Post-infusion Time Points Research Articles

A Native Whole Blood Thrombin Generation Assay Allows Discrimination of Whole Blood Samples with FVIII Levels below 1%

Monitoring the correction of abnormal bleeding tendencies during the treatment of patients with hemostatic disorders is essential to evaluate success of therapy. While single clotting factor assays provide valuable information, global coagulation assays are desirable to better understand the overall hemostatic condition of patients. In Hemophilia A, severity of the clotting defect is traditionally evaluated by determining FVIII activity using chromogenic or clotting assays. Evaluation of thrombin generation in plasma samples for the assessment of bleeding tendencies in hemophilic patients has been suggested. Discriminating between samples with FVIII levels below 1%, however, has been challenging using FVIII activity and thrombin generation assays. We previously reported a native whole blood thrombin generation assay (nWB-TGA) that uses recalcification of whole blood samples without the addition of tissue factor to initiate clotting. We have shown that this assay is sensitive to varying levels of FVIII in vitroand to platelet targeted FVIII gene therapy in a murine model of Hemophilia A. The objective of the present study was to determine if the nWB-TGA can be used to monitor Hemophilia A patients during FVIII therapy and if this assay allows discrimination of whole blood samples with FVIII levels below 1%.Using the nWB-TGA we evaluated thrombin generation in a severe hemophilia A patient carrying an intron 22 inversion. Numerous data points were obtained from 15 different FVIII infusions, each targeting a FVIII level of 50%. Samples collected at least 72 hours (hrs) post infusion (>6 half-lives, calculated FVIII levels <1%) significantly differed from healthy control samples in all thrombin generation parameters. Compared with healthy controls (6.9 ± 0.6 min; mean ± SEM) the hemophilic patient had a lag time (LT) of 24.8 ± 3.4 min. Peak time in healthy controls and the patient was 10.1 ± 1 min and 35 ± 5 min, peak thrombin was 528 ± 78 nM and 124 ± 20 nM, endogenous thrombin potential (ETP) was 1949 ± 117 nM and 1201 ± 50 nM, and thrombin generation rate was 196 ± 58 nM/min and 21 ± 6 nM/min, respectively. While previous studies on thrombin generation in plasma samples mainly reported on differences in peak thrombin and ETP, spiking of hemophilic blood with increasing concentration of recombinant FVIII in vitro revealed that the LT was FVIII dose dependent in our assay. When hemophilic blood was reconstituted with FVIII to a 2%, 5% and 50% level, the LT was 22.5 ± 1.6 min, 16.1 ± 1.7 min and 8.8 ± 0.6 min, respectively. All other thrombin generation parameters were FVIII dose dependent as well. A FVIII dependent LT was also apparent in vivo, when we monitored the patient after FVIII infusions. LT was 6.4 ± 0.2 min at 15 min, 8.5 ± 0.4 min at 24 hrs, and 13.8 ± 0.5 min at 48 hrs post FVIII treatment. We identified the lack of tissue factor as being key to a FVIII dose dependent LT. At all post infusion time points the LT was approximately 5 min when tissue factor was added to the assay. To our surprise, looking at specific time points equal to or greater than 72 hrs post infusion enabled us to discriminate these samples based on the LT (72 hrs: LT= 13.0 ± 0.6 min, 84 hrs: LT= 19.5 ± 0.8 min, 96 hrs: 36.0 ± 4.4 min). The ETP, commonly used as a variable parameter in previous thrombin generation reports, however, was not different among 72, 84 and 96 hrs post FVIII infusion samples. Strikingly, FVIII activity determined by chromogenic and one stage clotting assay was below detection limit (1% FVIII:C) in samples obtained 72 hrs post infusion or later. Thus, the patient in our study displayed considerable thrombin generation determined by the nWB-TGA at post FVIII infusion time points when FVIII levels were considered undetectable with currently available technology. Our data suggest that the different LT observed in 72, 84 and 94 hrs post infusion samples is possibly related to differences in residual FVIII levels below 1%.In conclusion, the nWB-TGA provides a useful tool to monitor efficacy of FVIII replacement therapy and might assist in tailoring individual FVIII treatment regimens. This close to physiological whole blood assay allows distinguishing blood samples with FVIII levels below 1% in vivo, and might help to explain the heterogeneity in bleeding phenotypes observed in severe hemophilia A patients. This assay may also be useful in assessing therapeutic benefit of “long acting” FVIII or FIX products. DisclosuresNo relevant conflicts of interest to declare.

