Molecular Hybridization of Naphthoquinones and Thiazoles: A Promising Strategy for Anticancer Drug Discovery

Abstract

Cancer remains one of the leading causes of morbidity and mortality worldwide, demanding the continuous search for novel and more selective chemotherapeutic agents. Quinones, particularly naphthoquinones, constitute a privileged class of redox-active compounds with well-documented antitumor activity. Likewise, thiazoles represent a heterocyclic scaffold widely explored in medicinal chemistry due to their broad pharmacophoric adaptability and diverse biological activities. In this context, this review comprehensively explores the chemical synthesis and anticancer potential of hybrid molecules combining the naphthoquinone and thiazole scaffolds. The hybridization of these pharmacophores has emerged as a powerful strategy to design multitarget antitumor agents. The review summarizes key synthetic methodologies, including Hantzsch, hetero Diels–Alder cycloaddition and multicomponent reactions, leading to structurally diverse hybrids. Particular emphasis is placed on derivatives exhibiting strong cytotoxic effects against a broad spectrum of cancer cell lines (e.g., OVCAR3, MCF-7, A549, HCT-116, HeLa, and Jurkat), low toxicity toward normal cells and well-defined mechanisms of action involving topoisomerase IIα, EGFR, STAT3, and CDK1 inhibition, as well as ROS generation and cell cycle arrest. Among these, certain hybrids displayed nanomolar potency and high selectivity indices, reinforcing their potential as promising lead compounds for anticancer drug development.

1. Introduction

Cancer comprises more than one hundred complex and heterogeneous diseases arising from the accumulation of genetic mutations that disrupt cellular growth control [1,2,3]. Mutation is the central event in carcinogenesis, affecting genes regulating cell growth, DNA repair machinery, and genes controlling apoptosis [4,5]. Cancer is one of the main representatives of the group of noncommunicable diseases [6], responsible for 19.3 million new cases and 10 million deaths in 2020, and the main public health problem in the world [7,8]. Despite great geographic variability, breast cancer is the most common type of cancer among women in approximately 80% of countries. In men, there is considerable international variability, with lung, prostate, and liver cancers being the most common [9].

Chemotherapy is one of the most commonly used methods and is often performed in combination with other types of treatment [10,11]. However, the toxicity of drugs used in chemotherapy in healthy tissues limits the dose that can be administered, just as pharmacokinetic factors limit the amount that can reach the tumor [11,12]. The most important limitation to the success of chemotherapy is tumor resistance to drugs [13,14]. This resistance may be intrinsic or acquired and can arise from several mechanisms, including increased drug efflux, altered drug metabolism, enhanced DNA repair, and target mutations. One of the most common resistance mechanisms is the expression of efflux pumps, such as P-glycoprotein, reducing intracellular drug [15,16,17]. Due to the heterogeneous nature of the tumor, the use of more than one drug, with different pharmacodynamic characteristics, for treatment becomes common [14,18].

Cancer is seen as a disease capable of causing a major impact on society due to its mortality and morbidity rates. Thus, the search for new anticancer molecules remains an urgent priority. Hybrid molecule strategies have emerged as a promising approach to rational drug design, whether in the search for substances with more potent or more selective activity [19,20]. These molecules can be obtained by mixing pharmacophoric groups or even by combining entire drugs [21]. The presence of two moieties with different biological targets provides the possibility of combining drugs into a molecule that exhibits greater efficacy, lower toxicity, and less vulnerability to the development of cancer resistance [22].

Within this context, quinones are widely recognized in the literature for their diverse biological activities, including antineoplastic [23,24,25,26,27,28], antiviral [29,30,31], antifungal [32,33,34], antibacterial [35,36,37], and antiparasitic [38,39,40] effects. Clinically relevant quinonoid drugs include the anthracyclines, such as daunorubicin (1) and doxorubicin (2) (Figure 1), one of the most widely used antitumor drugs in the world, as well as mitomycin C (3) [41].

Figure 1.

Examples of antitumor drugs containing quinonoid moiety.

Quinones can exert their biological activity through different mechanisms, which is a consequence of the structural diversity present in this family of substances. For example, the mechanisms of topoisomerase inhibition [42], inhibition of indoleamine-2,3-dioxygenase [43] inhibition of the P2X7R ion channel [27], DNA alkylation and inhibition of heat shock proteins have already been studied [44,45]. A mechanism of action that tends to be common to molecules in this family is the generation of reactive oxygen species (ROS) within the target cell, causing damage to DNA and other macromolecules, which culminates in apoptosis [46].

Quinones can undergo two-electron reduction to form hydroquinones (4), or one-electron reduction to form semiquinone radicals (5) through reductases. After this reduction, a cyclic process occurs in the presence of oxygen, which mainly produces superoxide radicals (O₂^•−). The superoxide radical is then converted to peroxide by the superoxide dismutase (SOD) enzyme. At this stage, the peroxide can be transformed into water and singlet oxygen by the catalase (CAT) enzyme, or it may undergo the Fenton reaction, forming hydroxyl radical (•OH). The ROS generated in this cycle creates oxidative stress in the cell, causing the scission of DNA molecules, unsaturated lipids, and proteins, and, consequently, leading to cell death (Scheme 1) [47].

