Coagulopathy In Acute Promyelocytic Leukemia Research Articles

Podoplanin Expression Measured By Flow Cytometry Accurately Identifies Patients with Acute Promyelocytic Leukemia at Diagnosis

Acute promyelocytic leukemia (APL) is a hematological emergency associated with high early mortality rates, thus requiring prompt diagnosis and treatment initiation. Currently, diagnostic suspicion relies on morphological criteria, which can be further corroborated by immunophenotypic analyses, until results based on molecular techniques confirm the diagnosis. However, while both cytology and immunophenotypic analyses are very important in this setting, their accuracy can be influenced by inherent morphological variation of blasts, as well as by gating and other technical issues related to flow cytometry. In this context, novel parameters that could contribute to the segregation of APL from other forms of acute myeloid leukemia (AML) would be desirable. Recently, Lavallée et al demonstrated that the aberrant expression of podoplanin (PDPN), a protein that mediates platelet activation in inflammatory contexts, by leukemic cells is as a relevant mediator of the coagulopathy of APL, as well potential biomarker of this condition based on the analysis of primary cells from patients with AML. The aim of this study was to prospectively and independently validate whether the surface expression of PDPN by leukemic promyelocytes measured by flow cytometry can contribute to the differential diagnosis between APL and other forms of AML. We also explored the association of PDPN expression with clinical and laboratory characteristics of these patients. The study population consisted of all consecutive patients diagnosed with APL by cytological, immunophenotypical and molecular techniques in our adult academic hematology hospital from 2020 to 2023. Exclusion criteria included the presence of hemodynamic instability or refusal to participate. The expression of PDPN was measured by flow cytometry using BD FACSCanto II™ cytometer. Analysis was performed in accordance with EuroFlow protocols, with a panel that includes CD45, CD33, CD34, MPO, CD117, CD13, HLA-DR, as well as an antibody against PDPN (PE anti-human Podoplanin Antibody - Catalog# 337004 - BioLegend). Lymphocytes were used as internal negative control for PDPN expression. Data were analyzed with Infinicyt™ software and reported both as the % of PDPN + cells or according to mean fluorescence intensity (MFI). Patients with other forms of AML diagnosed during the same period and matched for age, gender and peripheral blast count were used to compare the diagnostic accuracy of PDPN measurement. Clinical and laboratory data were obtained from the medical records. The study was approved by the Institutional Review Board and all participants provided written informed consent. In total, 19 consecutive patients with confirmed diagnosis of APL were compared with 18 patients with non-APL AML. Clinical and laboratory data from the study population are shown in table 1. The expression of PDPN in leukemic blasts was significantly higher in APL compared to non-APL AML, considering either the percentage of PDPN + cells or the MFI (table 1; figure 1a-b). When considered as a categorical variable, the expression of PDPN was strongly associated with the diagnosis of APL (Chi-square test= 19.169; P<0.001). PDPN expression was inversely correlated with platelet count (R S= -0.47; P=0.003) and fibrinogen (R S= -0.51; P=0.001), but not with other laboratory parameters such as PT and aPTT. No association of PDPN expression with the available clinical variables was observed, although it should be noted that our study is underpowered to confirm these findings. We next estimated the diagnostic accuracy of PDPN expression by flow cytometry by ROC analysis. As shown in figures 1c-d, the area under the ROC curve was higher than 0.9 for both analytical strategies. In conclusion, our prospective study demonstrated that measurement of the expression of PDPN by flow cytometry at diagnosis is capable to accurately segregate APL from other forms of AML. Additional studies are warranted to explore the clinical association of PDPN expression in larger and independent cohorts.

Predictors of very early death in acute promyelocytic leukemia: a retrospective real-world cohort study

Early death (ED) is still the major obstacle to cure in acute promyelocytic leukemia (APL). Most studies focus on 30-day ED; however, little is known on predictors of death before starting APL treatment (very early death — VED) and on predictors of 7-day ED, the period with most deaths due to thrombohemorrhagic diathesis. We hypothesized whether the severity of the coagulopathy of APL could predict VED and 7-day ED. We also aimed to evaluate other characteristics associated with these outcomes. We undertook a retrospective, single-center observational study including newly diagnosed APL patients admitted to our institution between January 2000 and November 2022. Baseline demographical, clinical, and laboratorial data were collected. Statistical analysis was performed using Stata. One hundred four patients were included. The VED rate was 4.8%. A DIC Score ≥ 7 (p = 0.045), serum creatinine > 1.5 mg/dL (p < 0.001%), a DIC Score ≥ 6 within 24 h (p = 0.009), and mechanical ventilation (p < 0.001) were associated with VED. The 7-day ED rate was 12.5%. High-risk (p = 0.007) and hypogranular APL (p = 0.029), DIC Score at diagnosis (p = 0.047), DIC Score ≥ 7 (p = 0.043), DIC Score ≥ 6 within 24 h (p = 0.025), PT prolongation > 6 s (p = 0.002), and creatinine > 1.5 mg/dL (p = 0.004) were associated with 7-day ED. However, only elevated creatinine emerged as an independent predictor of 7-day ED (OR 21.4; p = 0.008). Our study shows that in patients with APL, an elevated creatinine at diagnosis strongly predicts for 7-day ED. A DIC Score ≥ 7 and a Score that remains ≥ 6 within 24 h and a serum creatinine > 1.5 mg/dL significantly associated with VED.

