Dual Blockade of PD-L1 and AXL: A Novel Immunotherapeutic Approach for Ovarian and Cervical Cancer
-
Hossein Rahavi
,
Ahmad Najafi
,
Hossein Asgarian-Omran
,
Behrouz Aflatoonian
,
Mohsen Tehrani
,
Ehsan Farashahi-Yazd
Abstract
Tumor microenvironment modulators have produced durable effects in cancer treatment. Targeting immune checkpoint receptors, such as PD-L1, has demonstrated efficacy in eliciting antitumor responses. However, resistance to immune checkpoint blockers (ICBs) has constrained the efficacy of these therapies. Previous studies showed a link between the expression of AXL receptor tyrosine kinase and resistance to ICBs. Therefore, designing combination treatments with synergistic mechanisms to overcome ICB-based resistance is needed. In addition to antibody-based therapies, gene silencing with siRNAs has recently been explored to alter the cancer environment to enhance the immune response.
In this study, we targeted PD-L1 using an siRNA and AXL using a blocker (R428) in OVACAR-3 and CaSki cells, ovarian and cervical cancer cell lines, respectively, in the following groups: Scramble-siRNA, PD-L1-siRNA, Scramble-siRNA in conjunction with R428, PD-L1-siRNA in conjunction with R428, R428 monotherapy and untreated controls. Cell viability was assessed by MTT assay after 48 hours of treatment, and cisplatin sensitization was evaluated in resistant OVACAR-3 cells. Gene expression was analyzed by qRT-PCR, while flow cytometry quantified CD44+PD-L1+ populations, apoptosis (Annexin V/PI), and cell cycle distribution.
The results showed a significant decrease in cell proliferation, suppression of EMT-regulating genes, reduction of stemness in cancer cells, increased apoptosis and disruption of the cell cycle in the studied cell lines.
These findings suggest that simultaneous blockade of PD-L1 and AXL could serve as a novel tumor-suppressive strategy, especially for cancer patients resistant to ICBs.
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8. Bakir B, Chiarella AM, Pitarresi JR, Rustgi AK. EMT, MET, plasticity, and tumor metastasis. Trends Cell Biol. 2020;30(10):764–76.
9. Skinner HD, Sandulache VC, Ow TJ, Meyn RE, Yordy JS, Beadle BM, et al. AXL-PI3K-PD-L1 signaling axis and radiation resistance. Clin Cancer Res. 2017;23(11):2713–22.
10. Tsukita Y, Fujino N, Miyake K, Tsurudome Y, Araki T, Ohsaki Y, et al. Axl kinase drives immune checkpoint and chemokine signalling in lung adenocarcinomas. Mol Cancer. 2019;18(1):24.
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15. Li J, Wang J, Chen X, Zhao Y, Zhang L, Xu Y, et al. ICIs and TKIs in advanced colorectal cancer: Meta-analysis. Oncol Lett. 2024;27(4):153.
16. Wu Y, Chen W, Xu Z, Zhao Z, Wang X. PD-L1 distribution and perspectives for cancer immunotherapy. Front Immunol. 2019;10:2022.
17. Hu B, Weng Y, Xia XH, Liang XJ, Huang Y. Therapeutic siRNA: State of the art. Signal Transduct Target Ther. 2020;5(1):101.
18. Ozcan G, Ozpolat B, Coleman RL, Sood AK, Lopez-Berestein G. Development of siRNA-based therapeutics.
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19. Teo PY, Yang C, Guo X, Liu X, Shen J. PD-L1 siRNA delivery for ovarian cancer immunotherapy. Adv Healthc Mater. 2015;4(8):1180–9.
20. Kang SW, Choi HJ, Cho M, Shin H, Kim YK. Transcriptome profiling of ovarian cancer xenografts. Sci Rep. 2024;14(1):11894.
21. Lee EH, Kim JY, Jeong JH, Choi JW, Kim YJ. Axl blockade in HPV16E6-mediated cervical cancer. Sci Rep. 2017;7(1):5759.
