Behavior of Metastatic and Nonmetastatic Breast Tumors in Old Mice

Abstract

Breast cancer incidence and mortality increase with age. A better understanding of the biological behavior of metastatic and nonmetastatic breast tumors in older subjects may help to develop improved breast cancer therapies. In this study, we used syngeneic metastatic (4TO7cg) and nonmetastatic (64pT) mouse breast tumor models at three age levels to evaluate various characteristics that are considered to be important for effective anti-breast cancer immunotherapy. These included tumor size and growth, metastases, vascularization, gene expression levels of the tumor-associated antigen (TAA) Mage-b (homologous to human MAGE-B) in primary breast tumors and metastases, and the presence of CD4(+) and CD8(+) T cells in the inguinal lymph nodes at the site of the tumor. The primary breast tumors and metastases were generated by injection of mouse mammary tumor cell lines 4TO7cg or 64pT into a mammary fat pad of normal 3-, 9-, or 21/24-month old BALB/c mice. In the nonmetastatic breast tumor model, significantly smaller tumors were observed in old compared with young mice. This was associated with a significant increase in the percentage of CD8(+) T cells in inguinal lymph nodes and significantly higher Mage-b expression levels in the primary tumors at old age. In the metastatic (4TO7cg) breast tumor model, a less pronounced, not statistically significant, smaller tumor size was found in the old mice, without a difference in the percentage of CD8(+) T cells or Mage-b expression levels. However, in this mouse model almost all metastases showed high levels of Mage-b expression (2- to 3-fold higher than the primary tumors in the same animals) regardless of age. These results indicate that the metastatic and nonmetastatic breast tumor models could be useful model systems to analyze how breast cancer vaccines for humans can be tailored to old age.

Vaccination with Mage-b DNA induces CD8 T-cell responses at young but not old age in mice with metastatic breast cancer

“…In a previous study, we showed that Mage-b is highly expressed in 4TO7cg metastases and tumours in young and old mice (Gravekamp et al, 2004). We have also shown that Mage-b is highly expressed in 4T1 metastases and tumours of young mice .…”

“…These spontaneous tumours also express Mage-b (unpublished results). Age-related effects on the growth of 4TO7cg tumours and metastases have also been extensively analysed in a previous study (Gravekamp et al, 2004). In contrast to 4TO7cg, a dramatic effect of ageing was observed in the 4T1 model.…”

“…To study the efficacy of cancer vaccination against breast cancer metastases, two different preclinical metastatic breast tumour models were used, that is, 4TO7cg, which is a relatively mild model with 2 -20 metastases per mouse, and the much more aggressive 4T1 with 5 -300 metastases per mouse. Both models highly express Mage-b tumour-associated antigen (TAA) in their primary tumours and metastases (Gravekamp et al, 2004. Mouse Mage-b is homologous to human MAGE-B (Lurquin et al, 1997).…”

Mage-b vaccine delivered by recombinant Listeria monocytogenes is highly effective against breast cancer metastases

“…To avoid the generation of immune responses against all other TAA than Mage-b, we used 64pT instead 4T1 tumour cells in the restimulation assay. 64pT is a syngeneic breast tumour cell, highly expressing Mage-b (Gravekamp et al, 2004) and highly secreting IL-6 (3000 pg ml À1 ) (Gravekamp et al, 2008). Indeed, addition of anti-IL-6 antibodies to the in vitro restimulation assay significantly increased the production of IFNg, whereas the production of IFNg could not be induced in the same restimulation assay without anti-IL-6 antibodies ( Figure 6A).…”

“…Two days later, the number of IFNg-producing cells was determined by ELISPOT reader as described above. In addition, anti-IL-6 antibodies (50 mg ml À1 ) were either or not added to the wells with LNs (2 Â 10 5 ) and syngeneic 64pT tumour cells (1 Â 10 4 ; treated with mitomycin C; expressing highly Mage-b; Gravekamp et al, 2004), and secreting highly IL-6 (3000 pg ml À1 ) (Gravekamp et al, 2008), in 200 ml of RPMI containing 10% FBS. Two days later, the production of IFNg was determined by quantitative ELISA as described previously (Sypniewska et al, 2005).…”