The Antitumor Effect of a Non-Transforming E7 Protein Combined with a TLR7 Agonist
Abstract
Despite great efforts in developing peptide-based therapeutic vaccines against human papillomavirus (HPV)-induced cervical cancers, they have failed to elicit strong and sustainable immune responses. Here, we evaluated the vaccine potential of an HPV16 three mutant of E7 (E7GGG) (D21G/C24G/E26G) protein combined with Aldara (topical imiquimod) adjuvant in a TC-1 mouse tumor model.
The HPV16-E7GGG, with eliminated transforming properties but retained antigenicity, and E7 wild-type were inserted into pET28, expressed in the E coli system, and purified using Ni-NTA chromatography. The E7GGG and E7 wild-type proteins were combined with Aldara adjuvant and injected into C57BL mice.
We determined the ability of HPV16-E7GGG in combination with Aldara adjuvant to induce robust immune responses by IgG total development, IL-4, IL-17, and IFN-γ induction, CTL activity, and inhibit tumor growth in the murine TC-1 model in different immunized groups.
The generated recombinant HPV16-E7GGG induced humoral and cellular immune responses in a TH1-mediated pathway, specifically with the (E7GGG) (D21G/C24G/E26G) antigen combined with Aldara, which could be a suitable therapeutic vaccine candidate against HPV.
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8. Vaz M, Fahrenkrog B. The retinoblastoma tumor suppressor is required for the NUP98-HOXA9-induced aberrant nuclear envelope phenotype. Cells. Cells. 2021;10(11):2851.
9. Chabeda A, Yanez RJR, Lamprecht R, Meyers AE, Rybicki EP, Hitzeroth II. Therapeutic vaccines for high-risk HPV-associated diseases. Papillomavirus Res. 2018;5:46-58.
10. Skeate JG, Woodham AW, Einstein MH, Da Silva DM, Kast WM. Current therapeutic vaccination and immunotherapy strategies for HPV-related diseases. Hum Vaccin Immunother. 2016;12(6):1418-29.
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12. Tranoulis A, Laios A, Mitsopoulos V, Lutchman-Singh K, Thomakos N. Efficacy of 5% imiquimod for the treatment of Vaginal intraepithelial neoplasia—A systematic review of the literature and a meta-analysis. Eur J Obstet Gynecol Reprod Biol. 2017;218:129-36.
13. Trutnovsky G, Reich O, Joura EA, Holter M, Ciresa-König A, Widschwendter A, et al. Topical imiquimod versus surgery for vulvar intraepithelial neoplasia: a multicentre, randomised, phase 3, non-inferiority trial. Lancet. 2022;399(10337):1790-8.
14. Tambunlertchai S, Geary SM, Salem AK. Topically applied resiquimod versus imiquimod as a potential adjuvant in melanoma treatment. Pharmaceutics. 2022;14(10):2076.
15. Yu H, Cen J, Lin X, Cheng H, Seifert O. Imiquimod induced vitiligo‐like lesions-A consequence of modified melanocyte function. Immun Inflamm Dis. 2022;10(1):70-7.
16. Avcilar H, Eken A. Could imiquimod (Aldara 5% cream) or other TLR7 agonists be used in the treatment of COVID-19? Med Hypotheses. 2020;144:110202.
17. Yuan J, Ni G, Wang T, Mounsey K, Cavezza S, Pan X, Liu X. Genital warts treatment: beyond imiquimod. Hum Vaccin Immunother. 2018;14(7):1815-9.
18. Hasche D, Akgül B. Prevention and Treatment of HPV-Induced Skin Tumors. Cancers (Basel). 2023;15(6):1709.
19. Jansen MHE, Kessels JPHM, Nelemans PJ, Kouloubis N, Arits AHMM, van Pelt HPA, et al. Randomized trial of four treatment approaches for actinic keratosis. N Engl J Med. 2019;380(10):935-46.
20. T Tio DCKS, van Montfrans C, Ruijter CGH, Hoekzema R, Bekkenk MW. Effectiveness of 5% topical imiquimod for lentigo maligna treatment. Acta Derm. Acta Derm Venereol. 2019 Sep 1;99(10):884-8.
21. Hameed HI, Al-Mayahy MH. Insight into imiquimod skin permeation and increased delivery using microneedle pre-treatment. Eur. J. Pharm. Biopharm. 2019;139: 33–43.
