Evaluation of efficacy and safety of the anti-VWF Nanobody ALX-0681 in a preclinical baboon model of acquired thrombotic thrombocytopenic purpura.

ALX-0681 is a therapeutic Nanobody targeting the A1-domain of VWF. It inhibits the interaction between ultra-large VWF and platelet GpIb-IX-V, which plays a crucial role in the pathogenesis of thrombotic thrombocytopenic purpura (TTP). In the present study, we report the efficacy and safety profile of ALX-0681 in a baboon model of acquired TTP. In this model, acute episodes of TTP are induced by administration of an ADAMTS13-inhibiting mAb. ALX-0681 completely prevented the rapid onset of severe thrombocytopenia and schistocytic hemolytic anemia. After induction of TTP, platelet counts also rapidly recovered on administration of ALX-0681. This effect was corroborated by the full neutralization of VWF activity. The schistocytic hemolytic anemia was also halted and partially reversed by ALX-0681 treatment. Brain CT scans and post mortem analysis did not reveal any sign of bleeding, suggesting that complete neutralization of VWF by ALX-0681 under conditions of thrombocytopenia was not linked with an excessive bleeding risk. The results obtained in this study demonstrate that ALX-0681 can successfully treat and prevent the most important hallmarks of acquired TTP without evidence of a severe bleeding risk. Therefore, ALX-0681 offers an attractive new therapeutic option for acquired TTP in the clinical setting.

Antithrombotic drug candidate ALX-0081 shows superior preclinical efficacy and safety compared with currently marketed antiplatelet drugs.

Neutralizing the interaction of the platelet receptor gpIb with VWF is an attractive strategy to treat and prevent thrombotic complications. ALX-0081 is a bivalent Nanobody which specifically targets the gpIb-binding site of VWF and interacts avidly with VWF. Nanobodies are therapeutic proteins derived from naturally occurring heavy-chain-only Abs and combine a small molecular size with a high inherent stability. ALX-0081 exerts potent activity in vitro and in vivo. Perfusion experiments with blood from patients with acute coronary syndrome on standard antithrombotics demonstrated complete inhibition of platelet adhesion after addition of ALX-0081, while in the absence of ALX-0081 residual adhesion was observed. In a baboon efficacy and safety model measuring acute thrombosis and surgical bleeding, ALX-0081 showed a superior therapeutic window compared with marketed antithrombotics. Pharmacokinetic and biodistribution experiments demonstrated target-mediated clearance of ALX-0081, which leads to a self-regulating disposition behavior. In conclusion, these preclinical data demonstrate that ALX-0081 combines a high efficacy with an improved safety profile compared with currently marketed antithrombotics. ALX-0081 has entered clinical development.

A Phase 1 First-in-Human Study of FS118, a Tetravalent Bispecific Antibody Targeting LAG-3 and PD-L1 in Patients with Advanced Cancer and PD-L1 Resistance.

PURPOSE: This phase 1 study (NCT03440437) evaluated the safety, tolerability, pharmacokinetics (PK), and activity of FS118, a bispecific antibody-targeting LAG-3 and PD-L1, in patients with advanced cancer resistant to anti-PD-(L)1 therapy. PATIENTS AND METHODS: Patients with solid tumors, refractory to anti-PD-(L)1-based therapy, received intravenous FS118 weekly with an accelerated dose titration design (800 µg to 0.3 mg/kg) followed by 3+3 ascending dose expansion (1 to 20 mg/kg). Primary objectives were safety, tolerability, and PK. Additional endpoints included antitumor activity, immunogenicity, and pharmacodynamics. RESULTS: Forty-three patients with a median of three prior regimens in the locally advanced/metastatic setting, including at least one anti-PD-(L)1 regimen, received FS118 monotherapy. FS118 was well tolerated, with no serious adverse events relating to FS118 reported. No dose-limiting toxicities (DLT) were observed, and an MTD was not reached. The recommended phase 2 dose of FS118 was established as 10 mg/kg weekly. The terminal half-life was 3.9 days. Immunogenicity was transient. Pharmacodynamic activity was prolonged throughout dosing as demonstrated by sustained elevation of soluble LAG-3 and increased peripheral effector cells. The overall disease control rate (DCR) was 46.5%; this disease control was observed as stable disease, except for one late partial response. Disease control of 54.8% was observed in patients receiving 1 mg/kg or greater who had acquired resistance to PD-(L)1-targeted therapy. CONCLUSIONS: FS118 was well tolerated with no DLTs observed up to and including 20 mg/kg QW. Further studies are warranted to determine clinical benefit in patients who have become refractory to anti-PD-(L)1 therapy. See related commentary by Karapetyan and Luke, p. 835.

