Research Article: Pharmacokinetic parameters and mechanism of action of an efficient anti-Aβ single chain antibody fragment

Date Published: May 31, 2019

Publisher: Public Library of Science

Author(s): Gisela Esquerda-Canals, Joaquim Martí-Clúa, Sandra Villegas, Stephen D. Ginsberg.


The success of the targeting of amyloid-β (Aβ) oligomers through immunotherapy in Alzheimer’s disease (AD) mouse models has not been translated into the clinics. The use of single-chain variable fragments (scFvs) has been proposed to prevent the potential severe effects of full-length mAbs by precluding crystallizable fraction-mediated microglia activation. The efficacy of scFv-h3D6, a bapineuzumab-derived anti-Aβ scFv, has been extensively proven. In this work, we compared scFv-h3D6-EL, an elongated variant of the scFv-h3D6, with its original version to assess whether its characteristic higher thermodynamic stability improved its pharmacokinetic parameters. Although scFv-h3D6-EL had a longer half-life than its original version, its absorption from the peritoneal cavity into the systemic compartment was lower than that of the original version. Moreover, we attempted to determine the mechanism underlying the protective effect of scFv-h3D6. We found that scFv-h3D6 showed compartmental distribution and more interestingly crossed the blood–brain barrier. In the brain, scFv-h3D6 was engulfed by glial cells or internalized by Aβ peptide-containing neurons in the early phase post-injection, and was colocalized with the Aβ peptide almost exclusively in glial cells in the late phase post-injection. Aβ peptide levels in the brain decreased simultaneously with an increase in scFv-h3D6 levels. This observation in addition to the increased tumor necrosis factor-α levels in the late phase post-injection suggested that the engulfment of Aβ peptide/scFv-h3D6 complex extruded from large neurons by phagocytic cells was the mechanism underlying Aβ peptide withdrawal. The mechanism of action of scFv-h3D6 demonstrates the effectivity of Aβ-immunotherapy and lays the background for other studies focused on the finding of a treatment for AD.

Partial Text

The currently available therapies for Alzheimer’s disease (AD) include the use of cholinesterase inhibitors (donepezil, galantamine, and rivastigmine) that partially compensate the pathological reduction in acetylcholine and an NMDA receptor antagonist (memantine) that prevents the effect of the increased glutamate levels in the synaptic cleft [1]. Because these treatments palliate the symptoms of AD rather than targeting its underlying causes, the implementation of novel therapeutic strategies has become a necessity [2]. In this sense, several molecules have been designed for targeting amyloid-β (Aβ) peptide, the key component in AD [3,4]. Aβ peptide-directed immunotherapy is a promising approach because it focuses on capturing the Aβ peptide through active immunotherapy (by directing a patient’s immune response to different forms and/or fragments of the peptide) or through passive immunotherapy (by administrating antibodies or their derivatives that directly arrest the Aβ peptide) [5,6]. Passive immunotherapy is a safer option than active immunotherapy because it can be stopped immediately in case of any adverse reaction [7]. Several clinical trials are currently ongoing in this regard [8–10]. Bapineuzumab (Pfizer/Janssen) was the first mAb to reach phase III clinical trials; however, the occurrence of vasogenic edema and microhemorrhage resulted in the suspension of the studies in 2012 [11]. Similarly, solanezumab (Eli Lilly), albeit resulted safe, showed a benefit that was not higher than that associated with the palliative acetylcholinesterase inhibitors drugs, and the studies were terminated [12]. Unfortunately, this has also recently been the case for aducanumab (Biogen Idec) [13]. However, other mAbs, such as gantenerumab (Hoffman-La Roche) and crenezumab (Genentech), are currently under phase III clinical trials [10].

At present, only palliative treatment is available for AD, which interferes with the symptoms rather than the causes of AD and therefore cannot preclude the dramatic deterioration of patients with AD [2,8,9]. Determination of disease-modifying treatments involves enormous efforts and considerations for determining appropriate targets and mechanisms of action [41]. Molecular engineering and treatment designing play a key role in determining the suitability of a therapy. ScFv-h3D6 is already known to improve the first hallmarks of AD [27–29]. Moreover, molecular redesigning produced an elongated and thermodynamically more stable version of scFv-h3D6, i.e., scFv-h3D6-EL [25,26]. In the present study, we determined the pharmacokinetic parameters of scFv-h3D6 (scFv-h3D6-WT) and compared them with those of the thermodynamically improved version (scFv-h3D6-EL) to test whether the in vitro benefits of greater stability [25,26] were maintained in vivo. ScFv-h3D6-WT showed better absorption from the intraperitoneal cavity into the blood than scFv-h3D6-EL, whereas scFv-h3D6-EL exhibited a longer half-life than scFv-h3D6-WT. This indicated that the improved stability of scFv-h3D6-EL translated into its in vivo performance but its modified structure hindered its absorption, thus reducing its potential effects. Moreover, these results also highlighted the reciprocal requirement between pharmacokinetic parameters and molecular/treatment designing because, for example, a different administration route could be used to improve scFv-h3D6-EL absorption [42].