22. Demurtas OC, Massa S, Ferrante P, Venuti A, Franconi R, Giuliano G. A Chlamydomonas-derived Human Papillomavirus 16 E7 vaccine induces specific tumor protection. PLoS One. 2013;8(4):e61473.
23. Francis DM, Page R. Strategies to optimize protein expression in E. Curr Protoc Protein Sci. 2010;5(1):5.24.1-5.24.9.
24. Spriestersbach A, Kubicek J, Schäfer F, Block H, Maertens, B. Purification of his-tagged proteins. In: Methods in enzymology.2015;559:1–15.
25. Kielkopf CL, Bauer W, Urbatsch IL. Bradford assay for determining protein concentration. Cold Spring Harb Protoc. 2020;2020(4):102269.
26. Basukala O, Trejo-Cerro O, Myers MP, Pim D, Massimi P, Thomas M, et al. HPV-16 E7 Interacts with the Endocytic Machinery via the AP2 Adaptor μ2 Subunit. mBio. 2022;13(6):e0230222.
27. Mashhadi Abolghasem Shirazi, M, Roohvand F, Arashkia A. Preparation and in vivo anti-tumor evaluation of human papillomavirus E7 adjuvanted with Montanide ISA 266 as a vaccine candidate. Vaccine Res. 217;4, 29–33.
28. Chen OM, Kim K, Steele C, Wilmas KM, Aboul-Fettouh N, Burns C, et al. Advances in management and therapeutics of cutaneous basal cell carcinoma. Cancers (Basel). 2022;14(15):3720.
29. Franconi R, Massa S, Paolini F, Vici P, Venuti A. Plant-derived natural compounds in genetic vaccination and therapy for HPV-associated cancers. Cancers (Basel). 2020;12(11):3101.
30. Sciullo PD, Menay F, Cocozza F, Gravisaco MJ, Waldner CI, Mongini C. Systemic administration of imiquimod as an adjuvant improves immunogenicity of a tumor-lysate vaccine inducing the rejection of a highly aggressive T-cell lymphoma. Clin Immunol. 2019;203:154-61.
31. Kokcu A, Alper T, Ogur G, Akar OS, Kurtoglu Ozdes E, Altunkaynak BZ... Imiquimod Inhibits Proliferation of Serous Epithelial Ovarian Cancer Cells In Vitro: A Preliminary Study. Indian J Gynecol Oncol. 2020;18:1–7.
32. Agerholm R, Bekiaris V. Evolved to protect, designed to destroy: IL‐17‐producing γδ T cells in infection, inflammation, and cancer. Eur J Immunol. 2021;51(9):2164-2177
33. Farooq M, Batool M, Kim MS, Choi S. Toll-like receptors as a therapeutic target in the era of immunotherapies. Front Cell Dev Biol. 2021;9:756315.
34. Walter A, Schäfer M, Cecconi V, Matter C, Urosevic-Maiwald M, Belloni B, et al. Aldara activates TLR7-independent immune defence. Nat Commun. 2013;4:1560.
35. Wenzel J, Uerlich M, Haller O, Bieber T, Tueting T. Enhanced type I interferon signaling and recruitment of chemokine receptor CXCR3‐expressing lymphocytes into the skin following treatment with the TLR7‐agonist imiquimod. J Cutan Pathol. 2005;32(4):257-62.
36. Lopez PA, Denny M, Hartmann AK, Alflen A, Probst HC, von Stebut E, et al. Transcutaneous immunization with a novel imiquimod nanoemulsion induces superior T cell responses and virus protection. J Dermatol Sci. 2017;87(3):252-9.
37. Dong Z, Hu R, Du Y, Tan L, Li L, Du J, et al. Immunodiagnosis and immunotherapeutics based on human papillomavirus for HPV-induced cancers. Front Immunol. 2021;11:586796.
2. Yan F, Cowell LG, Tomkies A, Day AT. Therapeutic vaccination for HPV-mediated cancers. Curr Otorhinolaryngol Rep. 2023;11(1):44-61.
3. Akhatova A, Azizan A, Atageldiyeva K, Ashimkhanova A, Marat A, Iztleuov Y, et al. Prophylactic Human Papillomavirus Vaccination: From the Origin to the Current State. Vaccines (Basel). 2022;10(11):1912.
