Targeting Alpha-Fetoprotein (AFP)-MHC Complex with CAR T-Cell Therapy for Liver Cancer.

PURPOSE: The majority of tumor-specific antigens are intracellular and/or secreted and therefore inaccessible by conventional chimeric antigen receptor (CAR) T-cell therapy. Given that all intracellular/secreted proteins are processed into peptides and presented by class I MHC on the surface of tumor cells, we used alpha-fetoprotein (AFP), a specific liver cancer marker, as an example to determine whether peptide-MHC complexes can be targets for CAR T-cell therapy against solid tumors. EXPERIMENTAL DESIGN: We generated a fully human chimeric antigen receptor, ET1402L1-CAR (AFP-CAR), with exquisite selectivity and specificity for the AFP158-166 peptide complexed with human leukocyte antigen (HLA)-A*02:01. RESULTS: We report that T cells expressing AFP-CAR selectively degranulated, released cytokines, and lysed liver cancer cells that were HLA-A*02:01+/AFP+ while sparing cells from multiple tissue types that were negative for either expressed proteins. In vivo, intratumoral injection of AFP-CAR T cells significantly regressed both Hep G2 and AFP158-expressing SK-HEP-1 tumors in SCID-Beige mice (n = 8 for each). Moreover, intravenous administration of AFP-CAR T cells in Hep G2 tumor-bearing NSG mice lead to rapid and profound tumor growth inhibition (n = 6). Finally, in an established intraperitoneal liver cancer xenograft model, AFP-CAR T cells showed robust antitumor activity (n = 6). CONCLUSIONS: This study demonstrates that CAR T-cell immunotherapy targeting intracellular/secreted solid tumor antigens can elicit a potent antitumor response. Our approach expands the spectrum of antigens available for redirected T-cell therapy against solid malignancies and offers a promising new avenue for liver cancer immunotherapy. Clin Cancer Res; 23(2); 478-88. ©2016 AACR.

A mouse model based on replication-competent Tiantan vaccinia expressing luciferase/HIV-1 Gag fusion protein for the evaluation of protective efficacy of HIV vaccine.

BACKGROUND: Developing an effective vaccine against human immunodeficiency virus type 1 (HIV-1) remains a grand challenge after more than two decades of intensive effort. It is partially due to the lack of suitable animal models for screening and prioritizing vaccine candidates. In this study, we aim to develop a mice model to test HIV-1 vaccine efficacy. METHODS: We constructed a recombinant vaccinia expressing firefly luciferase and HIV-1 Gag fusion protein based on Tiantan strain, an attenuated but replication-competent poxvirus (rTTV-lucgag). By quantifying the luciferase activity as its read out, we defined the biodistribution of Tiantan strain poxvirus in mice inoculated intraperitoneally and attempted to apply this model to evaluate the HIV-1 vaccine efficacy. RESULTS: Our data demonstrated that the rTTV-lucgag was able to express high level of luciferase (< or = 10(6) relative luciferase units (RLU)/mg protein) and HIV-1 Gag (> 3 folds increase comparing to the control). After intraperitoneal inoculation, this virus had dominant replication in the ovary, uterus, and cervix of mice and the luciferase activities in those organs are significantly correlated with viral titers (r(2) = 0.71, P < 0.01). Pre-immunization with an HIV gag DNA vaccine reduced the luciferase activity in ovary from (6006 +/- 3141) RLU/mg protein in control group to (1538 +/- 463) RLU/mg protein in vaccine group (P = 0.1969). CONCLUSIONS: The luciferase activity in ovary could represent viral replication in vivo; this rTTV-lucgag/mice model may be suitable to assess the protective efficacy of cytotoxic T-cell responses to HIV Gag with less tedious work and high through-put.

[Effect of GBV-C/HIV coinfection on HIV/AIDS disease progression and HIV replication].

