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ACS Medicinal Chemistry Letters
ACS Med Chem Lett. 2016 April 14; 7(4): 385–390.
Published online 2016 February 16. doi:  10.1021/acsmedchemlett.6b00004
PMCID: PMC4834660

Development of Novel 1,2,3,4-Tetrahydroquinoline Scaffolds as Potent NF-κB Inhibitors and Cytotoxic Agents


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1,2,3,4-Tetrahydroquinolines have been identified as the most potent inhibitors of LPS-induced NF-κB transcriptional activity. To discover new molecules of this class with excellent activities, we designed and synthesized a series of novel derivatives of 1,2,3,4-tetrahydroquinolines (4ag, 5ah, 6ah, and 7ah) and bioevaluated their in vitro activity against human cancer cell lines (NCI-H23, ACHN, MDA-MB-231, PC-3, NUGC-3, and HCT 15). Among all synthesized scaffolds, 6g exhibited the most potent inhibition (53 times that of a reference compound) of LPS-induced NF-κB transcriptional activity and the most potent cytotoxicity against all evaluated human cancer cell lines.

Keywords: 1,2,3,4-Tetrahydroquinolines; NF-κB inactivation; in vitro cytotoxicity; human cancer cell lines

NF-κB is a lymphoid-specific protein that binds to the enhancer of kappa light chain in the nucleus of B cells; NF-κB was discovered by Sen and Baltimore.1 NF-κB is involved in the regulation of many immune and inflammatory responses, cellular growth, and apoptosis.2,3 At present, NF-κB and its signaling is one of the most exciting and extensively studied research fields since NF-κB dysregulation is associated with many diseases such as cancer, AIDS, asthma, arthritis, diabetes, and inflammatory bowel disease.46 Several natural and synthetic compounds, including some drugs, have been tested for their potential to inhibit NF-κB, but very few of them are suitable for anticancer therapy.7,8 Therefore, it has been suggested that the development of novel NF-κB inhibitors with antitumor and anti-inflammatory activities is most important. Our group has been involved in the development of novel potent NF-κB inhibitors911 with anticancer activity.

Besides, quinolines and tetrahydroquinolines are important ubiquitous structural motifs in biologically active natural products and pharmacologically relevant therapeutic agents.1216 However, Khan et al.17 demonstrated that tetrahydroquinolines can be used as NF-κB inhibitors as well as anti-HIV agents, anti-Parkinson’s diseases, etc. María José Abad et al.18 also tested quinoline-based compounds as modulators of HIV transcription through NF-κB and Sp1 inhibition. These distinct inhibitory activities have encouraged us to prepare such core motifs to test our hypothesis that their derivatives would act as most potent NF-κB inhibitors and have anticancer activity. We designed and synthesized different novel derivatives of 1,2,3,4-tetrahydroquinoline-2-carboxylic acid N-(substituted)phenyl amide and tested them as potential NF-κB inhibitors and also evaluated their cytotoxicities against six human cancer cell lines (NCI-H23, ACHN, MDA-MB-231, PC-3, NUGC-3, and HCT-15). Based on our previous reports,911 we wish to maintain amide functionality to the core motifs, and the effect of the synthesized molecules on NF-κB transcriptional activity was measured by using a reported procedure.19In vitro cytotoxicity assay was performed using the number of cells measured indirectly by the sulforhodamine B method according to the National Cancer Institute (USA) protocol.20

We commenced our synthesis from commercially available quinoline-2-carboxylic acid, which underwent amidation reaction with various substituted aromatic amines in the presence of the coupling reagent 1,1′-carbonyldiimidazole (CDI) in tetrahydrofuran at room temperature. This reaction afforded the 3ag series of N-(substituted)quinoline-2-carboxamides in good yields (Scheme 1). The 3ag derivatives were also converted to 4ag by Pd/C hydrogenation reaction (H2 balloon) in ethanol solvent at room temperature. To obtain the 5ah, 6ah, and 7ah series, we performed acylation reaction in the presence of triethylamine in anhydrous tetrahydrofuran (Scheme 2). All newly synthesized derivatives (3ag, 4ag, 5ah, 6ah, and 7ah) were confirmed by 1H and 13C NMR and mass spectra. In evaluation studies of inhibition of LPS-induced NF-κB transcriptional activity, we compared all synthesized derivatives (4ag, 5ah, 6ah, and 7ah) with the reference compound pyrrolidine dithiocarbamate (PDTC), which acts as an antioxidant and is a potent inhibitor of NF-κB activation,2125 and also with the lead compound KL-1156, which is an inhibitor of NF-κB translocation to the nucleus in LPS-stimulated RAW 264.7 macrophages.26

