Although ATRA and ATO are well-established treatment options for APL, the occurrence of various adverse effects still seriously affects the quality of life of patients. ATRA induces terminal differentiation of leukemia cells and significantly improves the prognosis of patients with APL. However, the continued use of drugs, such as ATRA, usually causes significant treatment-related toxicity, resulting in ATRA resistance, retinoic acid syndrome (RAS), hypercalcemia, and decreased plasma drug concentrations [51]. Natural drugs are usually characterized by low toxicity and multi-pathway action, and they are more biocompatible than chemical drugs. Therefore, natural products can be used as a potential complementary resource to chemotherapeutic drugs.

Small-molecule compounds with therapeutic potential for APLHonokiol

Magnolia officinalis and Magnolia obovata bark extracts have been utilized as traditional medicines in China and Japan for centuries. These extracts are commonly employed in traditional medicine to address various health conditions, offering sedative, antioxidant, anti-inflammatory, antibiotic, and spasmodic effects. Additionally, they exhibit significant anticancer potential while maintaining low toxicity towards healthy cells. Honokiol (HKL) and magnolol (MAG), as shown in Fig. 3, are natural lignans with multiple effects that can be extracted from Magnolia grandiflora. Their safety and efficacy have gained widespread recognition [52, 53].

Fig. 3

Structure of HKL and its analogue MAG

Although both HKL and MAG are active ingredients extracted from Magnoliaceae, NB4 cells are more sensitive to HKL than MAG, which significantly reduces the activity of NB4 cells. Interestingly, the pathway of HKL-induced NB4 cell death does not involve apoptotic features, such as caspase activation and nucleus fragmentation, and its apoptosis-inducing process is accompanied by increased reactive oxygen species (ROS), mitochondrial damage, and expansion of the endoplasmic reticulum by triggering the accumulation of misfolded and unfolded proteins. This induces extensive cytoplasmic vacuolization and NB4 cell apoptosis. As cancer cells can evade apoptotic cell death through a variety of adaptive mechanisms, HKL, which induces cancer cell death in a non-apoptotic manner, could be an important drug for the treatment of APL [54].

In addition to inducing apoptosis, HKL is effective when used in combination with other therapies. In the treatment of myeloid leukemia, the combination of HKL with low-concentration chemotherapeutic agents has significant synergistic cytotoxic effects, which can effectively reverse drug resistance and reduce drug toxicity [55]. Moreover, HKL has a significant synergistic effect with cytarabine for the treatment of AML, where it inhibits cell proliferation and induces apoptosis [56]. However, the effect of this regimen on APL has not been confirmed. In the treatment of APL, HKL counteracts the toxic effects of ATO on the cardiac mitochondria and exerts cardioprotective effects against ischemia/reperfusion chemistry-induced cardiotoxicity. This result was confirmed in a mouse model, in which mice pretreated with HKL demonstrated significant amelioration of ATO-induced myocardial apoptosis, cardiac dysfunction, and cardiac remodeling [57]. Therefore, the combination of ATO and HKL for the treatment of APL may achieve good safety.

Sesquiterpene lactones

Sesquiterpene lactones are a class of secondary metabolites. They are a large group of naturally occurring compounds with a wide range of notable biological properties, such as anti-inflammatory, anti-bacterial, and anti-tumor properties. Sesquiterpene lactones are mostly derived from the Compositae family, and families such as Cactaceae, Solanaceae, and Euphorbiaceae may also contain sesquiterpene lactones. Many of the active constituents of traditional medicinal plants used for various ailments, such as infections, inflammation, and headaches, contain sesquiterpene lactones [58]. Their main anti-tumor mechanisms include oxidative stress, iron death, induction of apoptosis, and cellular autophagy. Aucklandia lappa Decne. from the Asteraceae family, which has been used in the combinatorial treatment of leukemia in Xiangsha Six Gentlemen Decoction, contains the sesquiterpene lactone compound dehydrocostus lactone (DL). DL can enhance TNF-α-induced apoptosis and has anti-leukemia activity in vitro [59]. The natural products of sesquiterpene lactones not only inhibit drug-resistant tumor cells, but also present sensitizing and potentiating effects when used in combination with other drugs. However, current research on drug-resistant cells and drug combinations is still limited [58]. Sesquiterpene lactones, such as gaillardin and artesunate, also show therapeutic potential for APL. Chemical structures of the sesquiterpene lactone compounds DL, gaillardin, and artesunate are shown in Fig. 4.

