Khanh Nghiem

Selective serotonin reuptake inhibitors: measurement of effect on platelet function

Selective serotonin reuptake inhibitors (SSRIs) reduce platelet serotonin and are associated with increased gastrointestinal bleeding, an effect that is enhanced when taken with NSAIDs or aspirin. The best method to evaluate hemorrhagic events in patients taking SSRIs has not been determined. Platelet aggregation, which is not widely available, shows SSRI inhibition of platelet function; we tested whether a platelet function analyzer could detect SSRI inhibition of platelet function. Two groups of outpatients with mood disorders were recruited; each patient was taking a stable dose of either an SSRI or bupropion for at least 6 weeks. They were tested using the platelet function analyzer-100 (PFA-100; Dade International Inc, Miami, Fla) concomitantly with platelet aggregation. Fifty-eight patients were analyzed. We detected significant differences between the groups using aggregation methods with arachidonic acid (aggregation, P = 0.00001; release, P = 0.009) and collagen (aggregation, P = 0.016; release, P = 0.006). The PFA-100 did not detect differences between the groups or results outside the reference range. The PFA-100 does not detect the inhibitory effects of SSRIs on platelet function, but it can be used to direct evaluation of bleeding in a patient taking an SSRI. Abnormal PFA-100 results suggest additional evaluation for von Willebrand disease, other platelet inhibitory medications, or underlying intrinsic platelet dysfunction.

Incidence and risk factors of bleeding-related adverse events in patients with chronic lymphocytic leukemia treated with ibrutinib

Ibrutinib is associated with bleeding-related adverse events of grade ≤2 in severity, and infrequently with grade ≥3 events. To investigate the mechanisms of bleeding and identify patients at risk, we prospectively assessed platelet function and coagulation factors in our investigator-initiated trial of single-agent ibrutinib for chronic lymphocytic leukemia. At a median follow-up of 24 months we recorded grade ≤2 bleeding-related adverse events in 55% of 85 patients. No grade ≥3 events occurred. Median time to event was 49 days. The cumulative incidence of an event plateaued by 6 months, suggesting that the risk of bleeding decreases with continued therapy. At baseline, von Willebrand factor and factor VIII levels were often high and normalized on treatment. Platelet function measured via the platelet function analyzer (PFA-100™) was impaired in 22 patients at baseline and in an additional 19 patients on ibrutinib (often transiently). Collagen and adenosine diphosphate induced platelet aggregation was tested using whole blood aggregometry. Compared to normal controls, response to both agonists was decreased in all patients with chronic lymphocytic leukemia, whether on ibrutinib or not. Compared to untreated chronic lymphocytic leukemia patients, response to collagen showed a mild further decrement on ibrutinib, while response to adenosine diphosphate improved. All parameters associated with a significantly increased risk of bleeding-related events were present at baseline, including prolonged epinephrine closure time (HR 2.74, P=0.012), lower levels of von Willebrand factor activity (HR 2.73, P=0.009) and factor VIII (HR 3.73, P=0.0004). In conclusion, both disease and treatment-related factors influence the risk of bleeding. Patients at greater risk for bleeding of grade ≤2 can be identified by clinical laboratory tests and counseled to avoid aspirin, non-steroidal anti-inflammatory drugs and fish oils. ClinicalTrials.gov identifier NCT01500733

Interleukin-2 cycling causes transient increases in high-sensitivity C-reactive protein and D-dimer that are not associated with plasma HIV-RNA levels.

