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Eleftherios P Diamandis

Publications and source records attributed to Eleftherios P Diamandis.

At least 19 recordsLinked to original sources
PubMed2007

Mass spectrometry: uncovering the cancer proteome for diagnostics.

Despite impressive scientific achievements over the past few decades, cancer is still a leading cause of death. One of the major reasons is that most cancer patients are diagnosed with advanced disease. This is clearly illustrated with ovarian cancer in which the overall 5-year survival rates are only 20-30%. Conversely, when ovarian cancer is detected early (stage 1), the 5-year survival rate increases to 95%. Biomarkers, as tools for preclinical detection of cancer, have the potential to revolutionize the field of clinical diagnostics. The emerging field of clinical proteomics has found applications across a wide spectrum of cancer research. This chapter will focus on mass spectrometry as a proteomic technology implemented in three areas of cancer: diagnostics, tissue imaging, and biomarker discovery. Despite its power, it is also important to realize the preanalytical, analytical, and postanalytical limitations currently associated with this methodology. The ultimate endpoint of clinical proteomics is individualized therapy. It is essential that research groups, the industry, and physicians collaborate to conduct large prospective, multicenter clinical trials to validate and standardize this technology, for it to have real clinical impact.

Animals↗
PubMed2006

A potential role for multiple tissue kallikrein serine proteases in epidermal desquamation.

Desquamation of the stratum corneum is a serine protease-dependent process. Two members of the human tissue kallikrein (KLK) family of (chymo)tryptic-like serine proteases, KLK5 and KLK7, are implicated in desquamation by digestion of (corneo)desmosomes and inhibition by desquamation-related serine protease inhibitors (SPIs). However, the epidermal localization and specificity of additional KLKs also supports a role for these enzymes in desquamation. This study aims to delineate the probable contribution of KLK1, KLK5, KLK6, KLK13, and KLK14 to desquamation by examining their interactions, in vitro, with: 1) colocalized SPI, lympho-epithelial Kazal-type-related inhibitor (LEKTI, four recombinant fragments containing inhibitory domains 1-6 (rLEKTI(1-6)), domains 6-8 and partial domain 9 (rLEKTI(6-9')), domains 9-12 (rLEKTI(9-12)), and domains 12-15 (rLEKTI(12-15)), secretory leukocyte protease inhibitor, and elafin and 2) their ability to digest the (corneo)desmosomal cadherin, desmoglein 1. KLK1 was not inhibited by any SPI tested. KLK5, KLK6, KLK13, and KLK14 were potently inhibited by rLEKTI(1-6), rLEKTI(6-9'), and rLEKTI(9-12) with Ki values in the range of 2.3-28.4 nm, 6.1-221 nm, and 2.7-416 nm for each respective fragment. Only KLK5 was inhibited by rLEKTI(12-15) (Ki = 21.8 nm). No KLK was inhibited by secretory leukocyte protease inhibitor or elafin. Apart from KLK13, all KLKs digested the ectodomain of desmoglein 1 within cadherin repeats, Ca2+ binding sites, or in the juxtamembrane region. Our study indicates that multiple KLKs may participate in desquamation through cleavage of desmoglein 1 and regulation by LEKTI. These findings may have clinical implications for the treatment of skin disorders in which KLK activity is elevated.

Carrier Proteins↗
PubMed2006

Expression and functional characterization of the cancer-related serine protease, human tissue kallikrein 14.

Human tissue kallikrein 14 (KLK14) is a novel extracellular serine protease. Clinical data link KLK14 expression to several diseases, primarily cancer; however, little is known of its (patho)-physiological role. To functionally characterize KLK14, we expressed and purified recombinant KLK14 in mature and proenzyme forms and determined its expression pattern, specificity, regulation, and in vitro substrates. By using our novel immunoassay, the normal and/or diseased skin, breast, prostate, and ovary contained the highest concentration of KLK14. Serum KLK14 levels were significantly elevated in prostate cancer patients compared with healthy males. KLK14 displayed trypsin-like specificity with high selectivity for P1-Arg over Lys. KLK14 activity could be regulated as follows: 1) by autolytic cleavage leading to enzymatic inactivation; 2) by the inhibitory serpins alpha1-antitrypsin, alpha2-antiplasmin, antithrombin III, and alpha1-antichymotrypsin with second order rate constants (k(+2)/Ki) of 49.8, 23.8, 1.48, and 0.224 microM(-1) min(-1), respectively, as well as plasminogen activator inhibitor-1; and 3) by citrate and zinc ions, which exerted stimulatory and inhibitory effects on KLK14 activity, respectively. We also expanded the in vitro target repertoire of KLK14 to include collagens I-IV, fibronectin, laminin, kininogen, fibrinogen, plasminogen, vitronectin, and insulin-like growth factor-binding proteins 2 and 3. Our results indicate that KLK14 may be implicated in several facets of tumor progression, including growth, invasion, and angiogenesis, as well as in arthritic disease via deterioration of cartilage. These findings may have clinical implications for the management of cancer and other disorders in which KLK14 activity is elevated.

