Asthma is one of the most prevalent global health problems, with 300 million humans affected worldwide, including 26.5 million US residents. Asthma is characterized by chronic airway inflammation leading to airway remodeling, hyperactivity to external stimuli, and airway obstruction. First-line controller medications include inhaled corticosteroids that are supplemented in more severe disease with long-acting β2 adrenergic receptor agonists, oral corticosteroids, or other immune modulating agents. Fast acting bronchodilators can offer immediate symptom relief but have diminished effect when used chronically. One of the few alternatives for asthma patients who respond poorly to inhaled steroids is a leukotriene receptor antagonist. However, variations in leukotriene signaling genes may preclude efficacy in many patients. More recent alternatives are injectable monoclonal antibodies that target Th2-high asthmatic eosinophilia. However, high drug costs limit their use to only severe disease. Taken together, an increasing patient population, limited safe and effective treatment options, and high drug cost for biologics represent an unmet medical need for many asthma patients.
Volatile and intravenous anesthetic binding to GABAARs have long been known to dose dependently promote bronchodilation and attenuate bronchoconstriction, respectively. However, it has been a longstanding belief that any GABAergic contribution to airway tone was largely mediated by CNS mediated mechanisms. Importantly, our collaborative research team showed that selective ligands can bind directly to GABAARs on airway smooth muscle (ASM) (i.e., no CNS involvement) to increase the efficacy of the receptor’s Cl– ion channel, which in turn modulates intracellular Ca++ leading to muscle relaxation.(Gallos et al. 2015) We also showed that GABAAR are expressed in immune cells and can similarly be modulated, effecting intracellular Ca++ and reducing cell activation (e.g. cytokine expression in activated CD4+ T cells).(Forkuo et al. 2016) We further showed that an overlapping subset of GABAAR subunit isotypes are expressed on ASM and immune cells.(Mizuta et al. 2008, Yocum et al. 2017)
Bronchodilation ex vivo and in vivo was demonstrated for α4 subtype selective GABA(A) receptor ligands was described in several publications (Gallos et al. 2012, Yocum et al. 2016, Forkuo et al. 2016, Forkuo et al. 2017 and Jahan et al. 2017). In parallel, we introduced α5 subtype selective GABA(A) receptor ligands as potential asthma agents (Gallos et al. 2015, Forkuo et al. 2017, Forkuo et al. 2018). As depicted in Figure 1, we used a battery of pre-clinical criteria that included microsomal stability, pharmacokinetics, ability to cross the blood-brain barrier, ex vivo relaxation of constricted lung tissue, in vivo reduction of airway hyperresponsiveness, and the ability to reduce lung inflammation as criteria to advance compounds through our drug development pipeline.
Figure 1. Development of subtype selective GABA(A) receptor ligand as asthma treatment.eu
The breakthrough was achieved with compound MIDD0301 that did not cross the blood brain barrier and interacted with common GABAARs on ASM and immune cells with optimal potency in suppressing lung inflammation and relaxing ASM using several in vitro assays and in vivo animal asthma models when given orally and nebulized. In the presence of MIDD0301, ex vivo human lung strips showed smooth muscle relaxation comparable to mouse lung tissue supporting the translation of the animal studies to humans (Yocum et al. 2019 and Roni et al. 2022) A key innovation of PI301 is a new chemical entity that modulates a molecular target common to ASM and leukocytes, whereby a single small molecule can control ASM constriction and inflammation two clinical hallmarks of asthma.
PI301 has a favorable safety profile and employs a diazepine pharmacophore that, in contrast to all clinically used diazepines, acts peripherally without the induction of sensorimotor inhibition and did not induce respiratory depression in naïve mice (Zahn et al. 2020). We reported that orally administered PI301 causes no adverse systemic (gastrointestinal, hematological, and CNS) effects when given at 200 mg/kg daily for 28 days or a single 1,000 mg/kg bolus dose. In contrast to commonly used steroids, it does not cause systemic immune suppression as shown by no effect on T-dependent antibody responses or alteration in immune organ histology during 28-day dosing. In contrast to β-adrenergic receptor agonists, MIDD0301 can be taken orally without cardiovascular adverse effects. Bronchodilators, such as clenbuterol and bitolterol, were withdrawn from the market because of these adverse effects. We have shown in vitro that the affinity of MIDD0301 for the hERG channel is very low (2.7 % at 10 µM) and no other pharmacological targets were affected among 48 diverse receptors tested (NIMH PDSP). We also demonstrated that PI301 given orally to mice at 100 mg/kg did not alter systolic, diastolic, or mean blood pressure. We have developed a large-scale synthesis of MIDD0301 to support future clinical studies by optimizing purification methods and ensure retention of optical purity through the manufacturing process (Knutson et al. 2020).
Scheme 1. Synthesis of MIDD0301
Using this route, we generated several hundred grams of pure material and characterized the impurity profile of every step to comply with GMP regulations. Detailed studies pH dependent studies enabled us to develop stable and effective salt forms of MIDD0301 to enable controlled dissolution (Roni et al. 2020). Additional analytical work showed minimal phase 1 metabolism and conjugation with glucuronic acid and glucose in liver and kidney, respectively. The in vitro results supported by monitoring the formation of these metabolites in blood and lung as well as quantification of metabolites and parent compound in urine and feces after oral and intravenous administration (Roni et al. 2021) Pharmacokinetic studies were performed in several species with bioavailabilities ranging between 10-28% with T1/2 of about 100 min.
Recently, we developed a novel class of spiro-imidazodiazepines that are non-chiral and retain the same characteristics as MIDD0301 (Webb et al. 2023).