22. Mujoo K, Kipps TJ, Yang HM, Cherney B, Larrick JW. Emergence of cisplatin-resistant OVCAR-3 cells. Int J Gynecol Cancer. 2000;10(2):105–14.
23. Wang L, Xu W, Zhang L, Hu J, Xia X. Drug resistance in ovarian cancer: Mechanisms to clinical trials. Mol Cancer. 2024;23(1):66.
24. Nagasaki J, Ishino T, Togashi Y. Mechanisms of resistance to immune checkpoint inhibitors. Cancer Sci. 2022;113(10):3303–12.
25. Santoro A, Angi M, Camarda F, Dellino M, Bogani G. Role of PD-L1 in gynecological cancers. Gynecol Oncol. 2024;184:57–66.
26. Bhalla S, Gerber DE. AXL inhibitors: Status of clinical development. Curr Oncol Rep. 2023;25(5):521–9.
27. Liu Y, Li X, Zhang Y, Qian J, Yang L. AXL and the immunosuppressive microenvironment. Mol Cancer. 2025;24(1):11.
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29. Tanaka T, Fujiwara T, Saito Y, Kudo M, Nakamura M. Anti-PD-L1 promotes proliferation via PD-L1–AXL in liver cancer. Hepatol Int. 2024;18(3):984–97.
30. Yu W, Zhang X, Chen G, Xu R, Wang S. PD-L1 activates WIP and β-catenin in lung cancer. Cell Death Dis. 2020;11(7):506.
31. Boshuizen J, Vredevoogd DW, Krijgsman O, Ligtenberg MA, de Bruijn B, Jimenez CR, et al. AXL-ADC and ICB in melanoma and lung cancer. Cancer Res. 2021;81(7):1775–87.
32. Li C, Han X. Melanoma immunotherapy with PD-L1 siRNA and imatinib. Pharm Res. 2020;37(6):109.
33. Tian M, Wang H, Wang J, Zhang W, Chen Q. AXL inhibition sensitizes ovarian cancer to cisplatin. Acta Pharmacol Sin. 2021;42(7):1180–9.
34. Oien DB, Pathoulas CL, Ray U, Shi R, Conejo-Garcia JR, Lengyel E, et al. AXL inhibitor enhances mesothelioma cell death. Front Pharmacol. 2017;8:970.
35. Bastaki S, Bae DH, Mecham B, Richardson DR, Huang
ML. Codelivery of STAT3 and PD-L1 siRNA suppresses cancer. Life Sci. 2021;266:118847.
36. Han S, Wang H, Zhang C, Liu L, Li S. AXL inhibition suppresses esophageal carcinoma growth. Exp Ther Med. 2020;20(5):41.
37. Almozyan S, Colak D, Mansour F, Alaiya A, Ghebeh H. PD-L1 promotes stemness in breast cancer. Int J Cancer. 2017;141(7):1402–12.
38. Idowu MO, Kmieciak M, Dumur CI, Burton RS, Grimes MM, Powers CN, et al. CD44+/CD24−/low cells in triple-negative breast cancer. Hum Pathol. 2012;43(3):364–73.
39. Alwosaibai K, Zakaria Z, Yahaya BH, Abdullah H, Lim KP. PD-L1 and cancer stem cells in ovarian cancer. BMC Cancer. 2023;23(1):13.
40. Zhang J, Ding L, Tian Y, Li X, Yu S. CSC-like traits in cervical cancer and radioresistance. Gynecol Obstet Invest. 2019;84(2):174–82.
41. Woo SM, Min KJ, Kim S, Park JW, Kim DE, Kim SH, et al. AXL inhibitor R428 enhances TRAIL-induced apoptosis. Int J Mol Sci. 2019;20(13):–.
42. Li J, Xu J, Yan X, Jin K, Li W. PD-L1 knockdown in gastric cancer cells. Cell Physiol Biochem. 2017;41(3):907–20.
43. Mitra AK, Davis DA, Tomar S, Roy A, Gurler H, Xie JL, et al. In vivo growth of ovarian cancer cell lines. Gynecol Oncol. 2015;138(2):372–7.
