PDE4 inhibitor intermediate 1

Atopic dermatitis

Atopic dermatitis (AD) is the most common chronic inflammatory skin disease, with a lifetime prevalence of up to 20% and substantial effects on quality of life. AD is characterized by intense itch, recurrent eczematous lesions and a fluctuating course. AD has a strong heritability component and is closely related to and commonly co-occurs with other atopic diseases (such as asthma and allergic rhinitis). Several pathophysiological mechanisms contribute to AD aetiology and clinical manifestations. Impairment of epidermal barrier function, for example, owing to deficiency in the structural protein filaggrin, can promote inflammation and T cell infiltration. The immune response in AD is skewed towards T helper 2 cell-mediated pathways and can in turn favour epidermal barrier disruption. Other contributing factors to AD onset include dysbiosis of the skin microbiota (in particular overgrowth of Staphylococcus aureus), systemic immune responses (including immunoglobulin E (IgE)-mediated sensitization) and neuroinflammation, which is involved in itch. Current treatments for AD include topical moisturizers and anti-inflammatory agents (such as corticosteroids, calcineurin inhibitors and cAMP-specific 3',5'-cyclic phosphodiesterase 4 (PDE4) inhibitors), phototherapy and systemic immunosuppressants. Translational research has fostered the development of targeted small molecules and biologic therapies, especially for moderate-to-severe disease.

The Cyclic Nitronate Route to Pharmaceutical Molecules: Synthesis of GSK's Potent PDE4 Inhibitor as a Case Study

An efficient asymmetric synthesis of GlaxoSmithKline's potent PDE4 inhibitor was accomplished in eight steps from a catechol-derived nitroalkene. The key intermediate (3-acyloxymethyl-substituted 1,2-oxazine) was prepared in a straightforward manner by tandem acylation/(3,3)-sigmatropic rearrangement of the corresponding 1,2-oxazine-N-oxide. The latter was assembled by a (4 + 2)-cycloaddition between the suitably substituted nitroalkene and vinyl ether. Facile acetal epimerization at the C-6 position in 1,2-oxazine ring was observed in the course of reduction with NaBH₃CN in AcOH. Density functional theory (DFT) calculations suggest that the epimerization may proceed through an unusual tricyclic oxazolo(1,2)oxazinium cation formed via double anchimeric assistance from a distant acyloxy group and the nitrogen atom of the 1,2-oxazine ring.

Apremilast, a novel phosphodiesterase 4 (PDE4) inhibitor, regulates inflammation through multiple cAMP downstream effectors

Introduction: This work was undertaken to delineate intracellular signaling pathways for the PDE4 inhibitor apremilast and to examine interactions between apremilast, methotrexate and adenosine A2A receptors (A2AR).
Methods: After apremilast and LPS incubation, intracellular cAMP, TNF-α, IL-10, IL-6 and IL-1α were measured in the Raw264.7 monocytic murine cell line. PKA, Epac1/2 (signaling intermediates for cAMP) and A2AR knockdowns were performed by shRNA transfection and interactions with A2AR and A2BR, as well as with methotrexate were tested in vitro and in the murine air pouch model. Statistical differences were determined using one or two-way ANOVA or Student's t test. The alpha nominal level was set at 0.05 in all cases. A P value of < 0.05 was considered significant.
Results: In vitro, apremilast increased intracellular cAMP and inhibited TNF-α release (IC50=104nM) and the specific A2AR-agonist CGS21680 (1μM) increased apremilast potency (IC50=25nM). In this cell line, apremilast increased IL-10 production. PKA, Epac1 and Epac2 knockdowns prevented TNF-α inhibition and IL-10 stimulation by apremilast. In the murine air pouch model, both apremilast and MTX significantly inhibited leukocyte infiltration, while apremilast, but not MTX, significantly inhibited TNF-α release. The addition of MTX (1 mg/kg) to apremilast (5 mg/kg) yielded no more inhibition of leukocyte infiltration or TNF-α release than with apremilast alone.
Conclusions: The immunoregulatory effects of apremilast appear to be mediated by cAMP through the downstream effectors PKA, Epac1, and Epac2. A2AR agonism potentiated TNF-α inhibition by apremilast, consistent with the cAMP-elevating effects of that receptor. Because the A2AR is also involved in the anti-inflammatory effects of MTX, the mechanism of action of both drugs involves cAMP-dependent pathways and is therefore partially overlapping in nature.

Efficient synthesis of an apremilast precursor and chiral β-hydroxy sulfones via ketoreductase-catalyzed asymmetric reduction

Ketoreductase (KRED)-catalyzed asymmetric reduction of prochiral ketones is an attractive method to synthesize chiral alcohols. Herein, two KREDs LfSDR1-V186A/E141I and CgKR1-F92I with complementary stereopreference were identified towards reduction of apremilast prochiral ketone intermediate 1a. LfSDR1-V186A/E141I exhibited >99% conversion and 99.2% ee yielding an apremilast chiral alcohol intermediate ((R)-2a) at 50 g L^-1 substrate loading. Furthermore, we investigated the substrate scope of β-keto sulfones by using LfSDR1-V186A/E141I and CgKR1-F92I to produce both enantiomers of the corresponding β-hydroxy sulfones, with good-to-excellent conversion (up to >99%) and enantioselectivity (up to 99.9% ee) being obtained in most cases. Finally, the gram-scale synthesis of (R)-2a was performed by employing the crude enzyme of LfSDR1-V186A/E141I and BsGDH to afford the desired enantiomer with >99% conversion, 85.9% isolated yield and 99.2% ee. This study presents a biocatalytic strategy to synthesize chiral β-hydroxy sulfones.

Directed evolution of an amine transaminase for the synthesis of an Apremilast intermediate via kinetic resolution

Apremilast is an important active pharmaceutical ingredient that relies on a resolution to produce the key chiral amine intermediate. To provide a new catalytic and enzymatic process for Apremilast, we performed the directed evolution of the amine transaminase fromVibriofluvialis. Six rounds of evolution resulted in the VF-8M-E variant with > 400-fold increase specific activity over the wildtype enzyme. A homology model of VF-8M-E was built and a molecular docking study was performed to explain the increase in activity. The purified VF-8M-E was successfully applied to produce the key chiral amine intermediate in enantiopure form and 49% conversion via a kinetic resolution, representing a new enzymatic access towards Apremilast.