Chronically Elevated Interleukin-6 Disturbs the Coagulation Cascade in Mice

Abstract Introduction: Pro-inflammatory cytokines play an essential role as activators of the hemostatic system and in the regulation of physiological antithrombotic mechanisms. Interleukin-6 (IL-6) influences platelet production and platelet activation. It was associated with accelerated clotting and intravascular coagulation in tissue factor (TF)-driven murine thrombosis models. However, the precise role of myeloid cell-derived IL-6 on thrombosis formation and the hemostatic system is still unknown. Methods and Results: To better understand the role of IL-6 in thrombosis and the hemostatic system, we developed a new mouse strain with Cre-recombinase driven constitutive IL-6 expression specifically in myeloid cells (LysM-IL-6 OE, Control mice: IL-6 OE). LysM-IL-6 OE mice had a prolonged tail bleeding time and lacked venous thrombus formation induced by inferior vena cava (IVC) stenosis. There were no differences in D-Dimer levels in LysM-IL-6 OE mice neither on baseline level nor after IVC ligation. However, we found unstoppable post-operative bleedings in LysM-IL-6 OE. They showed a prolonged aPTT, a significantly increased INR and a prolonged thrombin converting time. The Factor V and IX expression were reduced, but von Willebrand factor, antithrombin and fibrinogen expression were up-regulated and could not explain the missing thrombus formation. We found significantly elevated erythrocyte sedimentation in line with erythrocytes aggregates, which seemed to be mediated by IL-6 and α2M. Most importantly, hepatic levels of thrombin inhibitor α2 macroglobulin (α2M) mRNA and protein were increased in LysM-IL-6 OE/+ mice compared to control mice. In parallel, Platelet erythrocyte interaction seemed to be essential in the development of the bleeding phenotype. Conclusions: These findings show the role of chronically elevated IL-6 in driving the accumulation of A2m on the surface of erythrocytes, thereby mediating a phenotype of increased bleeding complications. This work was supported by the DFG KA4035/1-1 and by the German Ministry for Education and Research (BMBF 01EO1503) Disclosures Lämmle: Takeda: Membership on an entity's Board of Directors or advisory committees; Ablynx: Membership on an entity's Board of Directors or advisory committees, Other: Travel Support, Speakers Bureau; Baxter: Other: Travel Support, Speakers Bureau; Alexion: Other: Travel Support, Speakers Bureau; Siemens: Other: Travel Support, Speakers Bureau; Bayer: Other: Travel Support, Speakers Bureau; Roche: Other: Travel Support, Speakers Bureau; Sanofi: Other: Travel Support, Speakers Bureau. Ruf: ARCA bioscience: Consultancy, Patents & Royalties; ICONIC Therapeutics: Consultancy; MeruVasimmune: Current holder of individual stocks in a privately-held company.

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The role of oral anticoagulant therapy in patients with acute coronary syndrome

Therapeutic Advances in Hematology ◽

10.1177/2040620717733691 ◽

2017 ◽

Vol 8 (12) ◽

pp. 353-366 ◽

Cited By ~ 12

Author(s):

Jae Youn Moon ◽

Deepa Nagaraju ◽

Francesco Franchi ◽

Fabiana Rollini ◽

Dominick J. Angiolillo

Keyword(s):

Acute Coronary Syndrome ◽

Anticoagulant Therapy ◽

Oral Anticoagulant ◽

Thrombus Formation ◽

Oral Anticoagulant Therapy ◽

Ease Of Use ◽

Coagulation Cascade ◽

Bleeding Complications ◽

Coronary Syndrome

Dual antiplatelet therapy (DAPT) with aspirin and a P2Y12 receptor antagonist represents the current standard of care to prevent atherothrombotic recurrences in patients with acute coronary syndrome (ACS). However, despite the use of DAPT, the recurrence rate of cardiovascular ischemic events still remains high. This persistent risk may be in part attributed to the sustained activation of the coagulation cascade leading to generation of thrombin, which may continue to play a key role in thrombus formation. The use of vitamin K antagonists (VKAs) as a strategy to reduce atherothrombotic recurrences after an ACS has been previously tested, leading to overall unfavorable outcomes due to the high risk of bleeding complications. The recent introduction of non-VKA oral anticoagulants (NOACs), characterized by a better safety profile and ease of use compared with VKA, has led to a reappraisal of the use of oral anticoagulant therapy for secondary prevention in ACS patients. The present article provides an overview of the rationale and prognostic role of oral anticoagulant therapy in ACS patients as well as recent updated clinical data, in particular with NOACs, in the field and future perspectives on this topic.

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Prognostic Role of IL-6 in POEMS Syndrome

Blood ◽

10.1182/blood-2021-149792 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2700-2700

Author(s):

Maryam Omar ◽

Rahma M Warsame ◽

Morie A. Gertz ◽

Francis K. Buadi ◽

Ronald Go ◽

...

Keyword(s):