Immune Reconstitution With Activated T Cells In Companion Canines With Lymphoma

Companion canine cancer provides a clinically-relevant immunotherapy model for human malignancies because of their large size, intra-species genetic diversity, genetic similarity of tumors to human oncology, and spontaneously occurring tumors that develop despite an intact immune system. Canine B-lineage non-Hodgkin lymphoma (NHL) is genetically, molecularly, and physiologically similar to human NHL and the various treatment modalities can be interchanged with exception of antibody-based chemotherapies. Canine NHL represents 25% of all canine cancers diagnosed in the United States among the more than one million pet dogs diagnosed with cancer annually. As in humans, the common presentation is Stage III or Stage IV disease. It is rarely curable in the out-bred dog and even with aggressive chemotherapy the median survival rate is less than a year. Current standard-of-care (SOC) combination chemotherapy, CHOP, induces remission in approximately 85% of dogs. Thus, the companion canine provides a model for human T-cell therapy and addresses an unmet need of dogs diagnosed with NHL. Our initial canine trial (Sci Rep. 2012;2:249) demonstrated a survival advantage upon reconstitution of the immune system with autologous ex vivo-activated CD3+CD8+ T cells after CHOP for pet dogs with NHL. These T cells were propagated on engineered artificial antigen presenting cells (aAPC), which are also used for our human T-cell trials. Our updated survival data shows significant improvement in overall survival (p = 0.01) and tumor-free survival (p = 0.0008) compared with matched historical controls (Figure). We also report initial data on an additional trial infusing donor- or patient-derived canine T cells after allogeneic or autologous hematopoietic stem-cell transplantation (HSCT), respectively. The first allogeneic and autologous recipients with NHL safely received 2 escalating doses of activated T cells and achieved an anti-tumor response. As in our first trial, granzyme B expression in the propagated T cells continues to significantly correlate with prognosis. A third trial tests whether steroid-resistant T cells can persist in pet dogs with NHL receiving systemic dosing of corticosteroids. Fluorescent dyes were used to distinguish infused T cells and demonstrated that steroid-resistant activated T cells survived compared to infused control T cells. In all trials, single-cell studies are underway to determine genetic factors concordant with the survival of infused T cells and treated companion canines. Global peripheral blood T-cell gene signatures, as quantitated by nanostring, were measured at 3 hours, 7 to 14 days, and 35 to 42 days post infusion. The different gene patterns between infused dogs and post infusion time points may reflect changes in en vivo T-cell activation and companion canine prognosis. In conclusion, the add-back of activated CD8+T cells in dogs with NHL can inform of the immunobiology of adoptive transfer of T cells in humans. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Emergency reversal of anticoagulation with vitamin K antagonists with 3-factor prothrombin complex concentrates in patients with major bleeding

Major bleeding is a serious and potentially fatal complication of treatment with vitamin K antagonists (VKAs). Prothrombin complex concentrates (PCCs) can substantially shorten the time needed to reverse VKA effects. To determine the efficacy and safety of 3-factor PCCs for the rapid reversal of VKAs in patients with major bleeding. Patients receiving VKAs and suffering from acute major bleeding were eligible for this prospective cohort study if their international normalized ratio (INR) was higher than or equal to 2.0. Stratified 35-50IUkg(-1) PCC doses were infused based on initial INR. A total of 126 patients (62 males; mean age: 74years, range 37-96years) were enrolled. The mean INR at presentation was 3.3 (range 2-11). At 30min after PCC administration the mean INR was 1.4 (range: 0.9-3.1), declining to less than or equal to 1.5 in 75% of patients. The benefit of PCC was maintained for a long time, since in 97% of all post-infusion time points through 96h the mean INR remained lower than or equal to 1.5 (mean: 1.19; range: 0.9-2.3). During hospitalization neither thrombotic complications nor significant adverse events were observed and 12 patients died (10%); none of the deaths was judged to be related to PCC administration. 3-factor PCC administration is an effective, rapid ad safe treatment for the urgent reversal of VKAs in patients with acute major bleeding. Broader use of PCC in this clinical setting appears to be appropriate.

