The biological hallmarks of cancer include uncontrolled cell proliferation and differentiation, dysregulated signaling pathways, genomic instability and metabolic reprogramming (1). These characteristics make the development of more effective cancer therapies a critical research priority. In recent years, the emergence of immunotherapy represents a major breakthrough in cancer treatment. As the fifth treatment modality following surgery, radiotherapy, chemotherapy and targeted therapy, immunotherapy works by re-establishing the tumor-immune cycle and restoring the body's normal anti-tumor immune response, thereby achieving tumor control and eradication (2,3). Immune checkpoint blockade therapy has been widely proven to be effective against various human tumors, while cell therapy has shown notable efficacy in hematological malignancies and remains in the clinical research stage for solid tumors. In the field of immune checkpoint inhibitors, the main approaches include: i) Monoclonal antibody therapy targeting programmed cell death protein 1 (PD-1) and its ligand PD-L1; ii) monoclonal antibody therapy targeting cytotoxic T-lymphocyte-associated protein 4 (CTLA-4); and iii) monoclonal antibody therapy targeting lymphocyte-activation gene 3. As for adoptive cell therapy, it primarily encompasses: i) Tumor-infiltrating lymphocyte therapy; ii) T cell receptor-engineered T cell therapy; iii) chimeric antigen receptor T cell (CAR-T) therapy; and iv) natural killer (NK) cell therapy (4).

PD-1/PD-L1 monoclonal antibody therapy has emerged as the most widely used immune checkpoint inhibitor in clinical practice, demonstrating notable efficacy across various malignancies (5). PD-1, also known as CD279, is a pivotal immune checkpoint molecule that plays a central role in maintaining immune homeostasis and regulating tumor immune evasion. PD-1 is predominantly expressed on the surface of B cells, T cells and NK cells, where it can specifically recognize and bind to two ligands expressed on tumor cells: i) PD-L1 (CD274); and ii) PD-L2 (CD273) (6). Unlike CTLA-4 which primarily regulates immune responses during the early stage of T cell activation, PD-1 predominantly suppresses effector T cell function in peripheral tissues and the tumor microenvironment during the effector phase (7). In the tumor microenvironment, the binding of PD-L1 expressed at high level on tumor cells to PD-1 on T cell surfaces induces conformational changes in PD-1. This leads to the exposure and phosphorylation of the immunoreceptor tyrosine-based inhibitory motif (ITIM) at Y223 and the immunoreceptor tyrosine-based switch motif (ITSM) at Y248. The phosphorylated ITSM preferentially recruits the SHP-2 protein tyrosine phosphatase, while the phosphorylated ITIM forms dimers with the SHP-1 protein tyrosine phosphatase. These molecular events collectively attenuate T cell activation signals, suppress T cell cytotoxic function and consequently mediate negative regulation of immune responses to maintain immune homeostasis (8–11). PD-L1 can also interact with the costimulatory molecule CD80, transmitting inhibitory signals to activated T cells (12). Currently, several PD-1/PD-L1 monoclonal antibodies such as nivolumab (13), pembrolizumab (14) and avelumab (15) have been widely used in clinical cancer treatment. Although these immune checkpoint inhibitors demonstrate notable efficacy against various solid tumors and hematological malignancies, acquired resistance remains a major clinical challenge (16).

Research has demonstrated that dysregulation of the ubiquitin-dependent protein degradation pathway represents a crucial molecular mechanism in cancer pathogenesis (17,18). The UPS is an essential protein degradation mechanism in cells. This system primarily works by ubiquitinating damaged, abnormal, or functionally completed regulatory proteins and directing their degradation by the proteasome. (19). The UPS consists of a series of enzymes: i) Ubiquitin-activating enzyme E1 activates ubiquitin molecules; ii) ubiquitin-conjugating enzyme E2 mediates ubiquitin transfer; and iii) ubiquitin ligase E3 specifically recognizes substrate proteins and completes ubiquitin tagging (Fig. 1). These enzymes work cooperatively to ultimately achieve targeted protein degradation via the proteasome pathway (20,21). Ubiquitinated substrates are classified into different ubiquitination pathways based on the types of polyubiquitin chains, with K48 and K63 being the two most widely studied ubiquitination forms (22). Among these, K48-linked ubiquitination is recognized to direct target proteins for degradation via the proteasome pathway (23,24). K63-linked ubiquitination is primarily involved in proteasome-independent signaling pathways, typically associated with positive regulatory processes such as protein stabilization, subcellular localization and functional activation, including critical biological processes such as endocytic trafficking, DNA replication and signal transduction (25). Moreover, this modification can also facilitate substrate protein degradation through the autophagy-lysosome pathway (26).