Scheme 1
Synthesis of 3ag and 4ag Series of Scaffolds
Scheme 2
Synthesis of 5ah, 6ah, and 7ah Series of Scaffolds

Initially, we screened the 4ag derivatives; they exhibited marginal inhibitory effects on NF-κB transcriptional activity (Table 1). After finding that the tetrahydroquinoline core motif was less potent, we next turned our attention mainly to the substitutions of R, R1, R2, R3, and R4, which resulted in 24 derivatives (5ah, 6ah, and 7ah; Table 1). Among them, 5e (IC50: 1.4 ± 0.71 μM), 6f (IC50: 0.90 ± 0.071 μM), 6g (IC50: 0.70 ± 0.071 μM), and 6h (IC50: 2.7 ± 0.42 μM) exhibited outstanding inhibitory effects (Figure Figure11) on LPS-induced NF-κB transcriptional activity in comparison with remaining derivatives (Table 1). After having the initial experimental results, structure–activity relationship development was initiated for the tetrahydroquinoline scaffold. The 1,2,3,4-tetrahydroquinoline-2-carboxamide motif was modified at two key positions interpreted as R, and substitutions were performed at the aromatic system (R1, R2, R3, and R4). In the first set of compounds, we performed different substitutions at R1, R2, R3, and R4, which resulted in 4ag analogues. Subsequently R1 = R2 = R3 = R4 = H (4a, IC50 60 μM) was prepared to study the effect of the core 1,2,3,4-tetrahydroquinoline-2-carboxamide motif. We also examined the electronic influence of different groups at R1, R2, R3, and R4, including electron-deficient (−CF3) and electron-rich groups (−OCH3 and −OH). Next, we introduced various acyl and aroyl groups at the 1-position of tetrahydroquinoline-2-carboxamides, which resulted in 5ah, 6ah, and 7ah analogues. Sizes of different aliphatic chains (−R) and substituents at the aromatic ring were then examined. The inhibitory effects of these compounds (4ag, 5ah, 6ah, and 7ah) depended on the nature of the substitution at the R-position and the substituents (R1, R2, R3, and R4) on the N-phenyl ring of tetrahydroquinoline-2-carboxamides. For instance, with R = an alkyl group (methyl, ethyl, propyl, or octyl), we did not notice any significant inhibitory effects, but with R = phenyl. The following compounds exhibited excellent inhibitory effects on LPS-induced NF-κB transcriptional activity (Figure Figure22): 5e (IC50: 1.4 ± 0.071 μM, about 26 and 37 times more potent than PDTC and KL-1156, respectively), 6f (IC50: 0.90 ± 0.071 μM, about 41 and 58 times more potent), 6g (IC50: 0.70 ± 0.071 μM, about 53 and 75 times), and 6h (IC50: 2.7 ± 0.42 μM, about 13 and 19 times). We also tested other substitutions at the R-position and different substituents at the R1, R2, R3, and R4 positions of the 1,2,3,4-tetrahydroquinoline core motif (Table 1). To explain the trend toward NF-κB inhibition interestingly, the electron-withdrawing group −CF3 at the R1 position resulted in outstanding inhibitory effects on LPS-induced NF-κB transcriptional activity in comparison with any electron-releasing group (for example, −OH). We also noticed that −I and +M effect groups (such as −Cl) at the second and third positions of the aryl group of R in combination with −CF3 at the R1 position also showed excellent inhibition of NF-κB transcriptional activity (6g, IC50: 0.70 ± 0.071 μM; 6f, IC50: 0.90 ± 0.071 μM). Overall, most of the synthesized compounds (4d, 4e, 4g, 5f, 5h, 6d, 7ac, and 7eh) were more potent inhibitors of NF-κB transcriptional activity than PDTC and KL-1156 (Figure Figure22). We next studied the in vitro cytotoxicity of the synthesized compounds and initially evaluated quinoline-2-carboxamide derivatives (3ag) with the human lung cancer cell line (NCI-H23) and doxorubicin (ADR) as a reference compound.