Phase 1 Clinical Trial of Autologous Mesenchymal Stem Cell Transplantation in Multiple Sclerosis (S30.002)

Objective: To describe the design and progress of an ongoing Phase 1 group-sequential trial of autologous mesenchymal stem cell (MSC) transplantation in multiple sclerosis (MS). Background MSCs have potent immunomodulatory, tissue-protective, and repair-promoting properties in vitro and in animal models. Clinical trials support the safety and efficacy of MSC transplantation in a number of human conditions. Published experience in MS is modest. Design/Methods: 24 participants with relapsing forms of MS, Expanded Disability Status Scale (EDSS) 3.0-6.5, clinical or MRI activity in the prior 2 years, and optic nerve involvement will be enrolled. Bone-marrow-derived MSCs are culture-expanded in DMEM / 10% fetal bovine serum / 10 ng/ml human fibroblast growth factor-2 then cryopreserved. After confirmation of release criteria, 1-2x10^6 MSCs/kg are administered IV. The primary outcomes are feasibility, safety, and tolerability with Data Safety Monitoring Committee review after every 4 participants. Exploratory efficacy endpoints comparing 2 months pre-infusion and 6 months post-infusion include relapses, disability (EDSS, MS Functional Composite, low-contrast letter acuity), MRI (T2 lesions, T1 lesions, gadolinium-enhancing lesions, whole brain and gray matter atrophy, diffusion tensor imaging, and magnetization transfer imaging), optical coherence tomography, visual evoked potentials, and patient self-reported health status. Ancillary immunologic mechanistic studies utilizing peripheral blood mononuclear cells (PBMCs) isolated at several pre- and post-infusion timepoints assess lymphocyte subsets by flow cytometry, T-cell responses to myelin antigens, and T-cell and B-cell regulatory functions. In vitro studies assess the mechanisms of MSC-induced immunomodulation and compare MSC-PBMC interactions in MS patients vs. controls. Results: 4 participants have been infused to date. Culture duration was 21-41 days, yielding cell doses for administration of 2x10^6 MSCs/kg (n=2) and 1.5x10^6 MSCs/kg (n=2). MSC cultures for 2 additional participants are in progress. Conclusions: Target enrollment by April 2012 is 10-12 participants. Updated study status will be presented. Supported by: Department of Defense, National Institutes of Health, Caroline Hurwitz Fund. Disclosure: Dr. Cohen has received personal compensation for activities with Biogen Idec, Eli Lilly & Company, Novartis, and Vaccinex. Dr. Cohen has received research support for activities with Biogen Idec, BioMS, Genzyme Corporation, Novartis, Synthon, and Teva Neuroscience. Dr. Bar-Or has received personal compensation for activities with Aventis Pharmaceuticals, Bayhill Therapeutics, Biogen Idec, Berlex Laboratories, Eli Lilly & Company, Genentech, Inc., GlaxoSmithKline, Ono Pharmaceutical, Diogenix, Roche Diagnostics Corporation, Merck Serono, Novartis, Teva Neuroscience. Dr. Bermel has received personal compensation for activities with Astellas, Biogen Idec, Novartis, and Teva Neuroscience as a consultant and/or speaker. Dr. Fisher has received personal compensation for activities with Biogen Idec. Dr. Fisher has received research support from Biogen Idec and Genzyme. Dr. Fox has received personal compensation for activities with Avanir, Biogen Idec, EMD Serono, and Novartis. Dr. Fox has received research support from Biogen Idec and Genentech, Inc. Dr. Gerson has received (royalty or license fee or contractual rights) payments from Osiris Therapeutics. Dr. Imrey has nothing to disclose. Dr. Lazarus has nothing to disclose. Dr. Planchon has nothing to disclose. Dr. Reese has nothing to disclose. Dr. Schwanger has nothing to disclose. Dr. Skaramagas has nothing to disclose.

