Antiinflammatory Pharmacology of Ensifentrine

Abstract

PURPOSE: Ensifentrine, a trequinsin analog, is an investigational dual inhibitor of PDE3 and 4 designed to enhance bronchodilator and anti-inflammatory effects locally in the airways and minimize possible adverse systemic effects through direct delivery via inhalation. Ensifentrine has shown bronchodilatory effects in clinical studies in patients with COPD and asthma (Singh 2020, Bjermer 2019). Anti-inflammatory effects including a significant reduction in inflammatory cells (e.g. neutrophils) have been shown with once-daily inhaled ensifentrine (e1mg) in sputum of healthy individuals following LPS challenge (Franciosi 2013). In enzyme activity assays of recombinant PDE3A, PDE3B, and PDE4B2, ensifentrine inhibited PDE activity with IC50s of 0.25, 0.29 and 290nM, respectively. The activity of ensifentrine against PDE4 aligns with positive control rolipram (130nM) and with reported functional activity of ensifentrine on PDE4-mediated inhibition of TNFa production in LPS-stimulated human monocytes (520nM) and proliferation of human mononuclear cells stimulated with PHA (460nM, Boswell-Smith 2006). While ensifentrine appears to be 1000-fold selective for PDE3 over PDE4 in vitro, it is well-recognized that several factors including dimerization, phosphorylation state and cross talk between PDEs limit the interpretation of PDE selectivity in assays using isolated or recombinant PDEs (Francis 2011). As an inhaled therapy delivered topically to the lung ate1mg/mL (e2000uM), pharmacologically relevant local concentrations of ensifentrine for PDE4 inhibition will be readily achieved. METHODS: Studies were conducted to demonstrate that at doses that produced bronchoprotective effects in vivo, ensifentrine could also inhibit antigen-induced inflammatory cell infiltration into the lungs and nose of allergic animals. Study 1 examined the anti-inflammatory effects of ensifentrine in ovalbumin (OVA) sensitized guinea pigs. Animals and were administered 0.3mg/mL inhaled ensifentrine immediately post antigen challenge. Study 2 examined the effect of ensifentrine on cell infiltration in nasal lavage in OVA sensitized guinea pigs. Ensifentrine was dosed 0.1, 0.3 & 0.5mg/mL via snout inhalation prior to antigen challenge. RESULTS: Study 1: ensifentrine produced significant inhibition of the recruitment of total cell numbers to BAL fluid 6 hours post antigen challenge. Study 2: guinea pigs dosed with ensifentrine showed a dose dependent inhibition of OVA-induced eosinophil and total cell recruitment into the nose (>80% inhibition at 1mg/mL dose). CONCLUSIONS: This in-vivo work supplements previously reported work and confirms that, at clinically relevant inhaled doses in animal models of lung inflammation, ensifentrine demonstrates robust anti-inflammatory activity. CLINICAL IMPLICATIONS: This further supports the dual mechanism of action of ensifentrine showing in vivo pharmacology consistent with both inhibition of PDE3 and 4.

Ensifentrine as a Novel, Inhaled Treatment for Patients with COPD

“…In enzyme activity assays of recombinant PDE3A, PDE3B, and PDE4B2, ensifentrine inhibited PDE activity with IC 50 values of 0.25, 0.29 and 290 nM, respectively. 26 The inhibitory activity of ensifentrine on PDE4 was aligned with the positive control rolipram (130 nM) 26 and with reported functional activity of ensifentrine on PDE4-mediated inhibition of TNFα production in lipopolysaccharide (LPS)-stimulated human monocytes (520 nM) and proliferation of human mononuclear cells stimulated with phytohemagglutinin (460 nM). 15 Ensifentrine has been shown to increase levels of cAMP in human neutrophils and CFBE41o-WT cells, demonstrating the ability of ensifentrine to inhibit PDE3 and PDE4 and produce expected effects in a cellular mechanistic assay.…”

Perspectives on Ensifentrine and Its Therapeutic Potential in the Treatment of COPD: Evidence to Date

“…6 Multiple in-vitro and in-vivo studies have demonstrated dose-dependent bronchodilator, bronchoprotective and anti-inflammatory effects of ensifentrine. 15,16 Multiple clinical studies have demonstrated similar effects of this drug when tested on healthy individuals and patients with asthma or COPD. In 4-week, phase II clinical trials, nebulized drug given twice a day alone or in combination with other bronchodilators, showed sustained bronchodilation and improvement in FEV1, improvement in symptoms judged by questionnaires, improvement in residual volumes and COPD symptoms.…”

“… 6 Multiple in-vitro and in-vivo studies have demonstrated dose-dependent bronchodilator, bronchoprotective and anti-inflammatory effects of ensifentrine. 15 , 16 …”

“…It also showed a dose-dependent inhibition of ovalbumin-induced eosinophil and total cell recruitment into the nasal passages, thereby reducing inflammation. 16 …”

Reply to Gan et al., to Calzetta et al., and to Poor

“…Ensifentrine is a novel PDE3/4 (phosphodiesterase 3 and 4) dual inhibitor, described as such because it potently inhibits each isolated recombinant enzyme ( 2 ) and shows functional activity in cellular and tissue assays consistent with PDE3 inhibitory effects (approximately 50% inhibition of contraction of tracheal preparations at a concentration of 100 nM) and PDE4 inhibitory effects (inhibition of TNFa production in human monocytes [half maximal inhibitory concentration (IC 50 ) = 520 nM] and proliferation of human mononuclear cells [IC 50 = 460 nM]) ( 2 , 3 ). Furthermore, in vivo pharmacology models with inhaled ensifentrine in guinea pigs also demonstrated that ensifentrine produced dose-dependent effects in animal models of lung inflammation at the same doses that produced bronchoprotective and smooth muscle effects ( 4 ). In healthy volunteers after lipopolysaccharide challenge, significant reductions in sputum inflammatory cells (e.g., neutrophils, macrophages, lymphocytes, and eosinophils) were observed after 6 days dosing with once-daily inhaled ensifentrine ( 5 ).…”