Fig. 4

Chemical structures of the sesquiterpene lactone compounds gaillardin and artesunate (ART), which consist of three isoprenoid units and usually have multiple biological properties

Gaillardin

Inula sesquiterpene lactones are a kind of sesquiterpene lactones extracted from Inula species. They have many pharmacological activities such as anti-inflammation, anti-asthma, anti-tumor, neuroprotection, and anti-allergy. The Inula genus has long been used in folk medicine to treat various ailments including kidney stones, urethral infections, jaundice, bronchitis, respiratory diseases, and cancer. It is widely utilized as a traditional medicine across Asia, the Middle East, Europe, and North America. In recent years, numerous studies have increasingly demonstrated the significance of these drugs as potential candidates for treating various types of cancers due to their strong anti-tumor activity [60,61,62]. Gaillardin, a sesquiterpene lactone isolated from the chloroform extract of Inula oculus-christi L., is toxic to a variety of cancer cells. It has been demonstrated that gaillardin induces cytotoxicity through the G0/G1 phase blockade and then apoptosis in a dose-dependent manner and that it has no significant cytotoxic effect on healthy cells, making it a promising anti-hematological malignancy medicine that could open new avenues for the treatment of APL [63].

In vitro experiments further support this idea. In a previous study, gaillardin dose-dependently induced apoptosis in APL cells. Gaillardin extracts at concentrations of 1, 4, and 5 μM induced early apoptosis in 10.5%, 19%, and 32% of NB4 cells, respectively, with an IC50 of approximately 7 μM after 48 h. Treatment of NB4 cells with gaillardin resulted in the upregulation of Bax transcripts and a decrease in Bcl-2 mRNA, in turn increasing the Bax/Bcl-2 transcription ratio. At the tissue level, Bcl-2 and Bax dimers formed, which initiated the release of cytochrome C from the mitochondria and activated caspase-3, ultimately leading to cell death [64]. In vivo experiments evaluating the treatment of APL with gaillardin are expected.

Artesunate

Artemisinin analogs, with their unique peroxy-bridge structure, have been shown to have significant therapeutic effects against Plasmodium falciparum, which causes malaria, and are not susceptible to drug resistance [65]. One of the herbs in the Chinese prescription Qinggu Powder is Artemisia annua L. In leukemia, artemisinin has been shown to induce cell cycle arrest [66]. Artesunate (ART) is a semi-synthetic derivative of artemisinin, which has the advantages of oral administration and good water solubility. It is widely used clinically for the treatment of malaria and has shown anti-tumor effects against a variety of hematologic tumors. Studies have shown that, after the treatment of NB4, HL-60, and NB4-R1 (retinoic acid-resistant strains) with 2, 10, or 20 μg/mL ART for 12, 24, or 48 h, cell proliferation was significantly inhibited in an obvious time- and concentration-dependent manner and the cells showed typical apoptotic morphology changes after 24 h. The mechanism of action of ART may be phosphorylation of the JNK pathway in the form of p-JNK, p-MKK4, and p-ATE-2, as well as inhibition of PI3K/AKT/mTOR pathway phosphorylation [67]. ART shows potential for the treatment of APL and retinoic acid-resistant APL, but its therapeutic efficacy needs to be further demonstrated.

Celastrol

Celastrol is a natural pentacyclic triterpenoid purified from the Celastraceae family. Celastrol possesses a variety of properties as a TCM, including anti-inflammatory and broad-spectrum anti-cancer properties [68]. Celastrol achieves its anti-malignant properties against hematological neoplasm through several pathways. First, celastrol is a potent low-molecular-weight inhibitor that induces myeloid differentiation and cancer cell apoptosis by inhibiting Myb activity. In combination with other compounds, the inhibitory effect of celastrol on cell proliferation can be enhanced [69]. Second, several studies have proven that celastrol can induce apoptosis in APL cells through the p53-activated mitochondrial pathway [70]. As shown in Fig. 5, the mRNA expression of caspase-9, caspase-3, and Bax was elevated after celastrol treatment, while the mRNA expression of p53 was not. The protein expression of cleaved caspase-9, cleaved caspase-3, Bax, and p53 was significantly elevated. After 24 and 48 h, inhibition of HL-60 cell proliferation occurred in a dose-dependent manner, with IC50 values of 0.48 and 0.55 μM at 24 and 48 h.

