Adipokines expression and epithelial cell polarity in normal and cancerous breast tissue.

Cell polarity is crucial for the correct structural and functional organization of epithelial tissue. Its disruption can lead to loss of the apicobasal polarity, alteration in the intracellular components, misregulation of the pathways involved in cell proliferation and cancer promotion. Very recent in vitro/in vivo findings demonstrated that obesity-associated alterations in tissue adipokines protein level negatively affect epithelial polarity. We performed an in silico study to investigate whether such alterations also occur in surgical samples. We aimed to explore the relationship among the expression of the genes coding for leptin (LEP), adiponectin (ADIPOQ), adipokine receptors (LEPR, ADIPOR1 and ADIPOR2), and a panel of polarity-associated genes in normal tissue from breast reduction mammoplasty, and a series of paired samples of histologically normal (HN) tissue and invasive cancer. Results indicated that, in normal tissue, the expression of adipokines and their receptors negatively correlated with that of the polarity-associated genes and GGT1, which codes for γ-glutamyl transferase (GGT) enzyme, a marker of cell distress and membrane disruption. This negative correlation progressively decreased in HN and cancerous tissue, and loss of correlation between ADIPOR2 and polarity-associated genes appeared the most noticeable alteration. Given the growing role of obesity in breast cancer etiology and the opposite action of leptin and adiponectin in epithelial tissue remodeling, ADIPOR2 loss could be addressed as a key mechanism leading to an unbalanced leptin stimulatory activity, subsequent cell polarity disruption and eventually tumor initiation, a finding that requires to be confirmed also at the protein level and with in vivo models.

Correction to: Observational study on the prognostic value of testosterone and adiposity in postmenopausal estrogen receptor positive breast cancer patients.

Following publication of the original article [1], the authors reported that the affiliation of author Annalisa Orenti was omitted.

Observational study on the prognostic value of testosterone and adiposity in postmenopausal estrogen receptor positive breast cancer patients.

BACKGROUND: Despite the clear endocrine-metabolic relationship between androgenic activity and adiposity, the role of androgens in breast cancer prognosis according to patient's adiposity is scarcely explored. Here, we aimed at investigating the prognostic value of circulating testosterone in association with patient's body mass index (BMI). METHODS: Circulating testosterone and BMI were evaluated at breast cancer diagnosis in 460 estrogen receptor (ER)-positive postmenopausal patients. Local relapse, distant metastasi(e)s and contralateral breast cancer were considered recurrence events. The Kruskal-Wallis test was performed to evaluate if testosterone levels differed within subgroups of categorical tumour characteristics. The Cox proportional hazard regression model was fitted to estimate the impact of standard prognostic factors on relapse-specific hazard ratio (HR). After backward selection, a model including continuous testosterone level, BMI categories (< 25, normal-weight; =25-30, overweight; ≥30 kg/m2, obese), tumour size and lymph nodes number was fitted. Furthermore, Cox models provided the relapse-specific HRs for median, third quartile and 95th percentile compared to the first quartile of testosterone levels, stratified by BMI categories. RESULTS: During a median follow up of 6.3 years, 45 patients relapsed. Testosterone levels significantly increased across BMI categories (p = 0.001). Both circulating testosterone and BMI were positively associated with disease free survival (p = 0.005 and p = 0.021, respectively). A significant interaction was found between testosterone and BMI (p = 0.006). For normal-weight women, testosterone concentration around median (0.403 ng/mL) or third quartile (0.532 ng/mL) showed a high significant HR of relapse (5.52; 95% CI:1.65-18.49 and 4.55; 95% CI:1.09-18.98, respectively). Overweight patients showed increased HR at increasing testosterone levels, reaching a significant high HR (4.68; 95% CI:1.39-15.70) for testosterone values of 0.782 ng/mL (95th percentile). For obese patients HR decreased (not significantly) at increased testosterone concentrations, explaining the interaction between testosterone levels and BMI categories. CONCLUSIONS: In ER-positive postmenopausal breast cancer patients, high testosterone levels are associated with worse prognosis in normal-weight and overweight women, whereas in obese seems to be associated with a better outcome. Although the results require further validation, they suggest that assessment of circulating testosterone and BMI could help to identify postmenopausal ER-positive patients at higher risk of relapse and potentially open new therapeutic strategies.

