Sewell Duane A, Shahabi Vafa, Gunn George R, Pan Zhen-Kun, Dominiecki Mary E, Paterson Yvonne
Department of Microbiology and the University of Pennsylvania Cancer Center, Philadelphia, Pennsylvania, USA.
Cancer Res. 2004 Dec 15;64(24):8821-5. doi: 10.1158/0008-5472.CAN-04-1958.
Previous work in our laboratory has established that the fusion of tumor-associated antigens to a truncated form of the Listeria monocytogenes virulence factor listeriolysin O (LLO) enhances the immunogenicity and antitumor efficacy of the tumor antigen when delivered by Listeria or by vaccinia. LLO contains a PEST sequence at the NH(2) terminus. These sequences, which are found in eukaryotic proteins with a short cellular half-life, target proteins for degradation in the ubiquitin-proteosome pathway. To investigate whether the enhanced immunogenicity conferred by LLO is due to the PEST sequence, we constructed new Listeria recombinants that expressed the HPV-16 E7 antigen fused to LLO, which either contained or had been deleted of this sequence. We then compared the antitumor efficacy of this set of vectors and found that Listeria expressing the fusion protein LLO-E7 or PEST-E7 were effective at regressing established macroscopic HPV-16 immortalized tumors in syngeneic mice. In contrast, Listeria recombinants expressing E7 alone or E7 fused to LLO from which the PEST sequence had been genetically removed could only slow tumor growth. Because CD8(+) T cell epitopes are generated in the ubiquitin-proteosome pathway, we also investigated the ability of the vaccines to induce E7-specific CD8(+) T cells in the spleen and to generate E7-specific tumor-infiltrating lymphocytes. A strong correlation was observed between CD8(+) T-cell induction and tumor homing and the antitumor efficacy of the Listeria-E7 vaccines. These findings suggest a strategy for the augmentation of tumor antigen-based immunotherapeutic strategies that may be broadly applicable.
我们实验室之前的工作已经证实，将肿瘤相关抗原与单核细胞增生李斯特菌毒力因子溶血素O（LLO）的截短形式融合，当通过李斯特菌或痘苗病毒递送时，可增强肿瘤抗原的免疫原性和抗肿瘤功效。LLO在NH₂末端含有一个PEST序列。这些序列存在于细胞半衰期较短的真核蛋白中，可将蛋白质靶向泛素-蛋白酶体途径进行降解。为了研究LLO赋予的增强免疫原性是否归因于PEST序列，我们构建了新的李斯特菌重组体，其表达与LLO融合的HPV-16 E7抗原，该LLO要么含有此序列，要么已被删除。然后我们比较了这组载体的抗肿瘤功效，发现表达融合蛋白LLO-E7或PEST-E7的李斯特菌能有效消退同基因小鼠中已形成的宏观HPV-16永生化肿瘤。相比之下，单独表达E7或表达与已通过基因手段去除PEST序列的LLO融合的E7的李斯特菌重组体只能减缓肿瘤生长。由于CD8⁺ T细胞表位是在泛素-蛋白酶体途径中产生的，我们还研究了这些疫苗在脾脏中诱导E7特异性CD8⁺ T细胞以及产生E7特异性肿瘤浸润淋巴细胞的能力。在CD8⁺ T细胞诱导、肿瘤归巢与李斯特菌-E7疫苗的抗肿瘤功效之间观察到了强烈的相关性。这些发现提示了一种增强基于肿瘤抗原的免疫治疗策略的方法，该方法可能具有广泛的适用性。