Fig. 5

Celastrol induces apoptosis in APL cells through the mitochondrial pathway, causing changes in cytokines

Celastrol has a satisfactory safety profile in the treatment of APL. In the nude mouse model of APL with tumor xenografts, there was no significant difference in the coefficients of the heart, liver, spleen, lungs, kidneys, brain, testes, and epididymides between the control and celastrol-treated groups of mice. There were no statistically significant differences between the alanine transaminase (ALT), aspartate transaminase (AST), blood urea nitrogen (BUN), and creatinine concentrations (CREA) of control mice and celastrol-treated mice, which were all within normal ranges, and no obvious histopathology was seen in the testes of mice, suggesting that celastrol has no toxic effects on the liver, kidneys, or reproductive system at a dose of 2 mg/kg [70]. Additional in vitro results showed that the ability of leukemia cells from two different AML mouse models to form colonies in the semisolid medium was inhibited by sub-micromolar concentrations of celastrol, but the proliferative capacity of normal hematopoietic progenitor cells from healthy mice was not inhibited under the same conditions. Similarly, colony formation assays performed on leukemic cells from patients with AML and cells from healthy donors confirmed that leukemic cell proliferation was significantly inhibited by celastrol, whereas healthy progenitor cells were unaffected [69]. These results suggest that celastrol has a good safety profile in the treatment of leukemia and is a useful alternative combination therapy.

Tanshinone IIA

Salvia miltiorrhiza Bunge is an important component of oral arsenic agent RIF. Tanshinone IIA (Tan IIA) is a diterpene quinone isolated from Salvia miltiorrhiza Bunge, which is the most abundant and structurally representative fat-soluble constituent of Salvia miltiorrhiza, and has been found to possess anti-tumor properties, such as inducing cell autophagy and apoptosis, and inhibiting tumor invasion and metastasis [71]. Many studies have proven that Tan IIA has the biological activity to treat APL by inducing apoptosis in the APL cell line NB4 [72]. It has also shown excellent therapeutic effects when combined with ATRA and ATO.

In a previous study, Tan IIA inhibited the growth of NB4 cells and induced the differentiation of NB4 cells, and these effects were gradually enhanced with an increase in drug concentration and a prolonged duration of action. Similar to As2O3 and ATRA, Tan IIA did not change the expression of PML-RARα mRNA, but it degraded the PML-RARα fusion protein and restored the expression of the PML protein. The optimal concentration to achieve this effect was 2.55 μmol/L [73]. In addition, Tan IIA induced NB4 cell autophagy to form the autophagic stream in a concentration- and time-dependent manner. The effects of Tan IIA on NB4 cells, evaluated after 48 h., included significantly reduced expression of the autophagy-related protein p62, and increased autophagy rate of NB4 cells. PI3K-Akt, and mTOR protein bands in NB4 cells were less pronounced than those in the control group after treatment with Tan IIA, which indicated that Tan IIA reduced the expression of the PI3K-I, Akt, and mTOR proteins in NB4 cells. In short, Tan IIA reduced the Akt and mTOR phosphorylation levels, and inhibited PI3K/Akt/mTOR signaling [74]. Thirdly, when Tan IIA was combined with ATO, the apoptosis and autophagy rates of NB4 cells were higher than those of the single-drug group. This may be due to the fact that Tan IIA-ATO can upregulate the expression of the apoptosis-specific protein caspase-3 and the autophagy-specific protein LC3-II in transplanted tumor tissues, as well as enhancing tumor cell apoptosis and autophagy. Therefore, the Tan IIA-ATO combination has a chemo-sensitizing effect on NB4 transplanted tumors. In addition, experiments have shown that the Tan IIA-ATO regimen causes no obvious pathological damage to the bone marrow, heart, liver, lungs, kidneys, lymph nodes, and other important tissues and organs in nude mice, demonstrating its safety [75]. Fourth, as shown in Fig. 6, Tan IIA still has a therapeutic effect in ATRA-resistant strains. MR-2 is an ATRA-resistant APL cell type. When Tan IIA was co-cultured with NB4 and MR-2 cells, Tan IIA caused differentiation and apoptosis of both cell types, which indicated that there was no cross-resistance between ATRA and Tan IIA. Tan IIA at 1.0 mg/L inhibited the proliferation of MR-2 cells and induced their transformation into granulocytes, which is the most effective way to prevent the proliferation of ATRA [76]. Tan IIA at 1.0 mg/L inhibited the proliferation of MR-2 cells and induced their differentiation to the mature stage of the granulosa lineage, and the effect was comparable to that of 0.5 mg/L Tan IIA, which induced the differentiation of NB4 cells [77]. From the above studies, it is clear that Tan IIA may be a promising clinical treatment for APL, especially for recurrent and drug-resistant patients.

