3.1. Furoxan Hybrids as Antiatherosclerotic Agents
Nian-Guang Li et al. [
37,
38] synthesized a series of furoxan–cinnamic acid hybrids as potential antiatherosclerotic drug candidates. Ferulic acid hybrids
17 and
18 (
Figure 7), as well as monoesters
9a–
c (
Figure 5), did not present better antioxidant activity than ferulic acid (IC
50 = 16.2 μM for DPPH, IC
50 = 17.6 μM for lipid peroxidation). Hybrid
18c, the most potent antioxidant, showed the same activity as
9c, with IC
50 values of 66.9 μM (DPPH) and 72.1 μM (lipid peroxidation).
Caffeic hybrids
19 and
20 (
Figure 7) presented a higher antioxidant activity (IC
50 = 13–39 μM in DPPH assay, IC
50 = 25–56 μM in lipid peroxidation assay) compared to ferulic hybrids
17 and
18. The phenolic hydroxy group seems to be significant for the antioxidant activity of the molecules. The most potent antioxidant agent was found to be hybrid
19a, with an IC
50 value of 13 μM in DPPH and 25 μM in lipid peroxidation. This hybrid combines a phelylfuroxan moiety and caffeic acid with no linker. In general, phenylfuroxan hybrids
19 had better antioxidant activity than phenylsulfonylfuroxan hybrids
20 [
38].
The vasodilating activity of the hybrids was measured through an in vitro vascular relaxation assay (organ bath) using PGF2a-precontracted porcine pulmonary arteries. The most active compounds belonged to the phenylsulfonylfuroxan series
18. In particular, hybrid
18c, with a higher antioxidant activity among the furoxan hybrids
17 and
18, was more potent than ISDN (EC
50 = 0.1123 μM), with an EC
50 value of 0.0928 μM, followed by hybrids
18b and
18d (which were as potent as ISDN), with EC
50 values of 0.1143 μM and 0.1039 μM, respectively [
37]. Phenylfuroxans of caffeic acid
19 demonstrated lower EC
50 values compared to the corresponding phenylsulfonylfuroxans
20 (EC
50 = 1.03–1.83 μM). The best vasodilating activity was shown by hybrid
19a (EC
50 = 0.12 μM), the molecule with the higher antioxidant activity [
38].
Hybrids 17 exhibited higher NO release potency (3.71–6.58 μM at 2 h) than mononitrates 9, but not as good as dinitro hybrids 12, with the exception of 17a, with a value of 10.70 μM at 2 h. Correlation observed between vasodilating activities and NO-releasing potency of the new ferulic molecules; e.g., phenylsulfonyl hybrids 18 were the most potent NO donors, with values of 22.06–27.53 μM at 2 h. In addition, caffeic hybrids 19, with the best vasodilating activities in comparison to hybrids 20, presented NO release in a range of 9.05–11.08 μM, and were as potent as ISDN. Phenylsulfonylfuroxans of caffeic acid values released were found from 2.29 to 3.82 μΜ.
3.2. Furoxan Hybrids as Anticancer Agents
Li et al. [
42] tested furoxan hybrids
17,
18, and
19a in 14 cancer cell lines. The most promising anticancer agents seemed to be compounds
18b–
d, presenting decreased IC
50 values (0.40–2.88 μM) against all the human cancer cells (
Table 3 and
Table 4). The results indicated that the linkers played an important role in the appearance of anticancer acti-vity. Caffeic hybrid
19a showed better values than ferulic hybrids
17, with the exception of the corresponding hybrid
17a.