Scheme 1.

Summary of the redox cycle for ROS generation.

Similarly, the thiazole scaffold is a crucial building block used in medicinal chemistry for the synthesis and preparation of new compounds with a variety of biological activities, such as antidiabetic [48,49], antihypertensive [50,51], analgesic [52,53], neuroprotective [54,55,56], antiviral [57,58], antiparasitic [59,60], antifungal [61,62], antimicrobial [63,64], and antiproliferative [65,66,67,68]. The thiazole moiety is present in a wide variety of pharmacological classes of antitumor drugs (Figure 2), such as dabrafenib (6), an inhibitor of enzyme B-RAF; tiazofurin (7), a dehydrogenase inhibitor; vosaroxin (8), a topoisomerase inhibition; dasatinib (9), a Bcr-Abl tyrosine kinase inhibitor; and utidelone (10), a microtubule inhibitor [69]. Due to its privileged structure, the thiazole ring can interact with multiple molecular targets relevant to cancer therapy [70], such as EGFR kinase inhibition [71], Akt (PKB) protein kinases inhibition [72], topoisomerase inhibition [73], and inhibition of tubulin polymerization [74].

Figure 2.

Examples of antitumor drugs containing the thiazole moiety.

Given the strong anticancer potential of both quinone and thiazole pharmacophores, combining these two privileged structures into hybrid molecules represents a logical and promising strategy for developing new chemical entities with enhanced biological activity.

2. Quinones–Thiazole Hybrids and Anticancer Activity

The first experiments for the synthesis of hybrid molecules of quinones and thiazoles were reported in the early 1970s, involving the between thiourea and excess 1,4-benzoquinone (11) or 1,4-naphthoquinones (12,13) to produce 2-amino-6-hydroxybenzothiazoles (14) and 2-amino-5-hydroxynaphtho[1,2-d]thiazoles (15), respectively (Scheme 2) [75].

Scheme 2.

First synthetic hybrids of quinones and thiazoles: (a) thiourea, EtOH, conc. HCl, rt.

Only many years later, in the early 1990s, the biological activity of these hybrids began to be studied when the substance BE-10988 (16) (Scheme 3), containing the quinone and thiazole moieties linked by a pyrrole ring, was isolated from the fungus Streptomyces sp. BA10988. Subsequent studies revealed that this compound presented inhibitory activity of the topoisomerase II enzyme [76].

Scheme 3.

First hybrids of indolequinones and thiazoles tested against carcinogenic strains. (a) BrCH₂COMe, EtOH, Δ; (b) H₂, Pd-C, EtOAc; (c) Fremy’s salt, aqueous acetone; (d) aziridine, methanol, rt; (e) 1. MeCSNH₂, KHCO₃, 2. (CF₃CO)₂O, pyridine; (f) NH₄OH, heat; (g) Lawesson’s reagent, toluene, Δ; (h) 1. Pd(OH)₂, H₂, EtOH, 2. Fremy’s salt.

Quinone 16 was tested against murine lymphoid neoplasm lines (P388/S) and analogous lines resistant to vincristine (P388/VCR) and doxorubicin (P388/ADR), demonstrating a marked inhibition of their growth, with IC₅₀ of 0.5, 0.4, and 2.0 μM, respectively [76]. This substance was properly characterized [77] and had its total synthesis carried out [78,79]. These findings motivated the synthesis of indolequinone derivatives varying the substituent linked to the thiazole ring, which had their cytotoxic activity tested against breast (SKBr3) and lung (A549 and PV9) cancer cell lines [80,81]. However, these molecules were not classified as topoisomerase II inhibitors, as they were at least 100-fold less potent than mitoxantrone.

Within this series, the derivatives 17 (IC₅₀ = 0.12 μM), 18 (IC₅₀ = 0.12 μM), and 19 (IC₅₀ = 0.2 μM) bearing aziridyl groups had comparable potency to etoposide and amsacrine against the SKBr3 (human breast) cell line. The derivatives containing the aziridyl group also showed greater cytotoxicity than 16 (IC₅₀ = 8.7 μM) against SKBr3. Because the thiazole ring was attached at different positions, different synthetic routes were developed for the production of these compounds (Scheme 2). Quinone 17 features a bond between position 2 of the thiazole and position 3 of the indolequinone. To construct this ring, the strategy adopted was to start with indole 20 (produced from 4-benzyloxy-5-methoxyindole-2-carbaldehyde in 5 steps [78]), which cyclized to 21 using bromoacetone. A reductive deprotection was performed, yielding 22, which was oxidized with Fremy’s salt to 23. Finally, the methoxy group was displaced by aziridine, yielding 17. Quinone 18 has a bond at carbon 4 of the thiazole ring. Synthesis of this molecule began from an indole with a chloroacetyl group 24 (produced from 4-benzyloxy-5-methoxyindole-2-carbaldehyde in 2 steps [78,81]), which, through a modified Hantzsch reaction, yielded the thiazole ring to give compound 25. The remainder of the route followed the same strategy as the previous methodology. To produce quinone 19, which has a bond at position 2 of the indolequinone ring, was necessary to obtain the indole-2-thiocarboxamide 28, starting from ester 29 (produced from 2-benzyloxy-3-methoxybenzaldehyde in 3 steps [82,83]) via amide 30. In this synthetic route, the thiazole ring was also produced by a Hantzsch reaction, following the same strategy used for 17 and 18.