The Coagulopathy of Acute Promyelocytic Leukemia: An Updated Review of Pathophysiology, Risk Stratification, and Clinical Management.

Acute promyelocytic leukemia (APL) has a well-established mechanism and a long-term prognosis that exceeds that of any other acute leukemia. These improving outcomes are due, in part, to all-trans retinoic acid (ATRA) and arsenic trioxide (ATO), two targeted and highly active agents in this disease. However, there remains a considerable morbidity and mortality risk in APL secondary to clinically significant hemorrhagic and/or thrombotic events. Prevention and treatment of these coagulopathic complications remain significant impediments to further progress in optimizing outcomes for patients with APL. Moreover, the relative rarity of APL hinders adequately powered randomized controlled trials for evaluating APL coagulopathy management strategies. This review draws from peer-reviewed works falling between initial descriptions of APL in 1957 and work published prior to January 2023 and provides an updated overview of the pathophysiology of hemorrhagic and thrombotic complications in APL, outlines risk stratification parameters, and compiles current clinical best practices. An improved understanding of the pathophysiologic mechanisms driving hemorrhage and thrombosis along with the completion of well-designed trials of management strategies will assist clinicians in developing interventions that mitigate these devastating complications in an otherwise largely curable disease.

Coagulopathy in Acute Promyelocytic Leukemia: Can We Go Beyond Supportive Care?

Acute promyelocytic leukemia (APL) is characterized by frequent complications due to a distinct coagulopathy. While advances in treatments have improved long-term survival, hemorrhagic and thrombotic complications remain the most common causes of death and morbidity. Improved understanding of the mechanisms of the coagulopathy associated with APL may lead to therapeutic interventions to mitigate the risk of hemorrhage and thrombosis.

Successful treatment of acute promyelocytic leukemia in a patient with prosthetic heart valves: a case report and review of the literature

BackgroundAlthough life-threatening hemorrhage is a usual manifestation of acute promyelocytic leukemia (APL), thrombotic events seem to be more common in APL compared to other subtypes of acute leukemia. The treatment and prophylaxis of thrombosis are controversial due to the high risk of bleeding caused by disseminated intravascular coagulation (DIC) and thrombocytopenia. To the best of our knowledge we report the first case of APL in a patient with prosthetic heart valves successfully treated with a combination of all-trans-retinoic acid (ATRA) and arsenic trioxide (ATO). We hope this case report helps clinicians to manage different spectra of coagulopathy in APL successfully.Case presentationA 38-year-old Asian man presented with diagnosis of APL confirmed by bone marrow biopsy. He was on warfarin due to prosthetic mitral and aortic valves. He was at risk of both hemorrhagic events due to DIC and life-threatening valve thrombosis. Our management regimen included unfractionated heparin adjusted according to the platelet count to prevent both valve thrombosis and bleeding events. The patient tolerated treatment well without any hemorrhagic or thrombotic events, and complete molecular remission was achieved by ATRA and ATO without the need for chemotherapeutic agents.ConclusionAlthough this case is exceptional, a precise evaluation may be needed to select the appropriate dose and type of anticoagulant to treat a patient with APL.

Determination of Protein C Antigen Levels in Hospitalized Patients with Blood Coagulative Defects

Protein C, a vitamin K-dependent anticoagulant serine protease, is involved in blood coagulation. Activated protein C inactivates Va and VIIIa in purified protein systems and stimulates fibrinolysis by indirectly increasing the level of circulating plasminogen activator. In this process, protein S serve as an important factor for activated protein C. In recent years, excess protein S drives cancer cell proliferation and cell survival through oncogenic receptor Axl (Anexelekto). We determined changes of plasma protein C antigen by using rocket immunoassay both in 50 healthy individuals and 103 distinct hospitalized patients. In healthy individuals protein C antigen(PC:Ag) ranges o.6439- 1.4752 µ/ml. The results showed that plasma protein C antigen was considerably high in 22 diabetes mellitus. In contrast, the PC:Ag was significantly decreased in 19 liver cirrhosis(p&lt; 0.001) and in closely line with serum albumin levels(p&lt; 0.05). In 31 acute leukemias, on the average, there was slightly lower values in PC:Ag, and accompanied with the distribution of significant decrease of PC:Ag values in 5 FAB M5 subtype and in 9 hyperleukocytic leukemias. However, the 3 acute promyelocytic leukemia (APL) with overt laboratory criteria of disseminated intravascular coagulation (DIC) had protein C concentration no lower than the remaining 2 patients with infectious DIC, which suggested the coagulopathy in APL might be due to mechanisms different from other forms of DIC.