44. Chowdhury S, Matin K, Ali M, Paul P, Rahman M. PD-L1 as a marker in thyroid cancer. Oncotarget. 2016;7(22):32318–28.
45. Ghebeh H, Tulbah A, Mohammed S, Elkum N, Bin Amer S, Alaiya A, et al. Doxorubicin modulates PD-L1 expression. Breast Cancer Res. 2010;12(4)\:R48.
46. Hua D, Zheng S, Xu G, Yuan X, Zhang B, Chen Y, et al. B7-H1 and regulatory T cells in colorectal cancer. World J Gastroenterol. 2012;18(9):971–8.
47. Satelli A, Batth IS, Brownlee Z, Mitra A, Zhou S, Rojas C, et al. Nuclear PD-L1 in circulating tumor cells. Sci Rep. 2016;6:28910.
48. Tang Y, Fang W, Zhang Y, Hong S, Kang S, Yan Y, et al. PD-L1, EGFR and prognosis in NSCLC. Oncotarget. 2015;6(16):14209–19.
49. Ukpo OC, Thorstad WL, Lewis JS Jr. B7-H1 in HPV-related oropharyngeal cancer. Head Neck Pathol. 2013;7(2):113–21.
50. Qu QX, Xie F, Huang Q, Zhang XH, Xie J, Wang XH, et al. PD-L1 expression in ovarian cancer. Cell Physiol Biochem. 2017;43(5):1893–906.
51. Parvez A, Alam M, Hamid AS, Suhail M, Ahmad A. PD-1 and PD-L1 in cancer immunotherapy. Front Immunol. 2023;14:1296341.
2. Stewart C, Ralyea C, Lockwood S. Ovarian cancer: An integrated review. Semin Oncol Nurs. 2019;35(2):151–6.
3. Jiang G, Wu Q, Li B. Evaluation of immunotherapy efficacy in gynecologic cancer. Front Immunol. 2023;14:1061761.
4. Wherry EJ, Kurachi M. Molecular and cellular insights into T cell exhaustion. Nat Rev Immunol. 2015;15(8):
486–99.
5. Akinleye A, Rasool Z. Immune checkpoint inhibitors of PD-L1 as cancer therapeutics. J Hematol Oncol. 2019;12(1):92.
6. Yang Y, Wang L, Han R, Wang Q, Zhang G, Liu H, et al. Protein tyrosine kinase inhibitor resistance in malignant tumors: Molecular mechanisms and future perspective. Signal Transduct Target Ther. 2022;7(1):329.
7. Zhu C, Wei Y, Wei X, Wang D, Zheng Z, Zhang Y, et al. A dual MET/AXL small-molecule inhibitor exerts efficacy against gastric carcinoma. MedComm (2020). 2020;1(1):103–18.
8. Bakir B, Chiarella AM, Pitarresi JR, Rustgi AK. EMT, MET, plasticity, and tumor metastasis. Trends Cell Biol. 2020;30(10):764–76.
9. Skinner HD, Sandulache VC, Ow TJ, Meyn RE, Yordy JS, Beadle BM, et al. AXL-PI3K-PD-L1 signaling axis and radiation resistance. Clin Cancer Res. 2017;23(11):2713–22.
10. Tsukita Y, Fujino N, Miyake K, Tsurudome Y, Araki T, Ohsaki Y, et al. Axl kinase drives immune checkpoint and chemokine signalling in lung adenocarcinomas. Mol Cancer. 2019;18(1):24.
11. Jin H, Wang L, Bernards R. Rational combinations of targeted cancer therapies. Nat Rev Drug Discov. 2023;22(3):213–34.
12. Yang C, Xia BR, Zhang ZC, Zhang YJ, Lou G. Immunotherapy for ovarian cancer: Adjuvant, combination, and neoadjuvant. Front Immunol. 2020;11:577869.
13. Iinuma K, Saito K, Hirano Y, Suzuki Y, Nagai K, Tanaka H, et al. Combination therapy in metastatic renal cell carcinoma. Cancers (Basel). 2023;15(3):–.