One of the first naphthoquinone–thiazole hybrids with antineoplastic activity, compounds 33 and 34, were synthesized with an amine bridge linking the two moieties [84]. The synthetic route for producing these molecules (Scheme 4) began with aldehyde 35 (produced from 1,5-naphthalenediol in 4 steps [85,86]), which was condensed with different 2-aminobenzothiazoles to form imines 36. The imines were reduced with LAH to give amines 37, which were subsequently oxidized to a mixture of naphthoquinones 33 and 34. Naphthoquinones 33 were selectively produced using cerium (IV) ammonium nitrate (CAN), and naphthoquinones 34 using chromium (IV) oxide.

Scheme 4.

Synthesis of hybrids of naphthoquinone and benzothiazoles 33 and 34. (a) 2-aminobenzothiazole, TEA, AcOH, Δ; (b) LAH, THF, rt; (c) CAN, MeCN, H₂O; (d) CrO₃, acetone, H₂SO₄, H₂O, rt; (e) NaH, THF, MeI, 0−5 °C; (f) succinic anhydride, toluene, Δ; (g) AlCl₃, DCM, 0–5 °C; (h) acyl chloride, DCM, TEA, 0−5 °C; ADR, adriamycin.

The molecules were tested against a lymphocytic leukemia cell line (L1210) and gastric carcinoma (SNU-1). In general, naphthoquinones 34 showed superior activity than their analogues 33, except for 33a, which exhibited the highest cytotoxic activity against both cell lines. The antineoplastic activity of 33a was comparable to that of adriamycin, motivating the synthesis of new naphthoquinone derivatives 38–41 with different substituents attached to the amine nitrogen. These derivatives were produced by nucleophilic attack of intermediate amine 37 on methyl iodide or acyl substitution on succinic anhydride [87].

The naphthoquinone moieties of 38 and 39 were produced as previously described, and their respective demethylation in 40 and 41 was performed using AlCl₃. Derivatives 38–41 were tested against L1210 and lymphoid neoplasm (P388). Within this series, 40 and 41, presenting free hydroxyls, displayed higher activity than their methoxylated analogues 38 and 39. Interestingly, contrary to what had been previously reported, derivative 40, in which the thiazole moiety is attached at position 2 of the naphthoquinone, had higher activity than its analogue 41, attached at position 6. In this context, derivative 40a showed a rate of growth inhibition (T/C) of 331% in murine S-180 sarcoma cells in the peritoneal cavity, higher than adramycin (234%). Ab initio calculations indicated that substances with greater electrophilicity at carbon 3 tended to show greater cytotoxic activity, as observed for compounds 40 [88]. Continuing the studies with this molecular framework, acyl groups were introduced into the phenolic hydroxyls of 40 using acyl chlorides, producing naphthoquinones 43 [89]. It was found that increasing the carbon chain up to propyl, as in 43a, enhanced antitumor activity, but continuing to increase the chain resulted in decreased potency.

In order to increase the planarity of the molecules to enhance the DNA intercalation process, derivatives 44 were developed, which feature a naphthoquinone fused with a 1H,3H-pyrrolo[1,2-c]thiazole moiety (Scheme 5) [90]. These molecules were produced by heating the corresponding thiazolidine (45) in acetic anhydride in the presence of 1,4-naphthoquinone (12) or juglone (46). The thiazolidines were acylated in situ, followed by a cyclodehydration. Finally, a 1,3-dipolar cycloaddition occurred with 12 or 46. Acylation of thiazolidine 47 with 4-fluorobenzoyl chloride was also performed, yielding 48 as a pure diastereoisomer. Compound 48 was then used to produce the naphthoquinone derivative 49. All derivatives 44 were isolated as pure R enantiomers. These molecules were tested against colorectal adenocarcinoma (WiDr) and melanoma (A375) cell lines but, except for 44a, were inactive against WiDr. Derivatives 44 with hydroxyls in the aromatic ring were inactive against both strains, which contrasts with their parent naphthoquinone, 46, that is active against WiDr (IC₅₀ = 8.8 μM) and A375 (IC₅₀ = 1.23 μM). The results demonstrated that the fusion of the quinone ring with thiazolidine resulted in decreased antineoplastic activity for these cell lines.

Scheme 5.

Synthesis of hybrids of naphthoquinones and fused 1H,3H-pyrrolo[1,2-c]thiazoles 44 and 49. (a) 12 or 46, Ac₂O, 120 °C; (b) pyridine, 4-fluoro-benzoyl chloride, 0 °C; (c) 2, Ac₂O, 120 °C.

Although studies on the antineoplastic activity of naphthoquinone-isothiazole hybrids are less common, the natural compound aulosirazole (50) (Scheme 6), after isolation from the algae Aulosira fertilissima, was identified as a promising antineoplastic molecule by exhibiting selective activity in solid tumors in the Corbett assay [91]. The antineoplastic potential of the molecular scaffold of 50 subsequently motivated investigations of this compound and its analogues as a potential inhibitor of the enzyme indoleamine-2,3-dioxygenase [92].