CLINICAL APPLICATION OF PLASMA PROTEIN C DETERMINATION

Objective: Protein C, a vitamin K-dependent coagulation factor, is involved in blood coagulation. Activated protein C inactivates Va and VIIIa and stimulates fibrinolysis. In this process, protein S serve as an important factor for activated protein C. Furthermore, excess protein S drives cancer cell proliferation and cell survival through oncogenic receptor Axl(Anexelekto). We determined ranges of protein C both in healthy individuals and distinct hospitalized patients.&#x0D; Methods: A total of 100 patients with different diagnostic diseases and 50 healthy individuals were included in their plasma protein C determination. A rabbit antibody against human protein C was used for the quantitative estimation of plasma protein C antigen by using rocket immunoassay.&#x0D; Results: In healthy individuals protein C antigen (PC:Ag) ranges o.6439- 1.4752 µ/ml. The mean coefficient of variation (CV) of length of rocket was calculated to be 12.45%. PC:Ag within laboratory variation was 11.47%. Plasma protein C antigen was destroyed at 56℃ for 30 minutes, whereas no significant decrease of protein C was found at 4℃ refrigerator for one week.&#x0D; Conclusion: The results showed that plasma protein C antigen was considerably high in 22 diabetes mellitus. On the other hand, the PC:Ag was significantly decreased in 19 liver cirrhosis(p&lt; 0.001) and was positively correlation with serum albumin levels(p&lt; 0.05). In 20 acute leukemias, on the average,there was slightly lower values in PC:Ag, and accompanied with significant decrease of PC:Ag in 5 M5 subtype and in 9 hyper- leukocytes acute leukemias. However, the 3 acute promyelocytic leukemia (APL) with overt laboratory picture of DIC(disseminated intravascular coagulation) had protein C concentration no lower than the remaining 2 patients with infectious DIC, which suggested the coagulopathy in APL might be due to mechanisms different from other forms of DIC.&#x0D; &#x0D; Peer Review History: &#x0D; Received 17 November 2020; Revised 12 Decembe; Accepted 1 January, Available online 15 January 2021&#x0D; UJPR follows the most transparent and toughest ‘Advanced OPEN peer review’ system. The identity of the authors and, reviewers will be known to each other. This transparent process will help to eradicate any possible malicious/purposeful interference by any person (publishing staff, reviewer, editor, author, etc) during peer review. As a result of this unique system, all reviewers will get their due recognition and respect, once their names are published in the papers. We expect that, by publishing peer review reports with published papers, will be helpful to many authors for drafting their article according to the specifications. Auhors will remove any error of their article and they will improve their article(s) according to the previous reports displayed with published article(s). The main purpose of it is ‘to improve the quality of a candidate manuscript’. Our reviewers check the ‘strength and weakness of a manuscript honestly’. There will increase in the perfection, and transparency.&#x0D; Received file: Comments of reviewer(s): &#x0D; Average Peer review marks at initial stage: 5.5/10&#x0D; Average Peer review marks at publication stage: 7.5/10&#x0D; Reviewer(s) detail:&#x0D; Dr. Idoko Alexander, Caritas University, Enugu, Nigeria, idokoalexander1@gmail.com&#x0D; Dr. Rawaa Souhil Al-Kayali, Aleppo University, Syria, rawah67@hotmail.com&#x0D; Similar Articles:&#x0D; THE ASSOCIATION BETWEEN LEVELS OF HEPCIDIN, IRON STATUS AND MICRO-INFLAMMATION MARKERS AMONG HAEMODIALYSIS &#x0D; COUMARIN ANALOGUES AS A POTENTIAL INHIBITOR OF LEISHMANIASIS: A MULTI-TARGETING PROTEIN INHIBITION APPROACH BY MOLECULAR DOCKING &#x0D; VITAMIN A, RETINOIC ACID AND TAMIBAROTENE, A FRONT TOWARD ITS ADVANCES: A REVIEW

Advances in the management of coagulopathy in acute promyelocytic leukemia

Since the introduction of all-trans retinoic acid and, more recently, arsenic trioxide into the therapy of acute promyelocytic leukemia (APL), significant improvements in patient outcomes have been achieved, and this disease has become the most curable subtype of acute myeloid leukemia. However, while primary leukemia resistance has virtually disappeared, a sizable fraction of APL patients still die before or during induction therapy. Hemorrhagic death still remains the major problem during this early phase of treatment and, to a lesser extent, deaths due to infection, differentiation syndrome and other causes. Patients with APL typically present with a range of laboratory abnormalities consistent with the diagnosis of disseminated intravascular coagulation and hyperfibrinolysis. This APL-associated coagulopathy, as a result of a dysregulation of the hemostatic system due to the imbalance between procoagulant, anticoagulant and profibrinolytic mechanisms, may show a variety of clinical manifestations, ranging from minimal bleeding or localized thrombosis to lethal or life-threatening hemorrhages or thrombotic events that sometimes occur concomitantly. Hemorrhagic events are the most common cause of death associated with APL coagulopathy, but thrombosis, a less recognized and probably underestimated life-threatening manifestation of the thrombo-hemorrhagic syndrome, is also a non-negligible cause of morbidity and mortality in patients with APL. In this article, we aim to discuss recent advances in the knowledge of pathogenesis, predictors of thrombo-hemorrhagic events, management of coagulopathy associated with APL and the controversial issues that still persist.