The TITAN trial--assessing the efficacy and safety of an anti-von Willebrand factor Nanobody in patients with acquired thrombotic thrombocytopenic purpura.

The Phase II TITAN trial is designed to assess the efficacy and safety of an anti-von Willebrand factor (vWF) Nanobody in patients with acquired thrombotic thrombocytopenic purpura (TTP). Nanobodies are a novel class of therapeutic proteins and are based on the smallest functional fragments of single-chain antibodies that occur naturally in the Camelidae family (Nanobody® and Nanobodies® are registered trademarks of Ablynx NV). With vWF implicated in the thrombotic process underlying TTP, an anti-vWF Nanobody may hold significant promise as adjunctive therapy to plasma exchange. Recruitment is currently ongoing, and aims to include a total of 110 patients from countries in Europe, the Middle East, Australia and Northern America.

Durable response of anaplastic thyroid carcinoma to FS118, a bispecific LAG-3/PD-L1 antibody, after checkpoint inhibitor progression: a case report.

Anaplastic thyroid cancer represents a rare, highly aggressive form of thyroid cancer with a poor prognosis and an overall survival ranging from 5 to 12 months. Unfortunately, treatment options remain limited, even for patients with a targetable driver mutation. Here, we present a case of a patient with a BRAF V600E-mutated, PD-L1 positive (tumor proportion score of 95%) anaplastic thyroid cancer refractory to standard therapies, including debulking surgery, followed by chemoradiation, who had further progressed on PD-1 monotherapy, and was unable to tolerate BRAF/MEK inhibition. Ongoing treatment with FS118, a bispecific LAG-3/PD-L1 antagonist, has afforded 3 years of disease control, including a late confirmed partial response, with excellent tolerability. Given this response, further investigation is required to delineate the mechanism by which dual PD-L1/LAG-3 blockade by FS118 overcomes initial PD-1 pathway resistance, and therefore, identify which patients are most likely to benefit. Simultaneously, expanded use should be considered for those with refractory disease, especially if PD-L1 positive. Insights Dual PD-L1/LAG-3 blockade may be an effective treatment strategy for refractory metastatic tumors, including anaplastic thyroid cancer.

Novel antiplatelet agents: ALX-0081, a Nanobody directed towards von Willebrand factor.

This manuscript reviews the studies performed with ALX-0081 (INN: caplacizumab), a Nanobody targeting von Willebrand factor, in the context of current antithrombotic therapy in coronary artery disease. ALX-0081 specifically inhibits platelet adhesion to the vessel wall, and may control platelet aggregation and subsequent clot formation without increasing bleeding risk. A substantial number of antithrombotics are aimed at this cascade; however, their generally indiscriminative mode of action can result in a narrow therapeutic window, defined by the risk for bleeding complications, and thrombotic events. Nonclinically, ALX-0081 compared favorably to several antithrombotics. In Phase I studies in healthy subjects and stable angina patients undergoing percutaneous coronary intervention (PCI), ALX-0081 was well tolerated, and effectively inhibited pharmacodynamic markers. Following these results, a phase II study was initiated in high-risk acute coronary syndrome patients undergoing PCI. Based on its mechanism of action, ALX-0081 is also being developed for acquired thrombotic thrombocytopenic purpura.

von Willebrand factor inhibition improves endothelial function in patients with stable angina.

