Absence of pharmacodynamic interaction between inclacumab and heparin in healthy smokers.

Inclacumab is a novel monoclonal antibody directed against P-selectin in development for the prevention and treatment of atherosclerotic cardiovascular diseases. It is likely to be used concomitantly with heparin in patients undergoing percutaneous coronary intervention. Coadministration of both drugs may potentially increase the bleeding risk associated with heparin. This crossover study evaluated the potential pharmacodynamic interaction between inclacumab and unfractionated heparin in 18 healthy smokers. Owing to the long elimination of inclacumab (half-life of approximately 18 days), a 2-period, one-sequence study design was used. Subjects received an intravenous bolus injection of unfractionated heparin (5000 IU) on days 1 and 8 and an intravenous infusion of inclacumab (20 mg/kg) on day 8. Blood samples were collected on days 1 and 8 for pharmacodynamic effects of unfractionated heparin (anti-FXa and anti-FIIa activities, activated partial thromboplastin time and tissue factor pathway inhibitor) and over 6 months for pharmacokinetics of inclacumab. Sixteen subjects were eligible for pharmacodynamic analysis. Inclacumab had no clinically significant pharmacodynamic interaction with unfractionated heparin. With the exception of the minor but statistically significant increase of the maximum effect [Emax] of anti-FIIa activity, pharmacodynamic parameters (areas under the effect curve [AUElast] and Emax of anti-FXa) were almost similar on days 1 and 8. The 90% confidence intervals of geometric mean ratios of day 8 to day 1 for AUElast and Emax were however all contained within bioequivalence boundaries. The data demonstrate that the anticoagulant effect of unfractionated heparin was not affected by the administration of inclacumab.

First-in-Man Study With Inclacumab, a Human Monoclonal Antibody Against P-selectin.

Inclacumab, a novel monoclonal antibody against P-selectin in development for the treatment and prevention of atherosclerotic cardiovascular diseases, was administered in an ascending single-dose study as intravenous infusion to evaluate safety, pharmacokinetics, and pharmacodynamics. Fifty-six healthy subjects were enrolled in this randomized, double-blind placebo-controlled study. Each dose level (0.03-20 mg/kg) was investigated in separate groups of 8 subjects (6 on inclacumab, 2 on placebo). Platelet-leukocyte aggregates, free/total soluble P-selectin concentration ratio, drug concentrations, bleeding time, platelet aggregation, antibody formation, and routine laboratory parameters were measured frequently until 32 weeks. Pharmacokinetic profiles were indicative of target-mediated drug disposition. Platelet-leukocyte aggregate inhibition and soluble P-selectin occupancy showed dose dependency and were strongly correlated to inclacumab plasma concentrations, with IC50 of 740 and 4600 ng/mL, respectively. Inclacumab was well tolerated by the majority of subjects and did neither affect bleeding time nor platelet aggregation. These findings allowed the investigation of the potential beneficial therapeutic use of inclacumab in patient study.

Preclinical characterization of XB002, an anti-tissue factor antibody-drug conjugate for the treatment of solid tumors.

Tissue factor (TF) is overexpressed in various cancers, where its expression is generally associated with poor disease outcomes. XB002 is an anti-TF antibody-drug conjugate designed to deliver a cytotoxic payload to TF-expressing tumors while minimizing adverse events related to disruption of TF function, notably bleeding. XB002 is composed of a zovodotin linker-payload conjugated to a monoclonal antibody (clone 25A3) that binds to TF with high affinity (KD = 0.86 nM). In vitro coagulation studies indicated that 25A3 did not interfere with the clotting cascade; at a 100 nM concentration, 25A3 had no effect on activation of coagulation factor X or thrombin generation. XB002 was internalized in TF-expressing cancer cell lines and displayed potent cytotoxic activity at sub-nanomolar concentrations. When evaluated in the HPAF-II xenograft model, XB002 (1.5 mg/kg, IV) given once weekly for 2 weeks induced complete regression with no tumor growth even at 5 weeks after the second dose. In murine patient-derived xenograft models, a single dose of XB002 (10 mg/kg, IV) inhibited tumor growth across multiple cancer models including bladder, cervical, gastric, head and neck squamous cell carcinoma (HNSCC), and non-small cell lung cancer. Further, complete tumor regression was observed in both the cervical and HNSCC models by 30 days post-treatment. In non-human primate models, XB002 showed exposure in the desired range and no evidence of bleeding or neutropenia. Taken together, these data demonstrate potential anti-tumor activity across a spectrum of oncology indications and strongly support its clinical development.