Amino Acid Sequence↗
PubMed2006

Proteolytic processing of human growth hormone by multiple tissue kallikreins and regulation by the serine protease inhibitor Kazal-Type5 (SPINK5) protein.

BACKGROUND: Human growth hormone (hGH) is naturally present in numerous isoforms, some of which arise from proteolytic processing in both the pituitary and periphery. The nature of the enzymes that proteolytically cleave hGH and the regulation of this process are not fully understood. Our objective is to examine if members of a newly discovered human tissue kallikrein family (KLKs) are expressed in the pituitary and if these enzymes can cleave hGH in-vitro. METHODS: Expression of 12 of the KLKs (KLKs 4-15) and serine protease inhibitor Kazal-type 5 (SPINK5) genes and their proteins in the pituitary was examined by RT-PCR and immunohistochemistry. Recombinant hGH was digested by various recombinant KLKs and fragments were characterized by N-terminal sequencing. SPINK5 recombinant fragments were used for inhibition of KLK activities. RESULTS: We here describe for the first time expression of numerous KLKs (KLKs 5-8, 10-14) and SPINK5 in the pituitary. KLK6 and SPINK5 appeared to be localized to hGH-producing cells. KLKs 4-6, 8, 13 and 14 were able to cleave hGH, yielding various isoforms, in vitro. Inhibitor SPINK5 fragments were able to suppress activity of KLKs 4, 5 and 14 in vitro. Based on these data, we propose a model for the proteolytic processing of hGH in the pituitary and the regulation of this system by SPINK5 inhibitory domains. We speculate that loss of SPINK5 inhibitory domains, as in the case of Netherton syndrome, may lead to proteolytic over-processing of hGH and to growth retardation. CONCLUSION: We conclude that many KLKs and SPINK5 are expressed in the pituitary. This serine protease-inhibitor system is likely to participate in the regulated proteolytic processing of hGH in the pituitary, leading to generation of hGH fragments. Our data suggest that KLKs 5, 6 and 14 might be involved in this process.

Carrier Proteins↗
PubMed2006

Development of an immunofluorometric assay for human kallikrein 15 (KLK15) and identification of KLK15 in tissues and biological fluids.

BACKGROUND: Human kallikrein 15 (KLK15) may have some utility as a prostate, ovarian, and breast cancer biomarker, based on previous studies, which examined mRNA levels of KLK15. The aim of this study was to develop analytical technology for human kallikrein 15, including recombinant protein, specific antibodies, and a sensitive and specific ELISA immunoassay. The assay was then used to examine levels of KLK15 in tissues and biological fluids. METHODS: We produced human, recombinant pro-KLK15 in HEK 293 cells. Recombinant KLK15 was purified with various chromatographic steps and used to immunize rabbits and mice for production of KLK15 polyclonal antibodies. We used these antibodies to develop a highly sensitive and specific KLK15 immunoassay and to study KLK15 expression in various tissues and biological fluids. RESULTS: Large amounts of pure, recombinant KLK15 have been produced and characterized. KLK15 mouse and rabbit polyclonal antibodies have been employed for development of a KLK15 immunoassay. This assay has a lower detection limit of 0.05 microg/L, and no cross-reactivity with any of the other fourteen kallikreins. Using this assay, KLK15 was detected in prostate, colon, and thyroid tissues, as well as in breast milk and seminal plasma. CONCLUSIONS: The KLK15 reagents developed here will allow for analysis of KLK15 protein expression levels in tissues and biological fluids, both normal and cancerous. This will expand upon previously characterized tissue KLK15 mRNA expression studies which suggested that KLK15 might be useful as a biomarker for breast, ovarian, and prostate cancer. KLK15 is another serine protease that is produced in prostate and other tissues and is secreted in seminal plasma and other fluids. Its physiological function needs to be further elucidated.