Overall Survival ◽

Board Of Directors ◽

Interleukin 6 ◽

Research Funding ◽

Statistical Significance ◽

Poems Syndrome ◽

Reference Ranges ◽

Time To Progression ◽

Advisory Committees

Abstract INTRODUCTION POEMS syndrome is a paraneoplastic disorder that results in multi-organ disease, and Vascular endothelial growth factor (VEGF) is associated with disease activity. There is evidence that the cytokine Interleukin- 6 (IL-6) stimulates VEGF production in several cancer cell lines. It has also been suggested that patients with POEMS syndrome experience symptoms improvement when IL-6 levels are low. Our study aims to understand the role of IL-6 and whether it is a contributor to disease activity in POEMS syndrome. METHODS We conducted a retrospective study of patients (pts) seen at Mayo Clinic within 1 year of diagnosis for POEMS syndrome with an IL-6 level within 3 months of diagnosis between 01/01/2011-05/31/2021 were included. Our database comprised 358 pts,7 were excluded because did not have official diagnosis of POEMS syndrome. Two hundred and ninety-nine pts did not have IL-6 testing within 90 days of diagnosis and were excluded. Clinical data was abstracted from our electronic medical record. Descriptive statistics were used for IL-6 levels. The majority of IL-6 laboratory testing for pts utilized the same reference range. A minority of pts had alternate reference ranges; thus, all IL-6 values and reference ranges were fold-corrected to have a unified reference to determine elevation in IL-6. High IL-6 was considered any value above the normal range, also analyzed pts in quartiles and deciles. Hematologic, VEGF and PET response criteria were utilized to group pts into responders or non-responders. Time to progression (TTP) and overall survival (OS) was calculated using the Kaplan Meier method. Differences between curves were by log rank. Statistical analysis was conducted via JMP software package (SAS, Cary, NC, USA). Univariate analysis was done to determine the prognostic value of IL-6. Statistical significance is considered with p-values <0.05. RESULTS 52 patients from among 352 met criteria and were included for analysis. Twenty-one patients (40%) had elevated IL-6 levels (IQR 5.67, 22.6) at the time of diagnosis. The fold elevation about normal was typically not very high, with a median of 3.1-fold elevation (IQR 1.3, 4.4; and range 1.0, 8.3). The percentage of males in the high IL-6 versus the normal Il-6 group were 90% versus 65%, p=0.03. Compared to normal IL-6, those with elevated IL-6 values had more instances of hepatomegaly (43% vs 16%, p=0.03), ascites (28% vs 6%, p=0 .04), abnormal DLCO (26% vs 3%, p=0.03), mixed bone lesions (57% vs 29%, p= 0.04), and lower serum albumin (range 2.3-3.5 vs 2.5-4.5 g/dL, p=0.0008).There was a trend for lower VEGF values among the high IL-6 group, but this did not meet statistical significance (median 250 vs 438 pg/mL). There was no significant difference in the time to progression between pts with high versus normal IL-6 levels (HR 1.36; p=0.456). Overall survival was significantly longer in pts with normal IL-6 compared to pts with high IL-6 levels (Median OS 67.4 vs 47.8 months, p=0.02). Forty-two pts were evaluated for VEGF response after treatment; there was a significant improvement in time to progression (85.5 vs 14.9 months, p=0.03) in pts who were VEGF responders. Among pts evaluable for a hematologic response (n=15), time to progression in pts with baseline high IL6 was longer in hematologic responders compared to non-responders (p=0.02). Conclusion Although OS was longer for pts with normal IL-6, elevated IL-6 at the time of diagnosis does not prevent pts from achieving disease remission. The study demonstrates that response to treatment rather than interleukin-6 levels in pts with POEMS syndrome is more prognostic. This study is an important initial step into understanding the utility of IL-6 in POEMS syndrome. Figure 1 Figure 1. Disclosures Gertz: Akcea Therapeutics, Alnylam Pharmaceuticals Inc, Prothena: Consultancy; Aurora Biopharma: Other: Stock option; Ionis Pharmaceuticals: Other: Advisory Board; AbbVie Inc, Celgene Corporation: Other: Data Safetly & Monitoring; Akcea Therapeutics, Ambry Genetics, Amgen Inc, Celgene Corporation, Janssen Biotech Inc, Karyopharm Therapeutics, Pfizer Inc (to Institution), Sanofi Genzyme: Honoraria. Kumar: Oncopeptides: Consultancy; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Beigene: Consultancy; Merck: Research Funding; Antengene: Consultancy, Honoraria; Tenebio: Research Funding; Novartis: Research Funding; Roche-Genentech: Consultancy, Research Funding; Carsgen: Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bluebird Bio: Consultancy; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Research Funding; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Consultancy, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding. Dingli: GSK: Consultancy; Novartis: Research Funding; Janssen: Consultancy; Sanofi: Consultancy; Apellis: Consultancy; Alexion: Consultancy. Lin: Janssen: Consultancy, Research Funding; Merck: Research Funding; Novartis: Consultancy; Legend: Consultancy; Juno: Consultancy; Bluebird Bio: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Vineti: Consultancy; Kite, a Gilead Company: Consultancy, Research Funding; Sorrento: Consultancy; Gamida Cell: Consultancy; Takeda: Research Funding. Kapoor: Karyopharm: Consultancy; Cellectar: Consultancy; BeiGene: Consultancy; Pharmacyclics: Consultancy; Sanofi: Consultancy; Amgen: Research Funding; Ichnos Sciences: Research Funding; Regeneron Pharmaceuticals: Research Funding; Glaxo SmithKline: Research Funding; Karyopharm: Research Funding; Sanofi: Research Funding; Takeda: Research Funding; AbbVie: Research Funding. Dispenzieri: Pfizer: Research Funding; Sorrento Therapeutics: Consultancy; Oncopeptides: Consultancy; Alnylam: Research Funding; Takeda: Research Funding; Janssen: Consultancy, Research Funding.

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Activated FVIII Released from FVIII/VWF Complex Facilitates Thrombus Development Under High Shear Flow Condition

Blood ◽

10.1182/blood.v128.22.83.83 ◽

2016 ◽

Vol 128 (22) ◽

pp. 83-83

Author(s):

Hiroaki Yaoi ◽

Yasuaki Shida ◽

Takehisa Kitazawa ◽

Kunihiro Hattori ◽

Midori Shima ◽

...

