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Effect of canarypox virus (ALVAC)-mediated cytokine expression on murine prostate tumor growth
M Kawakita1, G S Rao, J K Ritchey
1Division of Urologic Surgery, Washington University School of Medicine, St. Louis, MO, USA.
Background:
Canarypox virus, ALVAC, does not replicate in infected mammalian cells and has potential as a vector for gene therapy in the treatment of cancer.
Purpose:
Recombinant viruses carrying DNA sequences encoding interleukin 2 (ALVAC-IL-2), interferon gamma (ALVAC-IFN gamma), tumor necrosis factor-alpha (ALVAC-TNF-alpha), or the co-stimulatory molecule B7-1 (ALVAC-B7-1) were investigated as agents for the treatment of a newly defined mouse prostate tumor model.
Methods:
RM-1 mouse prostate cancer cells, which are syngeneic (i.e., same genetic background) to C57BL/6 mice, were used. The expression of foreign gene products in vitro in infected RM-1 cells was measured by immunoprecipitation, bioassay, or flow cytometry. The effects of foreign gene product expression on RM-1 tumor cell growth in C57BL/6 mice were measured after subcutaneous injection (in the back) of 5 x 10(5) uninfected or infected cells; measurements included determinations of time to a measurable tumor size, tumor size as a function of time, and survival. The induction of protective immunity by uninfected and infected RM-1 cells was tested by injection of lethally irradiated (70 Gy) cells and subsequent challenge with uninfected cells. The generation of cytotoxic T cells was monitored by use of a 51Cr release assay. Severe combined immunodeficient (SCID) mice were used to determine whether T or B lymphocytes were involved in ALVAC vector-mediated antitumor responses. Data were analyzed by use of Pearson's modification of the chi-squared test and Kaplan-Meier survival methods. Reported P values are two-sided.
Results:
The level of foreign gene product expression in ALVAC-infected RM-1 cells was dependent on the multiplicity of virus infection used; a multiplicity of five viruses per infected cell was chosen for subsequent experiments. RM-1 tumor growth in C57BL/6 mice was not affected by tumor cell expression of IL-2 alone, IFN gamma alone, or B7-1 alone; however, expression of TNF-alpha alone significantly delayed tumor growth at early time points (compared with parental ALVAC-infected tumors, P = .0001 at day 21 and P = .037 at day 28). Tumor cell expression of both TNF-alpha and IL-2 completely inhibited tumor growth in 60%-100% of treated mice. No protection against subsequent tumor challenge was detected in mice previously exposed to RM-1 cells expressing both TNF-alpha and IL-2. Cytotoxic T-lymphocyte activity toward RM-1 cells was not observed in C57BL/6 mice that rejected tumors. Tumor cell expression of TNF-alpha and IL-2 also resulted in tumor growth inhibition in SCID mice.
Conclusions:
RM-1 mouse prostate cancer cells are readily infected by ALVAC vectors, and foreign gene products are efficiently expressed. Inhibition of RM-1 tumor growth by tumor cell expression of TNF-alpha and IL-2 appears to involve nonspecific antitumor activity.
Insights
Canarypox virus (ALVAC) vectors expressing tumor necrosis factor-alpha and interleukin 2 effectively inhibited mouse prostate tumor growth. This gene therapy approach shows promise for cancer treatment by leveraging non-replicating viral vectors.
Area of Science:
- Oncolytic Virotherapy
- Cancer Gene Therapy
- Immunotherapy
Background:
- Canarypox virus (ALVAC) is a non-replicating viral vector with potential for cancer gene therapy.
- Recombinant ALVAC vectors were engineered to express therapeutic genes for cancer treatment.
Purpose of the Study:
- To evaluate ALVAC-vectored interleukin 2 (IL-2), interferon gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), and B7-1 in a mouse prostate tumor model.
- To assess the efficacy of these recombinant viruses in inhibiting tumor growth and inducing immune responses.
Main Methods:
- RM-1 mouse prostate cancer cells were infected with ALVAC vectors expressing specific genes.
- Tumor growth, survival, and immune responses (cytotoxic T-lymphocytes) were evaluated in immunocompetent (C57BL/6) and immunodeficient (SCID) mice.
- Gene product expression was confirmed using immunoprecipitation, bioassays, and flow cytometry.
Main Results:
- Tumor cell expression of TNF-α alone delayed tumor growth; combined TNF-α and IL-2 expression completely inhibited tumor growth in 60%-100% of mice.
- Tumor inhibition was observed in both immunocompetent and SCID mice, suggesting a non-specific antitumor mechanism.
- No protective immunity against subsequent tumor challenge was observed.
Conclusions:
- ALVAC vectors efficiently express foreign genes in RM-1 prostate cancer cells.
- Co-expression of TNF-α and IL-2 by tumor cells leads to significant tumor growth inhibition, likely through non-specific mechanisms.
- ALVAC-based immunotherapy holds potential for prostate cancer treatment.