Therapeutic Potential of 1,3,4-Thiadiazole Scaffolds in Cancer: Targeting Receptor Tyrosine Kinases, Apoptosis, and Epigenetic Pathways

Cancer is one of the leading causes of death worldwide, with nearly 19.3 million new cases and 10 million deaths reported in 2020 [1]. Despite progress in chemotherapy, radiotherapy, and immunotherapy, many treatments still suffer from poor selectivity, systemic toxicity, and the development of resistance. This created an urgent need for novel molecules with improved efficacy and safety [2]. Heterocycles play a central role in medicinal chemistry because of their structural diversity and ability to engage in multiple interactions with biological targets.  Among them, 1,3,4-thiadiazoles have attracted significant attention. These nitrogen–sulfur heterocycles display diverse pharmacological activities, including antimicrobial, anti-inflammatory, anticonvulsant, and notably, anticancer effects. Their chemical properties, such as lipophilicity, hydrogen-bonding ability, and metabolic stability, make them suitable scaffolds for drug design[3-5]. Recent studies show that thiadiazole derivatives act as multi-target anticancer agents. They inhibit receptor tyrosine kinases (RTKs) such as VEGFR-2[6], EGFR[7], FLT3[8], and BRAF[9], thereby suppressing tumor proliferation and angiogenesis. They also trigger apoptosis through caspase activation[10] and the inhibition of CDK1[11], leading to cell cycle arrest. In addition, thiadiazoles can function as histone deacetylase (HDAC) inhibitors[12], offering epigenetic regulation with therapeutic potential. Some analogues also target metabolic enzymes such as carbonic anhydrases[13], underscoring their broad activity profile.

Chemistry of 1,3,4-Thiadiazole

Thiadiazoles are an important group of heteroaromatic five-membered rings composed of two nitrogen atoms and one sulfur atom. Several positional isomers are known, such as 1,2,3-thiadiazole, 1,2,4-thiadiazole, 1,2,5-thiadiazole, and 1,3,4-thiadiazole(Figure 1). Among these, the 1,3,4-thiadiazole framework has attracted the greatest research interest. This nucleus demonstrates pronounced aromatic character but exhibits only weak basicity, which is attributed to the electron-withdrawing influence of the sulfur atom.

Figure 1. Isomers of thiadiazole.

The 1,3,4-thiadiazole ring demonstrates considerable stability in acidic aqueous media but is prone to ring cleavage under basic conditions. Owing to the strong electron-withdrawing influence of the nitrogen atoms, the ring system is highly electron-deficient, rendering it relatively unreactive toward electrophilic substitution while remaining more susceptible to nucleophilic attack. Substitution at the 2′ or 5′ positions, however, markedly enhances its reactivity, facilitating the formation of a wide range of derivatives. These unique structural and electronic features have contributed to the extensive use of 1,3,4-thiadiazole derivatives in both pharmaceutical development and materials science[14,15].While the biological significance of the 1,3,4-thiadiazole nucleus is well recognized, its ease of synthesis has further enhanced its role in drug discovery. Typically, the heterocyclic core is prepared through the cyclization of thiosemicarbazides with carboxylic acids or their derivatives [16], or via oxidative ring closure of thiohydrazides[17]. In addition, cyclocondensation reactions with aldehydes or ketones provide versatile routes that enable the incorporation of a wide variety of substituents [15].

Anticancer Potential of Thiadiazole Derivatives

The 1,3,4-thiadiazole scaffold is increasingly recognized as a privileged heterocyclic nucleus in anticancer research, due to its structural versatility and ability to engage multiple oncogenic pathways. Systematic reviews have highlighted its value in cancer drug discovery, particularly citing the 1,3,4-isomer as exceptionally productive for cytotoxic activity [18,19]. Key pharmacological mechanisms include: carbonic anhydrase (CA) inhibition, particularly CA IX and II [20]; cell-cycle arrest and apoptosis induction [21], and multi-target engagement [10]including CA IX, Src/Abl kinases, topoisomerase II, and DNA replication enzymes—underscoring the broad mechanistic reach of the scaffold. 1,3,4-Thiadiazole derivatives have been reported as HDAC inhibitors [23] with potential antitumor activity. Several recent synthetic and screening studies support these trends.

