Leukemia, a malignancy affecting hematopoietic stem cells, is linked to exposure to benzene, ionizing radiation, and genetic mutations. This disease presents with severe infections, anemia, and bleeding, resulting from the abnormal expansion of hematopoietic stem cells within the bone marrow, disrupting normal hematopoiesis (25). HHT acts as an anti-leukemia drug for leukemia treatment by mechanisms that are multifarious. Previous literature has reported that HHT can influence the expression levels of transcription factors, such as nuclear factor kappa B (NF-κB) (26), Forkhead box protein M1 (FOXM1) (27), specificity protein 1 (SP1) (28), the B-cell lymphoma-2 (Bcl-2)/Bcl-2-associated X protein (Bax) complex (29), the caspase family (30), telomerase (31), CDK2 (32), myosin-9 (33), and p53 (14). Furthermore, it can inhibit protein synthesis, block the cell cycle process, and disturb the activity of cell signaling pathways, resulting in many impacts on the proliferation and apoptosis of leukemia cells.

HHT functions as a translation inhibitor, acting by interacting with the ribosomal A site to inhibit the initiation and initial elongation steps of translation. This process induces cancer cell death (34–36). NF-κB-repressing factor (NKRF) is a transcriptional repressor of NF-κB, interacting with NF-κB to suppress its activity (37). The binding of HHT to NKRF disrupts the translocation of p65 and its binding to the MYC promoter, resulting in the downregulation of MYC transcriptional expression, subsequently suppressing the growth of t (7, 20) acute myeloid leukemia (AML) cells (26). In AML and chronic myeloma leukemia (CML) cell lines, HHT upregulates the expression of the cytoskeletal protein myosin-9 in a time-dependent manner (33). This upregulation arrests cells in the S and G2/M phases and increases the sensitivity of leukemia cells to HHT’s cytotoxic effects, ultimately inhibiting leukemic cell growth. Moreover, HHT enhances the binding affinity between CDK2 and tripartite motif 21 (Trim21) by specifically targeting the interaction between CDK2 and Cyclin A. This leads to the autophagic degradation of CDK2, thereby modulating the cell cycle of leukemia cells. Consequently, HHT significantly inhibits leukemia progression in both leukemia mouse models and human primary leukemia cells (32). FOXM1, an oncogenic transcription factor and member of the Forkhead family, is also a known target of HHT. FOXM1 is involved in cell cycle regulation, proliferation, invasion, vascular invasion, angiogenesis, oxidative stress, and inflammation (38). Targeting FOXM1 with HHT sensitizes K562 leukemia cells to the drug (27). Another study reported that HHT’s antitumor function is associated with the competitive action in which HHT can bind to SP1, which is one of the key transcriptional activators, in the TET1 promoter. SP1 is an essential transcriptional activator (39, 40). TET1 is a 5-methylcytosine hydroxylase which is critical for demethylation (41). HHT functions in the SP1/TET1/5hmC/FLT3 axis by inhibiting SP1-mediated TET1 transcription, which regulates the DNA epigenome of AML and modulates the growth of FLT3-mutant AML cells (28). Moreover, telomerase activity is essential for telomerase immortalize tumor cells and transform them to malignant status (42, 43). Around 80% of acute leukemia cells have increased telomerase activity (44, 45). Another research study showed that HHT induced apoptosis in human leukemic HL-60 cells by inhibiting their telomerase, indicating that telomerase may be a possible therapeutic target of leukemia treatment (31). By triggering the mTOR signaling pathway, HHT also suppresses the expression of the anti-apoptotic B-cell lymphoma 6 protein (BCL-6), which causes leukemic K562 cells to undergo apoptosis (46). HHT exerts a dual effect on K562 cells: it induces apoptosis through the activation of Caspase-3 and simultaneously triggers autophagy under continuous exposure to HHT. When combined with autophagy inhibitors such as 3-methyladenine (3-MA), the cytotoxic effects of HHT are significantly enhanced, and the inhibition of autophagy further potentiates HHT-induced apoptosis (47). The above results show that HHT is able to exert anti-leukemic effects by regulating the DNA epigenome, blocking the cell cycle, and inducing apoptosis.

