Immune checkpoints are critical in modulating immune responses and maintaining self-tolerance. Cancer cells can exploit these mechanisms to evade immune detection, making immune checkpoints attractive targets for cancer therapy. The introduction of immune checkpoint inhibitors (ICIs) has transformed cancer treatment, with monoclonal antibodies targeting CTLA-4, PD-1, and PD-L1 demonstrating clinical success. However, challenges such as immune-related adverse events, primary and acquired resistance, and high treatment costs persist. To address these challenges, it is essential to explore alternative strategies, including small-molecule and peptide-based inhibitors, aptamers, RNA-based therapies, gene-editing technologies, bispecific and multispecific agents, and cell-based therapies. Additionally, innovative approaches such as lysosome-targeting chimeras, proteolysis-targeting chimeras, and N-(2-hydroxypropyl) methacrylamide copolymers are emerging as promising options for enhancing treatment effectiveness. This review highlights significant advancements in the field, focusing on their clinical implications and successes.

Department of Cell Biology Faculty of Science Charles University Prague Czech Republic

Institute of Organic Chemistry and Biochemistry Czech Academy of Sciences Flemingovo n 2 Prague 6 Prague 16610 Czech Republic

See more in PubMed

Pardoll DM. The blockade of immune checkpoints in cancer immunotherapy. Nat Rev Cancer. 2012;12(4):252–64. PubMed PMC

Ribas A, Wolchok JD. Cancer immunotherapy using checkpoint blockade. Science. 2018;359(6382):1350–5. PubMed PMC

Farkona S, Diamandis EP, Blasutig IM. Cancer immunotherapy: the beginning of the end of cancer? BMC Med. 2016;14(1):73. PubMed PMC

Schoenfeld AJ, Hellmann MD. Acquired resistance to immune checkpoint inhibitors. Cancer Cell. 2020;37(4):443–55. PubMed PMC

Li S-J, Sun Z-J. Fueling immune checkpoint blockade with oncolytic viruses: current paradigms and challenges ahead. Cancer Lett. 2022;550: 215937. PubMed

Hu Z. Chapter 11—using CAR-NK cells to overcome the host resistance to antibody immunotherapy and immune checkpoint blockade therapy. In: Successes and Challenges of NK Immunotherapy, B. Bonavida and A. Jewett, editors. Academic Press; 2021. p. 193–212.

Topalian SL, Drake CG, Pardoll DM. Immune checkpoint blockade: a common denominator approach to cancer therapy. Cancer Cell. 2015;27(4):450–61. PubMed PMC

Andrews LP, Yano H, Vignali DAA. Inhibitory receptors and ligands beyond PD-1, PD-L1 and CTLA-4: breakthroughs or backups. Nat Immunol. 2019;20(11):1425–34. PubMed

Guo J, et al. Beyond inhibition against the PD-1/PD-L1 pathway: development of PD-L1 inhibitors targeting internalization and degradation of PD-L1. RSC Med Chem. 2024;15(4):1096–108. PubMed PMC

Zahavi D, Weiner L. Monoclonal antibodies in cancer therapy. Antibodies (Basel). 2020;9(3). PubMed PMC

Camacho LH. CTLA-4 blockade with ipilimumab: biology, safety, efficacy, and future considerations. Cancer Med. 2015;4(5):661–72. PubMed PMC

Darvin P, et al. Immune checkpoint inhibitors: recent progress and potential biomarkers. Exp Mol Med. 2018;50(12):1–11. PubMed PMC

Qin S, et al. Novel immune checkpoint targets: moving beyond PD-1 and CTLA-4. Mol Cancer. 2019;18(1):155. PubMed PMC

Arce Vargas F, et al. Fc effector function contributes to the activity of human anti-CTLA-4 antibodies. Cancer Cell. 2018;33(4):649-663.e4. PubMed PMC

Sato Y, et al. Fc-independent functions of anti-CTLA-4 antibodies contribute to anti-tumor efficacy. Cancer Immunol Immunother. 2022;71(10):2421–31. PubMed PMC