Serum levels of testosterone and SHBG in association with body mass index improve the predictive capability of consolidate tumor biomarkers in pre- and postmenopausal breast cancer patients.

OBJECTIVE: To investigate the contribution of serum levels of testosterone (TS) and sex hormone binding globulin (SHBG) in association with body mass index (BMI) as a surrogate marker of obesity, to the predictive capability of tumor size (T), lymph node (N) and estrogen receptor (ER) status and proliferative activity (TLI). METHODS: We investigated 120 women with primary breast cancer and median follow-up of 138 months. Serum levels of TS and SHBG and patient's BMI were evaluated before surgery. The contribution of TS, SHBG, their ratio (TS/SHBG) and BMI to the predictive capability of tumor-specific biomarkers was investigated by Harrell's c statistic. RESULTS: TS alone did not affect prognosis, whereas SHBG was protective in postmenopausal patients, in which BMI was associated with a progressive increase in the relapse-specific hazard ratio (HR). When in combination, TS, SHBG and BMI, affected prognosis in different ways depending on menopausal status. The best predictive capability (c = 0.78) was observed in postmenopausal patients when at the basic model (N + TLI) were added TS, BMI, TS * BMI interaction, with or without SHBG. In premenopause subgroup, the best predictive capability (c = 0.67) was provided by the basic model (N + TLI) plus TS and SHBG or their ratio, BMI and TS * BMI or TS/SHBG * BMI interaction. CONCLUSIONS: Patient-associated features such as BMI and serum levels of TS and SHBG can improve the predictive capability of consolidate tumor-specific biomarkers in both pre- and postmenopause, thus providing a relevant contribution to the decision-making process.

The role of maintenance proteins in the preservation of epithelial cell identity during mammary gland remodeling and breast cancer initiation.

During normal postnatal mammary gland development and adult remodeling related to the menstrual cycle, pregnancy, and lactation, ovarian hormones and peptide growth factors contribute to the delineation of a definite epithelial cell identity. This identity is maintained during cell replication in a heritable but DNA-independent manner. The preservation of cell identity is fundamental, especially when cells must undergo changes in response to intrinsic and extrinsic signals. The maintenance proteins, which are required for cell identity preservation, act epigenetically by regulating gene expression through DNA methylation, histone modification, and chromatin remodeling. Among the maintenance proteins, the Trithorax (TrxG) and Polycomb (PcG) group proteins are the best characterized. In this review, we summarize the structures and activities of the TrxG and PcG complexes and describe their pivotal roles in nuclear estrogen receptor activity. In addition, we provide evidence that perturbations in these epigenetic regulators are involved in disrupting epithelial cell identity, mammary gland remodeling, and breast cancer initiation.

Differential expression of genes involved in the epigenetic regulation of cell identity in normal human mammary cell commitment and differentiation.

The establishment and maintenance of mammary epithelial cell identity depends on the activity of a group of proteins, collectively called maintenance proteins, that act as epigenetic regulators of gene transcription through DNA methylation, histone modification, and chromatin remodeling. Increasing evidence indicates that dysregulation of these crucial proteins may disrupt epithelial cell integrity and trigger breast tumor initiation. Therefore, we explored in silico the expression pattern of a panel of 369 genes known to be involved in the establishment and maintenance of epithelial cell identity and mammary gland remodeling in cell subpopulations isolated from normal human mammary tissue and selectively enriched in their content of bipotent progenitors, committed luminal progenitors, and differentiated myoepithelial or differentiated luminal cells. The results indicated that, compared to bipotent cells, differentiated myoepithelial and luminal subpopulations were both characterized by the differential expression of 4 genes involved in cell identity maintenance: CBX6 and PCGF2, encoding proteins belonging to the Polycomb group, and SMARCD3 and SMARCE1, encoding proteins belonging to the Trithorax group. In addition to these common genes, the myoepithelial phenotype was associated with the differential expression of HDAC1, which encodes histone deacetylase 1, whereas the luminal phenotype was associated with the differential expression of SMARCA4 and HAT1, which encode a Trithorax protein and histone acetylase 1, respectively. The luminal compartment was further characterized by the overexpression of ALDH1A3 and GATA3, and the down-regulation of NOTCH4 and CCNB1, with the latter suggesting a block in cell cycle progression at the G2 phase. In contrast, myoepithelial differentiation was associated with the overexpression of MYC and the down-regulation of CCNE1, with the latter suggesting a block in cell cycle progression at the G1 phase.