In a few studies, HHT has been shown to have some potential effects in multiple myeloma (MM) and lymphoma treatment. Studies have demonstrated that HHT markedly enhances the anti-myeloma efficacy of BTZ in both in vitro multiple myeloma (MM) cell models and in vivo mouse xenotransplantation models through inhibition of the NF-κB signaling pathway (48). Furthermore, HHT exerts anti-tumor effects by inducing mitochondrial autophagy and mitochondrial dysfunction, with Parkin-dependent autophagy playing a crucial role in this process (49).

Approximately 40% of all malignant lymphoid neoplasms consist of diffuse large B-cell lymphoma (DLBCL), the most prevalent form of malignant lymphoma (50). A recent study has reported that HHT can induce apoptosis in DLBCL cells, which is done by decreasing the expression levels of the anti-apoptotic protein myeloid cell leukemia-1 (Mcl-1) and B-cell lymphoma-2 (Bcl-2) (51).

When HHT was co-administered with bortezomib (BTZ), it effectively reduced the expression of the anti-apoptotic protein Mcl-1, increased the levels of the pro-apoptotic protein NADPH oxidase activator (Noxa), and activated the pro-apoptotic protein Bcl-2 homologous antagonist/killer (Bak) (19). Either HHT, or HHT in combination with curcumin, was able to inhibit the growth and angiogenesis of lymphoma cells by targeting the Vascular Endothelial Growth Factor/Protein Kinase B (VEGF/AKT) signaling pathway (29).

Figure 3 shows the mechanisms of action of HHT in hematological tumors.

In the treatment of colorectal cancer, HHT exerts its therapeutic effects by regulating proliferation, apoptosis, and cell cycle-related signaling pathways. Ephrin type-B receptor 4 (EphB4) belongs to the Ephrin-B receptor family of receptor tyrosine kinases, which are involved in cell adhesion, migration, and angiogenesis, and EphB4 overexpression is closely associated with tumor invasion, metastasis, and prognosis (52, 53). HHT targets EphB4, inhibiting the activation of the Mitogen-Activated Protein Kinase/Extracellular Signal-Regulated Kinase 1/2 (MAPK/ERK1/2) and Phosphoinositide 3-Kinase (PI3K)/AKT pathways, while regulating the expression of cell cycle-related proteins (such as cyclin A2 and CDC2) and apoptosis-related proteins (including Bcl-2/Bax, Mcl-1, Bad, and caspases-3, 7, and 9), thereby effectively suppressing the progression of colorectal cancer cells (LoVo) (20). The PI3K/AKT signaling pathway is intricately involved in tumorigenesis, cellular proliferation, metastasis, apoptosis, epithelial-mesenchymal transition (EMT), metabolism, and chemoresistance (54–57). HHT modulates colorectal cancer cell proliferation, apoptosis, and xenograft growth by targeting the PI3K/AKT/mTOR signaling pathway (21, 58). In addition, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) promotes apoptosis by binding to TRAIL receptors on the surface of tumor cells (59, 60). HHT, as a sensitizer of TRAIL, has demonstrated significant potential in anticancer therapy. HHT markedly enhances the anticancer efficacy of TRAIL by downregulating the expression of anti-apoptotic proteins such as Mcl-1 and cFLIP, activating pro-apoptotic signaling pathways including JNK and p38, and synergizing with TRAIL to induce necroptotic cell death via the Receptor-Interacting Protein Kinase 1/Receptor-Interacting Protein Kinase 3/Mixed-Lineage Kinase Domain-Like Protein (RIPK1/RIPK3/MLKL) signaling pathway (61–63). In summary, HHT not only inhibits tumor cell proliferation, migration, and invasion, but also overcomes drug resistance in cancer cells, thereby promoting tumor cell death when combined with TRAIL.