Randomized Phase II Trial Comparing Bevacizumab Plus Carboplatin and Paclitaxel With Carboplatin and Paclitaxel Alone in Previously Untreated Locally Advanced or Metastatic Non-Small-Cell Lung Cancer.

PURPOSE: To investigate the efficacy and safety of bevacizumab plus carboplatin and paclitaxel in patients with advanced or recurrent non-small-cell lung cancer. PATIENTS AND METHODS: In a phase II trial, 99 patients were randomly assigned to bevacizumab 7.5 (n = 32) or 15 mg/kg (n = 35) plus carboplatin (area under the curve = 6) and paclitaxel (200 mg/m2) every 3 weeks or carboplatin and paclitaxel alone (n = 32). Primary efficacy end points were time to disease progression and best confirmed response rate. On disease progression, patients in the control arm had the option to receive single-agent bevacizumab 15 mg/kg every 3 weeks. RESULTS: Compared with the control arm, treatment with carboplatin and paclitaxel plus bevacizumab (15 mg/kg) resulted in a higher response rate (31.5% v 18.8%), longer median time to progression (7.4 v 4.2 months) and a modest increase in survival (17.7 v 14.9 months). Of the 19 control patients that crossed over to single-agent bevacizumab, five experienced stable disease, and 1-year survival was 47%. Bleeding was the most prominent adverse event and was manifested in two distinct clinical patterns; minor mucocutaneous hemorrhage and major hemoptysis. Major hemoptysis was associated with squamous cell histology, tumor necrosis and cavitation, and disease location close to major blood vessels. CONCLUSION: Bevacizumab in combination with carboplatin and paclitaxel improved overall response and time to progression in patients with advanced or recurrent non-small-cell lung cancer. Patients with nonsquamous cell histology appear to be a subpopulation with improved outcome and acceptable safety risks.

Rituximab pediatric drug development: Pharmacokinetic and pharmacodynamic modeling to inform regulatory approval for rituximab treatment in patients with granulomatosis with polyangiitis or microscopic polyangiitis.

Anti-neutrophil cytoplasmic antibody-associated vasculitides granulomatosis with polyangiitis (GPA) and microscopic polyangiitis (MPA) are rare, potentially organ- and life-threatening autoimmune conditions affecting adult and pediatric patients. An open-label phase II study was conducted to determine safe and effective dosing regimens of rituximab in pediatric patients with GPA/MPA. To determine the selection of an appropriate dose regimen in children for induction and maintenance, a population pharmacokinetic approach was used (nonlinear mixed-effect modeling), combining pediatric data with data from adults with GPA/MPA. The time course of B-cell depletion was assessed in both populations. The exposure-effect relationship was assessed by logistic regression. Twenty-five pediatric patients (80% female patients; age range, 6-17 years) were enrolled in the trial and received the induction regimen of intravenous rituximab 375 mg/m2 weekly for 4 weeks, which resulted in a similar exposure to that of adults. Based on pharmacokinetic modeling, a maintenance dosing regimen of 250 mg/m2 administered twice over 14 days followed by 250 mg/m2 every 6 months is expected to result in similar rituximab exposure as that of adults receiving the approved maintenance dose of 500 mg administered twice over 14 days followed by 500 mg every 6 months. The time course of B-cell depletion was similar between the pediatric and adult populations, supporting the similarities in response in both populations and allowing extrapolation to patients less than 6 years old. Using a partial extrapolation approach helped identify safe and effective dosing regimens of rituximab in pediatric patients with GPA/MPA and lead to regulatory approval.

Phase IIa Global Study Evaluating Rituximab for the Treatment of Pediatric Patients With Granulomatosis With Polyangiitis or Microscopic Polyangiitis.