Objective:To determine the effects of interleukin (IL)-2 treatment on inflammatory and thrombotic biomarkers in chronically HIV-infected adults receiving antiretroviral therapy. Methods:Cryopreserved plasma was evaluated retrospectively for C-reactive protein (CRP) and D-dimer at baseline, end of an IL-2 cycle, and long-term follow up from two randomized, controlled trials: 57 IL-2-naive adults receiving either three to six cycles of IL-2 as well as antiretroviral therapy (nucleoside analogues) or antiretroviral therapy alone for 12 months, and 40 IL-2-experienced adults on highly active antiretroviral therapy who either interrupted or continued therapy for 6 months after a baseline IL-2 cycle. High-sensitivity CRP (hsCRP) was measured by immunonephelometry (detection limit 0.175 mg/l) and D-dimer by latex agglutination (detection limit 0.20 mg/l). Median within-group differences and pre and post-IL-2 changes between groups were assessed via nonparametric Wilcoxon signed-rank and Mann–Whitney U-tests. Spearmans rank test was used to assess correlations between changes in hsCRP, D-dimer, and HIV-RNA viral load. Results:Significant increases in hsCRP (study 1: 138.6 mg/l; study 2: 58.9 mg/l) and D-dimer (study 1: 3.1 mg/l; study 2: 0.4 mg/l, all P < 0.0001) occurred by the end of the initial IL-2 cycle, returning to baseline by the end of study. No correlations were seen between changes in hsCRP or D-dimer and HIV-RNA, CD4 T-cell count, or proliferation (Ki67 expression). No thrombotic or cardiovascular serious adverse events occurred during these study periods. Conclusion:IL-2 dosing caused transient increases in plasma hsCRP and D-dimer levels, regardless of HIV-RNA viral load, suggesting the possibility of increased risk for thrombotic events.

Inhibitory effect of nitrite on coagulation processes demonstrated by thrombelastography

Nitric oxide (NO) can be generated by two-step reduction pathway in which nitrate is converted first into nitrite and then into NO via several mechanisms, as well as from arginine by endogenous nitric oxide synthase (NOS). We have recently shown that nitrite ions in the presence of erythrocytes inhibit platelet aggregation and activation, as measured by aggregometry and flow cytometric analysis of P-selectin, through its reduction to NO under partially deoxygenated conditions. In the current study, we investigated how nitrite may affect overall clotting processes via modulating platelet function using thrombelastography (TEG). We measured three major TEG parameters, reaction time (R, time to initial fibrin formation), α angle (velocity of clot growth) and maximum amplitude (MA, maximum clot strength) using blood from healthy volunteers. An NO donor (DEANONOate) showed inhibitory effects on all TEG parameters in platelet rich plasma (PRP) and whole blood, resulting in delayed R, decreased angle, and reduced MA in a dose dependent manner. Nitrite ions also exhibited inhibitory effects in whole blood at 20% hematocrit, and this was greatly enhanced under hypoxic conditions, being demonstrable at 0.1 μM concentration. Neither compound changed any TEG parameters in plasma. Our results suggest that nitrite affects overall blood clotting and that TEG may be used to follow this process. Further the physiological effects of factors which determine NO bioavailability, such as endogenous levels of blood and tissue nitrite, may be useful as biomarkers for predicting hemostatic potential.

Elevations in D-dimer and C-reactive protein are associated with the development of osteonecrosis of the hip in HIV-infected adults.

Background:A high incidence of nontraumatic osteonecrosis has been reported in HIV-infected patients. We investigated the levels of D-dimer and C-reactive protein (CRP) in a cohort of HIV-infected adults with and without osteonecrosis of the femoral head. Methods:Forty-three HIV-infected patients with osteonecrosis of the femoral head and a comparison group of 50 HIV-infected patients with negative MRI of the hips and for whom serial plasma samples were available were included. D-dimer and CRP levels were measured prior to and at the time of diagnosis for osteonecrosis patients, at the time of negative MRI of the hips for controls, and at least 6 months later for both groups. Results:Biomarker levels were elevated at the time of diagnosis in the osteonecrosis cohort compared with controls. Median D-dimer value was 0.32 &mgr;g/ml in the osteonecrosis group compared with less than 0.22 &mgr;g/ml in the control group (P = 0.016). For CRP, the corresponding values were 2.52 mg/l and 1.23 mg/l (P = 0.003). Postdiagnosis, D-dimer and CRP levels were also elevated in the osteonecrosis patients compared with controls. Linear regression demonstrated a rise in D-dimer levels from prediagnosis to diagnosis in the osteonecrosis patients whereas CRP levels did not change significantly over time. Conclusion:Compared to controls, patients who developed osteonecrosis had elevated levels of D-dimer and CRP at diagnosis. D-dimer levels increased whereas CRP levels did not change significantly from prediagnosis to diagnosis. These data suggest that patients with higher levels of inflammation are at an increased risk of osteonecrosis.