4. Garbuglia AR, Lapa D, Sias C, Capobianchi MR, Del Porto P. The Use of Both Therapeutic and Prophylactic Vaccines in the Therapy of Papillomavirus Disease. Front Immunol. 2020;11:188.
5. Yousefi Z, Aria H, Ghaedrahmati F, Bakhtiari T, Azizi M, Bastan R, Hosseini R, et al. An update on human papilloma virus vaccines: history, types, protection, and efficacy. Front Immunol. 2022;12:805695.
6. Rodriguez Rodriguez N, Alfonso Chaviano AB, Granadillo Rodríguez M, Batte Figueredo A, Torrens I. Obtaining a Recombinant Protein Based on the E7 Antigen of Human Papillomavirus Type 18. J. Cancer Sci Clin Ther. 2022;6:336-43.
7. Todorovic B, Hung K, Massimi P, Avvakumov N, Dick
FA, Shaw GS, et al. Conserved region 3 of human papillomavirus 16 E7 contributes to deregulation of the retinoblastoma tumor suppressor. J Virol. 2012;86;13313-23.
8. Vaz M, Fahrenkrog B. The retinoblastoma tumor suppressor is required for the NUP98-HOXA9-induced aberrant nuclear envelope phenotype. Cells. Cells. 2021;10(11):2851.
9. Chabeda A, Yanez RJR, Lamprecht R, Meyers AE, Rybicki EP, Hitzeroth II. Therapeutic vaccines for high-risk HPV-associated diseases. Papillomavirus Res. 2018;5:46-58.
10. Skeate JG, Woodham AW, Einstein MH, Da Silva DM, Kast WM. Current therapeutic vaccination and immunotherapy strategies for HPV-related diseases. Hum Vaccin Immunother. 2016;12(6):1418-29.
11. Liu D, Liu L, Li X, Wang S, Wu G, Che X. Advancements and challenges in peptide-based cancer vaccination: a multidisciplinary perspective. Vaccines. 2024;12(8):950.
12. Tranoulis A, Laios A, Mitsopoulos V, Lutchman-Singh K, Thomakos N. Efficacy of 5% imiquimod for the treatment of Vaginal intraepithelial neoplasia—A systematic review of the literature and a meta-analysis. Eur J Obstet Gynecol Reprod Biol. 2017;218:129-36.
13. Trutnovsky G, Reich O, Joura EA, Holter M, Ciresa-König A, Widschwendter A, et al. Topical imiquimod versus surgery for vulvar intraepithelial neoplasia: a multicentre, randomised, phase 3, non-inferiority trial. Lancet. 2022;399(10337):1790-8.
14. Tambunlertchai S, Geary SM, Salem AK. Topically applied resiquimod versus imiquimod as a potential adjuvant in melanoma treatment. Pharmaceutics. 2022;14(10):2076.
15. Yu H, Cen J, Lin X, Cheng H, Seifert O. Imiquimod induced vitiligo‐like lesions-A consequence of modified melanocyte function. Immun Inflamm Dis. 2022;10(1):70-7.
16. Avcilar H, Eken A. Could imiquimod (Aldara 5% cream) or other TLR7 agonists be used in the treatment of COVID-19? Med Hypotheses. 2020;144:110202.
17. Yuan J, Ni G, Wang T, Mounsey K, Cavezza S, Pan X, Liu X. Genital warts treatment: beyond imiquimod. Hum Vaccin Immunother. 2018;14(7):1815-9.
18. Hasche D, Akgül B. Prevention and Treatment of HPV-Induced Skin Tumors. Cancers (Basel). 2023;15(6):1709.
19. Jansen MHE, Kessels JPHM, Nelemans PJ, Kouloubis N, Arits AHMM, van Pelt HPA, et al. Randomized trial of four treatment approaches for actinic keratosis. N Engl J Med. 2019;380(10):935-46.
20. T Tio DCKS, van Montfrans C, Ruijter CGH, Hoekzema R, Bekkenk MW. Effectiveness of 5% topical imiquimod for lentigo maligna treatment. Acta Derm. Acta Derm Venereol. 2019 Sep 1;99(10):884-8.
21. Hameed HI, Al-Mayahy MH. Insight into imiquimod skin permeation and increased delivery using microneedle pre-treatment. Eur. J. Pharm. Biopharm. 2019;139: 33–43.
