Preclinical Safety Profile of a Depleting Antibody against CRTh2 for Asthma: Well Tolerated Despite Unexpected CRTh2 Expression on Vascular Pericytes in the Central Nervous System and Gastric Mucosa

Abstract

CRTh2 is expressed on immune cells that drive asthma pathophysiology. Current treatment options for severe asthma are inadequate and therapeutic antibody-mediated depletion of CRTh2-expressing cells represents a promising new therapeutic strategy. Here we report for the first time that CRTh2 is not only expressed on immune cells, but also on microvasculature in the central nervous system (CNS) and gastric mucosa in humans. Microvascular expression of CRTh2 raises a safety concern because a therapeutic antiCRTh2 antibody with enhanced depletion capacity could lead to vascular damage. To evaluate this safety risk, we characterized microvascular expression in human and in transgenic mice expressing human CRTh2 protein (hCRTh2.BAC.Tg) and found that CRTh2 is not localized to microvascular endothelium that is directly exposed to circulating therapeutic antibody, but rather, to pericytes that in the CNS are shielded from direct circulatory exposure by the blood-brain barrier. Immunohistochemical visualization of an intravenously administered antiCRTh2 antibody in transgenic mice revealed localization to microvascular pericytes in the gastric mucosa but not in the CNS, suggesting the blood-brain barrier effectively limits pericyte exposure to circulating therapeutic antibody in the CNS. Repeated dosing with a depleting antiCRTh2 antibody in hCRTh2.BAC.Tg mice revealed linear pharmacokinetics and no drug-related adverse findings in any tissues, including the CNS and gastric mucosa, despite complete depletion of CRTh2 expressing circulating eosinophils and basophils. Collectively, these studies demonstrate that the likelihood of drug-related CNS or gastrointestinal toxicity in humans treated with a therapeutic depleting antiCRTh2 antibody is low despite pericyte expression of CRTh2 in these tissues.

Automated Affinity Capture and On-Tip Digestion to Accurately Quantitate in Vivo Deamidation of Therapeutic Antibodies

“…In this report, we developed an automated high-throughput platform to perform on-tip affinity capture using MSIA D.A.R.T.’S tips (Thermo Fisher Scientific), with rapid digestion and peptide elution into a 96-well plate followed by LC–MS analysis. We applied the automated platform to study the levels of in vivo deamidated anti-CRTh2, a therapeutic antibody candidate targeting CRTh2 ( C hemoattractant R eceptor-homologous molecule expressed on Th2 cells). , CRTh2 is expressed on human immune cells such as eosinophils, basophils, type-2 innate lymphoid cells, and Th2 cells that mediate allergic inflammation and asthma pathogenesis. − Anti-CRTh2 has been shown in mouse models of Th2 inflammation to deplete CRTh2-expressing immune cells that are important in allergy and asthma, , thus has the potential to be a novel therapeutic for the management of allergic inflammatory diseases such as asthma.…”

G Protein–Coupled Receptors in Asthma Therapy: Pharmacology and Drug Action

“…One of them is a DP2-targeting antibody, which could deplete inflammatory cells, including eosinophils, ILC2, and T H 2 cells, and therefore may present a novel therapy for asthma (Huang et al, 2016b). Preclinical studies indicated a low risk of central nervous system or gastrointestinal toxicity associated with this antibody (Rajapaksa et al, 2016).…”