Antiretroviral Boosting Agent Cobicistat Increases the Pharmacokinetic Exposure and Anticoagulant Effect of Dabigatran in HIV-Negative Healthy Volunteers

Drug interactions between antiretroviral therapy and anticoagulant medications are of particular concern given that ≈50% of the current HIV population is >50 years of age. Moreover, HIV infection is characterized by a hypercoaguable state and premature immunologic aging, in which thromboembolic events may be as much as 10 times more prevalent than in the general population across all age spectra.1 Dabigatran was the first direct oral anticoagulant approved by the US Food & Drug Administration and is the only direct oral anticoagulant with a US Food & Drug Administration -approved specific reversal agent, idarucizumab. Unlike warfarin and many other direct oral anticoagulants, dabigatran is not a substrate, inhibitor, or inducer of cytochrome P450 metabolic enzymes. However, dabigatran is a substrate of Permeability-glycoprotein (P-gp) and renal multidrug and toxin extrusion-1 transporters. Cobicistat is a US Food & Drug Administration -approved antiretroviral-boosting agent that is coformulated with numerous fixed-dose combination antiretroviral products because of its inhibitory effects on cytochrome P450 3A4. Currently, ≈40% of all treatment-naive patients with HIV in the United States are initiated on a cobicistat-boosted antiretroviral regimen. In addition to cytochrome P450 3A4, cobicistat is also an inhibitor of both P-gp and multidrug and toxin extrusion-1 transporters.2 Thus, this study aimed to determine whether the …

Acquired haemophilia A after stem cell transplant for sickle cell disease: treatment with recombinant porcine factor VIII (OBI‐1) and tolerance induction with rituximab/prednisone

J. N. LOZIER,* K. NGHIEM,* M. LEE,† B. HODSDON,‡ G. JOE,‡ R. P. WEITZEL,§ J . F. TISDALE§ and M. HSIEH§ *Department of Laboratory Medicine, National Institutes of Health Clinical Center, Bethesda, MD; †UCLA School of Public Health, Los Angeles, CA; ‡Rehabilitation Medicine Department, National Institutes of Health Clinical Center; and §Molecular and Clinical Hematology Branch, NHLBI, NIH, Bethesda, MD, USA

Biochemical dynamics relevant to the safety of low-dose, intraclot alteplase for deep vein thrombosis

Intraclot tissue plasminogen activator (tPA) has been shown to be an effective treatment for deep vein thrombosis (DVT) (Radiology 2008;246:619 and J Vasc Interv Radiol 2011;22:1107). We sought to correlate pharmacokinetics of tPA, fibrinogen, fibrinolytic inhibitors, and D-dimers with the safety and efficacy of intraclot tPA. Thirty subjects received intraclot tPA for lower extremity DVT by infiltrating the thrombus with ≤10 mg doses tPA in an open-label study, using a pulse-spray catheter. We measured various parameters over 8 h following a first dose of tPA. Mean tPA levels of 75 units per mL (95% confidence interval 19-131 units/mL) were seen immediately after administration of a mean tPA dose of 8.0 mg (SD 1.5 mg). tPA levels returned to baseline within 2 h of completion of treatment. Plasminogen activator inhibitor-1 (PAI-1) was consumed following tPA treatment, but rose to levels significantly greater than baseline (P < 0.001). Fibrinogen decreased slightly, but remained >125 mg/dL for all subjects. α2-antiplasmin decreased from a mean of 115 units/mL to 56 units/mL after tPA administration (P < 0.001) and remained decreased for 8 h. Plasminogen at baseline (112 units/mL) decreased to 89 units/mL immediately after tPA administration (P < 0.001) and was unchanged thereafter. D-dimer levels were >20 μg/mL in all but 4 subjects, one of whom was the only one to fail to achieve clot lysis. The safety of low-dose, intraclot tPA is due to its short persistence in the circulation, lack of hypofibrinogenemia, and a reflexive rise of PAI-1. Subjects whose D-dimers remain <20 μg/mL are at risk of not achieving thrombolysis.