Impact of De Novo Cholesterol Biosynthesis on the Initiation and Progression of Breast Cancer.

Cholesterol (CHOL) is a multifaceted lipid molecule. It is an essential structural component of cell membranes, where it cooperates in regulating the intracellular trafficking and signaling pathways. Additionally, it serves as a precursor for vital biomolecules, including steroid hormones, isoprenoids, vitamin D, and bile acids. Although CHOL is normally uptaken from the bloodstream, cells can synthesize it de novo in response to an increased requirement due to physiological tissue remodeling or abnormal proliferation, such as in cancer. Cumulating evidence indicated that increased CHOL biosynthesis is a common feature of breast cancer and is associated with the neoplastic transformation of normal mammary epithelial cells. After an overview of the multiple biological activities of CHOL and its derivatives, this review will address the impact of de novo CHOL production on the promotion of breast cancer with a focus on mammary stem cells. The review will also discuss the effect of de novo CHOL production on in situ and invasive carcinoma and its impact on the response to adjuvant treatment. Finally, the review will discuss the present and future therapeutic strategies to normalize CHOL biosynthesis.

Interaction of de novo cholesterol biosynthesis and Hippo signaling pathway in ductal carcinoma in situ (DCIS)-comparison with the corresponding normal breast epithelium.

Background: Ductal carcinoma in situ (DCIS) is a non-obligate precursor to invasive breast cancer. However, if left untreated, about 50% of DCIS progress. Preventing such a progression is of paramount importance. Cumulative evidence indicated that the mevalonate cascade, the core of cholesterol biosynthesis, contributes to the regulation of the Hippo signaling pathway providing the isoprenoids required for GTPase activation, the nuclear accumulation of the Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ) coactivator, and the subsequent gene transcription and that the disruption of this cooperation associated with tumor progression. Methods: In this in silico study, we investigated whether such a disruption occurred already during the transformation of the normal mammary epithelium into DCIS. To this aim, we interrogated a publicly available dataset, and we explored the interrelationship of the genes involved in the de novo cholesterol biosynthesis and the association with those coding for the core components of the Hippo signaling pathway in a set of patient-matched samples of DCIS and corresponding histologically normal (HN) epithelium. Results: Most genes involved in cholesterol biosynthesis were more expressed in DCIS than in the corresponding HN epithelium. This differential expression was associated with a substantial change in their correlation profile. In particular, 3-hydroxy-3-methylglutaryl coenzyme-A reductase (HMGCR) and INSIG1 lost the positive association shown in the HN epithelium, and their negative association with LSS switched to a positive one. Also, GGPS1, which plays a crucial role in isoprenoids production, significantly changed its correlation profile. The positive association between GGPS1 and HMGCR or INSIG1 disappeared, whereas the positive association with SQLE, which drives the irreversible commitment to cholesterol, switched to a negative one in DCIS. Conclusions: Present findings corroborated the hypothesis that a dysfunctional mevalonate pathway possibly concurs with DCIS development by leading to abnormal production of isoprenoids, which in turn activate GTPases and promote YAP/TAZ nuclear translocation, and suggested the safe and low-cost treatment with statins as the possible winning strategy to contrast this metabolic dysfunction.

TP53 mutation, epithelial-mesenchymal transition, and stemlike features in breast cancer subtypes.

