Bringing androgens up a NOTCH in breast cancer

While it has been known for decades that androgen hormones influence normal breast development and breast carcinogenesis, the underlying mechanisms have only been recently elucidated. To date, most studies have focused on androgen action in breast cancer cell lines, yet these studies represent artificial systems that often do not faithfully replicate/recapitulate the cellular, molecular and hormonal environments of breast tumours in vivo. It is critical to have a better understanding of how androgens act in the normal mammary gland as well as in in vivo systems that maintain a relevant tumour microenvironment to gain insights into the role of androgens in the modulation of breast cancer development. This in turn will facilitate application of androgen-modulation therapy in breast cancer. This is particularly relevant as current clinical trials focus on inhibiting androgen action as breast cancer therapy but, depending on the steroid receptor profile of the tumour, certain individuals may be better served by selectively stimulating androgen action. Androgen receptor (AR) protein is primarily expressed by the hormone-sensing compartment of normal breast epithelium, commonly referred to as oestrogen receptor alpha (ERa (ESR1))-positive breast epithelial cells, which also express progesterone receptors (PRs) and prolactin receptors and exert powerful developmental influences on adjacent breast epithelial cells. Recent lineage-tracing studies, particularly those focussed on NOTCH signalling, and genetic analysis of cancer risk in the normal breast highlight how signalling via the hormone-sensing compartment can influence normal breast development and breast cancer susceptibility. This provides an impetus to focus on the relationship between androgens, AR and NOTCH signalling and the crosstalk between ERa and PR signalling in the hormone-sensing component of breast epithelium in order to unravel the mechanisms behind the ability of androgens to modulate breast cancer initiation and growth.Gerard A Tarulli, Lisa M Butler, Wayne D Tilley and Theresa E Hicke

Unraveling an identity for the androgen receptor-expressing mammary epithelial cell

Abstracts of the Endocrine Society of Australia Annual Scientific Meeting 2015, 23–26 August 2015Gerard A Tarulli, Geraldine Laven-Law, Wayne D Tilley, and Theresa E Hicke

New insights into lineage restriction of mammary gland epithelium using parity-identified mammary epithelial cells

10.1186/bcr3593Breast Cancer Research161-BCRR

Mammary-specific ablation of Cyp24a1 inhibits development, reduces proliferation and increases sensitivity to vitamin D

Active vitamin D (1,25(OH)2D) has been shown to regulate numerous cell processes in mammary cells. Degradation of 1,25(OH)2D is initiated by the mitochondrial enzyme, 25-hydroxyvitamin D 24-hydroxylase (CYP24 A1), and provides local control of 1,25(OH)2D bioactivity. Several reports of the association between elevated CYP24 A1 activity and breast cancer incidence, suggest that CYP24 A1 may be a target for therapeutic intervention. Whether CYP24 A1 activity within the mammary epithelium regulates 1,25(OH)2D levels and mammary gland development is yet to shown. We have used a conditional knockout of the Cyp24a1 gene specifically in the mammary epithelium to demonstrate reduced terminal end bud number, ductal outgrowth and branching during puberty and alveologenesis at early pregnancy, by inhibiting proliferation but not apoptosis in both basal and luminal MECs. In vitro study showed increased sensitivity of luminal MECs to lower levels of 1,25(OH)2D with the ablation of Cyp24a1 activity. In summary, Cyp24a1 within MECs plays an important role in modulating postnatal and pregnancy-associated mammary gland development which provides support for inhibiting CYP24 A1 as a potential approach to activating the vitamin D pathway in breast cancer prevention and therapy.Lei Sheng, Andrew G.Turner, Kate Barratt, Richard Kremer, Howard A.Morris, David F.Callen, Paul H.Anderson, Gerard A.Tarull

Gonadotropin suppression in men leads to a reduction in claudin-11 at the Sertoli cell tight junction

STUDY QUESTION Are Sertoli cell tight junctions (TJs) disrupted in men undergoing hormonal contraception? SUMMARY ANSWER Localization of the key Sertoli cell TJ protein, claudin-11, was markedly disrupted by 8 weeks of gonadotropin suppression, the degree of which was related to the extent of adluminal germ cell suppression. WHAT IS KNOWN ALREADY Sertoli cell TJs are vital components of the blood–testis barrier (BTB) that sequester developing adluminal meiotic germ cells and spermatids from the vascular compartment. Claudin-11 knockout mice are infertile; additionally claudin-11 is spatially disrupted in chronically gonadotropin-suppressed rats coincident with a loss of BTB function, and claudin-11 is disorganized in various human testicular disorders. These data support the Sertoli cell TJ as a potential site of hormonal contraceptive action. STUDY DESIGN, SIZE, DURATION BTB proteins were assessed by immunohistochemistry (n = 16 samples) and mRNA (n = 18 samples) expression levels in available archived testis tissue from a previous study of 22 men who had undergone 8 weeks of gonadotropin suppression and for whom meiotic and post-meiotic germ cell numbers were available. The gonadotropin suppression regimens were (i) testosterone enanthate (TE) plus the GnRH antagonist, acyline (A); (ii) TE + the progestin, levonorgestrel, (LNG); (iii) TE + LNG + A or (iv) TE + LNG + the 5α-reductase inhibitor, dutasteride (D). A control group consisted of seven additional men, with three archived samples available for this study. PARTICIPANTS/MATERIALS, SETTINGS, METHODS Immunohistochemical localization of claudin-11 (TJ) and other junctional type markers [ZO-1 (cytoplasmic plaque), β-catenin (adherens junction), connexin-43 (gap junction), vinculin (ectoplasmic specialization) and β-actin (cytoskeleton)] and quantitative PCR was conducted using matched frozen testis tissue. MAIN RESULTS AND THE ROLE OF CHANCE Claudin-11 formed a continuous staining pattern at the BTB in control men. Regardless of gonadotropin suppression treatment, claudin-11 localization was markedly disrupted and was broadly associated with the extent of meiotic/post-meiotic germ cell suppression; claudin-11 staining was (i) punctate (i.e. ‘spotty’ appearance) at the basal aspect of tubules when the average numbers of adluminal germ cells were <15% of control, (ii) presented as short fragments with cytoplasmic extensions when numbers were 15–25% of control or (iii) remained continuous when numbers were >40% of control. Changes in localization of connexin-43 and vinculin were also observed (smaller effects than for claudin-11) but ZO-1, β-catenin and β-actin did not differ, compared with control. LIMITATIONS, REASONS FOR CAUTION Claudin-11 was the only Sertoli cell TJ protein investigated, but it is considered to be the most pivotal of constituent proteins given its known implication in infertility and BTB function. We were limited to testis samples which had been gonadotropin-suppressed for 8 weeks, shorter than the 74-day spermatogenic wave, which may account for the heterogeneity in claudin-11 and germ cell response observed among the men. Longer suppression (12–24 weeks) is known to suppress germ cells further and claudin-11 disruption may be more uniform, although we could not access such samples. WIDER IMPLICATIONS OF THE FINDINGS These findings are important for our understanding of the sites of action of male hormonal contraception, because they suggest that BTB function could be ablated following long-term hormone suppression treatment