Early Intracranial Hemorrhages in Acute Promyelocytic Leukemia: Analysis of Neuroradiological and Clinico-Biological Parameters

Introduction In the last decades, survival outcomes tremendously improved in acute promyelocytic leukemia (APL) due to all‐trans retinoic acid (ATRA)/anthracycline‐based chemotherapy, and more recently ATRA/arsenic trioxide (ATO) combinations. Despite the excellent results, with response rates exceeding 90%, early death (ED), mainly due to hemorrhagic complications occurring during the first 30 days after diagnosis, remains an unsolved issue. The prevalence of ED is largely underestimated due to discrepancies between data deriving from clinical trials, reporting 3-10% rates, and real-life, where it ranges from 10 to 30% of cases, and still remains the dominant cause of poor outcome. The most common site of bleeding is the brain (65%), particularly in fatal cases. Many theories have been proposed to elucidate the pathophysiology of this complication, considering the coagulopathy of APL and the perturbed homeostasis as primum movens, but no specific neuroradiologic studies have been conducted so far. Moreover, there is no specific predictor for bleeding events in APL, with exception of increased white blood cell counts (WBC). Patient and Methods We retrospectively identified 38 patients with APL (23 males and 15 females with a median age of 51.83 years, range 11-82), consecutively diagnosed at our Institution between 2004 and 2019, and treated with standard ATRA-based induction (Table 1). Occurrence of ED and ICH was then correlated with clinico-biological parameters, selecting for commonly reported variables predictors of these complications. In addition, CT scans of the 5 patients who experienced an intracranial hemorrhage (ICH) were revised by an expert neuroradiologist, looking for radiographic predictors of poor outcome (Al-Mufti et al 2018). Results Approximately 13% (5/38) of our APL patients experienced an ICH during induction therapy, which was fatal in 3 cases, while the overall incidence of ED was about 10% (4/38, including 1 case of death due to a differentiation syndrome). Looking at clinico-biological variables, the only predictor of ICH was a lower albumin level at baseline (3.8 vs 4.2 gr/dl; p=0.022), as compared to patients who did not experience ICH. A high Sanz-risk score was present in 2 of 3 pts with a fatal outcome of ICH (67% of cases), together with a shorter activated partial thromboplastin time (aPTT, 0.75 vs 0.86 ratio)(Table 1). Looking at neuroradiological findings (Figure 1) the three fatal cases showed a wider ICH volume, with perilesional edema and, interestingly, a positive "swirl" sign, which is defined as a marker of ongoing extravasation of blood within a hematoma. Both cases with a favourable outcome had a cerebellar involvement, without perilesional edema, and no major involvement of basal ganglia or thalamus. None of the patients had radiological signs of arteriovenous malformations. Conclusions Our data remark the impact of lower albumin levels at baseline and of a high Sanz-risk category on ED in patients with APL. In APL cases with fatal ICH, the thrombo-hemorrhagic imbalance is more profound than in other AMLs and results in a more devastating clinical picture. Taking into account the limitations of the small sample size, we found that a wide hemorrhage volume with perilesional edema, and the presence of the "swirl" sign are characteristic of fatal cases, while arteriovenous malformations did not seem to play a predisposing role. Further studies including large series of patients are warranted to identify the characteristics and the possible strategies to ameliorate the outcome of ICH in APL. Disclosures Venditti: Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy; Astellas: Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees.

An unusual case of acute promyelocytic leukemia manifesting with pulmonary embolism

Venous thromboembolism is four fold greater in patients with cancer. 5-20% of acute promyelocytic leukemia is M3 (Hypergranular promyelocytic leukemia) variety. Hemorrhagic complications are more common than thrombosis; Coagulopathy in acute promyelocytic leukemia is a very complex disorder and is under reported. Here is a case report of a rare presentation of acute promyelocytic leukemia manifesting with pulmonary embolism.

Hypofibrinogenemia and Disseminated Intravascular Coagulation Rarely Complicate Treatment-Naive Acute Lymphoblastic Leukemia