Hock BD, et al. Functional effects of immune complexes formed between pembrolizumab and patient-generated anti-drug antibodies. Cancer Immunol Immunother. 2020;69(12):2453–64. PubMed PMC

Yin Q, et al. Immune-related adverse events of immune checkpoint inhibitors: a review. Front Immunol. 2023;14:1167975. PubMed PMC

Andrews A. Treating with checkpoint inhibitors-figure $1 million per patient. Am Health Drug Benefits. 2015;8(Spec Issue):9. PubMed PMC

MalekiVareki S. High and low mutational burden tumors versus immunologically hot and cold tumors and response to immune checkpoint inhibitors. J Immunother Cancer. 2018;6(1):157. PubMed PMC

Memon D, et al. Clinical and molecular features of acquired resistance to immunotherapy in non-small cell lung cancer. Cancer Cell. 2024;42(2):209-224.e9. PubMed PMC

Dobosz P, et al. Challenges of the immunotherapy: perspectives and limitations of the immune checkpoint inhibitor treatment. Int J Mol Sci. 2022;23(5). PubMed PMC

Chen L, et al. Development of small molecule drugs targeting immune checkpoints. Cancer Biol Med. 2024;21(5):382–99. PubMed PMC

Smith MC, Gestwicki JE. Features of protein-protein interactions that translate into potent inhibitors: topology, surface area and affinity. Expert Rev Mol Med. 2012;14: e16. PubMed PMC

Sasikumar PG, et al. PD-1 derived CA-170 is an oral immune checkpoint inhibitor that exhibits preclinical anti-tumor efficacy. Communications Biology. 2021;4(1):699. PubMed PMC

Zauderer M, et al. P2.06-07 phase 1 study of CA-170: first-in-class small molecule targeting VISTA/PD-L1 in patients with malignant pleural mesothelioma. J Thoracic Oncol. 2019;14(10):S757–S758.

ctri.nic.in. CTRI/2021/10/037699. Available from: https://ctri.nic.in/Clinicaltrials/showallp.php?mid1=60435&EncHid=&userName=erode%20cancer%20centre.

Lai F, et al. YPD-30, a prodrug of YPD-29B, is an oral small-molecule inhibitor targeting PD-L1 for the treatment of human cancer. Acta Pharm Sin B. 2022;12(6):2845–58. PubMed PMC

Hargadon KM, Johnson CE, Williams CJ. Immune checkpoint blockade therapy for cancer: an overview of FDA-approved immune checkpoint inhibitors. Int Immunopharmacol. 2018;62:29–39. PubMed

Koblish HK, et al. Characterization of INCB086550: a potent and novel small-molecule PD-L1 inhibitor. Cancer Discov. 2022;12(6):1482–99. PubMed PMC

MAX-10181. Available from: https://synapse.patsnap.com/drug/e1d01add4b354ae99168cc53d7778678?utm_source=chatgpt.com.

Wang Y, et al. Abstract 4046: orally active PD-L1 inhibitor MAX-10181 in combination with temozolomide effectively prolonged survival in GBM animal model study. Cancer Res. 2024;84(6_Supplement):4046.

A phase 1b/2 dose escalation/expansion study to evaluate the safety, tolerability, pharmacokinetics, and efficacy of GS-4224 in subjects with advanced solid tumors; 2019.