Differential Influence of the Antiretroviral Pharmacokinetic Enhancers Ritonavir and Cobicistat on Intestinal P-Glycoprotein Transport and the Pharmacokinetic/Pharmacodynamic Disposition of Dabigatran

ABSTRACT Dabigatran etexilate (DE) is a P-glycoprotein (P-gp) probe substrate, and its active anticoagulant moiety, dabigatran, is a substrate of the multidrug and toxin extrusion protein-1 (MATE-1) transporter. The antiretroviral pharmacokinetic enhancers, ritonavir and cobicistat, inhibit both these transporters. Healthy volunteers received single doses of DE at 150 mg alone, followed by ritonavir at 100 mg or cobicistat at 150 mg daily for 2 weeks. DE was then given 2 h before ritonavir or cobicistat. One week later, DE was given simultaneously with ritonavir or cobicistat. No significant increases in dabigatran pharmacokinetic (PK) exposure or thrombin time (TT) measures were observed with the simultaneous administration of ritonavir. Separated administration of ritonavir resulted in a mean decrease in dabigatran PK exposure of 29% (90% confidence interval [CI], 18 to 40%) but did not significantly change TT measures. However, cobicistat increased dabigatran PK exposure (area under the concentration-versus-time curve from time zero to infinity and maximum plasma concentration) by 127% each (90% CI, 81 to 173% and 59 to 196%, respectively) and increased TT measures (33% for the area-under-the-effect curve from time zero to 24 h [90% CI, 22 to 44%] and 51% for TT at 24 h [90% CI, 22 to 78%]) when given simultaneously with dabigatran. Similar increases were observed when cobicistat was administered separately by 2 h from the administration of dabigatran. In all comparisons, no significant increase in the dabigatran elimination half-life was observed. Therefore, it is likely safe to coadminister ritonavir with DE, while there is a potential need for reduced dosing and prudent clinical monitoring with the coadministration of cobicistat due to the greater net inhibition of intestinal P-gp transport and increased bioavailability. (This study has been registered at ClinicalTrials.gov under identifier NCT01896622.)

Identification of a novel mutation in HPS6 in a patient with hemophilia B and oculocutaneous albinism

PURPOSE Hemophilia B, an X-linked disease, manifests with recurrent soft tissue bleeding episodes. Hermansky-Pudlak syndrome, a rare autosomal recessive disorder, is characterized by oculocutaneous albinism and an increased tendency to bleed due to a platelet storage pool defect. We report a novel mutation in HPS6 in a Caucasian man with hemophilia B and oculocutaneous albinism. RESULTS The patient was diagnosed with hemophilia B at age 4months due to recurrent soft tissue bleeding episodes, and he was also diagnosed with Hermansky-Pudlak syndrome at 32years of age due to unexplained oculocutaneous albinism. His factor IX level was markedly reduced at 13%; whole exome and Sanger sequencing showed the Durham mutation in F9 (NM_000133.3). The diagnosis of Hermansky-Pudlak syndrome subtype 6 was established by demonstrating absence of platelet delta granules on whole mount electron microscopy, an abnormal secondary wave in platelet aggregation studies, and a novel homozygous c.1114 C>T (p.Arg372*) mutation in HPS6 (NM_024747.5) on exome analysis and Sanger sequencing. Clinical phenotyping revealed no evidence of recurrent or unusual infections, interstitial lung disease or pulmonary fibrosis, or neurological disorders. The patient was treated with fresh frozen plasma, recombinant factor IX, and aminocaproic acid. Treatment with desmopressin was added to his regimen after he was diagnosed with Hermansky-Pudlak syndrome. Treatment of bleeding episodes results in effective hemostasis, and the patient has not required platelet or blood product transfusions. CONCLUSIONS This report highlights the need to consider Hermansky-Pudlak syndrome as an etiology of oculocutaneous albinism even in patients with known hematologic disorders associated with bleeding. Identification of a novel mutation in HPS6 in an individual with hemophilia B shows that, although quite rare, patients may be diagnosed with two independent inherited bleeding disorders. No evidence of lung disease was found in this adult patient with Hermansky-Pudlak syndrome subtype 6.