Schiff Bases: Interesting Scaffolds with Promising Antitumoral Properties

: Schiff bases, named after Hugo Schiff, are highly reactive organic compounds broadly used as pigments and dyes, catalysts, intermediates in organic synthesis, and polymer stabilizers. Lots of Schiff bases are described in the literature for various biological activities, including antimalarial, antibacterial, antifungal, anti-inﬂammatory, and antiviral. Schiff bases are also known for their ability to form complexes with several metals. Very often, complexes of Schiff bases with metals and Schiff bases alone have demonstrated interesting antitumor activity. Given the innumerable vastness of data regarding antitumor activity of all these compounds, we focused our attention on mono-and bis-Schiff bases alone as antitumor agents. We will highlight the most signiﬁcant examples of compounds belonging to this class reported in the literature.


Introduction
Since their discovery by the German chemist Hugo Schiff [1], Schiff bases (imines), scaffolds with high chemical reactivity, and their metal complexes have been very well known for catalysis in various synthetic processes and for their biological properties. In therapy, Schiff bases and their metal complexes have been reported to manifest a wide range of biological activities [2,3] such as antimicrobial [4], ureases inhibitory [5], anti-inflammatory [6,7], anti-ulcerogenic [8], antioxidant [9][10][11], pesticidal, cytotoxic, and anticancer [12] including DNA damage [13][14][15]. Schiff bases have been also successfully used in scientific studies [16] as highly efficient and selective sensing materials for optical, electrochemical [17,18], and membrane sensors [19]. Zinc-Schiff bases have been proposed as carrier vehicles for the delivery of zinc to prostate cells. Indeed, the use of the membrane-penetrating peptide Novicidin connected to zinc-Schiff base has been studied as a therapeutic approach for prostate cancer [20]. Schiff base ligands, as some other organic small molecules [21], have received great attention from researchers thanks to their easy preparation and ability to form complexes with almost all metals, due to the electron-donating nitrogen in their base structure [22][23][24]. Several metal complexes, in which the metal is coordinated to various ligands, are able not only to stabilize the metal but also to modify its chemical and pharmaceutical properties and are receiving attention in medicinal chemistry [25][26][27][28][29][30]. The general structure of a Schiff base is shown in Figure 1, R 1 , R 2 and R 3 being an alkyl or aryl moiety. Schiff bases are particularly interesting in the field of antitumor agents [31][32][33][34] as many other small organic molecules (for instance, diarylureas [35], indoles [36,37], carbazoles [38], phthalimides [39], and so on [40,41]). The most salient and recent data on Schiff bases will be, herein, reviewed.

Mono-Schiff Bases
For instance, in a recent study, the introduction of Schiff bases in the N-phenylcarbazole/ triphenylamine modified half-sandwiched iridium(III) compounds determined an enhancement of antitumor activity of about 13 times that of the clinical cisplatin [42]. This review focused on studies of the last decades on mono-and bis-Schiff bases as antiproliferative agents, paying attention particularly on Schiff bases showing high activity (concentration which kills or inhibits cell viability by 50% (IC 50 ) in the range of micromolar to nanomolar).

Mono-Schiff Bases
Vicini et al. (2003) [43] studied a series of Schiff bases and tested their antiproliferative activity against a panel of human cell lines derived from hematological and solid tumors. The most interesting compounds were 1-3 ( Table 1). All of them inhibited the growth of leukemia cell lines, with IC 50 values ranging between 1.5 and 7 µM against human CD4 + lymphocytes (MT-4), human CD4 + acute T-lymphoblastic leukemia (CCRF-CEM), human splenic B-lymphoblastoid cells (WIL-2NS), and human acute B-lymphoblastic leukemia (CCRF-SB). The 2-Amino-6-mercaptopurine was used as reference drug (IC 50 between 0.1 and 0.5 µM). Particularly, compound 3 was also active against solid tumor-derived cell lines' skin melanoma and breast adenocarcinoma cells (IC 50 = 6 and 10 nM) against human skin melanoma SK-MEL-28 and human breast adenocarcinoma MCF-7 cell lines, respectively. The values for 2-amino-6-mercaptopurine were 5 and 4 µM, respectively. Zhou et al. (2007) [44] studied several imines bearing thiazole and triazole moieties and evaluated their antiproliferative activities against leukemia, stomach, and larynx cancer cell lines. The 2,4-dinitro substituted Schiff base 4 displayed high activity against HL-60, BGC-823 and Hep-2 cell lines, showing percentage inhibition of 91.97, 98.49, and 91.16%, respectively. Abdel- Hafez et al. (2009) [45] studied several Schiff bases as derivatives of xanthotoxin and evaluated their antitumor activities against cervical carcinoma (HeLa) and breast carcinoma (MCF 7) cell lines. The Schiff base 5 was inactive against MCF-7 cell line but was the most interesting against HeLa, showing an IC 50 value of 7.2 µM and a percent viability of 70% (xanthotoxin, 7.6 µM and 62%, respectively). Kraicheva et al. (2009) [46] studied three Schiff bases and evaluated their antiproliferative activity, using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltratrazolium bromide (MTT) assay, against four human leukemic cell lines, viz., LAMA-84 (peripheral chronic myeloid leukemia cells), K-562 (non-adherent chronic myelogenous leukemia cells of the erythroleukemia type), HL-60 (acute promyelocytic leukemia cells) and its multi-drug-resistant sub-line HL-60/Dox (multi-drug resistant acute myeloblastic leukemia cell line), characterized by the overexpression of MRP-1 protein (ABC-C1). Compound 6 showed antiproliferative activity (IC 50 = 39.9 µM, 29.9 µM, and 68.6 µM against LAMA-84, K-562, and HL-60/Dox, respectively), while compound 7 was less active (IC 50 = 251.9 µM, 212.9 µM, and 226.1 µM against LAMA-84, K-562, and HL-60/Dox, respectively). Both the investigated compounds were identified as capable of evoking the distinctly marked lower cytotoxic effects (with the IC 50 values over 400 µM) against the sensitive leukemic cell line HL-60 in a preliminary antitumor screening. Nawaz et al. (2009) [47] studied Schiff bases with ferrocene addition and evaluated their antitumor, antioxidant, and DNA-protecting activities. Antitumor activity was eval-uated by Potato disc tumor induction assay using Agrobacterium tumefaciens (At-10) to induce tumors on potato discs, that is, a prescreen assay and its results were in accordance with other commonly used in vitro antitumor assays. All the tested compounds inhibited tumor production for treatment of 1000, 100, and 10 µg/mL concentration at p < 0.05 (vincristine, used as positive control, showed 100% tumor inhibition at all concentrations tested). The inhibition was observed in a dose-dependent manner with the highest inhibition at 1000 µg/mL concentration. Moreover, the highest tumor growth inhibition of 71% was observed with ferrocene containing Schiff base 8, followed by 9 with 58% inhibition at 1000 µg/mL. IC 50 values were 20 and 563 µg/mL versus 0.003 µg/mL of vincristine. Zaheer et al. (2010) [48] studied several Schiff bases and tested their cytotoxic activity by the brine shrimp lethality assay. Medium lethal concentration (LD 50 [52] prepared series of 14 imines and studied the suppression of proliferation of different human cancer cell lines (HeLa (cervical carcinoma), SW620 (colorectal adenocarcinoma, metastatic), MiaPaCa-2 (pancreatic carcinoma), MCF-7 (breast epithelial adenocarcinoma, metastatic)) and