Keyword(s):

Board Of Directors ◽

Research Funding ◽

Thrombus Formation ◽

High Shear ◽

Flow Conditions ◽

Advisory Committees ◽

Platelet Surface ◽

Chugai Pharmaceutical ◽

Low Shear

Abstract Background : Factor VIII (FVIII) is protected by binding to its carrier protein, von Willebrand factor (VWF) in the circulation. VWF contributes to hemostasis particularly under high shear flow condition by extending its multimeric configuration. By contrast, coagulation factors including FVIII are thought to play a dominant role under low shear. Since FVIII binds to VWF unlike other coagulation factors, FVIII may contribute to thrombus formation even under high shear conditions. FVIII has to be released from VWF and activated to exert its coagulation function. However, the role of interaction between FVIII and VWF on hemostasis under flow conditions needs to be explored in addition to the protective function of VWF. Aims: To analyze the mechanism and role of FVIII and VWF interaction on hemostasis under flow conditions. Methods: Whole blood samples were obtained from patients with type 2A von Willebrand disease (VWD). FVIII, VWF, FVIII/VWF (final concentration; f.c. 1U/mL), FVIII plus ESH8 (f.c. 1 µM), and emicizumab (ACE910; f.c. 100 µg/mL) were added to the blood followed by perfusion into the collagen-coated flow chamber under controlled high (2,500s-1) and low (50s-1) shear conditions. ESH8 is an anti-FVIII antibody that interrupts the release of FVIII from VWF, and emicizumab is a bispecific antibody mimicking activated FVIII (FVIIIa). The concentration of emicizumab was chosen, as it was the highest dose in clinical study. After the perfusion, formed thrombus was fixed and immunostaining was performed to visualize platelets, VWF, thrombin, and FVIII. Thrombi were observed by using confocal laser scanning microscopy and the obtained images were analyzed by Image Pro Premier 3D. Initial thrombus formation was measured as surface coverage (SC) and thrombus development was measured as thrombus volume (TV). Results: SC and TV were impaired in VWD particularly under high shear (high: SC 8.3%, TV 3.9x103μm3, low: SC 2.7%, TV 2.9x103μm3). Addition of FVIII/VWF improved thrombus formation to normal level under both shear conditions (high: SC 37.3%, TV 2.4x104μm3, low SC 4.3%, TV 4.1x103μm3). Addition of VWF improved SC and TV under both shear conditions (high: 40.9%, 1.7x104μm3, low: SC 3.0%, TV 3.3x103μm3), suggesting that the VWF function was crucial under both shear. By contrast, addition of FVIII alone did not improve SC and TV under both shears (high: SC 9.9%, TV 5.0x103μm3, low: SC 2.4%, TV 3.1x103μm3). Since FVIII/VWF had a greater effect on TV than VWF alone, FVIII enhanced thrombus development under high shears, however, this effect required the presence of VWF. FVIII immunostaining demonstrated the binding of FVIII on platelet surface in the FVIII-added experiment and, therefore, FVIII binding alone was not sufficient to initiate coagulation. Since FVIII may need to be activated on the platelet surface in a timely fashion, it was speculated that the optimal delivery from VWF and activation of FVIII on platelets was required for hemostasis. To analyze the role of FVIII and VWF interaction on hemostasis under flow condition, ESH8 was used to modify FVIII/VWF binding by blocking FVIII release. Treatment of FVIII/VWF with ESH8 did not change SC but slightly impaired TV under high shear (high: SC 41.8%, TV 1.8 x104μm3). Thus, the presence of FVIII at thrombus was not enough and the release of FVIII from VWF and presumably activation of FVIII on the platelet surface might be essential for thrombus formation under high shear. Under low shear, both SC and TV were interrupted by ESH8 (SC 2.6%, TV 3.1x103μm3), suggesting the important role of FVIII and coagulation under low shear. Lastly, to analyze the impact of activated FVIII on thrombus formation under flow conditions, emicizumab was evaluated in this system. In contrast to FVIII, emicizumab alone improved thrombus formation (high: SC 26.0%, TV 1.1 x104μm3, low: SC 3.5%, TV 4.2 x103μm3). The rate of improvement in SC and TV was better under low shear than high shear, which implicated the coagulative role of emicizumab. Even under high shear, however, emicizumab enhanced thrombus formation. Therefore, the role of emicizumab in substituting for the activated form of FVIII might be able to rapidly initiate coagulation even under high shear. Conclusion: VWF delivers and releases FVIII in a timely fashion on platelet surface. FVIII has to become active on platelets for hemostasis and FVIIIa can accelerate thrombus formation even under high shear condition. Disclosures Yaoi: Chugai Pharmaceutical Co., Ltd.: Research Funding. Shida:Chugai Pharmaceutical Co., Ltd.: Research Funding. Kitazawa:Chugai Pharmaceutical Co.: Employment, Equity Ownership, Patents & Royalties; Sysmex Corporation: Patents & Royalties. Hattori:Chugai Pharmaceutical Co.: Employment, Equity Ownership, Patents & Royalties. Shima:Sysmex Corporation: Patents & Royalties, Research Funding; Chugai Pharmaceutical Co., Ltd.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding; F. Hoffmann-La Roche Ltd.: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Nogami:F. Hoffmann-La Roche Ltd.: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sysmex Corporation: Patents & Royalties, Research Funding; Chugai Pharmaceutical Co., Ltd.: Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding.

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Abstract 35: Plasma Fibronectin is a Vital Supportive Factor in Hemostasis and a Unique Self-Limiting Regulator in Thrombosis

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.34.suppl_1.35 ◽

2014 ◽

Vol 34 (suppl_1) ◽

Author(s):

Yiming Wang ◽

Adili Reheman ◽

Christopher Spring ◽

Jalil Kalantari ◽

Alexandra Marshall ◽

...