Amino Acid Sequence↗
PubMed2006

Unfavorable prognostic value of human kallikrein 7 quantified by ELISA in ovarian cancer cytosols.

BACKGROUND: Human tissue kallikrein 7 (gene, KLK7; protein, hK7) is a member of the kallikrein family of secreted serine proteases. Reports indicate that in ovarian cancer, KLK7 is significantly up-regulated at the mRNA level. The aim of this study was to determine whether hK7, measured quantitatively by ELISA in ovarian cancer cytosols, is a prognostic biomarker for ovarian cancer. METHODS: We used a newly developed ELISA with 2 monoclonal antibodies to quantify hK7 production in 260 ovarian tumor cytosols and correlated these data with various clinicopathologic variables and patient outcomes [progression-free survival (PFS) and overall survival (OS)] over a median follow-up period of 52 months. RESULTS: Median (range) hK7 concentration in ovarian tumor cytosols was 2.84 (0-32.8) ng/mg of total protein. Compared with healthy and benign ovarian tissues and nonovarian tumors that metastasized to the ovary, malignant ovarian tumor cytosols highly overproduced hK7 (P <0.001). We used the median value as the cutoff value to categorize tumors as hK7-positive and hK7-negative. Women with hK7-positive tumors most frequently had advanced-stage disease, higher tumor grade (G3), suboptimal debulking, and serous or undifferentiated histotype (P <0.001). Univariate analysis showed that hK7 positivity was associated with significantly shorter PFS (P = 0.01) but not OS. Kaplan-Meier survival curves confirmed an increased risk of relapse in women with hK7-positive tumors (P = 0.009). In multivariate analysis, hK7 was not significantly associated with either PFS or OS. CONCLUSIONS: hK7 is associated with other unfavorable characteristics of ovarian cancer, but it is not an independent prognosticator for ovarian cancer.

Adult↗
PubMed2006

Coordinated steroid hormone-dependent and independent expression of multiple kallikreins in breast cancer cell lines.

The regulation of gene expression by steroid hormones plays an important role in the normal development and function of many organs, as well in the pathogenesis of endocrine-related cancers. Previous experiments have shown that many kallikrein genes are under steroid hormone regulation in breast cancer cell lines. We here examine the coordinated expression of multiple kallikrein genes in several breast cancer cell lines after steroid hormone stimulation. Breast cancer cell lines were treated with various steroid hormones and kallikrein (KLK/hK) expression of hK3 (prostate-specific antigen, PSA), hK5, hK6, hK7, hK8, hK10, hK11, hK13, and hK14 was analyzed at the RNA level via RT-PCR and at the protein level by immunofluorometric ELISA assays. We identified several distinct hK hormone-dependent and hormone-independent expression patterns. Hormone-specific modulation of expression was seen for several kallikreins in BT-474, MCF-7, and T-47D cell lines. hK6 was specifically up-regulated upon estradiol treatment in all three cell lines whereas PSA expression was induced by dihydrotestosterone (DHT) and norgestrel stimulation in BT-474 and T-47D. hK10, hK11, hK13, and hK14 were specifically up-regulated by DHT in T-47D and by estradiol in BT-474 cells. Bioinformatic analysis of upstream proximal promoter sequences for these hKs did not identify any recognizable hormone-response elements (HREs), suggesting that the coordinated activation of these four hKs represents a unique expression "cassette", utilizing a common hormone-dependent mechanism. We conclude that groups of human hKs are coordinately expressed in a steroid hormone-dependent manner. Our data supports clinical observations linking expression of multiple hKs with breast cancer prognosis.

Breast Neoplasms↗
PubMed2006

Proteinase-activated receptors, targets for kallikrein signaling.

