RARα1 control of mammary gland ductal morphogenesis and wnt1-tumorigenesis

Introduction: Retinoic acid signaling pathways are disabled in human breast cancer suggesting a controlling role in normal mammary growth that might be lost in tumorigenesis. We tested a single receptor isotype, RAR alpha 1 (retinoic acid receptor isotype alpha, isoform 1), for its role in mouse mammary gland morphogenesis and mouse mammary tumor virus (MMTV)-wingless-related MMTV integration site 1 (wnt1)-induced oncogenesis. Methods: The role of RAR alpha 1 in mammary morphogenesis was tested in RAR alpha 1-knockout (KO) mice and in mammary tumorigenesis in bi-genic (RAR alpha 1/KO crossed with MMTV-wnt1) mice. We used whole mounts analysis, stem cells/progenitor quantification, mammary gland repopulation, quantitative polymerase chain reaction (Q-PCR), test of tumor-free survival, tumor fragments and cell transplantation. Results: In two genetic backgrounds (129/Bl-6 and FVB) the neo-natal RAR alpha 1/KO-mammary epithelial tree was two-fold larger and the pubertal tree had two-fold more branch points and five-fold more mature end buds, a phenotype that was predominantly epithelial cell autonomous. The stem/progenitor compartment of the RAR alpha 1/KO mammary, defined as CD24(low)/ALDH(high) (activity) was increased by a median 1.7-fold, but the mammary stem cell (MaSC)-containing compartment, (CD24(low)/CD29(high)), was larger (approximately 1.5-fold) in the wild type (wt)-glands, and the mammary repopulating ability of the wt-gland epithelium was approximately two-fold greater. In MMTV-wnt1 transgenic glands the progenitor (CD24(low)/ALDH(high) (activity)) content was 2.6-fold greater than in the wt and was further increased in the RAR alpha 1/KO-wnt1 glands. The tumor-free survival of RAR alpha 1/KO-wnt1 mice was significantly (P = 0.0002, Kaplan Meier) longer, the in vivo growth of RAR alpha 1/KO-wnt1 transplanted tumor fragments was significantly (P = 0.01) slower and RAR alpha 1/KO-wnt1 tumors cell suspension produced tumors after much longer latency. Conclusions: In vitamin A-replete mice, RAR alpha 1 is required to maintain normal mammary morphogenesis, but paradoxically, also efficient tumorigenesis. While its loss increases the density of the mammary epithelial tree and the content of luminal mammary progenitors, it appears to reduce the size of the MaSC-containing compartment, the mammary repopulating activity, and to delay significantly the MMTV-wnt1-mammary tumorigenesis. Whether the delay in tumorigenesis is solely due to a reduction in wnt1 target cells or due to additional mechanisms remains to be determined. These results reveal the intricate nature of the retinoid signaling pathways in mammary development and carcinogenesis and suggest that a better understanding will be needed before retinoids can join the armament of effective anti-breast cancer therapies.