3.1 Receptor Tyrosine Kinase Inhibition

Receptor tyrosine kinases (RTKs) are key regulators of cell proliferation, angiogenesis, and survival, and their dysregulation is a hallmark of multiple cancers. The 1,3,4-thiadiazole scaffold has shown strong potential as a kinase-directed pharmacophore due to its ability to fit within ATP-binding pockets and form hydrogen-bond interactions with hinge residues. Several derivatives targeting EGFR, VEGFR, and c-Met [35] have been reported, underscoring their versatility as RTK inhibitors. Ibrahim H. Eissaet al., reported the design and evaluation of a new thiadiazole analogue (TDA) targeting VEGFR-2 as a potential anticancer agent. TDA was evaluated through in silico studies (molecular docking, MD simulations, MM-PBSA, DFT, and ADMET) and in vitro assays. TDA exhibited promising VEGFR-2 inhibitory activity (IC?? = 0.65 μM), surpassing the standard sorafenib (IC?? = 1.08 μM). Cytotoxicity assays revealed potent growth inhibition against MCF-7 (IC?? = 7.63 μM) and HepG2 (IC?? = 11.60 μM) cancer cell lines.Mechanistic studies showed that TDA induced cell cycle arrest at the G0/G1 phase, upregulated pro-apoptotic Baxand downregulated anti-apoptotic Bcl-2, leading to apoptosis in MCF-7 cells. Additionally, TDA significantly inhibited cell migration and angiogenesis. Docking and simulation studies confirmed stable interactions with VEGFR-2 active site residues, consistent with the experimental findings.The authors concluded that TDA is a promising VEGFR-2inhibitor with strong anticancer potential, warranting further in vivo evaluation and structural optimization as a lead candidate for anticancer drug development[2]. Aram Farajiet al., reported a series of twenty-six quinazolin-4(3H)-one derivatives bearing a thiadiazole–urea moiety (final products, series 9a–z) inspired by sorafenib/SKLB1002 for anti-proliferative and anti-angiogenic evaluation on VEGFR-overexpressing models (PC3, HepG2, T47D, HUVEC). The design replaced sorafenib’s central phenyl with a 1,3,4-thiadiazole and varied lipophilic meta/para substitutions on the terminal phenyl-urea. Among PC3 cells, 9l was most potent (IC?? = 7.8 μM), with 9f, 9k, 9m also active; notably, removing a C-7 Cl on the quinazolinone improved PC3 potency. In HepG2 cells, 9k was strongest (IC?? = 6.2 μM), ~4× more active than sorafenib (25.0 μM), while 9l (14.5 μM) and 9m (14.6 μM) were promising; para-Cl outperformed meta-Cl (e.g., 9c vs 9k, 9t vs 9x), and Me groups minimized activity; CF? analogs (9g, 9u) were weak across lines. In T47D cells, only 9p (11.2 μM) and 9q (10.8 μM) approached sorafenib’s effect. Against HUVEC, 9t (4.5 μM) and 9y (6.1 μM) were most potent, with 9v (10.9 μM) and 9x (10.4 μM) moderate; here, adding C-7 Cl generally enhanced activity. Mechanistic assays showed Annexin V/PI apoptosis in HUVEC and moderate in-ovo anti-vascular effects (e.g., 9f reduced CAM vessel branches by 44%vssorafenib64%), while Western blots indicated 9f and 9y reduced VEGFR-2 phosphorylation[4]. Mahfam Moradiet al., reported the design, synthesis, and biological evaluation of a novel series of furo[2,3-d]pyrimidin-4-ylsulfanyl-1,3,4-thiadiazole derivatives as potential FLT3-ITD inhibitors for acute myeloid leukemia (AML). Several derivatives exhibited nanomolar FLT3-ITD inhibitory activity with strong selectivity over K562 cells. The most active compounds significantly suppressed FLT3 phosphorylation and downstream signaling molecules including STAT5 and ERK1/2 in FLT3-ITD–expressing cells. In Ba/F3 cells expressing