Figure 1
Efficacy of 4ag, 5ah, 6ah, and 7ah analogues in inhibition of NF-κB transcriptional activities.
Figure 2
In vitro efficacy of 4ag, 5ah, 6ah, and 7ah analogues in inhibiting growth of human cancer cell lines.
Table 1
Inhibitory Effect on LPS-Induced NF-κB Transcriptional Activity for 1,2,3,4-Tetrahydroquinolines
Table 2
Cytotoxicity of 3ag Series against Lung Cancer (NCI-H23) Cell Lines

As shown in Table 2, electron-withdrawing substituents such as −CF3 at the R2 and R4 positions of the phenyl ring (3g, GI50 1.44 ± 0.463 μM) resulted in the most potent cytotoxicity. The series 4ag, 5ah, 6ah, and 7ah were also evaluated for in vitro cytotoxicity against six human cancer cells: NCI-H23, ACHN (renal), MDA-MB-231 (breast), PC-3 (prostate), NUGC-3 (gastric), and HCT15 (colon) (Table 3). Any substitution on the phenyl ring was not beneficial, and only 4b (GI50 2.23 ± 0.455 μM) exhibited better cytotoxic activities against all tested cell lines than other analogues of the 4ag series (Table 3). To further confirm that the tetrahydroquinoline motif is beneficial for cytotoxicity, we executed acylation reaction with triethyl amine in tetrahydrofuran with 4b, 4e, and 4g; introduction of electron-rich or electron-withdrawing substituents at the R1 position afforded 5ah, 6ah, and 7ah analogues. As expected, these analogues had improved cytotoxicity against all tested cell lines (Figure Figure22), suggesting that substitutions at the R1 position and the first position of the tetrahydroquinoline motif are most important (Table 3). Compound 5e exhibited the highest cytotoxicity (Table 3) against all evaluated cell lines (NCI-H23, GI50 3.49 ± 0.999 μM; NUGC-3, GI50 3.78 ± 0.618 μM; HCT 15, GI50 3.83 ± 0.994 μM). The importance of an electron-withdrawing group was also confirmed by 6g and 6h derivatives, which have −CF3 and −Cl on both phenyl rings and exhibited outstanding cytotoxicity (6g, GI50 0.292 ± 0.111–0.797 ± 0.173 μM; 6h, GI50 0.307 ± 0.0.0941–0.839 ± 0.0610 μM) against all tested cell lines (Figure Figure22, Table 3). The −CF3 group at the R2 and R4 positions of the phenyl ring in 7g also resulted in potent cytotoxicity against all tested cell lines (0.420–1.19 μM; Table 3). Compound 7h also exhibited potent cytotoxicity against lung (NCI-H23 (GI50 0.889 ± 0.102 μM) and gastric (NUGC-3, GI50 1.66 ± 0.406 μM) cancer cell lines and moderate cytotoxicity against the other four cell lines (Table 3).

Table 3
Cytotoxicity against NCI-H23, ACHN, MDA-MB-231, PC-3 NUGC-3, and HCT-15 Cancer Cell Lines

In summary, we identified a new class of 1,2,3,4-tetrahydroquinolines as the most potent NF-κB inhibitors and potent cytotoxic agents against human cancer cell lines.

The first round of screening starting from the initial lead scaffold 4a led to the discovery of 6f, 6g, and 6h analogues. These new lead analogues inhibited LPS-induced NF-κB transcriptional activity 41, 53, and 13 times more potently, respectively, than the reference compound PDTC. Analogues 6f, 6g, and 6h have also shown the most potent in vitro cytotoxicity against all evaluated human cancer cell lines. Thus, 6f, 6g, 6h, and related analogues provide new chemical tools for development of pathway-selective NF-κB inhibitors with anticancer activity. Work on the enhancement of potency and pharmacological profiles of these probe molecules are underway.


This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF-2013R1A1A2009381), and Medical Research Center Program (2008-0062275).



nuclear factor kappa-light-chain-enhancer of activated B cells

Supporting Information Available

Supporting Information Available

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsmedchemlett.6b00004.

  • Synthetic procedures, characterization of final products, biological assay protocols, and data and pharmacology profiles (PDF)


The authors declare no competing financial interest.

Supplementary Material


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