Population pharmacokinetic-pharmacodynamic modeling of PB2452, a monoclonal antibody fragment being developed as a ticagrelor reversal agent, in healthy volunteers.

PB2452, a neutralizing monoclonal antibody fragment that binds the antiplatelet drug ticagrelor with high affinity, is being developed as a ticagrelor reversal agent. To identify a clinically useful intravenous (i.v.) reversal regimen, a semimechanistic exposure-response model was developed during the PB2452 first-in-human phase I study. From a randomized, double-blind, placebo-controlled, single-dose trial to evaluate the safety, efficacy, and pharmacokinetics (PKs) of PB2452 in 61 healthy volunteers pretreated with ticagrelor, sequential dose cohort data were used to build and refine an exposure-response model that combined population PK models for ticagrelor (TICA), ticagrelor active metabolite (TAM), and PB2452, and related their binding relationships to the PK of uncomplexed TICA and TAM which is predictive of platelet inhibition. Platelet function was assessed by multiple assays. The model was developed using Bayesian methods in NONMEM. Human PK and pharmacodynamic data from sequential dose cohorts were used to initially define and then refine model parameters. Model simulations indicated that an initial i.v. bolus of PB2452, followed by a high-rate infusion, and then a slower-rate infusion would provide immediate and sustained reversal of the antiplatelet effects of ticagrelor. Based on model predictions, a 6 g i.v. bolus followed by 6 g infused over 4 h and then 6 g over 12 h was identified and tested in study subjects and shown to provide complete reversal within 5 min of infusion onset that was sustained for 20-24 h. The model is predictive of the reversal profile of PB2452 and will inform future trials of PB2452.

Pharmacokinetics of Monoclonal Antibody and Antibody Fragments in the Mouse Eye Following Systemic Administration.

The ocular pharmacokinetics (PK) of antibody-based therapies are infrequently studied in mice due to the technical difficulties in working with the small murine eye. This study is the first of its kind to quantitatively measure the PK of variously sized proteins in the plasma, cornea/ICB, vitreous humor, retina, and posterior cup (including choroid) of the mouse and to evaluate the relationship between molecular weight (MW) and antibody biodistribution coefficient (BC) to the eye. Proteins analyzed include trastuzumab (150 kDa), trastuzumab-vc-MMAE (T-vc-MMAE, 155 kDa), F(ab)2 (100 kDa), Fab (50 kDa), and scFv (27 kDa). As expected, ocular PK mirrored the systemic PK as plasma was the driving force for ocular exposure. For trastuzumab, T-vc-MMAE, F(ab)2, Fab, and scFv, respectively, the BCs in the cornea/ICB were 0.610%, 0.475%, 1.74%, 3.39%, and 13.7%; the BCs in the vitreous humor were 0.0198%, 0.0427%, 0.0934%, 0.234%, and 5.56%; the BCs for the retina were 0.539%, 0.230%, 0.704%, 2.44%, and 20.4%; the BCs for the posterior cup were 0.557%, 0.650%, 1.47%, 4.06%, and 13.9%. The relationship between BC and MW was best characterized by a log-log regression in which BC decreased as MW increased, with every doubling in MW leading to a decrease in BC by a factor of 3.44 × , 6.76 × , 4.74 × , and 3.43 × in cornea/ICB, vitreous humor, retina, and posterior cup, respectively. In analyzing the disposition of protein therapeutics to the eye, these findings enhance our understanding of the potential for ocular toxicity of systemically administered protein therapeutics and may aid in the discovery of systemically administered protein therapeutics for ocular disorders.

Development of a human antibody fragment directed against the alpha folate receptor as a promising molecule for targeted application.