Keyword(s):

Therapeutic Potential ◽

Thrombus Formation ◽

Vena Cava ◽

Bleeding Complications ◽

Plasma Fibronectin ◽

Vessel Occlusion ◽

Abdominal Bleeding ◽

Inferior Vena Cava Occlusion ◽

Fibrin Network

The role of plasma fibronectin (pFn) in thrombosis/hemostasis is controversial. We previously demonstrated that pFn supports thrombosis in pFn-/- mice. Interestingly, depletion of pFn in fibrinogen (Fg)/VWF-/- mice resulted in enhanced thrombus formation, revealing a functional switch of pFn in the absence of Fg/VWF. However, the mechanism controlling this switch is unknown. Furthermore, the hemostatic function of pFn is unclear. To address these questions, we established Fg/pFn-/- and VWF/pFn-/- mice in addition to triple deficient mice (Fg/VWF/pFn-/-, TKO). TKO mice had significantly higher mortality (23.1% vs. 9.1%) and longer tail bleeding time than their pFn+ Fg/VWF-/- littermates. Autopsy revealed severe subcutaneous or abdominal bleeding. pFn depletion in Fg-/- mice also increased mortality from 5.5% to 25.6% and prolonged bleeding times; the latter effect was reversed with pFn infusion. Using intravital microscopy, we found that pFn deposition at the site of injury preceded significant platelet accumulation, suggesting that pFn is an efficient and rapid hemostatic factor. Based on thromboelastography, fibrin clots from pFn+ mice were significantly stronger than those from pFn-/- mice. Consistent with this, inferior vena cava occlusion was significantly delayed in pFn-/- mice. Under confocal microscopy, fluorescently-labeled pFn was actively recruited into the fibrin network in mouse and human plasma. Under electron microscopy, physiological levels of pFn doubled the diameter of fibrin fibers, suggesting that pFn contributes to lateral aggregation of fibrin protofibrils. Interestingly, pFn enhances platelet aggregation when linked with fibrin and inhibits aggregation when fibrin is absent. pFn therefore gradually switches from supporting hemostasis to inhibiting thrombosis and vessel occlusion following the fibrin gradient that decreases farther from the injured endothelium. Our data established pFn as an important supportive factor in hemostasis and a unique self-limiting regulator of thrombosis, suggesting the therapeutic potential of pFn transfusion in controlling bleeding complications, particularly for those receiving antithrombotic therapy for heart attack and stroke.

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Role of Remission Status and Prior Transplant in Optimizing Survival Outcomes Following Allogeneic Hematopoietic Stem Transplantation (HSCT) in Patients Who Received Inotuzumab Ozogamicin (INO) for Relapsed / Refractory (R/R) Acute Lymphoblastic Leukemia (ALL)

Blood ◽

10.1182/blood.v130.suppl_1.886.886 ◽

2017 ◽

Vol 130 (Suppl_1) ◽

pp. 886-886

Author(s):

Partow Kebriaei ◽

Matthias Stelljes ◽

Daniel J. DeAngelo ◽

Nicola Goekbuget ◽

Hagop M. Kantarjian ◽

...

Keyword(s):

Board Of Directors ◽

Survival Probability ◽

Research Funding ◽

Employment Equity ◽

Inotuzumab Ozogamicin ◽

Advisory Committees ◽

Allogeneic Hsct ◽

Post Transplant ◽

Equity Ownership

Abstract Introduction: Attaining complete remission (CR) prior to HSCT is associated with better outcomes post-HSCT. Inotuzumab ozogamicin (INO), an anti-CD22 antibody conjugated to calicheamicin, has shown significantly higher remission rates (CR/CRi and MRD negativity) compared with standard chemotherapy (SC) in patients (pts) with R/R ALL (Kantarjian et al. N Engl J Med. 2016). Pts treated with INO were more likely to proceed to HSCT than SC, which allowed for a higher 2-yr probability of overall survival (OS) than patients receiving SC (39% vs 29%). We investigated the role of prior transplant and proceeding directly to HSCT after attaining remission from INO administration as potential factors in determining post-HSCT survival to inform when best to use INO in R/R ALL patients. Methods: The analysis population consisted of R/R ALL pts who were enrolled and treated with INO and proceeded to allogeneic HSCT as part of two clinical trials: Study 1010 is a Phase 1/2 trial (NCT01363297), while Study 1022 is the pivotal randomized Phase 3 (NCT01564784) trial. Full details of methods for both studies have been previously published (DeAngelo et al. Blood Adv. 2017). All reference to OS pertains to post-HSCT survival defined as time from HSCT to death from any cause. Results: As of March 2016, out of 236 pts administered INO in the two studies (Study 1010, n=72; Study 1022, n=164), 101 (43%) proceeded to allogeneic HSCT and were included in this analysis. Median age was 37 y (range 20-71) with 55% males. The majority of pts received INO as first salvage treatment (62%) and 85% had no prior SCT. Most pts received matched HSCTs (related = 25%; unrelated = 45%) with peripheral blood as the predominant cell source (62%). The conditioning regimens were mainly myeloablative regimens (60%) and predominantly TBI-based (62%). Dual alkylators were used in 13% of pts, while thiotepa was used in 8%. The Figure shows post-transplant survival in the different INO populations: The median OS post-HSCT for all pts (n=101) who received INO and proceeded to HSCT was 9.2 mos with a 2-yr survival probability of 41% (95% confidence interval [CI] 31-51%). In patients with first HSCT (n=86) the median OS post-HSCT was 11.8 mos with a 2-yr survival probability of 46% (95% CI 35-56%). Of note, some patients lost CR while waiting for HSCT and had to receive additional treatments before proceeding to HSCT (n=28). Those pts who went directly to first HSCT after attaining remission with no intervening additional treatment (n=73) fared best, with median OS post-HSCT not reached with a 2-yr survival probability of 51% (95% CI 39-62%). In the latter group, 59/73 (80%) attained MRD negativity, and 49/73 (67%) were in first salvage therapy. Of note, the post-HSCT 100-day survival probability was similar among the 3 groups, as shown in the Table. Multivariate analyses using Cox regression modelling confirmed that MRD negativity during INO treatment and no prior HSCT were associated with lower risk of mortality post-HSCT. Other prognostic factors associated with worse OS included older age, higher baseline LDH, higher last bilirubin measurement prior to HSCT, and use of thiotepa. Veno-occlusive disease post-transplant was noted in 19 of the 101 pts who received INO. Conclusion: Administration of INO in R/R ALL pts followed with allogeneic HSCT provided the best long-term survival benefit among those who went directly to HSCT after attaining remission and had no prior HSCT. Disclosures DeAngelo: Glycomimetics: Research Funding; Incyte: Consultancy, Honoraria; Blueprint Medicines: Honoraria, Research Funding; Takeda Pharmaceuticals U.S.A., Inc.: Honoraria; Shire: Honoraria; Pfizer Inc.: Consultancy, Honoraria, Research Funding; Novartis Pharmaceuticals Corporation: Consultancy, Honoraria, Research Funding; BMS: Consultancy; ARIAD: Consultancy, Research Funding; Immunogen: Honoraria, Research Funding; Celgene: Research Funding; Amgen: Consultancy, Research Funding. Kantarjian: Novartis: Research Funding; Amgen: Research Funding; Delta-Fly Pharma: Research Funding; Bristol-Meyers Squibb: Research Funding; Pfizer: Research Funding; ARIAD: Research Funding. Advani: Takeda/ Millenium: Research Funding; Pfizer: Consultancy. Merchant: Pfizer: Consultancy, Research Funding. Stock: Amgen: Consultancy; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Seattle Genetics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Wang: Pfizer: Employment, Equity Ownership. Zhang: Pfizer: Employment, Equity Ownership. Loberiza: Pfizer: Employment, Equity Ownership. Vandendries: Pfizer: Employment, Equity Ownership. Marks: Pfizer: Consultancy, Honoraria, Speakers Bureau; Amgen: Consultancy, Honoraria, Speakers Bureau.