Serine proteinases like thrombin can signal to cells by the cleavage/activation of proteinase-activated receptors (PARs). Although thrombin is a recognized physiological activator of PAR(1) and PAR(4), the endogenous enzymes responsible for activating PAR(2) in settings other than the gastrointestinal system, where trypsin can activate PAR(2), are unknown. We tested the hypothesis that the human tissue kallikrein (hK) family of proteinases regulates PAR signaling by using the following: 1) a high pressure liquid chromatography (HPLC)-mass spectral analysis of the cleavage products yielded upon incubation of hK5, -6, and -14 with synthetic PAR N-terminal peptide sequences representing the cleavage/activation motifs of PAR(1), PAR(2), and PAR(4); 2) PAR-dependent calcium signaling responses in cells expressing PAR(1), PAR(2), and PAR(4) and in human platelets; 3) a vascular ring vasorelaxation assay; and 4) a PAR(4)-dependent rat and human platelet aggregation assay. We found that hK5, -6, and -14 all yielded PAR peptide cleavage sequences consistent with either receptor activation or inactivation/disarming. Furthermore, hK14 was able to activate PAR(1), PAR(2), and PAR(4) and to disarm/inhibit PAR(1). Although hK5 and -6 were also able to activate PAR(2), they failed to cause PAR(4)-dependent aggregation of rat and human platelets, although hK14 did. Furthermore, the relative potencies and maximum effects of hK14 and -6 to activate PAR(2)-mediated calcium signaling differed. Our data indicate that in physiological settings, hKs may represent important endogenous regulators of the PARs and that different hKs can have differential actions on PAR(1), PAR(2), and PAR(4).

Amino Acid Sequence↗
PubMed2006

Characterization of human kallikreins 6 and 10 in ascites fluid from ovarian cancer patients.

OBJECTIVES: Human kallikreins 6 (hK6) and 10 (hK10) are secreted serine proteases. We previously found that hK6 and hK10 are highly overexpressed in epithelial ovarian tumors and demonstrated that serum levels of hK6 and hK10 are valuable biomarkers for ovarian cancer diagnosis and prognosis. Our aim is to purify and characterize these two kallikreins from ascites fluid of ovarian cancer patients. METHODS: Protein concentrations of hK6 and hK10 in ovarian cancer ascites fluids were measured with ELISA-type immunoassays. hK6 and hK10 were purified from the ascites fluids with immunoaffinity columns, followed by reverse-phase high performance liquid chromatography. Purified hK6 and hK10 were then subjected to N-terminal sequencing. Enzymatic analyses were performed with synthetic fluorogenic peptides. RESULTS: hK6 and hK10 were present in ovarian cancer ascites fluid at concentrations ranging from 0.2-571 and 0.7-220 microg/l, respectively. The majority of hK6 and hK10 in the ascites fluids were present in the free (uncomplexed) form. Both hK6 and hK10 purified from the ascites fluid were zymogens with a molecular mass of 30 kDa. Purified hK6 exhibited trypsin-like enzymatic activity, whereas no enzymatic activity was observed for purified hK10. The enzymatic activity of hK6 could be suppressed by a neutralizing monoclonal antibody. CONCLUSIONS: The majority of hK6 secreted by the ovarian tumor cells into the ascites fluid are present in the uncomplexed, zymogen form, possessing weak trypsin-like enzymatic activity. All hK10 present in ovarian cancer ascites fluids are in the uncomplexed, zymogen form and have no detectable enzymatic activity.

Amino Acid Sequence↗
PubMed2006

In silico identification and Bayesian phylogenetic analysis of multiple new mammalian kallikrein gene families.

Kallikrein gene families have been identified previously in genomes of the human, the mouse, and the rat, and individual kallikrein-like genes have been found in many more species. This study presents the in silico identification of kallikrein gene families in the recently sequenced genomes of four additional mammalian species, the chimpanzee, the dog, the pig, and the opossum. Phylogenies were constructed with gene sequences from all seven mammalian families, using Bayesian analysis, which clarified the evolutionary relationships between these genes. Individual gene sequences, as well as concatenated constructs of multiple sequences, were used. Fifteen kallikrein genes were located in the chimpanzee (Pan troglodytes) genome, while only 14 were identified in the canine (Canis familiaris) genome as no orthologue to human KLK3 was found. Thirteen genes were identified from the pig (Sus scrofa) genome, which lacked homologues to KLK2 and KLK3, and 11 genes, orthologous to human KLK5 through KLK15, were found in the opossum (Monodelphis domestica) genome. No kallikrein genes were identified from the available genome sequences of the chicken (Gallus gallus) or African clawed frog (Xenopus tropicalis). Within the family of kallikreins several subfamilies were suggested by phylogenetic analysis. One consisted of KLK4, KLK5, and KLK14; another of KLK9, KLK11, and KLK15; a third of KLK10 and KLK12; a fourth of KLK6 and KLK13; and finally one of KLK8 and the classical kallikreins (KLK1, KLK2, and KLK3).