14. Li H, Li Y, Liu H, Shi Y, Zhang Y, Liang Q, et al. AXL targeting restores PD-1 blockade sensitivity in NSCLC. Cell Rep Med. 2022;3(3):100554.
15. Li J, Wang J, Chen X, Zhao Y, Zhang L, Xu Y, et al. ICIs and TKIs in advanced colorectal cancer: Meta-analysis. Oncol Lett. 2024;27(4):153.
16. Wu Y, Chen W, Xu Z, Zhao Z, Wang X. PD-L1 distribution and perspectives for cancer immunotherapy. Front Immunol. 2019;10:2022.
17. Hu B, Weng Y, Xia XH, Liang XJ, Huang Y. Therapeutic siRNA: State of the art. Signal Transduct Target Ther. 2020;5(1):101.
18. Ozcan G, Ozpolat B, Coleman RL, Sood AK, Lopez-Berestein G. Development of siRNA-based therapeutics.
Adv Drug Deliv Rev. 2015;87:108–19.
19. Teo PY, Yang C, Guo X, Liu X, Shen J. PD-L1 siRNA delivery for ovarian cancer immunotherapy. Adv Healthc Mater. 2015;4(8):1180–9.
20. Kang SW, Choi HJ, Cho M, Shin H, Kim YK. Transcriptome profiling of ovarian cancer xenografts. Sci Rep. 2024;14(1):11894.
21. Lee EH, Kim JY, Jeong JH, Choi JW, Kim YJ. Axl blockade in HPV16E6-mediated cervical cancer. Sci Rep. 2017;7(1):5759.
22. Mujoo K, Kipps TJ, Yang HM, Cherney B, Larrick JW. Emergence of cisplatin-resistant OVCAR-3 cells. Int J Gynecol Cancer. 2000;10(2):105–14.
23. Wang L, Xu W, Zhang L, Hu J, Xia X. Drug resistance in ovarian cancer: Mechanisms to clinical trials. Mol Cancer. 2024;23(1):66.
24. Nagasaki J, Ishino T, Togashi Y. Mechanisms of resistance to immune checkpoint inhibitors. Cancer Sci. 2022;113(10):3303–12.
25. Santoro A, Angi M, Camarda F, Dellino M, Bogani G. Role of PD-L1 in gynecological cancers. Gynecol Oncol. 2024;184:57–66.
26. Bhalla S, Gerber DE. AXL inhibitors: Status of clinical development. Curr Oncol Rep. 2023;25(5):521–9.
27. Liu Y, Li X, Zhang Y, Qian J, Yang L. AXL and the immunosuppressive microenvironment. Mol Cancer. 2025;24(1):11.
28. Yang M, Li X, Yu X, Hu W, Zhang J. Cancer immunotherapy resistance: Mechanisms and biomarkers. Cell Commun Signal. 2024;22(1):338.
29. Tanaka T, Fujiwara T, Saito Y, Kudo M, Nakamura M. Anti-PD-L1 promotes proliferation via PD-L1–AXL in liver cancer. Hepatol Int. 2024;18(3):984–97.
30. Yu W, Zhang X, Chen G, Xu R, Wang S. PD-L1 activates WIP and β-catenin in lung cancer. Cell Death Dis. 2020;11(7):506.
31. Boshuizen J, Vredevoogd DW, Krijgsman O, Ligtenberg MA, de Bruijn B, Jimenez CR, et al. AXL-ADC and ICB in melanoma and lung cancer. Cancer Res. 2021;81(7):1775–87.
32. Li C, Han X. Melanoma immunotherapy with PD-L1 siRNA and imatinib. Pharm Res. 2020;37(6):109.
33. Tian M, Wang H, Wang J, Zhang W, Chen Q. AXL inhibition sensitizes ovarian cancer to cisplatin. Acta Pharmacol Sin. 2021;42(7):1180–9.
34. Oien DB, Pathoulas CL, Ray U, Shi R, Conejo-Garcia JR, Lengyel E, et al. AXL inhibitor enhances mesothelioma cell death. Front Pharmacol. 2017;8:970.