Scheme 6.

Synthesis of hybrids of isothiazolonaphthoquinones 51. (a) chlorocarbonylsulphenyl chloride, N,N-dimethylurea or N,N-dibenzylurea or benzamide, MeCN or toluene, rt or reflux; (b) 1,4-naphthoquinone (12), xylene, reflux; (c) CAN, MeCN/H₂O (9:1), rt. LMB, leptomycin B. LY, LY294002.

Derivatives 51, which contain an isothiazole ring fused to a quinonoid core, were identified within a library of molecules designed as potential histone deacetylase (HDAC) inhibitors, an important class of targets in antineoplastic drug development [93]. Molecules 51 are structurally analogous to 50 and were able to decrease the viability of LN18 and T98 glioma cells at a concentration of 20 μM. FOXO protein activation is an interesting target for the development of bioactive molecules against tumors due to its tumor suppressive activity [94]. Hundreds of compounds were tested for their ability to induce nuclear translocation of the GFP-FOXO3a reporter protein in the U2OS (osteosarcoma) cell line at a concentration of 10 μM [95]. Among them, 51a induced nuclear FOXO translocation with an EC₅₀ of 1.5 μM. Interestingly, compound 51d, which lacks the two methyl groups attached to the nitrogen atom, did not induce nuclear accumulation of FOXO fluorescent reporter protein at 10 μM, suggesting that the dimethylamino group may be crucial for interaction with the biological target.

Derivatives 51 were produced from 1,3,4-oxathiazol-2-ones 52 (Scheme 6) [93], which were prepared from the reaction between ureas or benzamide (53) and chlorocarbonylsulfynyl chloride. Thermal decarboxylation of 52 generated nitrile sulfides that reacted with 1,4-naphthoquinone (12) in a 1,3-dipolar cycloaddition. Naphthoquinone 51b also underwent debenzylation with CAN, yielding 51d.

Aulosirazole (50) was also isolated from Nostoc sp. UIC 10771 [96]. Along with 50, two other related natural compounds, aulosirazole B (54) and aulosirazole C (55) (Scheme 6), were also identified in the cyanobacteria. Aulosirazole (50) and both analogues were tested against the ovarian cancer cell line OVCAR3. Aulosirazole (50) was the most potent molecule (IC₅₀ = 301 nM), followed by 54 (IC₅₀ = 600 nM) and 55 (IC₅₀ = 3.03 μM). Owing to its high potency, the ability of 50 to induce nuclear accumulation of FOXO3a was further investigated. This molecule induced a significant nuclear accumulation accompanied by a reduction in its cytosolic concentration of FOXO3a when compared to positive and vehicle controls.

Another natural compound with quinonoid and isothiazole moieties is pronqodine A (56) (Scheme 6), isolated from Streptomyces sp. MK832-95F2 [97]. This molecule inhibited bradykinin-induced prostaglandin (PG) release in a concentration-dependent manner in human synovial sarcoma (SW982) cells. Inhibition occurred with IC₅₀ values of 9 nM for PGE2 production, 19 nM for 6-keto-prostaglandin F1α, and 7 nM for PGD2. Data from mechanistic studies suggest that 56 acts as a bioreductant, inhibiting prostaglandin release in activated NQO1-expressing cells.

A study examined the synergistic anticancer potential of the isothiazolonaphthoquinone-based compound 57, an activator of FOXO nuclear–cytoplasmic shuttling, in combination with selinexor (58), an exportin-1 (XPO1) inhibitor, against breast cancer (Figure 3) [98]. Compound 57 promoted the nuclear accumulation of the transcription factor FOXO1, a tumor suppressor whose activity is tightly linked to its subcellular localization. In vitro assays on MCF-7 and MDA-MB-175 breast cancer cell lines revealed that 57 alone significantly enhanced FOXO1 nuclear localization, leading to inhibition of cell migration, induction of apoptosis, and downregulation of oncogenic proteins c-Myc and cyclin D1. When combined with selinexor (58), these effects were markedly potentiated, exhibiting strong synergism (combination index < 0.9). Mechanistically, the dual treatment promoted FOXO1 competition with TCF for β-catenin binding, resulting in suppression of the Wnt/β-catenin signaling pathway, a key regulator of tumor proliferation and metastasis. In vivo, co-administration of both compounds in MCF-7-derived xenografts led to superior tumor growth inhibition (>40% TGI) without systemic toxicity, outperforming either monotherapy.

Figure 3.

Chemical structures of isothiazolonaphthoquinone-based compound (57) and Selinexor (58).

Continuing with naphthoquinones with fused heterocycles, the fusion of the quinonoid ring with thiopyrano[2,3-d]thiazol-2-one led to the synthesis of derivatives 59 (Scheme 7) [99]. Thiazolidines 60 were prepared by treatment of 4-thioxo-2-thiazolidinone 61 with the corresponding aromatic aldehyde. Compounds 59 were synthesized by hetero-Diels–Alder reaction between 60 and 1,4-naphthoquinone (12). The synthesized hybrids were tested on several cell lines, with melanoma cell lines being the most sensitive, especially 59a. The superior activity of 59a was attributed to the presence of the hydroxyl in the para position, since its methylation decreased the antineoplastic activity.