FLT3-ITD or FLT3-ITD-F691L mutant, the potent derivatives overcame resistance to both sorafenib and quizartinib. Molecular docking studies indicated that these compounds bind tightly to the active site of FLT3 in a type II manner. The authors concluded that compound 49 in the series of furo[2,3-d]pyrimidin-1,3,4-thiadiazole derivatives represents promising FLT3-targeted leads for AML therapy, meriting further optimization [8]. SeragMI, Tawfik SS et al., reportedthe design and synthesis of novel 1,3,4-thiadiazole-based hybrids as epidermal growth factor receptor (EGFR) inhibitors for anticancer therapy. The compounds were screened for cytotoxicity against colon (HCT-116), liver (HepG-2), and breast (MCF-7) cancer cell lines, with WI-38 fibroblasts used as normal controls. Among the series, compound 9a exhibited superior activity compared to doxorubicin, while showing reduced toxicity toward normal cells. EGFR inhibitory assays identified compounds 4a, 6b, 8b, 9a, and 9d as the most potent, with 9a and 8b achieving IC?? values in the submicromolar range, approaching the activity of the reference inhibitor gefitinib. Mechanistic studies revealed that compound 9a induced cell cycle arrest at G2/M phase, enhanced apoptosis by ~28%, and triggered ROS-mediated mitochondrial apoptotic pathways. Furthermore, molecular docking supported strong binding interactions of 9a and 8b with the EGFR active site, corroborating the biological findings. Collectively, these results highlight 1,3,4-thiadiazole scaffolds as promising candidates for further optimization as selective EGFR-targeted anticancer agents[16]. Xie, Wang, Sun et al., developed a novel series of N-(1,3,4-thiadiazol-2-yl)benzamide derivatives containing a 6,7-methoxyquinoline scaffold as dual inhibitors of EGFR and HER-2. The design was inspired by Cabozantinib, a multi-kinase inhibitor used clinically but limited by significant toxicity. The synthetic pathway involved sequential condensation, O-alkylation, hydrolysis, cyclodehydration, and benzamide coupling, affording compounds YH-1 to YH-20. The derivatives were evaluated through kinase inhibition assays against EGFR, HER-2, HER-3, HER-4, and VEGFR-2, in vitro cytotoxicity assays on A549, H1975, MCF-7, and SK-BR-3 cells, and toxicity screening against normal Beas-2B and MCF-10A cells. Additional mechanistic studies included apoptosis assays, ROS measurement, cytochrome c release, angiogenesis assays, and in vivo evaluation in a SK-BR-3 xenograft model. Several derivatives selectively inhibited EGFR and HER-2 while showing weak activity against HER-3, HER-4, and VEGFR-2. The lead compound, YH-9, displayed potent dual inhibition with IC?? values of 29.3 nM for EGFR and 55.7 nM for HER-2, outperforming Cabozantinib in cell-based assays, particularly in SK-BR-3 breast cancer cells where it was about 5.5-fold more effective. YH-9 significantly suppressed tumor growth and angiogenesis in vivo while exhibiting lower systemic toxicity. Mechanistic studies revealed that YH-9 induced apoptosis by generating ROS and promoting cytochrome c release, while also decreasing VEGF and bFGF secretion, thereby exerting anti-angiogenic effects. Structure–activity relationship analysis indicated that halogenated phenyl derivatives were generally the most potent, with the ortho-bromophenyl substitution in YH-9 providing optimal activity. The authors concluded that YH-9 represents a potent, selective, and less toxic dual EGFR/HER-2 inhibitor with promising potential for further preclinical development [27].