Yang et al. (64) demonstrated the anti-fibrotic and anti-tumor effects of HHT through investigations employing a subcutaneous xenograft tumor model and a carbon tetrachloride (CCl4)-induced liver fibrosis model. The Hippo pathway regulates cell proliferation and organ size balance, crucial for maintaining normal tissue morphology and development (65). Activation of the Hippo pathway by HHT inhibits the transition from the G1 phase to the S phase of the cell cycle, promoting cell apoptosis and impacting cellular proliferation, migration, and invasion (66). EMT induced by β-catenin is a prerequisite for cell migration (67, 68). The expressions of matrix metalloproteinases (MMPs), EphB4 and β-catenin were downregulated in tumor tissues from HepG2 cells and Xenograft model tissues treated with HHT (69, 70). Furthermore, HHT suppressed the PI3K/AKT/Glycogen Synthase Kinase 3β (GSK3β) signaling pathway and decreased snail family transcriptional repressor 2 (Slug) expression, ultimately suppressing EMT in hepatocellular carcinoma cells (71). Ataxia-telangiectasia mutated (ATM) is essential for the repair of double-stranded DNA breaks (72). In PLC5 hepatocellular carcinoma cells treated with HHT, DNA damage is induced. Upon sensing this damage, ATM is activated, which subsequently leads to p53 activation. p53 activates p21, which blocks the cyclin A/cyclin-dependent kinase 2 (CDK2) complex, collectively driving cellular apoptosis, arresting cell cycle progression, and inhibiting cellular proliferation (73). Additionally, studies have shown that HHT induces a mitochondria-mediated intrinsic apoptotic pathway through the upregulation of transforming growth factor-β (TGF-β) expression, TNF, Fas cell surface death receptor (FAS), p38 mitogen-activated protein kinase (p38MAPK) and p53 in a human hepatoma cell line QGY-7703 (11).

Research results reveal that the anti-cancer efficacy of HHT is contingent upon its capability to stop the proliferation and viability of tumor cells through inhibiting protein synthesis, decreasing oncogenic expression of proteins in cancer cells, inducing apoptosis, and interfering with signal pathways. Nuclear factor erythroid 2-related factor 2 (NRF2) is essential for modulating the antioxidant and cellular defense responses through its role as a transcription factor (74). Inhibition of NRF2 transcription in HHT-treated A549 lung cancer cells was associated with increased sensitivity to the anticancer drug etoposide. This suggests that HHT may potentiate the efficacy of etoposide against A549 cells by suppressing NRF2 expression and disrupting NRF2-mediated antioxidant and cellular defense mechanisms (75). Transmembrane protein 16A (TMEM16A) is a membrane protein that functions as an ion channel and plays a role in regulating various physiological processes, and its high expression is linked to the proliferation, invasion, and metastasis of tumor cells (76). TMEM16A has been reported to promote tumorigenesis and cancer progression by activating the MAPK signaling pathway (77). HHT downregulated TMEM16A expression in a dose-dependent manner in lung cancer LA795 cell lines, which in turn reduced the phosphorylation of MEK1/2 and ERK1/2 in the MAPK pathway, affecting tumor cell proliferation, invasion, and cell cycle (78). The Kirsten rat sarcoma viral oncogene homolog (KRAS) is subject to frequent mutations in various types of cancer, especially in non-small cell lung cancer (NSCLC), where KRAS mutations dysregulate downstream cell signaling pathways such as PI3K/AKT and MAPK/ERK, resulting in abnormal activation of cellular proliferation and survival mechanisms (79, 80). It was shown that, in murine lung tumor models with Kras mutations G12D and G12C, HHT showed remarkable therapeutic efficiency and immunomodulatory effects. After treatment with HHT, interleukin-12 production in splenocytes was significantly decreased compared to the control group. HHT treatment also induced a notable increase in CD80, CD86 and CD69 expression on B220+ B cells. These data suggest that HHT acts by modulating immune cell biology and anti-tumor activity in KRAS-mutated lung tumors (81). Interleukin-6 (IL-6) is an oncogenic mediator of utmost importance that mediates immune and inflammatory responses, activates Signal Transducer and Activator of Transcription 3 (STAT3) in tumor cells, and its effects on oncogenesis, tumor immunosuppression, tumor angiogenesis, and metastasis are well established (82, 83). HHT treatment can affect IL-6 signaling by targeting Janus kinase 1 (JAK1) and STAT3, inhibit STAT3 nuclear translocation and transcriptional activity by antagonizing IL-6-induced STAT3 phosphorylation, and further regulated the anti-apoptotic Mcl-1 (84). These results demonstrate that HHT can act as an anti-tumor agent by interfering with the IL-6/JAK1/STAT3 signaling pathway.

