Supplementary Figure S1. A2/TRAMP Animals Develop Prostate Adenocarcinomas

Supplementary Figure Legends

Supplementary Figure S1. A2/TRAMP animals develop prostate adenocarcinomas

Homozygous HHDII-DR1 and TRAMP mice were crossed to generate F1 heterozygous

A2/TRAMP animals. Male A2/TRAMP mice were euthanized at the indicated ages, and genitourinary complexes (including bladder, prostate, and seminal vesicles) were collected and weighed. Solid bars represent average genitourinary complex mass of age groups, with the number of animals per group listed on the x-axis under the animal age.

Supplementary Figure S2. A2/TRAMP prostate adenocarcinomas were used to generate a syngeneic A2/TRAMP prostate cancer cell line

Prostate tumors were collected from 24 week-old A2/TRAMP mice and were used to generate prostate cancer cell lines as described in Materials and Methods. Panel A,

A2/TRAMP prostate cancer cell protein lysates were analyzed for the expression of the

AR and SV40 by Western blot. LNCaP cells (which express the AR and not SV40 large

T antigen) and Cos7 cells (which express the SV40 large T antigen and not the AR) were also included as controls. Expected sizes of proteins are indicated by arrows.

Panel B, A2/TRAMP prostate cancer cells were analyzed for human (left) and murine

(right) MHC class I expression by flow cytometry (MHC class I antibodies – black lines; isotype controls – grey lines).

Supplementary Figure S3. Individual color images of immunofluorescence staining of sixteen-week old A2/TRAMP prostates Individual sixteen-week old A2/TRAMP animals immunized with either pTVG4 or pTVG-

AR were analyzed for tumor development by histological analysis combined with immunofluorescence using antibodies specific for SV40 (green), laminin (red), and a

DAPI counterstain (blue). Shown are the individual color images for the three channels used to generate the merged images shown in Figure 3A-D (first row, Figure 3A, normal prostate; second row, Figure 3B, PIN; third row, Figure 3C, WDC; fourth row,

Figure 3D, MDC; fifth row, Figure 3D inset, MDC). The three channels used to generate the merged images are displayed by column: first column: DAPI; second column; FITC, third column; Texas Red; fourth column; merged images.

Supplementary Figure S4. Poorly-differentiated A2/TRAMP tumors do not express the androgen receptor, and have CD8+ T cells at the margins but not infiltrating the tumor

Panel A, prostates from pTVG4- or pTVG-AR-immunized animals with evidence of poorly-differentiated carcinoma were analyzed for expression of the androgen receptor

(n=5). Shown is a representative example of an animal immunized with pTVG4 demonstrating that these poorly-differentiated tumors lacked expression of the androgen receptor as detected by immunohistochemistry (10x magnification, scale bar=200μm), whereas adjacent normal, PIN, and WDC tumor foci were found to have robust androgen receptor expression. Panel B, prostates from immunized eighteen- week old A2/TRAMP animals with evidence of poorly-differentiated carcinoma were analyzed for infiltrating CD8+ T cells by immunohistochemistry. Shown is a representative example of an animal immunized with pTVG-AR (10x magnification scale bar=200μm; 20x inset magnification scale bar=100μm)