In another research study, Ming-Dong Lu et al. [
47] synthesized NO-releasing derivatives of cinnamic acids (
Figure 8) as antitumor agents. The hybrid compounds consisted of one furoxan moiety and a hydroxylcinnamic acid derivative, coupled with various alkyl amines as linkers. The new molecules were evaluated for their in vitro antitumor activity against hepatocellular carcinoma cells (SMMC-7721 and HepG2) and human breast cancer cells (MCF-7). As a result, none of the tested hybrids were a more potent inhibitor than Adriamycin in all three cancer cell lines; however, hybrid
21 demonstrated the best inhibitory activity, with IC
50 values of 6.1 μM (SMMC-7721), 7.3 μM (HepG2), and 3.8 μM (MCF-7). This hybrid proceeded to further studies and presented a selective cytotoxic effect. Specifically, it did not present important inhibition potency in nontumor cells (liver LO2 cells), but a high inhibition effect of HepG2 cell proliferation (2–16 μM), in a concentration-dependent manner. Hybrid
22, the ferulic analogue of hybrid
21, also displayed a significant antitumor activity, with IC
50 values of 7.9 μM (SMMC-7721), 8.2 μM (HepG2), and 6.5 μM (MCF-7). Moreover, hybrids
23 and
24 were found to be more potent than the corresponding NO donor moiety
25, with IC
50 values of 5.0–10.8 μM, as shown in
Table 5, suggesting the contribution of the furoxan moiety to the antitumor activity of hydroxycinnamic acids.
3.3. Furoxan Hybrids as Antidiabetic Agents
Metabolic syndrome is a cluster of conditions occurring together, including increased blood pressure, high blood sugar, abnormal cholesterol and triglyceride levels, raised fasting glucose, and central obesity. This disorder can lead to the development of cardiovascular disease (CVD), renal and liver diseases, and type 2 diabetes mellitus (T2DM). The systemic inflammatory process, associated with the metabolic syndrome, presents numerous deleterious effects that promote plaque activation and the appearance of atherosclerosis. It has been reported that antioxidants and radical scavengers prevent the formation of advanced glycation end-products (AGEs) formed after long sustained hyperglycemia [
48,
49].
Xie et al. combined cinnamic acids with NO donor groups as a therapeutic approach against metabolic syndrome [
50] and diabetes mellitus [
51]. In particular, phenylsulfonyl- and phenylfuroxan alcohols were linked with some natural cinnamic acid derivatives (
Figure 7 and
Figure 9). In vitro evaluation of phenylsulfonyl hybrids
18a,
20a,
26, and
27 against yeast a-glucosidase from Saccharomyces cerevisiae showed that hybrid
20a (IC
50 = 187.34 μM) was more potent than acarbose (IC
50 = 232.41 μM). Hybrids
18a and
26 were comparable to acarbose, with IC
50 values of 263.47 μM and 278.54 μM, respectively, while hybrid
27 presented an increased IC
50 value [
50].
In vitro evaluation of phenylfuroxan hybrids
17a,
19a,
28, and
29 against a-glucosidase (maltase and sucrase) from rat small intestine showed that only hybrids
17a (IC
50 = 301.73 μM in maltase and 223.36 μM in sucrase) and
19a (IC
50 = 123.36 μM in maltase and 155.03 μM in sucrase) exhibited any activity. None of the hybrids presented better inhibitory activity than acarbose (5.59 μM in maltase and 2.31 μM in sucrase), and hybrids
28 and
29 did not show any activity [
51].
The antiglycosylation activity of the compounds was determined in a bovine serum albumin (BSA)-methylglyoxal (MGO) system, which simulated the glycation reactions of the body. The inhibition of AGEs formation was increased by the hybrids
18a and
20a (IC
50 values of 0.985 and 0.158 mM, respectively), which presented higher activity than aminoguanidine (IC
50 = 1.510 mM). Phenylfuroxans
17a and
19a were not as potent as the control aminoguanidine, with IC
50 values of 3.259 mM and 3.037 mM, respectively. Moreover, hybrids
26,
27,
28, and
29 did not show any activity [
50,
51].