Results from a recent study revealed an increased expression of four anti-apoptotic proteins (Mcl-1, Bcl-2, survivin, XIAP) was observed typically in TNBC cells treated with HHT. There’s a link between the increase in apoptosis in HHT-treated breast cancer cells and the downregulation of the expression of these anti-apoptotic proteins (85). After HHT treatment for 24 hours, chromosomes were shattered, nuclei fragmented, and apoptotic vesicles were generated in MDA-MB-453 cells, indicating that HHT markedly inhibits the proliferation of MDA-MB-453 human breast cancer cells and induces apoptosis in these cells (12). There is increasing evidence showing that microRNAs (miRNAs), a type of small non-coding RNAs, play a crucial role in regulating tumor proliferation and metastasis (86). HHT induces the downregulation of miR-18a-3p and the subsequent inhibition of the AKT/mechanistic Target of Rapamycin (mTOR) pathway, further regulates the expression of Bax/Bcl-2, caspase-3/caspase-9, and poly (ADP-ribose) polymerase (PARP) (87). These downstream molecules are all directly related to apoptosis in breast cancer cells. Plett et al. (88) demonstrated that co-treatment with HHT and paclitaxel is an effective treatment for TNBC cells by enhancing anticancer activity and reducing drug toxicity. Recent studies indicate that HHT dose-dependently reduces the viability of MDA-MB-231 cells, decreases the proportion of CD44⁺/CD24^- cells, inhibits tumor sphere formation, and suppresses the expression of Octamer-binding transcription factor 4 (Oct4), CD44, SRY-box transcription factor 2 (Sox2), and Nanog Homeobox (Nanog). Additionally, HHT effectively reduces the stemness, migration, and invasion of triple-negative breast cancer (TNBC) cells by inhibiting the activation of the Hedgehog (HH)/Gli1 Family Zinc Finger 1 (Gli1) signaling pathway (89).

Integrins belong to a subfamily of αβ heterodimeric receptors, which embed the cell membrane and regulate tumor proliferation, progression, and metastasis by interacting with cell-cell and cell-matrix complexes (90, 91). Focal adhesion kinase/Src proto-oncogene (FAK/Src) is a hub of integrin-regulated cellular functions that activates the MAPK/ERK and PI3K/AKT signaling pathways (92). HHT significantly inhibits the proliferation of bladder cancer cells and downregulates the expression levels of FAK, Src, ERK, MEK, PI3K, and AKT via the suppression of integrin α5/β1 activity. This suggests that HHT can downregulate the MAPK/ERK and PI3K/AKT signaling pathways and inhibit the progression of tumor metastasis through inhibiting the integrin α5/β1-FAK/Src axis (93). Furthermore, HHT significantly inhibits glycoprotein synthesis in T-24 bladder cancer cells and promotes the accumulation of dolichol-linked oligosaccharides, thereby further suppressing glycosylation (10).

Research indicates that HHT prevents A375 melanoma cells from entering the G2/M phase by regulating the expression of cell cycle-related proteins such as cyclin B1 and CDK1. At the same time, HHT activates the mitochondrial apoptotic signaling pathway, modulating the expression of apoptosis-related proteins, including Bcl-2/Bax, cleaved-caspase 3 and cleaved-PARP, thereby effectively inducing apoptosis (94). In another study, HHT may inhibit the activation of the PI3K/AKT pathway by inhibiting the expression of IRS4 and then downregulating the expression level of cyclin E1/CDK2, thereby blocking the cell cycle in the G0/G1 phase and significantly suppressing the proliferation of A375R cells (95). DNA damage is essential for cell cycle regulation and apoptosis. DNA damage activates the ATM/Checkpoint Kinase 2 (Chk2) signaling pathway, resulting in G2/M phase cell cycle arrest (96). Aurora kinase A (Aurka) and cell division cycle 25c (Cdc25c) are the key regulators of cell division, and Polo-like kinase 1 (Plk1) activates Cdc25c, which mediates G2/M cell cycle progression and promotes mitosis (97). Studies have shown that HHT can cause DNA damage, activate the ATM/Chk2/P53 signaling axis, inhibit the Aurka/Plk1/Cdc25c signaling pathway, and induce G2/M phase cell cycle arrest (98).