Induction of Apoptosis and Cell Cycle Arrest

Apoptosis, or programmed cell death, is a tightly regulated process essential for maintaining tissue homeostasis and eliminating damaged or unwanted cells. In cancer, disruption of apoptotic signaling allows malignant cells to survive, proliferate, and resist conventional therapies. Restoring or enhancing apoptosis has therefore emerged as a key strategy in anticancer drug development. Multiple pathways govern apoptotic cell death, primarily the intrinsic (mitochondrial) and extrinsic (death receptor–mediated) cascades, both of which converge on caspase activation and execution of cellular dismantling. Recent advances in medicinal chemistry have highlighted heterocyclic scaffolds, such as thiadiazoles and thiazolidinones, as promising structural frameworks capable of modulating apoptotic targets. These compounds not only induce cell cycle arrest and oxidative stress but also interact with molecular regulators like Bcl-2 family proteins and caspases, offering opportunities for designing selective and potent anticancer agents [31, 32]. Janowska S, Khylyuk D et al., designed and synthesized 15 new 2,5-disubstituted 1,3,4-thiadiazoles (ST1–ST15) and evaluated them against MCF-7 and MDA-MB-231 breast cancer cells. The most active was ST10 [2-(2-trifluoromethylphenylamino)-5-(3-methoxyphenyl)-1,3,4-thiadiazole] with IC50 = 49.6 μM (MCF-7) and 53.4 μM (MDA-MB-231); ST8 and ST9 were moderate, and normal fibroblasts were less affected. [^3H]-thymidine assays confirmed antiproliferative effects. Docking (Topo IIβ, Caspase-3/8, Bcl-xL, Bcl-2, BAX) suggested a multitarget apoptosis-oriented mechanism, with Caspase-8/BAX interactions strongest for ST15 and ST10. SwissADME/admetSAR indicated Lipinski compliance and high intestinal absorption; some analogs had predicted carcinogenicity, not including ST10. The authors propose ST10-like scaffolds as apoptosis-targeting anticancer leads [10].  Janowska S, Khylyuk D et al., synthesized a focused set of 1,3,4-thiadiazoles bearing a 3-methoxyphenyl substituent (SCT-1/2/4/5/6) to probe substituent positioning. Overall activity was weak: SCT-4 was the best vsMCF-7 (cell viability 74% at 100 μM; DNA biosynthesis 70% at 100 μM) and SCT-5 the best vsMDA-MB-231 (viability 75%; DNA 71% at 100 μM). Fibroblasts remained ≥90% viable at 100 μM. In silico profiling indicated caspase-8–centric interactions; comparison with earlier analogs showed that placing 3-methoxyphenyl at C-5 of the thiadiazole (rather than on the anilide nitrogen) improves potency, aligning these scaffolds with apoptosis-targeting mechanisms [11]. Hekal M, Farag P et al., synthesized 24 novel 1,3,4-thiadiazole derivatives and evaluated their anticancer potential, focusing on cell-cycle effects, apoptosis/necrosis, molecular modeling against CDK1, and ADMET properties. They screened cytotoxicity across MCF-7, HCT-116, PC-3, and HepG2 cells with selectivity vs WI-38 fibroblasts, then performed flow-cytometric cell-cycle analysis and annexin V/PI assays; docking to CDK1 (PDB: 6GU7) was validated by redocking the co-crystallized inhibitor FB8, and in silico pharmacokinetics were profiled (Lipinski/Veber, BBB, absorption). Five derivatives (4, 6b, 7a, 7d, 19) were active; 4, 7a, and 7d were non-selective and excluded, while 6b and 19 showed IC50 < 10 µM and good selectivity (SI ≈ ~5 and ~6) with MCF-7 being most responsive. In MCF-7 cells, 19 induced G2/M arrest (≈23.3% vs 4.6% control) and increased early apoptosis (≈15%) and necrosis (≈15%), whereas 6b markedly raised the sub-G1 fraction (≈53.6%) consistent with necrosis (≈12.5%) and minimal apoptosis. Docking indicated 19 forms key H-bond/π-interactions (e.g., Leu83, Asp86, Lys89) and is a credible CDK1 inhibitor candidate; the protocol showed low RMSD on redocking FB8. ADMET predictions suggested low BBB penetration for 6b/19, high intestinal absorption, and no Lipinski violations for these hits. The authors conclude 6b and especially 19 are promising thiadiazole-based leads that act via G2/M arrest/apoptosis (19) or necrosis (6b) and warrant further optimization as anticancer agents [12].