their cytotoxic activity on normal human fibroblasts (WI38 normal diploid human fibroblasts) using the MTT assay. Compounds 17 and 18 exerted a strong non-specific antiproliferative effect on all cell lines tested and a concentration-dependent effect on HeLa and MCF-7 cell lines at micromolar concentrations (IC 50 = 4.73 and 3.24 µM on HeLa and 9.23 and 15.27 µM on MCF-7). However, they were also highly cytotoxic on human fibroblasts. Shaker et al. (2011) [53] synthesized surfactants containing Schiff bases with hydrocarbon chains of different lengths (from C12 to C18). In vitro anticancer cytotoxic activity of these compounds was investigated using EAC as a model system of mice cell tumor at different concentrations (25,50, and 100%) against liver carcinoma (HepG2), breast carcinoma (MCF-7), and colon carcinoma (HCT-116) cell lines. Compound 19, bearing a C14 hydrocarbon chain, caused the death of 95% of EAC cell at the highest concentration. The IC 50 values for compound 19 at different concentrations ranged from 1 to 10 mg/mL. It showed high activity in in vitro system on the tumor cell lines investigated and the highest cytotoxic effect on HepG2, HCT-116, and MCF-7, respectively, and SBC12 surfactant-affected tumor tissue at very low concentrations at values lower than their critical micelle concentration (cmc) values. Kraicheva et al. (2012) [54] synthesized two anthracene-containing Schiff bases, 9anthrylidene-p-toluidine (20) and 9-anthrylidene-furfurylamine (21), and tested their anticancer activities in vitro on a panel of human epithelial cancer cell lines (cell lines from ductal carcinoma of the breast with low and high metastatic potential, MCF-7 and MDA-MB-231, respectively; colostrum-derived myoepithelial cells, expressing polyoma virus large T-antigen, HBL-100 line; bladder carcinoma, 647-V; hepatocellular carcinoma, HepG2; colon carcinoma, HT-29; cervical carcinoma, HeLa). Compounds 20 and 21 showed high cytotoxic activity toward colon carcinoma HT-29 cell line (IC 50 = 0.08 and 0.20 mg/mL versus 0.58 mg/mL of doxorubicin). The authors also performed their safety testing, both in vitro (Neutral Red Uptake Assay, 3T3 NRU test) and in vivo on ICR mice for genotoxicity and antiproliferative activity. Both compounds were shown not to induce clearly expressed dose-effect clastogenic activities, in contrast to the alkylating agent Mitomycin C. Bae et al. (2012) [55] synthesized new Schiff bases and evaluated their anti-melanogenesis activity, in murine B16F10 melanoma cells, through the inhibition of tyrosinase. Compound 12 exhibited the most potent and non-competitive inhibition on mushroom tyrosinase, even better than the kojic acid used as positive reference (IC 50  Moreover, they suggested that this different selectivity could depend on the difference forms of the phosphatidylinositol 3-kinases (PI3Ks) present in the adopted cell models. This hypothesis was proven by the means of in silico and in vitro studies, indicating that the phosphoinositide 3-kinase α (PI3K α) is one of the targets of the compound 44, which influences the fundamental PI3K/Akt signaling pathway [66] leading, ultimately, to cancer cell apoptosis. At the same time, compound 44 reduces the expression of the Vascular Endothelial Growth factor (VEGF) in MCF-7 cells, suggesting a role in inhibiting the angiogenesis process. However, no evidence about the effects on normal cell lines has been reported. Hassan  showed a slight cytotoxic activity against HeLa (IC 50 = 56.7 and 20.8 µM, respectively, versus 5.13 µM of carboplatin) and BHK-21 cells (IC 50 = 32.2 and 60.2 µM, respectively). The mechanism of action for the active compound L5 was deepened, studying the pro-apoptotic mechanism by fluorescence microscopy, cell cycle analysis, caspase-9 and -3 activity, reactive oxygen species (ROS) production, and DNA binding. Compound 52 exhibited disintegrated cell membranes and condensed cellular protein, probably due to the lipids' and proteins' oxidation, suggesting that it could be a potent drug against cancer. Several Schiff bases of tetrahydrocurcumin have been recently reported by Mahal et al. (2019) [73] as potential anticancer agents. The in vitro anticancer activity was evaluated against three human cancer cell lines: human epithelial lung carcinoma (A549) and cervical cancer (HeLa) and human breast adenocarcinoma (MCF-7) cells. Most compounds exhibited moderate to good anticancer activity against all three tested cell lines and were significantly more active than tetrahydrocurcumin. The most interesting was compound 53 (IC 50 = 11.9, 12.7, 4.8 µM, against the three cell lines considered, respectively).  [78] studied several Schiff bases containing a benzothiazole nucleus and studied the DNA binding interaction with pBR322 plasmid DNA by means of electrophoretic mobility shift assay [79]. The anticancer study was performed using the MTT assay. Imine 58 showed 85.82% inhibition of MCF-7 cancer cell lines at a concentration of 200 µg/mL. It was less toxic to normal cells at the concentration required to produce the anticancer effect (IC 50 = 973 µg/mL).  [78] studied several Schiff bases containing a benzothiazole nucleus and studied the DNA binding interaction with pBR322 plasmid DNA by means of electrophoretic mobility shift assay [79]. The anticancer study was performed using the MTT assay. Imine 58 showed 85.82% inhibition of MCF-7 cancer cell lines at a concentration of 200 µg/mL. It was less toxic to normal cells at the concentration required to produce the anticancer effect (IC50 = 973 µg/mL).  a series of Schiff bases and evaluated their cytotoxicity against cancer cell lines (HeLa and MCF-7) and a normal cell line (BHK-21) by means of the MTT assay. Compounds 51 and 53 showed a slight cytotoxic activity against HeLa (IC50 = 56.7 and 20.8 µM, respectively, versus 5.13 µM of carboplatin) and BHK-21 cells (IC50 = 32.2 and 60.2 µM, respectively). The mechanism of action for the active compound L5 was deepened, studying the proapoptotic mechanism by fluorescence microscopy, cell cycle analysis, caspase-9 and -3 activity, reactive oxygen species (ROS) production, and DNA binding. Compound 52 exhibited disintegrated cell membranes and condensed cellular protein, probably due to the lipids' and proteins' oxidation, suggesting that it could be a potent drug against cancer. Several Schiff bases of tetrahydrocurcumin have been recently reported by Mahal et al.
(2019) [73] as potential anticancer agents. The in vitro anticancer activity was evaluated against three human cancer cell lines: human epithelial lung carcinoma (A549) and cervical cancer (HeLa) and human breast adenocarcinoma (MCF-7) cells. Most compounds exhibited moderate to good anticancer activity against all three tested cell lines and were significantly more active than tetrahydrocurcumin. The most interesting was compound 53 (IC50 = 11.9, 12.7, 4.8 µM, against the three cell lines considered, respectively  [78] studied several Schiff bases containing a benzothiazole nucleus and studied the DNA binding interaction with pBR322 plasmid DNA by means of electrophoretic mobility shift assay [79]. The anticancer study was performed using the MTT assay. Imine 58 showed 85.82% inhibition of MCF-7 cancer cell lines at a concentration of 200 µg/mL. It was less toxic to normal cells at the concentration required to produce the anticancer effect (IC50 = 973 µg/mL).  a series of Schiff bases and evaluated their cytotoxicity against cancer cell lines (HeLa and MCF-7) and a normal cell line (BHK-21) by means of the MTT assay. Compounds 51 and 53 showed a slight cytotoxic activity against HeLa (IC50 = 56.7 and 20.8 µM, respectively, versus 5.13 µM of carboplatin) and BHK-21 cells (IC50 = 32.2 and 60.2 µM, respectively). The mechanism of action for the active compound L5 was deepened, studying the proapoptotic mechanism by fluorescence microscopy, cell cycle analysis, caspase-9 and -3 activity, reactive oxygen species (ROS) production, and DNA binding. Compound 52 exhibited disintegrated cell membranes and condensed cellular protein, probably due to the lipids' and proteins' oxidation, suggesting that it could be a potent drug against cancer. Several Schiff bases of tetrahydrocurcumin have been recently reported by Mahal et al. [73] as potential anticancer agents. The in vitro anticancer activity was evaluated against three human cancer cell lines: human epithelial lung carcinoma (A549) and