New Generation of Meso and Antiprogestins (SPRMs) into the Osteoporosis Approach

Receptor activator of nuclear factor κB (RANK) and its ligand (RANKL) play key roles in bone metabolism and the immune system. The RANK/RANKL complex has also been shown to be critical in the formation of mammary epithelia cells. The female hormones estradiol and progesterone closely control the action of RANKL with RANK. Blood concentration of these sex hormones in the postmenopausal period leads to an increase in RANK/RANKL signaling and are a major cause of women’s osteoporosis, characterized by altered bone mineralization. Knowledge of the biochemical relationships between hormones and RANK/RANKL signaling provides the opportunity to design novel therapeutic agents to inhibit bone loss, based on the anti-RANKL treatment and inhibition of its interaction with the RANK receptor. The new generation of both anti- and mesoprogestins that inhibit the NF-κB-cyclin D1 axis and blocks the binding of RANKL to RANK can be considered as a potential source of new RANK receptor ligands with anti-RANKL function, which may provide a new perspective into osteoporosis treatment itself as well as limit the osteoporosis rise during breast cancer metastasis to the bone.

Cooperation of membrane-translocated syntaxin4 and basement membrane for dynamic mammary epithelial morphogenesis

Mammary epithelia undergo dramatic morphogenesis after puberty. During pregnancy, luminal epithelial cells in ductal trees are arranged to form well-polarized cystic structures surrounded by a myoepithelial cell layer, an active supplier of the basement membrane (BM). Here, we identified a novel regulatory mechanism in this process by using a reconstituted BM-based three-dimensional culture and aggregates of a model cell line EpH4, which had been manipulated for inducible expression of a t-SNARE protein syntaxin4, either in an intact or signal peptide-connected form, and those genetically deficient in syntaxin4. We found that cells extruded syntaxin4 upon stimulation with the lactogenic hormone, prolactin, which in turn accelerated the turnover of E-cadherin. In response to extracellular expression of syntaxin4, cell populations that were less affected by BM actively migrated and integrated into the BM-faced cell layer. Concurrently, the BM-faced cells, which were simultaneously stimulated with syntaxin4 and BM, acquired unique epithelial characteristics to undergo dramatic cellular arrangement for cyst formation. These results highlight the importance of the concerted action of extracellular syntaxin4 extruded by the lactogenic hormone and BM components in epithelial morphogenesis.

Mammary-specific expression of Trim24 establishes a mouse model of human metaplastic breast cancer

Conditional overexpression of histone reader Tripartite motif containing protein 24 (TRIM24) in mouse mammary epithelia (Trim24COE) drives spontaneous development of mammary carcinosarcoma tumors, lacking ER, PR and HER2. Human carcinosarcomas or metaplastic breast cancers (MpBC) are a rare, chemorefractory subclass of triple-negative breast cancers (TNBC). Comparison of Trim24COE metaplastic carcinosarcoma morphology, TRIM24 protein levels and a derived Trim24COE gene signature reveals strong correlation with human MpBC tumors and MpBC patient-derived xenograft (PDX) models. Global and single-cell tumor profiling reveal Met as a direct oncogenic target of TRIM24, leading to aberrant PI3K/mTOR activation. Here, we find that pharmacological inhibition of these pathways in primary Trim24COE tumor cells and TRIM24-PROTAC treatment of MpBC TNBC PDX tumorspheres decreased cellular viability, suggesting potential in therapeutically targeting TRIM24 and its regulated pathways in TRIM24-expressing TNBC.

Loss of Nitric Oxide Induces Fibrogenic Response in Organotypic 3D Co-Culture of Mammary Epithelia and Fibroblasts—An Indicator for Breast Carcinogenesis

Excessive myofibroblast activation, which leads to dysregulated collagen deposition and the stiffening of the extracellular matrix (ECM), plays pivotal roles in cancer initiation and progression. Cumulative evidence attests to the cancer-causing effects of a number of fibrogenic factors found in the environment, diseases and drugs. While identifying such factors largely depends on epidemiological studies, it would be of great importance to develop a robust in vitro method to demonstrate the causal relationship between fibrosis and cancer. Here, we tested whether our recently developed organotypic three-dimensional (3D) co-culture would be suitable for that purpose. This co-culture system utilizes the discontinuous ECM to separately culture mammary epithelia and fibroblasts in the discrete matrices to model the complexity of the mammary gland. We observed that pharmaceutical deprivation of nitric oxide (NO) in 3D co-cultures induced myofibroblast differentiation of the stroma as well as the occurrence of epithelial–mesenchymal transition (EMT) of the parenchyma. Such in vitro response to NO deprivation was unique to co-cultures and closely mimicked the phenotype of NO-depleted mammary glands exhibiting stromal desmoplasia and precancerous lesions undergoing EMT. These results suggest that this novel 3D co-culture system could be utilized in the deep mechanistic studies of the linkage between fibrosis and cancer.

Epigenetic changes with age primes mammary luminal epithelia for cancer initiation

SummaryAging causes molecular changes that manifest as stereotypical phenotypes yet aging-associated diseases progress only in certain individuals. At lineage-specific resolution, we show how stereotyped and variant responses are integrated in mammary epithelia. Age-dependent directional changes in gene expression and DNA methylation (DNAm) occurred almost exclusively in luminal cells and implicated genome organizers SATB1 and CTCF. DNAm changes were robust indicators of aging luminal cells, and were either directly (anti-)correlated with expression changes or served as priming events for subsequent dysregulation, such as demethylation of ESR1-binding regions in DNAm-regulatory CXXC5 in older luminal cells and luminal-subtype cancers. Variance-driven changes in the transcriptome of both luminal and myoepithelial lineages further contributed to age-dependent loss of lineage fidelity. The pathways affected by transcriptomic and DNAm changes during aging are commonly linked with breast cancer, and together with the differential variability found across individuals, influence aging-associated cancer susceptibility in a subtype-specific manner.

Deep Proteome Profiling of Human Mammary Epithelia at Lineage and Age Resolution

SUMMARYAge is the major risk factor in most carcinomas, yet little is known about how proteomes change with age in any human epithelium. We present comprehensive proteomes comprised of >9,000 total proteins, and >15,000 phosphopeptides, from normal primary human mammary epithelia at lineage resolution from ten women ranging in age from 19 to 68. Data were quality controlled, and results were biologically validated with cell-based assays. Age-dependent protein signatures were identified using differential expression analyses and weighted protein co-expression network analyses. Up-regulation of basal markers in luminal cells, including KRT14 and AXL, were a prominent consequence of aging. PEAK1 was identified as an age-dependent signaling kinase in luminal cells, which revealed a potential age-dependent vulnerability for targeted ablation. Correlation analyses between transcriptome and proteome revealed age-associated loss of proteostasis regulation. Protein expression and phosphorylation changes in the aging breast epithelium identify potential therapeutic targets for reducing breast cancer susceptibility.