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Multiple Myeloma Cells Induce Lipolysis in Adipocytes and Uptake Fatty Acids through Fatty Acid Transporter Proteins

Blood ◽

10.1182/blood-2020-142625 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 41-42

Author(s):

Cristina Panaroni ◽

Keertik Fulzele ◽

Tomoaki Mori ◽

Chukwuamaka Onyewadume ◽

Noopur S. Raje

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Fatty Acid ◽

Board Of Directors ◽

Metabolic Reprogramming ◽

Glycerol Production ◽

Advisory Committees ◽

Myeloma Cells ◽

Long Chain

Multiple myeloma (MM) originates in the bone marrow where adipocytes occupy 65% of the cellular volume in a typical myeloma patient. Cancer associated adipocytes support the initiation, progression, and survival of solid tumors via mechanisms including adipokine secretion, modulation of the tumor microenvironment, and metabolic reprogramming of cancer cells. Although MM cells are surrounded by abundant bone marrow adipocytes (BMAd), the nature of their interaction remains unclear. Recent studies have elucidated the role of BMAds in supporting the survival of MM cells, in part, through secreted adiponectin. Increased fatty acid (FA) metabolism may result in metabolic reprogramming of cancer cells impacting their growth and survival. Here, we hypothesize that MM cells extract FA from adipocytes for their growth. We first characterized mesenchymal stem cells (MSCs) from MGUS, smoldering MM (SMM), and newly diagnosed MM (NDMM) patients by flow cytometry analysis. MSCs showed significant increase in Pref1, leptin receptor and perilipin A, suggesting increased adipogenic commitment. MSCs from healthy donors (HD), MGUS, SMM, and NDMM patients were induced to differentiate into adipocytes and then co-cultured with human MM MM.1S cells. After 72 hr of co-culture, CyQUANT assay demonstrated significant increase in proliferation of MM.1S cells in the presence of BMAd from HD; this was further increased in the presence of BMAd from MGUS/SMM and NDMM. These data suggest that the BMAd support the growth of MM cells and this effect is more pronounced in patient derived BMAd. A PCR-array targeting lipid metabolism on BM fat aspirates showed significant deregulation of genes involved in FA synthesis and lipolysis. Taken together, our data suggest that BMAd in MM patients are altered to further support the aggressive expansion of MM cells. The proliferative-supportive role of adipocytes was further validated in co-culture of OP9 murine BM stromal preadipocytes with 5TGM1 murine MM cells. To study the bidirectional interaction of MM/ BMAd, mature OP9 adipocytes were co-cultured with 5TGM1 or human OPM2 MM cells for 24 hr. Intracellular lipid droplets were labelled with Deep Red LipidTox stain. The lipid droplet sizes were significantly decreased in the presence of both 5TGM1 and OPM2 cells compared to OP9 alone. The decrease in lipid size suggested that MM cells may induce lipolysis in adipocytes. Indeed, 24hr co-culture of 5TGM1 cells with OP9 mature adipocytes significantly increased lipolysis 3-fold as measured by glycerol secretion in conditioned media. Co-culture of OP9 adipocytes with other MM cell lines of human origin, MM.1S, INA6, KMS-12 PE, and OPM2 also significantly increased the glycerol production as much as 4-fold. Taken together these data indicate that MM cells induce lipolysis in adipocytes. In contrast, treatment of 5TGM1 cells with synthetic catecholamine isoproterenol did not induce lipolysis, or glycerol production, indicating lack of triglyceride storage. Next, we hypothesized that the free FAs released from adipocytes are taken up by MM cells for various biological processes. To test this, 5TGM1, MM.1S and OPM2 cells were incubated with BODIPY-C12 and BODIPY-C16, the BODIPY-fluorophore labelled 12-carbon and 16-carbon long chain FA. All MM cells showed saturated uptake of the FA within 10 minutes suggesting that MM cells have efficient FA transporters. To confirm this uptake, unstained 5TGM1, OPM2 and KMS12 PE cells were co-cultured with the LipidTox-labelled OP9 mature adipocytes. After 24 hours, flow cytometric analysis showed LipidTox signal in MM cells. These data demonstrate that FAs released by MM induced adipocyte lipolysis are taken up by MM cells. Long-chain FAs such as BODIPY-C12 and BODIPY-C16 are transported into cells through FA transporter protein (FATP) family of lipid transporters. We therefore analyzed patient samples which showed that CD138+ plasmacells and myeloma cells expressed high levels of FATP1 and FATP4 whereas, their expression was absent in lineage-sibling T-cells. Moreover, pretreatment with Lipofermata, a FATP inhibitor, was able to decrease the uptake of BODIPY-C12 and -C16 in 5TGM1 cells. Taken together, our data show that myeloma cells induce lipolysis in adipocytes and the released free FAs are then uptaken by myeloma cells through FATPs. Inhibiting myeloma cell induced lipolysis or uptake of FA through FATPs may be a potential anti-tumor strategy. Disclosures Fulzele: FORMA Therapeutics, Inc: Current Employment, Other: Shareholder of Forma Therapeutics. Raje:Amgen: Consultancy; bluebird bio: Consultancy, Research Funding; Caribou: Consultancy, Membership on an entity's Board of Directors or advisory committees; Immuneel: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Consultancy; Celgene: Consultancy; Immuneel: Consultancy; Janssen: Consultancy; Karyopharm: Consultancy; Takeda: Consultancy.