The in vitro antioxidant activity of the hybrids was tested using vitamin C as standard. DPPH and OH radical assays were used. Hybrid
20a was found to be more potent than vitamin C, with IC
50 of 0.042 mM (DPPH), 0.219 mM (OH radical), and 0.221 μM (anti-lipid peroxidation). Hybrid
18a displayed antioxidant activity comparable to vitamin C, with IC
50 of 0.181 mM (DPPH) and 0.669 mM (OH radical) and showed to be more potent than the control in antilipid peroxidation, with IC
50 of 5.132 μM. However, hybrids
26 and
27 did not present good or any activity, suggesting that the absence of phenolic hydroxyl groups decreased the activity [
50]. In the DPPH and OH radical assays, hybrid
19a, containing two free phenolic hydroxyl groups, was determined to be the more potent antioxidant compared to ascorbic acid, with an IC
50 of 0.0082 mM (DPPH) and 0.222 mM (OH radical). Hybrid
17a showed an IC
50 of 0.177 mM (DPPH) and 0.895 mM (OH radical), comparable to ascorbic acid activity. Removal of the methoxy group led to a reduced activity, since hybrid
28 showed an IC
50 value of 0.987 mM (DPPH) and no activity in the OH radical assay. Hybrid
29 did not present any activity in either assay [
51]. The results supported that hybrids with antioxidant activity have good antiglycation properties. Moreover, combination of these properties is more efficient in inhibiting the glycation reactions.
The antiplatelet aggregation activity of the hybrids was determined using adenosine diphosphate (ADP)-induced platelet aggregation in human platelet-rich plasma (PRP) through Born’s turbidimetric method. All phenylsulfonylfuroxan compounds were more potent than aspirin (5.96% inhibition at 1.5 μΜ), used as control. Hybrid
20a showed the highest inhibition of 73.54% at 1.5 μM, followed by hybrids
18a,
26, and
27, with an inhibition of 65.53%, 58.14%, and 56.44% at 1.5 μM, respectively [
50]. Phenylfuroxans
17a,
19a,
28, and
29 did not present higher activity than aspirin (28.11% at 0.15 mM), with 9.46%, 17.66%, 10.78%, and 12.66% inhibition at 0.15 mM, respectively [
51].
The NO-releasing activity of the new compounds was correlated to the antiplatelet aggregation activity. The potency of the hybrids to release NO was estimated using a Griess reagent assay. Compound
20a released a considerable amount of NO (45.01 μM), followed by hybrids
27,
18a, and
26 (40.01, 39.20, and 35.72 μM respectively). The results showed that phenolic hydroxyl may enhance the NO-release activity of the compounds [
50], whereas the presence of the sulfonyl group may enhance the antiplatelet aggregation activity of the compounds. Phenylfuroxan hybrids
17a,
19a,
28, and
29 released a lower amount of NO (14.20–14.80 μM) [
51].
The vasorelaxant effect of the hybrids was determined by the measurement of the concentration-dependent relaxations of the mesenteric arteries that were preconstructed with 60 mM of potassium chloride (KCl). Cumulative concentration of the NO donors
18a,
20a,
26, and
27 that were applied to the arteries was 10
−10–10
−3.5 M and 10
−8–10
−1 M for hybrids
17a,
19a,
28, and
29. Hybrid
26 showed the best activity, with a pIC
50 value of 6.171, close to that of SNP (pIC
50 = 6.786). Hybrids
27,
18a, and
20a were not as potent as the control SNP, with pIC
50 values of 5.875, 5.872, and 5.698, respectively [
50]. Phenylfuroxans displayed decreased activity in comparison to SNP (pIC
50 = 6.786). Hybrids
19a and
29 were the most potent, with pIC
50 values of 4.286 and 4.547, respectively, followed by hybrids
17a (pIC
50 = 3.778) and
28 (pIC
50 = 3.538). The vasodilative activities of the target compounds were similar to their antiplatelet aggregation abilities, and also may be positively correlated with their NO-releasing abilities [
51].