Alpha folate receptor (FRα) is currently under investigation as a target for the treatment of patients with non-small-cell lung cancer (NSCLC), since it is highly expressed in tumor cells but is largely absent in normal tissue. In this study, a novel human variable domain of a heavy-chain (VH) antibody fragment specific to FRα was enriched and selected by phage bio-planning. The positive phage clone (3A102 VH) specifically bound to FRα and also cross-reacted with FRß, as tested by ELISA. Clone 3A102 VH was then successfully expressed as a soluble protein in an E. coli shuffle strain. The obtained soluble 3A102 VH demonstrated a high affinity for FRα with affinity constants (Kaff) values around 7.77 ± 0.25 × 107 M-1, with specific binding against both FRα expressing NSCLC cells and NSCLC patient-derived primary cancer cells, as tested by cell ELISA. In addition, soluble 3A102 VH showed the potential desired property of a targeting molecule by being internalized into FRα-expressing cells, as observed by confocal microscopy. This study inspires the use of phage display to develop human VH antibody (Ab) fragments that might be well suited for drug targeted therapy of NSCLC and other FRα-positive cancer cells.

A Heterodimeric Antibody Fragment for Passive Immunotherapy Against Norovirus Infection.

Human noroviruses cause an estimated 685 million infections and 200 000 deaths annually worldwide. Although vaccines against GII.4 and GI.1 genotypes are under development, no information is available regarding vaccines or monoclonal antibodies to other noroviral genotypes. Here, we developed 2 variable-domain llama heavy-chain antibody fragment (VHHs) clones, 7C6 and 1E4, against GII.4 and GII.17 human noroviruses, respectively. Although 7C6 cross-reacted with virus-like particles (VLPs) of GII.17, GII.6, GII.3, and GII.4, it neutralized only GII.4 norovirus. In contrast, 1E4 reacted with and neutralized only GII.17 VLPs. Both VHHs blocked VLP binding to human induced pluripotent stem cell-derived intestinal epithelial cells and carbohydrate attachment factors. Using these 2 VHHs, we produced a heterodimeric VHH fragment that neutralized both GII.4 and GII.17 noroviruses. Because VHH fragments are heat- and acid-stable recombinant monoclonal antibodies, the heterodimer likely will be useful for oral immunotherapy and prophylaxis against GII.4 and GII.17 noroviruses in young, elderly, or immunocompromised persons.

Comparative evaluation of affibody- and antibody fragments-based CAIX imaging probes in mice bearing renal cell carcinoma xenografts.

Carbonic anhydrase IX (CAIX) is a cancer-associated molecular target for several classes of therapeutics. CAIX is overexpressed in a large fraction of renal cell carcinomas (RCC). Radionuclide molecular imaging of CAIX-expression might offer a non-invasive methodology for stratification of patients with disseminated RCC for CAIX-targeting therapeutics. Radiolabeled monoclonal antibodies and their fragments are actively investigated for imaging of CAIX expression. Promising alternatives are small non-immunoglobulin scaffold proteins, such as affibody molecules. A CAIX-targeting affibody ZCAIX:2 was re-designed with the aim to decrease off-target interactions and increase imaging contrast. The new tracer, DOTA-HE3-ZCAIX:2, was labeled with 111In and characterized in vitro. Tumor-targeting properties of [111In]In-DOTA-HE3-ZCAIX:2 were compared head-to-head with properties of the parental variant, [99mTc]Tc(CO)3-HE3-ZCAIX:2, and the most promising antibody fragment-based tracer, [111In]In-DTPA-G250(Fab')2, in the same batch of nude mice bearing CAIX-expressing RCC xenografts. Compared to the 99mTc-labeled parental variant, [111In]In-DOTA-HE3-ZCAIX:2 provides significantly higher tumor-to-lung, tumor-to-bone and tumor-to-liver ratios, which is essential for imaging of CAIX expression in the major metastatic sites of RCC. [111In]In-DOTA-HE3-ZCAIX:2 offers significantly higher tumor-to-organ ratios compared with [111In]In-G250(Fab')2. In conclusion, [111In]In-DOTA-HE3-ZCAIX:2 can be considered as a highly promising tracer for imaging of CAIX expression in RCC metastases based on our results and literature data.

Analysis of Progress and Challenges of EGFR-Targeted Molecular Imaging in Cancer With a Focus on Affibody Molecules.

