Superior therapeutic efficacy of alphavirus-mediated immunization against human papilloma virus type 16 antigens in a murine tumour model: effects of the route of immunization.

In our efforts to develop a strong, effective immune response against cervical carcinoma and premalignant disease, we study the use of recombinant Semliki Forest virus (SFV) encoding the oncoproteins E6 and E7 from high-risk human papilloma viruses (HPVs). Optimal immunization conditions are required for immunotherapeutic treatment of cervical cancer as it has been postulated that cervical cancer patients are immune-suppressed and/or immunologically tolerant for HPV. We previously generated an optimized construct encoding a fusion protein of HPV16 E6 and E7 and a translational enhancer (enhE6,7). Immunization of mice with SFV-enhE6,7 was shown to induce cytoxic T cell (CTL) responses and resulted in the eradication of established tumours. We now demonstrate, using HPV16-specific MHC class I tetramers, that high pCTL frequencies can be induced. However, this induction is strongly influenced by the route of immunization applied. Whilst in bulk CTL assays, requiring in vitro restimulation, CTL activity can be observed upon s.c., i.p., i.v. and i.m. immunization, detectable pCTL frequencies, without in vitro restimulation, are only induced upon i.m. and i.v. immunization. The route of immunization also strongly influences the dose of viral vector needed to induce CTLs and tumour therapy. As few as 5x10(4) SFV-enhE6,7, primed and boosted i.v., are needed to eradicate tumours in six out of seven mice treated. Furthermore, exponentially growing tumours of approximately 500 mm3 in size were seen to completely resolve and even tumours as large as 1500 mm3 decreased to one-third of their size. Apart from this potency, SFV vectors can safely be used for the expression of oncoproteins such as E6 and E7, since the viral RNA is not integrated in the host genome. Thus SFV-enhE6,7 meets with the criteria that a vaccine against cervical cancer should be safe and induce a very strong, long-lasting CTL response, strong enough to eradicate existing tumours.