Epigenetic Regulation

Histone Deacetylase (HDAC) Inhibition

Histone deacetylases (HDACs) are central to epigenetic regulation, where they influence chromatin structure and gene transcription. In cancer, abnormal HDAC activity often silences tumor suppressor genes and facilitates unchecked cell growth. Inhibiting HDACs with small-molecule HDAC inhibitors (HDACis) helps restore acetylation balance, reactivate apoptosis-related genes, and improve responsiveness to chemotherapy and immunotherapy, making them an established class of anticancer drugs. Resistance to conventional cancer therapies often stems from epigenetic reprogramming, which modifies gene expression without altering DNA sequences. Epigenetic therapies—such as HDAC and DNA methyltransferase inhibitors—can restore sensitivity to drugs by reactivating suppressed signaling pathways. When combined with targeted therapies, these agents provide a means to counteract resistance and enhance long-term treatment efficacy [29, 30]. Maji A, Himaja Aet al., designed and synthesized 21 novel 1,3,4-thiadiazole-2-yl-imino-thiazolidine-4-one hybrid molecules and assessed their anticancer properties through MTT assays across multiple cancer cell lines (MCF-7, PC-3, 4T1, MDA-MB-231, MOC2) and normal HEK-293 embryonic cells. All compounds demonstrated higher cytotoxic activity than BG45, a selective HDAC3 inhibitor used as a reference, and displayed greater selectivity toward cancer cells over normal fibroblasts. Among the series, compound 6e emerged as the most potent, with an IC?? of 3.85 μM against MCF-7 cells, surpassing BG45 in efficacy. Mechanistic investigations revealed that 6e induced apoptosis (~25.3%), caused cell cycle arrest at the G?/G? phase, and promoted intracellular ROS accumulation with nuclear fragmentation, consistent with apoptosis-mediated cytotoxicity. Collectively, the study highlights compound 6e as a promising lead scaffold, combining thiadiazole and thiazolidinonepharmacophores, with potential utility as a selective HDAC-modulating anticancer agent [26]. Chen Chenet al., reported multiple series (2a–l, 3a–x, 4a–x, 7a–l, 8a–l) of 2,5-diphenyl-1,3,4-thiadiazole hydroxamate derivatives designed as HDAC inhibitors with DNA-minor-groove binding capacity. The most active compounds in HeLa nuclear extract were 4j and 4k (IC?? = 38 nM and 57 nM), and, in isozyme assays, 4j and 4k inhibited HDAC1 more potently than SAHA (IC?? = 15 nM and 19 nMvs SAHA 70 nM), with comparable potency to SAHA on HDAC6/8 and negligible activity on HDAC4/5/7/9 at 1–10 µM. Compound 4j showed the strongest antiproliferative activity across tested solid and leukemia cell lines, induced apoptosis via caspase-3, and increased acetyl-histone H3 and acetyl-α-tubulin levels. In vivo, 4j significantly suppressed tumor growth in the MC38 model (TGI = 66.0% at 50 mg/kg; 85.2% at 100 mg/kg), outperforming SAHA (51.5% at 100 mg/kg). Biophysical assays and docking/MD confirmed minor-groove DNA binding and stronger HDAC1 engagement than SAHA. The authors concluded that 4j is a promising lead for further optimization as an antitumor HDAC/DNA-binding agent [12].

