Pharmacological blockage of the CXCR4-CXCL12 axis in endometriosis leads to contrasting effects in proliferation, migration, and invasion.

High levels of inflammatory factors including chemokines have been reported in peritoneal fluid and blood of women with endometriosis. CXCL12 mediates its action by interaction with its specific receptor, CXCR4, reported to be elevated in human endometriosis lesions and in the rat model of endometriosis. Activation of the CXCR4-CXCL12 axis increases cell proliferation, migration, and invasion of cancer cells. To obtain insights into the CXCR4 expression profile in lesions and endometrium, as well as functionality of the CXCR4-CXCL12 axis in endometriosis, we analyzed the expression of CXCR4 in tissues on a human tissue array and studied CXCL12-mediated activation of proliferation, invasion, and migration in vitro. We observed differences in levels of nuclear CXCR4 expression among lesion types, being higher in ovarian lesions. Endometriotic cell lines (12Z) showed higher levels of CXCR4, proliferative and migratory potential, and AKT phosphorylation/kinase activity compared to untreated control cells (endometrial epithelial cells). CXCL12 and endometriotic stromal cell-enriched media increased proliferation of non-endometriotic epithelial cells. CXCL12 caused a significant increase in 12Z cell invasion but had no effect on migration; AMD3100, a CXCR4-specific inhibitor, significantly increased invasion of 12Z cells but decreased their migration. However, treatment with CXCL12 plus AMD3100 significantly decreased invasion and migration of 12Z cells. In conclusion, the CXCR4-CXCL12 axis is functional in endometriosis cells, but the expression of CXCR4 varies among lesions. CXCL12 promoted proliferation, migration, and invasion of endometriotic cells, while inducing AKT phosphorylation and activity, but pharmacologically blocking this axis in the absence of the ligand induced their invasiveness.

Dysregulation of Lysyl Oxidase Expression in Lesions and Endometrium of Women With Endometriosis.

UNLABELLED: Lysyl oxidases (LOXs) are enzymes involved in collagen deposition, extracellular membrane remodeling, and invasive/metastatic potential. Previous studies reveal an association of LOXs and endometriosis. We aimed to identify the mechanisms activated by upregulation of lysyl oxidases (LOX) in endometriotic cells and tissues. We hypothesized that LOX plays a role in endometriosis by promoting invasiveness and epithelial to mesenchymal transition (EMT). METHODS: The LOX protein expression levels were measured by immunohistochemistry in lesions and endometrium on a tissue microarray (TMA) and in endometrial biopsies from patients and controls during the window of implantation (WOI). Estradiol regulation of LOX expression was determined by quantitative polymerase chain reaction (qPCR). Proliferation, invasion, and migration assays were performed in epithelial (endometrial epithelial cell), endometrial (human endometrial stromal cell), and endometriotic cell lines (ECL and 12Z). Pathway-focused multiplex qPCR was used to determine transcriptome changes due to LOX overexpression. RESULTS: LOX protein was differentially expressed in ovarian versus peritoneal lesions. During WOI, LOX levels were higher in luminal epithelium of patients with endometriosis-associated infertility compared to controls. Invasive epithelial cell lines expressed higher levels of LOX than noninvasive ones. Transfection of LOX into noninvasive epithelial cells increased their migration in an LOX inhibitor-sensitive manner. Overexpression of LOX did not fully induce EMT but the expression of genes related to fibrosis and extracellular matrix remodeling were dysregulated. CONCLUSIONS: This study documents that expression of LOX is differentially regulated in endometriotic lesions and endometrium. A role for LOX in mediating proliferation, migration, and invasion of endometrial and endometriotic cells was observed, which may be implicated in the establishment and progression of endometriotic lesions.

[Breast cancer in México: a 10-year trend analysis on incidence and age at diagnosis].