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Impact of Interim FDG-PET (iPET) in the Outcomes of Patients with Aggressive B-Cell Lymphomas Receiving DA-EPOCH-R

Blood ◽

10.1182/blood-2019-122393 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 2898-2898

Author(s):

Vania Phuoc ◽

Leidy Isenalumhe ◽

Hayder Saeed ◽

Celeste Bello ◽

Bijal Shah ◽

...

Keyword(s):

B Cell ◽

Board Of Directors ◽

Fdg Pet ◽

Research Funding ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Advisory Committees ◽

End Of Treatment

Introduction: 2-[18F] fluoro-2-deoxy-D-glucose positron emission tomography (FDG-PET) remains the standard of care for baseline and end of treatment scans for aggressive non-Hodgkin lymphomas (NHLs). However, the role of interim FDG-PET remains not as well defined across aggressive NHLs, especially in the era of high-intensity chemoimmunotherapy. Interim FDG-PET (iPET) can serve as an early prognostic tool, and prior studies evaluating the utility of iPET-guided treatment strategies primarily focused on diffuse large B-cell lymphomas (DLBCL) and frontline R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone). Classification criteria systems assessing response also differ between studies with no clear consensus between use of Deauville criteria (DC), International Harmonization Project (IHP), and the ΔSUVmax method. Methods: This study evaluates our institutional experience with iPET during treatment with DA-EPOCH ± R (dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin with or without Rituximab) in aggressive NHLs. We retrospectively evaluated 70 patients at Moffitt Cancer Center who started on DA-EPOCH ± R between 1/1/2014 to 12/31/2018 for aggressive NHLs. Response on interim and end-of-treatment (EOT) scans were graded per DC, IHP, and ΔSUVmax methods, and progression free survival (PFS) probability estimates were calculated with chi-square testing and Kaplan Meier method. PFS outcomes were compared between interim negative and positive scans based on each scoring method. Outcomes were also compared between groups based on interim versus EOT positive or negative scans. Results: We identified 70 patients with aggressive NHLs who received DA-EPOCH ± R at our institute. The most common diagnoses were DLBCL (61%) followed by Burkitt's lymphoma (10%), primary mediastinal B-cell lymphoma (9%), plasmablastic lymphoma (7%), gray zone lymphoma (6%), primary cutaneous large B-cell lymphoma (1%), primary effusion lymphoma (1%), and other high-grade NHL not otherwise specified (3%). Of the 43 patients with DLBCL, 21/43 (49%) had double hit lymphoma (DHL) while 7/43 (16%) had triple hit lymphoma (THL), and 3/43 (7%) had MYC-rearranged DLBCL while 2/43 (5%) had double expressor DLBCL. Thirty nine out of 70 (56%) were female, and median age at diagnosis was 58.39 years (range 22.99 - 86.86 years). Most patients had stage IV disease (49/70, 70%), and 43/70 (61%) had more than one extranodal site while 45/70 (64%) had IPI score ≥ 3. Forty-six out of 70 (66%) received central nervous system prophylaxis, most with intrathecal chemotherapy (44/70, 63%). Fifty-five out of 70 (79%) had iPET available while 6/70 (9%) had interim computerized tomography (CT) scans. Fifty-six out of 70 (80%) had EOT PET, and 4/70 (6%) had EOT CT scans. Sustained complete remission occurred in 46/70 (66%) after frontline DA-EPOCH ± R (CR1), and 12/70 (17%) were primary refractory while 5/70 (7%) had relapse after CR1. Four of 70 (6%) died before cycle 3, and 3/70 (4%) did not have long-term follow-up due to transition of care elsewhere. Median follow-up was 15.29 months (range 0.85 - 60.09 months). There was significantly better PFS observed if iPET showed DC 1-3 compared to DC 4-5 (Χ2=5.707, p=0.0169), and PFS was better if iPET was negative by IHP criteria (Χ2=4.254, p=0.0392) or ΔSUVmax method (Χ2=6.411, p=0.0113). Comparing iPET to EOT PET, there was significantly better PFS if iPET was negative with EOT PET negative (iPET-/EOT-) compared to iPET positive with EOT negative (iPET+/EOT-), and iPET+/EOT+ and iPET-/EOT+ had worse PFS after iPET-/EOT- and iPET+/EOT- respectively. This pattern in iPET/EOT PFS probability remained consistent when comparing DC (Χ2=30.041, p<0.0001), IHP (Χ2=49.078, p<0.0001), and ΔSUVmax method (Χ2=9.126, p=0.0104). These findings fit clinical expectations with positive EOT scans indicating primary refractory disease. There was no significant difference in PFS when comparing DLBCL versus non-DLBCL (Χ2=3.461, p=0.0628) or DHL/THL versus non-DHL/THL diagnoses (Χ2=2.850, p=0.0914). Conclusion: Our findings indicate a prognostic role of iPET during treatment with DA-EPOCH ± R for aggressive NHLs. Significant differences in PFS were seen when graded by DC, IHP, and ΔSUVmax methods used in prior studies and when comparing interim versus EOT response. Larger studies are needed to confirm these findings. Disclosures Bello: Celgene: Speakers Bureau. Shah:Novartis: Honoraria; AstraZeneca: Honoraria; Spectrum/Astrotech: Honoraria; Adaptive Biotechnologies: Honoraria; Pharmacyclics: Honoraria; Jazz Pharmaceuticals: Research Funding; Incyte: Research Funding; Kite/Gilead: Honoraria; Celgene/Juno: Honoraria. Sokol:EUSA: Consultancy. Chavez:Janssen Pharmaceuticals, Inc.: Speakers Bureau; Genentech: Speakers Bureau; Kite Pharmaceuticals, Inc.: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees.