A rhabdoid tumor (RT) is a type of childhood cancer characterized by a poor prognosis (99). Primary RTs can occur in the brain, kidney, or various soft tissues (100). RT cell lines have been found to be highly sensitive to HHT, and this sensitivity may be associated with low expression of B-cell lymphoma-extra large (BCL-XL), a significant BCL-2 family member involved in apoptotic regulation. Therefore, the low expression of BCL-XL in RT cell lines may make them more sensitive to treatment with HHT (101).

Glioblastoma (GBM) is categorized as the primary brain tumor with the highest degree of aggressiveness in adults (102). Platelet-derived growth factor receptor alpha (PDGFRα) is crucial for embryogenesis and organogenesis (103). Activation of PDGFRα initiates multiple signaling pathways, including JAK-STAT (104–106). STAT3 is considered to be one of the hallmarks of GBM aggressiveness and contributes to tumor development and progression in several ways, such as inducing cell proliferation, inhibiting the apoptotic process, and promoting tumor cell migration and invasion (107). HHT treatment of GBM can inactivate STAT3 signaling by inhibiting PDGFRα phosphorylation and PDGFRα/Ras homolog family member A (RhoA)/Rho-associated coiled-coil-containing protein kinase (ROCK) axis to reduce glioblastoma cell proliferation, cytoskeletal remodeling, and migration (108).

Figure 4 illustrates the molecular mechanisms of Homoharringtonine (HHT) in malignant tumors.

Inflammation is a well-recognized hallmark of cancer, and a considerable body of evidence suggests that dysregulation of inflammatory pathways plays a key role in carcinogenesis (109). Inflammatory processes induce elevated levels of pro-inflammatory molecules, such as cytokines (IL-6, IL-2), transcription factors (NF-κB, STAT3), and protein kinase B (AKT), which contribute to cancer initiation and progression (110–112). Studies have shown that HHT exerts its immunomodulatory effects by interacting with a variety of immune mediators, influencing gene transcription, and regulating cellular functions. Key molecules associated with HHT’s immunomodulation will be discussed below. Research has said that HHT affects the activity of certain major transcription factors such as STAT3, NF-κB, and NRF2 which are important in regulating the signaling pathways of pro-inflammatory cytokines and growth factors (113, 114). Moreover, inhibition of STAT3 phosphorylation and nuclear translocation has been demonstrated in HHT-treated cells, which blocks cell growth and metastasis mediated by STAT3 (84, 115). HHT reduces the expression of anti-apoptotic genes regulated by STAT3, such as Bcl-2 and Mcl-1, thereby affecting cell growth and survival in cancer cells (116). NF-κB, an inflammatory transcription factor, is essential for the regulation of a range of genes and biological processes containing immunity, inflammation, cell proliferation and survival (117–121). Previous studies revealed that HHT can inhibit NF-κB activation, which could prevent the inflammatory responses mediated by NF-κB and tumor generation (122). In addition to interfering with transcription factors, HHT also has been shown to have an effect on the production of various cytokines and growth factors. Pro-inflammatory cytokines, such as tumor necrosis factor alpha (TNF-α) and interleukins, have a role in inflammatory processes and growth of neoplastic diseases. HHT has been shown to have an effect on the expression of IL-1, IL-6, and TNF-α. The effect of HHT on downregulating these cytokines affects cell proliferation in general (22).

HHT has been shown to inhibit viral replication in several models of viral infection, impeding protein synthesis and reducing virus production and spread. HHT has also been reported to inhibit coronaviruses, such as SARS-CoV-2, porcine epidemic diarrhea virus (PEDV) (123), and mouse hepatitis virus (MHV) (124), herpesviruses, including varicella-zoster virus (VZV) (125) and herpes simplex virus type 1 (HSV-1) (123), and Rhabdoviruses, such as vesicular stomatitis virus (VSV), pseudorabies virus (PRV), Newcastle disease virus (NDV) (123), and many others.

