Bridged tetrahydroisoquinolines as selective NADPH oxidase 2 (Nox2) inhibitors

Abstract

(1SR,4RS)-3,3-Dimethyl-1,2,3,4-tetrahydro-1,4-(epiminomethano)naphthalenes were synthesized in 2-3 steps from commercially available materials and assessed for specificity and effectiveness across a range of Nox isoforms. The N-pentyl and N-methylenethiophene substituted analogs 11g and 11h emerged as selective Nox2 inhibitors with cellular IC₅₀ values of 20 and 32 μM, respectively.

Figure1

Representation of Nox2 structure and activation.

Figure 2

Nonspecific or indirect inhibitors of Nox2 activity.

Figure 3

Tetrahydroisoquinoline hit 7 (CID3323417) and related analogs 8 (CID3160422) and 9 (CID4005560) from a screen of a subset of compounds from the UPCMLD library.

Figure 4

Concentration-response testing of 7 in (A) COS-Nox2 cells, (B) lysed COS-Nox2 cells (cell-free preparation), and (C) COS-Nox4 cells. (A) Effect of 7 on Nox2 activity was measured in whole COS-Nox2 cells stimulated by 5 μM PMA using L-012 and Amplex® Red (AR). Data are expressed as % of vehicle control and represent the mean ± SEM of 6 independent experiments. (B) COS-Nox2 cell lysate was preincubated with various concentrations of 7 for 15 min at 25 °C. After the addition of 130 μM LiDS, O₂^•− production was initiated by the addition of 180 μM NADPH and measured by the initial linear rate of SOD-inhibitable cytochrome C reduction. O₂^•− production is expressed as nmol O₂^•− /min/10⁷ cell equivalents. (C) COS-Nox4 cells were pretreated with various concentrations of 7 for 15 min. Initial rate of H₂O₂ production was measured using Amplex® Red. Data are expressed as the mean ± SEM of 7 independent experiments.

Figure 5

Concentration response analysis of compounds 11g (A) and 11h (B) for inhibition of Nox1, 2, 3, 4, and 5. IC₅₀ values were calculated using the non-linear regression for three parameters analysis which assumes a Hill slope = 1. (See Table 1 caption and Supplementary Information for experimental details.)

Scheme 1

Synthesis of hit 7 and related analog 8, and reductive amination of these building blocks to form tertiary amines 11a-h.

Scheme 2

Synthesis of biaryl analogs 12a-d via Suzuki cross-coupling with aryl bromides.

Scheme 3

Reduction of alkene 7 to alkane 13 and sulfonamide formation to form analogs 14a and b.