Despite the importance of TGFα and ErbB-mediated pathways in refractory clinical breast cancers, in vivo
models to explore these relationships during tumorigenesis have not been available. Here, we employed the novel NRL promoter, which drives uniform expression in mammary epithelial cells independently of estrogens and ovariectomy, to demonstrate that TGFα-induced tumorigenesis was strongly moderated by prophylactic ovariectomy. Although ER transcripts declined with tumorigenesis, end stage lesions remained ER+, but did not express PR. The declining but continued ER expression and associated lack of detectable PR observed in these lesions models the class of aggressive ER+/PR− breast tumors in women that prove refractory to hormonally-based therapeutics, and which often contain higher levels of growth factor ligands and/or receptors compared to ER + /PR + tumors (Arpino et al., 2005
The NRL-TGFα model shares many features with ER+/PR− clinical breast cancers. ER is present in epithelial and nonepithelial mammary cells, increasing in hyperplasias (Lee et al., 2006
). In addition, reduction in ER and, more radically, PR transcripts in NRL-TGFα mice parallels clinical findings in which breast cancers expressing TGFα and ErbB1 had reduced ER (Bieche et al., 2003
Despite diminishing ER mRNA in NRL-TGFα mice, prophylactic ovariectomy significantly reduced tumor incidence and increased latency. Early dependence on ovarian hormones despite elevated growth factor expression contrasts with findings in MMTV-neu/ErbB2 mice; ovariectomy at similar ages did not alter tumor incidence or latency (Hewitt et al., 2002
) suggesting constitutively active ErbB2 exerts more powerful growth/survival signals and early ovarian steroid independence, compared to TGFα mediated ErbB1 signaling. Cellular and tissue constraints which attentuate biological responses to TGFα may be responsible. Recent findings revealing distinctive molecular profiles for ER− tumors containing ErbB1 versus ErbB2 (Creighton et al., 2006
) may shed light on these differences.
Cystic papillary tumors in NRL-TGFα mice resemble those of other TGFα transgenic mice (Humphreys and Hennighausen, 2000
; Rose-Hellekant and Sandgren, 2000
; Arendt et al., 2006
). Concurrent expression of heterozygous p53 results in aggressive solid tumors of varying types reflecting the tumor spectrum of p53+/− BALB/c mice (Blackburn et al., 2003
; Jerry et al., 2000
; Kuperwasser et al., 2000
), although relative proportions cannot be determined due to low numbers of p53+/− mice in the present study. The B6 background delayed tumor latency in NRL-TGFα mice, similar to that observed previously in WAP-TGFα mice (Rose-Hellekant et al., 2002
) and p53+/− (BALB/c:B6) F1 mice (Jerry et al., 2000
), suggesting that B6 alleles protect against mammary cancer. The increased incidence, shortened latency and increased tumor grade conferred by p53 heterozygosity in the NRL-TGFα model is similar to other transgenic mice expressing mammary oncogenes concurrently with heterozygous, null or mutated p53 (e.g. Wnt-1, c-myc, c-neu, ras, DES, BRCA1 and BRCA2; reviewed by Blackburn and Jerry, 2002
). In combination with heterozygosity at the p53 locus, TGFα tumors were ER−, consistent with accelerated tumor progression facilitated by genomic instability or selective progression of a distinct tumor precursor subpopulation. Both ER+ and ER− tumors have been observed in other mouse models with one or more dysfunctional p53 alleles (Medina and Kittrell, 2003
; Zhang et al., 2005
ER and PR immunohistochemical status of NRL-TGFα tumors is similar to WAP-TGFα tumors (unpublished), and tumors from MMTV-TGFα × MMTV-c-myc bitransgenic mice also were ER+ using ligand binding assay (Amundadottir et al., 1995
). The presence of ER in end stage NRL-TGFα tumors suggests that estrogen may continue to be an important mitogen even at later stages of tumorigenesis, although the sensitivity of tumor growth to estrogens or antiestrogens has not been tested in this model. However, the 40-fold reduction in PR mRNA and lack of detectable protein observed in these tumors strongly suggests that progesterone plays little or no role in established tumors, which may reflect tumor cellular origin or a cause-effect relationship between changes in ER status with tumor progression.
In summary, NRL-TGFα mice have the following characteristics: (1) Females develop proliferative hyperplasias, complex cystic and solid adenomatous tumors similar to clinical breast lesion histotypes. (2) Most early and end stage lesions are ER+, but become PR− very early, thus resembling the ER+/PR− tumor type found in 25% of women with breast cancer. (3) Tumor development can be impaired by prophylactic ovariectomy. (4) Finally, coupling a TGFα growth environment with monoallelic p53 results in aggressive ER− carcinomas. These data indicate not only a role for TGFα in breast carcinogenesis, but suggest an intricate relationship between TGFα, ER and PR in the NRL-TGFα mouse model reflecting many aspects of ER + /PR− breast carcinogenesis in women, and thus can be used to dissect underlying mechanisms of tumorigenesis in this resistant tumor type.