Introduction: Hypofibrinogenemia (HF) in acute leukemia has largely been attributed to disseminated intravascular coagulation (DIC). Pathological alterations in hemostasis through various mechanisms by circulating leukemia cells have been implicated in the pathogenesis of DIC, particularly in acute promyelocytic leukemia (APL) (Tallman et al., Leuk Lymphoma, 1993). In adult acute lymphoblastic leukemia (ALL), DIC is rare at presentation, but is significantly more common during induction chemotherapy, with varying degrees of reported morbidity and mortality (Higuchi et al., Leuk Lymphoma, 2005; Sarris et al., Leuk Lymphoma, 1996). Conversely, hyperfibrinogenemia at presentation has also been observed, possibly due to increased synthesis as an acute phase reactant in the setting of inflammation (Al-Mondhiry H et al., Cancer, 1975). Discrepant reports in plasma fibrinogen levels and potentially serious hemostatic complications led us to investigate the incidence of HF and DIC among adults with untreated ALL at our center.Methods: Using a DataLine search and ICD-9 and ICD-10 codes (204.00, 204.01, 204.02, C91.00, C91.01, C91.02), all patients aged 18 years or older with a diagnosis of ALL treated at Memorial Sloan Kettering Cancer Center (MSKCC) between August 30, 1989 and April 27, 2018 were retrospectively identified. HF was defined as a fibrinogen level < 190 mg/dL. The diagnosis of DIC was made using the scoring system for overt DIC from the International Society on Thrombosis and Haemostasis (Toh et al., J Thromb Haemost, 2007). Patients who received any chemotherapy, including asparaginase and glucocorticoids, or who had a potential alternative cause of acquired HF such as sepsis or significant liver disease prior to their initial encounter were excluded. Data collection was approved by the MSKCC Institutional Review Board.Results: A total of 275 adult patients with treatment-naive ALL were identified. The median age at presentation was 44 years. The study population consisted of 122 patients with B-ALL, 81 with T-ALL and 72 with ALL of unspecified lineage. Of these, 187 had a fibrinogen level obtained prior to initiation of therapy. Eight patients (4.3%) had HF at initial assessment with a mean level of 141 mg/dL. Of these 8 patients, 5 had B-ALL, 2 had T-ALL and 1 had ALL of unspecified lineage. Four of these 8 patients required fresh frozen plasma (FFP) and/or cryoprecipitate during their hospitalization for prevention of overt coagulopathy. Three of the 4 patients who required FFP and/or cryoprecipitate went on to receive an asparaginase containing induction regimen. One patient with B-ALL met criteria for overt DIC without clinical complications. One patient with HF experienced WHO grade 2 epistaxis and hematochezia. No deaths in our study population were attributable to DIC at presentation.Discussion: Our study demonstrated that HF and DIC in treatment-naive ALL are uncommon. The rare incidence of DIC in our population is consistent with prior studies in adult ALL (Sarris et al., Blood, 1992). Our findings are also similar to those reported in patients with non-APL acute myeloid leukemia (Weltermann et al., Leukemia, 1998). In contrast, most patients with APL exhibit HF (Lou et al., Leuk Res, 2016). Proposed mechanisms of coagulopathy in APL include primary activation of fibrinolysis, as well as expression of tissue factor and cancer procoagulant by the malignant promyelocytes themselves; these mechanisms appear to be significantly less prevalent in ALL (Breen et al., Br J Haematol, 2012). DIC, when present at initial assessment, is generally subclinical and seldom leads to morbidity and mortality. Although the incidence of HF and DIC was low at diagnosis in our study population, we recommend pre-treatment screening for DIC to identify patients with overt coagulopathy at presentation and to establish baseline values, given the risk of coagulopathic complications during induction chemotherapy for ALL, particularly among patients receiving asparaginase products (Truelove et al., Leukemia, 2012). Further characterization of the ALL microenvironment in patients developing HF and DIC may elucidate mechanisms of procoagulant expression and increased fibrinolysis in this subgroup. Assessing predictors and clinical implications of baseline HF in adults with ALL in a larger series may help to guide early supportive management. DisclosuresGeyer:Dava Oncology: Honoraria. Park:Adaptive Biotechnologies: Consultancy; AstraZeneca: Consultancy; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Shire: Consultancy; Kite Pharma: Consultancy; Novartis: Consultancy; Pfizer: Consultancy; Juno Therapeutics: Consultancy, Research Funding. Mantha:Janssen Pharmaceuticals: Research Funding; Heidell, Pittoni, Murphy & Bach, LLP: Consultancy; GLG: Consultancy. Tallman:Orsenix: Other: Advisory board; Daiichi-Sankyo: Other: Advisory board; Cellerant: Research Funding; BioSight: Other: Advisory board; AROG: Research Funding; AbbVie: Research Funding; ADC Therapeutics: Research Funding.

Mechanisms and management of coagulopathy in acute promyelocytic leukemia

Acute promyelocytic leukemia (APL) is a subtype of leukemia which is associated with unique and distinctive coagulopathy. In the absence of treatment it is rapidly fatal and even after initiation of therapy the major cause of early mortality is related to hemorrhagic complications. The coagulopathy can be exacerbated with the start of treatment. In the absence of early hemorrhage related deaths the probability of cure exceeds 90% in low and intermediate risk patients and 80% even in high risk patients, highlighting the importance of understanding the pathophysiology of this complication and instituting prompt and appropriate management strategies. The coagulopathy in APL is complex and results from a combination of thrombocytopenia, disseminated intravascular coagulation and hyperfibronlysis. Recently the effect of all-trans retinioc acid (ATRA) induced ETosis on exacerbating coagulopathy in the first few days after starting therapy with this agent raises the potential for potentially novel strategies to reduce the risk of hemorrhage. Currently management is mainly related to rapid initiation of therapy with ATRA along with appropriate and adequate replacement of blood products to correct the coagulopathy. There is limited role for the use of low dose anti-coagulants and anti-fibrinolytic agents in the initial management of this disease. There is limited data on the use of rFVIIa or the use of global tests of hemostasis in the management of this condition.

The impact of oral arsenic and all-trans-retinoic acid on coagulopathy in acute promyelocytic leukemia

The aim of our study was to evaluate the impact of oral arsenic (the realgar-indigo naturalis formula, RIF) and all-trans retinoic acid (ATRA) on coagulopathy in acute promyelocytic leukemia (APL) compared with intravenous arsenic trioxide (ATO) and ATRA during induction. Mitoxantrone was added to all the patients at a dose of 1.4mg/m2 per day for 5–7 days. D-dimer levels, prothrombin time (PT), fibrinogen (Fbg) levels and the platelet count were comparably analyzed among 83 newly diagnosed APL patients treated with RIF (n=45) or with ATO (n=38). Since induction therapy with RIF and ATRA, the median levels of Fbg, PT and platelets were recovered to the normal range within 4days, 10days and 28days, respectively. The last day of platelet and plasma transfusion was day 12 (range: 0–24 days) and day 3 (range: 0–27 days), respectively. Among the 42 patients with a disseminated intravascular coagulation (DIC) score=4, the consumption of transfused platelets was less in the RIF group than that in the ATO group (P=0.037). In the 17 patients with a DIC score <4, prompt recovery of Fbg levels (P=0.028) was observed in the RIF group compared with that in the ATO group (P=0.401). RIF and ATO showed similar effects on the recovery of coagulopathy in APL patients. RIF had a potential beneficial effect in accelerating the recovery of thrombocytopenia and hypofibrinogenemia for subclinical DIC patients.