Odegard JM, et al. Oral PD-L1 inhibitor GS-4224 selectively engages PD-L1 high cells and elicits pharmacodynamic responses in patients with advanced solid tumors. J Immunother Cancer. 2024;12(4). PubMed PMC

Hu Z, et al. PCC0208025 (BMS202), a small molecule inhibitor of PD-L1, produces an antitumor effect in B16–F10 melanoma-bearing mice. PLoS ONE. 2020;15(3): e0228339. PubMed PMC

Park JJ, et al. Checkpoint inhibition through small molecule-induced internalization of programmed death-ligand 1. Nat Commun. 2021;12(1):1222. PubMed PMC

Shiravand Y, et al. Immune checkpoint inhibitors in cancer therapy. Curr Oncol. 2022;29(5):3044–60. PubMed PMC

Sui X, et al. Peptide drugs: a new direction in cancer immunotherapy. Cancer Biol Med. 2023;21(3):198–203. PubMed PMC

Randomized, double-blinded, placebo-controlled, single ascending dose study to evaluate the pharmacokinetics, safety, tolerability and pharmacodynamics of BMS-986189 in healthy subjects; 2016.

Nimjee SM, Rusconi CP, Sullenger BA. Aptamers: an emerging class of therapeutics. Annu Rev Med. 2005;56:555–83. PubMed

Kohlberger M, Gadermaier G. SELEX: critical factors and optimization strategies for successful aptamer selection. Biotechnol Appl Biochem. 2022;69(5):1771–92. PubMed PMC

Zhou J, Rossi J. Aptamers as targeted therapeutics: current potential and challenges. Nat Rev Drug Discovery. 2017;16(3):181–202. PubMed PMC

Huang Y, et al. Aptamer-based immune checkpoint inhibition for cancer immunotherapy. ChemBioChem. 2025;26(1): e202400599. PubMed

Brown A, et al. Development of Better Aptamers: Structured Library Approaches, Selection Methods, And Chemical Modifications. Angew Chem Int Ed. 2024;63(16): e202318665. PubMed

Fu J, et al. Novel bispecific aptamer targeting PD-1 and nucleolin for cancer immunotherapy. Cancer Nanotechnology. 2023;14(1):27.

Camorani S, et al. Aptamer targeted therapy potentiates immune checkpoint blockade in triple-negative breast cancer. J Exp Clin Cancer Res. 2020;39(1):180. PubMed PMC

Sun Y, et al. Bispecific aptamer-based recognition-then-conjugation strategy for PD1/PDL1 axis blockade and enhanced immunotherapy. ACS Nano. 2022;16(12):21129–38. PubMed

Huang B-T, et al. A CTLA-4 antagonizing DNA aptamer with antitumor effect. Molecular Therapy Nucl Acids. 2017;8:520–8. PubMed PMC

De Mey W, et al. RNA in cancer immunotherapy: unlocking the potential of the immune system. Clin Cancer Res. 2022;28(18):3929–39. PubMed PMC

Zabeti Touchaei A, Vahidi S. MicroRNAs as regulators of immune checkpoints in cancer immunotherapy: targeting PD-1/PD-L1 and CTLA-4 pathways. Cancer Cell International. 2024;24(1):102. PubMed PMC

Kanasty R, et al. Delivery materials for siRNA therapeutics. Nat Mater. 2013;12(11):967–77. PubMed

Ali Zaidi SS, et al. Engineering siRNA therapeutics: challenges and strategies. J Nanobiotechnol. 2023;21(1):381. PubMed PMC

Rostami F, et al. PDL1 targeting by miR-138-5p amplifies anti-tumor immunity and Jurkat cells survival in non-small cell lung cancer. Sci Rep. 2024;14(1):13542. PubMed PMC

Zhao L, et al. The tumor suppressor miR-138-5p targets PD-L1 in colorectal cancer. Oncotarget. 2016;7(29):45370–84. PubMed PMC

Yin M, Zhang Z, Wang Y. Anti-tumor effects of miR-34a by regulating immune cells in the tumor microenvironment. Cancer Med. 2023;12(10):11602–10. PubMed PMC

Deng S, et al. p53 downregulates PD-L1 expression via miR-34a to inhibit the growth of triple-negative breast cancer cells: a potential clinical immunotherapeutic target. Mol Biol Rep. 2023;50(1):577–87. PubMed

Klicka K, et al. The role of miR-200 family in the regulation of hallmarks of cancer. Front Oncol. 2022;12: 965231. PubMed PMC