SARS-CoV-2 is the coronavirus that causes COVID-19. Inhibiting SARS-CoV-2 replication is important for controlling viral infection and disease progression. Studies have demonstrated that HHT can significantly reduce the level of SARS-CoV-2 viral replication (126–130). PEDV-N protein is the nucleocapsid protein of PEDV, which mediates the S-phase host cell block and is crucial for PEDV infection and replication (131). The level of PEDV-N protein in PEDV was significantly reduced after treatment with HHT, effectively reducing the viral load of PEDV-infected cells and animals (123). HHT also successfully inhibits MHV replication in mice by inhibiting viral translocation (124). VZV, a member of the herpesviridae family, is the etiological agent responsible for both varicella (chickenpox) and herpes zoster (shingles) (17). HHT significantly inhibits the mRNA levels of VZV lytic genes, suggesting that HHT may affect the replication and transmission of VZV by inhibiting the expression and replication of VZV lytic genes, in addition to blocking the ribosomal function (125). Eukaryotic translation initiation factor 4E (eIF4E) is a key factor in the initiation of protein synthesis. Phosphorylation of eIF4E regulates the assembly of the eIF4F complex, thereby promoting the translation and replication of certain viruses and affecting the selective specificity of cellular translation (132–135). Dong et al. (123) showed that HHT can antagonize the phosphorylation level of eIF4E in Vero and HeLa cells. Moreover, embryos, chickens, and piglets treated with HHT demonstrated reduced susceptibility to NDV and PEDV infections, indicating that HHT may influence viral replication by inhibiting the phosphorylation of eIF4E. In addition, HHT has been shown to be effective against Hepatitis B, bovine viral diarrhea (136), Ebola (137), and Chikungunya (138) viruses.

Fibroblast proliferation is considered critical in the formation of fibrotic scar adhesions, while inducing fibroblast apoptosis proves effective in preventing such adhesions (139). The PI3K/AKT/mTOR signaling pathway is pivotal in controlling fibroblast proliferation and apoptosis in mammalian cells (140). Sun et al. (141) have shown that the phosphorylation levels of PI3K, AKT and mTOR in fibroblasts were significantly decreased after HHT treatment for 24 hours. This illustrates that inhibition of the PI3K/AKT/mTOR pathway may be one of mechanisms by which HHT induces apoptosis in fibroblasts and mitigates intra-articular fibrosis after surgery. HHT is also known to inhibit intraocular fibroplasia and vitreous proliferation (142, 143) and may prevent the development of retinal detachment in vivo ( 144). Similarly, there is evidence that HHT inhibits the proliferation of Human Retro-ocular Fibroblasts (HROF), and the rate of cytostatic inhibition is significantly correlated with HHT concentration (145). In addition, clinical studies have shown that HHT significantly inhibits corneal haze in photorefractive keratectomy (PRK) (146, 147) and is an effective and relatively safe adjunct to glaucoma filtration surgery (148).

Neuroinflammation refers to the inflammatory response occurring within the central nervous system that results from a variety of pathologic insults, including infection, trauma, ischemia, and toxic accumulation (149). Jiang et al. (115) demonstrated that HHT alleviates neuroinflammation in amyloid precursor protein/presenilin 1 (APP/PS1) mice by disrupting the STAT3 pathway, thereby slowing the progression of Alzheimer’s disease.

TGF-β inhibits allergic inflammation (150), and HHT exerts anti-inflammatory effects by inducing Ser423/425 phosphorylation of smad3, which activates the TGF-β signaling pathway (18). In addition, HHT inhibits allergic inflammation by regulating the NF-κB/miR-183-5p/B-cell translocation gene 1 (BTG1) axis (122).

Several drug delivery systems have been developed to address the distinctive characteristics of HHT in terms of low solubility and to improve solubilization, stabilization, and targetability to optimize pharmacological activities. A range of drug delivery systems, including nanoparticles, polymeric encapsulation and liposomes, have been employed for HHT delivery. By encapsulating HHT within a delivery vehicle, HHT solubilization, stability, pharmacokinetics, and bioavailability have been improved, and targeted delivery to tumors has been realized.