INTRODUCTION: Breast cancer is an important public health problem. Some countries have achieved a downward trend while in others, continues ascending. In México, information on incidence and age at diagnosis is isolated in time, and knowledge on trend analysis is lacking. OBJECTIVE: To examine the 2003-2012 trend of the incidence rate and age at diagnosis of breast cancer in the northeast of México. We also analyze the trend of positivity to nodes, hormone receptors and HER2; and its association with age at diagnosis. MATERIAL AND METHODS: This is an epidemiological study of breast cancer patients in a tertiary care hospital in Monterrey, México (n = 3,488). Only new cases with a histology report were included; if this was not available, the cytology result was considered. Trend analysis was performed using the JoinPoint regression program Version 3.5. RESULTS: The breast cancer incidence rate increased from 26.7 to 49.8 per 100,000 between 2003 and 2011 (p < 0.05). The adjusted rate showed an annual percentage rate of change of +6.2% (95%CI 4.2, 8.2). The mean age was 55.7 ± 13.7 years and remained stable over time. Nodes, hormone receptors and HER2 positivity rate also remained stable over time. Age < 50 years increased twice the risk for positivity to nodes (OR 2.0, 95%CI 1.4, 2.7), ER-PR- (OR 1.8, 95% CI 1.4, 2.4) and ER-PR-HER2- (OR 1.9, 95%CI 1.5, 2.5). CONCLUSIONS: The 10-year analysis showed a significant upward trend. This study represents a first effort in our country, for determining patterns on incidence and age at diagnosis of breast cancer, as well as that of biomarkers.

HDAC1 and HDAC2 are differentially expressed in endometriosis.

Epigenetic mechanisms have been ascribed important roles in endometriosis. Covalent histone modifications at lysine residues have been shown to regulate gene expression and thus contribute to pathological states in many diseases. In endometriosis, histone deacetylase inhibition (HDACi) resulted in reactivation of E-cadherin, attenuation of invasion, decreased proliferation of endometriotic cells, and caused lesion regression in an animal model. This study was conducted to assess basal and hormone-regulated gene expression levels of HDAC1 and HDAC2 (HDAC1/2) in cell lines and protein expression levels in tissues. Basal and steroid hormone-regulated HDAC1/2 gene expression levels were determined by quantitative polymerase chain reaction in cell lines and tissues. Protein levels were measured by immunohistochemistry (IHC) in tissues on an endometriosis tissue microarray (TMA). Basal HDAC1/2 gene expression levels were significantly higher in endometriotic versus endometrial stromal cells, which was confirmed by Western blot analysis. Estradiol (E2) and progesterone (P4) significantly downregulated HDAC1 expression in endometrial epithelial cells. Levels of HDAC2 were upregulated by E2 and downregulated by E2 + P4 in endometrial stromal cells. Hormone modulation of HDAC1/2 gene expression was lost in the endometriotic cell line. Immunohistochemistry showed that HDAC1/2 proteins were expressed in a substantial proportion of lesions and endometrium from patients, and their expression levels varied according to lesion localization. The highest proportion of strong HDAC1 immunostaining was seen in ovarian, skin, and gastrointestinal lesions, and of HDAC2 in skin lesions and endometrium from patients with endometriosis. These studies suggest that endometriosis etiology may be partially explained by epigenetic regulation of gene expression due to dysregulations in the expression of HDACs.

Basal and steroid hormone-regulated expression of CXCR4 in human endometrium and endometriosis.