Fig. 6

Tan IIA is therapeutically effective against both NB4 and MR-2, without cross-resistance

Oleanolic acid and its derivatives

Oleanolic acid (OA), a pentacyclic triterpenoid that is ubiquitous in the plant kingdom, is the main active ingredient of Akebia quinate (Thunb.) Decne. in Longdan Xiegan Decoction, Forsythia suspensa (Thunb.) Vahl in Lianhua Decoctio and Hedyotis diffusa. OA has been endowed with an extensive variety of biological properties and therapeutic potential through its complex and multi-factorial mechanisms, and it has received much attention from the scientific community because of its biological activity in a wide range of diseases. OA and related triterpenes have a wide range of pharmacological properties, but their therapeutic potential has only been partially exploited to date [78]. The anti-cancer potential of bioactive triterpenes in vitro and in vivo models, including in the treatment of APL, has been widely discussed [79].

OA enhances the differentiation of APL cells and prevents the development of leukemia in mice [80]. Firstly, OA and its analog ursolic acid (UA) significantly inhibited the proliferation of HL-60 cells in a concentration- and time-dependent manner from 0 to 72 h after treatment. The number of non-living cells was higher for cells cultured at high OA and UA concentrations of 80–100 μM. At non-cytotoxic concentrations, OA had a more significant differentiation-inducing effect on HL-60 cells, and when combined with low-dose ATRA, OA increased the differentiation rate of HL-60 cells, whereas UA had no significant effect on the differentiation of ATRA. In a mouse model of leukemia, OA increased the survival time and decreased the infiltration of leukemia cells into the liver and kidneys.

The structures of OA and its derivatives are shown in Fig. 7. The OA derivatives DIOXOL and HIMOXOL may also be therapeutically effective against HL-60 cells, its overexpressing subline HL-60/AR, and its multidrug-resistant subline ABCC1. DIOXOL and HIMOXOL are the most potent semi-synthetic OA derivatives against human APL cells [81]. Cell cycle analyses of 5–20 μM DIOXOL and HIMOXOL treatment for 24 h showed the presence of sub-G1 cell populations, indicating DNA fragmentation of dead cells. Among them, DIOXOL was the most effective at inducing apoptosis in HL-60 cells, and higher concentrations of DIOXOL (10 μM and 20 μM) activated apoptosis to a greater degree than HIMOXOL. DIOXOL significantly reduced p65 nuclear factor kappa-B (NF-κB) and inhibited its translocation to the nucleus to activate the apoptotic program. A 70% reduction in intracellular NF-κB subunit content was observed in samples treated with 20 μM DIOXOL. HIMOXOL is the most effective compound against drug-resistant HL-60/AR cells. It can inhibit ABCC1 transporter function in a short period and reduce ABCC1 protein expression over a longer period. HIMOXOL at concentrations of 5 and 10 μM were able to act at the transcriptional level, leading to significant reductions in ABCC1 transcripts of approximately 30% and 50%. HIMOXOL was also more effective at reducing the amount of Bcl-2. Bcl-2 was reduced by 15% when HIMOXOL was used at a concentration of 10 μM, which was increased to 70% when HIMOXOL was used at a concentration of 20 μM. OA and its derivatives could be used as part of an initial screen of potential synergistic anti-leukemic agents for ATRA, providing a direction for new APL drug development.