Among them, a co-delivery system of homoharringtonine and doxorubicin (183),high proportion PEG of long-circulating HHT liposomes (LCL-HHT-H-PEG) (184),and magnetic Fe₃O₄ nanoparticles (HHT-MNP-Fe₃O₄) (185) all promoted apoptosis, inhibited cell proliferation, and exhibited significant cytotoxicity. Long-circulating PEGylated liposomes loaded with HHT (LC-Lipo-HHT) have good biocompatibility and a high safety profile, reducing HHT irritation in the vasculature (186).Compared to DNR/HHT co-delivery liposomes without folic acid modification (DH-LP), folic acid-modified DNR and HHT concomitantly transmitted liposomes (FA-DH-LP) have stronger cell toxicity and a better capacity to target tumors (187). In addition, HHT-loaded PLGA-SS-PEG nanodrugs demonstrate enhanced therapeutic efficacy and reduced toxicity (24). The application of these nanoparticle drugs significantly improves the intracellular uptake efficiency and efficacy of therapeutic agents while reducing side effects.

A variety of combination drug delivery regimens and delivery systems are used to treat hematologic diseases, which may improve drug absorption, cytotoxicity, and safety compared to drug administration alone or in combination with drugs. The combination of HHT and other drugs, such as imatinib, bortezomib, and apatinib, can lead to a synergistic effect, which would enhance their anticancer effects against leukemia. These regimens with HHT have multi-target and multi-pathways acting on leukemia cells, which can increase the therapeutic effect of the drugs and reduce the occurrence of drug resistance.