OBJECTIVE: To assess the safety, tolerability, pharmacokinetics, and efficacy of rituximab (RTX) in pediatric patients with granulomatosis with polyangiitis (GPA) or microscopic polyangiitis (MPA). METHODS: The Pediatric Polyangiitis Rituximab Study was a phase IIa, international, open-label, single-arm study. During the initial 6-month remission-induction phase, patients received intravenous infusions of RTX (375 mg/m2 body surface area) and glucocorticoids once per week for 4 weeks. During the follow-up period, patients could receive further treatment, including RTX, for GPA or MPA. The safety, pharmacokinetics, pharmacodynamics, and exploratory efficacy outcomes with RTX were evaluated. RESULTS: Twenty-five pediatric patients with new-onset or relapsing disease were enrolled at 11 centers (19 with GPA [76%] and 6 with MPA [24%]). The median age was 14 years (range 6-17 years). All patients completed the remission-induction phase. During the overall study period (≤4.5 years), patients received between 4 and 28 infusions of RTX. All patients experienced ≥1 adverse event (AE), mostly grade 1 or grade 2 primarily infusion-related reactions. Seven patients experienced 10 serious AEs, and 17 patients experienced 31 infection-related AEs. No deaths were reported. RTX clearance correlated with body surface area. The body surface area-adjusted RTX dosing regimen resulted in similar exposure in both pediatric and adult patients with GPA or MPA. Remission, according to the Pediatric Vasculitis Activity Score, was achieved in 56%, 92%, and 100% of patients by months 6, 12, and 18, respectively. CONCLUSION: In pediatric patients with GPA or MPA, RTX is well tolerated and effective, with an overall safety profile comparable to that observed in adult patients with GPA or MPA who receive treatment with RTX. RTX is associated with a positive risk/benefit profile in pediatric patients with active GPA or MPA.

Clinical pharmacokinetics of bevacizumab in patients with solid tumors.

OBJECTIVE: To characterize the population pharmacokinetics of bevacizumab and the influence of demographic factors, disease severity, and concomitantly used chemotherapy agents on it's pharmacokinetic behavior. PATIENTS AND METHODS: Data from eight clinical trials with bevacizumab administered by intravenous infusion were included. A total of 4,629 bevacizumab concentrations from 491 patients with solid tumors, who received bevacizumab doses ranging from 1 to 20 mg/kg at a dosing frequency ranging from weekly to every 3 weeks, were analyzed using a nonlinear mixed-effects modeling approach (NONMEM). RESULTS: The best structural model was a two-compartment model with first-order elimination. In the final model, estimated clearance (CL) and central compartment volume of distribution (Vc) were 0.207 L/day and 2.39 L for a typical female. The terminal half-life estimate was approximately 20 days for both men and women. Body weight and gender were the most significant covariates to explain interpatient variability for CL and Vc. Clearance was 26% faster in men than in women. Patients with low serum albumin and high serum alkaline phosphatase had 19 and 23% faster CL, respectively, than a typical patient. Consistent with the long elimination half life, simulations showed that similar steady-state exposures can be maintained when the weekly mg/kg dose rate is maintained, therefore allowing administration of bevacizumab to coincide with the frequency of administration of the cytotoxic agents. CONCLUSION: The PK parameters were consistent with those of other IgG molecules. The results support dosing bevacizumab on a once every 2 weeks or once every 3 weeks dosing schedule on a mg/kg basis.

Randomized phase II trial comparing bevacizumab plus carboplatin and paclitaxel with carboplatin and paclitaxel alone in previously untreated locally advanced or metastatic non-small-cell lung cancer.