Promyelocytic extracellular chromatin exacerbates coagulation and fibrinolysis in acute promyelocytic leukemia

AbstractDespite routine treatment of unselected acute promyelocytic leukemia (APL) with all-trans-retinoic acid (ATRA), early death because of hemorrhage remains unacceptably common, and the mechanism underlying this complication remains elusive. We have recently demonstrated that APL cells undergo a novel cell death program, termed ETosis, which involves release of extracellular chromatin. However, the role of promyelocytic extracellular chromatin in APL-associated coagulation remains unclear. Our objectives were to identify the novel role of ATRA-promoted extracellular chromatin in inducing a hypercoagulable and hyperfibrinolytic state in APL and to evaluate its interaction with fibrin and endothelial cells (ECs). Results from a series of coagulation assays have shown that promyelocytic extracellular chromatin increases thrombin and plasmin generation, causes a shortening of plasma clotting time of APL cells, and increases fibrin formation. DNase I but not anti-tissue factor antibody could inhibit these effects. Immunofluorescence staining showed that promyelocytic extracellular chromatin and phosphatidylserine on APL cells provide platforms for fibrin deposition and render clots more resistant to fibrinolysis. Additionally, coincubation assays revealed that promyelocytic extracellular chromatin is cytotoxic to ECs, converting them to a procoagulant phenotype. This cytotoxity was blocked by DNase I by 20% or activated protein C by 31%. Our current results thus delineate the pathogenic role of promyelocytic extracellular chromatin in APL coagulopathy. Furthermore, the remaining coagulation disturbance in high-risk APL patients after ATRA administration may be treatable by intrinsic pathway inhibition via accelerating extracellular chromatin degradation.

FLT3 Mutations in Pediatric Acute Promyelocytic Leukemia; A Report from the Children's Oncology Group AAML0631 Trial

BackgroundSurvival for acute promyelocytic leukemia (APL) has improved dramatically in recent decades through use of targeted therapy with all-trans retinoic acid (ATRA) and arsenic trioxide (ATO). Early deaths, principally due to complications of APL coagulopathy, remain a significant barrier to cure. White blood cell (WBC) count at diagnosis is a well validated risk marker for early death and relapse. While APL is characterized by t(15;17), mutations of the FLT3 gene are prevalent. Prior reports in both adult and pediatric APL have demonstrated variable impact of FLT3 mutation status on outcomes. In a pediatric cohort from the Intergroup Study CALGB 9710, our group previously found an association of FLT3 mutations with early death. The more recent Children's Oncology Group (COG) APL trial AAML0631 included addition of two courses of arsenic trioxide (ATO) during consolidation. Relapse rate on this study was low, but survival, particularly for the high risk patients, was impacted by early deaths. In a prior analysis we reported that a significant proportion of patients on AAML0631 experienced clinically significant bleeding and clotting events. Here we report our analysis of the prevalence of FLT3 mutations and their association with outcomes for pediatric APL patients treated on the AAML0631 study.MethodsDiagnostic bone marrow samples were collected from consenting patients with newly diagnosed APL enrolling onto COG AAML0631. FLT3 mutation testing of these bone marrow samples included evaluation for internal tandem duplications (FLT3/ITD) of the juxta-membrane domain and point mutations of the activating loop (FLT3/ALM) of the tyrosine kinase domain. FLT3 mutation status was correlated with clinically significant bleeding/clotting, early death during induction and relapse. Significance of correlation was determined using the chi-square test and relapse risk for patients who achieved CR by end of Induction 1 was calculated using cumulative incidence.Eligible patients were age ≥ 2 years and < 22 years and required confirmation of PML-RARA by PCR testing. Risk group was defined by WBC at diagnosis: standard risk (SR) WBC < 10,000 and high risk (HR) WBC ≥ 10,000. Treatment regimen has been previously reported [Kutny et al, ASH Abstract, 2015].ResultsThere were 101 evaluable patients on AAML0631 including 35 HR APL and 66 SR APL. A total of 94 patients had specimens available for FLT3 mutation testing. Patients with unknown FLT3 status (3 HR and 4 SR) were excluded from the following analyses. FLT3 mutations were present in 43% (N=40) including 35% (N=33) with FLT3/ITD, 4% (N=4) with FLT3/ALM, and 3% (N=3) with both FLT3/ITD and FLT3/ALM. The median WBC was 24,450 for FLT3 mutant APL compared to 7,130 for FLT3 wild type, and the rate of FLT3 mutation was significantly higher in HR APL compared to SR APL (66% vs. 31%, P=0.001). Four patients died during induction (all HR APL with WBC of 16200-173800), but early death did not correlate with FLT3 mutation (2/4 patients had FLT3 mutations). We analyzed association of FLT3 mutations with prevalence of clinically significant (Grade III-IV) bleeding or clotting events during induction. There was no difference in prevalence of coagulopathy events (N= 19) between those with and without FLT3 mutations (18% vs. 22%, P=0.57).While OS and EFS were not significantly different, the relapse rate from end of Consolidation 1 was significantly higher at 6% (N=3) in FLT3 mutant patients versus 0% in FLT3 wild-type (P=0.039) [Figure 1].ConclusionsFLT3 mutations are highly prevalent (43%) in pediatric APL. The COG AAML0631 study represents the second largest group of pediatric APL patients with outcome data treated on a cooperative group trial and to our knowledge the current report is the largest group of pediatric APL patients evaluated for outcome based on FLT3 mutation status. This study failed to show an association of FLT3 mutations with early death or bleeding/clotting events in induction. With ATO consolidation, the relapse rate was low on this trial. However, it is of great interest that the relapse rate following ATO consolidation was significantly higher in FLT3 mutant patients. Evaluation of a larger (international) cohort of pediatric APL patients is warranted to further define of role of FLT3 in relapse risk, but the current results give further support to consideration of integrating FLT3 inhibitors into relapsed/refractory APL treatment. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Whatʼs new in the pathogenesis of the coagulopathy in acute promyelocytic leukemia?