Metabolic Enzyme Inhibition

Carbonic Anhydrase Inhibition

Carbonic anhydrases (CAs), particularly isoforms IX and XII, are key enzymes that regulate intracellular pH and support tumor survival in hypoxic environments. Inhibiting these isoforms disrupts tumor acid–base homeostasis, reduces metastasis, and enhances sensitivity to standard cancer treatments. Sulfonamide-based CA inhibitors are among the most studied chemical classes with demonstrated anticancer potential. Altered metabolism is a hallmark of cancer, enabling tumor cells to rapidly adapt and sustain uncontrolled growth. Beyond carbonic anhydrases, enzymes involved in glycolysis, glutamine utilization, and lipid biosynthesis represent attractive intervention points. Blocking these pathways can slow tumor progression and work synergistically with other treatments to overcome therapeutic resistance [33, 34]. MujahidAbaset al., reported a novel series of sulfonamide-based 2,5-disubstituted-1,3,4-thiadiazole derivatives (5a–j) designed as potential carbonic anhydrase (CA) inhibitors. Among these, compound 5h exhibited the most potent CA inhibitory activity (IC?? = 0.60 ± 0.02 μM) compared with the standard acetazolamide (IC?? = 0.984 ± 0.12 μM). Enzyme kinetics revealed that 5h acts as a mixed-type inhibitor, with inhibition constants Ki = 2.91 μM and Ki′ = 3.88 μM, indicating a preference for competitive binding. Compound 5h also demonstrated significant free radical scavenging activity (30.98%) and anticancer potential against MCF-7 cells, inhibiting 40% of cell growth at a concentration of 125 μM. DNA binding studies (UV-Vis and cyclic voltammetry) revealed a weak binding affinity (Kb = 30–32 M?¹), suggesting that its anticancer activity primarily arises from carbonic anhydrase inhibition rather than DNA interaction. The authors concluded that derivative 5h could serve as a lead structure for the development of more potent carbonic anhydrase inhibitors [1]. Eissa IH, Elkady H et al., synthesized a series of novel thiadiazole–sulfonamide derivatives designed as dual inhibitors of epidermal growth factor receptor (EGFR) and carbonic anhydrase IX (CA-IX), targeting selective anticancer activity. The compounds were evaluated for cytotoxicity against MDA-MB-231 and MCF-7 breast cancer cell lines, with Vero cells serving as normal controls to assess selectivity. Enzymatic inhibition assays confirmed significant suppression of both EGFR and CA-IX, using erlotinib and acetazolamide as standards. Mechanistic studies demonstrated induction of apoptosis through modulation of BAX/Bcl-2 expression, caspase-8 and caspase-9 activation, supported by flow cytometry and cell cycle arrest analyses. Complementary molecular docking and 200 ns molecular dynamics (MD) simulations validated stable binding interactions with EGFR and CA-IX active sites. Additionally, DFT calculations and in silico ADMET profiling predicted favorable pharmacokinetics, drug-likeness, and safety profiles. These findings suggest that thiadiazole–sulfonamides hold strong promise as dual-targeted anticancer agents with optimized efficacy and reduced toxicity [25]. Karaku? S, Ba?ç? E et al., reported the synthesis and evaluation of a new series of thiosemicarbazides (3a–i) and their corresponding 1,3,4-thiadiazoles (4a–d) as potential anticancer agents and carbonic anhydrase (CA) inhibitors. Among them, compound 4d showed the strongest antiproliferative activity, with IC?? = 0.58 ± 0.02 μM against U87 glioblastoma cells, outperforming acetazolamide (IC?? = 0.984 ± 0.12 μM). Compound 3b also exhibited strong cytotoxicity against both U87 and HeLa cells. Mechanistic studies revealed that 3a, 3b, 4a, and 4d induced apoptosis via caspase-3 activation, Bax/Bcl-2 modulation, and ROS production. Additionally, selected derivatives inhibited carbonic anhydrase IX (CA IX) activity, supporting their dual mechanism of anticancer action. Molecular docking confirmed favorable binding within the CA IX active site. The authors concluded that compound 4d is a promising lead scaffold fordual anticancer and CA-inhibitory drug development.