Mouse models of advanced spontaneous metastasis for experimental therapeutics

Abstract

An enduring problem in cancer research is the failure to reproduce highly encouraging preclinical therapeutic findings using transplanted or spontaneous primary tumours in mice in clinical trials of patients with advanced metastatic disease. There are several reasons for this, including the failure to model established, visceral metastatic disease. We therefore developed various models of aggressive multi-organ spontaneous metastasis after surgical resection of orthotopically transplanted human tumour xenografts. In this Opinion article we provide a personal perspective summarizing the prospect of their increased clinical relevance. This includes the reduced efficacy of certain targeted anticancer drugs, the late emergence of spontaneous brain metastases and the clinical trial results evaluating a highly effective therapeutic strategy previously tested using such models.

Figure 1. The metastatic cascade

Metastatically competent primary tumours grow, invade the local host tissue and eventually shed tumour cells into the circulation. These cells travel to and colonize distant organs, and their subsequent growth at the secondary sites constitutes metastatic disease^,. Spontaneous metastasis assays involve establishing a primary tumour that is allowed to grow and spread in the host, whereas experimental metastasis assays circumvent the initial growth and invasion stages as a result of directly injecting tumour cells into the circulation. Other assays mimic aspects of metastatic growth by seeding tumour cells at the secondary site (for example, by intrasplenic injection of colorectal cancer cells, which directly targets the cells to the liver where they grow as metastases). Figure is modified, with permission, from REF. © (1982) Amerian Association for the Advancement of Science.

Figure 2. The selection of metastatically aggressive subpopulations of tumour cells often requires rounds of in vivo selection

For example, a primary tumour of the human melanoma cell line WM239 was established by the injection of the cells subdermally (orthotopically) into severe combined immunodeficient (SCID) mice, allowed to grow and establish itself for 3 weeks, and was then surgically removed (to prevent the rapidly growing primary tumour from causing end point termination of the experiment). Several months later the mice showed evidence of lung metastases — from which the subline 113/6-4L was derived. In some studies, these cells were re-implanted into a new host for an additional round of selection. This type of in vivo enrichment procedure was also used to derive, for example, the highly metastatic 231/LM2-4 variant of the human breast cancer cell line MDA-MB-231 (REF. 34). By using a similar selection procedure, the H2N/met2.hCG metastatic variant was derived from MDA-MB-231 cells that previously expressed ERBB2 by gene transduction and which were then transfected to express and secrete the β-subunit of human choriogonadotropin (β-hCG) protein, which can be used as a surrogate molecular marker of disease burden and response to therapy^,. The treatment of mice with established 113/6-4L spontaneous metastases using a metronomic doublet vinblastine (Vbl) and CTX regimen led to prolonged survival and to the eventual emergence of brain metastases. Isolation of a brain metastases resulted in the derivation of a variant subpopulation that spontaneously metastasizes to the brain without any therapeutic intervention to prolong the survival times of mice. Figure is modified, with permission, from REF. © (2008) American Association for Cancer Research.

Figure 3. Differential response of metastases and primary tumours to therapy

A disparity between primary tumours (part a) and metastasis (part b) in terms of response to therapy was observed with the human breast cancer model 231/LM2-4 treated with the combination of metronomic daily cyclophosphamide (CTX) and the 5-Fluorouracil (5-FU) oral prodrug UFT (tegafur plus uracil). Thus, primary tumour growth assays involving treatment showed no antitumour effect by combining UFT with CTX, and did not foreshadow the UFT and CTX doublet metronomic therapy (part a). Figure is modified, with permission, from REF. © (2006) American Association for Cancer Research.

Figure 4. Treatment of visceral metastatic disease can result in the appearance of metastases to the brain

a | Mice with established spontaneous metastases from the highly metastatic variant 113/6-4L derived from the WM239 melanoma were treated using a metronomic doublet vinblastine (Vbl) and cyclophosphamide (CTX) regimen (FIG. 2), and this led to prolonged survival and to the eventual emergence of the brain metastases. b | A therapeutic benefit was not observed when the Vbl and CTX regimen was administered to 113/6-4L grown as primary tumours. c | Image of brain metastasis from this experiment. Figure is modified, with permission, from REF. © (2008) American Association for Cancer Research.