Acute promyelocytic leukemia (APL) is associated with a complex coagulopathy. In spite of substantial recent improvements in treatment regimens, hemorrhagic death remains the main cause of induction failure. In this review, we delineate recent understanding of the pathophysiology and management of the hemorrhagic diathesis of APL. Laboratory and clinical data suggest that the malignant leukocytes mediate the hemorrhagic diathesis associated with APL through multiple mechanisms which lead to a combination of consumptive coagulopathy and primary hyperfibrinolysis. Exposure of tissue factor and Annexin II by the leukemic blasts is the main determinants of these processes. Promyelocyte-derived microparticles have recently been implicated in the coagulopathy as well. Total white cell count and platelet count have emerged as good general predictors of hemorrhagic death, along with the different routine hemostatic parameters. Prompt treatment with all-trans retinoic acid, with or without arsenic trioxide, is the most important step in preventing bleeding complications. Repletion of coagulation factors and platelets with blood products remains the mainstay of supportive treatment, whereas the role of recombinant soluble thrombomodulin is currently being investigated. The coagulopathy of APL is multifactorial, with both disseminated intravascular coagulation and primary hyperfibrinolysis mediated largely by the malignant leukocytes.

Tissue Factor Pathway Inhibitor (TFPI) May be Another Important Factor in the Coagulopathy in Acute Promyelocytic Leukemia (APL)

The characteristic coagulopathy in Acute Promyelocytic Leukemia (APL) is unique among the leukemias and thrombotic and bleeding complications remain the major causes of early deaths. APL promyelocytes express tissue factor (TF) which, after activation by phospholipids, forms a complex with factor VII and converts factor X to activated factor X. TF plays a central role in the pathogenesis of APL coagulopathy, for the procoagulant activity of lysates from freshly isolated APL cells is mainly attributed to TF. During periods of increased apoptosis of promyelocytes, the procoagulant activity is correspondingly intensified. In addition, APL cells may also induce TF procoagulant activity of endothelial cells through their secretion of IL-1b. The activity of the complex FVIIa-TF can be inhibited by the tissue factor pathway inhibitor (TFPI), but its role in the APL-associated coagulopathy remains unknown. This study aimed to determine the time-course of TFPI levels in patients with APL at diagnosis and during the first two-weeks of treatment with ATRA and anthracyclines. Twenty patients with de novo APL (12 males, age ranging from 17 to 72 years) and 20 healthy blood donors (age and sex matched) were included in this study. Peripheral blood (PB) samples were collected at diagnosis, and at D3, D7 and D15 of the induction therapy with ATRA and daunorubicin. The following plasma concentrations were determined: Thrombin-Antithrombin complex (TAT) (Enzygnost¨ TAT micro, Dade Behring/Siemens, IL, EUA), total TFPI (Asserachrom¨ total TFPI, Diagnostica Stago, Frana) and free TFPI (Asserachrom¨ free TFPI, Diagnostica Stago, Frana). The plasma concentrations of TAT were significantly higher in APL patients at presentation, D3 and D7 compared to controls, suggesting that the pathologic activation of coagulation was reversed by D15 of treatment Figure 1). The concentrations of free-TFPI were significantly higher in APL samples compared to controls in all time-points (Figure 2). In contrast, the concentrations of the truncated form of TFPI (estimated by subtracting the free-TFPI values from total-TFPI concentration) did not vary significantly between the groups. Since, free-TFPI (intact form) has been proven to present higher capacity to inhibit the activated factor X and VIIa-TF proteases than truncated-TFPI (bound to lipoprotein), we hypothesized that our findings confirm the relevance of free-TFPI in APL coagulopathy. To test whether the administration of rh-TFPI may revert aberrant activation of coagulation in APL patients, we developed a murine model using the IV infusion of NB4 cells. Similarly to the reported in APL patients, we observed an increase on TAT levels in mice injected with 5x106 NB4 cells [(60.7 ng/ml (24 - 116 ng/ml) vs 11.5 ng/ml (2.2 - 27.8 ng/ml), p<0.05)]. In contrast, murine TFPI levels decreased 30 minutes after the infusion of NB4 cells [30 ng/ml (5.8 - 217 ng/ml) vs 202 ng/ml (46 - 354 ng/ml)], suggesting that it was rapidly consumed. Unfortunately, it was not possible to distinguish between truncated and intact TFPI forms in mice. Intravenous infusion of recombinant human (rh) TFPI at the dose of 10mg/Kg significantly decreased the TAT levels (Figure 3) and increased TFPI levels in the murine model. In conclusion, our data on clinical samples and pre-clinical data using a murine model suggest that treatment with rh-TFPI may be a strategy to counteract APL-associated coagulopathy. [Display omitted] [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Cerebral venous and sinus thrombosis in a patient with acute promyelocytic leukemia during all-trans retinoic acid induction treatment