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Bleeding Complications in Patients with Cancer and COVID 19- Analysis from the COVID 19and Cancer Consortium (CCC19) Registry

Blood ◽

10.1182/blood-2021-154314 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 4997-4997

Author(s):

Surbhi Shah ◽

Shuchi Gulati ◽

Ang Li ◽

Julie Fu ◽

Vaibhav Kumar ◽

...

Keyword(s):

Mechanical Ventilation ◽

Board Of Directors ◽

Stock Options ◽

Research Funding ◽

Tumor Board ◽

Bleeding Complications ◽

Advisory Committees ◽

Icu Admission ◽

Patients With Cancer ◽

Privately Held

Abstract Background : Patients (pts) with COVID-19 are reported to have increased risk of venous thromboembolism yet bleeding has been an under recognized complication. Rates of bleeding remain unexamined in all patients especially in pts with cancer and COVID-19. Aim: To estimate the incidence of bleeding complication in patients with cancer and COVID 19 Methods: The CCC19 international registry (NCT04354701) aims to investigate complications of COVID-19 in pts with cancer. Our aim was to investigate the frequency of bleeding in hospitalized adult pts with cancer andCOVID-19, enrolled between March 16, 2020 and Feb 8, 2021. The incidence of bleeding complications was captured as defined by CCC19 and included both major and non major bleeding . Associated baseline clinic-pathologic prognostic factors and outcomes such as need for mechanical ventilation, intensive care unit (ICU) admission and mortality rates were assessed Results :3849 pts met analysis inclusion criteria. Bleeding was reported in 276 (7%) pts with median age of 70years; incidence was 6.6 % in females and 7.6 % in males, 6.5% in non-Hispanic white pts, 8.2 % in non-Hispanic Black pts, and 7.8 % in Hispanic pts. 74% had solid cancer and 29% had hematologic malignancies, 33% had received anti-cancer therapy in preceding 30 days, and 8% had surgery within 4weeks. In pts taking antiplatelet or anticoagulant medications at baseline, 7.2% developed bleeding. Need for mechanical ventilation, ICU admission, 30-day mortality, and total mortality were significantly higher in those with bleeding complications compared to those without, p<0.05 Conclusion : We describe the incidence of bleeding in a large cohort of pts with cancer and COVID-19. Bleeding events were observed in those with adverse outcomes including mechanical ventilation, ICU admission, and high mortality; the overall mortality of 43% in patients with bleeding complications is especially notable. This important complication may reflect underlying COVID-19 pathophysiology as well as iatrogenic causes. Figure 1 Figure 1. Disclosures Kumar: Diagnostica Stago: Honoraria. Zon: AMAGMA AND RLZ: Consultancy, Current holder of individual stocks in a privately-held company. Byeff: Pfizer, BMS, Takeda,Teva, Merck, United health: Consultancy, Current equity holder in publicly-traded company, Current holder of stock options in a privately-held company. Nagaraj: Novartis: Research Funding. Hwang: astrazaneca,Merck,bayer, Genentech: Consultancy, Research Funding. McKay: Myovant: Consultancy; Bayer: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees; Exelixis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Calithera: Membership on an entity's Board of Directors or advisory committees; Tempus: Research Funding; Merck: Consultancy, Membership on an entity's Board of Directors or advisory committees; Tempus: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Dendreon: Consultancy; Caris: Other: Serves as a molecular tumor board ; Vividion: Consultancy; Sorrento Therapeutics: Consultancy; Bayer: Research Funding. Warner: Westat, Hemonc.org: Consultancy, Current holder of stock options in a privately-held company. Connors: Pfizer: Honoraria; CSL Behring: Research Funding; Alnylam: Consultancy; Bristol-Myers Squibb: Honoraria; takeda: Honoraria; Abbott: Consultancy. Rosovsky: Janssen: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Inari: Consultancy, Membership on an entity's Board of Directors or advisory committees; Dova: Consultancy, Membership on an entity's Board of Directors or advisory committees.