Endometriosis is associated with activation of local and systemic inflammatory mechanisms, including increased levels of chemokines and other proinflammatory cytokines. We have previously reported increased gene expression of chemokine receptor 4 (CXCR4), the receptor for CXCL12, in lesions of the rat model of endometriosis. The CXCR4-CXCL12 axis has been shown to have both immune (HIV infection, lymphocyte chemotaxis) and nonimmune functions, including roles in tissue repair, angiogenesis, invasion, and migration. There is evidence indicating that these mechanisms are also at play in endometriosis; therefore, we hypothesized that activation of the CXCR4-CXCL12 axis could be responsible, at least in part, for the survival and establishment of endometrial cells ectopically. Immunohistochemistry (IHC) showed that CXCR4 protein levels were significantly higher in endometriotic lesions compared to the endometrium of controls. Next, we determined basal gene and protein expression of CXCR4 and CXCL12 and regulation by estradiol (E2) and/or progesterone (P4) in endometrial cell lines using quantitative polymerase chain reaction (qPCR), and Western blots. Basal CXCR4 gene expression levels were higher in epithelial versus stromal cells; conversely, CXCL12 was expressed at higher levels in stromal vs epithelial cells. CXCR4 gene expression was significantly downregulated by ovarian steroid hormones in endometrial epithelial. These data suggest that steroid modulation of CXCR4 is defective in endometriosis, although the specific mechanism involved remains to be elucidated. These findings have implications for future therapeutic strategies specifically targeting the inflammatory component in endometriosis.

The role of DNA repair capacity in melanoma skin cancer risk in a population chronically exposed to high levels of sunlight.

Puerto Rican residents are exposed to some of the highest levels of environmental ultraviolet radiation in the world; paradoxically, the melanoma incidence in Puerto Rico is lower than that of the US mainland. The overall objective of this case-control pilot study was to test the hypotheses that (1) persons with melanoma have a significantly lower DNA repair capacity (DRC) in relation to controls matched by age, (2) decline in DRC is associated with vertical depth of melanoma invasion, and (3) DRC is associated with anatomical tumor location. Controls (n  =  124) were examined by dermatologists; cases (n  =  62) were histopathologically confirmed. The mean DRC ± 1 SE of controls was 6.46% ± 0.3. Melanoma patients (n  =  62) had a mean decrease in DRC of 3% (6.25% ± 0.5), which was not statistically different from controls (P  =  0.697). No significant differences in DRC were evident in participants with either in situ or malignant melanoma tumors; neither were such differences evident when evaluating anatomical location of tumors (ie, non-sun-exposed versus sun-exposed). DRC generally declined in participants with increased depth of melanoma tumor penetration when compared with controls and those with small in situ tumors. These findings should be examined in a larger-scale population study that includes participants with more advanced metastatic melanoma.

Early-onset of sporadic basal-cell carcinoma: germline mutations in the TP53, PTCH, and XPD genes.

BACKGROUND: Basal cell carcinoma (BCC) is the most common non-melanoma skin cancer in the Western world. The objective of this study was to examine together germline mutations in the TP53, PTCH, and XPD genes as risk factors for developing BCC at a young age. We hypothesized that mutations in these genes significantly increase the risk of early-onset BCC (< or = 35 years). METHODS: The PCR, DNA sequencing and Restriction Fragment Length Polymorphisms methods were utilized to study eight Puerto Rican patients with a confirmed diagnosis of BCC before age 35. RESULTS: A novel germline mutation (T:A transversion) was identified at the exon 4, codon 50 of the TP53 gene of one BCC patient. No other mutations were found at the TP53 or PTCH genes. The presence of the XPD mutant allele is associated with a seven-fold increase in risk (OR = 7.0, p = 0.03) for developing BCC prior to age 35. In addition, the DNA Repair Capacity (DRC) of these BCC patients showed a 47% reduction that was significant in relation to age-matched controls (p = 0.021). However, the XPD mutant allele was not associated with the decrease in DRC observed in BCC participants. CONCLUSIONS: The evaluated population presented BCC before age 35, a phenomenon that is so rare as to make very difficult the study of this subpopulation with a larger sample size. The results of this study, suggest that the XPD Lys751Gln polymorphism may have a significant role in the development of early-onset BCC in the Puerto Rican population.

UV dose determines key characteristics of nonmelanoma skin cancer.