Scheme 7.

Synthesis of hybrids 59 and 62. (a) ArCHO, AcONa, AcOH, 100 °C; (b) 1,4-naphthoquinone (12), AcOH, hydroquinone (cat.), reflux; (c) aromatic aldehyde, amberlyst−15, 100 °C.

Another series of compounds featuring a six-membered ring fused to the quinonoid ring, 62, was synthesized via a multicomponent reaction between lawsone (63), aromatic aldehydes, and 2-amino-benthiazole (64) (Scheme 7) [100]. The compounds were tested against hepatocellular carcinoma (HepG2) and cervical cancer (HeLa) cell lines. Although none of the derivatives had hydroxyl groups in the aromatic ring, it was found that the presence of electron-donating groups increased the cytotoxic activity. Additionally, the results suggested that the rotation of the aromatic ring at position 13 may be important for the activity, since 2,5-disubstituted derivatives had lower cytotoxicity.

In the context of 1,4-naphthoquinones with thiopyrano[2,3-d]thiazol-2-one, new derivatives of type 59 were reported by Lozynsky and co-workers (Scheme 8), with particular emphasis on compound 59b, which was obtained through a multicomponent reaction involving 61, phenylpropionaldehyde, and 1,4-naphthoquinone (12) [101]. This derivative exhibited the most pronounced biological effects within the series, especially against leukemia cell lines, such as Jurkat (IC₅₀ = 0.76 µM), comparable to the reference drug doxorubicin (IC₅₀ = 0.67 µM), and TPH-1 (IC₅₀ = 7.94 µM), being approximately twice as potent as doxorubicin (IC₅₀ = 13.97 µM). Moreover, compound 59b induced pro-apoptotic cytomorphological alterations, mitotic catastrophe in treated KB3-1 cells, necrotic cell death, and demonstrated direct interaction with DNA.

Scheme 8.

Synthesis of hybrids 59 and 65. (a) aldehyde or ketone, EDDA, EtOH, reflux, 10 min; (b) 1,4-naphthoquinone (12) or juglone (46), AcOH, hydroquinone (cat.), reflux; (c) phenylpropionaldehyde, 1,4-naphthoquinone (12) or juglone (46), EDDA, MeCN, reflux. DOX, doxorubicin.

Subsequently, the same research group conducted further studies employing juglone (46) as a synthetic platform to obtain a new series of analogs related to 59, featuring a hydroxyl group on the aromatic ring derived from the naphthoquinone moiety (Scheme 8) [102]. From a biological perspective, compound 65b did not exhibit cytotoxic activity comparable to its counterpart 59b. Although not tested across all the same cancer cell lines, the presence of the hydroxyl group proved detrimental to activity, as observed for MCF-7 (59b: IC₅₀ = 8.94 µM; 65b: IC₅₀ > 50 µM) and HCT-116 p53 (−/−) (59b: IC₅₀ = 12.34 µM; 65b: IC₅₀ > 50 µM). Interestingly, this trend was reversed for the pair of compounds 59c and 65c, both bearing R = furan-2-yl, where 65c exhibited superior cytotoxicity across all tested cell lines compared to its non-hydroxylated analog 59c. Notably, 65c emerged as the most potent derivative of the new series, particularly against the HCT-116 cell line, with an IC₅₀ of 0.60 µM, comparable to the reference drug doxorubicin (IC₅₀ = 0.58 µM) and approximately 10-fold more active than its corresponding non-hydroxylated analog 47c (IC₅₀ = 5.44 µM).

Continuing the investigation of juglone-bearing thiopyrano[2,3-d]thiazole derivatives, Kozak and co-workers report the synthesis and biological evaluation of two compounds of type 65 obtained through a regioselective hetero-Diels–Alder reaction between juglone (46) and substituted 4-thioxothiazolidin-2-ones, similar to that illustrated in Scheme 8 [103]. Both compounds exhibited favorable in silico ADMET properties and strong binding affinities in molecular docking studies, particularly toward CDK2, JAK2, and MAPK8, which are key targets involved in cancer proliferation and apoptosis. In vitro assays demonstrated potent and selective cytotoxicity of 65d and 65e against colorectal adenocarcinoma cell lines HT-29 and DLD-1, with IC₅₀ values ranging from 1.9 to 10.4 µM, while exhibiting low toxicity toward pseudonormal and normal human cells. Mechanistic studies revealed that both compounds induced significant ROS generation, S-phase and G2/M cell cycle arrest, and activation of both intrinsic and extrinsic apoptotic pathways via caspase-3/7, -8, -9, and -10 activation.

