Placenta-derived gp96 as a multivalent prophylactic cancer vaccine

Placenta-derived gp96 as a multivalent prophylactic cancer vaccine SUBJECT AREAS: CANCER PREVENTION ADJUVANTS PROTEIN VACCINES ANTIGEN PRESENTATION Received 8 January 2013 Accepted 22 May 2013 Published 6 June 2013 Correspondence and Bao Zhao1,2*, Yanzhong Wang1*, Bo Wu1, Shan Liu3, Erjie Wu1, HongXia Fan1, MingMing Gui1, Lizhao Chen1, Changfei Li1, Ying Ju1, Wei Zhang3 & Songdong Meng1,2 1 CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences (CAS), Beijing, China, 2School of Life Sciences, University of Science and Technology of China, Hefei, China, 3Detection Center of Tumor Biology, Cancer Institute & Hospital, Chinese Academy of Medical Sciences, Beijing, China. A major challenge for designing prophylactic cancer vaccines is to define immunogenic and safe cancer antigens. Given the striking similarity of antigen expression patterns between cancer and embryonic tissues, we defined a prototype strategy of using placenta-derived heat shock protein gp96, which induces prophylactic anti-tumor T cell responses. Immunization with placental gp96 provided partial protection and long-term (at least 3 months) anti-tumor immunity against growth of transplantable melanoma or breast tumors in mice, elicited total protection against 7, 12-dimethylbenz(a)-anthracene (DMBA)-induced mammary tumors in rats, and significantly reduced the occurrence and growth of autochthonous breast tumors in HER2 transgenic mice. Placental gp96 activated HER2- and MUC1-specific T cell responses through binding to tumor-associated antigens. Our results reveal the novel immunogenicity of placental gp96 and its potential use as a multivalent cancer vaccine. requests for materials should be addressed to S.D.M. (mengsd@im. ac.cn) * These authors contributed equally to this work. D espite the remarkable progress in understanding the causes of cancer and the significant advances in cancer therapy in recent years, the disease persists, and the incidence of cancer is increasing worldwide1. Unlike the highly efficient prophylactic vaccines against infectious diseases, therapeutic vaccines against cancer that do not cause unacceptable autoimmune disorders have not been as effective, eliciting only incremental therapeutic effects2. The simplest explanation may be that using therapeutic vaccines to treat established tumors is equivalent to the unsuccessful approach of using hepatitis B virus (HBV) or human papilloma virus (HPV) vaccines to treat chronic HBV or HPV infection. Several count-back mechanisms have been reported for the involvement of tumor escape and immune suppression, likely driven by long-term tumor development, establishment, and growth, including impaired T cell responses, immune tolerance, and the suppressive tumor microenviroment, which likely act synergistically3. This underlines the need to develop a prophylactic vaccine approach for cancer that will provide low-cost and highly efficiency rationales. A major challenge for designing prophylactic cancer vaccines is to define immunogenic and safe cancer antigens that can serve as targets for effective vaccines, including tumor-specific antigens and proteins overexpressed on the tumor but not on normal tissues. At present, only a limited number of cancer antigens have been found with few successes4–6. It is well documented that most solid tumor types express embryonic antigens to varying extents, and there is striking similarity of antigen expression between cancer and embryonic tissues, which provides the potential to target embryonic components as an effective strategy to prevent the appearance of cancers7. Indeed, vaccination with embryonic or stem cell antigens leads to a potent protective immune response against cancers8–10. As a temporary organ that perform nutrient and waste product exchanges between the mother and fetus, the placenta also displays higher cancer-associated gene expression, including IGF2, HIF-2a, GPC3, pregnancy-associated plasma protein A (PAPP-A), and MUC111. As a member of the heat shock protein (HSP) 90 family, gp96 has the unique ability to associate with antigenic peptides, presents these loaded antigens to both MHC class I and class II molecules, and activates specific T cells12,13. Our previous studies show that in hepatitis B virus (HBV)-infected liver cancer, gp96 binds virus-derived peptides and activates specific CTL responses by antigen presentation14,15. Moreover, recent studies provide compelling evidence of macrophages and dendritic cells activation by gp96 through interaction with a subset of Toll-like receptors (TLRs) or CD9116–19. Clinical trials using autologous gp96-peptide complexes as therapeutic vaccines have been initiated for treatment of a range of tumors with modest antitumor effects20. Based on the observations above, the aim of this study was to investigate whether placenta-derived gp96 (P-gp96) induces prophylactic anti-tumor T cell responses. SCIENTIFIC REPORTS | 3 : 1947 | DOI: 10.1038/srep01947 1 www.nature.com/scientificreports Results We first tested the ability of a placenta-derived gp96 vaccine to prevent tumors using tumor challenge assays. Gp96 protein was extracted from the placenta or liver tissues of C57BL/6 mice as previously described21. C57BL/6 mice were subcutaneously immunized three times with placenta-derived gp96 (P-gp96), gp96derived from liver (L-gp96), or PBS (no immunization) as a control. One week after the last immunization, mice were subcutaneously challenged with B16-F10 melanoma cells (53104 cells/ mouse). Compared to L-gp96 or PBS, immunization with P-gp96 significantly inhibited tumor growth, decreasing tumor volume by approximately 49 or 45% at day 30 (both P , 0.01) (Fig. 1a). Pgp96 immunization also dramatically enhanced the survival of tumor burdened mice through 50 days of observation (Fig. 1b). All PBS- or L-gp96-treated mice died within 40 days, whereas half of the P-gp96-treated mice survived at day 50. We further examined if P-gp96 could initiate a B16-specific T cell response. As seen in Fig. 1c, ELISPOT assay revealed that P-gp96 immunization resulted in ,1-fold increase of tumor-specific T cells compared to L-gp96 immunization. Additionally, as assessed by the killing assay using B16 cells as target cells, P-gp96 effectively elicited CTL with higher cytotoxicity than L-gp96 (P , 0.01) (Fig. 1d). Figure 1 | Immunization with the P-gp96 vaccine induces antitumor T cell responses. Female C57BL/6 mice (a–d) or BALB/c mice (e–h) were immunized three times with P-gp96, L-gp96, or PBS. One week after the third immunization, the mice were subcutaneously challenged with 53104 B16F10 cells or 63105 TUBO cells. (a, e) Tumor burden was measured at 2-day intervals. (b, f) Kaplan-Meier plot of mouse survival. (c, g) Splenocytes from immunized mice were stimulated with B16-F10 (c) or TUBO (g) whole cell lysates antigens or BSA for background evaluation and assayed by IFN-c ELISPOT. (d, h (...truncated)