Evaluating VHH-E7 Immunotherapy for HPV Tumors

To study HPV-induced tumors, age-matched, 6-week-old female C57BL/6J mice were inoculated with 3×10⁵ C3.43 cells. When tumors were palpable (~14 days later; see average tumor growth in Supplemental Figure 5), mice were treated intraperitoneally with VHH_CD11b-E7 plus adjuvant or adjuvant only. The adjuvant was 50 μg polyinosinic-polycytidylic acid (Poly(I:C) (Sigma Aldrich) and 50 μg anti-mouse CD40 (clone 1C10; Southern Biotech) as used previously.^{22 (link)} For mice inoculated with both C3.43 and B16 cells, C3.43 cells were injected 6 days prior to inoculation with 1×10⁵ B16 melanoma cells as B16 cells grow faster in vivo. Mice were treated with VHH_CD11b-E7 plus adjuvant or adjuvant only at ~14 days post C3.43 challenge (8 days post B16 challenge). Seven days later, mice were given the ⁶⁴Cu-labeled HPV E7 synTac by retroorbital injection and imaged by PET the following day (8 days post treatment). Following imaging, mice were euthanized and splenocytes and tumor cells were harvested, enumerated, and used for ELISpot analyses where indicated.