The 26S proteasome is a multi-subunit proteolytic complex composed of a 20S core particle (CP) and a 19S regulatory particle (RP), which specifically recognizes polyubiquitin-tagged proteins and degrades them into short peptides (27). The 19S RP performs three key functions: i) Recognizing ubiquitinated substrate proteins; ii) regulating the deubiquitination process; and iii) delivering ubiquitinated proteins to the 20S CP. The 20S CP is a barrel-shaped proteolytic core containing active catalytic sites, where the final protein degradation occurs (28,29). Deubiquitinating enzymes (DUBs) can reverse protein ubiquitination, with their primary functions including: i) Maintaining cellular free ubiquitin levels; ii) releasing substrate proteins from the ubiquitin-proteasome degradation pathway; and iii) protecting target proteins from degradation (22). Research has revealed an increasingly clear connection between ubiquitination and cancer immunotherapy. Tumor cells can modulate the UPS to stabilize immune checkpoint protein expression and suppress immune-related protein function, thereby evading immune surveillance and promoting tumor progression.

In recent years, the regulatory role of the UPS in cancer immunotherapy has received growing attention. The present review summarized the key molecular mechanisms of UPS involvement in tumor immune regulation and discusses potential therapeutic strategies to enhance immunotherapeutic efficacy.

The UPS is the core mechanism for maintaining protein homeostasis within cells. During tumor immune evasion, UPS mainly regulates the dynamic balance of immune checkpoint molecules, antigen presentation processes and immunosuppressive cells through ubiquitination tagging and proteasomal degradation (64). Studies have confirmed that small-molecule drugs targeting the UPS play an important role in improving the therapeutic effects of tumor immune checkpoint inhibitors (Table I) (51,52,60). In pancreatic cancer, DUB-IN-2 inhibits the deubiquitination function of USP8, reduces the expression level of PD-L1 and thereby reverses the immunosuppressive state in the tumor microenvironment (52). In gastric cancer, the use of the small-molecule inhibitors Almac4 and P5091 targeting USP7 can inhibit the deubiquitination activity of USP7, thereby suppressing the proliferation of cancer cells. In addition, these two inhibitors can also downregulate the expression level of PD-L1 and enhance the anti-tumor immune response (51). In melanoma and colorectal cancer, the small-molecule inhibitor OTUB2-IN-1 can notably inhibit the DUB activity of OTUB2 and reduce the expression level of PD-L1 in tumor cells in a dose-dependent manner, thereby promoting antitumor immune function (60).

Recent studies have further revealed that specific small-molecule compounds can modulate the stability of PD-L1 through ubiquitination pathways. In non-small cell lung cancer, compound #25 can enhance the anti-tumor immune response by inhibiting the Skp2-mediated K63 ubiquitination process of PD-L1 (44). The small-molecule agonist AK087 of ITCH can effectively promote the ubiquitination and degradation process of PD-L1 mediated by ITCH and notably inhibit the resistance of tumors to PD-1/PD-L1 treatment (39).

In addition, some small-molecule drugs have shown great potential in the field of tumor immunotherapy. OTUB1/USP8-IN-1 is a dual inhibitor targeting OTUB1 and USP8, which can effectively inhibit the functions of these two DUBs and is a potential small-molecule drug for tumor immunotherapy (65). Natural compounds also play a marked role in the field of tumor immunotherapy. Lu et al (66) found that the natural compound gentiopicroside can inhibit the DUB activity of USP22, thereby reducing the expression level of PD-L1 in lung adenocarcinoma and enhancing the body's anti-tumor immune ability. Curcumin, as a natural dietary supplement, has been proven to have the potential for anti-tumor immunity. In TNBC, curcumin can inhibit the DUB activity of CSN5, reduce the stability of PD-L1, induce the ubiquitination and degradation of PD-L1 and thereby enhance the immune system's ability to attack tumor cells (62). In lung cancer, CSN5 can promote the deubiquitination process of PD-L1, thereby inducing tumor immune evasion. The natural compound berberine can inhibit the DUB activity of CSN5, reduce the expression level of PD-L1 and thus enhance the body's anti-tumor immune ability (67). In colorectal and lung cancers, demethylzeylasteral specifically binds to USP22 and induces its degradation, thereby promoting ubiquitin-dependent proteasomal degradation of PD-L1 and ultimately enhancing T cell-mediated antitumor immune responses (68).

Small-molecule inhibitors targeting the UPS have shown great potential in the field of tumor immunotherapy and are very likely to be one of the important strategies for cancer treatment in the future.