22. Demurtas OC, Massa S, Ferrante P, Venuti A, Franconi R, Giuliano G. A Chlamydomonas-derived Human Papillomavirus 16 E7 vaccine induces specific tumor protection. PLoS One. 2013;8(4):e61473.
23. Francis DM, Page R. Strategies to optimize protein expression in E. Curr Protoc Protein Sci. 2010;5(1):5.24.1-5.24.9.
24. Spriestersbach A, Kubicek J, Schäfer F, Block H, Maertens, B. Purification of his-tagged proteins. In: Methods in enzymology.2015;559:1–15.
25. Kielkopf CL, Bauer W, Urbatsch IL. Bradford assay for determining protein concentration. Cold Spring Harb Protoc. 2020;2020(4):102269.
26. Basukala O, Trejo-Cerro O, Myers MP, Pim D, Massimi P, Thomas M, et al. HPV-16 E7 Interacts with the Endocytic Machinery via the AP2 Adaptor μ2 Subunit. mBio. 2022;13(6):e0230222.
27. Mashhadi Abolghasem Shirazi, M, Roohvand F, Arashkia A. Preparation and in vivo anti-tumor evaluation of human papillomavirus E7 adjuvanted with Montanide ISA 266 as a vaccine candidate. Vaccine Res. 217;4, 29–33.
28. Chen OM, Kim K, Steele C, Wilmas KM, Aboul-Fettouh N, Burns C, et al. Advances in management and therapeutics of cutaneous basal cell carcinoma. Cancers (Basel). 2022;14(15):3720.
29. Franconi R, Massa S, Paolini F, Vici P, Venuti A. Plant-derived natural compounds in genetic vaccination and therapy for HPV-associated cancers. Cancers (Basel). 2020;12(11):3101.
30. Sciullo PD, Menay F, Cocozza F, Gravisaco MJ, Waldner CI, Mongini C. Systemic administration of imiquimod as an adjuvant improves immunogenicity of a tumor-lysate vaccine inducing the rejection of a highly aggressive T-cell lymphoma. Clin Immunol. 2019;203:154-61.
31. Kokcu A, Alper T, Ogur G, Akar OS, Kurtoglu Ozdes E, Altunkaynak BZ... Imiquimod Inhibits Proliferation of Serous Epithelial Ovarian Cancer Cells In Vitro: A Preliminary Study. Indian J Gynecol Oncol. 2020;18:1–7.
32. Agerholm R, Bekiaris V. Evolved to protect, designed to destroy: IL‐17‐producing γδ T cells in infection, inflammation, and cancer. Eur J Immunol. 2021;51(9):2164-2177
33. Farooq M, Batool M, Kim MS, Choi S. Toll-like receptors as a therapeutic target in the era of immunotherapies. Front Cell Dev Biol. 2021;9:756315.
34. Walter A, Schäfer M, Cecconi V, Matter C, Urosevic-Maiwald M, Belloni B, et al. Aldara activates TLR7-independent immune defence. Nat Commun. 2013;4:1560.
35. Wenzel J, Uerlich M, Haller O, Bieber T, Tueting T. Enhanced type I interferon signaling and recruitment of chemokine receptor CXCR3‐expressing lymphocytes into the skin following treatment with the TLR7‐agonist imiquimod. J Cutan Pathol. 2005;32(4):257-62.
36. Lopez PA, Denny M, Hartmann AK, Alflen A, Probst HC, von Stebut E, et al. Transcutaneous immunization with a novel imiquimod nanoemulsion induces superior T cell responses and virus protection. J Dermatol Sci. 2017;87(3):252-9.
37. Dong Z, Hu R, Du Y, Tan L, Li L, Du J, et al. Immunodiagnosis and immunotherapeutics based on human papillomavirus for HPV-induced cancers. Front Immunol. 2021;11:586796.
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| Issue | Articles in Press | |
| Section | Original Article(s) | |
| Keywords | ||
| Epitopes Human papillomavirus 16 Imiquimod Uterine cervical neoplasms Vaccines | ||
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How to Cite
1.
Mashhadi Abolghasem Shirazi M, Sadat SM, Hashemi Goradel N, Arashkia A. The Antitumor Effect of a Non-Transforming E7 Protein Combined with a TLR7 Agonist. Iran J Allergy Asthma Immunol. 2025;:1-10.