Basal cell carcinoma (BCC) and squamous cell carcinoma (SCC), known as nonmelanoma skin cancer (NMSC), are the most common cancers worldwide. Although many factors are involved in the pathogenesis of NMSC, UV radiation is an important risk factor. A fundamental question in skin cancer research is whether varying doses of total UV radiation influence key characteristics of NMSC. The hypothesis that differences in UV doses influence the BCC/SCC ratio, number of tumors, and anatomic location of the tumor was investigated in 311 participants having 326 tumors and with exposure to a broad range of UV doses. An epidemiologic questionnaire was given to each participant soliciting detailed information on exposure to solar radiation. Environmental UVA and UVB doses were measured continually for 6 years at a permanent UV monitoring station. The total ratio of BCC/SCC was 3.5. Participants who received low and high UV doses had a BCC/SCC ratio of 4.2. Those who received very high UV doses had a ratio of 2.1. A very high UV dose was also associated with the doubling of the total number of tumors per person and a significantly increased risk of having SCC, a more aggressive malignancy. Tumors in sun-exposed areas (on the body) were more common in participants who received high and very high UV doses. The tumors in sun-protected areas were associated with exposure to lower levels of UV. This large-scale population study provides evidence that varying doses of UV radiation have a profound influence on key characteristics of NMSC.

DNA repair and breast carcinoma susceptibility in women.

BACKGROUND: Breast carcinoma is the most common cancer and the second leading cause of cancer-related deaths among women. The disease represents approximately 31% of all cancers in Puerto Rican women. Several DNA repair pathways are involved in preventing carcinogenesis. The current study evaluated the hypothesis that a reduced DNA repair capacity (DRC) is a susceptibility factor for breast carcinoma. METHODS: A retrospective case-control clinical study was performed to compare age-matched DRC in 33 women with histopathologically confirmed breast carcinoma (cases) and 47 cancer-free women (controls). DRC was measured using a host cell reactivation assay with a luciferase reporter gene and then transfected into human peripheral lymphocytes. A questionnaire was used to solicit breast carcinoma risk factors. RESULTS: Women with breast carcinoma had a mean DRC of 5.6% +/- 0.5 standard error of the mean (SEM). Cancer cases had a 36% reduction (P<0.001) in DRC when compared with the control group (DRC=8.7% +/- 0.7 SEM). Younger participants with breast carcinoma were found to have a more significant reduction in DRC when compared with age-matched controls. Family (odds ratio [OR]=4.1), maternal lineage (OR=5.5), and maternal (OR=12.4) history of breast carcinoma were found to be the only statistically significant (P<0.05) risk factors associated with the disease. CONCLUSIONS: The findings supported the hypothesis that a low DRC is a susceptibility factor for breast carcinoma. A 1% decrease in DRC corresponded to a 22% increase in breast carcinoma risk. To the authors' knowledge, the current study was the first to directly determine the DRC of women with breast carcinoma. Because DRC is an independent risk factor for breast carcinoma, the DRC of women may be a useful marker in predicting susceptibility.

DNA repair and nonmelanoma skin cancer in Puerto Rican populations.

BACKGROUND: UV radiation is a risk factor for nonmelanoma skin cancer (NMSC). The relation between DNA damage and oncogenesis suggests that diminished DNA repair capacity (DRC) is involved in tumorigenesis. OBJECTIVE: The purpose of this study was to test the hypothesis that a low DRC is a susceptibility factor for the development of NMSC in Puerto Rico. METHODS: A case-control retrospective clinical study was done to compare the age-adjusted DRC in participants with and without NMSC. DRC was measured using a host cell reactivation assay with a luciferase reporter gene irradiated with UV light and transfected into human peripheral lymphocytes. An epidemiologic questionnaire was used to solicit risk factors. RESULTS: The mean (+/-2 SE) DRC of 177 control patients without skin cancer was 8.6% +/- 0.7. Participants (280) with NMSC had a 42% lower DRC (5.0% +/- 0.3). CONCLUSION: A low DRC is a susceptibility factor for NMSC.