Within the quinone family, anthraquinones stand out for their antineoplastic capacity [104], as evidenced by the anthracycline class of chemotherapeutics widely used for the treatment of various types of cancer. The recognized antineoplastic activity of anthraquinones, combined with evidence that fusion with heterocyclic moieties, including imidazoles, generates bioactive compounds of interest, inspired the synthesis of anthraquinone derivatives fused to thiazole 66 [105]. Scheme 9 describes the synthesis process of these derivatives. The amino intermediate 67 was obtained by a Curtius rearrangement from rhein (68) via the acyl azide intermediate 69. Selective iodination of 67 using iodine and silver sulfate afforded intermediate 70. The thiazole moiety of 71 was constructed by the reaction of 70 with different amines in a medium containing carbon disulfide. Finally, demethylation of 71 to give 66 occurred using EtSH/AlCl₃ or BBr₃ at room temperature. Most of the compounds tested against lung cancer (A549) and cervical cancer (HeLa) cell lines exhibited higher cytotoxicity than the starting material rhein (56), with 66a and 66b being the most potent. Within this series, derivatives containing more than one heteroatom in the side chain of the thiazole ring generally showed better activities, possibly due to improved aqueous solubility.

Scheme 9.

Synthesis of thiazole-fused anthraquinones 66. (a) NaH, MeI, DMF, ice bath, overnight; (b) NaOH in H₂O/EtOH, 50 °C, 1 h; (c) Et₃N, DPPA, DMF, rt, 1 h; (d) dioxane, reflux, 30 min; (e) NaOH in H2O, reflux, 4 h; (f) I₂, Ag₂SO₄, ethylene glycol, rt, 3 h; (g) amine, CS₂, K₂CO₃, DMF, 115 °C, 6 h; (h) EtSH, AlCl₃, DCM, rt, 10 h or BBr₃, DCM, rt, 10 h.

Shikonin (72) is a naphthoquinone with excellent antineoplastic activity [106,107,108], and evidence suggests that esterification of its aliphatic hydroxyl enhances this activity [109,110]. Considering these factors, hybrids 73 of shikonin (72) and thiazoles 74 connected by an ester bridge were synthesized [111]. Initially, the thiazole moiety of 74 was constructed by a cyclization between different aryl nitriles (75) and L-cysteine (Scheme 10). Then, the esterification between the acidic group and the aliphatic hydroxyl was performed using 4-dimethyaminopyridine (DMAP) and N,N’-dicyclohexylcarbodiimide (DCC). The presence of the dihydrothiazole group in 73, in general, translated into a lower antineoplastic activity when compared to 72, but conferred greater selectivity toward tumor cell lines. Among the 60 derivatives prepared, compound 73a showed greater cytotoxicity against the HeLa cell line (IC₅₀ = 3.14 μM) in addition to a lower activity against the non-tumor cell line VERO (IC₅₀ = 92.2 μM) in comparison to 72 (IC₅₀ HeLa = 5.75 μM; IC₅₀ VERO = 6.76 μM). Docking and confocal microscopy experiments were performed, which indicated that 73a could bind to tubulin at the paclitaxel binding site, leading to tubulin polymerization and mitotic rupture.

Scheme 10.

Synthesis of thiazole derivatives of shikonin 73. (a) L-cysteine, NaHCO₃, MeOH/H₂O, NaOH, rt; (b) L-cysteine, NaHCO₃, EtOH, 100 °C; (c) Shikonin (72), DMAP, DCC, DCM, rt, 12 h.

The biological activities of naphthoquinone and thiazole hybrids have already been demonstrated throughout this review. In particular, derivatives 59 [99] inspired the synthesis of thiazole derivatives 76 of dichlone (77) [112]. The synthesis began with a Hantzsch condensation between α-haloketones 78 and thiourea, producing 2-aminothiazoles 79. These intermediates then underwent nucleophilic vinylic substitution in 77, providing derivatives 76 (Scheme 11). These molecules were tested against HeLa and SH-SY5 tumor cell lines. Against the SH-SY5 cell line, it was observed that electron-withdrawing groups were associated with greater antineoplastic activity, as was the case for derivative 76a, R = p-NO₂Ph (IC₅₀ = 0.004 μM), when compared with 76b, R = Ph (IC₅₀ = 1.8 μM). However, these derivatives did not demonstrate considerable activity against HeLa cells.

Scheme 11.

Synthesis of thiazole derivatives of dichlone 76. (a) thiourea, EtOH, 70 °C, 1 h; (b) 77 (dichlone), K₂CO₃, DMF, rt, 5 h. CHL, chloroquine.

A series of naphtho[2′,3′:4,5]thiazolo[3,2-a]pyrimidine hybrids 80 was successfully synthesized through molecular hybridization of thiazolopyrimidine and naphthoquinone scaffolds. The synthetic strategy started with the preparation of 6-aryl-2-mercapto-1,6-dihydropyrimidine-5-carbonitriles (81) via modified Biginelli condensation of ethyl cyanoacetate, diverse aromatic aldehydes (82), and thiourea, employing potassium carbonate as the base [113]. The target derivatives (80) were subsequently obtained via the reaction of thiazolopyrimidines (81) with dichlone (77), as shown in Scheme 12.

Scheme 12.

Synthesis of naphtho[2′,3′:4,5]thiazolo[3,2-a]pyrimidine hybrids 80. (a) thiourea, ethyl cyanoacetate, K₂CO₃, EtOH, reflux, 2 h; (b) 77 (dichlone), DMF, rt, 12 h. DOX, doxorubicin and ERL, erlotinib.