cervical cancer (HeLa) and human breast adenocarcinoma (MCF-7) cells. Most compounds exhibited moderate to good anticancer activity against all three tested cell lines and were significantly more active than tetrahydrocurcumin. The most interesting was compound 53 (IC50 = 11.9, 12.7, 4.8 µM, against the three cell lines considered, respectively  [78] studied several Schiff bases containing a benzothiazole nucleus and studied the DNA binding interaction with pBR322 plasmid DNA by means of electrophoretic mobility shift assay [79]. The anticancer study was performed using the MTT assay. Imine 58 showed 85.82% inhibition of MCF-7 cancer cell lines at a concentration of 200 µg/mL. It was less toxic to normal cells at the concentration required to produce the anticancer effect (IC50 = 973 µg/mL).              antiproliferative activity toward three human carcinoma HepG2 (liver), MCF-7 (breast), and RPE-1 (normal retina pigmented epithelium) cell lines using MTT assay. The results showed that compound 67 was found to be the active candidate against HepG2 (IC50 = 84.2 µM versus 25.3 µM of doxorubicin) and MCF-7 cells (IC50 = 99.4 µM versus 20.9 µM of doxorubicin), while compound 68 was found to be the most active of the series against RPE-1 cells (IC50 = 127.7 µM versus 19.1 µM of doxorubicin). and RPE-1 (normal retina pigmented epithelium) cell lines using MTT assay. The results showed that compound 67 was found to be the active candidate against HepG2 (IC50 = 84.2 µM versus 25.3 µM of doxorubicin) and MCF-7 cells (IC50 = 99.4 µM versus 20.9 µM of doxorubicin), while compound 68 was found to be the most active of the series against RPE-1 cells (IC50 = 127.7 µM versus 19.1 µM of doxorubicin).  [80] showed that compound 67 was found to be the active candidate against HepG2 (IC50 = 84.2 µM versus 25.3 µM of doxorubicin) and MCF-7 cells (IC50 = 99.4 µM versus 20.9 µM of doxorubicin), while compound 68 was found to be the most active of the series against RPE-1 cells (IC50 = 127.7 µM versus 19.1 µM of doxorubicin).

Summary
Schiff bases have long attracted researchers due to their chemical reactivity and to the broad range of pharmacological activities that they exert as such or complexed with metals, including antibacterial, antifungal, anti-inflammatory, antioxidant, and anticancer. They are also employed as versatile tools in several applications such as fluorescent turn-on/turn-off sensors for the determination of diverse analytes. Their easy preparation and capability of forming complexes with almost all metals make them interesting compounds in medicinal chemistry. Recently, several organic compounds bearing Schiff base structure or their complexes with metals were used as effective drugs against cancer. Metal complexes' actions are multiple, depending on the broad range of coordination numbers, geometries, and kinetic properties and, after the worldwide use of cisplatin, different papers reported the importance of Schiff bases' anticancer actions in metal complexes. The reviewed paper indicated that the use of these compounds offered better anticancer properties with respect to the reference molecules, viz., cisplatin, doxorubicin, and vincristine, for instance, both in in vitro and in vivo. It is worthy to note that these compounds displayed the anticancer effects against a very broad variety of tumor cell models, solid or liquid, without hampering, in the most cases, the growth of the normal cells used as control. Furthermore, Schiff bases may target different intracellular regulator enzymes, together with the already known interactions with nuclear DNA, producing cancer cells' death by apoptosis. Finally, different evidence about their ability to modulate the intracellular redox equilibrium, strongly associated with tumor prevention, onset, and progression, have been revealed, confirming the multiple actions exerted by these molecules. In this review, studies regarding mono-and bis-Schiff bases with potent antitumor activity on several cell lines were reviewed. In the future, the study of structure-activity relationships of Schiff bases against cancer cells may help in synthesizing new and effective antitumor agents derived by modification of the already studied imines.