HHT and bortezomib (BTZ) are two commonly used anticancer drugs that show potential for synergistic effects in the management of leukemia and other malignancies. In the cells cultured in vitro, the synergistic use of HHT and BTZ enhanced the cytotoxicity of BTZ against K562 leukemia cells, reduced the expression of the anti-apoptotic proteins Bcl-2 and Mcl-1, while enhancing the expression of the pro-apoptotic protein Bax (15). In DLBCL and mantle cell lymphoma (MCL) cells, HHT and BTZ synergistically enhanced the expression levels of pro-apoptotic proteins Noxa and Bak, while concurrently downregulating the expression of the anti-apoptotic protein Mcl-1 (19). The above results suggest that HHT in combination with BTZ can affect cell growth by regulating apoptosis-related proteins. Additionally, Zhang et al. (14) reported that the combination of HHT and BTZ influenced the proliferation of SKM-1 cells, a myelodysplastic syndrome (MDS) cell line, by interfering with the AKT and NF-κB signaling pathways, upregulating the expression of their downstream oncogene p53, and decreasing the expression of miR-3151. In addition, the combination of HHT and decitabine (DAC) induced apoptosis in the MDS cell line SKM-1 by upregulating the expression of pro-apoptotic proteins caspase-3 and caspase-9, while downregulating the expression of the anti-apoptotic protein BCL-XL (30). Co-treatment with HHT and imatinib (IM) significantly suppressed proliferation and induced apoptosis in CML K562 cells. A prior study showed that HHT and IM synergistically inhibited the expression level of Zinc finger X-linked protein and interfered with the PI3K/AKT pathway (188), Bcl-6 expression (189), and p210 protein expression and its kinase activity (190). Co-treatment with HHT and IM also augmented the sensitivity of K562 cells to IM and the anti-tumor activity of imatinib by blocking the EphB4/RhoA pathway (191). Furthermore, BCR-ABL is a critical oncogene that mediates the malignant proliferation observed in hematopoietic stem and progenitor cells in CML (192). HHT induces autophagy and promotes ubiquitination of BCR-ABL in CML cells. The ubiquitinated BCR-ABL binds to p62, facilitating the degradation of the BCR-ABL protein and inducing apoptosis in imatinib-resistant CML K562G cells (193). FLT3-ITD leads to constitutive activation and autophosphorylation of FLT3, triggers the activation of various intracellular signaling pathways, promotes independent cell proliferation, and is crucial in the development and progression of AML (194, 195). Specific inhibition of FLT3 kinase activity represents a crucial strategy in the treatment of AML. HHT can be administered in conjunction with other therapeutic agents to treat leukemia associated with FLT3-ITD mutations by modulating cell signaling pathways, inducing apoptosis, and affecting stem cell properties. HHT in combination with quizartinib has been documented to exhibit anti-leukemic properties through the modulation of the FLT3-AKT-c-Myc signaling pathway and the reduction of side population and aldehyde dehydrogenase-positive cells possessing leukemic stem cell properties (196). HHT in combination with abivertinib affects leukemia cell growth by targeting the phosphorylation of the BTK and PI3K pathways and downregulating the expression of p-FLT3 and p-STAT5 (197). HHT in combination with heat shock protein 90 (HSP90) inhibitors synergistically reduces FLT3 expression and inhibits downstream signaling pathways, including STAT5, AKT, ERK, and 4E-BP1, demonstrating efficacy in treating FLT3-ITD-positive acute myeloid leukemia (AML) (198). The combination treatment of HHT and apatinib can exert its anti-leukemic effects by inhibiting the VEGFR-2 signaling pathway (199). HHT and gilteritinib can upregulate Ubiquitin Conjugating Enzyme E2 L6 (UBE2L6) together, promoting the degradation of Mcl-1 through the ubiquitin-proteasome pathway, and bring an effective drug target for FLT3-ITD mutated leukemia (200). Bcl-2 is a potent anti-apoptotic protein pivotal in mediating resistance to chemotherapy (201, 202). HHT combined with a Bcl-2 inhibitor antagonizes the FLT3-STAT5 pathway by downregulating the expression of Bcl-2 and Mcl-1 (203) and also interferes with the PI3K/AKT/GSK3β pathway to effectively treat leukemia (204). AML1-ETO is one of the hallmark features of t (7, 20) AML. Given its pivotal role in the pathogenesis of AML, especially in the M2 subtype, inhibiting AML1-ETO has become an important therapeutic strategy for treating AML (205). Further studies have demonstrated that the combination of HHT with aclarubicin and cytarabine significantly induces apoptosis in t (7, 20) type AML cells. This effect is mediated through caspase-3-dependent cleavage of AML1-ETO, thereby exerting an antitumor effect (206). Evidence suggests that HHT also has anti-leukemic effects when combined with oridonin (207), triptolide (208), arsenic trioxide (209–211), matrine (212), and AG490 (a JAK2 inhibitor) (213). In addition, HHT combined with curcumin significantly suppresses the proliferation, migration, and angiogenesis of lymphoma cells by inhibiting the phosphorylation of VEGFR2 and AKT, which reduces the signaling of angiogenin 1 (ANG-1), matrix metalloproteinase 2 (MMP2), and matrix metalloproteinase 9 (MMP9) (29). The mechanisms by which HHT synergizes with other drugs are illustrated in Figure 5.

With the in-depth investigation into the anti-cancer mechanisms of HHT, researchers have refined its molecular structure to enhance antitumor efficacy and mitigate toxicity, particularly in the treatment of CML and other myeloid malignancies (7, 13, 214). Clinical studies have shown that semi-synthetic homoharringtonine (sHHT) significantly reduces BCR-ABL transcript levels, particularly in Philadelphia chromosome-positive (Ph+) CML patients with poor response to imatinib, with excellent tolerability (169). Additionally, semi-synthetic homoharringtonine (ssHHT) demonstrates favorable pharmacokinetics and low inter-patient variability in patients with advanced AML (215). Furthermore, BS-HH-002, a novel HHT derivative, has demonstrated significantly enhanced anti-tumor activity through structural optimization. Specifically, in the treatment of pancreatic cancer, it effectively inhibits cell proliferation and induces apoptosis by degrading the anti-apoptotic protein MCL-1. Additionally, BS-HH-002 exhibits superior pharmacokinetic properties and circumvents the cardiotoxicity issues associated with traditional HHT (216). With the ongoing refinement of semi-synthetic HHT and its derivatives, coupled with the advancement of clinical research, the potential of HHT in treating various tumors is anticipated to be more extensively explored. Moreover, by developing innovative drug delivery routes and systems, it is possible to further address the solubility limitations of traditional HHT, thereby enhancing its bioavailability.