PURPOSE: To investigate the efficacy and safety of bevacizumab plus carboplatin and paclitaxel in patients with advanced or recurrent non-small-cell lung cancer. PATIENTS AND METHODS: In a phase II trial, 99 patients were randomly assigned to bevacizumab 7.5 (n = 32) or 15 mg/kg (n = 35) plus carboplatin (area under the curve = 6) and paclitaxel (200 mg/m(2)) every 3 weeks or carboplatin and paclitaxel alone (n = 32). Primary efficacy end points were time to disease progression and best confirmed response rate. On disease progression, patients in the control arm had the option to receive single-agent bevacizumab 15 mg/kg every 3 weeks. RESULTS: Compared with the control arm, treatment with carboplatin and paclitaxel plus bevacizumab (15 mg/kg) resulted in a higher response rate (31.5% v 18.8%), longer median time to progression (7.4 v 4.2 months) and a modest increase in survival (17.7 v 14.9 months). Of the 19 control patients that crossed over to single-agent bevacizumab, five experienced stable disease, and 1-year survival was 47%. Bleeding was the most prominent adverse event and was manifested in two distinct clinical patterns; minor mucocutaneous hemorrhage and major hemoptysis. Major hemoptysis was associated with squamous cell histology, tumor necrosis and cavitation, and disease location close to major blood vessels. CONCLUSION: Bevacizumab in combination with carboplatin and paclitaxel improved overall response and time to progression in patients with advanced or recurrent non-small-cell lung cancer. Patients with nonsquamous cell histology appear to be a subpopulation with improved outcome and acceptable safety risks.

Preclinical pharmacokinetics of Ranibizumab (rhuFabV2) after a single intravitreal administration.

PURPOSE: Ranibizumab (rhuFab V2; Lucentis, Genentech, South San Francisco, CA) is a humanized monoclonal antibody fragment designed to bind all forms of VEGF, thereby blocking vessel permeability and angiogenesis in neovascular age-related macular degeneration. This study evaluated the pharmacokinetic (PK) and serum bioavailability of ranibizumab after a single intravitreal (ITV) or intravenous (IV) dose in cynomolgus monkeys. METHODS: Monkeys received ranibizumab as either a bilateral ITV dose (500 or 2000 microg/eye; n = 6/group) or a single IV dose (1000 or 4000 microg/animal; n = 4/group). After ITV administration, ranibizumab concentrations were measured in several ocular compartments and in serum for 10 days and, after IV administration, for 48 hours. Pharmacokinetic parameters were estimated by compartmental and noncompartmental methods. RESULTS: Ranibizumab cleared in parallel from all ocular compartments, with a terminal half-life of approximately 3 days. It distributed rapidly to the retina (6-24 hours), and concentrations were approximately one third that in the vitreous. After ITV injection, bioavailability (F) was 50% to 60%. Serum concentrations were very low, reflecting wider distribution and faster clearance when ranibizumab reached the serum. After IV administration, the terminal half-life was approximately 0.5 day. CONCLUSIONS: This study demonstrates that ranibizumab has a PK profile that is favorable for its clinical use in treating neovascular AMD by monthly ITV injection.

Prevention of experimental choroidal neovascularization with intravitreal anti-vascular endothelial growth factor antibody fragment.

OBJECTIVE: To evaluate the safety and efficacy of intravitreal injections of an antigen-binding fragment of a recombinant humanized monoclonal antibody directed toward vascular endothelial growth factor (rhuFab VEGF) in a monkey model of choroidal neovascularization (CNV). METHODS: In phase 1 of the study, each animal received intravitreal injections, 500 microg per eye, of rhuFab VEGF in one eye (prevention eye), while the contralateral eye received rhuFab VEGF vehicle (control eye) at 2-week intervals. On day 21, laser photocoagulation was performed to induce CNV. In phase 2, the vehicle-treated eye was crossed over and both eyes received 500 microg of rhuFab VEGF beginning 21 days following laser-induced injury at days 42 and 56. The eyes were monitored by ophthalmic examinations, color photographs, and fluorescein angiography. RESULTS: rhuFab VEGF did not cause any ocular hemorrhages. All eyes treated with rhuFab VEGF developed acute anterior chamber inflammation within 24 hours of the first injection that resolved within 1 week, and this inflammation was less severe with subsequent injections. The incidence of CNV, defined angiographically, was significantly lower in the prevention eyes than the control eyes (P<.001). Subsequent treatments were associated with less leakage in eyes with established CNV that were crossed over from the control eyes to the treatment eyes (P =.001). CONCLUSIONS: Intravitreal rhuFab VEGF injections prevented formation of clinically significant CNV in cynomolgus monkeys and decreased leakage of already formed CNV with no significant toxic effects. CLINICAL RELEVANCE: This study provides the nonclinical proof of principle for ongoing clinical studies of intravitreally injected rhuFab VEGF in patients with neovascular age-related macular degeneration.

