capacity. However, it remains unclear whether the requirement for MYC-MIZ1 complexes for cell expansion is retained in lymphoma, similar to what we observed in GC B cells.

We first investigated MIZ1 expression in primary samples of B cell lymphoma sub-types and found it to be ubiquitous in BL (100% of cases; 14/14), whereas virtually absent in low-grade Follicular Lymphoma (7% of cases, 4/58). Roughly 42% of DLBCL cases (36/85; ) co-expressed MYC and MIZ1 and that was associated with increased cell proliferation assessed by Ki67. To investigate the role of MYC-MIZ1 complexes in lymphomagenesis, we generated compound mutant mice overexpressing in GC B cells wild-type MYC or a MYC mutant that cannot interact with MIZ1 (MYC VD) in combination with PI3K. MYC VD cannot repress MIZ1 target genes but displays normal interaction with MAX and transcriptional activation. As shown previously (Sander et. al., Cancer Cell 2012) MYC synergised with PI3K for BL development; however, overexpression of MYC VD plus PI3K had significantly delayed disease development and developed instead plasma cell hyperplasia. Analysis of pre-tumoral cells by single cell RNAseq revealed massive expansion of a unique GC B cell cluster only in mice carrying MYC and PI3K and this cluster had the highest suppression of MYC-MIZ1 target genes among all GC B cell clusters.

Taken together, this data indicates that the fraction of cells in which MYC and MIZ1 are co-expressed represents a "sweet spot" for B cell lymphomagenesis; and that the transcriptional repressive complex formed by MYC and MIZ1 is crucial for GC B lymphomagenesis.

Disclosures

Toboso-Navasa: Benevolent AI: Current Employment. Calado: Myricx Pharma: Consultancy, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company, Patents & Royalties: Cancer Treatments. WO patent WO 2020/128475 A1 (2020).
NK Cells of Acute Myeloid Leukemia Patients Exhibit Exhausted Phenotype with Impaired Functional Activity
Authors: Zhigarev Dmitry , MacFarlane Alexander W. , Drenberg Christina Diane , Nejati Reza , Varshavsky Asya , Campbell Kerry S.
Published: November 2021

Acute myeloid leukemia (AML) accounts for about one third of all leukemias, with half of all leukemia deaths. The low 5-year survival rate and manifestation of this deadly disease in old age reinforce the need for new safe and effective AML therapies. Considering the exceptional role of immune cells in the recognition and elimination of tumor cells, one of these methods is immunotherapy. However, for the development of immunotherapeutic approaches, it is necessary to clearly understand the role of certain effector cells in the pathogenesis of the disease, as well as to have the knowledge about the phenotypic characteristics of these cells.

Natural killer (NK) cells play important roles in innate cancer immunosurveillance, and some published data indicate that the antitumor function of NK cells is reduced in AML patients. To expand upon previous work, we performed a comprehensive analysis of the phenotypic and functional characteristics of NK cells in previously untreated AML patients that took into account a wide variety of biomarkers. The goals of this study were to define the phenotypic and functional differences in NK cells from AML patients and healthy donors and determine how these parameters affect outcome of the disease.

We used 14-color flow cytometry to assess more than 30 measurable markers on NK cells from the peripheral blood of 33 untreated AML patients and age-matched healthy controls. In addition, NK cells were tested for interferon-γ responses under antibody-dependent cellular cytotoxicity conditions. Results in AML patients were then compared to healthy donors.

We found that the NK cells of patients with AML have a significantly lower capacity to secrete IFN-γ and showed numerous signs of an exhausted phenotype, as compared to healthy controls. These included increased surface expression of CD39, PD-1, and LILRB1 on NK cells from AML patients. Interestingly, surface expression of the TIGIT checkpoint receptor did not differ between AML patients and healthy donors, but surface expression of the activating receptor DNAM-1, which shares the same ligands on tumor cells, was decreased. All of these data confirm that the NK cells of AML patients are functionally impaired.

