Modeling Human Prostate Cancer Metastasis in Mice via Resection of Subcutaneous Allografts

Abstract

The 5-year survival rate for patients diagnosed with distant metastatic prostate cancer in the United States is 30.6%. Therefore, there is a great need to develop in vivo model systems to study prostate cancer metastasis and to test potential therapeutics. Most murine prostate cancer metastatic models involve intracardiac or intraosseous implantation of cancer cells, which bypass the early stages of tumor cell migration and invasion. Herein we provide a detailed protocol for a novel method of resecting subcutaneous prostate cancer allografts in immunocompetent mice to produce spontaneous metastases and describe a pilot study using this method of tumor resection. Intact male FVB/NCrl mice (n = 9) were inoculated subcutaneously with Myc-CaP cells. Tumors were surgically resected, and mice were monitored for tumor recurrence. Animals were euthanized or died, and a full set of tissues was collected for histopathologic examination. Tumors took an average of 44 days (range 23-61) to reach 1.7 cm in any direction. All tumors were resectable, and resection of the tumors increased the study length by 70 days (range 30-121). One mouse was euthanized early of an unrelated cause, and of eight remaining mice, four developed tumor recurrence at the site of resection. One mouse developed bone metastases, one mouse developed metastases to the abdominal cavity, and two mice showed signs of local invasion. This study demonstrates that resection of subcutaneous Myc-CaP cell allografts in mice results in local tumor recurrence and the development of distant metastases, providing a new model system to study prostate cancer metastasis in vivo.

Keywords: FVB mouse7; Myc-CaP3; allograft4; metastasis2; mouse model5; prostate cancer1; resection6.

Figure 1

Summary of pilot study. Nine mice were inoculated with Myc-CaP cells. Tumors were resected from all nine mice once tumor size reached 1.7 cm in any direction, an average of 44 days (range 23–61 days) after engraftment. One mouse was euthanized due to dermatitis of an unknown cause. Four of the remaining eight mice had recurrent tumors of which two showed signs of local invasion. Two of the remaining four mice had distant metastasis. Seven mice were euthanized on average 70 days (range 30–121 days) post-resection when recurrent tumors reached 1.7 cm, tumors became ulcerated, or mice were moribund. One mouse was found dead.

Figure 2

Confirmation that tumors were derived from Myc-CaP cells. H&E staining of tumor cells from resected, recurred, and metastatic tumors demonstrate abundant cytoplasmic expression of human MYC and nuclear AR protein expression, consistent with Myc-CaP cells. Scale bar = 2 mm (top), 50 µm (bottom).

Figure 3

Gross and histologic findings for Mouse #3. This animal was found dead with (A) 3 ml of serosanguinous fluid and approximately 20 tan nodules up to 1 cm in diameter in the abdominal cavity, which expresses (B) both human MYC mRNA and AR protein expression. Scale bar = 5 mm (top), 50 µm (bottom). (C) H&E showed neoplastic cells present within the thoracic and abdominal cavities, and human MYC expression confirmed that these tumors were derived from Myc-CaP cells (scale bar = 5 mm). Arrows show neoplastic cells. (D) Human MYC-positive cells invaded the pancreas and were present intravascularly. Scale bar = 200 µm.

Figure 4

Gross and histologic findings for Mouse #5. (A) This animal presented clinically with a hard swelling of the left front leg. (B) The swelling was due to a well-demarcated, 1 cm, white mass that involved the humerus. Scale bar = 20 mm. (C) Sheets of neoplastic cells surround the bone; they multifocally disrupt the cortex (arrow). Scale bar = 500 µm. The medullary cavity is occupied by necrotic cells (arrowheads) and (D) nests of neoplastic cells (arrow). Scale bar = 200 µm. (E) Sub-periosteally, bone trabeculae radiate from the cortex and are surrounded by neoplastic cells. Scale bar = 100 µm.

Figure 5

Gross and histologic findings for Mouse #8. This mouse was euthanized when the recurrent tumor measured 1.7 cm in size. (A) Subcutaneous nodules were located in the pelvic region, over the right thigh. (B) H&E showed skeletal muscle invasion (arrows) of MYC-positive neoplastic cells. Scale bar = 2 mm. (C) MYC-positive cells are observed between myofibers (arrowhead) and completely entrapping myofibers (arrow). Scale bar = 100 µm.