Epidermal growth factor receptor (EGFR)-targeted cancer therapy requires an accurate estimation of EGFR expression in tumors to identify responsive patients, monitor therapeutic effect, and estimate prognosis. The EGFR molecular imaging is an optimal method for evaluating EGFR expression in vivo accurately and noninvasively. In this review, we discuss the recent advances in EGFR-targeted molecular imaging in cancer, with a special focus on the development of imaging agents, including epidermal growth factor (EGF) ligand, monoclonal antibodies, antibody fragments, Affibody, and small molecules. Each substrate or probe, whether it is an endogenous ligand, antibody, peptide, or small molecule labeled with fluorochrome or radionuclide, has unique advantages and limitations. Antibody-based probes have high affinity but a long metabolic cycle and therefore offer poor imaging quality. Affibody molecules promise to surpass antibody-based probes due to their small size, stable chemical properties, and high affinity to the target. Small-molecule probes are safe, have favorable pharmacokinetics, and show high affinity and specificity, in addition to having an ideal size, but are inadequate for delayed imaging after injection due to their fast clearance.

Evaluation of antibody fragment properties for near-infrared fluorescence imaging of HER3-positive cancer xenografts.

In vivo imaging is influenced by the half-life, tissue penetration, biodistribution, and affinity of the imaging probe. Immunoglobulin G (IgG) is composed of discrete domains with known functions, providing a template for engineering antibody fragments with desired imaging properties. Here, we engineered antibody-based imaging probes, consisting of different combinations of antibody domains, labeled them with the near-infrared fluorescent dye IRDye800CW, and evaluated their in vivo imaging properties. Antibody-based imaging probes were based on an anti-HER3 antigen binding fragment (Fab) isolated using phage display. Methods: We constructed six anti-HER3 antibody-based imaging probes: a single chain variable fragment (scFv), Fab, diabody, scFv-CH3, scFv-Fc, and IgG. IRDye800CW-labeled, antibody-based probes were injected into nude mice bearing FaDu xenografts and their distribution to the xenograft, liver, and kidneys was evaluated. Results: These imaging probes bound to recombinant HER3 and to the HER3-positive cell line, FaDu. Small antibody fragments with molecular weight <60 kDa (scFv, diabody, and Fab) accumulated rapidly in the xenograft (maximum accumulation between 2-4 h post injection (hpi)) and cleared primarily through the kidneys. scFv-CH3 (80 kDa) had fast clearance and peaked in the xenograft between 2-3 hpi and cleared from xenograft in a rate comparable to Fab and diabody. IgG and scFv-Fc persisted in the xenografts for up to 72 hpi and distributed mainly to the xenograft and liver. The highest xenograft fluorescence signals were observed with IgG and scFv-Fc imaging probes and persisted for 2-3 days. Conclusion: These results highlight the utility of using antibody fragments to optimize clearance, tumor labeling, and biodistribution properties for developing anti-HER3 probes for image-guided surgery or PET imaging.

Tumor uptake of pegylated diabodies: Balancing systemic clearance and vascular transport.

The accumulation, dissemination and clearance of monoclonal antibody-based therapeutics or imaging reagents targeting tumor associated antigens is governed by several factors including affinity, size, charge, and valency. Tumor targeting antibody fragments have distinct advantages over intact monoclonal antibodies such as enhanced penetration within the tumor and rapid accumulation but are subject to rapid clearance. Polyethylene glycol (PEG)-modified antibody fragments can provide a way to balance tumor penetration and accumulation with improved serum persistence. In this study, we use a diabody, the dimeric antibody fragment, targeting the 5T4 antigen to assess the impact of PEGs of distinct size and shape on tumor accumulation and pharmacokinetics (PK). We show that PEG-modified diabodies improved the PK of the parental diabody from a half-life of 40 min to over 40 h for the higher molecular weight PEG conjugated diabodies. This improvement correlates with the increasing hydrodynamic size of pegylated diabodies, and can serve as a better predictor of the PK behavior of pegylated molecules than molecular weight alone. Tumor uptake profiles determined by quantitative PET imaging differed significantly based on PEG size and shape with diabody-PEG5K showing peak accumulation early on, but with the larger diabody-PEG20K showing better sustained tumor uptake at later time points. In addition, we demonstrate that a diabody-PEG20K-B with a hydrodynamic radius (Rh) of 6 nm had superior tumor uptake than the larger diabody-PEG40K-B with Rh of 12 nm, indicating that beyond 6 nm, larger pegylated diabodies have a slower tumor uptake rate while having comparable clearance kinetics. Our data demonstrate that pegylated diabodies with Rh of ~6 nm have an optimal size and PK profile for tumor uptake. Understanding the impact of pegylation on PK and tumor uptake could facilitate the development of pegylated diabodies as therapeutics.