During the occurrence and development of tumors, tumor cells can evade the strict surveillance of the immune system through a variety of complex mechanisms. Among these mechanisms, the activation of immune checkpoint pathways is one of the core mechanisms by which tumors achieve immune evasion. At present, anti-PD-1/PD-L1 therapy, as a representative of immune checkpoint inhibitor therapy, is one of the most widely used tumor immunotherapy strategies in clinical practice (5). Although anti-PD-1/PD-L1 therapy has shown good efficacy in some patients, a considerable number of patients still do not respond to the treatment after receiving it and even develop acquired resistance due to anti-PD-1/PD-L1 therapy (69). Therefore, elucidating further the potential molecular mechanisms underlying tumor resistance to anti-PD-1/PD-L1 and exploring effective combination therapy strategies are of great significance for improving the therapeutic efficacy of anti-PD-1/PD-L1 and prolonging the survival of patients (Table II).

The UPS, as a crucial molecular mechanism responsible for protein degradation and stabilization within cells, plays a pivotal role in regulating various biological processes such as cell cycle progression, signal transduction networks and immune response reactions. It is one of the main pathways mediating protein degradation or stabilization inside cells (70,71). The UPS has the ability to precisely recognize and selectively tag damaged, abnormal, or function-completed proteins with ubiquitin ‘tags’. Subsequently, the proteasome recognizes these tagged proteins and precisely regulates the degradation process of the proteins (19). In recent years, with the continuous deepening of research, an increasing number of studies have shown that the UPS plays a crucial role in the pathological and physiological processes of tumor proliferation, invasion, metastasis, immune regulation and drug resistance (72–74). It has been confirmed that the UPS can mediate tumor immune evasion and drug resistance induced by immune checkpoint inhibitor therapy by regulating the ubiquitination level of PD-1/PD-L1 (31,51,52). In-depth exploration of the potential molecular mechanisms by which the UPS regulates PD-1/PD-L1 will not only help improve the efficacy of tumor immunotherapy, but also provide a solid theoretical basis for the development of novel therapeutic strategies.

In recent years, targeting the UPS for disease treatment has become an important direction in drug development, showing great therapeutic potential in a number of fields such as autoimmune diseases and neurodegenerative diseases and cancer (75,76). Studies have shown that targeting the ubiquitination process of PD-1/PD-L1 mediated by the UPS has achieved notable effects in inhibiting tumor immune evasion and improving tumor resistance to anti-PD-1/PD-L1 therapy. For example, the small-molecule agonist AK087 of ITCH can effectively promote the ubiquitination and degradation of PD-L1 mediated by ITCH and markedly inhibit the resistance of tumors to anti-PD-1/PD-L1 treatment (39). The USP8 inhibitor DUB-IN-2 can effectively inhibit the deubiquitination process of PD-L1 mediated by USP8. This action notably inhibits tumor immune evasion and can effectively improve the efficacy of tumor immunotherapy (53). The small-molecule compound ML364 directly binds to USP2 and inhibits its deubiquitinase activity (77). The UPS not only determines the protein stability of PD-L1, but also forms a ‘positive-negative feedback’ loop with the IFN-γ/JAK-STAT signaling pathway through key nodes including TRIM25/SOCS1/USP18. Targeting this regulatory loop can amplify or suppress IFN-γ signaling across different cancer types, thereby guiding combination strategies between UPS inhibitors and immune checkpoint inhibitors (78,79).

This therapeutic model targeting the UPS has a unique mechanism of action. Instead of directly blocking protein functions, it regulates the stability and degradation of proteins from the source, thereby affecting the levels of abnormal proteins. Meanwhile, this therapeutic model has precise targeting ability, which can reduce the biological toxicity caused by broad-spectrum inhibition. Due to tissue specificity, the functions of E3 ubiquitin ligases also vary. The ‘functional switch’ of the same E3 ubiquitin ligase in different cancers is collectively determined by protein expression in the tumor microenvironment, signaling pathways and post-translational modifications. For instance, the mutational inactivation and dysregulated expression of SPOP in various types of cancer can affect its E3 ubiquitin ligase activity. Notably, Skp2-mediated PD-L1 K63 ubiquitination in non-small cell lung cancer depends on LKB1 inactivation (Fig. 2), whereas BRAF inhibitors in melanoma suppress PD-L1 ubiquitination by inhibiting the ERK-GSK3β-β-TrCP axis (41,80). This explains the reason Skp2 inhibitors (compound #25) can enhance the efficacy of PD-1 antibodies in non-small cell lung cancer, while melanoma requires combined MAPK inhibition to relieve β-TrCP suppression. For example, preclinical data on the USP7 inhibitor P5091 in gastric cancer showed that H. pylori-positive patients (with concomitant USP7 overexpression) had a 3.2-fold higher response rate compared with negative patients (P<0.01), suggesting that future trials should stratify patients based on microbiome-UPS co-mutation status (51). By contrast, a phase II trial of the CSN5 inhibitor curcumin in TNBC (trial no. NCT00094445) demonstrated limited efficacy due to the lack of screening for CSN5-high populations, highlighting the necessity of biomarker-guided therapy. Although UPS-targeted drugs such as USP8 inhibitors may develop resistance due to mutations or compensatory pathways, current evidence suggests that such resistance mechanisms are independent of PD-1/PD-L1 ubiquitination regulation. Future studies should explore whether resistance to UPS-targeted drugs upregulates PD-L1 through non-UPS pathways such as through transcriptional reprogramming or exosome release, thereby indirectly leading to immunotherapy failure. It is recommend that future studies employ cancer-specific organoid models to validate UPS-targeting drugs, thereby mimicking the effect of stromal cells on ubiquitination regulation within the tumor microenvironment.