Pharmacological control of platelet-leukocyte interactions by the human anti-P-selectin antibody inclacumab--preclinical and clinical studies.

BACKGROUND AND OBJECTIVE: Elevated levels of platelet-leukocyte aggregates (PLAs) have been reported in several cardiovascular diseases and suggested to contribute to disease pathology. Our aim was to characterize the effects of inclacumab, a novel human anti-P-selectin antibody, on the interactions between leukocytes and platelets in preclinical and clinical studies. EXPERIMENTAL APPROACHES: Dual-label flow cytometry was used to detect the effect of inclacumab on agonist-induced platelet-leukocyte/platelet-monocyte aggregates in cynomolgus monkeys and humans, following ex vivo and in vivo administration. Platelet-dependent leukocyte activation and leukocyte adhesion to a platelet monolayer were also investigated after ex vivo administration of inclacumab to human blood. RESULTS: Treatment of cynomolgus monkeys with inclacumab profoundly inhibited thrombin receptor-activating peptide (TRAP) or adenosine diphosphate (ADP)-induced PLAs with an IC50 (<2 µg/mL) similar to the in vitro spiking experiments. Maximal inhibition of PLAs persisted for ≥28 days following single dose of inclacumab. In human blood, inclacumab was about 2-fold more potent in inhibiting TRAP-induced PLAs (IC50: 0.7 µg/mL) compared to monkeys. PLA formation was suppressed independently of the inducing platelet agonist. Inclacumab also inhibited the activation of the leukocyte integrin Mac-1 and leukocyte adhesion to a platelet monolayer under flow conditions. In clinical studies, inclacumab inhibited TRAP-induced PLA formation in a dose-dependent manner following single and multiple dose administration to healthy volunteers. It also reduced elevated circulating PLA levels in patients with peripheral arterial disease. CONCLUSION: By inhibiting platelet-leukocyte interactions, demonstrated in multiple preclinical and clinical studies, inclacumab may provide an effective treatment for cardiovascular diseases.

Pharmacokinetics and retinal distribution of ranibizumab, a humanized antibody fragment directed against VEGF-A, following intravitreal administration in rabbits.

PURPOSE: Ranibizumab (Lucentis) is a humanized antigen-binding fragment designed to inhibit all isoforms and active degradation products of vascular endothelial growth factor A (VEGF-A); it is in clinical development for the treatment of neovascular age-related macular degeneration (AMD). This study evaluated its pharmacokinetics (PK) and retinal distribution in rabbits when administered intravitreally (ITV). METHODS: A total of 27 New Zealand white rabbits received a single bilateral ITV injection of ranibizumab 625 muicrog/eye (Group 1, n = 24) or I-labeled ranibizumab 625 microg/eye, 22.5 microCi/eye (Group 2, n = 3). Ranibizumab concentration was determined in the vitreous, aqueous humor, and serum up to 60 days postdose by enzyme-linked immunosorbent assay in Group 1. Group 2 eyes were microautoradiographed on days 1-4. RESULTS: Ranibizumab has a terminal half-life of 2.9 days in the ocular compartments. Systemic exposure was low, measuring less than 0.01% of vitreous exposure when comparing AUC0-t values. Microautoradiography analysis demonstrated that ranibizumab penetrated all retinal layers, reaching the choriocapillaris on days 1, 2, and 4. CONCLUSIONS: This study demonstrates that following ITV injection, ranibizumab has a vitreous half-life of 2.9 days with minimal systemic exposure. Ranibizumab rapidly penetrates through the retina to reach the choroid, supporting its clinical development for neovascular AMD.

Ranibizumab inhibits multiple forms of biologically active vascular endothelial growth factor in vitro and in vivo.

