Carboplatin/paclitaxel, E7-vaccination and intravaginal CpG as tri-therapy towards efficient regression of genital HPV16 tumors

Abstract

High-risk human papillomavirus (HPV) are responsible for genital and oral cancers associated with the expression of the E6/E7 HPV oncogenes. Therapeutic vaccines targeting those oncogenes can only partially control tumor progression, highlighting the necessity to investigate different treatment strategies. Using the genital orthotopic HPV16 TC-1 model, herein we sequentially investigated in progressively more stringent settings the effects of systemic administration of carboplatin/paclitaxel (C + P) chemotherapy combined with HPV16-E7 synthetic long peptide (E7LP) vaccination, followed by intravaginal immunostimulation with the synthetic toll-like-receptor-9 agonist CpG. Our data show that systemic delivery of C + P prior to E7LP vaccination significantly increased mice survival. This survival benefit was associated with both reduced genital tumor growth at the time of vaccination, and a decreased infiltration of Ly6G myeloid cells and tumor-associated macrophages. Adding intravaginal CpG, which results in increased E7-specific CD8 T cells locally, to E7LP vaccination and the chemotherapy formed a tri-therapy, which significantly increased mice survival as compared to any of the dual treatments. When the tri-therapy was further refined by using a recently optimized nanoparticle-conjugated E7LP vaccine, even larger end-stage genital-TC-1 tumors responded, with 90% of mice showing a survival benefit as compared to 30% of mice with the tri-therapy involving the traditional E7LP 'liquid' vaccine. C + P is commonly used to treat cervical cancer patients and its combination with E7/E6 vaccination is currently being tested in a phase I/II trial (NCT02128126). Our data suggests that new vaccine formulations combined with local immunostimulation and standard-of-care chemotherapy have promise to further benefit patients with HPV-associated cancer.

Keywords: Carboplatin/paclitaxel; E7 long peptide and nanoparticle vaccination; HPV genital cancer; Intravaginal CpG immunostimulation.

Conflict of interest statement

Ethics approval

All animal experiments were performed in accordance with Swiss law and with approval of the Cantonal Veterinary Office of Canton de Vaud, Switzerland.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

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Figures

Fig. 1

Effect of C + P on tumor size and E7LP vaccination. Groups of female mice bearing TC-1 genital tumors received i.p. 1 dose of Carboplatin (40 mg/kg) at day 8 and 2 doses of paclitaxel (20 mg/kg at days 8/9 and/or s.c. vaccination with E7 vaccine (E7LP) at day 13. Mice survival (a) and tumor luminescence before vaccination (b) are shown. C: Flow-cytometry analysis of TC-1 genital tumors untreated or treated with C + P as indicated on the graphs is shown at day 13. Individual and mean % among CD45+ of myeloid cells (CD45+CD11b+), gMDSC (CD45+CD11b+Ly6G+), mMDSC (CD45+CD11b+Ly6C+) and macrophages (CD45+CD11b+Ly6G−Ly6C−F4/80+) are shown. Significant differences are shown following a log rank test (a) or a Student t-test (b-c): * = p < 0.05, ** = p < 0.01, *** = p < 0.001, **** = p < 0.0001

Fig. 2

Intravaginal (ivag) immunostimulation with CpG after C + P and E7LP vaccination resulted in an effective tri-therapy. Groups of female mice bearing TC-1 genital tumors received different treatments as indicated on the graphs. C + P (days 8/9) and/or vaccination with E7LP (day 15) and /or ivag administration of CpG (at days 20, 23 and 26). Tumor luminescence before vaccination or before ivag CpG (a), percentages of tumor free mice before ivag CpG (b) and after ivag CpG (days 26–29) (c) and mice survival (d) are shown. Total number of mice/group (n) are indicated (b-c). Significant differences following one-way Anova +Tukey’s post-test (a), contingency Chi-square test (b-c) or log rank test (d) are shown: * = p < 0.05, ** = p < 0.01, *** = p < 0.001, **** = p < 0.0001

Fig. 3

Including the nanoparticle vaccine (NP-E7LP) in the tri-therapy can efficiently regress very large TC-1 genital tumors. Groups of female mice bearing TC-1 genital tumors received C + P (days 8/9) and vaccination with E7LP or with NP-E7LP (at day 18) followed by ivag administration of CpG at days 23, 26 and 29. Tumor luminescence before vaccination (a), E7-specific T cells responses in PBMC one week after vaccination (b) and mice survival (c) are shown. Significant differences following a student t-test (b) or a log rank test (c) are shown: * = p < 0.05, ** = p < 0.01