ALX-0081 is a novel nano-antibody inhibiting von Willebrand factor (vWF). We evaluated whether direct inhibition of vWF by ALX-0081 improves endothelial function. Stable patients (pts, n = 55) with single vessel disease undergoing percutaneous coronary intervention (PCI) were randomized to ALX-0081 (n = 38) or placebo (n = 17). vWF inhibition was assessed by vWF antigen level (vWF:Ag) and activity by ristocetin test (vWF:RiCo). Endothelial function was assessed before (BL), 6 h and 24 h after PCI by: (a) endothelial peripheral arterial tonometry (Endoscore); (b) endothelial microparticles (EMPs) by flow cytometry. vWF:Ag and vWF:RiCo decreased within 1 h from ALX-0081. In the placebo group, no significant Endoscore changes occurred from BL to 24 h. In ALX-0081 group, Endoscore increased from BL to 24 h (p = 0.014). A decrease in EMPs was observed after ALX-0081 (p < 0.01), while no changes occurred in placebo pts. An inhibition of vWF with ALX-0081 significantly improves peripheral endothelial function.

Relation of endothelial function to residual platelet reactivity after clopidogrel in patients with stable angina pectoris undergoing percutaneous coronary intervention.

Platelet reactivity is greater in patients with stable angina and with more extensive peripheral vascular atherosclerosis. We sought to evaluate whether impaired peripheral microcirculatory endothelial function might correlate with platelet reactivity after clopidogrel and therefore predispose to an unfavorable outcome after percutaneous coronary intervention (PCI). In 52 consecutive patients with stable angina undergoing elective PCI, endothelial function was assessed by (1) endothelial peripheral arterial tonometry (measuring the "Endoscore"); (2) the von Willebrandt factor antigen level and ristocetin co-factor activity. Basal platelet reactivity was assessed by soluble P-selectin. Patients then received a 600-mg clopidogrel loading dose > or = 12 hours before PCI. A blood sample was withdrawn 12 hours later, but before PCI, to assess platelet reactivity using the P2Y12 reaction unit and percentage of P2Y12 inhibition with the point-of-care VerifyNow P2Y12 assay. Troponin T was assessed 24 hours after PCI. The Endoscore inversely correlated with von Willebrandt factor antigen activity (r = -0.52, p = 0.0001) and soluble P-selectin concentration (r = -0.36, p = 0.021), suggesting greater platelet reactivity with increased impaired endothelial function. After clopidogrel, the Endoscore correlated directly with the percentage of P2Y12 inhibition (r = 0.36, p = 0.009) and inversely with the P2Y12 reaction unit (r = -0.41, p = 0.002), suggesting greater residual platelet reactivity with more impaired endothelial function. The average Endoscore was significantly lower in patients with troponin T elevation (troponin positive group 0.267 + or - 0.091) than in patients without troponin T elevation (troponin negative group 0.508 + or - 0.041, p = 0.015 vs troponin positive). In conclusion, an impaired endothelial response before clopidogrel was associated with greater platelet reactivity after clopidogrel. This link might explain the unfavorable PCI outcomes in patients with more severe endothelial impairment.

The development of nanobodies for therapeutic applications.

Evolution has been continuously honing the design of antibodies to function as specific molecular markers that are able to alert the immune system to the presence of pathogenic antigens, and to recruit complement- and Fc receptor-bearing effector cells. During the past 25 years, the versatility of antibodies has been applied to several therapeutic applications. The development of new technologies, combined with data obtained using a new generation of antibody reagents, have allowed the adaptation of the design of antibodies to better match drug development requirements. Nanobodies are therapeutic proteins derived from the heavy-chain variable (VHH) domains that occur naturally in heavy-chain-only Ig molecules in camelidae. These VHH domains are the smallest known antigen-binding antibody fragments. Nanobodies can be easily produced in prokaryotic or eukaryotic host organisms, and their unique biophysical and pharmacological characteristics render these molecules ideal candidates for drug development. This review describes the structural properties of nanobodies and focuses on their unique features, which distinguishes these molecules from other antibody formats and small-molecule drugs. Possible therapeutic applications of nanobodies are discussed and data from phase I clinical trials of the novel 'first-in-class' anti-thrombotic agent ALX-0081 (Ablynx NV) are presented.