Emergency reversal of anticoagulation with a three-factor prothrombin complex concentrate in patients with intracranial haemorrhage.

Intracranial haemorrhage is a serious and potentially fatal complication of oral anticoagulant therapy. Prothrombin complex concentrates can substantially shorten the time needed to reverse the effects of oral anticoagulants. The aim of this study was to determine the efficacy and safety of a prothrombin complex concentrate for rapid reversal of oral anticoagulant therapy in patients with intracranial haemorrhage. Patients receiving oral anticoagulant therapy and suffering from acute intracranial haemorrhage were eligible for this prospective cohort study if their International Normalised Ratio (INR) was higher than or equal to 2.0. The prothrombin complex concentrate was infused at doses of 35-50 IU/kg, stratified according to the initial INR. Forty-six patients (25 males; mean age: 75 years; range 38-92 years) were enrolled. The median INR at presentation was 3.5 (range, 2-9). At 30 minutes after administration of the prothrombin complex concentrate, the median INR was 1.3 (range, 0.9-3), and the INR then declined to less than or equal to 1.5 in 75% of patients. The benefit of the prothrombin complex concentrate was maintained for a long time, since the median INR remained lower than or equal to 1.5 (median, 1.16; range, 0.9-2.2) at 96% of all post-infusion time-points up to 96 hours. No thrombotic complications or significant adverse events were observed during hospitalisation; six patients (13%) died, but none of these deaths was judged to be related to administration of the prothrombin complex concentrate. Prothrombin complex concentrates are an effective, rapid and safe treatment for the urgent reversal of oral anticoagulation in patients with intracranial haemorrhage. Broader use of prothrombin complex concentrates in this clinical setting appears to be appropriate.

Use of a Population Pharmacokinetic Model of ADVATE In Pediatric and Adult Patients with Hemophilia A Permits Limited Blood Sampling for Individual Dose Tailoring.

AbstractAbstract 1416 Introduction:A population pharmacokinetic (PK) model of a recombinant FVIII (rFVIII) was established on ADVATE® (Antihemophilic Factor (Recombinant), Plasma/Albumin-Free Method) studies in pediatric and adult patients with hemophilia A. The objective of this analysis was to evaluate the effect of reduced PK sampling time points on the estimated PK parameters in the population PK model. Patients and Methods:Plasma FVIII activity PK data were collected for 3 ADVATE® clinical trials in previously treated patients: 184 full PK data sets (11 time points) for 100 adults/adolescents, aged 10 to 65 years, and from 52 reduced sample PK data sets (5 time points) for 52 children, aged 1 to 6 years. A population PK analysis was conducted on a two-compartment structure model and the covariate effect of age and weight was explored.Four reduced sampling scenarios from the full 10 post-infusion sampling time points, were investigated: 1) Reduced to 4 (1 hr, 9 hr, 24 hr, and 48 hr), 2) Reduced to 3 (6 hr, 24 hr, and 48 hr), 3) Reduced to 2 (6 hr and 24 hr), and 4) Reduced to 1 sampling time points (24 hr post-infusion). After applying the reduced sampling on a random 10% of sampling set at a time in the population PK model, the differences in model estimates and individual PK estimates between full and reduced sampling, were evaluated. Results:The two-compartment population PK model adequately described the data. Clearance (CL) was significantly correlated with age and body weight and central volume of distribution was also related with body weight.Absolute deviations (%) from the estimates using full PK sampling in the Individual PK estimates (CL, Vss, and Half-life) using each of the reduced sampling time points were showed in the below table.Median Absolute Deviation % from Full SamplingReduced to 4Reduced to 3Reduced to 2Reduced to 1Clearance [mL/(Kg*hr)]2.793.153.065.34Volume [dL/kg]2.233.883.944.20Half-life [hr]2.673.754.034.43 Conclusions:It appears that PK parameters estimated using population PK model are robust to reduced sampling time points. Accurate measurement of PK on reduced samples gives patients and clinicians the opportunity to design treatment regimens that are better tailored to individuals. Disclosures:Oh:Baxter: Employment. Björkman:Baxter: Consultancy; Octapharma: Consultancy. Schroth:Baxter: Employment. Fritsch:Baxter: Employment. Collins:NovoNordisk: Consultancy, Honoraria, The EACH2 registry was funded by Novonordisk; Baxter Healthcare: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Fischer:Baxter: Consultancy; NovoNordisk: Consultancy. Blanchette:Bayer: Consultancy; Baxter: Research Support. Casey:Baxter: Employment. Spotts:Baxter: Employment. Ewenstein:Baxter Bioscience: Employment.