Anti-CTLA-4 therapy requires an Fc domain for efficacy.

Ipilimumab, a monoclonal antibody that recognizes cytotoxic T lymphocyte antigen (CTLA)-4, was the first approved "checkpoint"-blocking anticancer therapy. In mouse tumor models, the response to antibodies against CTLA-4 depends entirely on expression of the Fcγ receptor (FcγR), which may facilitate antibody-dependent cellular phagocytosis, but the contribution of simple CTLA-4 blockade remains unknown. To understand the role of CTLA-4 blockade in the complete absence of Fc-dependent functions, we developed H11, a high-affinity alpaca heavy chain-only antibody fragment (VHH) against CTLA-4. The VHH H11 lacks an Fc portion, binds monovalently to CTLA-4, and inhibits interactions between CTLA-4 and its ligand by occluding the ligand-binding motif on CTLA-4 as shown crystallographically. We used H11 to visualize CTLA-4 expression in vivo using whole-animal immuno-PET, finding that surface-accessible CTLA-4 is largely confined to the tumor microenvironment. Despite this, H11-mediated CTLA-4 blockade has minimal effects on antitumor responses. Installation of the murine IgG2a constant region on H11 dramatically enhances its antitumor response. Coadministration of the monovalent H11 VHH blocks the efficacy of a full-sized therapeutic antibody. We were thus able to demonstrate that CTLA-4-binding antibodies require an Fc domain for antitumor effect.

Polymer Cancerostatics Targeted with an Antibody Fragment Bound via a Coiled Coil Motif: In Vivo Therapeutic Efficacy against Murine BCL1 Leukemia.

A BCL1 leukemia-cell-targeted polymer-drug conjugate with a narrow molecular weight distribution consisting of an N-(2-hydroxypropyl)methacrylamide copolymer carrier and the anticancer drug pirarubicin is prepared by controlled radical copolymerization followed by metal-free click chemistry. A targeting recombinant single chain antibody fragment (scFv) derived from a B1 monoclonal antibody is attached noncovalently to the polymer carrier via a coiled coil interaction between two complementary peptides. Two pairs of coiled coil forming peptides (abbreviated KEK/EKE and KSK/ESE) are used as linkers between the polymer-pirarubicin conjugate and the targeting protein. The targeted polymer conjugate with the coiled coil linker KSK/ESE exhibits 4× better cell binding activity and 2× higher cytotoxicity in vitro compared with the other conjugate. Treatment of mice with established BCL1 leukemia using the scFv-targeted polymer conjugate leads to a markedly prolonged survival time of the experimental animals compared with the treatment using the free drug and the nontargeted polymer-pirarubicin conjugate.

Targeted cancer therapy through antibody fragments-decorated nanomedicines.

Active targeting in cancer nanomedicine, for improved delivery of agents and diagnose, has been reviewed as a successful way for facilitating active uptake of theranostic agents by the tumor cells. The application of a targeting moiety in the targeted carrier complexes can play an important role in differentiating between tumor and healthy tissues. The pharmaceutical carriers, as main part of complexes, can be polymeric nanoparticles, micelles, liposomes, nanogels and carbon nanotubes. The antibodies are among the natural ligands with highest affinity and specificity to target pharmaceutical nanoparticle conjugates. However, the limitations, such as size and long circulating half-lives, hinder reproducible manufacture in clinical studies. Therefore, novel approaches have moved towards minimizing and engineering conventional antibodies as fragments like scFv, Fab, nanobody, bispecific antibody, bifunctional antibody, diabody and minibody preserving their functional potential. Different formats of antibody fragments have been reviewed in this literature update, in terms of structure and function, as smart ligands in cancer diagnosis and therapy of tumor cells.