Neovascular age-related macular degeneration (AMD) is the leading cause of blindness in older adults in the Western world. Ranibizumab (Lucentis), a humanized antibody fragment directed against vascular endothelial growth factor (VEGF-A), was recently approved by the US Food and Drug Administration (FDA) for the treatment of neovascular AMD. The objective of this study was to characterize the binding affinity and pharmacological activity of ranibizumab for 3 biologically active forms of VEGF-A: VEGF165, VEGF121, and VEGF110. The apparent equilibrium binding affinity of ranibizumab for VEGF-A molecules was determined by Biacore analysis; the biological activity of VEGF-A was demonstrated in a human umbilical vein endothelial cell (HUVEC) proliferation-inhibition assay. Inhibition of VEGF-A-induced vascular permeability by ranibizumab was assessed in vivo using hairless guinea pigs and a modified Miles assay. Ranibizumab was capable of binding to recombinant human VEGF165, VEGF121, and VEGF110 (KD < or = 192 pM), inhibiting VEGF-A-induced HUVEC proliferation in a concentration-dependent manner. Ranibizumab also exerted potent dose-dependent inhibition (IC(50) of 0.4-1.2 nM) of the vascular permeability-enhancing activity of VEGF165, VEGF121, and VEGF110 in the Miles assay. In conclusion, these results show that ranibizumab is capable of binding to and specifically inhibiting the activities of 3 biologically active forms of VEGF-A. As VEGF-A plays a pivotal role in the pathogenesis of neovascular AMD, ranibizumab activity, as demonstrated in this study, supports its clinical utility in the treatment of this disease.

Concomitant administration of bevacizumab, irinotecan, 5-fluorouracil, and leucovorin: nonclinical safety and pharmacokinetics.

Bevacizumab (Avastin) is a humanized monoclonal antibody against vascular endothelial growth factor approved for use in combination with 5-fluorouracil (5-FU)-based chemotherapy for first-line treatment of metastatic colorectal cancer. The Saltz regimen (irinotecan/5-FU/leucovorin [LV]) is a first-line treatment for this indication. The objective of this study was to evaluate the safety of bevacizumab when administered concomitantly with the Saltz regimen to cynomolgus monkeys, and to determine if the pharmacokinetics of bevacizumab, irinotecan, SN38 (the active metabolite of irinotecan), or 5-FU were affected by combined administration. Male cynomolgus monkeys were intravenously administered the Saltz regimen (125 mg/m2 irinotecan, 500 mg/m2 5-FU, 20 mg/m2 LV) alone (n = 4) or concomitantly with 10 mg/kg bevacizumab (n = 5) on days 1 and 8. All animals survived to euthanasia on day 15. Adverse effects associated with the Saltz regimen included diarrhea and neutropenia. Macroscopically, two animals from each group had small thymus glands that correlated microscopically with lymphoid depletion. Myeloid hypoplasia and/or erythroid hyperplasia was observed in the sternal bone marrow of most animals. These effects were considered to be associated with the Saltz regimen; concomitant bevacizumab administration did not alter the severity of these findings. Irinotecan and 5-FU were observed to be rapidly eliminated (t1/2 = 1 h and 0.5 h, respectively). Although the number of animals in each group was small and no statistical comparison between groups was performed, bevacizumab did not affect the disposition of either agent. These results indicate that bevacizumab can be safely administered in combination with the Saltz regimen without pharmacokinetic interaction.

Design, construction, and in vitro analyses of multivalent antibodies.

Some Abs are more efficacious after being cross-linked to form dimers or multimers, presumably as a result of binding to and clustering more surface target to either amplify or diversify cellular signaling. To improve the therapeutic potency of these types of Abs, we designed and generated Abs that express tandem Fab repeats with the aim of mimicking cross-linked Abs. The versatile design of the system enables the creation of a series of multivalent human IgG Ab forms including tetravalent IgG1, tetravalent F(ab')2, and linear Fab multimers with either three or four consecutively linked Fabs. The multimerized Abs target the cell surface receptors HER2, death receptor 5, and CD20, and are more efficacious than their parent mAbs in triggering antitumor cellular responses, indicating they could be useful both as reagents for study as well as novel therapeutics.