Field trial on glucose-induced insulin and metabolite responses in Estonian Holstein and Estonian Red dairy cows in two herds

BackgroundInsulin secretion and tissue sensitivity to insulin is considered to be one of the factors controlling lipid metabolism post partum. The objective of this study was to compare glucose-induced blood insulin and metabolite responses in Estonian Holstein (EH, n = 14) and Estonian Red (ER, n = 14) cows.MethodsThe study was carried out using the glucose tolerance test (GTT) performed at 31 ± 1.9 days post partum during negative energy balance. Blood samples were obtained at -15, -5, 5, 10, 20, 30, 40, 50 and 60 min relative to infusion of 0.15 g/kg BW glucose and analysed for glucose, insulin, triglycerides (TG), non-esterified fatty acids (NEFA), cholesterol and β-hydroxybutyrate (BHB). Applying the MIXED Procedure with the SAS System the basal concentration of cholesterol, and basal concentration and concentrations at post-infusion time points for other metabolites, area under the curve (AUC) for glucose and insulin, clearance rate (CR) for glucose, and maximum increase from basal concentration for glucose and insulin were compared between breeds.ResultsThere was a breed effect on blood NEFA (P < 0.05) and a time effect on all metabolites concentration (P < 0.01). The following differences were observed in EH compared to ER: lower blood insulin concentration 5 min after glucose infusion (P < 0.05), higher glucose concentration 20 (P < 0.01) and 30 min (P < 0.05) after infusion, and higher NEFA concentration before (P < 0.01) and 5 min after infusion (P < 0.05). Blood TG concentration in ER remained stable, while in EH there was a decrease from the basal level to the 40th min nadir (P < 0.01), followed by an increase to the 60th min postinfusion (P < 0.01).ConclusionOur results imply that glucose-induced changes in insulin concentration and metabolite responses to insulin differ between EH and ER dairy cows.

Prothrombin Complex Concentrates for Urgent Anticoagulation Reversal in Patients with Intracranial Haemorrhage

Background: Intracranial haemorrhage (ICH) is a serious and potentially fatal complication of oral anticoagulant therapy (OAT). Prothrombin complex concentrates (PCCs) produce a rapid and effective reversal of OAT effects, but little evidence exists on their efficacy and safety in the management of ICH in patients on OAT. Aim:To evaluate the efficacy and safety of PCCs for the rapid reversal of OAT in patients with ICH. Methods: Patients suffering from acute ICH while receiving OAT were eligible for this prospective cohort study if their international normalized ratio (INR) was ≥2.0. Stratified 35–50 IU kg^–1 PCC doses were infused based on initial INR. Results: A total of 92 patients (50 males; mean age 74 years, range 34–92 years) were included. The median INR at presentation was 3.3 (range 2–9). At 30 min after PCC administration the median INR was 1.4 (range 0.9–3.1), declining to ≤1.5 in 75% of patients. The benefit of PCC was maintained for a long time, since in 98% of all post-infusion time points through 96 h the median INR remained ≤1.5 (median 1.19; range 0.9–2.3). During hospitalization neither thrombotic complications nor significant adverse events were observed and 11 patients died (11.9%). None of the deaths was judged to be related to PCC administration. Conclusions: PCC administration is an effective, rapid and safe treatment for the urgent reversal of OAT in patients with ICH. Broader use of PCC in this clinical setting appears to be appropriate.