Cerebral venous and sinus thrombosis is a rare cerebrovascular disorder, which seldom represents a complication of acute promyelocytic leukemia (APL). As a part of the coagulopathy of APL, thrombosis is a less recognized and underestimated life-threatening manifestation and is overshadowed by the more obvious bleeding complications. Here, we described a 28-year-old woman with APL who developed massive thrombosis of the cerebral sinuses while on induction treatment with all-trans retinoic acid. On the basis of this report, the potential pathogenic mechanisms and the diagnosis based on magnetic resonance imaging (MRI) combined with magnetic resonance venogram (MRV) are discussed. Early anticoagulant therapy contributed to the progressive dissolution of the thrombosis, as documented by MRI, with the complete disappearance of neurological signs without sequelae. Given the increasing recognition of thromboembolic events in APL, the use of prophylactic anticoagulation during induction therapy may need to be redefined.

Disseminated intravascular coagulation in acute promyelocytic leukaemia and its impact on the induction failure: a single centre study.

Life-threatening coagulopathy associated with acute promyelocytic leukemia (APL) has been the defining clinical characteristic and is an important risk factor for fatal haemorrhage and early death. Pathogenesis of coagulopathy in APL is complex and mainly includes disseminated intravascular coagulation (DIC). The study was done to see the status of DIC and its impact on the outcome of APL in our setting. Among the total 60 patients, induction mortality rate was 30% and remission rate was 70%. The main cause of induction mortality was bleeding that accounts for 66.7% of mortality. DIC was present among 32 out of 60 patients (53.33%). Induction mortality has significant relationship to DIC as the induction mortality rate is 47% in patients with DIC and 11% in patient without DIC (P value 0.0009). Induction motality rate in low, intermediate and high risk group is 6.70%, 24% and 58% respectively (p value < 0.0001). Finally, risk group subclassification revealed presence of DIC in high risk group has the highest early mortality rate.

PML-RARa Regulated Vesiculation Of Acute Promyelocytic Leukemia Cells

BackgroundIn acute promyelocytic leukemia (APL), a subtype of acute myeloid leukemia (AML), severe bleeding tendency coincides with upregulation of tissue factor (TF), fibrinogenopenia and disseminated intravascular coagulation (DIC). In APL, the reciprocal translocation between chromosomes 15 and 17 results in the oncogenic fusion between promelocytic leukemia gene (PML) and retinoic acid receptor alpha (RARa). Targeting the activity of PML-RARa using targeted therapy combined with chemotherapy produces a high rate of complete remission, and reversal of tissue factor (TF)-related coagulopathy. Oncogenic transformation, therapeutic responses, coagulopathy and interaction of malignant cells with their vascular microenvironment has been linked, at least in part, to the emission and uptake of extracellular vesicles (EVs, including ectosomes/microparticles and exosomes). Little is known about these processes in APL. PurposeIn this study, we explored the role of oncogenic PML-RARa in vesiculation of APL cells and the resulting changes in their procoagulant/angiogenic properties. We hypothesized that severe coagulopathy in APL may propagate to stromal (endothelial) cells via the exchange of TF-containing vesicles. MethodEV emission by NB4 cells derived from an APL patient with t(15;17) was measured by Nanoparticle Tracking Analysis (NTA). Ectosomes and exosomes were separated by ultracentrifucation, and the content of PML-RARa and TF in these EVs were studied at both RNA (RT-PCR) and protein and activity levels (Western, ELISA, TF-PCA). Transfer of EVs between NB4 cells and endothelium (HUVEC) was tested using membrane labeling with the PKH26 dye and flow cytometry. PML-RARa and TF in transfer to HUVECs were studied at both in RNA level and protein level, and bioassays (invasion, angiogenesis) were used to assess the consequences. ResultsPML-RARa controls cellular vesiculation as all-trans retinoic acid (ATRA) alters emission of EVs by NB4 cells leading to preponderance of exosomes. These EVs contain both PML-RARa and TF transcripts, but only TF protein is detected in the EV cargo and transferred to endothelial cells. NB4-derived EVs render endothelial cells TF-positive and procoagulant and changes their angiogenic properties. Surprisingly, the migratory phenotype of endothelial cells is inhibited by co-culture with leukemic cells and by exposure to leukemic EVs. The nature of these effects is being studied in vitro and in vivo. ConclusionNB4 cells shed EVs as a function of their differentiation and oncogenic status. Unlike oncogenic receptors in cancer cells, PML-RARa protein is not detected in leukemic EVs, but exerts an indirect effect on their production and cargo. APL-derived EVs may render endothelial cells procoagulant. In spite of the pro-angiogenic phenotype of APL cells, their EVs inhibit migration of endothelial cells. The significance of EV interactions between leukemic and non-leukemic cells is being investigated. Disclosures:No relevant conflicts of interest to declare.