Xuan J, et al. Sequence requirements for miR-424-5p regulating and function in cancers. Int J Mol Sci. 2022;23(7). PubMed PMC

Li T, et al. CRISPR/Cas9 therapeutics: progress and prospects. Signal Transduct Target Ther. 2023;8(1):36. PubMed PMC

Liu Z, et al. Recent advances and applications of CRISPR-Cas9 in cancer immunotherapy. Mol Cancer. 2023;22(1):35. PubMed PMC

Pacesa M, Pelea O, Jinek M. Past, present, and future of CRISPR genome editing technologies. Cell. 2024;187(5):1076–100. PubMed

Hennig SL, et al. Evaluation of mutation rates, mosaicism and off target mutations when injecting Cas9 mRNA or protein for genome editing of bovine embryos. Sci Rep. 2020;10(1):22309. PubMed PMC

A phase I clinical trial of PD-1 knockout engineered T cells treating patients with advanced non-small cell lung cancer. In: C.L. Chengdu MedGenCell, editor; 2016.

Lu Y, et al. Safety and feasibility of CRISPR-edited T cells in patients with refractory non-small-cell lung cancer. Nat Med. 2020;26(5):732–40. PubMed

Wang Z, et al. Phase I study of CRISPR-engineered CAR-T cells with PD-1 inactivation in treating mesothelin-positive solid tumors. J Clin Oncol. 2020;38(15_suppl):3038.

Labrijn AF, et al. Bispecific antibodies: a mechanistic review of the pipeline. Nat Rev Drug Discovery. 2019;18(8):585–608. PubMed

Buzzetti M, Gerlinger M. Assessing the toxicity of bispecific antibodies. Nat Biomed Eng. 2024;8(4):339–40. PubMed

Zhou Y, et al. Immunogenicity assessment of bispecific antibody-based immunotherapy in oncology. J Immunother Cancer. 2022;10(4). PubMed PMC

Phase II trial of XmAb20717 in patients with advanced biliary tract cancers. In: Xencor I, editor; 2022.

An open label, multicenter, dose escalation and expansion, phase 1 study to evaluate safety, pharmacokinetics, and preliminary anti-tumor activity of RO7121661, a PD-1/TIM-3 bispecific antibody, in patients with advanced and/or metastatic solid tumors; 2018.

A 3-arm, randomized, blinded, active-controlled, phase II study of RO7121661, a PD1-TIM3 bispecific antibody and RO7247669, a PD1-LAG3 bispecific antibody, compared with Nivolumab in participants with advanced or metastatic squamous cell carcinoma of the esophagus; 2021.

A phase 1 dose escalation and expansion study of ABL501, a bispecific antibody of PD-L1 and LAG-3 as a single agent in subjects with any progressive, locally advanced (unresectable) or metastatic solid tumors; 2021.

Sung E, et al. LAG-3xPD-L1 bispecific antibody potentiates antitumor responses of T cells through dendritic cell activation. Mol Ther. 2022;30(8):2800–16. PubMed PMC

Pang X, et al. Cadonilimab, a tetravalent PD-1/CTLA-4 bispecific antibody with trans-binding and enhanced target binding avidity. mAbs. 2023;15(1):2180794. PubMed PMC

Cadonilimab (PD-1/CTLA-4 Bi-specific antibody) combined with chemoradiotherapy in high-risk locoregionally-advanced nasopharyngeal carcinoma: a randomized, controlled, multicenter, phase 3 Clinical Trial, I. Akeso Pharmaceuticals, Editor; 2022.

Cao F, et al. An open-label, single-center phase II trial of cadonilimab (an anti-PD-1/CTLA-4 bispecific antibody) in combination with platinum-based dual-drug neoadjuvant chemotherapy for locally advanced, resectable head and neck squamous cell carcinoma. J Clin Oncol. 2024;42(16_suppl):6044.