Biological evaluation revealed that several derivatives, particularly 80a, 80c, and 80i, exhibited superior cytotoxicity against MCF-7, A549, and HCT-116 cancer cell lines when compared with doxorubicin and erlotinib, used as reference drugs. Hybrid 80i demonstrated the most potent cytotoxic activity, with IC₅₀ values ranging from 1.55 to 2.29 µM, making it approximately three times more active than doxorubicin and up to four times more active than erlotinib, as shown in Scheme 12. Mechanistic studies demonstrated that these hybrids act as dual inhibitors of topoisomerase IIα and EGFR. They function as catalytic inhibitors of topo IIα while effectively suppressing EGFR kinase activity, with compound 80a showing nanomolar potency (IC₅₀ = 16.03 nM). Furthermore, these derivatives induced apoptosis through upregulation of p53 and pro-apoptotic protein Bax, downregulation of anti-apoptotic protein Bcl-2, and activation of caspases-7 and -9, most prominently for compound 80i. Molecular docking supported strong binding interactions with the ATPase domain of topo IIα and the active site of EGFR, in line with the observed multitarget activity. Collectively, these findings identify compounds 80a, 80c, and 80i as promising dual-acting anticancer prototypes with potential for further development as clinically relevant multitarget agents.

A diverse library of thiazole derivatives incorporating the N-2,5-dimethylphenylthioureido acid scaffold was synthesized, leading to the identification of compound 83, a naphthoquinone-fused thiazole with promising anticancer potential [114]. Structurally, the fusion of the thiazole core with a naphthoquinone moiety conferred interesting biological activity, underscoring the importance of extended conjugation and redox-active substituents in enhancing cytotoxic properties. Compound 83 exhibited broad-spectrum anticancer activity, significantly reducing the viability of both pulmonary adenocarcinoma (A549) and colorectal adenocarcinoma (Caco-2) cell lines to 14% and 13.1%, respectively, after 24 h of exposure, compared to untreated control (Scheme 13).

Scheme 13.

Synthesis of thiazole-fused 1,4-naphthoquinone 83. (a) 77 (dichlone), sodium acetate, glacial acetic acid, rt, 24 h, then 70–80 °C, 12 h.

A new series of naphthoquinothiazole derivatives was designed and synthesized through scaffold modification of napabucasin (88), aiming to enhance STAT3 inhibition and redox modulation [115]. Among the compounds, 85e demonstrated the most potent anticancer activity, with IC₅₀ values of 46.3 nM, 66.4 nM, and 53.8 nM against HCT116, HepG2, and HeLa cells, respectively, while maintaining low cytotoxicity toward normal fibroblasts, resulting in a high selectivity index. Mechanistic studies revealed that 85e induces S-phase arrest, apoptosis, and strong intracellular ROS production, accompanied by suppression of AKT phosphorylation. Furthermore, 85e effectively inhibited STAT3 phosphorylation at Tyr705, with direct binding confirmed by SPR and CETSA assays, and docking simulations showed stable interactions with the STAT3 SH2 domain. Importantly, in vivo experiments using HCT116 xenografts confirmed that 85e significantly suppressed tumor growth without observable toxicity (Scheme 14).

Scheme 14.

Synthesis of naphthoquinothiazole derivatives 85. (a) Na₂S·9H₂O, EtOH/H₂O, 90 °C; (b) Glyoxylic acid, 0 °C; (c) NH₂R, HATU, TEA, DMF.

A high-throughput screening of 5700 compounds from the National Cancer Institute (NCI) chemical library using Schizosaccharomyces pombe deletion mutants enabled the identification of several potential anticancer agents. Among these, compound 89 emerged as particularly promising because it exhibited lower IC₅₀ values against S. pombe and demonstrated higher cytotoxicity toward yeast cells than toward mammalian cells, suggesting that these compound may selectively target yeast-specific pathways and hold potential for further development as an anticancer agents [116].

Inspired by these results, Yanik and coworkers synthesized a series of 2-aminonaphtho[2,3-d][1,3]thiazole-4,9-dione derivatives (90) via the condensation of 2-amino-3-bromo-1,4-naphthoquinone (91) with various isothiocyanates under CuO nanoparticle-catalyzed conditions, forming of thiourea intermediates that cyclized to yield the desired heterocycles (Scheme 15) [117].

Scheme 15.

Synthesis of 2-aminonaphtho[2,3-d][1,3]thiazole-4,9-dione derivatives 90. (a) NH₃, H₂O, dioxane, 2.5 h, rt; (b) isothiocyanate derivatives, CuO nanoparticles, K₂CO₃, H₂O, 24 h, reflux.

The compounds were evaluated for their cytotoxic potential against MDA-MB-231 (breast), HeLa (cervical), and MKN-45 (gastric) cancer cell lines using MTT assays. Among them, compounds 90b and 90d displayed the most potent antiproliferative activity, with IC₅₀ values of 0.269 μM (HeLa) and 0.276 μM (MDA-MB-231), respectively, and high selectivity index values. Compound 90a also showed strong inhibition in HeLa (IC₅₀ = 0.336 μM) and MKN-45 (IC₅₀ = 8.769 μM) cells. Molecular docking studies suggested favorable binding of the ligands to the human DNA topoisomerase IIβ (hDNA TopoIIβ) active site, stabilized by hydrogen bonding and cation–π interactions, supporting a potential mechanism of TopoIIβ inhibition.