Altered p53 protein is prevalently associated with the pathologic class of triple-negative breast cancers and loss of p53 function has recently been linked to the induction of an epithelial-mesenchymal transition (EMT) and acquisition of stemness properties. We explored the association between TP53 mutational status and expression of some genes involved in the canonical TGF-ß signaling pathway (the most potent EMT inducer) and in two early EMT associated events: loss of cell polarity and acquisition of stemness-associated features. We used a publicly accessible microarray dataset consisting of 251 p53-sequenced primary breast cancers. Statistical analysis indicated that mutant p53 tumors (especially those harboring a severe mutation) were consistent with the aggressive class of triple-negative cancers and that, differently from cell cultures, surgical tumors underexpressed some TGF-ß related transcription factors known as involved in EMT (ID1, ID4, SMAD3, SMAD4, SMAD5, ZEB1). These unexpected findings suggest an interesting relationship between p53 mutation, mammary cell dedifferentiation, and the concomitant acquisition of stemlike properties (as indicated by the overexpression of PROM1 and NOTCH1 genes), which improve tumor cells aggressiveness as indicated by the overexpression of genes associated with cell proliferation (CDK4, CDK6, MKI67) and migration (CXCR4, MMP1).

Cytoplasmic Trop-1/Ep-CAM overexpression is associated with a favorable outcome in node-positive breast cancer.

OBJECTIVE: Trop-1/Ep-CAM modulates growth and survival of transformed cells, and it is highly expressed in most carcinomas including breast cancer. Only membranous staining is typically considered in evaluating Trop-1/epithelial cell adhesion molecule (Ep-CAM) expression in tumor cells. However, there is evidence of retention of Trop-1/Ep-CAM, as functionally incompetent molecules, in intra-cytoplasmic vesicles. Hence, we investigated whether cytoplasmic immunostaining may have an independent clinical significance with respect to membranous staining. METHODS: Membranous and cytoplasmic Trop-1/Ep-CAM expression was immunohistochemically investigated in 642 unilateral breast cancers from patients with a 99-month median follow-up. Multiple correspondence analysis was used to investigate the association between Trop-1/Ep-CAM and other biological variables. The impact of Trop-1/Ep-CAM expression on the patient's outcome was evaluated as event-free survival by the Kaplan-Meier method and proportional hazard Cox model. RESULTS: While tumors with intermediate/strong membranous staining were mostly associated with concomitant cytoplasmic Trop-1/Ep-CAM expression (97%), tumors with weak-to-nil membranous staining showed intermediate/high cytoplasmic expression in 23% of cases. Cytoplasmic overexpression was associated with a favorable outcome, especially in node-positive patients, regardless of the adjuvant therapy received. CONCLUSION: Trop-1/Ep-CAM expression may have different clinical implications according to its subcellular localization.

De novo cholesterol biosynthesis: an additional therapeutic target for the treatment of postmenopausal breast cancer with excessive adipose tissue.

The onset and development of breast cancer in postmenopausal women are associated with closely related individual-dependent factors, including weight gain and high levels of circulating androgens. Adipose tissue is the most peripheral site of aromatase enzyme synthesis; therefore, the excessive accumulation of visceral fat results in increased androgens aromatization and estradiol production that provides the microenvironment favorable to tumorigenesis in mammary epithelial cells expressing estrogen receptors (ERs). Moreover, to meet the increased requirement of cholesterol for cell membrane assembly and the production of steroid hormones to sustain their proliferation, ER-positive cells activate de novo cholesterol biosynthesis and subsequent steroidogenesis. Several approaches have been followed to neutralize the de novo cholesterol synthesis, including specific enzyme inhibitors, statins, and, more recently, metformin. Cumulating evidence indicated that inhibiting cholesterol biosynthesis by statins and metformin may be a promising therapeutic strategy to block breast cancer progression. Unlike antiestrogens and aromatase inhibitors (AIs) which compete for binding to ER and inhibit androgens aromatization, respectively, statins block the production of mevalonic acid by inhibiting the activity of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, and metformin hampers the activation of the sterol regulatory element-binding protein 2 (SREBP2) transcription factor, thus inhibiting the synthesis of several enzymes involved in cholesterol biosynthesis. Noteworthy, statins and metformin not only improve the prognosis of overweight patients with ER-positive cancer but also improve the prognosis of patients with triple-negative breast cancer, the aggressive tumor subtype that lacks, at present, specific therapy.

