Development of the clinical candidate PBD-C06, a humanized pGlu3-Aβ-specific antibody against Alzheimer’s disease with reduced complement activation

Abstract

In clinical trials with early Alzheimer's patients, administration of anti-amyloid antibodies reduced amyloid deposits, suggesting that immunotherapies may be promising disease-modifying interventions against Alzheimer's disease (AD). Specific forms of amyloid beta (Aβ) peptides, for example posttranslationally modified Aβ peptides with a pyroglutamate at the N-terminus (pGlu3, pE3), are attractive antibody targets, due to pGlu3-Aβ's neo-epitope character and its propensity to form neurotoxic oligomeric aggregates. We have generated a novel anti-pGlu3-Aβ antibody, PBD-C06, which is based on a murine precursor antibody that binds with high specificity to pGlu3-Aβ monomers, oligomers and fibrils, including mixed aggregates of unmodified Aβ and pGlu3-Aβ peptides. PBD-C06 was generated by first grafting the murine antigen binding sequences onto suitable human variable light and heavy chains. Subsequently, the humanized antibody was de-immunized and site-specific mutations were introduced to restore original target binding, to eliminate complement activation and to improve protein stability. PBD-C06 binds with the same specificity and avidity as its murine precursor antibody and elimination of C1q binding did not compromise Fcγ-receptor binding or in vitro phagocytosis. Thus, PBD-C06 was specifically designed to target neurotoxic aggregates and to avoid complement-mediated inflammatory responses, in order to lower the risk for vasogenic edemas in the clinic. Alzheimer's disease (AD) is a neurodegenerative disease that accounts for 60-70% of dementia patients. Progressive memory loss, and cognitive dysfunction and premature death 3-9 years after diagnosis are typical for AD 1 and 140 million AD patients or related dementia patients are projected to require treatment and care in 2050 worldwide, generating a major socioeconomic burden 2-5. Currently, the disease is often treated symptomatically with drugs that transiently improve neuropsychiatric symptoms by antagonizing acetylcholinesterase (AChE) or N-Methyl-D-aspartic acid (NMDA) receptor alone or in combinations 6. In contrast, disease-modifying therapies (DMTs) are new therapeutics, which aim at preventing or slowing AD progression by targeting the disease-causing mechanisms of AD 7-9. Most DMTs against AD in late stage clinical development either target amyloid-or tau-related pathologies. These two histopathological hallmarks are identified either as extracellular plaques of aggregated amyloid β (Aβ) peptides or as intracellular aggregation of hyperphosphorylated tau proteins in neurofibrillary tangles 10,11. According to the amyloid hypothesis of AD 11 , proteolytic cleavage of the amyloid precursor protein (APP) results in the generation and accumulation of soluble Aβ monomers that further aggregate into soluble oligomers and fibrils of different sizes and functionalities 12. These soluble aggregates may aggregate further into larger

Mutual structural effects of unmodified and pyroglutamylated amyloid β peptides during aggregation

“…Aβ 1–40 is more abundant in human brain than Aβ 1–42 , but the latter aggregates faster and is more toxic, and the Aβ 1–42 /Aβ 1–40 ratio correlates with AD severity 5,12,20–26 . N‐terminally truncated and pyroglutamylated forms of Aβ (pEAβ), especially pEAβ 3–40 and pEAβ 3–42 , accumulate in AD and Down syndrome brains in significant quantities (up to 50% of total Aβ), 27–32 possess augmented cytotoxicity and form hypertoxic assemblies with unmodified Aβ 7,13,33–38 . Analysis of senile plaques identified both individual pEAβ and unmodified Aβ deposits, as well as heterogeneous, intermixed deposits 28 .…”

Combination of the Glutaminyl Cyclase Inhibitor PQ912 (Varoglutamstat) and the Murine Monoclonal Antibody PBD-C06 (m6) Shows Additive Effects on Brain Aβ Pathology in Transgenic Mice

“…Based on the results of previous studies [ 27 ], we selected a dose of 0.8 g/kg PQ912 in chow, which translates in this study to a daily dose of ≈140 mg/kg. The pGlu-specific antibody m6 (murine PBD-C06, IgG2a) was also investigated in detail in previous studies [ 34 , 35 ], however, these studies were not performed in the hAPPsl×hQC transgenic mice, which harbor an increased pGlu-Aβ formation. Hence, we first performed a dose-finding study with the m6 antibody to be able to select subtherapeutic doses of both treatments for the combination experiment.…”

“…The mouse IgG2a variant of the murine anti-pGlu3Aβ antibody PBD-C06 (PBD-C06.02; m6) has been produced as described previously [ 34 , 59 ].…”

“…Accordingly, inhibition of QC has been shown to attenuate AD-like symptoms in mice dose-dependently [ 27 ] and Varoglutamstat, a first in-class QC inhibitor, has been proven safe and showed signs of efficacy in clinical phase 1 and 2 studies [ 32 , 33 ]. In contrast to the suppression of pGlu-formation by inhibition of QC, the targeting with antibodies aims at clearance of pGlu-Aβ after formation and/or the blocking of aggregation [ 34 ]. Hence, both treatment paradigms address the same goal by apparently different and independent molecular strategies.…”

Ultrasound-mediated blood-brain barrier disruption improves anti-pyroglutamate3 Aβ antibody efficacy and enhances phagocyte infiltration into brain in aged Alzheimer’s disease-like mice

“…We obtained a murine anti-pGlu3 amyloid-β IgG2a mAb (07/2a) from Vivoryon Therapeutics AG (Halle, Germany) for this study. We previously characterized and determined in vivo and ex vivo the pathological efficacy and in vivo behavioral improvement using the IgG2a mAb, 07/2a, and its IgG1 version, 07/1, in APP/PS1dE9 Tg mice, a model of Alzheimer's disease amyloidosis [20][21][22][23]. Here, we investigated whether FUS-mediated BBB disruption combined with an anti-pGlu3 Aβ mAb, 07/2a, can improve both clearance of amyloid plaques and cognition in aged APP/PS1dE9 mice without increasing the incidence of microhemorrhage.…”