We also noted that the frequency of CD57 + NKG2C + KIR + memory-like "adaptive" NK cells was greater in the blood of AML patients. The proportion of adaptive NK cells did not correlate with the age of donors. Phenotypic features of this cell subpopulation from the AML patients included significantly increased expression levels of CD56 and significantly lower expression of the activating receptor DNAM-1.

Disclosures

Zhigarev: Janssen R&D: Research Funding; Immunitas Therapeutics: Consultancy; Tavotek Biotherapeutics: Consultancy. Drenberg: Janssen R&D: Current Employment. Campbell: Janssen R&D: Research Funding.
Anti-Leukemic Properties of the Kinase Inhibitor OTSSP167 in T Cell Acute Lymphoblastic Leukemia
Authors: Bridges Cory Seth , Chen Taylor J , Puppi Monica , Lacorazza H. Daniel
Published: November 2021

The remission rates of T-cell acute lymphoblastic leukemia (T-ALL), a hematological malignancy in children, have improved through the years. Despite this advance, 10-20% of the patients do not respond to current therapy, and relapse leukemia is the leading cause of death in children with cancer. Our previous work showed aberrant activation of the actionable MAP2K7-JNK pathway in pediatric T-ALL and proposed this pathway as a target for pharmacological inhibition. Here we demonstrate the anti-leukemic capacity of the kinase inhibitor OTSSP167 in T-ALL via MAP2K7 inhibition. OTSSP167, a MELK inhibitor shown to also inhibit MAP2K7, showed a dose-dependent cytotoxicity in a panel of T-ALL cell lines with IC50 in the nanomolar range (10-50 nM). OTSSP167 induced apoptosis and G2/M cell cycle arrest that correlated with increased cleavage of PARP and caspase 3 and elevated levels of cyclin B1 and phosphorylated CDC2. Immunoblot blot analysis shows OTSSP167 treatment inhibited the MAP2K7-JNK-ATF2 pathway, in addition to MELK, in a panel of T-ALL cell lines. Biochemical kinase assay shows inhibition of MAP2K7 kinase activity with an IC50 of 160 nM. Furthermore, OTSSP167 was able to inhibit MAP2K7 activated by treatment of T-ALL cells with sorbitol. This kinase inhibitor shows low toxicity in mice (10 mg/Kg) and controls leukemia burden in cell-based xenograft model with significant reduction on the expansion of KOPT-K1 cells and improved survival (P=0.003). Furthermore, OTSSP167 was able to control leukemia in patient-derived xenograft (PDX) model and to target leukemia-initiating cells in the NOTCH1-induced T-ALL mouse model. Altogether, OTSSP167 is a potent MAP2K7 inhibitor with low toxicity that emerges as a potential adjuvant drug to increase treatment response and reduce relapses in pediatric T-ALL.

Disclosures

No relevant conflicts of interest to declare.
Decitabine Combined with Minimally Myelosuppressive Therapy for Induction of Remission of Pediatric High-Risk Acute Myeloid Leukemia with Chromosome 5q Deletion. a Report of Three Cases
Authors: Cheng Shengqin , Xiao Peifang , Wang Juxiang , Gao Li , Zheng Jiajia , Hu Yixin , Ding Xin , Ling Jing , Lu Qin , Pan Jian , Li Bohan , Lu Jun , Wang Yi , Ribeiro Raul C , Hu Shaoyan
Published: November 2021

Cases of pediatric acute myeloid leukemia (AML) with complex karyotypes including chromosome 5 abnormalities are rare and have a dismal prognosis. Management of AML with monosomy 5/del(5q) has not been uniform. We treated three adolescents with this AML subtype with combined low-dose cytarabine and mitoxantrone, concurrent with decitabine and G-CSF, for remission induction. Decitabine was also included in the conditioning regimen before hematopoietic cell transplantation (HCT). All three patients attained complete remission after treatment with this combination. The treatment was well tolerated, and the patients are alive and free of disease at 3.6, 3.2, and 3.0 years after HCT, respectively. Our experience suggests that HCT is required for the eradication of pediatric AML, and possibly MDS, with complex karyotypes including del(5q). Decitabine combined with regimens of low myelotoxicity for remission induction represents an alternative approach to decrease the risk of complications associated with post-remission chemotherapy before HCT.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.