In recent years, a large number of studies have been devoted to the various pathways that non-classically regulate cell death, mainly focusing on oxidative cellular damage. Ferroptosis, an iron-dependent mode of programmed cell death proposed by Stockwell in 2012, is morphologically and biochemically distinct from apoptosis, autophagy, and necrosis. It plays a key role in many diseases, including cancer, acute kidney injury, cardiovascular, neurodegenerative, and hepatic diseases. It is also reported that ferroptosis is a critical regulator of drug resistance [1]. Ferroptosis, mainly regulated by iron metabolism and lipid peroxidation signaling, is characterized by the accumulation of intracellular iron, leading to massive lipid peroxidation and ultimately cell death. Therefore, induction of ferroptosis has become a novel potential therapeutic strategy for cancer therapy.

Glutathione peroxidase 4 (GPX4), also known as phospholipid hydroperoxide glutathione peroxidase (PHGPx), has the ability to scavenge membrane lipid hydroperoxide products. It is an important member of the selenium-containing GPX family with selenocysteine in the catalytic active site. As shown in Fig. 1A, the anti-ferroptosis enzyme GPX4 is critical for protecting cells from oxidative damage and maintaining membrane lipid bilayer homeostasis by converting the toxic lipid hydroperoxides (LOOH) that arise from iron-catalyzed oxidation of polyunsaturated fatty acids into non-toxic lipid alcohols. As a key regulator of ferroptosis, GPX4 is highly expressed in a variety of tumor types, suggesting that GPX4 is a potential target for cancer treatment. Targeting GPX4 to induce ferroptosis for cancer treatment can be achieved by (1) mutation of selenocysteine to cysteine (reducing the activity of GPX4 by 90%), and (2) inhibition of GPX4 activity or degradation of GPX4. Very recently, various small molecule inhibitors have been reported to have GPX4 inhibitory activity, such examples including RSL3, ML162, ML210, JKE-1674, and BCP-T.A (Fig. 1B). However, there are currently no specific GPX4 small-molecule inhibitors or ferroptosis inducers available in the clinic. As a result, the development of novel and effective ferroptosis inducers is still needed.

Although ferroptosis is considered a distinctive form of cell death compared to apoptosis, recent studies revealed an interplay between ferroptosis and apoptosis through the endoplasmic reticulum (ER) stress signaling pathway [2,3]. In addition to strong inhibition of tumor growth, small-molecule ferroptosis inducers also enhanced the sensitivity of chemotherapeutic drugs [4]. A combination of chemotherapeutic drugs such as doxorubicin, tmozolomide, and cisplatin, with ferroptosis inducer erastin resulted in a remarkable synergistic effect on tumor treatment [5]. Besides, multiple studies have shown that simultaneous targeting of apoptosis and ferroptosis can effectively overcome or bypass chemoresistance. For instance, Liu et al. reported platinum (IV) complexes could intervene oxaliplatin resistance in colon cancer via inducing ferroptosis and apoptosis [6]. By inducing the apoptosis/ferroptosis hybrid pathway, ent-kaurane derivative showed great potential in sensitizing CDDP resistance towards A549/CDDP cells [7]. Similarly, the co-delivery of the apoptosis and ferroptosis agents has also been demonstrated to be a promising strategy to overcome or evade the resistance in chemotherapy-induced apoptotic pathways, with examples including but not limited to: Fu et al. developed a US-activatable DFHHP nanomedicine that can overcome chemoresistance and suppress tumor growth by inducing collaborative apoptosis and ferroptosis of tumor cells [8]. Chen et al. also proposed an active-targeting small-molecular self-assembly nano-prodrug for the co-delivery of chemotherapeutics (CPT), ferrocene (Fc), and GPX4 inhibitor (RSL3) that showed potential in overcoming the chemo-resistance via the combination of apoptosis and ferroptosis therapy [9]. These findings suggested a synergistic interaction between ferroptosis and apoptosis for cancer treatment and combating drug-resistance. Most of the reported GPX4 inhibitors have poor selectivity and pharmacokinetics, but they can be used as a lead compound for further development [[10], [11], [12], [13]]. Among them, ferroptosis inducer ML162 showed significant GPX4 inhibitory activity and relatively high anti-tumor potency at the cellular level. Based on ML162, many GPX4 inhibitors or degraders with more excellent properties have been developed. For example, Luo et al. reported a GPX4 degrader (dGPX4) that showed a five-fold enhancement of ferroptosis induction efficiency compared to that of ML162 [14], suggesting that ML162 is an important lead compound for further modification.

The quinone moiety is a crucial apoptosis-induced pharmacodynamic scaffold that is presented in many currently used anticancer drugs, such as the anthracycline antibiotics (daunorubicin, doxorubicin, idarubicin, and mitoxantrone), dactinomycin and mitomycin-C [15]. Inspired by the ferroptosis-inducing potential of ML162 and the active anticancer effect of quinone derivatives, herein we designed and synthesized a series of ML162-quinone conjugates by using pharmacophore hybridization and bioisosterism strategies, with the aim of obtaining more active anticancer agents via the ferroptosis and apoptosis dual cell death processes. The purpose of designing ferroptosis and apoptosis dual agents is mainly based on the fact that much evidence suggests that the combination of apoptosis and ferroptosis represents a highly appealing strategy for eradicating tumor cells and addressing apoptosis-related drug resistance [[16], [17], [18], [19], [20]]. We evaluated the antiproliferative effect of all the synthesized compounds on the ferroptosis-sensitive HT1080 fibrosarcoma cells. Subsequently, ferroptosis- and apoptosis-related phenotype evaluations were conducted on the most promising one (GIC-20). Finally, the antiproliferative and drug-resistant reversal effects were preliminarily evaluated on pancreatic cancer cells.