Summary
Schiff bases have long attracted researchers due to their chemical reactivity and to the broad range of pharmacological activities that they exert as such or complexed with metals, including antibacterial, antifungal, anti-inflammatory, antioxidant, and anticancer. They are also employed as versatile tools in several applications such as fluorescent turn-on/turn-off sensors for the determination of diverse analytes. Their easy preparation and capability of forming complexes with almost all metals make them interesting compounds in medicinal chemistry. Recently, several organic compounds bearing Schiff base structure or their complexes with metals were used as effective drugs against cancer. Metal complexes' actions are multiple, depending on the broad range of coordination numbers, geometries, and kinetic properties and, after the worldwide use of cisplatin, different papers reported the importance of Schiff bases' anticancer actions in metal complexes. The reviewed paper indicated that the use of these compounds offered better anticancer properties with respect to the reference molecules, viz., cisplatin, doxorubicin, and vincristine, for instance, both in in vitro and in vivo. It is worthy to note that these compounds displayed the anticancer effects against a very broad variety of tumor cell models, solid or liquid, without hampering, in the most cases, the growth of the normal cells used as control. Furthermore, Schiff bases may target different intracellular regulator enzymes, together with the already known interactions with nuclear DNA, producing cancer cells' death by apoptosis. Finally, different evidence about their ability to modulate the intracellular redox equilibrium, strongly associated with tumor prevention, onset, and progression, have been revealed, confirming the multiple actions exerted by these molecules. In this review, studies regarding mono-and bis-Schiff bases with potent antitumor activity on several cell lines were reviewed. In the future, the study of structure-activity relationships of Schiff bases against cancer cells may help in synthesizing new and effective antitumor agents derived by modification of the already studied imines.

Summary
Schiff bases have long attracted researchers due to their chemical reactivity and to the broad range of pharmacological activities that they exert as such or complexed with metals, including antibacterial, antifungal, anti-inflammatory, antioxidant, and anticancer. They are also employed as versatile tools in several applications such as fluorescent turn-on/turn-off sensors for the determination of diverse analytes. Their easy preparation and capability of forming complexes with almost all metals make them interesting compounds in medicinal chemistry. Recently, several organic compounds bearing Schiff base structure or their complexes with metals were used as effective drugs against cancer. Metal complexes' actions are multiple, depending on the broad range of coordination numbers, geometries, and kinetic properties and, after the worldwide use of cisplatin, different papers reported the importance of Schiff bases' anticancer actions in metal complexes. The reviewed paper indicated that the use of these compounds offered better anticancer properties with respect to the reference molecules, viz., cisplatin, doxorubicin, and vincristine, for instance, both in in vitro and in vivo. It is worthy to note that these compounds displayed the anticancer effects against a very broad variety of tumor cell models, solid or liquid, without hampering, in the most cases, the growth of the normal cells used as control. Furthermore, Schiff bases may target different intracellular regulator enzymes, together with the already known interactions with nuclear DNA, producing cancer cells' death by apoptosis. Finally, different evidence about their ability to modulate the intracellular redox equilibrium, strongly associated with tumor prevention, onset, and progression, have been revealed, confirming the multiple actions exerted by these molecules. In this review, studies regarding mono-and bis-Schiff bases with potent antitumor activity on several cell lines were reviewed. In the future, the study of structure-activity relationships of Schiff bases against cancer cells may help in synthesizing new and effective antitumor agents derived by modification of the already studied imines.