Animals↗
PubMed2006

Effect of testosterone administration on serum and urine kallikrein concentrations in female-to-male transsexuals.

BACKGROUND: Concentrations of human tissue kallikreins (hKs), a group of 15 secreted serine proteases found in many tissues, are modulated by steroid hormones in cancer cell lines. To gain insight into in vivo kallikrein regulation we measured kallikrein concentrations in serum and urinary tissue in female-to-male transsexuals before and after testosterone administration. METHODS: We collected blood and urine samples before treatment and after 4 and 12 months from 28 female-to-male transsexuals who received 250 mg of testosterone esters intramuscularly every 2 weeks. We used ELISA assays to measure multiple kallikreins in serum and urine. RESULTS: After testosterone administration, serum testosterone concentrations increased by approximately 15-fold. Serum kallikrein concentrations increased dramatically for hK3 (prostate-specific antigen) and increased moderately for hK2, hK5, hK6, hK7, hK8, hK10, and hK11. In urine, we noted major increases for hK3 and hK2 only. For all other kallikrein concentrations, we observed no considerable changes. CONCLUSIONS: We conclude that, in serum and urine of female-to-male transsexuals after testosterone administration, hK3 (prostate-specific antigen) and to a lesser extent hK2 concentrations increase dramatically, but concentration of other kallikreins increase either moderately in serum (hK5, hK6, hK7, hK8, hK10, and hK11) or not at all in either serum (hK4, hK13, hK14) or urine (hK4, hK5, hK6, hK7, hK8, hK10, hK11, hK13, hK14).

Adolescent↗
PubMed2006

Three new serum markers for prostate cancer detection within a percent free PSA-based artificial neural network.

BACKGROUND: We aimed to evaluate the value of macrophage inhibitory cytokine 1 (MIC-1), human kallikrein 11 (hK11) migration inhibitor factor (MIF) in comparison to prostate-specific antigen (PSA) and % fPSA and also to develop a % fPSA-based ANN with the new input factors to determine whether these additional markers can further eliminate unnecessary prostate biopsies. METHODS: Serum samples from 371 patients with prostate cancer (PCa, n=135) or benign prostate hyperplasia (BPH, n=236) within the PSA range 0.5-20 microg/L were analyzed for total PSA, free PSA, MIC-1, hK11, and MIF. 'Leave one out' ANN models with these variables and prostate volume were constructed and compared to logistic regression (LR) and all single parameters. RESULTS: The discriminatory power of MIC-1, hK11, and MIF was less than that for PSA despite significant differences in BPH compared to PCa patients. At 90% and 95% sensitivity, the artificial neural networks (ANNs) were only significantly better than % fPSA if prostate volume was included. CONCLUSIONS: ANNs with the novel input factors of MIC-1, MIF, and/or hK11 and additional use of prostate volume demonstrated significant advantage compared with % fPSA and tPSA and may lead to a reduction in unnecessary prostate biopsies.

Biomarkers, Tumor↗
PubMed2006

Serum and urine tissue kallikrein concentrations in male-to-female transsexuals treated with antiandrogens and estrogens.

BACKGROUND: The expression of human tissue kallikrein genes is regulated by steroid hormones, but most studies have been conducted with cancer cell lines. Our purpose was to examine serum and urinary tissue kallikrein concentration changes in male-to-female transsexuals before and after treatment with antiandrogens and estrogens. METHODS: Thirty-five male-to-female transsexuals receiving cyproterone acetate and estrogens (orally or transdermally) were included in this study. Serum and urine samples were collected before initiation of therapy and 4 and 12 months post therapy. ELISAs were used to measure multiple kallikreins in serum and urine. RESULTS: After antiandrogen and estrogen therapy, serum testosterone concentrations decreased dramatically, as did serum and urinary concentrations of human glandular kallikrein (hK2) and prostate-specific antigen (PSA; hK3). Statistically significant but relatively small changes in serum and urinary concentrations of many other kallikreins were also seen. Kallikreins in serum and urine were correlated before and after treatment. CONCLUSIONS: The concentrations of hK2 and hK3, but not of any other kallikreins, decrease dramatically after combined antiandrogen and estrogen treatment in male-to-female transsexuals. The smaller responses of the other kallikreins presumably reflect their expression in multiple tissues.

