Blood myeloid cells are known to be dysregulated in coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2. It is unknown whether the innate myeloid response differs with disease severity and ...whether markers of innate immunity discriminate high-risk patients. Thus, we performed high-dimensional flow cytometry and single-cell RNA sequencing of COVID-19 patient peripheral blood cells and detected disappearance of non-classical CD14LowCD16High monocytes, accumulation of HLA-DRLow classical monocytes (Human Leukocyte Antigen - DR isotype), and release of massive amounts of calprotectin (S100A8/S100A9) in severe cases. Immature CD10LowCD101−CXCR4+/− neutrophils with an immunosuppressive profile accumulated in the blood and lungs, suggesting emergency myelopoiesis. Finally, we show that calprotectin plasma level and a routine flow cytometry assay detecting decreased frequencies of non-classical monocytes could discriminate patients who develop a severe form of COVID-19, suggesting a predictive value that deserves prospective evaluation.
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•Patients with severe COVID-19 accumulate HLA-DRLow monocytes and immature neutrophils in blood/lungs•Calprotectin level positively correlates with neutrophil count and disease severity•Loss of non-classical monocytes could identify high risk of severe COVID-19
An analysis of patients with severe COVID-19 reveals immature neutrophil and non-classical monocyte pools, with levels of the protein calprotectin linked to disease severity.
Loss-of-function mutations affecting one or both copies of the
Ten-Eleven-translocation (
TET)
2 gene have been described in various human myeloid malignancies. We report that inactivation of
Tet2 in ...mouse perturbs both early and late steps of hematopoiesis including myeloid and lymphoid differentiation in a cell-autonomous manner, endows the cells with competitive advantage, and eventually leads to the development of malignancies. We subsequently observed
TET2 mutations in human lymphoid disorders.
TET2 mutations could be detected in immature progenitors endowed with myeloid colony-forming potential. Our results show that the mutations present in lymphoid tumor cells may occur at both early and later steps of lymphoid development and indicate that impairment of TET2 function or/and expression predisposes to the development of hematological malignancies.
►
TET2 inactivation in mice affects HSC homeostasis and differentiation ►
TET2 inactivation in mice leads to a CMML-like disease ►
TET2 is mutated in human B and T cell lymphomas
APR-246 is a promising new therapeutic agent that targets p53 mutated proteins in myelodysplastic syndromes and in acute myeloid leukemia (AML). APR-246 reactivates the transcriptional activity of ...p53 mutants by facilitating their binding to DNA target sites. Recent studies in solid cancers have found that APR-246 can also induce p53-independent cell death. In this study, we demonstrate that AML cell death occurring early after APR-246 exposure is suppressed by iron chelators, lipophilic antioxidants and inhibitors of lipid peroxidation, and correlates with the accumulation of markers of lipid peroxidation, thus fulfilling the definition of ferroptosis, a recently described cell death process. The capacity of AML cells to detoxify lipid peroxides by increasing their cystine uptake to maintain major antioxidant molecule glutathione biosynthesis after exposure to APR-246 may be a key determinant of sensitivity to this compound. The association of APR-246 with induction of ferroptosis (either by pharmacological compounds, or genetic inactivation of SLC7A11 or GPX4) had a synergistic effect on the promotion of cell death, both in vivo and ex vivo.
•In myelodysplastic syndromes (MDSs), the disease-initiating cell is a rare hematopoietic stem cell (HSC) that transmits the genetic abnormalities to its myeloid and lymphoid progeny.•The ...heterogeneity of MDS phenotypes is related to the diversity of genetics, the various combinations and order of mutations in cooperating genes, and the variegation of clonal hematopoietic hierarchy.•A mutation in a granulomonocytic progenitor may mark the transformation into acute myeloid leukemia.
Myelodysplastic syndromes (MDS) are hematopoietic stem cell (HSC) disorders in which recurrent chromosome abnormalities and gene mutations define a clonal hematopoiesis. The MDS-initiating cell is a rare HSC which transmits the genetic abnormalities to its myeloid and lymphoid progeny. The heterogeneity of MDS phenotypes could be linked to the diversity of genetic events involving epigenetic regulators, chromatin modifiers, splicing factors, transcription factors and signaling adaptors, the various combinations and order of mutations in cooperating genes, and the variegation of clonal hematopoietic hierarchy. Usually, epigenetic and splicing gene mutations occur first. A combination of one epigenetic event with a splicing gene alteration is frequent. The HSC compartment is invaded by a dominant and few minor clones organized linearly or with a branched architecture. The dominant clone containing the first initiating mutations produces myeloid and lymphoid lineages in transplanted immune-deficient mice. The mutations confer a selective advantage to myeloid progenitors at the expense of lymphoid progenitors. In the context of differentiation, one mutation may favor the amplification of granulo-monocytic progenitor, which drives the transformation into acute myeloid leukemia. Understanding the hierarchy of mutations provides insights on the mechanism of transformation. Investigation of mutation pattern and distribution along the hematopoietic tree may influence the therapeutic decision for targeted therapy.
Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is presenting as a systemic disease associated with vascular inflammation and endothelial ...injury. Severe forms of SARS-CoV-2 infection induce acute respiratory distress syndrome (ARDS) and there is still an ongoing debate on whether COVID-19 ARDS and its perfusion defect differs from ARDS induced by other causes. Beside pro-inflammatory cytokines (such as interleukin-1 β IL-1β or IL-6), several main pathological phenomena have been seen because of endothelial cell (EC) dysfunction: hypercoagulation reflected by fibrin degradation products called D-dimers, micro- and macrothrombosis and pathological angiogenesis. Direct endothelial infection by SARS-CoV-2 is not likely to occur and ACE-2 expression by EC is a matter of debate. Indeed, endothelial damage reported in severely ill patients with COVID-19 could be more likely secondary to infection of neighboring cells and/or a consequence of inflammation. Endotheliopathy could give rise to hypercoagulation by alteration in the levels of different factors such as von Willebrand factor. Other than thrombotic events, pathological angiogenesis is among the recent findings. Overexpression of different proangiogenic factors such as vascular endothelial growth factor (VEGF), basic fibroblast growth factor (FGF-2) or placental growth factors (PlGF) have been found in plasma or lung biopsies of COVID-19 patients. Finally, SARS-CoV-2 infection induces an emergency myelopoiesis associated to deregulated immunity and mobilization of endothelial progenitor cells, leading to features of acquired hematological malignancies or cardiovascular disease, which are discussed in this review. Altogether, this review will try to elucidate the pathophysiology of thrombotic complications, pathological angiogenesis and EC dysfunction, allowing better insight in new targets and antithrombotic protocols to better address vascular system dysfunction. Since treating SARS-CoV-2 infection and its potential long-term effects involves targeting the vascular compartment and/or mobilization of immature immune cells, we propose to define COVID-19 and its complications as a systemic vascular acquired hemopathy.
Objectives
To identify which level of D-dimer would allow the safe exclusion of pulmonary embolism (PE) in COVID-19 patients presenting to the emergency department (ED).
Methods
This retrospective ...study was conducted on the COVID database of Assistance Publique – Hôpitaux de Paris (AP-HP). COVID-19 patients who presented at the ED of AP-HP hospitals between March 1 and May 15, 2020, and had CTPA following D-dimer dosage within 48h of presentation were included. The D-dimer sensitivity, specificity, and positive and negative predictive values were calculated for different D-dimer thresholds, as well as the false-negative and failure rates, and the number of CTPAs potentially avoided.
Results
A total of 781 patients (mean age 62.0 years, 53.8% men) with positive RT-PCR for SARS-Cov-2 were included and 60 of them (7.7%) had CTPA-confirmed PE. Their median D-dimer level was significantly higher than that of patients without PE (4,013 vs 1,198 ng·mL
−1
,
p
< 0.001). Using 500 ng·mL
−1
, or an age-adjusted cut-off for patients > 50 years, the sensitivity and the NPV were above 90%. With these thresholds, 17.1% and 31.5% of CTPAs could have been avoided, respectively. Four of the 178 patients who had a D-dimer below the age-adjusted cutoff had PE, leading to an acceptable failure rate of 2.2%. Using higher D-dimer cut-offs could have avoided more CTPAs, but would have lowered the sensitivity and increased the failure rate.
Conclusion
The same D-Dimer thresholds as those validated in non-COVID outpatients should be used to safely rule out PE.
Key Points
•
The median D-dimer level was significantly higher in COVID-19 patients with PE as compared to those without PE (4,013 ng·mL
−1
vs 1,198 ng·mL
−1
respectively, p
<
0.001).
•
Using 500 ng·mL
−1
, or an age-adjusted D-dimer cut-off to exclude pulmonary embolism, the sensitivity and negative predictive value were above 90%.
•
Higher cut-offs would lead to a reduction in the sensitivity below 85% and an increase in the failure rate, especially for patients under 50 years.
Background
Coronavirus disease 2019 (COVID-19) is a respiratory disease associated with endotheliitis and microthrombosis.
Objectives
To correlate endothelial dysfunction to in-hospital mortality in ...a bi-centric cohort of COVID-19 adult patients.
Methods
Consecutive ambulatory and hospitalized patients with laboratory-confirmed COVID-19 were enrolled. A panel of endothelial biomarkers and von Willebrand factor (VWF) multimers were measured in each patient ≤ 48 h following admission.
Results
Study enrolled 208 COVID-19 patients of whom 23 were mild outpatients and 189 patients hospitalized after admission. Most of endothelial biomarkers tested were found increased in the 89 critical patients transferred to intensive care unit. However, only von Willebrand factor antigen (VWF:Ag) scaled according to clinical severity, with levels significantly higher in critical patients (median 507%, IQR 428–596) compared to non-critical patients (288%, 230–350,
p
< 0.0001) or COVID-19 outpatients (144%, 133–198,
p
= 0.007). Moreover, VWF high molecular weight multimers (HMWM) were significantly higher in critical patients (median ratio 1.18, IQR 0.86–1.09) compared to non-critical patients (0.96, 1.04–1.39,
p
< 0.001). Among all endothelial biomarkers measured, ROC curve analysis identified a VWF:Ag cut-off of 423% as the best predictor for in-hospital mortality. The accuracy of VWF:Ag was further confirmed in a Kaplan–Meier estimator analysis and a Cox proportional Hazard model adjusted on age, BMI, C-reactive protein and
d
-dimer levels.