Passive Immunization of Farmed Fish.

The fast growth and potential of global aquaculture has necessitated the adoption of sustainable and welfare-oriented therapeutics and prophylactic strategies. Knowledge gathered from studies about maternal passive immunity in fish and fish-to-fish passive immunization experiments supports the concept of using therapeutic Abs (of piscine and other vertebrate origin) in aquaculture. Traditional Ab formats (IgG, IgM) are expensive and laborious to produce; however, the introduction of new rAb fragments and single-domain Abs have reinvigorated the concept of passive immunization. This review will focus primarily on farmed salmonids (salmon and trout) within a comparative context and will give an overview of the basic principles and scientific premises for the passive immunization strategy, including existing and emerging Ab therapeutics.

Treatment with hyperimmune equine immunoglobulin or immunoglobulin fragments completely protects rodents from Ebola virus infection.

Recent successes with monoclonal antibody cocktails ZMapp(TM) and MIL77 against Ebola virus (EBOV) infections have reignited interest in antibody-based therapeutics. Since the production process for monoclonal antibodies can be prolonged and costly, alternative treatments should be investigated. We produced purified equine antisera from horses hyperimmunized with EBOV virus-like particles, and tested the post-exposure efficacy of the antisera in a mouse model of infection. BALB/c mice were given up to 2 mg of purified equine antisera per animal, at 30 minutes, 1 or 2 days post-infection (dpi), in which all animals survived. To decrease the possibility of serum sickness, the equine antisera was digested with pepsin to generate F(ab')2 fragments, with in vitro neutralizing activity comparable to whole immunoglobulin. Full protection was achieved with when treatment was initiated at 1 dpi, but the suboptimal protection observed with the 30 minute and 2 dpi groups demonstrate that in addition to virus neutralization, other Fc-dependent antibody mechanisms may also contribute to survival. Guinea pigs given 20 mg of antisera or F(ab')2 at or starting at 1 or 2 dpi were also fully protected from EBOV infection. These results justify future efficacy studies for purified equine products in NHPs.

Photo-thermal therapy of bladder cancer with Anti-EGFR antibody conjugated gold nanoparticles.

The aim of this study was to enhance the effectiveness of photo thermal therapy (PTT) in the targeting of superficial bladder cancers using a green light laser in conjunction with gold nanoparticles (GNPs) conjugated to antibody fragments (anti-EGFR). GNPs conjugated with anti-EGFR-antibody fragments were used as probes in the targeting of tumor cells and then exposed to a green laser (532nm), resulting in the production of sufficient thermal energy to kill urothelial carcinomas both in vitro and in vivo. Nanoparticles conjugated with antibody fragments are capable of damaging cancer cells even at relatively very low energy levels, while non-conjugated nanoparticles would require an energy level of 3 times under the same conditions. The lower energy required by the nanoparticles allows this method to destroy cancerous cells while preserving normal cells when applied in vivo. Nanoparticles conjugated with antibody fragments (anti-EGFR) require less than half the energy of non-conjugated nanoparticles to kill cancer cells. In an orthotopic bladder cancer model, the group treated using PTT presented significant differences in tumor development.

Emerging site-specific bioconjugation strategies for radioimmunotracer development.