Differential Expression of the Genes Coding for Adipokines and Epithelial Cell Polarity Components in Women With Low and High Mammographic Density.

BACKGROUND: Women with extensive mammographic density (MD) are more likely to develop breast cancer than women with low MD because of a high epithelial component associated with a high proportion of stromal cells. To elucidate the biological association between high MD and risk of breast cancer, we compared the expression of a panel of genes coding for leptin, adiponectin, and some component of cell polarity and adherens junction complexes in dense and non-dense breast tissue. METHODS: We interrogated a public dataset composed by 120 specimens of normal breast tissue with MD evaluation. The differential expression of the selected genes in the 2 MD subgroups was assessed by the Wilcoxon test, whereas Kruskal-Wallis test evaluated the differential expression of single genes in the fatty, epithelium, or nonfatty compartment. Spearman's correlation measured the relationship among genes in the subset with the highest epithelium proportion. RESULTS: In high MD, the expression level of PARD6B, CRB3, PATJ, LLGL2, CDH1, and MARVELD2 significantly lowered in tissues with the highest epithelium proportion, whereas, in low MD, the expression level of the genes increased with the increasing of the epithelium proportion. In the low MD subgroup, LEP correlated negatively with PRKCZ and DLG3, whereas, in high MD, such correlation was not observed. CONCLUSIONS: The expression of the genes governing cell polarity establishment and cell-cell adhesion assembly differed significantly in the epithelial component of dense and non-dense breasts. The correlation pattern between LEP and PRKCZ or DLG3 agrees with the role of leptin in cell polarity disruption.

p53 status identifies two subgroups of triple-negative breast cancers with distinct biological features.

OBJECTIVE: Despite the clinical similarities triple-negative and basal-like breast cancer are not synonymous. Indeed, not all basal-like cancers are negative for estrogen receptor, progesterone receptor and HER2 expression while triple-negative also encompasses other cancer types. P53 protein appears heterogeneously expressed in triple-negative breast cancers, suggesting that it may be associated with specific biological subgroups with a different outcome. METHODS: We comparatively analyzed p53 expression in triple-negative tumors from two independent breast cancer case series (633 cases from the University of Ferrara and 1076 cases from the University of Nottingham). RESULTS: In both case series, p53 protein expression was able to subdivide the triple-negative cases into two distinct subsets consistent with a different outcome. In fact, triple-negative patients with a p53 expressing tumor showed worse overall and event-free survival. CONCLUSIONS: The immunohistochemical evaluation of p53 expression may help in taming the currently stormy relationship between pathological (triple-negative tumors) and biological (basal breast cancers) classifications and in selecting patient subgroups with different biological features providing a potentially powerful prognostic contribution in triple-negative breast cancers.

Epithelial cell polarity and tumorigenesis: new perspectives for cancer detection and treatment.

Loss of cell-cell adhesion and cell polarity is commonly observed in tumors of epithelial origin and correlates with their invasion into adjacent tissues and formation of metastases. Growing evidence indicates that loss of cell polarity and cell-cell adhesion may also be important in early stage of cancer. In first part of this review, we delineate the current understanding of the mechanisms that establish and maintain the polarity of epithelial tissues and discuss the involvement of cell polarity and apical junctional complex components in tumor pathogenesis. In the second part we address the clinical significance of cell polarity and junctional complex components in cancer diagnosis and prognosis. Finally, we explore their potential use as therapeutic targets in the treatment of cancer.

Impact of sex hormones dysregulation and adiposity on the outcome of postmenopausal breast cancer patients.

Epidemiological studies demonstrated that, in postmenopausal women, high circulating levels of testosterone, especially when associated with weight gain, positively correlated with an increased risk of breast cancer because of the augmented production of oestrogen via testosterone aromatization in the adipose tissue. Besides, growing evidence suggests that sulfatase can increase the tissue concentration of bioactive estradiol through the reconversion of estrone sulfate, thus providing a favourable milieu for epithelial cells expressing the oestrogen receptor. In this review, we will discuss how the "obesity-insulin-testosterone" connection and the abnormal production of bioactive oestrogen - as a result of the conversion of the androgens by aromatase and the estrone reconversion by sulfatase-, may affect the response to hormone therapy and the outcome of postmenopausal breast cancer patients, and how a combined therapy including metformin, anti-inflammatory drugs, and aromatase/sulfatase inhibitors could successfully improve patient's outcome.