Several research groups have recently reported that persistent GB virus C (GBV-C) co-infected with human immunodeficiency virus (HIV) leads to slower AIDSs disease progression than HIV-1 infection alone. However, these findings were not confirmed by several other studies. To investigate the association between GBV-C replication and plasma HIV loads and CD4+ T cell counts, 203 HIV-1 positive former blood/plasma donors(FBDs) were enrolled from Fuyang city of Anhui Province in China. Plasma specimens were collected from them and were tested for GBV-C using RT-PCR and ELISA. Out of 203 specimens, 52 (25.6%) cases were positive for GBV-C, including 35 male (67.3%) and 17 female (32.7%) cases. No significant association was identified between GBV-C infection and CD4+ T-cell counts or between GBV-C infection and HIV viral loads. Since all the subjects studied were naive to ART, the influence of therapy on AIDS disease progression was ruled out in this study. Overall, our data indicated that HIV-1 positive male FBDs were prone to be infected, GBV-C coinfection with HIV-1 does not significantly influence HIV/AIDS disease progression during the late stage of chronic HIV-1 infection.

[Intranasal priming with HIV DNA vaccine and systemic boosting with recombinant vaccinia induce vigorous immune responses: experiment with mice].

OBJECTIVE: To explore the strategy to raise both mucosal and systemic anti-HIV-1 immunity. METHODS: Eighteen BALB/c rats were randomly divided into 2 groups, experimental group and control group. The experimental group were further subdivided into 4 subgroups of 3 mice: 3-dose HIV DNA vaccine group, 3-dose DNA vaccine + cholera toxin (CT) adjuvant subgroup, 1-dose recombinant Tiantan strain vaccinia-based vaccine subgroup, and 3-dose DNA vaccine + CT adjuvant + Tiantan strain vaccinia-based vaccine subgroup. The control group was subdivided into 2 subgroups of 3 mice: 3-dose DNA blank vector subgroup, and 3-dose DNA blank vector + Tiantan strain vaccinia-based vaccine subgroup. Intranasal administration of DNA vaccine-based vaccine (10 microg) was done on the days 0, 14, and 28 as the mucosal priming, and recombinant Tiantan vaccinia (1 x 10(7) PFU) was injected intramuscularly as systemic boosting on the day 42. On the day 56 the mice were killed and specimens of serum, nasopharynx wash, lung wash, and spleen were collected and splenocytes were isolated. Splenocytes were added into the phosphate-buffered saline with anti-mouse interferon-gamma (IFN-gamma) envelop antibody to count the number of spot-forming cells (SFCs). Indirect ELISA was used to detect the HIV-1 specific antibody in the nasopharynx wash and lung wash. Immunohistochemistry was used to detect the intracellular staining of IFN-gamma in the splenocytes. RESULTS: The number of spot forming cells in the HIV-1 DNA vaccine + CT adjuvant group was (14 +/- 11) SFCs/10(6) splenocytes, significantly more than that of the HIV-1 DNA vaccine group [(2 +/- 1) SFCs/10(6) spleen cells (P < 0.01). The number of SFCs of the 1-dose DNA-vaccine subgroup was [(30 +/- 18) SFCs/10(6) spleen cells], significantly higher than that of the only DNS vaccine group (P < 0.01). The number of SFCs of DNA vaccine + CT adjuvant + recombinant Tiantan vaccinia-based vaccine was (61 +/- 35) SFCs/10(6) splenocytes, significantly higher than those of the other groups (all P < 0.01). Flow cytometry showed that the rate of HIV-1 Gag specific CD8(+) T cell was 1.8% +/- 1.4%. The value of specific IgG of the DNA vaccine + adjuvant + Tiantan vaccinia-based vaccine was 1.50 +/- 0.30, significantly higher than those of the blank vector, single-dose Tiantan vaccinia-based vaccine, and single-dose DNA vaccine + CT adjuvant subgroups (0.42 +/- 0.02, 0.74 +/- 0.13, and 0.75 +/- 0.02 respectively, all P < 0.05). In different subgroups the levels of specific IgA in the lung wash were all higher than those in the nasopharynx wash. The levels of specific IgA in the lung and nasopharynx wash of the DNA vaccine + CT adjuvant subgroup were higher than those of the other subgroups whether or not with boosting of Tiantan. The specific IgA levels of the groups enhanced by Tiantan vaccinia-based vaccine were all significantly higher than those of the corresponding subgroups without enhancement (all P < 0.01). The IgA level of lung wash of the DNA vaccine + CT adjuvant subgroup was 1.82 +/- 0.76, significantly higher than that of the one-dose Tiantan vaccinia-based vaccine group (0.52 +/- 0.19, P < 0.05). CONCLUSION: The vaccination modality of mucosal priming and systemic boosting induces both mucosal and systemic immune responses.