Adult↗
PubMed2006

Overexpression of kallikrein 10 (hK10) in uterine serous papillary carcinomas.

OBJECTIVE: Kallikrein 10 is a secreted serine protease recently implicated in the growth and invasion of several human tumors. The goal of this study was to investigate the expression and secretion levels in vitro and in vivo of kallikrein 10 in uterine serous papillary carcinoma, a highly aggressive variant of endometrial tumor. STUDY DESIGN: Human kallikrein 10 gene expression levels were evaluated in 11 snap-frozen uterine serous papillary carcinoma biopsies and 6 normal endometrial cell biopsies by real-time polymerase chain reaction. Secretion of kallikrein 10 protein by 10 primary tumor cultures including 3 uterine serous papillary carcinomas, 2 endometrioid carcinomas, and 5 ovarian serous papillary tumors was measured using a sensitive ELISA. Finally, kallikrein 10 concentration in 75 serum and plasma samples from 22 healthy women, 20 women with benign diseases, 21 women with endometrioid carcinomas, and 12 uterine serous papillary carcinoma patients was studied. RESULTS: Kallikrein 10 gene expression levels were significantly higher in uterine serous papillary carcinoma when compared with normal endometrial cell biopsies (mean copy number by real time polymerase chain reaction = 743 versus 1.4; uterine serous papillary carcinoma versus endometrioid carcinoma: P < .02). In vitro kallikrein 10 secretion was detected in all primary uterine serous papillary carcinoma cell lines tested (mean = 2.7 microg/L), and the secretion levels were not significantly different to those found in primary ovarian serous papillary tumor cultures (mean 4.2 microg/L). In contrast, no kallikrein 10 secretion was detectable in primary endometrioid carcinomas. Kallikrein 10 serum and plasma concentrations (microg/L; mean +/- SEM) among normal healthy females (0.6 +/- 0.04), patients with benign diseases (0.6 +/- 0.06), and patients with endometrioid carcinomas (0.7 +/- 0.06) were not significantly different. In contrast, serum and plasma kallikrein 10 values in uterine serous papillary carcinoma patients (1.2 +/- 0.1) were significantly higher than those in the non-cancer group (P = .002), benign group (P = .002), and endometrioid carcinoma patients (P = .005). CONCLUSION: Kallikrein 10 is highly expressed in uterine serous papillary carcinoma, and it is released in the plasma and serum of uterine serous papillary carcinoma patients. Kallikrein 10 may represent a novel biomarker for uterine serous papillary carcinoma.

Adult↗
PubMed2006

Effects of long-term androgen administration on breast tissue of female-to-male transsexuals.

Our aim was to examine the effects of androgen administration on breast tissue histology of female-to-male transsexuals and to study the immunohistochemical expression of three human tissue kallikreins, hK3 (PSA), hK6, and hK10. We studied 23 female-to-male transsexuals who were treated with injectable testosterone for 18-24 months. We also used 10 control female breast tissues. All tissues were fixed in buffered formalin, embedded in paraffin, and examined by hematoxylin-eosin staining and immunohistochemical staining for PSA, hK6, and hK10. Females treated with androgens exhibited similar involutionary changes as those seen in breast of menopausal women, such as marked reduction of glandular tissue, involution of the lobuloalveolar structures, and prominence of fibrous connective tissue, but presence of only small amounts of fat tissue. Fibrocystic lesions were generally not observed. In immunohistochemistry, in control breast tissues, we found moderate to strong cytoplasmic immunoexpression of hK6 and hK10 in the epithelial ductal and lobuloalveolar structures, but myoepithelial cells were negative. Luminal secretions were also positive. In menopausal breast, the immunoexpression of hK6 and hK10 was weaker and focal. No control case showed immunoexpression for PSA. In female-to-male transsexuals, one case showed focal PSA cytoplasmic immunoexpression in the epithelium of moderately involuting lobules. Long-term administration of androgens in female-to-male transsexuals causes marked reduction of glandular tissue and prominence of fibrous connective tissue. These changes are similar to those observed at the end-stage of menopausal mammary involution.

Androgens↗