Epidermal Overexpression of Stratum Corneum Chymotryptic Enzyme in Mice: A Model for Chronic Itchy Dermatitis

Identification of tissue-specific mechanisms involved in the pathophysiology of inflammatory skin diseases could offer new possibilities to develop effective therapies with fewer systemic effects. The serine protease stratum corneum chymotryptic enzyme is preferentially expressed in cornifying epithelia. We have previously reported on increased expression of the stratum corneum chymotryptic enzyme in psoriasis. Here is reported an increased epidermal expression of stratum corneum chymotryptic enzyme also found in chronic lesions of atopic dermatitis. Transgenic mice expressing human stratum corneum chymotryptic enzyme in suprabasal epidermal keratinocytes were found to develop pathologic skin changes with increased epidermal thickness, hyperkeratosis, dermal inflammation, and severe pruritus. The results suggest that stratum corneum chymotryptic enzyme may be involved in the pathogenesis of inflammatory skin diseases, and that stratum corneum chymotryptic enzyme and related enzymes should be evaluated as potential targets for new therapies

Heat shock protein-90-alpha, a prolactin-STAT5 target gene identified in breast cancer cells, is involved in apoptosis regulation

Introduction The prolactin-Janus-kinase-2-signal transducer and activator of transcription-5 (JAK2-STAT5) pathway is essential for the development and functional differentiation of the mammary gland. The pathway also has important roles in mammary tumourigenesis. Prolactin regulated target genes are not yet well defined in tumour cells, and we undertook, to the best of our knowledge, the first large genetic screen of breast cancer cells treated with or without exogenous prolactin. We hypothesise that the identification of these genes should yield insights into the mechanisms by which prolactin participates in cancer formation or progression, and possibly how it regulates normal mammary gland development. Methods We used subtractive hybridisation to identify a number of prolactin-regulated genes in the human mammary carcinoma cell line SKBR3. Northern blotting analysis and luciferase assays identified the gene encoding heat shock protein 90-alpha (HSP90A) as a prolactin-JAK2-STAT5 target gene, whose function was characterised using apoptosis assays. Results We identified a number of new prolactin-regulated genes in breast cancer cells. Focusing on HSP90A, we determined that prolactin increased HSP90A mRNA in cancerous human breast SKBR3 cells and that STAT5B preferentially activated the HSP90A promoter in reporter gene assays. Both prolactin and its downstream protein effector, HSP90α, promote survival, as shown by apoptosis assays and by the addition of the HSP90 inhibitor, 17-allylamino-17-demethoxygeldanamycin (17-AAG), in both untransformed HC11 mammary epithelial cells and SKBR3 breast cancer cells. The constitutive expression of HSP90A, however, sensitised differentiated HC11 cells to starvation-induced wild-type p53-independent apoptosis. Interestingly, in SKBR3 breast cancer cells, HSP90α promoted survival in the presence of serum but appeared to have little effect during starvation. Conclusions In addition to identifying new prolactin-regulated genes in breast cancer cells, we found that prolactin-JAK2-STAT5 induces expression of the HSP90A gene, which encodes the master chaperone of cancer. This identifies one mechanism by which prolactin contributes to breast cancer. Increased expression of HSP90A in breast cancer is correlated with increased cell survival and poor prognosis and HSP90α inhibitors are being tested in clinical trials as a breast cancer treatment. Our results also indicate that HSP90α promotes survival depending on the cellular conditions and state of cellular transformation

Mammary tumor formation and prostate hyperplasia in prolactin transgenic mice. Functional analysis of receptor specificity

Breast cancer and prostate hyperplasia are two very common diseases in humans, influenced by several different hormones, including sex hormones. The pituitary hormones growth hormone (GH) and prolactin (PRL) have also been demonstrated to have stimulatory effects on breast and prostate cells. It has even been speculated that GH and PRL might be important for the development of breast cancer and prostate hyperplasia in humans. Human-GH (hGH) can activate both GH-receptors (GHR) and PRL-receptors (PRLR) in humans and rodents. GHR and PRLR have been demonstrated in the mammary gland and prostate in several species, including humans. In order to study the potential role of GHR and PRLR stimulation in mammary tumor formation and prostate hyperplasia transgenic mice were used. Receptor specificity was investigated by using GHs and PRL from different species having specific binding to the GHR or the PRLR. Stimulation of both the GHR and the PRLR in hGH transgenic mice resulted in development of mammary adenocarcinomas at 7-9 months of age. To study the importance of selective GHR activation for tumor formation, bovine-GH (bGH) transgenic mice were used. No mammary tumors were observed in bGH transgenic mice displaying elevated levels of bGH and elevated insulin-like growth factor I (IGF-I). Rat-PRL (rPRL) transgenic mice were generated with the aim to selectively activate the PRLR. All of the rPRL transgenic female animals from two separate lines, displaying normal IGF-I levels, developed mammary adenocarcinomas at an age of 11-15 months. One of the lines had rPRL levels 4 times higher than normal peak values and the other levels in the normal range of endogenous PRL. Cell lines established from a tumor from a transgenic mouse expressing low serum levels of PRL produced rPRL and expressed PRLR. Furthermore, in organ culture experiments, an auto/paracrine function of rPRL produced from the transgene was demonstrated.Male rPRL transgenic mice developed prostate hyperplasia at an age of 10-15 months. The prostate weights were increased approximately 20 times and the DNA content 4-5 times compared with controls. Histologically, the prostate glands in the transgenic mice were distended from secretion and the amount of interstitial tissue was increased. The levels of testosterone and IGF-I were increased in the PRL transgenic animals. In bGH transgenic mice, displaying elevated IGF-I levels, the prostate gland was only slightly larger compared with normal mice, indicating that the effect of PRL was not primarily mediated through elevated plasma IGF-I levels. In summery, activation of the PRLR but not the GHR is important for induction of mammary tumors in female mice. Rat-PRL produced locally in the mammary gland from rPRL transgenic mice could stimulate mammary differentiation in a paracrine way in vitro, suggesting that locally produced PRL might be of importance for tumor development. In mice, PRL is involved in development of prostate hyperplasia, acting directly on the prostate or via increased plasma levels of testosterone

Development of pure prolactin receptor antagonists

Prolactin (PRL) promotes tumor growth in various experimental models and leads to prostate hyperplasia and mammary neoplasia in PRL transgenic mice. Increasing experimental evidence argues for the involvement of autocrine PRL in this process. PRL receptor antagonists have been developed to counteract these undesired proliferative actions of PRL. However, all forms of PRL receptor antagonists obtained to date exhibit partial agonism, preventing their therapeutic use as full antagonists. In the present study, we describe the development of new human PRL antagonists devoid of agonistic properties and therefore able to act as pure antagonists. This was demonstrated using several in vitro bioassays, including highly sensitive assays able to detect extremely low levels of receptor activation. These new compounds also act as pure antagonists in vivo, as assessed by analyzing their ability to competitively inhibit PRL-triggered signaling cascades in various target tissues (liver, mammary gland, and prostate). Finally, by using transgenic mice expressing PRL specifically in the prostate, which exhibit constitutively activated signaling cascades paralleling hyperplasia, we show that these new PRL analogs are able to completely revert PRL-activated events. These second generation human PRL antagonists are good candidates to be used as inhibitors of growth-promoting actions of PRL