Zheng Q, et al. Lysosome-targeting chimera (LYTAC): a silver bullet for targeted degradation of oncogenic membrane proteins. MedComm Oncol. 2024;

Ji P, et al. Lysosome-targeting bacterial outer membrane vesicles for tumor specific degradation of PD-L1. Small. 2024:e2400770. PubMed

Ahn G, et al. Elucidating the cellular determinants of targeted membrane protein degradation by lysosome-targeting chimeras. Science. 2023;382(6668):eadf6249. PubMed PMC

Li S, et al. PROTACs: novel tools for improving immunotherapy in cancer. Cancer Lett. 2023;560: 216128. PubMed

Khan S, et al. PROteolysis targeting chimeras (PROTACs) as emerging anticancer therapeutics. Oncogene. 2020;39(26):4909–24. PubMed PMC

Békés M, Langley DR, Crews CM. PROTAC targeted protein degraders: the past is prologue. Nat Rev Drug Discovery. 2022;21(3):181–200. PubMed PMC

Wang H, et al. HIP1R targets PD-L1 to lysosomal degradation to alter T cell–mediated cytotoxicity. Nat Chem Biol. 2019;15(1):42–50. PubMed

Chytil P, Kostka L, Etrych T. HPMA copolymer-based nanomedicines in controlled drug delivery. J Pers Med. 2021;11(2). PubMed PMC

Li L, et al. Inhibition of immunosuppressive tumors by polymer-assisted inductions of immunogenic cell death and multivalent PD-L1 crosslinking. Adv Funct Mater. 2020;30(12). PubMed PMC

Zamani MR, et al. Polymer-based antibody mimetics (iBodies) target human PD-L1 and function as a potent immune checkpoint blocker. J Biol Chem. 2024;300(6): 107325. PubMed PMC

Lu SJ, Feng Q. CAR-NK cells from engineered pluripotent stem cells: off-the-shelf therapeutics for all patients. Stem Cells Transl Med. 2021;10(Suppl 2):S10-s17. PubMed PMC

Li Y, et al. Human iPSC-derived natural killer cells engineered with chimeric antigen receptors enhance anti-tumor activity. Cell Stem Cell. 2018;23(2):181-192.e5. PubMed PMC

Daher M, Rezvani K. Outlook for new CAR-based therapies with a focus on CAR NK cells: what lies beyond CAR-engineered T cells in the race against cancer. Cancer Discov. 2021;11(1):45–58. PubMed PMC

Wang F, et al. Combined treatment with anti-PSMA CAR NK-92 cell and anti-PD-L1 monoclonal antibody enhances the antitumour efficacy against castration-resistant prostate cancer. Clin Transl Med. 2022;12(6): e901. PubMed PMC

Strassheimer F, et al. CAR-NK cell therapy combined with checkpoint inhibition induces an NKT cell response in glioblastoma. Br J Cancer. 2025;132(9):849–60. PubMed PMC

Zhi L, et al. CAR-NK cells with dual targeting of PD-L1 and MICA/B in lung cancer tumor models. BMC Cancer. 2025;25(1):337. PubMed PMC

Hosseinalizadeh H, et al. Emerging combined CAR-NK cell therapies in cancer treatment: finding a dancing partner. Molecular Ther. 2025. PubMed PMC

Lovatt C, Parker AL. Oncolytic viruses and immune checkpoint inhibitors: the “hot” new power couple. Cancers. 2023;15(16):4178. PubMed PMC

LaRocca CJ, Warner SG. Oncolytic viruses and checkpoint inhibitors: combination therapy in clinical trials. Clin Transl Med. 2018;7(1): e35. PubMed PMC

Sivanandam V, et al. Oncolytic viruses and immune checkpoint inhibition: the best of both worlds. Molecular Ther Oncolytics. 2019;13:93–106. PubMed PMC

Zhang T, et al. Talimogene laherparepvec (T-VEC): a review of the recent advances in cancer therapy. J Clin Med. 2023;12(3):1098. PubMed PMC