35. Bastaki S, Bae DH, Mecham B, Richardson DR, Huang
ML. Codelivery of STAT3 and PD-L1 siRNA suppresses cancer. Life Sci. 2021;266:118847.
36. Han S, Wang H, Zhang C, Liu L, Li S. AXL inhibition suppresses esophageal carcinoma growth. Exp Ther Med. 2020;20(5):41.
37. Almozyan S, Colak D, Mansour F, Alaiya A, Ghebeh H. PD-L1 promotes stemness in breast cancer. Int J Cancer. 2017;141(7):1402–12.
38. Idowu MO, Kmieciak M, Dumur CI, Burton RS, Grimes MM, Powers CN, et al. CD44+/CD24−/low cells in triple-negative breast cancer. Hum Pathol. 2012;43(3):364–73.
39. Alwosaibai K, Zakaria Z, Yahaya BH, Abdullah H, Lim KP. PD-L1 and cancer stem cells in ovarian cancer. BMC Cancer. 2023;23(1):13.
40. Zhang J, Ding L, Tian Y, Li X, Yu S. CSC-like traits in cervical cancer and radioresistance. Gynecol Obstet Invest. 2019;84(2):174–82.
41. Woo SM, Min KJ, Kim S, Park JW, Kim DE, Kim SH, et al. AXL inhibitor R428 enhances TRAIL-induced apoptosis. Int J Mol Sci. 2019;20(13):–.
42. Li J, Xu J, Yan X, Jin K, Li W. PD-L1 knockdown in gastric cancer cells. Cell Physiol Biochem. 2017;41(3):907–20.
43. Mitra AK, Davis DA, Tomar S, Roy A, Gurler H, Xie JL, et al. In vivo growth of ovarian cancer cell lines. Gynecol Oncol. 2015;138(2):372–7.
44. Chowdhury S, Matin K, Ali M, Paul P, Rahman M. PD-L1 as a marker in thyroid cancer. Oncotarget. 2016;7(22):32318–28.
45. Ghebeh H, Tulbah A, Mohammed S, Elkum N, Bin Amer S, Alaiya A, et al. Doxorubicin modulates PD-L1 expression. Breast Cancer Res. 2010;12(4)\:R48.
46. Hua D, Zheng S, Xu G, Yuan X, Zhang B, Chen Y, et al. B7-H1 and regulatory T cells in colorectal cancer. World J Gastroenterol. 2012;18(9):971–8.
47. Satelli A, Batth IS, Brownlee Z, Mitra A, Zhou S, Rojas C, et al. Nuclear PD-L1 in circulating tumor cells. Sci Rep. 2016;6:28910.
48. Tang Y, Fang W, Zhang Y, Hong S, Kang S, Yan Y, et al. PD-L1, EGFR and prognosis in NSCLC. Oncotarget. 2015;6(16):14209–19.
49. Ukpo OC, Thorstad WL, Lewis JS Jr. B7-H1 in HPV-related oropharyngeal cancer. Head Neck Pathol. 2013;7(2):113–21.
50. Qu QX, Xie F, Huang Q, Zhang XH, Xie J, Wang XH, et al. PD-L1 expression in ovarian cancer. Cell Physiol Biochem. 2017;43(5):1893–906.
51. Parvez A, Alam M, Hamid AS, Suhail M, Ahmad A. PD-1 and PD-L1 in cancer immunotherapy. Front Immunol. 2023;14:1296341.
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| Keywords | ||
| Axl receptor tyrosine kinase Cervical cancer Molecular targeted therapies Ovarian cancer Programmed death ligand 1 siRNA | ||
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How to Cite
1.
Rahavi H, Najafi A, Asgarian-Omran H, Aflatoonian B, Tehrani M, Farashahi-Yazd E. Dual Blockade of PD-L1 and AXL: A Novel Immunotherapeutic Approach for Ovarian and Cervical Cancer. Iran J Allergy Asthma Immunol. 2025;:1-17.