Gene expression profile of normal breast tissue and body mass index.

BACKGROUND: In human breast, adipose tissue represents up to 80% of the total volume and plays a critical role in mammary gland remodeling. Given the emerging role of obesity in breast cancer growth and development, we explored the relationship between body mass index (BMI), as a proxy of woman's obesity status, and the expression in normal breast tissue from healthy women of a selected panel of genes, known to be involved in mammary gland homeostasis. METHODS: Two independent publicly available datasets, composed of 180 specimens of normal breast tissue from reduction mammoplasty were interrogated. Differential gene expression among BMI classes was evaluated by ANOVA, and partial correlation coefficient was used to assay the correlation between genes controlling for BMI. RESULTS: Despite the differences in microarray platforms and analytical procedures, the two datasets shared a core of 9 genes differentially expressed in BMI classes and significantly correlated with BMI. Four (44%) of these genes belong to the functional class of cytokines and cytokine receptors (IL1R1, IL2RA, IL12A, and IL12RB2). The others belong to the functional class of the epigenetic regulation (MEDAG and SETD7), signal transduction (STAT1), cell adhesion (ITGAV), and enzymatic activity (STS). CONCLUSIONS: Although exploratory, present findings are in agreement with the role of inflammation modulators in the homeostasis of normal breast tissue and the believe that an increase in body adipose tissue may have a potentially dangerous local effect, through the increased expression of inflammation-related genes and the establishment of a low-grade chronic inflammation.

Recurrence and mortality dynamics for breast cancer patients undergoing mastectomy according to estrogen receptor status: different mortality but similar recurrence.

(Cancer Sci 2010; 101: 826-830) The purpose was to ascertain whether the recurrence risk patterns for patients with estrogen receptor (ER)-positive (P) and ER-negative (N) breast cancer support the ER-related clinical divergence suggested by the observed different mortality patterns and gene expression profiles. Both recurrence and death were considered in a series of 771 patients undergoing mastectomy. ER status was available for 539 patients. The hazard rates for recurrence and mortality throughout 15 years of follow-up were assessed. The recurrence dynamics displays a bimodal pattern for both ERP and ERN tumors with comparable peak timings. The two curves cross during the 3rd year. By contrast, the mortality dynamics are definitely different for ERP and ERN tumors: during the early follow-up period ERN patients have their highest mortality risk, while ERP patients have their lowest mortality risk. The two curves cross during the 5th year. In spite of the different mortality dynamics, the recurrence dynamics do not demonstrate a major distinction in timing between ERP and ERN breast cancers, suggesting that the metastasis development process following mastectomy is apparently similar for both ER categories. The observed differences in the mortality risk are plausibly attributable to ER-related factors influencing the clinical course from recurrence to death. These clinical findings apparently contradict the occurrence of two different types of breast cancer, notwithstanding the distinct epidemiological, clinical, and molecular features linked to ERP and ERN tumors, although ER levels may concur to establish the event risk levels.

Recurrence and mortality according to estrogen receptor status for breast cancer patients undergoing conservative surgery. Ipsilateral breast tumour recurrence dynamics provides clues for tumour biology within the residual breast.

BACKGROUND: The study was designed to determine how tumour hormone receptor status affects the subsequent pattern over time (dynamics) of breast cancer recurrence and death following conservative primary breast cancer resection. METHODS: Time span from primary resection until both first recurrence and death were considered among 2825 patients undergoing conservative surgery with or without breast radiotherapy. The hazard rates for ipsilateral breast tumour recurrence (IBTR), distant metastasis (DM) and mortality throughout 10 years of follow-up were assessed. RESULTS: DM dynamics displays the same bimodal pattern (first early peak at about 24 months, second late peak at the sixth-seventh year) for both estrogen receptor (ER) positive (P) and negative (N) tumours and for all local treatments and metastatic sites. The hazard rates for IBTR maintain the bimodal pattern for ERP and ERN tumours; however, each IBTR recurrence peak for ERP tumours is delayed in comparison to the corresponding timing of recurrence peaks for ERN tumours. Mortality dynamics is markedly different for ERP and ERN tumours with more early deaths among patients with ERN than among patients with ERP primary tumours. CONCLUSION: DM dynamics is not influenced by the extent of conservative primary tumour resection and is similar for both ER phenotypes across different metastatic sites, suggesting similar mechanisms for tumour development at distant sites despite apparently different microenvironments. The IBTR risk peak delay observed in ERP tumours is an exception to the common recurrence risk rhythm. This suggests that the microenvironment within the residual breast tissue may enforce more stringent constraints upon ERP breast tumour cell growth than other tissues, prolonging the latency of IBTR. This local environment is, however, apparently less constraining to ERN cells, as IBTR dynamics is similar to the corresponding recurrence dynamics among other distant tissues.

Body mass index and γ-glutamyl transferase expression in normal and cancerous breast tissue.

BACKGROUND: Localized to cell membrane, γ-glutamyl transferase (GGT) is a reliable marker for the evaluation of cell distress occurring in several pathological conditions including obesity, metabolic syndrome, and cancer. In particular, high GGT serum levels are associated with breast cancer incidence and progression. METHODS: The tissue expression of GGT1, the gene coding for GGT, was investigated in silico in a large case series of paired samples of breast cancer and adjacent histologically normal (HN) tissue, and in a collection of healthy breast tissues from reduction mammoplasty. The association of GGT1 with patient's body mass index (BMI), and the relationship between GGT1 and a panel of genes involved in apoptosis, IGF-1 signaling, or coding for adipokines and adipokine receptors were also investigated. RESULTS: GGT1 expression was significantly higher in tumor than in the adjacent HN tissue (P = 0.0002). Unexpectedly, the expression of GGT1 was inversely associated with BMI in normal and HN tissue, whereas no correlation was found in cancerous tissue. In all tissues, GGT1 correlated positively with TP53 and negatively with BCL2 and LEPR, whereas only in normal and HN tissue GGT1 correlated positively with IGF1R. The linear regression model, adjusted for BMI, showed no confounding effect on any correlation, except for the correlation of GGT1 with LEPR in normal tissue from healthy women. CONCLUSIONS: Even if present results provide interesting insights on the still elusive mechanism(s) underlying the association between obesity and epithelial cell proliferation, possibly promoting neoplastic transformation, such relationship deserves further investigation in other independent datasets.

Conditional independence relations among biological markers may improve clinical decision as in the case of triple negative breast cancers.

The associations existing among different biomarkers are important in clinical settings because they contribute to the characterisation of specific pathways related to the natural history of the disease, genetic and environmental determinants. Despite the availability of binary/linear (or at least monotonic) correlation indices, the full exploitation of molecular information depends on the knowledge of direct/indirect conditional independence (and eventually causal) relationships among biomarkers, and with target variables in the population of interest. In other words, that depends on inferences which are performed on the joint multivariate distribution of markers and target variables. Graphical models, such as Bayesian Networks, are well suited to this purpose. Therefore, we reconsidered a previously published case study on classical biomarkers in breast cancer, namely estrogen receptor (ER), progesterone receptor (PR), a proliferative index (Ki67/MIB-1) and to protein HER2/neu (NEU) and p53, to infer conditional independence relations existing in the joint distribution by inferring (learning) the structure of graphs entailing those relations of independence. We also examined the conditional distribution of a special molecular phenotype, called triple-negative, in which ER, PR and NEU were absent. We confirmed that ER is a key marker and we found that it was able to define subpopulations of patients characterized by different conditional independence relations among biomarkers. We also found a preliminary evidence that, given a triple-negative profile, the distribution of p53 protein is mostly supported in 'zero' and 'high' states providing useful information in selecting patients that could benefit from an adjuvant anthracyclines/alkylating agent-based chemotherapy.