Fig. 7

Structure of OA and its derivatives, which all have therapeutic potential for APL

Active extracts from plantsNatural seaweed extracts—fucoidan

Fucoidan is a high molecular-weight, fucose-based, sulfated polysaccharide extracted from the brown macroalgae. It is a natural component of seaweed and is found in the cell walls of a range of brown seaweeds. Fucoidan is a heterogeneous sulfated polysaccharide containing sulfated L-fucose with 34–44% fucose content, which has immunomodulatory and anti-tumor effects [82, 83].

In a previous study, fucoidan inhibited the proliferation and induced the apoptosis of the APL cell lines NB4 and HL-60 via both endogenous and exogenous pathways. The proliferation of NB4 and HL-60 cells was inhibited in a dose-dependent manner, and the cell proliferation of HL-60 and NB4 cells decreased to less than 10% at fucoidan concentrations of 50 and 25 μg/mL, respectively. After treatment of NB4 and HL-60 cells with 100 μg/mL fucoidan for 48 h, the percentage of sub-G0/G1 cells in the dead cell population increased significantly in a time-dependent manner, and fucoidan significantly increased apoptosis in both cell lines. After 10 days of inoculation of NB4 cells into seven nude mice in each of the two groups, five in the control group developed subcutaneous tumors, whereas only two in the fucoidan group developed subcutaneous tumor masses, and no other toxicity was observed in either group [84]. These findings collectively suggest that fucoidan significantly delays tumor growth.

The conventional therapy for APL is ATRA-ATO, but drug resistance or RAS may occur with long-term use of this regimen. Some findings suggest that, by adding fucoidan to the standard APL regimen, the number of resistant cells in patients who respond to ATRA can be limited [85]. Fucoidan combined with the ATRA-ATO regimen synergistically induced NB4 cell differentiation, as evidenced by increased CD11b expression and G0/G1 blockade. In vitro findings showed that a portion of cells remained undifferentiated when cells were treated with ATRA alone or ATRA-ATO, whereas almost all cells underwent differentiation when fucoidan was combined with ATRA-ATO. CD44 expression in APL cells was reduced when mouse tumor cuts were treated with fucoidan combined with ATRA, implying that the use of this regimen may decelerate the spread of cancer cells in patients with APL. The use of fucoidan as a supplement to standard APL therapy may represent a promising new strategy for APL management.

Crocin and crocetin from saffron

Saffron is derived from Crocus sativus L., which is a valuable medicinal plant in many traditional medicinal cultures. There are around 75 species of crocus in the world, and saffron is the only species available for medicinal use. It is mainly distributed in Southern Europe and Iran, and also planted in China [86]. Saffron has more than 200 active ingredients, including pigments, flavonoids, phenolic acids, and fatty acids, amongst others [87]. Gardenia jasminoides Ellis, commonly used in TCM, is one of the components of Longdan Xiegan Decoction, and also contains similar components. The Compendium of Materia Medica describes saffron as “The smell is sweet, flat, and non-toxic. Indications: Heart worries and stagnation, persistent Qi stagnation, and promoting blood circulation. Long-serving brings joy and treats palpitations.” Among the active ingredients, crocin (CRO) and crocetin (CRT) have great potential for the treatment of APL. The activity mechanism of CRO and CRT is shown in Fig. 8.