Retrovirus Insertion Site Analysis Following In Vivo Drug Selection in a Phase I Clinical Trial Utilizing G156A MGMT for Enhanced Chemotherapy of Advanced Malignancies.

The G156A mutant of the DNA repair gene O6-methylguanine DNA-methyltransferase (MGMT) confers hematopoietic resistance to O6-benzylguanine (BG) combined with DNA-alkylating agents BCNU or temozolomide, and allows for selective in vivo expansion with drug administration of murine hematopoietic progenitors transduced with G156A MGMT retrovirus. Here we report our latest findings on retroviral vector copy number and insertion site analysis following drug treatment from a Phase I clinical trial utilizing MGMT-mediated chemoprotection for enhanced treatment of advanced solid tumors. Seven patients have entered the trial and 6 have completed the cell infusion process. For all patients, autologous CD34+ cells were transduced ex vivo with an MFG retroviral vector containing the G156A MGMT gene (packaged with PG13 by the National Gene Vector Laboratory, Ken Cornetta, Director) in the presence of the fibronectin fragment CH-296 and the cytokines SCF, Tpo, and Flt-3 ligand for 72 hours with three additions of retroviral supernatant. At 72 hours following patient treatment with BG and BCNU, cells were re-infused. Prior to infusion, the average vector copy number by quantitative real-time PCR analysis for six patients was 0.34 copies per genome, with an average of 24% of CFUs transduced by standard PCR for G156A MGMT, and an average of 9% of CD34+ cells expressing the MGMT transgene by flow cytometry. In one patient with metastatic melanoma we have further analysis of insertions. For this patient, the pre-infusion vector copy number of the bulk CD34+ population was 0.54 copies per genome by real-time PCR, with 27% of CFUs transduced and 8% of CD34+ cells expressing the MGMT transgene prior to infusion. Linear amplification-mediated (LAM)-PCR analysis of retroviral insertion sites in pre-infusion CFUs from this patient confirmed a polyclonal population, with an average of 1.6 retroviral insertions per positive CFU. In this patient, BG (120 mg/m2) and BCNU (33 mg/m2) were administered at 6 weeks post-infusion, and temozolomide (300 mg/day for 5 days) was administered at 13 weeks. Peripheral blood (PB) and bone marrow (BM) granulocyte and mononuclear cells (MNCs) were collected at weeks 5, 11, 15, and 16 for DNA and CFU analysis. Vector copy number at all post-infusion time points was below the limit of detection of SYBR Green probe-based real-time PCR (<100 copies of G156A MGMT per 5000 genomes). LAM-PCR detected the vector in post-treatment samples based on an internal vector control band present in BM MNCs at week 11 and in BM granulocytes at week 16, although specific insertion sites were not detected. Standard PCR revealed 1 out of 100 CFUs from week 11 BM MNCs contained the vector, with 2 out of 30 CFUs from week 15 PB MNCs. LAM-PCR in a subset of week 11 CFUs confirmed a single insertion site present in the same PCR-positive CFU. Sequence analysis of clonal vector insertions pre- and post-infusion is ongoing, and thus far a number of sites have been characterized, adding to the emerging database of clinical retroviral insertions. These are the first data to show emergence of transduced mutant MGMT cells after nonmyeloablative conditioning in humans and suggest that despite a low frequency of vector-marked hematopoietic cells, clinical in vivo drug selection can be observed.

Expression of therapeutic levels of factor VIII in hemophilia A mice using a novel adeno/adeno-associated hybrid virus.