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Weight Loss Predicts Inferior Outcome in Polycythemia Vera Patients

Blood ◽

10.1182/blood-2020-134801 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 22-23

Author(s):

Albert Jang ◽

Hussein Hamad ◽

Sarvari Venkata Yellapragada ◽

Iberia R. Sosa ◽

Gustavo A. Rivero

Keyword(s):

Weight Loss ◽

High Risk ◽

Blood Cell ◽

Polycythemia Vera ◽

Board Of Directors ◽

Clinical Variable ◽

Advisory Committees ◽

Wbc Count

Background: Conventional risk factors for inferior outcomes in polycythemia vera (PV) include elevated hematocrit, white blood cell (WBC) count, age, and abnormal karyotype. Weight loss adversely impacts survival in cancer patients. JAK2 myeloproliferative neoplasms (MPN) upregulate tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and IL-8 and induce decreased leptin levels leading to weight loss. The impact of weight loss in PV patients receiving best supportive care (i.e. frontline hydroxyurea [HU] therapy, phlebotomy) on overall survival (OS) is largely unknown. In this study, we seek to investigate: (1) differential effect on survival for weight loss, and (2) variables with predictive value for weight loss among JAK2 inhibitor-naïve PV patients. Methods: After IRB approval, 46 patients at the Michael E. DeBakey VA Medical Center diagnosed with PV between 2000 and 2016 were selected for analysis. Our outcome of interest was OS among PV patients exhibiting weight loss versus patients who maintained, gained weight or had minor weight loss. To objectively estimate weight changes overtime, the difference between baseline BMI [BMI-B] at the time of diagnosis and BMI at last follow-up (BMI-L) was obtained for each patient. Survival analysis was performed for PV patients exhibiting more than 10% weight loss (>10%) versus all other patients (less than 10% loss, stable and increased weight) (<10%) over time. Kaplan-Meier (KM) method was used to determine OS. Cox regression model was performed to assess independent role of different variables including age, blood cell counts and ferritin level Statistical analysis was performed using SAS software. Results: Median BMI loss was 10% (0.03-36.72%); 33/46 (71.7%) and 13/46 (28.2%) patients developed <10% and >10% BMI loss, respectively. Baseline characteristics are summarized in Table 1. Median BMI at last follow up was 21 for PV patients exhibiting >10% BMI loss and 27.7 for PV patients exhibiting <10% BMI loss (p<0.01). Median age was higher among patients exhibiting >10% BMI loss (68 vs 56 y, respectively, p=0.006). A non-significant clinical trend for higher WBC was observed among patients losing >10% BMI (10.9 vs 7.6 K/uL, p=0.08). Median Hemoglobin (Hb), hematocrit (Hct) and ferritin were intriguingly lower in the >10% loss group at 16 vs 18.3 g/dL (p=0.01), 49.3 vs 54.2% (p=0.04) and 29.8 vs 50.6 ng/mL (p=0.09) respectively, while median RDW was higher at 18 vs 15.1% (p=0.01). OS was 9125 days vs 5364 days, in patients with <10% and >10% BMI loss, respectively (p=0.02, HR=0.20; CI 95% 0.04-0.84) (Figure 1). On multivariate analysis, age (hazard ratio [HR], 1.34; p<0.02) and WBC count (HR, 1.57; p<0.01), were predictive of OS. Conclusions: A subgroup of PV patients exhibit progressive weight loss. Over 10% BMI reduction is associated with decreased survival, suggesting that "early weight loss" is an independent clinical variable that predicts high risk PV. While a larger study is needed to validate this observation, this small study highlights the role of leukocytosis, advanced age and weight loss in PV. Confirmation of the observations reported here could unveil an important role for pharmacologic and/or dietary interventions to improve survival among high-risk PV patients. Disclosures Rivero: agios: Membership on an entity's Board of Directors or advisory committees; celgene: Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees.

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NADPH Oxidases Are Required for Full Platelet Activation In Vitro and Thrombosis In Vivo but Dispensable for Plasma Coagulation and Hemostasis

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvbaha.120.315565 ◽

2020 ◽

Author(s):

Dina Vara ◽

Reiner K. Mailer ◽

Anuradha Tarafdar ◽

Nina Wolska ◽

Marco Heestermans ◽

...

Keyword(s):

Thrombus Formation ◽

Single Gene ◽

Coagulation Cascade ◽

Intracellular Cgmp ◽

Antithrombotic Effects ◽

Tail Tip ◽

Synthase Inhibitor

Objective: Using 3KO (triple NOX [NADPH oxidase] knockout) mice (ie, NOX1 −/− /NOX2 −/− /NOX4 −/− ), we aimed to clarify the role of this family of enzymes in the regulation of platelets in vitro and hemostasis in vivo. Approach and Results: 3KO mice displayed significantly reduced platelet superoxide radical generation, which was associated with impaired platelet aggregation, adhesion, and thrombus formation in response to the key agonists collagen and thrombin. A comparison with single-gene knockouts suggested that the phenotype of 3KO platelets is the combination of the effects of the genetic deletion of NOX1 and NOX2, while NOX4 does not show any significant function in platelet regulation. 3KO platelets displayed significantly higher levels of cGMP—a negative platelet regulator that activates PKG (protein kinase G). The inhibition of PKG substantially but only partially rescued the defective phenotype of 3KO platelets, which are responsive to both collagen and thrombin in the presence of the PKG inhibitors KT5823 or Rp-8-pCPT-cGMPs, but not in the presence of the NOS (NO synthase) inhibitor L-NG-monomethyl arginine. In vivo, triple NOX deficiency protected against ferric chloride–driven carotid artery thrombosis and experimental pulmonary embolism, while hemostasis tested in a tail-tip transection assay was not affected. Procoagulatory activity of platelets (ie, phosphatidylserine surface exposure) and the coagulation cascade in platelet-free plasma were normal. Conclusions: This study indicates that inhibiting NOXs has strong antithrombotic effects partially caused by increased intracellular cGMP but spares hemostasis. NOXs are, therefore, pharmacotherapeutic targets to develop new antithrombotic drugs without bleeding side effects.

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