Although some molecular drugs targeting the UPS have achieved preliminary progress in improving tumor immunotherapy, the number of those in clinical verification stage is still limited. As of November 2024, <20 UPS-targeting agents have entered clinical stages globally, with most concentrated in the proteolysis-targeting chimera (PROTAC). Notably, UPS modulators specifically targeting PD-1/PD-L1 remain in Phase I or earlier development (81). High expression of E3 ubiquitin ligases such as CRBN and VHL in the liver and kidneys often leads to hematological and renal toxicity, Excessive PROTACs may form nonfunctional binary complexes, disrupting UPS activity and causing drug ‘rebound effects’ (82). Dong et al (83) encapsulated VPS18/11 inhibitors such as RD-N into lung-targeted nanoparticles, effectively reversing tumor resistance and suppressing metastasis. Similar strategies could enable tissue-specific delivery of deubiquitinase inhibitors, minimizing off-target effects and systemic toxicity. UPS gene mutations or compensatory pathway activation can result in adaptive resistance during long-term treatment (84). Real-time drug concentration tracking, combined with artificial intelligence and machine learning-based predictive models, may optimize pharmacokinetic profiles and address long-term adaptive resistance.

The PROTAC technology, as an emerging therapeutic strategy for targeted protein degradation in recent years, has gradually attracted widespread attention (85,86). The PROTAC consists of three parts: i) A target protein-binding ligand; ii) an E3 ligase ligand; and iii) a linker. Its main mechanism of action is to induce the binding of the E3 ligase to the target protein, mediate the ubiquitination of the target protein and subsequently promote the degradation of the target protein (87). This technology has overcome the limitation of the ‘undruggable’ status of certain proteins and can reversibly regulate the expression levels of proteins over time. However, PROTAC drugs usually have a relatively large molecular weight, which leads to limited bioavailability. Therefore, the drug design of PROTAC still needs further optimization.

Conventional PROTACs suffer from high molecular weight, poor membrane permeability and low oral bioavailability, leading to weak in vitro-in vivo association and limited clinical translation. To overcome these hurdles, Sun et al (88) developed tumor microenvironment (TME)-responsive enzyme-activated click-forming PROTACs (ENCTACs). By exploiting cathepsin B overexpressed in >90% of solid tumors as a biological trigger, an orthogonal cleavage-click reaction assembles the active degrader in situ, selectively eliminating the epigenetic regulator BRD4 and consequently downregulating PD-L1 to remodel the immune microenvironment. In the 4T1 TNBC mouse model, ENCTACs achieved a 65% tumor-growth inhibition, a three-fold deeper tissue penetration and negligible systemic toxicity compared with traditional PROTACs. In parallel, a recent study conjugated a CD47 antibody with a folate ligand to create a Folate Receptor Targeting Chimera (FRTAC). Leveraging the high folate-receptor expression on cancer cells, FRTAC drives CD47 into lysosomal degradation via receptor-mediated endocytosis, markedly potentiating macrophage-mediated phagocytosis while sparing normal tissues (89). At present, multiple PROTAC drugs for tumor treatment have entered the clinical stage, such as ARV-110 for prostate cancer (90) and ARV-471 for ER⁺ breast cancer (91). Looking ahead, next-generation PROTAC platforms that integrate TME-specific activation with nanoparticle-based delivery are poised to surmount the dual barriers of cellular permeability and off-target toxicity, offering unprecedented opportunities for cancer immunotherapy.

In conclusion, the present review discussed the potential molecular mechanisms by which the UPS plays a role in tumor immune evasion and resistance to anti-PD-1/PD-L1 therapy and has summarized the potential targeted drugs that can inhibit tumor immune evasion and overcome resistance to immune checkpoint therapy by targeting the UPS. Future research should focus on the following key aspects: i) Further in-depth exploration of the regulatory mechanisms of the UPS on various immune checkpoints in different tumor types and immune microenvironments; ii) optimization of the design of targeted UPS drugs to improve their targeting ability and bioavailability; and iii) development of combined treatment regimens of immune checkpoint inhibitors and targeted UPS drugs to enhance the synergistic therapeutic effect. These research directions will lay a solid theoretical foundation for the development of the next-generation precision immunotherapy strategies.