We have generated an E1a/E1b/E3-deleted adeno/adeno-associated (Ad/AAV) hybrid virus driven by a small nuclear RNA (pHU1-1) promoter for expression of a B domain-deleted (Thr761-Asn1639) factor VIII transgene (FVIIIDelta761-1639). Productive replication of Ad/AAV/FVIIIDelta761-1639 in AAV rep-expressing cells resulted in generation of monomeric and dimeric mini-adenoviral (mAd) replicative forms that retained the AAV integration elements (mAd/FVIIIDelta761-1639). In vitro studies using Ad/AAV/FVIIIDelta761-1639 generated approximately 2-logs greater FVIII activity than mAd/FVIIIDelta761-1639. To determine its capacity for in vivo excision and/or genomic integration, Ad/AAV/FVIIIDelta761-1639 was injected by tail vein into three groups of hemophilia A mice (2 x 10(11) vp [n = 3]; 4 x 10(11) vp [n = 3]; 8 x 10(11) vp [n = 3]), with clear concentration-dependent increase in FVIII activity (range 160-510 mU/ml; plasma activity 16%-51% of normal). Peak activity was seen by Day (D) 5, with slow return to baseline by D28 (0.1-0.9% activity); in only 3/9 mice was loss of FVIII activity associated with development of anti-FVIII antibodies. Quantitative-PCR using genomic DNA isolated from D28 liver, spleen, heart, lungs, and kidney demonstrated the highest concentration in liver (approximately 10 genomes/cell), with little to no organ toxicity at early (D5 or 6) or late (D28) post-infusion time points. There was no evidence for spontaneous transgene excision or genomic integration in vivo as evaluated by quantitative PCR and genomic blotting. These data establish (i) the feasibility and applicability of developing high-titer Ad/AAV hybrid viruses for FVIII delivery using a small cellular promoter, (ii) the potential utility of this virus for generation of "gutted" monomeric and dimeric mAD/FVIII retaining AAV integration elements, and (iii) that the development of strategies for regulated Rep68/78 co-expression may provide a novel approach for excision, integration, and long-term FVIII transgene expression.

Pharmacodynamic profile of short-term readministration of abciximab in healthy subjects

Objective The purpose of the study was to establish a rebolus regimen for abciximab that restores pharmacologic glycoprotein (GP) IIb/IIIa receptor blockade within a short time frame (up to 48 hours) after completion of an initial treatment. Methods and Results The study was a single-center, nonrandomized, open-label dose escalation trial in healthy volunteers (n = 30). Each subject received a 0.25 mg/kg bolus and a 0.125 μg/kg per minute infusion of abciximab, followed by incremental bolus doses of the agent at 15-minute intervals up to 48 hours (10 per group) after completion of the infusion, (maximal cumulative rebolus dose of 0.25 mg/kg). Pharmacodynamic measurements (GP IIb/IIIa receptor blockade, turbidimetric and whole blood platelet aggregation with use of a rapid platelet function assay [RPFA]) were obtained at periodic intervals during and after administration of the abciximab bolus and infusion. At the time of the first rebolus, pharmacodynamic measurements were attained immediately before administration of each rebolus and 15 minutes after the last rebolus dose. In subjects who received reboluses 12 hours after infusion, a cumulative dose of 0.05 mg/kg restored >80% blockade of GP IIb/IIIa receptors and >80% inhibition of turbidimetric (5 and 20 μmol/L adenosine diphosphate) and RPFA aggregation in 10 of 10 subjects. At 24 hours after treatment, a cumulative abciximab bolus dose of 0.1 mg/kg restored >80% blockade of all 4 pharmacodynamic measurements in 10 of 10 subjects. At 48 hours after treatment, a cumulative bolus dose of 0.15 mg/kg restored >80% blockade of all 4 pharmacodynamic measurements in 10 of 10 subjects. Conclusions A fraction of the bolus of abciximab restored pharmacologic (>80%) GP IIb/IIIa receptor blockade when readministered at various postinfusion time points. These observations suggest that in the setting where acute readministration of abciximab is required less than a full bolus dose of the agent is warranted. (Am Heart J 2002;143:87-94.)