Conclusion
VWF:Ag is a relevant predictive factor for in-hospital mortality in COVID-19 patients. More than a biomarker, we hypothesize that VWF, including excess of HMWM forms, drives microthrombosis in COVID-19.
Specific and reciprocal interactions with the bone marrow microenvironment (BMM) govern the course of hematological malignancies. Matrix metalloproteinase-9 (MMP-9), secreted by leukemia cells, ...facilitates tumor progression via remodeling of the extracellular matrix (ECM) of the BMM. Hypothesizing that leukemias may instruct the BMM to degrade the ECM, we show, that MMP-9-deficiency in the BMM prolongs survival of mice with BCR-ABL1-induced B-cell acute lymphoblastic leukemia (B-ALL) compared with controls and reduces leukemia-initiating cells. MMP-9-deficiency in the BMM leads to reduced degradation of proteins of the ECM and reduced invasion of B-ALL. Using various in vivo and in vitro assays, as well as recipient mice deficient for the receptor for tumor necrosis factor (TNF) α (TNFR1) we demonstrate that B-ALL cells induce MMP-9-expression in mesenchymal stem cells (MSC) and possibly other cells of the BMM via a release of TNFα. MMP-9-expression in MSC is mediated by activation of nuclear factor kappa B (NF-κB) downstream of TNFR1. Consistently, knockdown of TNF-α in B-ALL-initiating cells or pharmacological inhibition of MMP-9 led to significant prolongation of survival in mice with B-ALL. In summary, leukemia cell-derived Tnfα induced MMP-9-expression by the BMM promoting B-ALL progression. Inhibition of MMP-9 may act as an adjunct to existing therapies.
Several prognostic scoring systems have been proposed for chronic myelomonocytic leukemia (CMML), a disease in which some gene mutations-including ASXL1-have been associated with poor prognosis in ...univariable analyses. We developed and validated a prognostic score for overall survival (OS) based on mutational status and standard clinical variables.
We genotyped ASXL1 and up to 18 other genes including epigenetic (TET2, EZH2, IDH1, IDH2, DNMT3A), splicing (SF3B1, SRSF2, ZRSF2, U2AF1), transcription (RUNX1, NPM1, TP53), and signaling (NRAS, KRAS, CBL, JAK2, FLT3) regulators in 312 patients with CMML. Genotypes and clinical variables were included in a multivariable Cox model of OS validated by bootstrapping. A scoring system was developed using regression coefficients from this model.
ASXL1 mutations (P < .0001) and, to a lesser extent, SRSF2 (P = .03), CBL (P = .003), and IDH2 (P = .03) mutations predicted inferior OS in univariable analysis. The retained independent prognostic factors included ASXL1 mutations, age older than 65 years, WBC count greater than 15 ×10(9)/L, platelet count less than 100 ×10(9)/L, and anemia (hemoglobin < 10 g/dL in female patients, < 11g/dL in male patients). The resulting five-parameter prognostic score delineated three groups of patients with median OS not reached, 38.5 months, and 14.4 months, respectively (P < .0001), and was validated in an independent cohort of 165 patients (P < .0001).
A new prognostic score including ASXL1 status, age, hemoglobin, WBC, and platelet counts defines three groups of CMML patients with distinct outcomes. Based on concordance analysis, this score appears more discriminative than those based solely on clinical parameters.
Chronic myelomonocytic leukemia (CMML) is a myelodysplastic syndrome/ myeloproliferative neoplasm whose diagnosis is currently based on the elevation of peripheral blood monocytes to >1 × 109/L, ...measured for ≥3 months. Diagnosis can be ambiguous; for example, with prefibrotic myelofibrosis or reactive monocytosis. We set up a multiparameter flow cytometry assay to distinguish CD14+/CD16− classical from CD14+/CD16+ intermediate and CD14low/CD16+ nonclassical monocyte subsets in peripheral blood mononucleated cells and in total blood samples. Compared with healthy donors and patients with reactive monocytosis or another hematologic malignancy, CMML patients demonstrate a characteristic increase in the fraction of CD14+/CD16− cells (cutoff value, 94.0%). The associated specificity and sensitivity values were 95.1% and 90.6% in the learning cohort (175 samples) and 94.1% and 91.9% in the validation cohort (307 samples), respectively. The accumulation of classical monocytes, which demonstrate a distinct gene expression pattern, is independent of the mutational background. Importantly, this increase disappears in patients who respond to hypomethylating agents. We conclude that an increase in the fraction of classical monocytes to >94.0% of total monocytes is a highly sensitive and specific diagnostic marker that rapidly and accurately distinguishes CMML from confounding diagnoses.
•An increase in the classical monocyte subset to >94% of total monocytes discriminates CMML from other monocytoses with high specificity.•This characteristic increase in classical monocytes disappears in CMML patients who respond to hypomethylating agents.
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