Prostate carcinoma in transgenic Lewis rats - a tumor model for evaluation of immunological treatments

Abstract

Transgenic rodent models of prostate cancer have served as valuable preclinical models to evaluate novel treatments and understand malignant disease progression. In particular, a transgenic rat autochthonous model of prostate cancer using the SV40 large T antigen expressed under a prostate-specific probasin promoter was previously developed as a model of androgen-dependent prostate cancer (TRAP). In the current report, we backcrossed this strain to the Lewis strain, an inbred rat strain better characterized for immunological analyses. We demonstrate that Lewis transgenic rats (Lew-TRAP) developed prostate adenocarcinomas with 100% penetrance by 25 weeks of age. Tumors were predominantly androgen-dependent, as castration prevented tumor growth in the majority of animals. Finally, we demonstrate that Lew-TRAP rats could be immunized with a DNA vaccine encoding a human prostate tumor antigen (prostatic acid phosphatase) with the development of Lewis strain-specific T-cell responses. We propose that this Lew-TRAP strain, and prostate tumor cell lines derived from this strain, can be used as a future prostate cancer immunotherapy model.

Keywords: Lewis rat; prostate cancer; transgenic; vaccine.

Figure 1

Probasin/SV40 TAg+ transgenic rats on both Sprague Dawley and Lewis background strains develop prostate tumors with age. Probasin/SV40 TAg+ transgenic rats and non-transgenic littermate controls in both Sprague Dawley (SD-TRAP, panel A) and Lewis (Lew-TRAP, panel B) strain backgrounds underwent necropsy at 4, 10, 12, 15, 20, 25, 30, and 35 weeks of age. Rats were weighed and the genitourinary complexes were excised and weighed. Each dot represents the ratio of genitourinary complex mass to body mass for an individual rat at each time point. Closed circles represent SV40 TAg+ transgenic rats (SV40 TAg+) and the open circles represent the non-transgenic littermates (WT). Comparisons of the means between groups were made with an unpaired t-test and P-values are shown

Figure 2

Representative prostate tissue sections from Lew-TRAP rats at multiple time points. Shown are representative prostate tissue sections (stained with hematoxylin and eosin) from Lew-TRAP rats. Panel A. Atypical hyperplasia noted at 4 weeks old. Panel B: PIN observed at 12 weeks of age; Panel C. Moderately-to well-differentiated adenocarcinoma observed at 15 weeks old; Panel D. Poorly-differentiated carcinoma observed at 35 weeks of age; Panels E and F. Lew-TRAP prostate tissue obtained at 30 weeks of age following castration at 20 weeks, with no tumor observed in one animal (panel E), and poorly-differentiated tumor observed in another (panel F). Prostate tumors from 25-week old animals were stained for SV40 TAg (panel G) or androgen receptor (panel H)

Figure 3

Characterization of prostate tumor cell lines derived from Lew-TRAP rats. Prostate tumor cell lines were established from the ventral lobe of a 32-week old, Lew-TRAP rat. The expression of SV40 TAg (Panel A) was detected by Western blot. [Lane 1: mouse TRAMP cell line (6); lanes 2–4 individual Lew-TRAP prostate cell lines.] The arrow indicates the expected size of the TAg band. Panel B: Prostatic acid phosphatase activity in lysates of the same individual Lew-TRAP cell lines or the AR-deficient human prostate cancer cell line, DU145 (negative control). Shown is the tartrate-inhibited acid phosphatase activity per mg protein lysate. The data are representative of two to three replicate experiments

Figure 4

I-mmunization of Lew-TRAP rats with a DNA vaccine encoding PAP elicited a PAP-specific immune response. Seven-week old, male Lew-TRAP rats received six intradermal immunizations with 100 μg of pTVG4 (n=5) or pTVG-HP (n=6) at weekly intervals. Splenocytes were cultured with media alone, human PAP protein (PAP), PAP peptides (HP_201–215 or RP_200–214), or PHA for 72 hours, and then pulsed with BrdU for 8–12 hours. Cells were then assessed for antigen-specific CD8+ (panel A) or CD4+ (panel B) proliferation by flow cytometry. Supernatants of splenocyte cultures were assessed for IFNγ release by quantitative ELISA (Panel C). Each dot represents the frequency of BrdU+ T cells of each rat or the average IFNγ release of each rat as determined in triplicate. Comparisons of the means between the groups were made with an unpaired t-test and P-values are shown. Sera from immunized rats were evaluated for the presence of anti-human PAP or anti-OVA (negative control) antibodies using an indirect ELISA (Panel D). Each point represents the average absorbance difference and standard deviation from the sera of pTVG4 (n=5) and pTVG-HP (n=6) immunized rats