INTRODUCTION: Radioimmunotracers are a promising class of companion diagnostics for precision medicine. They are composed of an antibody-based targeting agent and a radiolabeled imaging probe. Together with the tendency towards the use of small antibody-derived fragments, the employed conjugation method is gaining increasing attention. Conventional bioconjugation methods result in heterogeneous tracer populations of which the single elements can differ in immunoreactivity, pharmacokinetic behavior and stability. Site-specific conjugation strategies try to overcome these shortcomings and facilitate radioimmunotracer delivery, characterization and manufacturing. AREAS COVERED: An overview is provided of site-specific conjugation strategies for use in radioimmunotracer development. Currently applied strategies are discussed, together with other emerging site-specific conjugation methods that are applicable to diabodies, single-chain variable fragments (scFvs) and camelid single-domain antibody-fragments (sdAbs or nanobodies). EXPERT OPINION: The ultimate goal of site-specific bioconjugation strategies is to allow precise control over the conjugation site, to result in homogenous tracer populations, and to be versatile in use with different imaging probes. Chemoenzymatic methods appear to be promising in this respect.

Camelid heavy chain only antibody fragment domain against ß-site of amyloid precursor protein cleaving enzyme 1 inhibits ß-secretase activity in vitro and in vivo.

UNLABELLED: Accumulation and aggregation of the amyloid-ß (Aß) peptide is associated with Alzheimer's disease (AD). Aß is generated from the amyloid precursor protein by the successive action of two membrane-associated processing enzymes: ß-secretase or ß-site of amyloid precursor protein cleaving enzyme 1 (BACE1) and γ-secretase. Inhibition of one or both of these enzymes prevents Aß generation and the accompanying Aß accumulation. Antigen binding fragments from camelid heavy chain only antibodies (VHHs) were found to exert excellent enzyme inhibition activity. In the present study, we generated VHHs against BACE1 by active immunization of Lama glama with the recombinant BACE1 protein. Two classes of VHHs were selected from a VHH-phage display library by competitive elution with a peptide encoding the Swedish mutation variant of the BACE1 processing site. One VHH was found to inhibit the enzyme activity of BACE1 in vitro and in cell culture, whereas two other VHHs were found to stimulate BACE1 activity under the same conditions in vitro. Furthermore, an in vivo study with a transgenic AD mouse model, using intracisternal injection of the inhibitory VHH, led to acute reduction of the Aß load in the blood and brain. This inhibitory VHH may be considered as a candidate molecule for a therapy directed towards reduction of Aß load and prevention of AD progression. Both the inhibitory and stimulatory VHH may be useful for improving our understanding of the structure-function relationship of BACE1, as well as its role in AD progression. DATABASE: The GenBank sequence accession numbers are KR363186 for VHH B1a; KR363187 for VHH B3a; and KR363188 for VHH B5a.

Investigation of Fragment Antibody Stability and Its Release Mechanism from Poly(Lactide-co-Glycolide)-Triacetin Depots for Sustained-Release Applications.

Achieving long-term drug release from polymer-based delivery systems continues to be a challenge particularly for the delivery of large hydrophilic molecules such as therapeutic antibodies and proteins. Here, we report on the utility of an in situ-forming and injectable polymer-solvent system for the long-term release of a model antibody fragment (Fab1). The delivery system was prepared by dispersing a spray-dried powder of Fab1 within poly(lactide-co-glycolide) (PLGA)-triacetin solution. The formulation viscosity was within the range 1.0 ± 0.3 Pa s but it was injectable through a 27G needle. The release profile of Fab1, measured in phosphate-buffered saline (PBS), showed a lag phase followed by sustained-release phase for close to 80 days. Antibody degradation during its residence within the depot was comparable to its degradation upon long-term incubation in PBS. On the basis of temporal changes in surface morphology, stiffness, and depot mass, a mechanism to account for the drug release profile has been proposed. The unprecedented release profile and retention of greater than 80% of antigen-binding capacity even after several weeks demonstrates that PLGA-triacetin solution could be a promising system for the long-term delivery of biologics.

Resimmune, an anti-CD3ε recombinant immunotoxin, induces durable remissions in patients with cutaneous T-cell lymphoma.

Resimmune is a second-generation recombinant immunotoxin composed of the catalytic and translocation domains of diphtheria toxin fused to two single chain antibody fragments reactive with the extracellular domain of CD3ε. We gave intravenous infusions of Resimmune 2.5 - 11.25 µg/kg over 15 minutes to 30 patients (25 with cutaneous T-cell lymphoma, 3 with peripheral T-cell lymphoma, 1 with T-cell large granular lymphocytic leukemia and 1 with T-cell prolymphocytic leukemia) in an inter-patient dose escalation trial. The most common adverse events were fever, chills, hypotension, edema, hypoalbuminemia, hypophosphatemia, and transaminasemia. Among the 25 patients with cutaneous T-cell lymphoma, there were nine responses for a response rate of 36% (95% CI, 18%-57%) including four complete remissions (16%, 95% CI, 5%-36%). The durations of the complete remissions were 72+, 72+, 60+ and 38+ months. There were five partial remissions lasting 3, 3, 3+, 6+ and 14 months. Of 17 patients with a modified skin weighted assessment tool score <50, 17 patients with stage IB/IIB, and 11 patients with both a score <50 and stage IB/IIB, nine (53%), eight (47%), and eight (73%) had responses, respectively. Further studies of Resimmune in patients with low tumor burden, stage IB-IIB cutaneous T-cell lymphoma are warranted. This trial is registered at clinicaltrials.gov as #NCT00611208.

First-in-human trial of the safety, pharmacokinetics and immunogenicity of a PEGylated anti-CD40L antibody fragment (CDP7657) in healthy individuals and patients with systemic lupus erythematosus.

OBJECTIVE: The objective of this paper is to investigate the safety, pharmacokinetics (PK) and immunogenicity of CDP7657, a PEGylated anti-CD40L antibody fragment, in healthy individuals and patients with systemic lupus erythematosus (SLE). METHODS: This randomized, double-blind, single-dose, dose-escalation phase I study consisted of two parts. In part 1, 28 healthy individuals received CDP7657 IV (0.004-5 mg/kg) or placebo. In part 2, 17 patients with SLE received CDP7657 IV (5-60 mg/kg) or placebo. The CDP7657:placebo ratio was 3:1. RESULTS: Adverse events (AEs) were reported by 76% of healthy individuals and 100% of patients with SLE treated with CDP7657; most were mild or moderate in intensity. Two healthy individuals reported serious AEs (SAEs), one of which was considered treatment related (infusion-related reaction; 5 mg/kg cohort). One patient with SLE (60 mg/kg cohort) experienced three SAEs, one of which was considered treatment related (herpes zoster infection). No thromboembolic events were reported. CPD7657 exposure increased in a dose-proportional manner. Low anti-CDP7657 antibody titres were detected in the majority of CDP7657-treated participants with no apparent impact on the PK of CDP7657. CONCLUSION: Single doses of CDP7657 showed predictable PK in healthy individuals and patients with SLE and were well tolerated, with no safety signals of concern. These findings support further investigation of CDP7657 as a therapy for SLE.

Human monoclonal antibody fragments targeting matrilin-3 in growth plate cartilage.

PURPOSE: Many genetic disorders, including chondrodysplasias, and acquired disorders impair growth plate function, resulting in short and sometimes malformed bones. There are multiple endocrine and paracrine factors that promote chondrogenesis at the growth plate, which could potentially be used to treat these disorders. Targeting these growth factors specifically to the growth plate might augment the therapeutic skeletal effect while diminishing undesirable effects on non-target tissues. METHODS: Using yeast display technology, we selected single-chain variable antibody fragments that bound to human and mouse matrilin-3, an extracellular matrix protein specifically expressed in cartilage tissue. The ability of the selected antibody fragments to bind matrilin-3 and to bind cartilage tissue in vitro and in vivo was assessed by ELISA and immunohistochemistry. RESULTS: We identified antibody fragments that bound matrilin-3 with high affinity and also bound with high tissue specificity to cartilage homogenates and to cartilage structures in mouse embryo sections. When injected intravenously in mice, the antibody fragments specifically homed to cartilage. CONCLUSIONS: Yeast display successfully selected antibody fragments that are able to target cartilage tissue in vivo. Coupling these antibodies to chondrogenic endocrine and paracrine signaling molecules has the potential to open up new pharmacological approaches to treat childhood skeletal growth disorders.