Fig. 8

The bioactive substances CRO and CRT in saffron. CRO inhibits the proliferation and tumorigenicity of HL-60 cells. CRT has anti-oxidant and anti-apoptotic properties and can significantly reduce oxidative stress in ATO-induced nephrotoxicity

CRO is the main water-soluble carotenoid in saffron extract. It has anti-tumor activity against many human tumors [88]. Studies have shown [89] that CRO at a certain concentration range (0.625–10 mg/mL) significantly inhibits the proliferation of HL-60 cells, and with an increase in the CRO concentration from 0.625 to 5 mg/mL, the percentage of apoptotic cells increases significantly, and this effect is time-dependent. In the nude mouse HL-60 cell model, the tumor formation time in the experimental group (6.25 mg/kg CRO) was significantly longer than in the other groups, and the tumor formation time in the experimental group (25 mg/kg CRO) was longer than in the control group and the experimental group (100 mg/kg CRO). Compared with the control group, the rate of change in tumor weight and tumor size was significantly suppressed in mice treated with 6.25 and 25 mg/kg CRO. Moreover, Bcl-2 protein expression was reduced and Bax protein expression was elevated in the tumor. The above findings prove that CRO inhibits the proliferation and tumorigenicity of HL-60 cells.

In addition to its therapeutic potential, CRO in combination with ATO reduces ATO-induced cardiotoxicity [90]. CRO administration not only reduces QTc interval prolongation, cardiac enzymes, and troponin T, but it also improves histopathological results. The expression of Bax and caspase-3 in the myocardium of rats treated with CRO was significantly decreased compared to when rats without CRO. CRO appears to reduce ATO-induced myocardial pathological changes, and the therapeutic effect of CRO appears to be dose-dependent. Similarly, CRT may be protective against ATO-induced renal injury [91]. CRT has anti-oxidant and anti-apoptotic properties, and therefore it can significantly reduce oxidative stress in ATO-induced nephrotoxicity. In one study, ATO-induced histopathological changes in the kidneys of rats showed glomerular destruction, tubular cell swelling, interstitial fibrosis with inflammatory cell infiltration, and nephrocyte atrophy and necrosis. Treatment with 25 or 50 mg/kg CRT significantly reduced the morphological changes in the kidney induced by ATO. From the above, it is reasonable to propose that CRO and CRT are ideal choices as combined treatments with ATO, and the usefulness of these combinations should be further investigated for clinical application.

Green tea extract

Tea is one of the most popular drinks in the world. Originating from China, tea was introduced to the world thousands of years ago via the Silk Road. The production of green tea involves decoction or steaming of freshly harvested leaves to inactivate polyphenol oxidase and other enzymes that prevent fermentation/oxidation, preserving the active chemical properties [92]. As one of the most consumed beverages worldwide, green tea has been the focus of much research, and its polyphenolic compounds have been shown to have many benefits for human health. Catechins are the main components extracted from green tea leaves and are present in about 30% of dried green tea, which includes epigallocatechin gallate (EGCG), epicatechin gallate (ECG), epigallocatechin (EGC), and epicatechin (EC), as shown in Fig. 9. Catechins are inexpensive, safe, and can be administered orally. Catechins, especially EGCG, have multifaceted effects that make them attractive candidates for the prevention and treatment of leukemia and myelodysplastic syndrome [92, 93].

Fig. 9

The four main active substances of catechins: EC, ECG, EGC, and EGCG

A previous study showed that green tea extract reduced leukocytes and immature cells (progenitor cells) in the peripheral blood, bone marrow, and spleen of leukemic mice while increasing mature cells in the bone marrow. An important observation in leukemic mice is an increase in the number of leukocytes, and treatment with 250 mg/kg green tea extract for 4 days decreased the percentage of leukocytes while decreasing the percentage of immature cells and increasing the percentage of mature cells. These results suggest that green tea extract has anti-leukemic proliferative effects in vivo by inhibiting malignant clonal expansion [94]. In addition, catechins can have anti-leukemic activity by inducing apoptosis. In another study, NB4 cells were inoculated subcutaneously in nude mice, and 10 mM catechin was used as the only drinking water of the mice for 10 days. Tumor size was significantly reduced in the treated group, and no tumor infiltration was detected in any organ at necropsy. The PML-RARα fusion protein was degraded after treatment of primary leukemia cells with 100 and 150 μM catechin for 24 h [93]. This is a strong rationale supporting the therapeutic potential of catechins for APL.

Of the four major catechins, EGCG is the most abundant and potent polyphenolic compound in green tea extract, accounting for 50–75% of total catechins [92]. Due to its long half-life, the compound is rapidly absorbed and distributed in all tissues [