A new series of thiophene-based heterocycles incorporating thioureido substituents was synthesized, among which only ethyl (Z)-2-((1-allyl-4,5-dioxo-4,5-dihydronaphtho[1,2-d]thiazol-2(1H)-ylidene)amino)-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylate (93) has the naphthoquinone core. Compound 93 was obtained by reaction of intermediate 94 and 2,3-dihydro-2,3-epoxy-1,4-naphthoquinone (95) [118]. In vitro assays revealed that these derivatives exhibited strong growth inhibition toward HCT-116 cells (61.55914% for 93). However, the IC₅₀ was not reported for compound 93 (Scheme 16).

Scheme 16.

Synthesis of ethyl (Z)-2-((1-allyl-4,5-dioxo-4,5-dihydronaph tho[1,2-d]thiazol-2(1H)-ylidene)amino)-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylate 93. (a) 95, EtOH, reflux, 5 h.

A series of paracyclophanyl–thiazole hybrids incorporating a naphthoquinone scaffold was designed and synthesized as potential cyclin-dependent kinase 1 (CDK1) inhibitors, exhibiting pronounced antiproliferative activity against melanoma cells. The target compounds (96) were synthesized via the Eschenmoser coupling reaction of N-substituted thioureas (97) with dichlone (77), affording the desired conjugates in satisfactory yields (Scheme 17) [119].

Scheme 17.

Synthesis of paracyclophanyl–thiazole naphthoquinone hybrids 96. (a) 64, PhP₃, Et₃N, CH₃CN, reflux, 10–14 h. DIN, dinaciclib.

Comprehensive biological screening conducted by the NCI demonstrated that these derivatives exhibited strong cytotoxic profiles across a broad spectrum of cancer cell lines, with 96c, 96d, and 96e inducing complete growth inhibition in several panels, notably melanoma, colon, and CNS cancers. Among them, compound 96c emerged as the most potent, displaying an IC₅₀ value of 0.81 μM against the SK-MEL-5 melanoma cell line, markedly superior to the reference CDK inhibitor dinaciclib (IC₅₀ = 5.97 μM). Mechanistic investigations revealed that 96c selectively inhibited CDK1 (IC₅₀ = 54.8 nM), promoted cell-cycle arrest at the G₂/M phase, and induced apoptosis via caspase-3 overexpression and phospho-Tyr15 downregulation. Molecular docking studies further supported these findings, highlighting strong hydrogen-bonding interactions with GLN137 and ILE15, together with enhanced π–π stacking interactions mediated by the paracyclophane moiety.

A series of 1,4-naphthoquinone–thiazole hybrids (98) was synthesized through the cyclization of 1,4-naphthoquinone thioureas (99) with α-bromoketones in good yields (Scheme 18) [120]. The hybrids were evaluated for antiproliferative activity against PC3 human prostate cancer cells, with compounds 98f (IC₅₀ = 20.10 µM) and 98e (IC₅₀ = 21.14 µM) showing the highest cytotoxic effects. Hoechst staining confirmed that 98f induces apoptosis in a dose-dependent manner. Inverse molecular docking and molecular dynamics simulations suggested that the anticancer mechanism of 98f is likely mediated through p38 MAP-kinase inhibition, with stable binding confirmed over 300 ns. Additionally, solubility studies revealed that derivatives bearing a fluorine atom at R₁ and a tert-butyl substituent at R display improved solubility and biological performance.

Scheme 18.

Synthesis of 1,4-naphthoquinone–thiazole hybrids 98. (a) isothiocyanates, acetone, reflux, 18 h; (b) α-bromoketones, acetone, reflux, 12 h. DOX, doxorubicin.

3. Conclusions

In summary, naphthoquinone–thiazole hybrids clearly demonstrate their relevance as privileged scaffolds in anticancer drug discovery. The combination of the redox-active naphthoquinone core with the pharmacophoric versatility of the thiazole ring has generated a structurally diverse range of compounds with notable cytotoxic profiles. From a medicinal chemistry perspective, molecular hybridization has proven to be an effective strategy for designing multitarget agents capable of overcoming common limitations of conventional chemotherapeutics, such as resistance and systemic toxicity. Continued exploration of structural modifications, mechanistic pathways and biological validations will be essential to advance these hybrid frameworks toward clinically relevant anticancer agents.

Author Contributions

Conceptualization, L.G.C.d.M., T.B.S. and D.R.d.R.; writing—original draft preparation, L.G.C.d.M. and T.B.S.; writing—review and editing, L.G.C.d.M., T.B.S. and D.R.d.R.; supervision, D.R.d.R.; funding acquisition, D.R.d.R. All authors have read and agreed to the published version of the manuscript.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

No new data were created or analyzed in this study. Data sharing is not applicable to this article.

Conflicts of Interest

The authors declare no conflicts of interest.

Funding Statement

The authors thank the agencies that finance our research: CNPq, CAPES, and FAPERJ. CNPq grants (306892/2022-7) and FAPERJ grants (E-26/211.068/2019, E-26/201.318/2021; E-26/204.289/2024) are gratefully acknowledged. This study was financed in part by the Coordination for the Improvement of Higher Education Personnel—Brazil (CAPES) -Finance Code 001.