title: Effect of canarypox virus (ALVAC)-mediated cytokine expression on murine prostate tumor growth.
abstract: 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.
  
  
  OBJECTIVE
  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.
authors.affiliation: Division of Urologic Surgery, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: M
authors.initials: M
authors.lastname: Kawakita
authors.forename: G S
authors.initials: GS
authors.lastname: Rao
authors.forename: J K
authors.initials: JK
authors.lastname: Ritchey
authors.forename: D K
authors.initials: DK
authors.lastname: Ornstein
authors.forename: M A
authors.initials: MA
authors.lastname: Hudson
authors.forename: J
authors.initials: J
authors.lastname: Tartaglia
authors.forename: E
authors.initials: E
authors.lastname: Paoletti
authors.forename: P A
authors.initials: PA
authors.lastname: Humphrey
authors.forename: T J
authors.initials: TJ
authors.lastname: Harmon
authors.forename: T L
authors.initials: TL
authors.lastname: Ratliff
mesh_terms: D000818:Animals; D018150:Avipoxvirus; D018122:B7-1 Antigen / Q000096:biosynthesis; D016207:Cytokines / Q000096:biosynthesis*; D004195:Disease Models, Animal; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D015967:Gene Expression Regulation, Viral; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D007167:Immunotherapy / Q000379:methods*; D007371:Interferon-gamma / Q000096:biosynthesis; D007376:Interleukin-2 / Q000096:biosynthesis; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016513:Mice, SCID; D011471:Prostatic Neoplasms / Q000276:immunology* / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
chemical_list: D018122:B7-1 Antigen; D016207:Cytokines; D007376:Interleukin-2; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma