Cell-based Hyper-interleukin 6 or Hyper-interleukin 11 secreting vaccines combined with low dose cyclophosphamide in an orthotopic murine prostate cancer model

Abstract

Background: Cell based vaccines encoding Hyper-IL-6 (H6) and Hyper-IL-11 (H11) present high activity in murine melanoma and renal cancer model. We evaluated the efficacy of cellular vaccines modified with H6 or H11 combined with cyclophosphamide in orthotopic murine prostate cancer model.

Material and methods: TRAMP cells were transduced with H6 and H11 cDNA (TRAMP-H6 and TRAMP-H11). An orthotopic TRAMP model based on the implantation of TRAMP cells into the dorsolateral lobe of the prostate of C57BL6/J mice was employed. The efficacy of TRAMP-H6 and TRAMP-H11 vaccines evaluated in the therapeutic setting was compared with the TRAMP cells modified with a mock transduced E1-deleted adenoviral vector (TRAMP-AdV) and non-modified irradiated TRAMP cells (TRAMP IRR) in relation to naive (non-immunized) mice. In the next experimental groups mice vaccinated with TRAMP-H6 and TRAMP-H11 received cyclophosphamide (CY). Detection of immune cells in the spleen in mice receiving vaccines combined with CY was evaluated.

Results: Modification of TRAMP cells with H6 increased the efficacy of TRAMP-based whole-cell vaccine. The highest response rate was observed in mice receiving TRAMP-H6 alone and combined with CY. Vaccination with TRAMP-H6 alone and combined with CY and TRAMP H11 combined with CY extended median OS of mice bearing orthotopic TRAMP tumors in therapeutic setting. Low dose CY administered alone demonstrated some antitumor activity in employed model. TRAMP-H6 or TRAMP-H11 combined with CY strongly augmented generation of CD8+, CD4+ T lymphocytes and memory T cells. Immunization with TRAMP combined with or without CY suppressed generation of T regulatory cells.

Conslusions: Prostate cancer vaccines modified with H6 or H11 induce prostate tumour regression and increase mice survival by stimulating the immune system. Cyclophosphamide added to modified TRAMP vaccines demonstrated clinical benefit of treated mice and enhanced anti-tumour immune response.

Keywords: cancer vaccine; cyclophosphamide; immunotherapy; murine model; prostate cancer.

Fig. 1

Overall survival of mice treated in the orthotopic prostate cancer model. Mice were inoculated with TRAMP-WT cells into the prostate. Subsequently, the animals were vaccinated with TRAMP-H6 with or without cyclophosphamide, TRAMP-H11 with or without cyclophosphamide, TRAMP-AdV or TRAMP IRR. An additional group received cyclophosphamide alone. Control mice did not receive any treatment (untreated). A) Treatment with TRAMP-H6 elongated median OS compared to untreated controls (p = 0.01). There was a trend towards elongated median OS between TRAMP-H6 compared to TRAMP-H11, TRAMP IRR or TRAMP-AdV, but the differences were not statistically significant. Administration of CY alone was linked with longer survival compared to TRAMP-IRR, TRAMP-AdV, TRAMP-H11, and untreated control although the differences were not statistically significant. B) Mice receiving TRAMP-H6 + CY demonstrated the longest median OS. Combining TRAMP-H6 with CY was linked with elongation of median OS compared to TRAMP-H6 alone and CY alone, but the difference was not significant (p = 0.15 and p = 0.34, respectively). The median OS in the TRAMP-H6 + CY group was significantly longer compared to control mice (p = 0.01) (not shown in graph). C) Mice treated with TRAMP H11 + CY presented longer median OS compared to untreated controls (p = 0.03). Addition of CY to TRAMP-H11 compared to TRAMP-H11 elongated median OS of the treated mice but the difference was not statistically significant (p = 0.32)

Fig. 2

Response assessment in murine prostate cancer model treated in the therapeutic setting. Twenty-four hours after inoculation of TRAMP-WT cells into the prostate mice were vaccinated with TRAMP-H6, TRAMP-H11, TRAMP-AdV or TRAMP IRR. Additionally, two groups received TRAMP-H6 and TRAMP-H11 combined with CY. One group was treated with CY alone. Control mice did not receive any treatment (untreated). Twelve weeks after the beginning of vaccination mice were sacrificed and tumor volume was assessed. The lowest mean tumor volume was observed in mice receiving TRAMP-H6, TRAMP-H6 + CY, TRAMP-H11, TRAMP-H11 + CY and CY alone

Fig. 3

Influence of repeated immunization (six times with 4-day interval) with TRAMP-H6, TRAMP-H11, TRAMP IRR with or without CY (administered i.p. 24 h before each vaccination) on memory T lymphocytes: A) CD8+ CD62L^low, B) CD4+ CD62L^low. An additional group received CY alone. Control mice were not vaccinated and did not receive CY. Ten days after the last vaccination mice were sacrificed and spleens were excised, minced, pooled (five mice per group), stained and analyzed by flow cytometry

Fig. 4

Flow cytometric analyses of T regulatory lymphocytes (CD4 + CD25 + FoxP3 + ) assessed in the spleen after vaccination with TRAMP-H6, TRAMP-H11, TRAMP IRR with or without CY (administered i.p. 24 h before each vaccination). An additional group received CY alone. Control mice were not vaccinated and did not receive CY